RESUMEN
Long noncoding RNAs (lncRNAs), a group of transcripts, have been revealed to be critical participants in regulating multiple biological processes of malignant tumors. The knowledge of NPPA-AS1 (a new lncRNA) in cancer research is hardly known. Thus, it is of urgent need to study the underlying role of NPPA antisense RNA 1 (NPPA-AS1) in cervical cancer (CC). In this study, NPPA-AS1 was discovered to be lowly expressed and upregulation of it impaired cell proliferation and migration in CC. Besides, downregulation of it led to opposite results. Molecular mechanism assays uncovered that increased expression of NPPA-AS1 could inactivate Wnt/ß-catenin signaling pathway in CC. In addition, NPPA-AS1 was found to negatively interact with miR-302e whereas positively correlate with dickkopf-1 (DKK1, an inhibitor of Wnt pathway) in CC. Besides, loss of function assay illuminated that miR-302e inhibition restrained cell proliferation and migration in CC. Subsequent rescue assays confirmed that NPPA-AS1 acted as a competing endogenous RNA in CC by sponging miR-302e to upregulate DKK1 expression. Finally, the RE-1 silencing transcription factor (REST) was testified to function as a transcription suppressor of NPPA-AS1 in CC. In brief, REST-repressed NPPA-AS1 regulates CC progression by modulating miR-302e/DKK1/Wnt/ß-catenin signaling pathway.
Asunto(s)
Regulación Neoplásica de la Expresión Génica , Péptidos y Proteínas de Señalización Intercelular/metabolismo , ARN Largo no Codificante/genética , Proteínas Represoras/metabolismo , Neoplasias del Cuello Uterino/patología , Proteína Wnt1/metabolismo , beta Catenina/metabolismo , Animales , Apoptosis , Factor Natriurético Atrial/antagonistas & inhibidores , Factor Natriurético Atrial/genética , Biomarcadores de Tumor/genética , Biomarcadores de Tumor/metabolismo , Proliferación Celular , Femenino , Humanos , Péptidos y Proteínas de Señalización Intercelular/genética , Masculino , Ratones , Ratones Desnudos , MicroARNs/genética , Persona de Mediana Edad , Pronóstico , ARN sin Sentido/genética , Proteínas Represoras/genética , Tasa de Supervivencia , Células Tumorales Cultivadas , Neoplasias del Cuello Uterino/genética , Neoplasias del Cuello Uterino/metabolismo , Proteína Wnt1/genética , Ensayos Antitumor por Modelo de Xenoinjerto , beta Catenina/genéticaRESUMEN
Atrial and brain natriuretic peptides (ANP and BNP) exist in the central nervous system and modulate neuronal function, although the locus of actions and physiological mechanisms are still unclear. In the present study we used rat spinal sacral dorsal commissural nucleus (SDCN) and hippocampal 'synaptic bouton' preparations, to record both spontaneous and evoked glycinergic inhibitory postsynaptic currents (sIPSCs and eIPSCs) in SDCN neurons, and the evoked excitatory postsynaptic currents (eEPSCs) in hippocampal CA3 neurons. ANP potently and significantly reduced the sIPSC frequency without affecting the amplitude. ANP also potently reduced the eIPSCs amplitude concurrently increasing the failure rate and the paired pulse ratio response. These ANP actions were blocked by anantin, a specific type A natriuretic peptide receptor (NPR-A) antagonist. The results clearly indicate that ANP acts directly on glycinergic presynaptic nerve terminals to inhibit glycine release via presynaptic NPR-A. The ANP effects were not blocked by the membrane permeable cGMP analog (8Br-cGMP) suggesting a transduction mechanisms not simply related to increasing cGMP levels in nerve terminals. BNP did not affect on glycinergic sIPSCs and eIPSCs. Moreover, both ANP and BNP had no effect on glutamatergic EPSCs in hippocampal CA3 neurons. The results indicate a potent and selective presynaptic inhibitory action of ANP on glycinergic transmission in spinal cord sensory circuits.
Asunto(s)
Factor Natriurético Atrial/farmacología , Glicina/metabolismo , Neuronas/efectos de los fármacos , Terminales Presinápticos/efectos de los fármacos , Médula Espinal/citología , 4-Aminopiridina/farmacología , 6-Ciano 7-nitroquinoxalina 2,3-diona/farmacología , Anestésicos Locales/farmacología , Animales , Animales Recién Nacidos , Factor Natriurético Atrial/antagonistas & inhibidores , GMP Cíclico/análogos & derivados , GMP Cíclico/farmacología , Antagonistas de Aminoácidos Excitadores/farmacología , Potenciales Postsinápticos Excitadores/efectos de los fármacos , Hipocampo/citología , Potenciales Postsinápticos Inhibidores/efectos de los fármacos , Lidocaína/análogos & derivados , Lidocaína/farmacología , Péptidos Cíclicos/farmacología , Bloqueadores de los Canales de Potasio/farmacología , Terminales Presinápticos/fisiología , Ratas , Ratas Wistar , Tionucleótidos/farmacologíaRESUMEN
PURPOSE: The retinal pigment epithelium (RPE) expresses aquaporin-1 (AQP1) and components of the natriuretic peptide signaling pathway. We hypothesized that stimulation of the natriuretic signaling pathway in RPE with atrial natriuretic peptide (ANP) and with membrane-permeable analogs of cGMP would induce a net apical-to-basal transport of fluid. METHODS: The hypothesis was tested using human RPE cultures that retain properties seen in vivo. Confluent monolayers were treated with ANP or membrane-permeable cGMP analogs in the presence of anantin, H-8, and an AQP1 inhibitor, AqB013. Fluid movement from the apical to basal chambers was measured by weight and used to calculate net fluid transport. RESULTS: Our results demonstrated a 40% increase in net apical-to-basal fluid transport by ANP (5 µM) that was inhibited completely by the ANP receptor antagonist anantin and a 60% increase in net apical-to-basal fluid transport in response to the extracellularly applied membrane-permeable cGMP analog pCPT-cGMP (50 µM), which was not affected by the protein kinase G inhibitor H-8. The aquaporin antagonist AqB013 (20 µM) inhibited the cGMP-stimulated RPE fluid flux. CONCLUSIONS: The effect of cGMP is consistent with an enhancement of the net fluid flux in RPE mediated by AQP1 channels. Pharmacologic activation of cGMP signaling and concomitant stimulation of fluid uptake from the subretinal space could offer insights into a new approach to treating or reducing the risk of retinal detachment.
Asunto(s)
Acuaporina 1/metabolismo , GMP Cíclico/farmacología , Epitelio Pigmentado de la Retina/efectos de los fármacos , Agua/metabolismo , Animales , Acuaporina 1/antagonistas & inhibidores , Factor Natriurético Atrial/antagonistas & inhibidores , Factor Natriurético Atrial/farmacología , Transporte Biológico Activo , Western Blotting , Permeabilidad de la Membrana Celular/efectos de los fármacos , Células Cultivadas , GMP Cíclico/análogos & derivados , Proteínas Quinasas Dependientes de GMP Cíclico/metabolismo , Relación Dosis-Respuesta a Droga , Humanos , Isoquinolinas/farmacología , Péptidos Cíclicos/farmacología , Epitelio Pigmentado de la Retina/metabolismo , Xenopus laevisRESUMEN
Atrial natriuretic peptide (ANP) has been recently identified as a modulator of acute lung injury (ALI) induced by pro-inflammatory agonists. While previous studies tested effects of exogenous ANP administration, the role of endogenous ANP in the course of ALI remains unexplored. This study examined regulation of ANP and its receptors NPR-A, NPR-B and NPR-C by LPS and involvement of ANP receptors in the modulation of LPS-induced lung injury. Primary cultures of human pulmonary endothelial cells (EC) were used in the in vitro tests. Expression of ANP and its receptors was determined by quantitative RT-PCR analysis. Agonist-induced cytoskeletal remodeling was evaluated by immunofluorescence staining, and EC barrier function was characterized by measurements of transendothelial electrical resistance. In the murine model of ALI, LPS-induced lung injury was assessed by measurements of protein concentration and cell count in bronchoalveolar lavage fluid (BAL). LPS stimulation significantly increased mRNA expression levels of ANP and NPR-A in pulmonary EC. Pharmacological inhibition of NPR-A augmented LPS-induced EC permeability and blocked barrier protective effects of exogenous ANP on LPS-induced intercellular gap formation. In contrast, pharmacological inhibition of ANP clearance receptor NPR-C significantly attenuated LPS-induced barrier disruptive effects. Administration of NPR-A inhibitor in vivo exacerbated LPS-induced lung injury, whereas inhibition of NPR-C suppressed LPS-induced increases in BAL cell count and protein content. These results demonstrate for the first time opposite effects of NPR-A and NPR-C in the modulation of ALI and suggest a compensatory protective mechanism of endogenous ANP in the maintenance of lung vascular permeability in ALI.
Asunto(s)
Permeabilidad Capilar , Células Endoteliales/metabolismo , Lipopolisacáridos , Lesión Pulmonar/prevención & control , Pulmón/irrigación sanguínea , Arteria Pulmonar/metabolismo , Receptores del Factor Natriurético Atrial/metabolismo , Animales , Factor Natriurético Atrial/antagonistas & inhibidores , Factor Natriurético Atrial/metabolismo , Líquido del Lavado Bronquioalveolar/química , Líquido del Lavado Bronquioalveolar/citología , Permeabilidad Capilar/efectos de los fármacos , Células Cultivadas , Citoesqueleto/metabolismo , Modelos Animales de Enfermedad , Impedancia Eléctrica , Células Endoteliales/efectos de los fármacos , Células Endoteliales/patología , Técnica del Anticuerpo Fluorescente , Antagonistas de Hormonas/farmacología , Humanos , Pulmón/patología , Lesión Pulmonar/inducido químicamente , Lesión Pulmonar/genética , Lesión Pulmonar/metabolismo , Lesión Pulmonar/patología , Masculino , Ratones , Ratones Endogámicos C57BL , Arteria Pulmonar/efectos de los fármacos , Arteria Pulmonar/patología , ARN Mensajero/metabolismo , Receptores del Factor Natriurético Atrial/efectos de los fármacos , Receptores del Factor Natriurético Atrial/genética , Reacción en Cadena de la Polimerasa de Transcriptasa Inversa , Factores de Tiempo , Factor de Necrosis Tumoral alfa/metabolismoRESUMEN
MDMA (3,4-methylenedioxymethamphetamine) induces thermogenesis in a mitochondrial uncoupling protein 3-dependent manner. There is evidence that this hyperthermia is mediated in part by the lipolytic release of free fatty acids, that subsequently activate uncoupling protein 3 in skeletal muscle mitochondria. We hypothesize that atrial natriuretic peptide (ANP), a strong lipolytic mediator, may contribute to the induction and maintenance of MDMA-induced thermogenesis. The specific aims of this study were to (1) determine if ANP is released following MDMA administration, and (2) use the ANP receptor antagonist, Anantin, to ascertain the role of ANP in MDMA-induced hyperthermia. ANP levels were measured in plasma at baseline, 10, 20, 30 and 60 min following MDMA (40 mg/kg, sc) administration in 16 male Sprague-Dawley rats. A robust increase in ANP was seen within 10 min of MDMA administration. ANP levels returned to baseline at 20 min and then gradually rose over the 60 min monitoring period. The administration of Anantin (40 mg, ip), 15 min before and after MDMA, significantly attenuated the MDMA-induced hyperthermia. We conclude that ANP signaling contributes to the hyperthermia induced by MDMA.
Asunto(s)
Factor Natriurético Atrial/sangre , Fiebre/inducido químicamente , Alucinógenos/toxicidad , N-Metil-3,4-metilenodioxianfetamina/toxicidad , Pirógenos/toxicidad , Animales , Antipiréticos/uso terapéutico , Factor Natriurético Atrial/antagonistas & inhibidores , Regulación de la Temperatura Corporal/efectos de los fármacos , Fiebre/sangre , Fiebre/prevención & control , Alucinógenos/antagonistas & inhibidores , Masculino , N-Metil-3,4-metilenodioxianfetamina/antagonistas & inhibidores , Péptidos Cíclicos/uso terapéutico , Pirógenos/antagonistas & inhibidores , Distribución Aleatoria , Ratas , Ratas Sprague-Dawley , Receptores del Factor Natriurético Atrial/antagonistas & inhibidores , Transducción de Señal/efectos de los fármacos , Simpatomiméticos/antagonistas & inhibidores , Simpatomiméticos/toxicidad , Factores de TiempoRESUMEN
Caveolae may act as mechanosensors and function as binding sites for calcium ions. The intracaveolar localization of atrial natriuretic peptide (ANP) derived from the direct interaction of atrial granules with caveolae has been demonstrated. The aim of this study was to define the effect of caveolae on ANP secretion induced by stretch and angiotensin II. The isolated perfused beating atria from Sprague-Dawley rats were used. To disrupt caveolae, 10mM methyl-ß-cyclodextrin (MbCD) was applied for 1h and the number of caveoli were markedly decreased. MbCD increased basal ANP secretion and atrial diastolic pressure. The molecular profile of ANP in perfusate from control atria showed mainly one major peak corresponded to synthetic ANP whereas that from MbCD-treated atria showed two major immunoreactive peaks corresponded to synthetic rat ANP and proANP. High atrial stretch induced by elevating the height of outflow catheter from 5 cm H2O to 7.5 cm H2O increased atrial contractility and ANP secretion. The response of ANP secretion to high stretch was attenuated in MbCD-pretreated atria. Pretreatment with MbCD abolished angiotensin II-induced suppression and losartan-induced stimulation of ANP secretion. However, the effect of angiotenisin (1-7) on ANP secretion was not altered by MbCD treatment. The expression of angiotensin II type 1 receptor protein was reduced by MbCD treatment. These data suggest that caveolae are essential for angiotensin II type 1 receptor-mediated ANP secretion and relate to the processing of proANP.
Asunto(s)
Factor Natriurético Atrial , Atrios Cardíacos/metabolismo , Miocardio/metabolismo , Precursores de Proteínas , Receptor de Angiotensina Tipo 1/metabolismo , beta-Ciclodextrinas/farmacología , Angiotensina II/farmacología , Bloqueadores del Receptor Tipo 1 de Angiotensina II/farmacología , Animales , Factor Natriurético Atrial/agonistas , Factor Natriurético Atrial/antagonistas & inhibidores , Factor Natriurético Atrial/biosíntesis , Factor Natriurético Atrial/metabolismo , Presión Sanguínea , Cateterismo , Caveolas/metabolismo , Cromatografía Líquida de Alta Presión , Atrios Cardíacos/efectos de los fármacos , Bombas de Infusión , Losartán/farmacología , Masculino , Contracción Miocárdica/efectos de los fármacos , Contracción Miocárdica/fisiología , Técnicas de Cultivo de Órganos , Perfusión , Precursores de Proteínas/agonistas , Precursores de Proteínas/antagonistas & inhibidores , Precursores de Proteínas/biosíntesis , Precursores de Proteínas/metabolismo , Radioinmunoensayo , Ratas , Ratas Sprague-Dawley , Receptor de Angiotensina Tipo 1/genéticaRESUMEN
Somatostatin is a cyclic-14 amino acid peptide which mainly distributed in digestive system and brain. Somatostatin receptor (SSTR) is a G-protein coupled receptor and all five SSTR subtypes are expressed in cardiomyocytes. The aim of this study was to investigate the effect of somatostatin on atrial natriuretic peptide (ANP) secretion and its signaling pathway. Somatostatin (0.01 and 0.1nM) decreased ANP secretion in isolated beating rat atrium in a dose-dependent manner. But atrial contractility and translocation of extracellular fluid were not changed. Somatostatin-induced decrease in ANP secretion was significantly attenuated by the pretreatment with CYN 154806 (SSTR type 2 antagonist; 0.1µM), but not by BIM 23056 (SSTR type 5 antagonist; 0.1µM) and urantide (urotensin II receptor antagonist; 0.1µM). When pretreated with an agonist for SSTR type 2 (Seglitide, 0.1nM) and SSTR type 5 (L 817818, 0.1nM), only Seglitide reduced ANP secretion similar to that of somatostatin. The suppressive effect of somatostatin on ANP secretion was attenuated by the pretreatment with an inhibitor for adenylyl cyclase (MDL-12330A, 5µM) or protein kinase A (KT 5720, 0.1µM). In diabetic rat atria, the suppressive effect of somatostatin on ANP secretion and concentration was attenuated. Real time-PCR and western blot shows the decreased level of SSTR type 2 mRNA and protein in diabetic rat atria. These data suggest that somatostatin decreased ANP secretion through SSTR type 2 and an attenuation of suppressive effect of somatostatin on ANP secretion in diabetic rat atria is due to a down-regulation of SSTR type 2.
Asunto(s)
Función Atrial/efectos de los fármacos , Factor Natriurético Atrial , Atrios Cardíacos , Contracción Miocárdica/efectos de los fármacos , Receptores de Somatostatina , Somatostatina/farmacología , Animales , Factor Natriurético Atrial/antagonistas & inhibidores , Factor Natriurético Atrial/metabolismo , Western Blotting , Proteínas Quinasas Dependientes de AMP Cíclico/antagonistas & inhibidores , Proteínas Quinasas Dependientes de AMP Cíclico/metabolismo , Diabetes Mellitus Experimental/fisiopatología , Líquido Extracelular/efectos de los fármacos , Atrios Cardíacos/efectos de los fármacos , Atrios Cardíacos/metabolismo , Hemodinámica/efectos de los fármacos , Iminas/farmacología , Masculino , Contracción Miocárdica/fisiología , Oligopéptidos/farmacología , Técnicas de Cultivo de Órganos , Péptidos Cíclicos/farmacología , Reacción en Cadena de la Polimerasa , Inhibidores de Proteínas Quinasas/farmacología , ARN Mensajero/análisis , Ratas , Ratas Sprague-Dawley , Receptores de Somatostatina/agonistas , Receptores de Somatostatina/antagonistas & inhibidores , Receptores de Somatostatina/metabolismo , Transducción de Señal/efectos de los fármacos , Somatostatina/antagonistas & inhibidoresRESUMEN
Reactive oxygen species (ROS) play a role in cardiovascular diseases such as hypertension and heart failure. The objective of the present study was to investigate the role of endogenous ROS in atrial hemodynamics and ANP secretion in isolated perfused beating rat atria. Pyrogallol (a generator of superoxide anion, 0.1, 1mM) or hydrogen peroxide (0.1, 1, 10, 30mM) was perfused into atria paced at 1.2Hz. Pyrogallol and hydrogen peroxide stimulated ANP secretion and concentration in a dose-dependent manner and dramatically decreased atrial contractility and translocation of extracellular fluid. The stimulatory effect of pyrogallol and hydrogen peroxide on ANP secretion was attenuated by the pretreatment with ascorbic acid (an antioxidant; 1mM) and cariporide (an inhibitor of the Na(+)/H(+) exchanger; 1µM) but negative inotropic effect was not changed. U120 (a MAPK(erk) pathway inhibitor; 10µM) attenuated the stimulatory effect of hydrogen peroxide on ANP secretion. However, U120 augmented negative inotropic effect and stimulatory effect of ANP concentration induced by pyrogallol. Antioxidant such as N-acetyl cystein, gallate, propyl gallate, or ellagic acid did not cause any significant changes in atrial parameters. These results suggest that intracellular - formed ROS stimulates ANP secretion partly through activation of MAPK(erk) pathway and Na(+)/H(+) exchanger.
Asunto(s)
Factor Natriurético Atrial , Líquido Extracelular/efectos de los fármacos , Atrios Cardíacos , Hemodinámica/efectos de los fármacos , Contracción Miocárdica/efectos de los fármacos , Estrés Oxidativo/efectos de los fármacos , Acetilcisteína/farmacología , Animales , Ácido Ascórbico/farmacología , Factor Natriurético Atrial/antagonistas & inhibidores , Factor Natriurético Atrial/metabolismo , Enfermedades Cardiovasculares/fisiopatología , Guanidinas/farmacología , Atrios Cardíacos/efectos de los fármacos , Atrios Cardíacos/metabolismo , Peróxido de Hidrógeno/efectos adversos , Masculino , Quinasas de Proteína Quinasa Activadas por Mitógenos/antagonistas & inhibidores , Contracción Miocárdica/fisiología , Técnicas de Cultivo de Órganos , Galato de Propilo/farmacología , Pirogalol/efectos adversos , Ratas , Ratas Sprague-Dawley , Especies Reactivas de Oxígeno , Intercambiadores de Sodio-Hidrógeno/antagonistas & inhibidores , Sulfonas/farmacologíaRESUMEN
AIMS: Although a beta-adrenoceptor (beta-AR) blockade-induced increase in plasma atrial natriuretic peptide (ANP) levels is implicated in the therapeutic significance of beta-AR antagonists, the role of beta-AR in the regulation of ANP release is not clearly defined. The purpose of the present study was to define the role of beta-AR subtypes and the mechanisms responsible for regulation of atrial ANP release. MAIN METHODS: Experiments were performed in isolated perfused beating rabbit atria, including measurement of atrial contractile response, cAMP efflux, and atrial myocyte ANP release. KEY FINDINGS: beta-AR activation with (-)-isoproterenol decreased ANP release concomitantly with increases in cAMP efflux concentration, atrial dynamics, stroke volume and pulse pressure in a concentration-dependent manner. The ANP response was inversely related to the change in cAMP efflux concentrations. The isoproterenol-induced decrease in ANP release was inhibited by beta(1)-AR blockade with CGP 20712A but not by beta(2)-AR blockade with ICI 118551. The isoproterenol-induced decrease in ANP release was attenuated by the L-type Ca(2+) channel antagonist nifedipine and the cAMP-dependent protein kinase inhibitor KT5720. SIGNIFICANCE: These findings suggest that beta(1)-AR activation decreases ANP release via cAMP- and Ca(2+)-dependent mechanisms.
Asunto(s)
Factor Natriurético Atrial/metabolismo , Calcio/metabolismo , AMP Cíclico/metabolismo , Atrios Cardíacos/metabolismo , Receptores Adrenérgicos beta 1/metabolismo , Agonistas de Receptores Adrenérgicos beta 1 , Agonistas de Receptores Adrenérgicos beta 2 , Agonistas Adrenérgicos beta/farmacología , Animales , Factor Natriurético Atrial/antagonistas & inhibidores , Canales de Calcio Tipo L/metabolismo , Técnicas In Vitro , Isoproterenol/farmacología , Conejos , Receptores Adrenérgicos beta 2/metabolismoRESUMEN
Rapid non-genomic effects of 17beta-estradiol, the principal circulating estrogen, have been observed in a wide variety of cell types. Here we investigate rapid signaling effects of 17beta-estradiol in rat hepatocytes. We show that, above a threshold concentration of 1 nm, 17beta-estradiol, but not 17alpha-estradiol, stimulates particulate guanylyl cyclase to elevate cGMP, which through activation and plasma membrane recruitment of protein kinase G isoform Ialpha, stimulates plasma membrane Ca(2+)-ATPase-mediated Ca(2+) efflux from rat hepatocytes. These effects are extremely rapid in onset and are mimicked by a membrane-impermeant 17beta-estradiol-BSA conjugate, suggesting that 17beta-estradiol acts at the extracellular face of the plasma membrane. We also show that 17beta-estradiol binds specifically to the intact hepatocyte plasma membrane through an interaction that is competed by an excess of atrial natriuretic peptide but also shows many similarities to the pharmacological characteristics of the putative gamma-adrenergic receptor. We, therefore, propose that the observed rapid signaling effects of 17beta-estradiol are mediated either through the guanylyl cyclase A receptor for atrial natriuretic peptide or through the gamma-adrenergic receptor, which is either itself a transmembrane guanylyl cyclase or activates a transmembrane guanylyl cyclase through cross-talk signaling.
Asunto(s)
Calcio/metabolismo , Membrana Celular/enzimología , Proteínas Quinasas Dependientes de GMP Cíclico/metabolismo , GMP Cíclico/metabolismo , Estradiol/farmacología , Estrógenos/farmacología , Hepatocitos/metabolismo , Animales , Factor Natriurético Atrial/antagonistas & inhibidores , Factor Natriurético Atrial/farmacología , ATPasas Transportadoras de Calcio/metabolismo , Células Cultivadas , Proteína Quinasa Dependiente de GMP Cíclico Tipo I , Antagonismo de Drogas , Activación Enzimática/efectos de los fármacos , Femenino , Guanilato Ciclasa/metabolismo , Ratas , Ratas Wistar , Receptores Adrenérgicos/metabolismoRESUMEN
Up-regulation of atrial natriuretic peptide (ANP) mRNA in the kidneys in several disorders has been demonstrated; however, evidence that ANP synthesized by the kidney exerts a local function has never been produced. Therefore, we investigated whether endogenous ANP could modulate high glucose-stimulated TGF-beta1, collagen type I and nuclear factor-kappaB (NF-kappaB) in NRK-52E cells using transfection of ANP and ANP small interfering RNA (siANP). NRK-52E cells were grown with or without transfection with ANP plasmid; cells were also transfected with ANP siRNA or control siRNA. These cells were then stimulated with a high glucose concentration to modulate ANP, TGF-beta1, collagen type I, NF-kappaB and IkappaB-alpha, and the results showed that ANP, TGF-beta1, collagen type I and NF-kappaB significantly increased in untransfected cells, and the transfection of ANP significantly attenuated high glucose-activated TGF-beta1, collagen I and NF-kappaB expression. ANP siRNA knocked-down ANP but significantly increased TGF-beta1 and collagen I under normal glucose conditions; ANP siRNA decreased IkappaB-alpha but strongly enhanced high glucose-activated TGF-beta1, collagen type I and NF-kappaB. In contrast, medium from ANP-transfected cells attenuated high glucose-activated TGF-beta1 and collagen type I expression in NRK-52E cells transfected with siANP. In conclusion, our results demonstrated that siANP increased activation of TGF-beta1, collagen type I and NF-kappaB in NRK-52E cells under high glucose conditions, and medium from ANP-transfected cells attenuated high glucose-activated TGF-beta1 and collagen type I. This is the first study to demonstrate the auto/paracrine action of endogenous ANP in renal tubular cells on the attenuation of hyperglycemia-activated TGF-beta1 and NF-kappaB expression. J. Cell. Physiol. 219: 776-786, 2009. (c) 2009 Wiley-Liss, Inc.
Asunto(s)
Factor Natriurético Atrial/metabolismo , Túbulos Renales/metabolismo , Animales , Factor Natriurético Atrial/antagonistas & inhibidores , Factor Natriurético Atrial/genética , Secuencia de Bases , Línea Celular , Colágeno Tipo I/genética , Colágeno Tipo I/metabolismo , Medios de Cultivo Condicionados , Cartilla de ADN/genética , Expresión Génica/efectos de los fármacos , Silenciador del Gen , Glucosa/farmacología , Túbulos Renales/citología , Túbulos Renales/efectos de los fármacos , FN-kappa B/genética , FN-kappa B/metabolismo , ARN Interferente Pequeño/genética , Ratas , Proteínas Recombinantes/genética , Proteínas Recombinantes/metabolismo , Transfección , Factor de Crecimiento Transformador beta1/genética , Factor de Crecimiento Transformador beta1/metabolismoRESUMEN
Atrial natriuretic peptide (ANP) is an endogenous peptide hormone that is synthesized and secreted by the myocardium in health and disease. Although the bioactivity of this molecule has been studied extensively, cellular mechanisms governing its processing and secretion are not fully understood. Through a yeast two-hybrid screen of a cDNA library made from tissue of a failing human heart, we have discovered that the precursor of ANP, natriuretic peptide precursor (NPPA), physically interacts with the N-terminus of apoptosis signal-regulating kinase 1 (ASK1), a kinase believed to be involved in the pathogenesis of heart failure. We demonstrated that NPPA is a substrate of ASK1 in an in vitro kinase assay. Indirect immunofluorescence microscopy shows that, when expressed in Hela cells, ASK1 and NPPA exhibit distinct, but overlapping, staining patterns, suggesting partial colocalization in cells. Additionally, coexpressing wild-type ASK1 with NPPA in Hela cells led to reduced levels of NPPA in the culture medium, suggesting that ASK1 negatively impacts NPPA processing and/or secretion. This negative effect was less pronounced when a dominant-negative allele of ASK1 with deficient kinase activity was coexpressed with NPPA. Because both ASK1 and ANP are associated with pathologic cardiac hypertrophy, their interaction may have pathophysiological and therapeutic relevance.
Asunto(s)
Factor Natriurético Atrial/antagonistas & inhibidores , Factor Natriurético Atrial/metabolismo , MAP Quinasa Quinasa Quinasa 5/metabolismo , Factor Natriurético Atrial/fisiología , Regulación hacia Abajo/genética , Células HeLa , Humanos , MAP Quinasa Quinasa Quinasa 5/fisiología , Precursores de Proteínas/biosíntesis , Precursores de Proteínas/genética , Precursores de Proteínas/fisiología , Procesamiento Proteico-Postraduccional/genéticaRESUMEN
The aim of the present study was to determine the effects of increased cAMP levels in response to pituitary adenylate cyclase-activating polypeptide 27 (PACAP27) on atrial atrial natriuretic peptide (ANP) secretion in rabbit atria. A perfused beating atrial model was used in the present study and cAMP efflux and ANP levels in atrial perfusates were measured by radioimmnoassay. At 100 nmol/L, PACAP27 increased cAMP production, which resulted in subsequent inhibition of ANP secretion. Nicardipine (1.0 micromol/L), an L-type Ca2+ channel blocker, attenuated inhibition of ANP secretion by PACAP27. Staurosporine (1.0 micromol/L), a non-specific protein kinase inhibitor, and H-89 (1.0 micromol/L), a cAMP-dependent protein kinase A (PKA) inhibitor, completely blocked the inhibition of ANP secretion in response to PACAP27 but had no effect on PACAP27-induced increases in cAMP. In conclusion, the results suggest that increased cAMP levels in response to PACAP27 negatively regulate ANP secretion via the adenylate cyclase-cAMP-PKA signalling pathway in rabbit atria and that L-type Ca2+ channels may be involved, in part, in the regulation of ANP secretion by cAMP.
Asunto(s)
Factor Natriurético Atrial/antagonistas & inhibidores , Factor Natriurético Atrial/metabolismo , AMP Cíclico/biosíntesis , Corazón/metabolismo , Polipéptido Hipofisario Activador de la Adenilato-Ciclasa/fisiología , Animales , AMP Cíclico/fisiología , Corazón/efectos de los fármacos , Atrios Cardíacos/efectos de los fármacos , Atrios Cardíacos/metabolismo , Perfusión , Conejos , Transducción de Señal/efectos de los fármacos , Transducción de Señal/fisiologíaRESUMEN
Disturbances of volume-regulating mechanisms have already been implicated in the pathophysiology of eating disorders like anorexia or bulimia nervosa with the peptide hormones vasopressin and atrial natriuretic peptide (ANP) being of special interest. Aim of the present study was to investigate, whether the expression of the corresponding genes was altered and if so, if these changes could be explained by epigenetic mechanisms such as DNA methylation. We analyzed blood samples of 46 women suffering from anorexia (n=22) or bulimia nervosa (n=24) as well as of 30 healthy controls. Peripheral mRNA expression and DNA methylation of the vasopressin and the ANP precursor genes were assessed using real-time PCR. We found significantly lower levels of ANP mRNA in patients with eating disorders. This downregulation was accompanied by a hypermethylation of the ANP gene promoter in the bulimic subgroup. We did not find differences regarding expression or methylation of the vasopressin gene. ANP mRNA expression was inversely associated with impaired impulse regulation. We conclude that epigenetic mechanisms may contribute to the known alterations of ANP homeostasis in women with eating disorders.
Asunto(s)
Factor Natriurético Atrial/antagonistas & inhibidores , Factor Natriurético Atrial/metabolismo , Regulación hacia Abajo/genética , Epigénesis Genética/genética , Trastornos de Alimentación y de la Ingestión de Alimentos/genética , Conducta Impulsiva/genética , ARN Mensajero/biosíntesis , Vasopresinas/biosíntesis , Adolescente , Adulto , Factor Natriurético Atrial/genética , Metilación de ADN/genética , Trastornos de Alimentación y de la Ingestión de Alimentos/metabolismo , Femenino , Regulación de la Expresión Génica/genética , Humanos , Conducta Impulsiva/metabolismo , Persona de Mediana Edad , Regiones Promotoras Genéticas , ARN Mensajero/antagonistas & inhibidores , Vasopresinas/genéticaRESUMEN
Worsening renal function in the setting of human acute heart failure (AHF) predicts poor outcomes, such as rehospitalization and increased mortality. Understanding potential renoprotective mechanisms is warranted. The guanylate cyclase (GC) enzymes and their second messenger cGMP are the target of two important circulating neurohumoral systems with renoprotective properties. Specifically, natriuretic peptides (NP) released from the heart with AHF target particulate GC in the kidney, while the nitric oxide (NO) system is an activator of renal soluble GC. We hypothesized that both systems are essential to preserve renal excretory and hemodynamic function in AHF but with distinct roles. We investigated these roles in three groups of anesthetized dogs (6 each) with AHF induced by rapid ventricular pacing. After a baseline AHF clearance, each group received intrarenal vehicle (control), N(G)-monomethyl-l-arginine (l-NMMA), a competitive NO inhibitor (50 microg.kg(-1).min(-1)) or a specific NP receptor antagonist, HS-142-1 (0.5 mg/kg). We observed that intrarenal l-NMMA decreased renal blood flow (RBF) without significant decreases in glomerular filtration rate (GFR), urinary sodium excretion (UNaV), or urinary cGMP. In contrast, HS-142-1 resulted in a decrease in UNaV and cGMP excretion together with a reduction in GFR and an increase in distal fractional tubular sodium reabsorption. We conclude that in AHF, the NP system plays a role in maintaining sodium excretion and GFR, while the function of NO is in the maintenance of RBF. These studies have both physiological and therapeutic implications warranting further research into cardiorenal interactions in this syndrome of AHF.
Asunto(s)
Guanilato Ciclasa/metabolismo , Insuficiencia Cardíaca/metabolismo , Riñón/enzimología , Animales , Factor Natriurético Atrial/antagonistas & inhibidores , Factor Natriurético Atrial/metabolismo , GMP Cíclico/metabolismo , Perros , Riñón/efectos de los fármacos , Masculino , Polisacáridos/farmacología , omega-N-Metilarginina/farmacologíaRESUMEN
Previously, our laboratory demonstrated that cardiac mast cell degranulation induces adverse ventricular remodeling in response to chronic volume overload. The purpose of this study was to investigate whether atrial natriuretic peptide (ANP), which is known to be elevated in chronic volume overload, causes cardiac mast cell degranulation. Relative to control, ANP induced significant histamine release from peritoneal mast cells, whereas isolated cardiac mast cells were not responsive. Infusion of ANP (225 pg/ml) into blood-perfused isolated rat hearts produced minimal activation of cardiac mast cells, similar to that seen in the control group. ANP also did not increase matrix metalloproteinase-2 activity, reduce collagen volume fraction, or alter diastolic or systolic cardiac function compared with saline-treated controls. In a subsequent study to evaluate the effects of natriuretic peptide receptor antagonism on volume overload-induced ventricular remodeling, anantin was administered to rats with an aortocaval fistula. Comparable increases of myocardial MMP-2 activity in treated and untreated rats with an aortocaval fistula were associated with equivalent decreases in ventricular collagen (P < 0.05 vs. sham-operated controls). Cardiac functional parameters and left ventricular hypertrophy were unaffected by anantin. We conclude that ANP is not a cardiac mast cell secretagogue and is not responsible for the cardiac mast cell-mediated adverse ventricular remodeling in response to volume overload.
Asunto(s)
Fístula Arteriovenosa/complicaciones , Factor Natriurético Atrial/metabolismo , Degranulación de la Célula , Liberación de Histamina , Hipertrofia Ventricular Izquierda/metabolismo , Mastocitos/metabolismo , Miocardio/metabolismo , Animales , Aorta Abdominal/cirugía , Fístula Arteriovenosa/metabolismo , Fístula Arteriovenosa/patología , Fístula Arteriovenosa/fisiopatología , Líquido Ascítico/citología , Líquido Ascítico/metabolismo , Factor Natriurético Atrial/antagonistas & inhibidores , Factor Natriurético Atrial/farmacología , Degranulación de la Célula/efectos de los fármacos , Colágeno/metabolismo , Modelos Animales de Enfermedad , Liberación de Histamina/efectos de los fármacos , Hipertrofia Ventricular Izquierda/etiología , Hipertrofia Ventricular Izquierda/patología , Hipertrofia Ventricular Izquierda/fisiopatología , Técnicas In Vitro , Masculino , Mastocitos/efectos de los fármacos , Metaloproteinasa 2 de la Matriz/metabolismo , Miocardio/enzimología , Miocardio/patología , Péptidos Cíclicos/farmacología , Ratas , Ratas Sprague-Dawley , Vena Cava Inferior/cirugía , Función Ventricular Izquierda , Remodelación VentricularRESUMEN
Soluble guanylate cyclase is a heterodimeric enzyme with a prosthetic heme group that, on binding of its main ligand, NO, generates the second messenger cGMP. Unlike conventional nitrovasodilators, the novel direct NO- and heme-independent soluble guanylate cyclase activator BAY 58-2667 is devoid of non-cGMP actions, lacks tolerance development, and preferentially activates NO-insensitive heme-free or oxidized soluble guanylate cyclase. BAY 58-2667, therefore, represents a novel therapeutic advance in mediating vasodilation. To date, its cardiorenal actions in congestive heart failure (CHF) are undefined. We, therefore, hypothesized that BAY 58-2667 would have beneficial preload- and afterload-reducing actions in experimental severe CHF together with renal vasodilating properties. We assessed the cardiorenal actions of intravenous administration of 2 doses of BAY 58-2667 (0.1 and 0.3 microg/kg per minute, respectively) in a model of tachypacing-induced severe CHF. In CHF, BAY 58-2667 dose-dependently reduced mean arterial, right atrial, pulmonary artery, and pulmonary capillary wedge pressure (from baseline 19+/-1 to 12+/-2 mm Hg). Cardiac output (2.4+/-0.3 to 3.2+/-0.4 L/min) and renal blood flow increased. Glomerular filtration rate and sodium and water excretion were maintained. Consistent with cardiac unloading, atrial and B-type natriuretic peptide decreased. Plasma renin activity (P=0.31) and aldosterone remained unchanged (P=0.19). In summary, BAY 58-2667 in experimental CHF potently unloaded the heart, increased cardiac output and renal blood flow, and preserved glomerular filtration rate and sodium and water excretion without further neurohumoral activation. These beneficial properties make direct soluble guanylate cyclase stimulation with BAY 58-2667 a promising new therapeutic strategy for cardiovascular diseases, such as heart failure.
Asunto(s)
Benzoatos/farmacología , Guanilato Ciclasa/metabolismo , Insuficiencia Cardíaca/fisiopatología , Hemo/metabolismo , Receptores Citoplasmáticos y Nucleares/metabolismo , Animales , Factor Natriurético Atrial/antagonistas & inhibidores , Benzoatos/administración & dosificación , Vasos Sanguíneos/efectos de los fármacos , Sistema Cardiovascular/efectos de los fármacos , Sistema Cardiovascular/fisiopatología , Perros , Relación Dosis-Respuesta a Droga , Insuficiencia Cardíaca/metabolismo , Riñón/irrigación sanguínea , Riñón/efectos de los fármacos , Riñón/fisiopatología , Masculino , Péptido Natriurético Encefálico/antagonistas & inhibidores , Oxidación-Reducción , Guanilil Ciclasa Soluble , VasodilataciónRESUMEN
OBJECTIVE: To assess the effect of short-term treatment with GH on left ventricular contractility and remodeling, after the development of heart failure in cardiomyopathic hamsters (CMH). DESIGN: Two groups of 200-day-old UM-X7.1 CMH received daily subcutaneous injections of recombinant bovine (rb) GH (1mg/kg/day) or 0.9% NaCl for 40 days. Golden Syrian hamsters (GSH) were used as controls. At 240-day-old, the hamsters were randomly subjected to (i) assessment of left ventricular systolic function in a Langendorff perfused mode followed by the determination of the passive diastolic pressure-volume relationship and morphometric measurements; (ii) assessment of left ventricular mRNA expression of genes belonging to the fetal gene program including atrial (ANP) and brain (BNP) natriuretic peptides and cardiac myosin heavy chain isoforms and of the circulating levels of the natriuretic peptides. RESULTS: Hearts from CMH were hypertrophied and dilated (p<0.05) compared to hearts from GSH, along with a approximately 10-fold increase in the circulating ANP and BNP levels. Left ventricular BNP and ANP mRNAs were elevated by 2- and 3-fold, respectively, compared to GSH. rbGH reduced both ANP mRNA and ANP circulating levels by 34% (p<0.01) but did not significantly modulate BNP levels. This effect was associated with a preserved systolic function and reverse remodeling as assessed by a leftward shift of the passive diastolic pressure-volume relationship indicating reduced ventricular dilatation. CONCLUSIONS: The data show that a short-term administration of GH in the terminal phase of the disease confers cardioprotection by attenuating systolic dysfunction and by inducing beneficial reverse remodeling.
Asunto(s)
Factor Natriurético Atrial/antagonistas & inhibidores , Cardiomiopatías/tratamiento farmacológico , Cardiotónicos/uso terapéutico , Hormona del Crecimiento/uso terapéutico , Insuficiencia Cardíaca/tratamiento farmacológico , Función Ventricular Izquierda/efectos de los fármacos , Animales , Factor Natriurético Atrial/sangre , Factor Natriurético Atrial/metabolismo , Cardiomiopatías/complicaciones , Cardiomiopatías/genética , Cardiotónicos/farmacología , Bovinos , Cricetinae , Hormona del Crecimiento/farmacología , Insuficiencia Cardíaca/etiología , Ventrículos Cardíacos/química , Ventrículos Cardíacos/efectos de los fármacos , Ventrículos Cardíacos/metabolismo , Factor I del Crecimiento Similar a la Insulina/metabolismo , Mesocricetus , Contracción Miocárdica/efectos de los fármacos , Cadenas Pesadas de Miosina/metabolismo , Péptidos Natriuréticos/sangre , Péptidos Natriuréticos/metabolismo , Isoformas de Proteínas/metabolismoRESUMEN
BACKGROUND: Atrial natriuretic peptide and long acting natriuretic peptide have anticancer effects in human prostate adenocarcinomas. MATERIALS AND METHODS: Atrial natriuretic peptide, long acting natriuretic peptide and cyclic GMP's effects on MEK 1/2 kinase were examined in human prostate adenocarcinoma cells. RESULTS: Atrial natriuretic peptide and long acting natriuretic peptide decreased the activation of MEK 1/2 over a concentration range of 0.01 microM to 10 microM. Long acting natriuretic peptide and atrial natriuretic peptide (each 10 microM) inhibited the phosphorylation of MEK 1/2 kinase 97% (p < 0.00001) and 88% (p < 0.00001), respectively. The inhibition of MEK 1/2 was maximal at two hours, and ceased by four hours, secondary to both peptides. The ability of peptides to inhibit MEK 1/2 was inhibited by cyclic GMP antibody and cyclic GMP itself inhibited MEK 1/2 phosphorylation by 93%. CONCLUSION: Atrial natriuretic peptide and long acting natriuretic peptide both inhibit MEK 1/2 kinase mediated via cyclic GMP as part of their anticancer mechanism(s) of action.
Asunto(s)
Adenocarcinoma/tratamiento farmacológico , Factor Natriurético Atrial/farmacología , MAP Quinasa Quinasa 1/metabolismo , MAP Quinasa Quinasa 2/metabolismo , Neoplasias de la Próstata/tratamiento farmacológico , Adenocarcinoma/enzimología , Anciano , Animales , Anticuerpos/farmacología , Especificidad de Anticuerpos , Factor Natriurético Atrial/antagonistas & inhibidores , Línea Celular Tumoral , GMP Cíclico/inmunología , GMP Cíclico/metabolismo , Relación Dosis-Respuesta a Droga , Interacciones Farmacológicas , Activación Enzimática/efectos de los fármacos , Humanos , MAP Quinasa Quinasa 1/antagonistas & inhibidores , MAP Quinasa Quinasa 2/antagonistas & inhibidores , Masculino , Ratones , Ratones Desnudos , Fragmentos de Péptidos/antagonistas & inhibidores , Fragmentos de Péptidos/farmacología , Fosforilación/efectos de los fármacos , Neoplasias de la Próstata/enzimologíaRESUMEN
The vertebrate heart possesses autoregulatory mechanisms enabling it first to sense and then to adapt its force of contraction to continually changing demands. The molecular components of the cardiac mechanical stretch sensor are mostly unknown but of immense medical importance, since dysfunction of this sensing machinery is suspected to be responsible for a significant proportion of human heart failure. In the hearts of the ethylnitros-urea (ENU)-induced, recessive embryonic lethal zebrafish heart failure mutant main squeeze (msq), we find stretch-responsive genes such as atrial natriuretic factor (anf) and vascular endothelial growth factor (vegf) severely down-regulated. We demonstrate through positional cloning that heart failure in msq mutants is due to a mutation in the integrin-linked kinase (ilk) gene. ILK specifically localizes to costameres and sarcomeric Z-discs. The msq mutation (L308P) reduces ILK kinase activity and disrupts binding of ILK to the Z-disc adaptor protein beta-parvin (Affixin). Accordingly, in msq mutant embryos, heart failure can be suppressed by expression of ILK, and also of a constitutively active form of Protein Kinase B (PKB), and VEGF. Furthermore, antisense-mediated abrogation of zebrafish beta-parvin phenocopies the msq phenotype. Thus, we provide evidence that the heart uses the Integrin-ILK-beta-parvin network to sense mechanical stretch and respond with increased expression of ANF and VEGF, the latter of which was recently shown to augment cardiac force by increasing the heart's calcium transients.