Aldosterone induces inflammatory cytokines in penile corpus cavernosum by activating the NF-κB pathway / 亚洲男科学杂志(英文版)

Emerging evidence indicates that aldosterone and mineralocorticoid receptors (MRs) are associated with the pathogenesis of erectile dysfunction. However, the molecular mechanisms remain largely unknown. In this study, freshly isolated penile corpus cavernosum tissue from rats was treated with aldosterone, with or without MRs inhibitors. Nuclear factor (NF)-kappa B (NF-κB) activity was evaluated by real-time quantitative PCR, luciferase assay, and immunoblot. The results demonstrated that mRNA levels of the NF-κB target genes, including inhibitor of NF-κB alpha (IκB-α), NF-κB1, tumor necrosis factor-alpha (TNF-α), and interleukin 6 (IL-6), were higher after aldosterone treatment. Accordingly, phosphorylation of p65/RelA, IκB-α, and inhibitor of NF-κB kinase-β was markedly increased by aldosterone. Furthermore, knockdown of MRs prevented activation of the NF-κB canonical pathway by aldosterone. Consistent with this finding, ectopic overexpression of MRs enhanced the transcriptional activation of NF-κB by aldosterone. More importantly, the MRs antagonist, spironolactone blocked aldosterone-mediated activation of the canonical NF-κB pathway. In conclusion, aldosterone has an inflammatory effect in the corpus cavernosum penis, inducing NF-κB activation via an MRs-dependent pathway, which may be prevented by selective MRs antagonists. These data reveal the possible role of aldosterone in erectile dysfunction as well as its potential as a novel pharmacologic target for treatment.

Espironolactona aumenta los niveles circulantes de angiotensina-(1-9) y revierte el remodelado cardíaco en la hipertensión arterial experimental / Spironolactone increases circulating levels of angiotensin-(1-9) and reverts cardiac remodeling in experimental hypertension

Previamente hemos demostrado que la eficacia de enalapril, candesartán y de fasudil -inhibidor de la vía RhoA/ROCK,- en el tratamiento del remodelado cardíaco en la hipertensión (HTA) e infarto al miocardio está mediada por aumento en los niveles circulantes del péptido vasoactivo angiotensina [Ang]-(1-9). Sin embargo, no hay información disponible si el antagonista del receptor de mineralocorticoide, espironolactona (espiro) disminuye el remodelado cardíaco aumentando los niveles circulantes de Ang-(1-9). El objetivo de este trabajo fue determinar si espironolac-tona disminuye el remodelado cardíaco aumentando los niveles circulantes de Ang-(1-9) en la hipertensión arterial experimental. Métodos. Estudio comparativo de 3 grupos experimentales. Se utilizaron ratas Sprague Dawley macho (150 ± 10 grs) unifrectomizadas tratadas con desoxi-corticosterona (DOCA, 60mg/Kg 2 veces sem, im) por 6 semanas. Como controles (Sham) se usaron ratas unifrectomizadas. A partir de la 3° semana las ratas DOCA con HTA> 140 mmHg fueron randomizadas a recibir vehículo o espiro (100 mg/kg día, gavage) por 3 sem. Al finalizar el tratamiento se determinó la presión arterial sistólica (PAS), masa corporal, peso del corazón (PC) y masa cardíaca relativa al largo de la tibia (MCR, mg ventrículos/LT*100). El grado de hipertrofia car-diomiocitaria se determinó midiendo el área y perímetro de los cardiomiocitos y la fibrosis por el contenido de colágeno en cortes teñidos con rojo picrosirio. Resultados (promedio ± ES): Conclusión: Espironolactona disminuye la PAS y aumenta los niveles circulantes de Ang-(1-9). Este aumento en los niveles circulantes de Ang-(1-9) se asocia con una disminución significativa de la hipertrofia y la fibrosis cardiaca hipertensiva. Este nuevo efecto de espironolactona en los niveles circulantes de Ang-(1-9), - péptido vasoactivo de la vía paralela del sistema renina-angiotensina-aldosterona,- podría contribuir al efecto antihipertensivo y disminución del daño cardiaco en la hipertensión y remodelamiento cardiovascular y renal patológico. Estos hallazgos pueden tener relevancia terapéutica en términos que Ang-(1-9) podría disminuir el daño cardiovascular patológico.

We have previously demonstrated that the efficacy of enalapril, candesartan and fasudil,- RhoA / ROCK inhibitor-, in the treatment of cardiac remodeling in hypertension (HT) and myocardial infarction is mediated by an increase in circulating levels of the vasoactive peptide angiotensin (Ang) -(1-9). However, it is not known whether the mineralocorticoid receptor antagonist, spironolactone (spiro) decreases cardiac remodeling by increasing the circulating levels of Ang- (1-9). The aim of this study was to determine whether spironolactone decreases cardiac remodeling by increasing circulating levels of Ang-(1-9) in experimental hypertension. Methods. Comparative study of 3 experimental groups. Unifirectomized male Sprague Dawley rats (150 ± 10 grams) were treated with deoxycorticos-terone (DOCA, 60 mg / kg 2 times a week, im) for 6 weeks. Unifirectomized rats were used as controls (Sham). At 3rd week after surgery, DOCA rats with HTA> 140 mmHg were randomized to receive vehicle or spironolactone (Spiro, 100 mg / kg day, gavage) for 3 weeks. At the end of treatment, systolic blood pressure (SBP), body mass (BM), heart weight (HW) and relative cardiac mass to the tibia length (MCR, mg ventricles / LT * 100) were determined. The degree of cardiomyocyte hypertrophy was determined by measuring the area and perimeter of cardiomyocytes and fibrosis by collagen content in sections stained with picrosirius red. Results (mean ± ES): Conclusion: Spironolactone decreases systolic blood pressure and increases circulating levels of Ang-(1-9). This increase in circulating levels of Ang- (1-9) was associated with a significant decrease in hypertrophy and hypertensive cardiac fibrosis. This new effect of spironolactone on the circulating levels of Ang- (1-9) - vasoactive peptide of the parallel pathway of the re-nin-angiotensin-aldosterone system - could contribute to the antihypertensive effect and decrease of cardiac damage in HT and cardiovascular remodeling and renal disease. These findings may have therapeutic relevance supporting that Ang-(1-9) may decreases pathologic cardiovascular damage.

Influence of antihypertensive drugs on aortic and coronary effects of Ang-(1-7) in pressure-overloaded rats

This study investigated the influence of antihypertensive drugs, such as angiotensin-converting enzyme inhibitors (ACEIs), AT1 receptor blockers (ARBs), voltage-gated L-type calcium channel blockers, and mineralocorticoid receptor antagonists (MRAs), on the effects of angiotensin-(1-7) [Ang-(1-7)] on aorta and coronary arteries from pressure-overloaded rats. Pressure overload was induced by abdominal aortic banding (AB). To evaluate the role of antihypertensive drugs on the effect of Ang-(1-7), AB male Wistar rats weighing 250–300 g were treated with vehicle or low doses (5 mg·kg-1·day-1, gavage) of losartan, captopril, amlodipine, or spironolactone. Isolated aortic rings and isolated perfused hearts under constant flow were used to evaluate the effect of Ang-(1-7) in thoracic aorta and coronary arteries, respectively. Ang-(1-7) induced a significant relaxation in the aorta of sham animals, but this effect was reduced in the aortas of AB rats. Chronic treatments with losartan, captopril or amlodipine, but not with spironolactone, restored the Ang-(1-7)-induced aorta relaxation in AB rats. The coronary vasodilatation evoked by Ang-(1-7) in sham rats was blunted in hypertrophic rats. Only the treatment with losartan restored the coronary vasodilatory effect of Ang-(1-7) in AB rat hearts. These data support a beneficial vascular effect of an association of Ang-(1-7) and some antihypertensive drugs. Thus, this association may have potential as a new therapeutic strategy for cardiovascular diseases.

Efecto de la administración de espironolactona sobre la pérdida de podocitos y la progresión de la nefropatía diabética experimental / Effects of spironolactone administration on the podocytes loss and progression of experimental diabetic nephropathy

Objetivos. Evaluar el efecto de espironolactona (SPL) sobre la pérdida de los podocitos durante la progresión de la nefropatía diabética (ND) experimental. Materiales y métodos. Aleatoriamente un grupo de ratas macho Holtzman recibieron estreptozotocina (grupo diabético) o citrato buffer (grupo control). Las ratas diabéticas fueron tratadas con SPL (50 mg/kg/día). El área glomerular y la celularidad fueron evaluadas por métodos histomorfométricos. La lesión y pérdida de podocitos fue evaluada por la expresión de desmina y Wt-1, respectivamente. La expresión génica del TGF-β1 se evaluó mediante RT-PCR. Resultados. Los niveles de glucosa, el área glomerular, la expansión mesangial y el contenido de colágeno se incrementaron significativamente en las ratas diabéticas. La administración de SPL previno estos cambios sin modificar los niveles de glucosa. La inmunotinción para Wt-1 se redujo significativamente, mientras que la inmunotinción para desmina se incrementó drásticamente en las ratas diabéticas. El tratamiento con SPL previno el incremento de expresión de desmina y la pérdida de expresión de Wt-1. Asimismo, la administración de SPL previno el incremento de la expresión del mRNA del TGF-β1 en las ratas diabéticas. Conclusiones. El tratamiento con SPL, a través de efectos glucosa independientes, atenúa la perdida de podocitos y la progresión de los cambios morfológicos de la ND. Los presentes resultados sugieren que estos efectos son mediados, al menos en parte, por la inhibición de la la expresión del mRNA del TGF-β1.

Objectives. Evaluate the effect of spironolactone (SPL) on the loss of podocytes during the progression of experimental diabetic nephropathy (DN). Materials and methods. A group of male Holtzman rats randomly received streptozotocin (diabetic group) or a buffer citrate (control group). Diabetic rats were treated with SPL (50 mg/kg/day). The glomerular area and the cellularity were evaluated by histomorphometric methods. The injury and loss of podocytes was assessed by desmin expression and Wt-1, respectively. The gene expression of TGF-β1 was assessed by RT-PCR. Results. Glucose levels, the glomerular area, the mesangial expansion and collagen content increased significantly in diabetic rats. The administration of SPL prevented these changes without changing glucose levels. Immunostain for Wt-1 decreased significantly while immunostain for desmin increased dramatically in diabetic rats. Treatment with SPL prevented the increase of desmin expression and the loss of Wt-1 expression. Furthermore, the administration of SPL prevented the increase of TGF-β1 mRNA expression in diabetic rats. Conclusions. Treatment with SPL, through independent glucose effects, reduces the loss of podocytes and the progression of DN morphological changes. These results suggest that these effects are mediated, at least in part, by the inhibition of TGF-β1 mRNA expression.

Effect of aldosterone on the amplification of oncolytic vaccinia virus in human cancer lines / 대한간학회지

BACKGROUND/AIMS: JX-594 is an oncolytic virus derived from the Wyeth vaccinia strain that causes replication-dependent cytolysis and antitumor immunity. Starting with a cross-examination of clinical-trial samples from advanced hepatocellular carcinoma patients having high levels of aldosterone and virus amplification in JX-594 treatment, we investigated the association between virus amplification and aldosterone in human cancer cell lines. METHODS: Cell proliferation was determined by a cell-counting-kit-based colorimetric assay, and vaccinia virus quantitation was performed by quantitative polymerase chain reaction (qPCR) and a viral plaque assay. Also, the intracellular pH was measured using a pH-sensitive dye. RESULTS: Simultaneous treatment with JX-594 and aldosterone significantly increased viral replication in A2780, PC-3, and HepG2 cell lines, but not in U2OS cell lines. Furthermore, the aldosterone treatment time altered the JX-594 replication according to the cell line. The JX-594 replication peaked after 48 and 24 hours of treatment in PC-3 and HepG2 cells, respectively. qPCR showed that JX-594 entry across the plasma membrane was increased, however, the changes are not significant by the treatment. This was inhibited by treatment with spironolactone (an aldosterone-receptor inhibitor). JX-594 entry was significantly decreased by treatment with EIPA [5-(N-ethyl-N-isopropyl)amiloride; a Na+/H+-exchange inhibitor], but aldosterone significantly restored JX-594 entry even in the presence of EIPA. Intracellular alkalization was observed after aldosterone treatment but was acidified by EIPA treatment. CONCLUSIONS: Aldosterone stimulates JX-594 amplification via increased virus entry by affecting the H+ gradient.

Aldosterone Modulates Cell Proliferation and Apoptosis in the Neonatal Rat Heart

In the present study, we investigated whether and how the mineralocorticoid receptor antagonist spironolactone affects cardiac growth and development through apoptosis and cell proliferation in the neonatal rat heart. Newborn rat pups were treated with spironolactone (200 mg/kg/d) for 7 days. The cell proliferation was studied by PCNA immunostaining. The treatment with spironolactone decreased proliferating myocytes by 32% (P<0.05), and reduced myocytes apoptosis by 29% (P<0.05). Immunoblot and immunohistochemistry for the expression of p38, p53, clusterin, TGF-beta2, and extracellular signal-regulated kinase were performed. In the spironolactone group, p38, p53, clusterin, and TGF-beta2 protein expression was significantly decreased (P<0.05). These results indicate that aldosterone inhibition in the developing rat heart induces cardiac growth impairment by decreasing proliferation and apoptosis of myocytes.

Aldosterone receptor antagonists induce favorable cardiac remodeling in diastolic heart failure patients / Antagonistas de receptores de aldosterona inducen remodelación cardiaca favorable en pacientes con insuficiencia cardiaca diastólica

Background. Serum levels of aldosterone in heart failure are increased up to 20 times compared to normal subjects. After an acute myocardial infarction, aldosterone increases progressively as well as interstitial fibrosis and collagen synthesis from cardiac fibroblasts, forming a patchy heterogeneous interstitial collagen matrix that affects ventricular function. Even if angiotensine converting enzyme inhibitors (ACEI) or angiotensin II receptor antagonists (ARA) can reduce aldosterone levels early during treatment, they increase again after a 12 week treatment. The aim of this study was to evaluate the changes in structure and function of the left ventricle in symptomatic (NYHA I-III) diastolic heart failure patients receiving an aldosterone receptor antagonist. Methods. Twenty-eight subjects with diastolic heart failure, on BB, ACEI and/or ARA were randomized to receive spironolactone (group A) on a mean dose of 37.5 mg once a day (n =14, age 63.7 ± 21.6 years and body mass index, BMI 27.5 ± 9.4), or not (group B, n = 14, Age 64.8 ± 11.9, BMI 26.9 ± 4.7). All patients were followed-up for a mean of 13.79 ± 0.99 months. Results. Group A showed a 42.8% ischemic origin of heart failure, while in group B was 55% (p = 0.2). No other co-morbidities were significativelly different among both groups. Mean percentage of changes by echocardiogram was as follows: Interventricular septum (IVS) -12.2 ± 11% vs. 1.3 ± 15.2 (p = 0.03), pulmonary systolic artery pressure (PSAP was 0.99 ± 3.8% vs. 10.5 ± 9.1, p = 0.05). Other parameters did not show statistically significant differences. Conclusion. Aldosterone receptor antagonists reduce or avoid increasing of PSAP and inducing a favorable remodeling of the left ventricle, especially in the IVS in diastolic heart failure patients.

Antecedentes. En pacientes con insuficiencia cardiaca existen aumentos de aldosterona hasta 20 veces mayores que en sujetos control. Después de un infarto miocárdico la aldosterona aumenta progresivamente, así como la fibrosis intersticial y la síntesis de colágena por fibroblastos cardiacos, provocando parches intersticiales heterogéneos en la matriz de colágena que afecta la función ventricular. El tratamiento inicial con inhibidores de enzima convertidora de angiotensina (IECA) y/o antagonistas de receptores de angiotensina II (ARA) puede reducir estos niveles; sin embargo, aumentan nuevamente después de 12 semanas de tratamiento. El propósito de este estudio fue evaluar los cambios estructurales y funcionales en el ventrículo izquierdo en pacientes con insuficiencia diastólica tratados con ARA angiotensina (NYHAI-III). Métodos. Veintiocho pacientes con insuficiencia cardiaca diastólica en tratamiento con BB, IECA y/o ARA se aleatorizaron a recibir una dosis media de 37.5 mg una vez al día de espironolactona (grupo A) (n - 14, edad 63.7 ± 0 21.6 años e índice masa corporal IMC 27.5 ± 9.4), o no (grupo B, n =14, edad 64.8 ± 11.9, IMC 26.9 ± 4.7). Todos los pacientes fueron seguidos por 13.79 ± 0.99 meses. Resultados. De los pacientes del grupo A, 42.8% y el 55 del grupo B (p = 0.2), tenían cardiopatía isquémica. No se encontraron diferencias significativas en otras comorbilidades. El porcentaje promedio de cambios en el ecocardiograma se observó en septum interventricular (SIV) -12.2 ± 11% vs. 1.3 ± 15.3% (p = 0.02), y la presión sistólica de la arteria pulmonar (PSAP, 0.99 ± 3.8% vs. 10.5 ± 9.1, p = 0.05, para los grupos A y B, respectivamente). Los otros parámetros no mostraron diferencias estadísticamente significativas. Conclusión. El tratamiento con antagonistas de receptores de aldosterona disminuye o limita aumentos de PSAP e inducen una remodelación favorable del ventrículo izquierdo, especialmente del SIV en pacientes con insuficiencia cardiaca diastólica.

Inhibitory effect of enalapril on serum aldosterone in essential hypertension

El objetivo del presente estudio fue evaluar en pacientes hipertensos esenciales, el efecto inhibitorio agudo y crónico del maleato de enalapril sobre la aldosterona plasmática. Diez pacientes con hipertensión arterial esencial leve y moderada (7 varones y 3 mujeres), con edades comprendidas entre 15 y 60 años, recibieron, luego de un período placebo de 28 días, 20mg de maleato de enalapril en una sola toma diaria. Durante el período placebo y a las 24, 48 y 72h de iníciado el tratamiento se determinó la actividad renínica plasmática (ARP) y los níveles de angiotensina II (AII) y aldosterona plasmática (A) por radioinmunoensayo, midiéndose asimismo la presión arterial media (PAM) utilizando un equipo automático de presión arterial (Critikon, Dynamap). Se comprobó un descenso significativo de la PAM de 120,6ñ4mmHg a 104,4ñ3, 102,6ñ2 y 100,5ñ1mmHg a las 24,48 y 72h, respectivamente (p<0,01). La ARP se incrementó de 0,68ñ0,6ng/ml/h a 9,33ñ2; 9,67ñ2 y 10,5ñ2ng/ml/h (p<0,05) y la AII descendió de 70,8ñ18pg/ml a 35,1ñ5; 27,6ñ3 y 25ñ4pg/ml (p<0,05)en el mismo período. La A descendió de 141,9ñ8 a 94,7ñ5pg/ml a las 24h (p<0,05), para retornar a las 48 y 72h a los niveles previos al tratamiento (106,7ñ6 y 118,5ñ7pg/ml). Ocho pacientes continuaron recibiendo enalapril 20mg/día durante un período de 6 meses al cabo del cual se determinó nuevamente AII y A previo y a las 2, 4 y 6h luego de la dosis diaria de enalapril. Se comprobó un descenso de la AII de 38,7ñ9 a 29ñ11pg/ml a las 2h (p<0,05), a 21ñ5pg/ml a las 4h (p<0,02) y a 17,5ñ4pg/ml a las 6h (p<0,01) de administrada la dosis de enalapril. La A descendió también de 170,14 a 106,3ñ8; 98,7ñ7 y 89,3ñ5pg/ml a las 2,4 y 6h de recibida la droga (p<0,01). En 7 de los 8 pacientes estudiados, se obtuvo una correlación individual entre AII y A que osciló en un rango de r entre 0,69 y la unidad. Se concluye que el enalapril mantiene el efecto inhibitorio sobre la enzima convertidora en el largo plazo al persistir reducidos los niveles de AII a los 6 meses de tratamiento. La tendencia a la recuperación de los niveles de aldosterona observados a las 48 y 72h y su posterior retorno a losvalores iniciales se debería a unasupersensibilidad del receptor de AII, aunque no podría descartarse la participación de la ACTH