The renin angiotensin aldosterone system is the dominant hormonal regulatory system controlling sodium balance and therefore blood pres- sure homeostasis. Abnormal modulation of this system is implicated in the pathogenesis of hypertension and end organ injury. We have previously developed the Cyp1a1-Ren2 transgenic rat to model an- giotensin II (ANG II) dependent hypertension. In this model hyper- tension causes renal injury, predominantly in the preglomerular vas- culature. The susceptibility to renal injury has a genetic component. A consomic/congenic study identified angiotensin converting enzyme (Ace) as an important modifer. However, renal injury is unlikely to be in uenced by a single gene. In this thesis it was hypothesised that examination of a renal microar- ray to compare the relative expression in F344 (susceptible) and Lewis (relatively protected) strains would reveal further genetic factors me- diating renal injury susceptibility. Genome wide expression analysis confirmed that Ace was a key modifier gene. Furthermore, the puriner- gic receptors P2x7 and P2x4 were identified as additional candidates. Gene and protein expression of these P2X receptors were both higher in F344 compared with Lewis. Immunohistochemistry localised P2X7 and P2X4 to the renal vasculature and tubules: the expression pattern was similar in both strains but became distinct in the renal medulla. F344, but not Lewis, responded to acute antagonism of P2X7 and P2X4. F344 showed a significant drop in blood pressure but maintained renal blood ow, indicative of tonic renal vasoconstriction. When ANG II was infused into F344 rats, there was a modest increase in blood pressure and an impairment of the pressure-natriuresis mecha- nism but no overt injury. Blood oxygenation-level dependent magnetic resonance imaging of the kidney identified a decrease in renal R2* sig- nal following P2X7 and P2X4 antagonism in ANG II infused F344 rats. P2X7/4 receptor activation reduces oxygenation and suppresses pressure-natriuresis. These effects are pro-biotic and may underpin susceptibility to renal injury.

京都大学 / 新制・課程博士 / 博士(医学) / 甲第24084号 / 医博第4860号 / 新制||医||1059(附属図書館) / 京都大学大学院医学研究科医学専攻 / (主査)教授 柳田 素子, 教授 長船 健二, 教授 黒田 知宏 / 学位規則第4条第1項該当 / Doctor of Medical Science / Kyoto University / DFAM

Elevated circulating Ang II activates neurons in the forebrain cardiovascular regulatory areas to cause sympatho-excitation and hypertension. We hypothesized that circulating Ang II causes neuronal activation in the SFO and thereby activates efferent pathways to the PVN, and chronically causes activation of aldosterone production in magnocellular neurons in PVN and SON, which amplifies neuronal activation in the PVN and central sympatho-excitatory pathways. The aim of the present study was to determine the pattern of neuronal activation in forebrain nuclei by circulating Ang II and to elucidate where in the hypothalamus Ang II may stimulate aldosterone biosynthesis. Dose related effects of circulating Ang II on BP were first assessed. Wistar rats instrumented with telemetry probes were infused subcutaneously with Ang II 150 and 500 ng/kg/min for 14 days. The subcutaneous infusion of Ang II at 150 ng/kg/min increased blood pressure gradually up to 20 mmHg and at 500 ng/kg/min up to 60 mmHg. Ang II at 500 ng/kg/min increased plasma Ang II by 4-fold. To assess effects of circulating Ang II on CNS pathways, Wistar rats were implanted subcutaneously with minipumps infusing 150 and 500 ng/kg/min Ang II for 1, 4 and 14 days. Three patterns of neuronal activation were observed by sc infusion of Ang II. The SFO was activated during the first day and remained activated for 4 days, but at 14 days showed diminished activation. MnPO did not show significant activation during the first day but, after several days the activation was high and then less by 14 days. Parvocellular PVN (pPVN), magnocellular PVN (mPVN) and SON showed an initial activation that increased over time. Chronic intracerebroventricular infusion of an aldosterone synthase inhibitor or a mineralocorticoid receptor (MR) blocker attenuated the increase in Fra expression in PVN but not SON, and prevented the decrease in SFO after 14 days infusion of Ang II. A significant increase in mRNA expression of steroidogenic acute regulatory protein (StAR), a rate limiting enzyme in aldosterone production was found in glia cells of PVN and SFO assessed by rt-PCR after 3 days subcutaneous infusion of Ang II at 500 ng/kg/min. Total expression of aldosterone synthase (CYP11B2) mRNA was increased in SFO, MnPO, SON and PVN after 3 days of infusion of Ang II. After 14 days no significant changes were observed in the expression of StAR or CYP11B2 mRNA. In comparison, in adrenal StAR mRNA expression increased after 3 days but no longer after 14 days. In contrast, CYP11B2 mRNA expression in adrenal increased after both 3 and 14 days of infusion. These findings may support our hypothesis that chronic elevation of circulating Ang II increases neuronal activity in CVOs, presumably leading to activation of the PVN and SON to induce an increase in aldosterone production in magnocelular PVN and SON. In the second phase activation of CVOs appears to diminish, but an aldosterone-dependent amplifying mechanisms, causes sustained activation of the PVN and thereby hypertension.

Elevated circulating Ang II activates neurons in the forebrain cardiovascular regulatory areas to cause sympatho-excitation and hypertension. We hypothesized that circulating Ang II causes neuronal activation in the SFO and thereby activates efferent pathways to the PVN, and chronically causes activation of aldosterone production in magnocellular neurons in PVN and SON, which amplifies neuronal activation in the PVN and central sympatho-excitatory pathways. The aim of the present study was to determine the pattern of neuronal activation in forebrain nuclei by circulating Ang II and to elucidate where in the hypothalamus Ang II may stimulate aldosterone biosynthesis. Dose related effects of circulating Ang II on BP were first assessed. Wistar rats instrumented with telemetry probes were infused subcutaneously with Ang II 150 and 500 ng/kg/min for 14 days. The subcutaneous infusion of Ang II at 150 ng/kg/min increased blood pressure gradually up to 20 mmHg and at 500 ng/kg/min up to 60 mmHg. Ang II at 500 ng/kg/min increased plasma Ang II by 4-fold. To assess effects of circulating Ang II on CNS pathways, Wistar rats were implanted subcutaneously with minipumps infusing 150 and 500 ng/kg/min Ang II for 1, 4 and 14 days. Three patterns of neuronal activation were observed by sc infusion of Ang II. The SFO was activated during the first day and remained activated for 4 days, but at 14 days showed diminished activation. MnPO did not show significant activation during the first day but, after several days the activation was high and then less by 14 days. Parvocellular PVN (pPVN), magnocellular PVN (mPVN) and SON showed an initial activation that increased over time. Chronic intracerebroventricular infusion of an aldosterone synthase inhibitor or a mineralocorticoid receptor (MR) blocker attenuated the increase in Fra expression in PVN but not SON, and prevented the decrease in SFO after 14 days infusion of Ang II. A significant increase in mRNA expression of steroidogenic acute regulatory protein (StAR), a rate limiting enzyme in aldosterone production was found in glia cells of PVN and SFO assessed by rt-PCR after 3 days subcutaneous infusion of Ang II at 500 ng/kg/min. Total expression of aldosterone synthase (CYP11B2) mRNA was increased in SFO, MnPO, SON and PVN after 3 days of infusion of Ang II. After 14 days no significant changes were observed in the expression of StAR or CYP11B2 mRNA. In comparison, in adrenal StAR mRNA expression increased after 3 days but no longer after 14 days. In contrast, CYP11B2 mRNA expression in adrenal increased after both 3 and 14 days of infusion. These findings may support our hypothesis that chronic elevation of circulating Ang II increases neuronal activity in CVOs, presumably leading to activation of the PVN and SON to induce an increase in aldosterone production in magnocelular PVN and SON. In the second phase activation of CVOs appears to diminish, but an aldosterone-dependent amplifying mechanisms, causes sustained activation of the PVN and thereby hypertension.

Thesis (Ph. D.)--University of Washington, 2001. / Vita. Includes bibliographical references (leaves 84-100).

Elevated circulating Ang II activates neurons in the forebrain cardiovascular regulatory areas to cause sympatho-excitation and hypertension. We hypothesized that circulating Ang II causes neuronal activation in the SFO and thereby activates efferent pathways to the PVN, and chronically causes activation of aldosterone production in magnocellular neurons in PVN and SON, which amplifies neuronal activation in the PVN and central sympatho-excitatory pathways. The aim of the present study was to determine the pattern of neuronal activation in forebrain nuclei by circulating Ang II and to elucidate where in the hypothalamus Ang II may stimulate aldosterone biosynthesis. Dose related effects of circulating Ang II on BP were first assessed. Wistar rats instrumented with telemetry probes were infused subcutaneously with Ang II 150 and 500 ng/kg/min for 14 days. The subcutaneous infusion of Ang II at 150 ng/kg/min increased blood pressure gradually up to 20 mmHg and at 500 ng/kg/min up to 60 mmHg. Ang II at 500 ng/kg/min increased plasma Ang II by 4-fold. To assess effects of circulating Ang II on CNS pathways, Wistar rats were implanted subcutaneously with minipumps infusing 150 and 500 ng/kg/min Ang II for 1, 4 and 14 days. Three patterns of neuronal activation were observed by sc infusion of Ang II. The SFO was activated during the first day and remained activated for 4 days, but at 14 days showed diminished activation. MnPO did not show significant activation during the first day but, after several days the activation was high and then less by 14 days. Parvocellular PVN (pPVN), magnocellular PVN (mPVN) and SON showed an initial activation that increased over time. Chronic intracerebroventricular infusion of an aldosterone synthase inhibitor or a mineralocorticoid receptor (MR) blocker attenuated the increase in Fra expression in PVN but not SON, and prevented the decrease in SFO after 14 days infusion of Ang II. A significant increase in mRNA expression of steroidogenic acute regulatory protein (StAR), a rate limiting enzyme in aldosterone production was found in glia cells of PVN and SFO assessed by rt-PCR after 3 days subcutaneous infusion of Ang II at 500 ng/kg/min. Total expression of aldosterone synthase (CYP11B2) mRNA was increased in SFO, MnPO, SON and PVN after 3 days of infusion of Ang II. After 14 days no significant changes were observed in the expression of StAR or CYP11B2 mRNA. In comparison, in adrenal StAR mRNA expression increased after 3 days but no longer after 14 days. In contrast, CYP11B2 mRNA expression in adrenal increased after both 3 and 14 days of infusion. These findings may support our hypothesis that chronic elevation of circulating Ang II increases neuronal activity in CVOs, presumably leading to activation of the PVN and SON to induce an increase in aldosterone production in magnocelular PVN and SON. In the second phase activation of CVOs appears to diminish, but an aldosterone-dependent amplifying mechanisms, causes sustained activation of the PVN and thereby hypertension.

Diss. (sammanfattning) Uppsala : Univ., 2003. / Härtill 4 uppsatser.

Thesis (M.S.)--Youngstown State University, 1995. / Includes bibliographical references (leaves 69-71).

Orientador: Heitor Moreno Junior / Dissertação (mestrado) - Universidade Estadual de Campinas, Faculdade de Ciencias Medicas / Made available in DSpace on 2018-08-12T13:57:46Z (GMT). No. of bitstreams: 1 Lacchini_Riccardo_M.pdf: 3791091 bytes, checksum: 8936fbea9ef58bfd48f550316c33f122 (MD5) Previous issue date: 2008 / Resumo: Introdução: Hipertensão Refratária se caracteriza por quadro de hipertensão concomitante ao uso de 3 classes diferentes de anti-hipertensivos, sendo, pelo menos um destes, diurético. Existem fatores genéticos que influenciam mecanismos de controle da pressão arterial, podendo ter papel importante na hipertensão refratária. O sistema Renina-Angiotensina-Aldosterona desempenha um papel central na gênese da hipertensão. Isto torna interessante o estudo das influências genéticas sobre sistemas envolvidos com a produção e transporte da aldosterona. Este trabalho apresenta o estudo de um polimorfismo na região promotora do gene CYP11b2 (troca de timina por citosina na posição -344 do gene), que codifica a aldosterona sintase, enzima responsável pelo último passo na síntese de aldosterona pelas glândulas supra-renais. Secundariamente, o trabalho apresenta o estudo de um polimorfismo no exon 26 do gene da p-glicoproteína (troca de citosina por timina na posição 3435 do gene). Esta proteína está envolvida com resistência a diversos medicamentos na terapêutica (o gene é chamado Muliple Drug Resistance-1, MDR-1), e recentemente foi associada a alterações na excreção e captação tecidual de aldosterona. Objetivos: O estudo visa determinar se existe associação entre o alelo T do polimorfismo -344C/T no gene CYP11b2 e a resistência ao tratamento anti-hipertensivo. Secundariamente, é avaliada, de maneira preliminar, a participação do alelo C do gene MDR-1 no fenótipo de hipertensão refratária. Materiais e Métodos: Foram avaliados 340 indivíduos, dos quais 88 eram portadores de HAR que preencheram os requisitos de inclusão no estudo, 142 indivíduos eram hipertensos moderados e responsivos a tratamento farmacológico (HT). Os demais indivíduos (n = 110) constituíram o grupo controle. Foram coletadas amostras de sangue para exames laboratoriais (dentre eles, quantificação de aldosterona, cortisol, renina) e extração de DNA. Foi medida a espessura da íntima média de carótida (EIM) dos pacientes através de ultra-som. Foi estimada a Velocidade de Onda de Pulso carótida-femural (VOP) dos pacientes. O DNA foi extraído pelo método de Salting Out. O polimorfismo do gene CYP11b2 foi genotipado por PCR(Polymerase Chain Reaction) seguido por digestão enzimática com a enzima Hae-III. O polimorfismo do gene MDR-1 foigenotipado através de PCR alelo específico. Resultados: Foi encontrada associação do alelo T do gene CYP11b2 com Hipertensão (p<0,05), porém não houve diferença entre os grupos hipertensos responsivos e hipertensos resistentes. Não houve diferença estatística nas concentrações plasmáticas de aldosterona, cortisol e renina; EIM e VOP nos grupos de alelos C e T. Não foi encontrada relação entre a distribuição alélica do gene MDR-1 e hipertensão tratada ou resistente. Foi detectada associação significativa entre alelo C do gene MDR-1 e pressão arterial diastólica (PAD) aumentada em hipertensos refratários. Conclusões: há associação entre o alelo T do gene CYP11b2 e hipertensão, mas não há associação com resistência farmacológica aos anti-hipertensivos. Não houve influência do alelo T sobre parâmetros clínicos e de remodelamento vascular. O alelo C do gene MDR-1 parece estar associado a uma maior PAD no grupo de hipertensos refratários. / Abstract: Introduction: Resistant hypertension is characterized by elevated blood pressure concurrent with the use of 3 different anti-hypertensive drugs, being one of them a diuretic. There are genetic factors that interfere in the arterial pressure control system and may have a significant role in resistant hypertension. Renin-Angiotensin-Aldosterone System plays a major role in hypertension genesis, therefore the systems that control the production and transport of aldosterone are interesting targets for genetic association studies. This work presents the study of a polymorphism (-344 C/T) in CYP11b2 gene. This gene translates aldosterone synthase, an enzyme responsible for endogenous synthesis of aldosterone on adrenal glands. A secondary study on a polymorphism (3435C/T) in MDR-1 gene was performed. This gene translates p-glicoprotein, an efflux-pump related to several drug resistance phenotypes. Recently it was reported that tissue uptake and excretion of aldosterone is mediated by p-glicoprotein. Objectives: The study aims to evaluate the association between allele T of CYP11b2 and resistance to anti-hypertension treatment. As a second objective it has been performed an initial evaluation of the influence of C allele of MDR-1 on resistant hypertension phenotype. Materials and Methods: In this study, 340 subjects were evaluated: 88 patients presented Refractory Arterial Hypertension (RAH), 142 were responsive to anti-hypertension treatment (RT), and the remaining was the control group (n=110). Blood samples were collected for laboratory examinations (plasmatic aldosterone, cortisol, renin) and DNA extraction. It was examined the carotid intima-media thickness (IMT) using ultra-sound, and carotid-femoral pulse wave velocity (PWV). DNA was extracted using the salting out method. CYP11b2 genotyping was performed through restriction fragment length polymorphism (polymerase chain reaction (PCR) followed by endonuclease digestion). PCR products were digested with Hae III enzyme. MDR-1 genotyping was performed through allele specific PCR. Results: It was found association between allele T of CYP11b2 and hypertension (p<0,005), but no difference has been detected between RT and RAH groups. No statistical difference was detected in plasmatic aldosterone, cortisol and renin concentrations, neither in PWV nor in IMT when compared C and T allele groups. It was found no relationship between alleleC of MDR-1 and hypertension, treated or resistant, although there was a significant association between allele C of this gene and elevated diastolic blood pressure in the RAH group. Conclusions: There is association between allele T of CYP11b2 gene and hypertension, but there is no relationship with pharmacological resistance to antihypertensive treatment. No influence was detected of allele T regarding clinical parameters or remodeling indicators. Allele C of MDR-1 gene seems to be associated with an elevated diastolic blood pressure in resistant hypertensive subjects. / Mestrado / Mestre em Farmacologia

Orientador: Heitor Moreno Junior / Tese (doutorado) - Universidade Estadual de Campinas, Faculdade de Ciencias Medicas / Made available in DSpace on 2018-08-13T00:33:13Z (GMT). No. of bitstreams: 1 Girioli_SamiraUbaid_D.pdf: 2811337 bytes, checksum: bc9374b0216cba3ee57f1b107392c0e0 (MD5) Previous issue date: 2009 / Resumo: Segundo o VII JNC e V Diretrizes Brasileiras de Hipertensão Arterial, a hipertensão arterial resistente (HAR) é definida como sendo a elevação persistente dos níveis pressóricos (= 140/90 mmHg) a despeito de tratamento farmacológico tríplice, pleno, incluindo diuréticos, em pacientes com boa adesão e sem causas secundárias de hipertensão arterial ou pseudo-hipertensão. O excesso ou escape de aldosterona pode ocorrer após tratamento crônico com IECA, BRA ou diuréticos tiazídicos, comumente utilizados nestes pacientes ou por hiperaldosteronismo primário não diagnosticado. A espironolactona tem sido apontada como fármaco indispensável no tratamento de HAR por melhorar os níveis pressóricos, mas pouco se sabe sobre a pressão arterial (PA), função endotelial e remodelamento cardiovascular avaliados simultaneamente após sua utilização. O objetivo deste estudo foi avaliar a pressão arterial, a função endotelial, o remodelamento cardiovascular e a função diastólica em hipertensos refratários antes e após a utilização de antagonista de aldosterona, espironolactona, em associação ao tratamento anti-hipertensivo otimizado e individualizado em centro de referência em HAR por 6 meses. Após triagem e adesão rigorosa, 71 pacientes encaminhados ao nosso serviço como hipertensos resistentes foram seqüencialmente incluídos no estudo, após avaliação e caracterização como hipertensos resistentes (HAR, n=39) ou hipertensos controlados (HAC, n=32). O grupo controle foi constituído por pacientes normotensos (CONT, n=37). Anti-hipertensivos eram fornecidos integralmente. Ecocardiograma, Teste de vasodilatação mediada pelo fluxo (VMF), espessura íntima-média de carótidas (IMT), dosagens bioquímicas de concentrações de renina e aldosterona plasmáticas, sódio e potássio séricos e urinários foram realizados na fase inicial (pré-espiro), seguida por associação de espironolactona 25mg/dia ao tratamento anti-hipertensivo por 6 meses (apenas para os grupos HAR e HAC) . Subsequentemente, estes parâmetros foram reavaliados (fase pós-espiro). Após 6 meses de espironolactona, os grupos HAR e HAC apresentaram redução da PA (p<0,001 para PAS e PAD). O grupo HAR apresentou melhora da função endotelial dependente (p<0,001) e independente do endotélio (p=0,007). Os grupos HAR e HAC apresentaram melhora da hipertrofia ventricular esquerda p<0,001) e melhora da função diastólica (índice Kappa HAR: 0,219 e índice Kappa HAC: 0,392) em 7,69% e 15,62%, respectivamente. Os grupos HAR e HAC apresentaram elevação da aldosterona plasmática (p<0,05). A otimização do tratamento da HAR com a associação de espironolactona em pequenas doses, reduz PA, melhora função endotelial, melhora hipertrofia ventricular esquerda e função diastólica a despeito da presença de excesso ou "escape" de aldosterona. / Abstract: According to JOINT VII and V Brazilian Guidelines of Hypertension, resistant hypertension (RH) is defined, as a persistent elevation of blood pressure (= 140/90 mmHg) despite the use of a triple drug regimen at maximal doses including a diuretic, in patients with good compliance and without secondary causes of hypertension or pseudo-hypertension. Aldosterone excess or "escape" can occur after chronic treatment with ACEI, ARB or thiazide-type diuretics, commonly used by these patients or in undiagnosed primary aldosteronism. Spironolactone is thought to be an essential drug in RH treatment because it improves blood pressure, but there are few available data about blood pressure, endothelial function and cardiovascular remodeling evaluated simultaneously with its use. The aim of this study was to assess blood pressure, endothelial function, cardiovascular remodeling and diastolic function in resistant hypertensive patients before and after the use of an aldosterone antagonist, spironolactone, for a six-month period in association with individualized and optimized antihypertensive treatment in a RH reference clinic. After confirming compliance to treatment, 71 hypertensive patients attending our service were included in the study as resistant hypertensive (RH, n = 39) or controlled hypertensive (CH, n = 32) patients. The control group was formed of normotensive patients (CONT, n = 37). All antihypertensive drugs were supplied. Echocardiography, flow-mediated vasodilation, carotid intima-media wall thickness, plasma renin and aldosterone concentrations, plasma and urinary sodium and potassium concentrations were measured at baseline (pre-spironolactone phase), followed by an association of 25 mg/d spironolactone with existing antihypertensive drugs for 6 months (only to RH and CH groups). Subsequently these parameters were reassessed (post-spironolactone phase). After 6 months of spironolactone, the RH and CH groups presented with a reduction in blood pressure (p<0.001 for both systolic and diastolic blood pressures). The RH group had improved endothelial function, both endothelium-dependent (p<0.001) and independent (p = 0.007). RH and CH groups presented with improved left ventricular hypertrophy (p<0.001) and diastolic function (Kappa index RH: 0.219 and Kappa index CH: 0.392) in 7.69% and 15.62%, respectively. There were increases in aldosterone concentrations for RH and CH groups (p< 0.05). Optimized RH treatment with low doses of spironolactone reduces blood pressure and improves endothelial function, left ventricular hypertrophy and diastolic function despite of aldosterone excess or "escape". / Doutorado / Doutor em Farmacologia