No description available.

The effects of the antihypertensive agent Losartan (Lo), on acute exercise performance was assessed in six sedentary, hypertensive women. The purpose, benefits and potential risks of the study were explained to each subject and their informed consent received. In a double blinded crossover design subjects were randomized to 7 days of (Lo) 50 mg, once every morning or placebo (Pl). Subjects reported to the laboratory for an exercise trial on the 7th treatment day. They received the final treatment dose 2.5 hours before the exercise trial. Blood samples for analysis of plasma renin activity (PRA) and Angiotensin II (Ang II) were obtained 15 min before the exercise trial began. In each trial, the subject rested for 15 min in a seated position on the stationary cycle. Hemodynamic and respiratory measurements were obtained. They began exercise at a workload equivalent to 45% VO2 pk for 15 min, immediately followed by a progression of 30 Watt*2 min-1 until volitional fatigue. Measurements included: blood pressure, heart rate, respiratory gas exchange, cardiac output (Q) and rate of perceived exertion (RPE). Total peripheral resistance index (TPRI), stroke volume index (SVI) and rate pressure product (RPP) were calculated. Compared to the pre-administration conditions, 1 week of Losartan treatment significantly reduced (p< .05) resting MAP, SBP and DBP in these subjects. Losartan treatment did not modify submaximal exercise HR, Q, VO2 or RPE. The RPP also was not different between the Lo and Pl trials at rest (p >.05), but was reduced at peak exercise with Lo treatment (p<.05). Losartan significantly reduced calculated TPRI at rest (p< .05) in comparison with Pl (12%) but not during steady-state exercise. Circulating plasma levels of Ang II and PRA were significantly higher with Lo (p<.05). In conclusion, Losartan, a new antihypertensive medication, reduced BP without altering exercise performance in hypertensive women. Losartan is an appropriate first line antihypertensive agent to use in treatment of hypertensive individuals who wish to participate in a regular exercise program. / Ph. D.

In the course of this study, several endogenous compounds and model substances were used to mimic the conditions in patients suffering from hypertension. As endogenous compounds, angiotensin II and aldosterone were chosen. As model substances, 4-nitroquinoline-1-oxide (NQO), hydrogen peroxide and phorbol 12-myristate 13-acetate (PMA) were selected. Benfotiamine as well as &#945;-tocopherol proved in the course of the experiments to be able to prevent angiotensin II-induced formation of oxidative DNA strand breaks and micronuclei. This could be due to a prior inhibition of the release of reactive oxygen species and is in contrast to results which were achieved using thiamine. Furthermore, experiments in which cells were pre-incubated with benfotiamine followed by incubation with NQO showed that benfotiamine was not able to prevent the induction of oxidative stress. The hypothesis that benfotiamine has, like &#945;-tocopherol, direct antioxidative capacity was fortified by measurements in cell free systems. In brief, a new working mechanism for benfotiamine in addition to the ones already known could be provided. In the second part of the study, angiotensin II was shown to be dose-dependently genotoxic. This effect is mediated via the angiotensin II type 1 receptor (AT1R) which. Further experiments were extended from in vitro settings to the isolated perfused kidney. Here it could be shown that angiotensin II caused vasoconstriction and DNA strand breaks. Co-perfusion of kidneys with angiotensin II and candesartan prevented vasoconstriction and formation of strand breaks. DNA strand break formation due to mechanical stress or hypoxia could be ruled out after additional experiments with the thromboxane mimetic U 46619. Detailed investigation of the DNA damage in vitro revealed that angiotensin II induces single strand breaks, double strand breaks and 8-hydroxydeoxyguanosine (8-oxodG)-adducts as well as abasic sites. Investigations of the effects of aldosterone-treatment in kidney cells showed an increase of oxidative stress, DNA strand breaks and micronuclei which could be prevented by the steroidal mineralocorticoid receptor antagonist eplerenone. Additional experiments with the non-steroidal mineralocorticoid receptor antagonist (S)-BR-4628 revealed that this substance was also able to prevent oxidative stress and genomic damage and proved to be more potent than eplerenone. In vivo, hyperaldosteronism was imitated in rats by aid of the deoxycorticosteroneacetate (DOCA) salt model. After this treatment, levels of DNA strand breaks and chromosomal aberrations in the kidney could be observed. Furthermore, an increase in the release of ROS could be measured. Treatment of these animals with spironolactone , BR-4628 and enalaprile revealed that all antagonists were effective BR-4628 was the most potent drug. Finally, rosuvastatin was investigated. In HL-60 cells phorbol 12-myristate 13-acetate caused oxidative stress. Rosuvastatin was able to prevent the release of ROS and subsequent oxidative DNA damage when co-incubated with PMA. Furthermore, not only an inhibition of PMA-induced oxidative stress but also inhibition of the unspecific release of ROS induced by hydrogen peroxide was observable. Addition of farnesyl pyrophosphate (FPP), geranylgeranyl pyrophosphate (GGPP), and mevalonate, intermediates of the cholesterol pathway, caused only a marginal increase of oxidative stress in cells treated simultaneously with PMA and rosuvastatin, thus indicating the effect of rosuvastatin to be HMG-CoA-reductase-independent. Investigation of the gene expression of subunits of NAD(P)H oxidase revealed a down-regulation of p67phox following rosuvastatin-treatment. Furthermore, it could be shown that rosuvastatin treatment alone or in combination with PMA increased total glutathione levels probably due to an induction of the gene expression and enzyme activity of &#947;-glutamylcysteine synthetase (&#947;-GCS). / Im Zuge dieser Studie wurden sowohl endogene Substanzen als auch Modellsubstanzen eingesetzt, um die pathologischen Verhältnisse in Patienten, die an Bluthochdruck leiden, zu imitieren. Als endogene Substanzen wurden Angiotensin II und Aldosteron ausgewählt. Als Modellsubstanzen wurden 4-Nitrochinolin-1-oxid (NQO), Wasserstoffperoxid und Phorbol-12-myristat-13-gewählt. Der erste Teil dieser Arbeit beschäftigt sich mit zwei Vitaminen, nämlich Benfotiamin und &#945;-Tocopherol. Sowohl Benfotiamin als auch &#945;-Tocopherol zeigten im Laufe der Experimente, dass sie in der Lage sind, durch Angiotensin II verursachte DNA-Strangbrüche und chromosomale Aberrationen zu verhindern. Dies ist möglicherweise auf eine ebenfalls beobachtbare vorausgegangene Inhibition der Freisetzung reaktiver Sauerstoffspezies zurückzuführen. Zusammenfassend konnte ein neuer Wirkmechanismus für Benfotiamin vorgestellt werden. Im zweiten Teil dieser Studie konnte nachgewiesen werden, dass Angiotensin II eine dosisabhängige Gentoxizität verursacht. Dieser Effekt wird durch den Angiotensin II-Rezeptor Typ 1 vermittelt. Im weiteren Verlauf der Studie wurden die in vitro Experimente auf das Modell der isolierten perfundierten Mäuseniere ausgeweitet. Hier konnte gezeigt werden, dass Angiotensin II Vasokonstriktion und DNA-Strangbrüche verursacht. Co-Perfusion der Nieren mit Angiotensin II und Candesartan verhinderte hingegen die Vasokonstriktion und die Bildung von DNA-Strangbrüchen. Die Verursachung von Strangbrüchen durch mechanischen Stress oder Hypoxie konnte ausgeschlossen werden. Die Untersuchung der ex vivo beobachteten DNA-Schäden in vitro ließ erkennen, dass Angiotensin II Einzelstrangbrüche, Doppelstrangbrüche, die Bildung des DNA-Addukts 8-OxodG und abasische Stellen induziert. Ein Reparatur-Comet Assay, parallel durchgeführt mit der Messung des phosphorylierten Histons 2AX (&#947;-H2AX) über 24 h, zeigte eine vollständige Reparatur der Einzelstrangbrüche, wohingegen die Zahl der Doppelstrangbrüche in diesem Zeitraum sogar zunahm. Untersuchungen der Effekte, die eine Aldosteron-Behandlung auf Nierenzellen hat, zeigten einen Anstieg des oxidativen Stress, der DNA Strangbrüche und der Mikrokerne. Diese Effekte konnten durch Eplerenon verhindert werden. Weitere Experimente mit dem nicht-steroidalen Mineralocorticoid Rezeptor-Antagonisten (S)-BR-4628 zeigten, dass auch diese Substanz oxidativen Stress und DNA Schäden verhindern konnte, im Gegensatz hierzu hatte das (R)-Isomer, das keine Aktivität am Mineralocorticoid Rezeptor zeigt, keine präventiven Effekte. In vivo wurde der Hyperaldosteronismus mit Hilfe des Deoxycorticosteronacetat- (DOCA) Salzmodells nachgeahmt. Unter dieser Behandlung konnten Level an DNA-Strangbrüchen und chromosomalen Aberrationen beobachtet werden. Des Weiteren konnten in den DOCA-Tieren erhöhte Level an oxidativem Stress gemessen werden. Wurden die Versuchstiere zusätzlich zur DOCA-Behandlung mit Spironolacton, BR-4628 und dem Enalapril behandelt, konnte gezeigt werden, dass BR-4628 potenter war als Spironolacton Enalapril. Zuletzt wurde mit Rosuvastatin eine Substanz untersucht, die die antioxidative Abwehr der Zellen aktivieren kann. In der humanen Leukämie-Zelllinie HL-60 verursachte Phorbol-12-myristat-13-acetat (PMA) oxidativen Stress. Rosuvastatin war in der Lage, die Freisetzung von ROS und daraus resultierende DNA-Strangbrüche bei Co-Inkubation mit PMA zu verhindern. Außerdem konnte gezeigt werden, dass Rosuvastatin nicht nur PMA-induzierten oxidativen Stress, sondern auch die unspezifische Wasserstoffperoxid-induzierte Freisetzung von ROS verhinderte. Die Untersuchung der Genexpression von Untereinheiten der NAD(P)H Oxidase ergab, dass p67phox nach Rosuvastatin-Behandlung herabreguliert wurde. Behandlung mit Rosuvastatin allein oder zusammen mit PMA konnte außerdem die Glutathion-Spiegel erhöhen. Dies ist vermutlich auf die Induktion der Genexpression und der Enzymaktivität der &#947;-Glutamylcystein-Synthetase (&#947;-GCS), des Schrittmacherenzyms des Glutathionsystems, zurückzuführen.

Die Entstehung atherosklerotischer Gefässwandveränderungen ist ein chronischer Prozess, welcher durch vaskuläre pro-atherosklerotische Wachstumsfaktoren und anti-atherosklerotisch wirkende Modulatoren in der Gefässwand reguliert wird. Angiotensin II ist sowohl in in-vitro als auch in in-vivo Studien als ein zentraler pro-atherosklerotischer Wachstumsfaktor identifiziert worden. Pro-atherosklerotische in-vitro Effekte von Angiotensin II resultieren in einer massiven Zunahme atherosklerotischer Läsionen durch Angiotensin II im Tiermodell. Die pro-atherosklerotische Funktion von Angiotensin II wird parallel durch klinische Daten unterstützt, in denen gezeigt wird, dass die Angiotensin II Blockade durch ACE-Inhibitoren oder AT1-R Antagonisten zu einer Reduktion atherosklerotischer Gefässwandveränderungen führt, was letztendlich in einer Senkung der kardiovaskulären Mortalität resultiert. Demgegenüber stehen multiple atherosklerose-hemmende Faktoren in der Gefässwand. In der Gruppe dieser Faktoren spielen die nukleären Hormonrezeptoren der Peroxisome Proliferator-Activated Receptors (PPARs) eine wichtige pathophysiologische und klinische Rolle. Aktiviert durch endogene Liganden supprimieren PPARs pro-atherosklerotische Abläufe in der Gefässwand. Eine zusätzliche Aktivierung wird durch exogene, synthetische Liganden erreicht, welche in der Therapie metabolischer Erkrankungen bereits klinisch eingesetzt werden. Zwei PPAR Isoformen sind in der Gefässwand überwiegend exprimiert, PPARa und PPARg. Beide Isoformen vermitteln anti-atherosklerotische Effekte durch die transkriptionelle Regulation unterschiedlicher Zielgene. Wir untersuchten die PPAR-vermittelte Regulation dieser Zielgene und konnten neue molekulare Wirkmechanismen, welche den anti-atherosklerotischen Wirkungen von PPARa- und PPARg-Liganden zugrunde liegen identifizieren. In weiterführenden Arbeiten konnten wir zeigen, dass multiple bidirektionale Interaktionen zwischen Angiotensin II und PPARs bestehen, welche einen wesentlichen Einfluss auf die Entstehung und Progression atherosklerotischer Gefässveränderungen haben. / The development of atherosclerosis is a chronic process, which is tightly regulated by pro-atherosclerotic vascular growth factors and anti-atherosclerotic modulators in the vessel wall. Angiotensin II has been identified in in-vitro and in-vivo studies as a central pro-atherosclerotic growth factor. Angiotensin II-infusion in mice results in a profound induction of atherosclerotic lesions in the aorta. In parallel, clinical studies have demonstrated that blockade of the renin-angiotensin-system by AT1R-antagonists or ACE-inhibitors significantly lowers the incidence of atherosclerotic diseases. The pro-atherosclerotic actions of angiotensin II are counterbalanced by multiple anti-atherosclerotic modulators present in the vessel wall. The peroxisome proliferators-activated receptors (PPARs), including the isoforms alpha and gamma, are ligand-activated nuclear hormone receptors, present in the vascular wall, and exerting major anti-atherosclerotic actions. They function as transcriptional regulators of target genes involved in atherogenesis. We investigated PPAR-mediated target gene expression in vascular and inflammatory cells, and identified new anti-atherosclerotic mechanisms of these receptors. Finally we characterized bidirectional interactions between angiotensin II and PPARs, which might play a major role in the development of atherosclerosis in humans.

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.

No description available.

The locus coeruleus (LC) of the rat was lesioned by microinjection of selective neurotoxins into the brainstem. 6‐Hydroxydopamine (6‐OHDA), 3 μg/μl, given unilaterally at two sites 0.6 mm apart on the rostro‐caudal axis of the LC, was used to lesion catecholamine‐containing neuronal elements. Ibotenic acid, 2.5 μg/0.5 μl, administered similarly was used to lesion nerve cell bodies. Two weeks after administration of the neurotoxin, lesion efficacy was determined based on the norepinephrine content of the cerebral cortex ipsi‐ and contralateral to the lesion. 6‐OHDA lesions of the LC caused a 46% reduction in ipsilateral cortical norepinephrine and a 60% reduction in specific 125I‐[Sar1, Ile8]‐angiotensin II (125I‐SIAII) binding in the LC. Ibotenic acid lesions of the LC caused a 73% reduction in ipsilateral cortical norepinephrine and a 81% reduction in specific 125I‐SIAII binding in the LC. These results indicate that All receptor binding sites in the LC are localized on noradrenergic nerve cell bodies or their dendritic and axonal ramifications within the LC.

Previous studies have used new angiotensin II (AII) receptor subtype selective compounds to localize AII receptor subtypes within discrete rat brain nuclei. The purpose of this autoradiographic study was to extend these preliminary findings and provide a comprehensive analysis of AII binding sites in 22 rat brain nuclei and the anterior pituitary, to include estimates of the binding affinity for 125I sar1 ile8 AII (125I SIAII) at each nucleus, and determine the fractional distribution of each subtype at each nucleus. Estimates of K(D), in separate experiments revealed that AT1 nuclei had a consistently higher affinity for 125I SIAII than AT2 nuclei (0.66 vs. 2.55 nM). Displacement of subsaturating concentrations of 125I SIAII by 10-8-10-4 M DuP753 (selective for the AT1 subtype) or PD123177 (selective for the AT2 subtype) indicated that approximately half of the brain regions surveyed contained predominantly AT1 sites and half contained predominantly AT2 sites. Binding was partially displaced by both compounds in several regions and two site analyses were performed to estimate the distribution of subtypes within each nucleus. The data were then corrected for differential occupancy by 125I SIAII. Brain nuclei associated with cardiovascular or dipsogenic actions of AII, e.g., subfornical organ, organum vasculosum of the lamina terminalis, median preoptic nucleus, nucleus of the solitary tract and area postrema, contained pure, or almost pure, populations of AT1 receptors. The functions of AII in brain regions containing predominantly AT2 binding sites, e.g., thalamus, colliculi, inferior olive and locus ceruleus, remain undefined. Thus, AII binding sites in the rat brain have been differentiated into two subtypes with similar characteristics to those reported in peripheral tissues. However, the unexpected finding that they can be differentiated on the basis of their affinity for 125I SIAII raises questions concerning their coidentity with peripheral receptor subtypes.

Prolylcarboxypeptidase (PRCP, EC3.4.16.2) ist ein ubiquitär exprimiertes Enzym, mit höchster Expression im Maushirn. Es spaltet spezifisch die letzte carboxyterminale Aminosäure von Substraten, deren vorletzte Aminosäure ein Prolin ist. Seine bisher publizierten Substrate Angiotensin II (AngII) und alpha Melanocortin Stimulierendes Hormone (alphaMSH) legen eine Rolle von PRCP in der Entwicklung von kardiovaskulären und metabolischen Krankheiten nahe. Um die in vivo Funktion von PRCP zu studieren, wurde eine Knockout Maus generiert (PRCP-/-). Metabolischer Phänotyp: PRCP-/- Mäuse zeigten generell ein reduziertes Körpergewicht, selbst wenn sie über Monate mit einer Hochfettdiät versorgt wurden. Erhöhte Plasmaleptin Werte und Proopiomelanocortin (pomc) Expression in knockout Hypothalami wiesen auf eine wichtige Rolle von PRCP in der Regulation von Futteraufnahme und Energiehomöostase hin. Eines der Genprodukte von pomc ist alphaMSH, welches im Hypothalamus die Futteraufnahme terminiert. Die carboxyterminale Struktur dieses Neuropeptids erfüllt alle Voraussetzungen, um von PRCP gespalten zu werden. Zudem konnte prcp Promotoraktivität in den selben Hirnstrukturen gezeigt werden, in denen auch alphaMSH-Wirkung beschrieben wurde. Eine mögliche Funktion von PRCP wäre somit die Inaktivierung des Appetitzüglers alphaMSH im Hypothalamus. Kardiovasculärer Phänotyp: Zunächst erwiesen sich zirkulierende Ang-Peptide in PRCP-/- Mäusen als unverändert. Jedoch konnte ein erhöhtes Niveau des Degradationsproduktes Ang1-7 in der Niere gezeigt werden. Die Entdeckung einer erhöhten Enzymaktivität von Angiotensin Converting Enzyme 2 (ACE2) in PRCP-/- Nieren, wurde als Kompensation der fehlenden PRCP Aktivität in PRCP-/- Nieren interpretiert. bot einen Erklärungsansatz für dieses Ergebnisse. Es wird davon ausgegangen, daß ACE2 die fehlende PRCP Aktivität in knockout Mäusen kompensiert. Das es sich hierbei um eine lokale begrenzte Kompensation handeln muß, zeigten der erhöhte Blutdruck und Herzrate, sowie die milde Herzhypertrophie. Da spezifische prcp Promotoraktivität in Hirnnuclei gefunden wurde, die in die Kontrolle der Herzfrequenz und des Blutdrucks involviert sind, wird eine regulatorische Funktion von Hirnstamm-PRCP auf Herzrhytmus und Blutdruck vermutet. / Prolylcarboxypeptidase (PRCP, EC3.4.16.2) is an enzyme specifically cleaving the last carboxy-terminal amino acid from substrates containing a penultimate proline. Its known potential substrates are linked to cardiovascular and metabolic phenomenon. To analyse the in vivo function of this enzyme a PRCP knockout mouse was generated. Homozygous knockout mice are viable but show tendency of decreased life span. In mice prcp expression is present in all tissues tested with very specific localizations of prcp promotor activity to distinct brain areas within the cortex, hippocampus, hypothalamus and the brain stem. The metabolic phenotype of PRCP deficient mice is characterized by low body weight even when feeding the animals a high fat diet. The increased plasma leptin levels and elevated expression of proopiomelanocortin gene (pomc) found in knockout hypothalami suggests an involvement of PRCP in the regulation of food intake and energy homeostasis. One of the gene products of pomc is alpha-melanocortin stimulating hormone (alphaMSH) that is terminating feeding when released from hypothalamic POMC neurons. Its carboxy-terminal structure is fitting the cleavage preferences of PRCP. Prcp promotor activities are localized in arcuate nucleus and paraventricular nucleus, brain areas of known alphaMSH signalling, supporting a role of PRCP in the degradation of central alphaMSH. The impact of PRCP on angiotensin II (AngII) metabolism was studied by determining the level of AngII and its degradation product Ang1-7 in blood and tissues. But instead of increased AngII levels due to the missing degradation enzyme in knockout mice, Ang1-7 levels were increased in kidney. These results were explainable by the increased activity of angiotensin converting enzyme 2 (ACE2) found in kidney. Probably ACE2 is compensating the lack of PRCP in the knockout mouse. Nevertheless, blood pressure and heart rate of PRCP knockout mice was increased. The mild hypertension was accompanied by mild hypertrophy of the hearts. Prcp promotor activity was found in brain stem an area important for regulation of blood pressure and heart rate suggesting that central PRCP regulates blood pressure.

In der vorliegenden Arbeit wurde der Einfluss der humanen AT2-Rezeptorexpression und -stimulation auf den Collagen I alpha 2-mRNA-Gehalt und die Migration von porcinen kardialen Fibroblasten untersucht, um die Frage zu klären, ob AT2-Rezeptoren in kultivierten kardialen Fibroblasten AT1-antagonistische antifibrotische und migrationshemmende Effekte auf den Collagen I alpha 2-mRNA-Gehalt bzw. die Migration ausüben. Um die Funktion der AT2-Rezeptoren in der Zellkultur untersuchen zu können, wurde die AT2-cDNA durch adenovirale Transduktion in die Fibroblasten übertragen und so der AT2-Rezeptor überexprimiert. Mittels RT-PCR wurden die relativen Änderungen im Collagen I alpha 2-mRNA-Gehalt in TGF-beta1- bzw. TGF-beta1 plus Ang II-stimulierten Fibroblasten im Vergleich zur unstimulierten Kontrolle bestimmt. Alle Werte wurden auf ein Referenzgen (beta-Actin) bezogen. Die AT2-Stimulation änderte den relativen Collagen I alpha 2-mRNA-Gehalt der Fibroblasten nicht signifikant gegenüber den Antisense-(Ad5TA2)-transduzierten Fibroblasten. In der modifizierten Boyden-Kammer wurde der AT2-vermittelte Effekt von Ang II, hPDGF-BB sowie der Kombination beider Stoffe auf die Migration untersucht. Die alleinige Stimulation von AT2-Rezeptoren mit Ang II verhinderte die Migration gegenüber nichttransduzierten Fibroblasten. In Kombination mit hPDGF-BB änderte Ang II die Migration in AT2-überexprimierenden Fibroblasten nicht gegenüber den Antisense-(Ad5TA2)-transduzierten Fibroblasten. Bei ausschließlicher Stimulation durch hPDGF-BB wurde aber in AT2-exprimierenden Fibroblasten eine signifikant geringere Migration als in Antisense-(Ad5TA2)-transduzierten Fibroblasten festgestellt. Die zugrunde liegende Hypothese, dass AT2-Expression und Stimulation den relativen Collagen I alpha 2-mRNA-Gehalt hemmt, konnte in den vorliegenden Experimenten nicht bestätigt werden. Dies ließ keine inhibitorische AT2-vermittelte Wirkung von Ang II im Bezug auf den TGF-beta1-induzierten Collagen I alpha 2-mRNA-Gehalt erkennen. Dagegen führte die Ang II-Stimulation überexprimierter AT2-Rezeptoren zu einer verringerten Migration und vermittelte so einen AT1-antagonistischen Effekt. / In this work the influence of expression and stimulation of the human AT2 receptor on Collagen I alpha 2-mRNA-content and migration of porcine cardiac fibroblastst was tested to clarify the question if AT2 receptors promote AT1 antagonistic antifibrotic and antimigratory effects on collagen I alpha 2-mRNA content and migration. To examine the AT2 receptor function in the cell culture AT2 cDNA was transferred into fibroblasts by adenoviral transduction and the AT2 receptor was overexpressed. Through the use of RT-PCR the relative changes in collagen I alpha 2-mRNA content in TGF-beta1 stimulated and TGF-beta1 plus Ang II stimulated fibroblasts were assayed and compared to the unstimulated control. All values were referred to a reference gene (beta-actin). Stimulation of AT2 receptors did not change the relative collagen I alpha 2-mRNA content of the fibroblasts significantly compared to antisense-(Ad5TA2) transduced fibroblasts. In the modified Boyden-chamber the AT2 mediated effect of Ang II, hPDGF-BB and the combination of both on migration was assessed. The stimulation of AT2 receptors with Ang II inhibited migration compared to nontransduced fibroblasts. In combination with hPDGF-BB Ang II did not change the migration in AT2 overexpressing fibroblasts compared to antisense-(Ad5TA2)-transduced fibroblasts. In the case of exclusive stimulation of AT2-expressing fibroblasts with hPDGF-BB a significantly lower migration was found compared to antisense-(Ad5TA2)-transduced fibroblasts. The underlying therory that AT2 expression and stimulation inhibits the relative collagen I alpha 2-mRNA content could not be confirmed in this work. This did not reveal an inhibitory AT2 mediated effect of Ang II in respect to the TGF-beta1 induced collagen I alpha 2-mRNA content. In contrast to that Ang II stimulation of overexpressed AT2 receptors led to a decreased migration and mediated an AT1 antagonistic effect.