Ein intaktes und funktionsfähiges Endothel ist eine entscheidende Vorraussetzung für ein funktionierendes Herz-Kreislaufsystem. Eine Dys-funktion des Endothels ist ein zentrales Glied in der Pathogenese zahlreicher Gefäßkrankheiten (Hypertonie, Atherosklerose, u.a.). Mehrere Mediatoren, welche die vasodilatierende und vasokontraktile Wirkungen des Endothels regulieren, sind für ein physiologisches Zusammenspiel der vernetzten Regel-kreise des Endothels verantwortlich. Ziel dieser Arbeit war es, den Einfluss von Östrogenen (Ethinylöstradiol) und Gestagenen (Medroxy-Progesteron-Acetat, MPA) bzw. von Drospirenon (aus Ethinylöstradiol und Gestagen) bei ovarektomierten Ratten auf das Gefäß-endothel und die Vasomotorik zu untersuchen. Hierfür wurden isolierte, vorgespannte Segmente der Aorta descendens der Versuchstiere in der Organkammer untersucht. Es erfolgte die Untersuchung der Endothel-abhängigen Gefäßreaktion als dosisabhängige Relaxation auf Acetylcholin mit und ohne Vorinkubation mit Diclofenac bzw. L-NAME, sowie der Endothel-abhängigen Gefäßreaktion auf Sodium-Nitroprussid. Zusätzlich wurde die dosisabhängige Kontraktion auf Norepinephrin bei Vorinkubation mit Diclofenac bzw. L-NAME evaluiert. Die Endothel-abhängige Relaxation auf Acetylcholin ergab bei den mit Drospirenon sowie Spironolacton behandelten, ovarektomierten Tieren ein Relaxationsverhalten, wie es auch typisch für unbehandelte Wildtyp-Ratten ist. Bei den Placebo-behandelten Tieren zeigte sich jedoch eine deutliche Beeinträchtigung der Endothel-abhängigen Relaxation, welche sich durch eine Präinkubation mit dem Arachidonsäure-Synthese-Hemmer Diclofenac egali-sieren liess. Eine stärkere Einschränkung der Endothel-abhängigen Relaxation zeigte sich bei den mit MPA-behandelten Tieren, bei denen sich auch durch Präinkubation mit Diclofenac keine Normalisierung der Gefäßreaktion erzielen ließ. Eine Vorbehandlung mit dem NOS-Inhibitor L-NAME führte zu einem vollständigen Ausfall der Endothel-abhängigen Relaxation mit einer bei steigender Ach-Dosierung auftretenden „paradoxen“ Kontraktion. Bei den mit MPA-behandelten Ratten war dieser Effekt nicht zu beobachten. Die Endothel-unabhängige Relaxation auf SNP war in allen Gruppen ver-gleichbar, mit Ausnahme der Tiere, die MPA erhielten. Bei diesen Tieren war die Endothel-unabhängige Gefäßantwort beeinträchtigt. Die Kontraktion auf Norepinephrin bei Präinkubation mit Diclofenac fiel in allen Gruppen einheitlich aus. Auch bei Präinkubation mit L-NAME zeigte sich in allen Versuchsgruppen eine vergleichbare Gefäßreaktion, wobei die Kontraktion hier in allen Gruppen diskret stärker ausfiel, als ohne Gegenwart von L-NAME. Zusammenfassend konnte in dieser Arbeit gezeigt werden, dass Ethinyl-östradiol zu keiner funktionell relevanten Beeinträchtigung der Endothelfunktion führt. Medroxy-Progesteron-Acetat hingegen führte in dieser Versuchs-anordnung zu einer deutlichen Einschränkung der Endothelfunktion und beeinträchtigte die Relaxationsfähigkeit der Gefäße erheblich. Im Gegensatz zu Progesteron konnte unter den Bedingungen dieser Arbeit zumindest kein Endothel-schädigender Effekt für Östrogene nachgewiesen werden. Inwieweit Östrogene – über längere Zeit appliziert – in der Lage sind, eine „Endothel-protektive“ Wirkung zu entfalten, kann jedoch im Rahmen der vorliegenden Versuchsanordnung nicht abschließend beurteilt werden. / An intact and functional endothel is important for the function of the regulation of blood pressure. A dysfunction of the endothel ist one of the most important figures in the pathogenese of several diseases of the vessels. Several mediators, which regulates the vasodilatatioon and vasoconstriction of the endothel are important for the function of the endothel. In this topic, we investigated the influence of estrogene and gestagene on the endothelial function in ovarectomized female rats. Therefore isolated segments of the aorta descendens of the rats are investigated. We looked for the reaction of the endothel in endothel-dependent, and in endothel-independent reaction of the vessels. The endothel-dependent relaxation of acetylcholine showed an normal relaxation, either by the rats with drospirenone and by the rats with spironolactone treatment. The animals withe placebo-treatment showed and much more worse relaxation. The endothel-independent relaxation on SNP was in all groups the same, only by the rats, which were treated with MPA. In this group the reaction of the endothel was much more worse. The contraction of norepinephrine by preincubation with diclofenac was in all groups similar. Although by preincubation with L-NAME was the reaction in all groups the same, the contraction was in all groups a little bit more than without L-NAME. Finally in this study we coud show, that estrogenes have no negative influence on the endothelial function. Medroxy-progesterone-acetate showed in this study an very negative influence of the endothelial function, and the relaxation of the endothel was by that treatment very bad. In this study we couldnt find any effect of negative influence of estrogene treatment on the endothelial function. If estrogenes - over a longer period applicated - may show protective effects on the endothelial function must be investigated in other studies.

ETB-Rezeptoren nehmen innerhalb der endothelialen Regulationsprozesse eine zentrale Rolle ein. In der vorliegenden Arbeit wurde der Frage nachgegangen, welchen Einfluss eine Salzbelastung auf den Blutdruck und die vaskuläre Funktion von ETB-Rezeptor-Knockout-Mäusen hat. In diesem Zusammenhang wurden männliche ETB-Rezeptor-Knockout-Mäuse parallel mit Wildtyp-Kontroll-Mäusen 15 Tage lang mit Standard- bzw. salzreichem Futter gehalten. Der systolische Blutdruck wurde ebenfalls dokumentiert. Nach 15 Tagen wurde den narkotisierten Tieren die Aorta descendens entnommen. An isolierten Aortenringen wurden in der Organkammer die Endothel-abhängige und -unabhängige vaskuläre Funktion untersucht. Die ETB-Rezeptor defizienten Mäuse bleiben – unter einer Haltung mit Standardfutter – normotensiv. Eine Hypertonie entwickeln die Tiere erst bei Verabreichung von salzreichem Futter. Die Endothel-abhängige Gefäßfunktion ist jedoch nicht nur bei den hypertensiven Tieren verändert, sondern bei allen ETB-Rezeptor defizienten Mäusen – unabhängig von Salzgehalt der Nahrung und Blutdruck. / The ETB receptor is involved in endothelial function. In the present study, we analysed whether salt alters endothelial function in rescued ETB receptor-deficient mice. Adult ETB-deficient mice were kept in parallel with wild-type control animals for 15 days on standard or salt-enriched chow, respectively. Systolic blood pressure was measured also and endothelium-dependent and endothelium-independent vascular function was assessed in isolated aortic rings. Systolic blood pressure increased on salt-enriched chow in ETB receptor-deficient mice, but neither in wild-type mice on high-salt diet nor in ETB receptor-deficient mice on standard chow.

Os relógios biológicos são fundamentais para a sincronização do comportamento dos organismos a mudanças no fotoperíodo. Todas as alterações rítmicas são determinantes para a sobrevivência da espécie uma vez que elas prevêem que os ajustes internos coincidam com a fase mais propícia do ciclo ambiental, permitindo aos organismos a capacidade de sincronizar esses eventos internos com os ciclos ambientais. Muitos desses ritmos biológicos são claramente associados ao ciclo claro-escuro, sendo este ciclo de grande importância para as espécies que possuem algum tipo de pigmento fotossensível. Os melanóforos de Xenopus laevis são fotossensíveis, respondendo à luz com dispersão dos grânulos de melanina, devido à presença de duas melanopsinas, Opn4x e Opn4m. As células pigmentares dos vertebrados heterotérmicos respondem com migração pigmentar a uma variedade de agentes, incluindo as endotelinas. Em peixes teleósteos, ETs induzem a agregação pigmentar em melanóforos, enquanto que em anfíbios, ET-3 induz a dispersão de grânulos de pigmentos em melanóforos de Xenopus laevis e de Rana catesbeiana, através da ativação de receptores ETc. Propusemos determinar o padrão temporal de expressão dos genes das melanopsinas e do receptor ETc em melanóforos dérmicos de X. laevis em cultura, bem como os efeitos temporais e dose- dependentes da endotelina sobre essa expressão. Demonstramos, através de ensaios de PCR quantitativo, que o tratamento de 12C:12E , somado a uma troca de meio, assim como o de endotelina-3 10-9 e 10-8M em escuro constante, foi capaz de sincronizar a expressão de Opn4x e Opn4m. Entretanto, o receptor ETc parece não ser sincronizado pelo ciclo claro-escuro, ou pelo tratamento hormonal. Dependendo da dose utilizada e do ZT analisado, ET-3 pode promover um aumento ou inibição da expressão gênica de Opn4x, Opn4m e ETc, indicando uma modulação de forma dose-dependente. Além disso, pode atuar como um agente sincronizador da expressão dos transcritos das melanopsinas. / The biological clocks are critical for synchronizing the behavior of organisms to changes in photoperiod. All rhythmic changes are crucial to the survival of the species since they provide for internal adjustments to coincide with the phase of the cycle most favorable. Many of these biological rhythms are clearly associated with the light-dark cycle, of major importance for species that have some type of photosensitive pigment. Melanophores of Xenopus laevis are photosensitive, responding to light with dispersion of melanin granules, due to the presence of two melanopsins, Opn4x and Opn4m. The pigment cells of ectothermic vertebrates respond with pigment migration to a variety of agents including the endothelins. In teleost fish, ETs induce pigment aggregation in melanophores, whereas in amphibians, ET-3 induces the dispersion of pigment granules in melanophores of Xenopus laevis and Rana catesbeiana, by activation of ETc. We proposed to determine the temporal pattern of gene expression of the ETc receptor and melanopsins in dermal melanophores of X. laevis in culture as well as the effects of endothelin-3 on the temporal expression of the 3 genes. Using quantitative PCR, we demonstrated that 12L: 12D regimen, combined with medium changes, as well as the treatment with 10-9 and 10-8M endothelin-3, was able to synchronize the expression of Opn4x and Opn4m. However, ETc receptor seems not to be synchronized by light-dark cycle, or hormone treatment. Depending on the dose and the ZT, ET-3 may promote an increase or inhibition of gene expression of Opn4x, Opn4m and ETc, indicating a dose-dependent modulatory effect. In addition, endothelin-3 may also act as a synchronizing agent of the melanopsins transcripts.

Inflammation has incompletely characterized effects on cardiopulmonary vascular reactivity. Sepsis is a major inflammatory disease characterized by two main vasomotor complications, generalized vasodilation with hyporesponsiveness to vasoconstrictors and pulmonary hypertension. The main aim of this study is to examine the molecular mechanisms involved in cardiopulmonary vascular reactivity changes induced by the powerful inflammatory stimulus lipopolysaccharide (LPS). Pulmonary and aortic rings from male Wistar rats (250-300g) were isolated and incubated for 20 h in culture medium (DMEM+10% FBS) with or without LPS (E. coli O55:B5, 10 μg.ml⁻¹). The effect of organ culture and LPS type, concentration and incubation time in addition to tissue contraction to endothelin-1 (ET-1), phenylephrine, 80 mM KCl, and U46619; and relaxation responses to ACh, sodium nitroprusside (SNP), 8-pCPT-cGMP, BAY 41-2272, T-0156, nifedipine, SKF-96365, Ro-31-8425, and Y-27632 were measured by standard organ bath techniques. Nitric oxide (NO) production was measured by the Griess method and SNP-induced cGMP production was measured by ELISA. mRNAs expression levels of eNOS, iNOS, ET-1, ETA and ETB were measured by qRT-PCR and the expression levels of PKC, sGCα₁, sGCβ₁ and PDE5 and phosphorylation of MLC₂₀, ROKα, CPI-17 and MYPT1 were measured by immunoblotting. The effect of endothelium removal, indomethacin, trolox, external Ca²⁺ removal, 1400W, ODQ, glibenclamide, iberiotoxin and cycloheximide in addition to changes in intracellular Ca²⁺ ([Ca²⁺]i) in aortic vascular smooth muscle cells (VSMCs) induced by ET-1 were also measured. LPS selectively induced vascular hyporeactivity to different vasoconstrictors in rat aorta but not in the pulmonary artery, which is not due to organ culturing and is not affected by changing the LPS type, but is enhanced by increasing LPS concentration or the incubation time. This aortic hypocontractility to ET-1 is largely mediated by NO-independent activation of sGC and depends on external Ca²⁺ influx through non-VOCCs, but not on ET-1 receptor expression or Ca²⁺ sensitization. In addition, this aortic hyporeactivity to ET-1 is dependent on protein synthesis. The pulmonary artery is not affected because LPS induces a desensitization of the sGC/cGMP dependent pathway by decreasing protein expression levels of sGCβ₁, and hence sGC activity, and increasing PDE5 activity. Neither the endothelium, cyclooxygenase, reactive oxygen species nor K⁺ channels are involved in these LPS effects. Therefore, it is likely that both Ca²⁺ homeostasis and the sGC/cGMP pathway play important roles in vasomotor complications in sepsis. sGC and/or PDE5-selective inhibitors, together with manipulating VSMC [Ca²⁺]i, could be important in controlling systemic and pulmonary vasomotor complications in sepsis.

Pigment patterns of zebrafish are a beautiful example in which to study key processes of vertebrate development such as neural crest cell migration and patterning of neural crest-derived cell types. This can for example be achieved by characterizing mutants like parade in which pigment cell development is abnormal. Here, we present a chemico-genetic study of the pigment pattern mutant parade; uniquely, this mutant displays ectopic pigment cells in the ventral medial pathway of the trunk but the characteristic stripe pattern of zebrafish embryos is unaffected. Using a positional cloning approach, we have identified the parade gene as the cell surface receptor ednra2. This was further confirmed in transient knock-down assays. Combined sequencing data from three different parade alleles strongly indicates that the mutation disrupts ednra2 receptor function by deleting C-terminal regulatory and structural residues. To expand the available molecular tools in pigment cell research, notably to chemically dissect the parade phenotype, we have participated in small molecule screening for inhibitors of pigment cell development. From this, we have isolated 57 compounds which robustly alter the development of melanophores and iridophores in wild-type embryos; 26 of these compounds additionally affect the parade phenotype, primarily by rescuing the ectopic pigment cells. Notably, chemical rescue has shown that the MEK pathway is important for the development of the parade phenotype. Our study therefore adds to our understanding of pigment pattern formation in zebrafish embryos and reveals novel functions for ednra2 in dorso-ventral patterning and cell type specification of neural crest derivatives.

A common feature of glomerular disease is a protein leak into the urine. Proteinuria occurs in kidney disease and is an important risk factor for cardiovascular disease (CVD). ET‐1 is a potent vasoconstrictor/pressor peptide that is up‐regulated in CVD and many forms of inflammatory renal diseases. The actions of ET‐1 are mediated via two G‐protein coupled receptors, the ETAR which serves primarily in the pro‐hypertensive actions of ET‐1 and is often considered as the main pathological receptor subtype, with the ETBR serving to clear circulating ET‐1. Antagonism of one or both of receptors has been shown to be of clinical benefit in the treatment of hypertension. This research demonstrated a beneficial effect of selective ETAR antagonism using Sitaxsentan in a rat model of GN. ETAR blockade reduced blood pressure and importantly reduced glomerular inflammation as assessed by glomerular macrophage (Mϕ) infiltration. Further, we aimed to demonstrate that Mϕ, key mediators of inflammation are activated by ET‐1 to adopt a pro‐inflammatoy phenotype. However, early studies demonstrated that ET‐1 does not activate Mϕ as hypothesised. Mϕ were more phagocytic, and ET‐1 was chemokinetic for macrophages, an ETBR medicated event. ET‐1 was also removed by Mϕ, suggesting a potential regulatory role of Mϕ in the ET system. This phenomenon led to inclusion of additional in vivo studies to investigate the role of Mϕ in the regulation of ET‐1 and its pressor effects. These effects were investigated in a murine model of Mϕ ablation using CD11b‐DTR mice. These experiments determined in vivo that Mϕ ablation augments pressor responses to ET‐1, suggesting that Mϕ are required to regulate ET‐1. In vitro, Mϕ remove ET‐1 by several mechanisms involving proteolytic degradation of the peptide and ETBR mediated clearance, demonstrating a potential mechanism for the in vivo observation. Furthermore, proteinuria is believed to be due to damage or effacement of specialized visceral glomerular epithelial cells or podocytes. We identified in vitro that the ETAR mediates ET‐1 induced human podocyte cell effacement by actin cytoskeleton aberrations and slit‐diaphragm protein down-regulation, ET‐1 and pro‐inflammatory cytokine production. This thesis provides evidence to support our initial hypotheses that selective ETAR antagonism ameliorates proteinuric renal disease via its effects on podocytes and macrophages. Continued studies both in vitro and in vivo will strengthen the body of evidence to promote the therapeutic use of ETR antagonists in inflammatory renal disease.

Cardiovascular disease (CVD) is highly prevalent in chronic kidney disease (CKD) patients. Whilst this can in part be explained by the high incidence of traditional CVD risk factors such as hypertension and diabetes evident in CKD patients, recent focus has been on non-traditional risk factors and their role in CVD progression. These include endothelial dysfunction, arterial stiffness, inflammation and oxidative stress. The potent vasoconstrictor endothelin-1 (ET-1) has been implicated in the pathogenesis of CKD and the CVD associated with it. Further understanding of the mechanisms by which it contributes to CKD and CVD pathogenesis, specifically its interactions with non-traditional risk factors are still required. Additionally, the potential applications of ET antagonists in renal disease have not been fully explored. This thesis aims to investigate the role of ET-1 in the development of renal disease and the associated inflammation, hypertension and vascular dysfunction through a series of in vitro, in vivo and clinical studies. I have demonstrated using in vitro techniques that murine macrophages (Mϕ) express both endothelin A (ETA) and endothelin B (ETB) receptors but that ET-1 does not elicit either a classical pro-inflammatory or alternative anti-inflammatory phenotype in Mϕ. I was however, able to show that M display chemokinesis towards ET-1 and M ETB receptors provide a novel clearance mechanism for ET-1 through receptor mediated dynamin-dependent endocytosis In an in vivo study I investigated whether ET-1 mediates the progressive renal injury after renal ischaemia reperfusion injury (IRI) that leads to the development of CKD. I demonstrated that endothelin A receptor antagonism provided long term beneficial effects reducing blood pressure and preventing progressive kidney injury, inflammation, and the development of fibrosis resulting from an episode of acute kidney injury (AKI). Similar benefits were observed with calcium channel blockade, suggesting hypertension may mediate some of the long term effects of renal IRI and anti-hypertensive treatments could prevent the development of CKD after AKI. Finally, in a clinical study I showed for the first time that CKD patients lack the diurnal variation in arterial stiffness that is seen in matched subjects without CKD. Alteration in the circadian variation of the ET-1 system may contribute to this. In summary, my studies have furthered our understanding of the role of ET-1 in CKD progression and the cardiovascular risk associated with it. Mϕ were shown to express both ET receptors and a novel mechanism of ET-1 clearance was observed in Mϕ. Using an in vivo model of AKI I was able to identify ETA receptor antagonism as a novel therapeutic agent in preventing the development of CKD caused by AKI where data are limited. Finally, alterations in the circadian rhythm of the cardiovascular system is emerging as an important factor in disease pathogenesis. Here the diurnal variation in arterial stiffness was described for the first time in a group of CKD patients and matched controls.

Tight control of blood glucose and blood pressure (BP) reduces cardiovascular risk in early Type 1 diabetes mellitus (T1DM). Increased BP normally increases renal medullary perfusion and sodium excretion. This is called acute pressure natriuresis. Inadequate acute pressure natriuresis disrupts circadian regulation of BP, which predicts hypertension. The peptide, endothelin-1 (ET-1), regulates BP via ETA and ETB receptors. ETA receptor antagonists reduce BP and restore its circadian rhythm. Two hypotheses were investigated. First, that acute pressure natriuresis is impaired in early T1DM, prior to established nephropathy, and this is associated with elevated BP. Second, that the mechanism is an ETA receptor-mediated blunting of medullary perfusion which can be reversed with insulin and ETA receptor antagonism. Experimental acute pressure natriuresis was induced in young, early T1DM (2-3 weeks post streptozotocin) Sprague Dawley rats and healthy controls. Despite maintaining glomerular filtration rate, early T1DM suppressed urinary flow (UV, 22.9±2.9 v. 93.7±11.1μl/min/gkw) and sodium excretion (UNaV, 3.2±0.7 v. 22.7±3.3μmol/min/gkw) rates by >80%, and reduced gradients of pressure diuresis (linear, 1.9 to 0.3) and natriuresis (non-linear k, 0.05 to 0.01) curves. Insulin treatment lowered blood glucose (16.8±1.8 to 9.3±0.6mmol/l) and restored gradients of the responses. Tissue and urine analyses did not suggest structural nephropathy. In early T1DM rats, changes in BP on radiotelemetry were consistent with impaired circadian regulation of BP and precursors of hypertension: 24-hour diastolic BP rose (92.3±0.4 to 97.1±0.5mmHg), and the circadian dip in diastolic BP fell (6±1 to 2±1%). Atrasentan (ETA receptor antagonist, 5mg/kg/day orally) reduced diastolic dipping in early T1DM (3±1 to 1±1%) while additional ETB receptor antagonism (A-192621, 10mg/kg/day orally) reversed this, suggesting that ETA, and not ETB receptors, mediate impairment of acute pressure natriuresis. To address this, renal blood flow was measured during experimental acute pressure natriuresis and ET receptor antagonism. Early T1DM suppressed the normal rise in medullary perfusion (flux, 227.2±26.7 v. 115.4±10.3%) by ~90%. Suppressed medullary flux was unaffected by insulin (112.2±6.8%), despite restoration of UV and UNaV. In controls, atrasentan reduced UV (15.7±4.9 v. 38.6±6.2μl/min/gkw), UNaV (1.7±0.5 v. 16.7±1.4μmol/min/gkw), FENa (3.4±1.4 v. 15.0±2.4%) and medullary flux (122.2±26.7%) by 60 to 90% of control values, while A-192621 increased UNaV (26.6±6.9μmol/min/gkw) and FENa (21.6±3.4%), but not medullary flux, by ~50%. ET receptor antagonism did not modify early T1DM+/-insulin effects. Diabetic status had no effect on renal ET-1 and ET receptor expression. These results support the first hypothesis but disprove the second. Early T1DM blunts medullary perfusion and acute pressure natriuresis, and increases diastolic BP. Insulin restores natriuresis but not medullary flow. Therefore, targeting medullary perfusion may reduce cardiovascular risk in early T1DM, but this is not achievable with selective ETA receptor antagonists. Novel natriuretic (ETA) and anti-natriuretic (ETB) roles for ET receptors, which are not apparent in early T1DM during severe, experimental rises in BP, appear to contribute to daily regulation of BP, and may preclude the use of selective ETA receptor antagonists in T1DM prior to nephropathy.

- / Background: Endothelin-1 (ET-1), a strong vasoconstrictive substance acting via stimulation of specific receptors (ET-A and ET-B), has been implicated in the development of renal scarring. Activation of endothelin system was observed in experimental models of glomerular diseases and this was attributed to the toxic action of proteinuria to the tubular epithelial cells. However, we have not enough information about the role of endothelin system in human glomerular diseases and in renal diseases without proteinuria like obstructive nephropathy. The aim of this study was to examine the endothelin system in patients with primary glomerular diseases and in experimental animals with unilateral ureteric obstruction. Patients and Methods: Thirty-seven patients with different types of primary glomerulonephritides and 14 controls were included in the study. Patients presented by either nephrotic syndrome (n=25) or mild proteinuria (<1g/24h, n=12). The expression of ET-A and ET-B receptors in the renal tissue was examined immunohistochemically. At the time of biopsy, urinary ET-1 was determined by RIA. Experimental animals and Methods: Twenty –day old opossum pups (n=6) underwent surgical ligation of the left ureter. Sham operated animals, non-operated controls and normal human kidneys were also used. Animals were sacrificed at 2 (n=2), 3 (n=1), 4 (n=1), 5(n=1) and 8 (n=1) weeks post surgery and their kidneys were examined. Sham operation was performed at equivalent times in pups that served as control. The expression of ET-A and ET-B receptors in the renal tissue was examined immunohistochemically. Results: The expression of both receptors was mainly localized within tubular epithelial cells and was significantly higher in patients with glomerulonephritides compared to controls. The expression of ET-B receptors was higher in nephrotic compared to non-nephrotic patients while no difference was observed in the expression of ET-A receptors. Urinary excretion of ET-1 was increased in patients compared to healthy subjects (579±146 ng/24h vs. 410±78 ng/24h, p<0.01) and it was higher in nephrotic compared to non-nephrotic patients (617±167 ng/24h vs. 485±71 ng/24h, p<0.05). A significant positive correlation of the excreted ET-1 with the degree of proteinuria (r= 0.338, p<0.05) and the extent of immunostaining for ET-B receptors (r=0.427, p<0.05) was observed. The expression of ET-B receptors and the excretion of ET-1 were significantly decreased in patients who present remission of the nephrotic syndrome under immunosuppressive therapy. In tubular epithelial cells of the experimental animals there was a temporal increase in the expression of ET-A receptors with duration of obstruction while there was no significant difference between the expression of ET-B receptors in obstructed kidneys and controls. Conclusions: this study provides evidence that the endothelin system is activated in renal diseases and proteinuria seems to be related only in part to this activation. Further investigation is needed to ascertain if the activation of endothelin system has a causative role in the progression of renal diseases.

Motivation: The prevalence of obesity is the highest among African women in South Africa. Since obesity is a major cardiovascular risk factor, African women in South Africa could be regarded as a high risk group. However, investigations on obesity-related hypertension are limited in this population group. The associations of body fat distribution and hormones such as leptin and endothelin-1 with cardiovascular function have not yet been determined in these women. It has been determined that endothelin-1 is a role player in the development and/or maintenance of hypertension in various population groups, especially African Americans. Endothelin-1 has also been found to be involved in obesity-related hypertension in non-African population groups. It has been indicated that the obesity-related hormone, leptin, also plays a role in obesity-related hypertension, especially in African Americans. Leptin levels have been found to be higher in obese hypertensive African American women compared to an obese normotensive control group. Since the above-mentioned two hormones playa prominent role in obesity and hypertension in African American and non-African population groups, the lack of data on African women in South Africa serves as motivation to conduct this investigation. Aim: To investigate obesity-related hypertension in African women through the determination of associations between various anthropometric and endocrinological variables with cardiovascular, especially vascular function. Methodology: Manuscripts presented in Chapters 2, 3 and 4 made use of data from the POWIRS (Profiles of Obese Women suffering from the Insulin Resistance Syndrome) I project where African women were selected from a government institution in the North West Province. A group of 98 women were divided into lean normotensive, overweight/obese normotensive and overweight/obese hypertensive groups. Anthropometric and cardiovascular measurements were taken and the lipid profile, leptin and endothelin-1 levels determined. The analysis of covariance (ANCOVA) was used to show significant differences between groups while adjusting for age. Partial correlation coefficients were used to show associations between various variables while adjusting for age. Stepwise linear regression analysis was also used to show associations between variables. The study presented in Chapter 5 made use of both POWIRS I and II, which are studies including Africans and Caucasians, respectively. The methodology of the two studies was the same. All subjects gave informed consent in writing and the Ethics Committee of the North-West University approved the study. The reader is referred to the "Materials and Methods" section of Chapters 2-5 for a more elaborate description of the subjects, study design and analytical methods used in each article. vii Results and conclusions of the individual manuscripts > Results from Chapter 2 showed that the volume loading effect associated with obesity was present in both overweight/obese normotensive and overweight/obese hypertensive groups, however, the accommodating effect observed in the overweight/obese normotensive group was absent in the overweight/obese hypertensive group due to decreased vascular function. This was confirmed by a high pulse pressure. Decreased vascular functioning was associated with the abdominal skin fold. This suggests that abdominal subcutaneous fat may either be a marker of visceral fat, or may in itself contribute to increased cardiovascular risk in Africans. > Results from Chapter 3 showed a negative result. Plasma endothelin-1 levels were similar for the lean normotensive, overweight/obese normotensive and overweight/obese hypertensive groups. After re-dividing the groups into normotensive and hypertensive, and then into lean and overweight/obese, still no differences could be obtained. Additionally, no correlations could be obtained between endothelin-1 and cardiovascular function in any of the groups. These findings suggest that endothelin-1 is not implicated in obesity-related hypertension in African women. > In Chapter 4, leptin levels were elevated in both overweight/obese normotensive and hypertensive groups compared to the lean normotensive group. However, leptin levels did not differ between the two overweight/obese groups. Even though leptin levels were the same, leptin was directly and positively associated with systolic blood pressure and pulse pressure and negatively with arterial compliance only in the overweight/obese hypertensive group, independent of obesity, insulin resistance, hyperinsulinemia and age. > In Chapter 5 the volume loading, as well as the accommodating effect, that is, decreased total peripheral resistance and increased arterial compliance, was present in both African and Caucasian obese groups compared to their lean controls. Even though leptin levels, body mass index and age were similar for both African and Caucasian obese groups, the accommodating effect seemed to be more prominent in the obese Caucasian group, explaining a lower diastolic blood pressure compared to the obese African group. Leptin showed a favourable negative association with diastolic blood pressure and total peripheral resistance in the obese Caucasian group, but not in the obese African group. This may indicate that leptin predominantly exerts pathological influences on obese African women, as determined previously in Chapter 4. / Thesis (Ph.D. (Physiology))--North-West University, Potchefstroom Campus, 2005.