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1.
PLoS One ; 9(4): e93473, 2014.
Artículo en Inglés | MEDLINE | ID: mdl-24695532

RESUMEN

BACKGROUND: Apelin, the endogenous ligand for the G protein-coupled apelin receptor, is an important regulator of the cardiovascular homoeostasis. We previously demonstrated that apelin is one of the most potent endogenous stimulators of cardiac contractility; however, its underlying signaling mechanisms remain largely elusive. In this study we characterized the contribution of protein kinase C (PKC), extracellular signal-regulated kinase 1/2 (ERK1/2) and myosin light chain kinase (MLCK) to the positive inotropic effect of apelin. METHODS AND RESULTS: In isolated perfused rat hearts, apelin increased contractility in association with activation of prosurvival kinases PKC and ERK1/2. Apelin induced a transient increase in the translocation of PKCε, but not PKCα, from the cytosol to the particulate fraction, and a sustained increase in the phosphorylation of ERK1/2 in the left ventricle. Suppression of ERK1/2 activation diminished the apelin-induced increase in contractility. Although pharmacological inhibition of PKC attenuated the inotropic response to apelin, it had no effect on ERK1/2 phosphorylation. Moreover, the apelin-induced positive inotropic effect was significantly decreased by inhibition of MLCK, a kinase that increases myofilament Ca2+ sensitivity. CONCLUSIONS: Apelin increases cardiac contractility through parallel and independent activation of PKCε and ERK1/2 signaling in the adult rat heart. Additionally MLCK activation represents a downstream mechanism in apelin signaling. Our data suggest that, in addition to their role in cytoprotection, modest activation of PKCε and ERK1/2 signaling improve contractile function, therefore these pathways represent attractive possible targets in the treatment of heart failure.


Asunto(s)
Péptidos y Proteínas de Señalización Intercelular/metabolismo , Sistema de Señalización de MAP Quinasas/fisiología , Contracción Muscular/fisiología , Contracción Miocárdica/fisiología , Proteína Quinasa C-epsilon/metabolismo , Animales , Apelina , Ventrículos Cardíacos/metabolismo , Ventrículos Cardíacos/fisiopatología , Masculino , Quinasa de Cadena Ligera de Miosina/metabolismo , Fosforilación/fisiología , Proteína Quinasa C-alfa/metabolismo , Ratas , Ratas Sprague-Dawley , Transducción de Señal/fisiología
2.
J Biol Chem ; 287(3): 1847-60, 2012 Jan 13.
Artículo en Inglés | MEDLINE | ID: mdl-22117073

RESUMEN

Obesity and insulin resistance are associated with chronic, low grade inflammation. Moreover, regulation of energy metabolism and immunity are highly integrated. We hypothesized that energy-sensitive coactivator peroxisome proliferator-activated receptor γ coactivator 1-α (PGC-1α) and AMP-activated protein kinase (AMPK) may modulate inflammatory gene expression in liver. Microarray analysis revealed that PGC-1α up-regulated expression of several cytokines and cytokine receptors, including interleukin 15 receptor α (IL15Rα) and, even more importantly, anti-inflammatory interleukin 1 receptor antagonist (IL1Rn). Overexpression of PGC-1α and induction of PGC-1α by fasting, physical exercise, glucagon, or cAMP was associated with increased IL1Rn mRNA and protein expression in hepatocytes. Knockdown of PGC-1α by siRNA down-regulated cAMP-induced expression of IL1Rn in mouse hepatocytes. Furthermore, knockdown of peroxisome proliferator-activated receptor α (PPARα) attenuated IL1Rn induction by PGC-1α. Overexpression of PGC-1α, at least partially through IL1Rn, suppressed interleukin 1ß-induced expression of acute phase proteins, C-reactive protein, and haptoglobin. Fasting and exercise also induced IL15Rα expression, whereas glucagon and cAMP resulted in reduction in IL15Rα mRNA levels. Finally, AMPK activator metformin and adenoviral overexpression of AMPK up-regulated IL1Rn and down-regulated IL15Rα in primary hepatocytes. We conclude that PGC-1α and AMPK alter inflammatory gene expression in liver and thus integrate energy homeostasis and inflammation. Induction of IL1Rn by PGC-1α and AMPK may be involved in the beneficial effects of exercise and caloric restriction and putative anti-inflammatory effects of metformin.


Asunto(s)
Proteínas Quinasas Activadas por AMP/metabolismo , Metabolismo Energético , Mediadores de Inflamación/metabolismo , Proteína Antagonista del Receptor de Interleucina 1/biosíntesis , Hígado/metabolismo , Proteínas de Unión al ARN/metabolismo , Transactivadores/metabolismo , Factores de Transcripción/metabolismo , Proteínas Quinasas Activadas por AMP/genética , Animales , Proteína C-Reactiva/biosíntesis , Proteína C-Reactiva/genética , Restricción Calórica , Células Cultivadas , Activadores de Enzimas/farmacología , Ayuno/metabolismo , Regulación de la Expresión Génica/efectos de los fármacos , Regulación de la Expresión Génica/genética , Técnicas de Silenciamiento del Gen , Haptoglobinas/biosíntesis , Haptoglobinas/genética , Hepatocitos/metabolismo , Hepatocitos/patología , Hipoglucemiantes/farmacología , Resistencia a la Insulina/genética , Proteína Antagonista del Receptor de Interleucina 1/genética , Hígado/patología , Masculino , Metformina/farmacología , Ratones , Ratones Endogámicos DBA , Obesidad/genética , Obesidad/metabolismo , Obesidad/terapia , PPAR alfa/genética , PPAR alfa/metabolismo , Coactivador 1-alfa del Receptor Activado por Proliferadores de Peroxisomas gamma , Condicionamiento Físico Animal , Proteínas de Unión al ARN/genética , Ratas , Receptores de Interleucina-15/biosíntesis , Receptores de Interleucina-15/genética , Transactivadores/genética , Factores de Transcripción/genética
3.
Biochem Pharmacol ; 82(12): 2008-15, 2011 Dec 15.
Artículo en Inglés | MEDLINE | ID: mdl-21933665

RESUMEN

The pregnane X receptor (PXR), a xenobiotic-sensing nuclear receptor plays a major role in regulation of drug metabolism but also modulates hepatic energy metabolism. PXR interacts with and represses several important transcription factors and coactivators regulating key enzymes in energy metabolism. Much less is known about how energy sensing cellular factors regulate PXR function. In this study we have investigated the effect of two major regulators of hepatic energy homeostasis, the transcriptional coactivator, peroxisome proliferator-activated receptor γ coactivator 1 alpha (PGC-1α) and the NAD-dependent deacetylase protein, sirtuin 1 (SIRT1) on PXR expression and function. Fasting induces PXR expression in liver. Furthermore, glucagon and PGC-1α overexpression upregulate PXR expression level in mouse primary hepatocytes suggesting that PGC-1α, in addition to coactivation of PXR, also transcriptionally regulates PXR gene. Knockdown of peroxisome proliferator-activated receptor α by siRNA attenuates PGC-1α mediated induction of PXR mRNA. PGC-1α overexpression alone has no effect on cytochrome P450 (CYP) 3A11 expression but potentiates induction by pregnenolone-16α-carbonitrile (PCN). Pyruvate, a nutrient signal activating SIRT1 abolishes synergistic induction of CYP3A11 by PCN and PGC-1α. Knockdown of SIRT1 prevented this effect of pyruvate. Downregulation of CYP7A1 by PCN was not affected by PGC-1α or pyruvate. Mammalian two hydrid assays indicate that pyruvate and SIRT1 interfere with interaction of PXR and PGC-1α. This may be mediated by well established PGC-1α deacetylation by SIRT1. However, we show by immunoprecipitation that SIRT1 also interacts with PXR. Thus we show that two fasting activated pathways PGC-1α and SIRT1 differentially modify PXR expression and function.


Asunto(s)
Receptores de Esteroides/metabolismo , Sirtuina 1/metabolismo , Factores de Transcripción/metabolismo , Animales , Células Cultivadas , Regulación de la Expresión Génica/fisiología , Hepatocitos/efectos de los fármacos , Hepatocitos/metabolismo , Humanos , Masculino , Ratones , Reacción en Cadena de la Polimerasa , Receptor X de Pregnano , Ácido Pirúvico/farmacología , ARN Mensajero/genética , ARN Mensajero/metabolismo , Ratas , Receptores de Esteroides/genética , Sirtuina 1/genética , Factores de Transcripción/genética
4.
J Physiol ; 589(Pt 11): 2669-86, 2011 Jun 01.
Artículo en Inglés | MEDLINE | ID: mdl-21486818

RESUMEN

Recent studies have demonstrated that changes in the activity of calcium-calmodulin-dependent protein kinase II (CaMKII) induce a unique cardiomyocyte phenotype through the regulation of specific genes involved in excitation-contraction (E-C)-coupling. To explain the transcriptional effects of CaMKII we identified a novel CaMKII-dependent pathway for controlling the expression of the pore-forming α-subunit (Cav1.2) of the L-type calcium channel (LTCC) in cardiac myocytes. We show that overexpression of either cytosolic (δC) or nuclear (δB) CaMKII isoforms selectively downregulate the expression of the Cav1.2. Pharmacological inhibition of CaMKII activity induced measurable changes in LTCC current density and subsequent changes in cardiomyocyte calcium signalling in less than 24 h. The effect of CaMKII on the α1C-subunit gene (Cacna1c) promoter was abolished by deletion of the downstream regulatory element (DRE), which binds transcriptional repressor DREAM/calsenilin/KChIP3. Imaging DREAM-GFP (green fluorescent protein)-expressing cardiomyocytes showed that CaMKII potentiates the calcium-induced nuclear translocation of DREAM. Thereby CaMKII increases DREAM binding to the DRE consensus sequence of the endogenous Cacna1c gene. By mathematical modelling we demonstrate that the LTCC downregulation through the Ca2+-CaMKII-DREAM cascade constitutes a physiological feedback mechanism enabling cardiomyocytes to adjust the calcium intrusion through LTCCs to the amount of intracellular calcium detected by CaMKII.


Asunto(s)
Transporte Activo de Núcleo Celular/fisiología , Canales de Calcio Tipo L/metabolismo , Proteína Quinasa Tipo 2 Dependiente de Calcio Calmodulina/metabolismo , Regulación de la Expresión Génica/fisiología , Proteínas de Interacción con los Canales Kv/metabolismo , Miocitos Cardíacos/metabolismo , Proteínas Represoras/metabolismo , Animales , Animales Recién Nacidos , Bencilaminas/farmacología , Sitios de Unión/genética , Canales de Calcio Tipo L/genética , Proteína Quinasa Tipo 2 Dependiente de Calcio Calmodulina/antagonistas & inhibidores , Proteína Quinasa Tipo 2 Dependiente de Calcio Calmodulina/genética , Línea Celular , Línea Celular Tumoral , Células Cultivadas , ADN/metabolismo , Regulación hacia Abajo/genética , Fenómenos Electrofisiológicos/fisiología , Acoplamiento Excitación-Contracción/fisiología , Retroalimentación Fisiológica/fisiología , Expresión Génica/efectos de los fármacos , Expresión Génica/genética , Proteínas de Interacción con los Canales Kv/genética , Ratones , Modelos Biológicos , Miocitos Cardíacos/efectos de los fármacos , Péptido Natriurético Encefálico/genética , Técnicas de Placa-Clamp , Mutación Puntual/genética , Regiones Promotoras Genéticas/genética , Ratas , Ratas Endogámicas , Proteínas Represoras/genética , Eliminación de Secuencia/genética , Sulfonamidas/farmacología , Transfección , Regulación hacia Arriba/genética
5.
J Mol Cell Cardiol ; 50(6): 1008-16, 2011 Jun.
Artículo en Inglés | MEDLINE | ID: mdl-21382378

RESUMEN

Sarcoplasmic reticulum (SR) Ca(2+)-ATPase (SERCA2a) is an essential component of cardiomyocyte excitation-contraction (EC)-coupling. Suppression of SERCA2a expression induces contractile dysfunction and has been reported in various forms of ischemic cardiac disease as well as in hypobaric hypoxia. The present study investigated whether SERCA2a expression is regulated by hypoxia in embryonic mouse cardiomyocytes and explored the underlying mechanism. We show that in cultured embryonic cardiomyocytes hypoxia (1% O(2)) induce time-dependent downregulation of SERCA2a expression. This mechanism manifested as specific changes in cardiac myocyte calcium signals induced by reduced expression and activity of SERCA2a. Chemical activation of hypoxia-inducible factor-1 (HIF-1) by DFO or overexpression of normoxia-stabile HIF-1α (HIF-1α/VP16) suppressed endogenous SERCA2a expression as well as the activity of the SERCA2a-promoter-luciferase reporter. Analysis of the SERCA2a promoter found two putative HIF-1 binding HRE-sites. Site-specific promoter mutagenesis revealed that co-operative HIF-1 binding to both of these hypoxia response elements on the SERCA2a promoter is required for expressional suppression. This mechanism establishes a link between oxygen supply and calcium activity in embryonic cardiac myocytes that is exploited in cardiac development, and further may offer a possible explanation for the functional depression of SERCA2a seen in ischemic and hypoxic myocardium.


Asunto(s)
Regulación del Desarrollo de la Expresión Génica , Factor 1 Inducible por Hipoxia/metabolismo , Miocitos Cardíacos/enzimología , Elementos de Respuesta/fisiología , ATPasas Transportadoras de Calcio del Retículo Sarcoplásmico/metabolismo , Animales , Secuencia de Bases , Sitios de Unión/genética , Calcio/metabolismo , Hipoxia de la Célula , Células Cultivadas , Citosol/metabolismo , Orden Génico , Proteína Vmw65 de Virus del Herpes Simple/genética , Proteína Vmw65 de Virus del Herpes Simple/metabolismo , Factor 1 Inducible por Hipoxia/genética , Ratones , Modelos Biológicos , Oxígeno/metabolismo , Regiones Promotoras Genéticas/genética , Transporte de Proteínas , Elementos de Respuesta/genética , ATPasas Transportadoras de Calcio del Retículo Sarcoplásmico/genética , Homología de Secuencia de Ácido Nucleico
6.
J Mol Cell Cardiol ; 50(5): 884-93, 2011 May.
Artículo en Inglés | MEDLINE | ID: mdl-21320508

RESUMEN

Increased production of reactive oxygen species (ROS) has been linked to the pathogenesis of contractile dysfunction in heart failure. However, it is unclear whether ROS can regulate physiological cellular processes in the myocardium. Here, we characterized the role of endogenous ROS production in the acute regulation of cardiac contractility in the intact rat heart. In isolated perfused rat hearts, endothelin-1 (ET-1, 1nmol/L) stimulated ROS formation in the left ventricle, which was prevented by the antioxidant N-acetylcysteine and the NAD(P)H oxidase inhibitor apocynin. N-acetylcysteine, the superoxide dismutase mimetic MnTMPyP, and apocynin significantly attenuated ET-1-mediated inotropic effect, which was accompanied by inhibition of extracellular signal regulated kinase 1/2 (ERK1/2) phosphorylation. Moreover, the mitochondrial K(ATP) channel blocker 5-HD, and the mitochondrial large conductance calcium activated potassium channel blocker paxilline, but not the sarcolemmal K(ATP) channel blocker HMR 1098 attenuated the inotropic response to ET-1. However, ET-1-induced ROS generation was not abolished by inhibiting mitochondrial K(ATP) channel opening. In contrast to ET-1 stimulation, the positive inotropic effect of ß(1)-adrenergic receptor agonist dobutamine (250nmol/L) was significantly augmented by N-acetylcysteine and apocynin. Moreover, dobutamine-induced phospholamban phosphorylation was markedly enhanced by apocynin. In conclusion, NAD(P)H oxidase-derived ROS play a physiological role in the acute regulation of cardiac contractility in the intact rat heart. Our results reveal that ET-1-induced increase in cardiac contractility is partially dependent on enhanced ROS generation, which in turn, activates the ERK1/2 pathway. On the other hand, ß-adrenergic receptor-induced positive inotropic effect and phospholamban phosphorylation is enhanced by NAD(P)H oxidase inhibition.


Asunto(s)
Especies Reactivas de Oxígeno/metabolismo , Animales , Western Blotting , Proteínas de Unión al Calcio/metabolismo , Endotelina-1/farmacología , Etidio/análogos & derivados , Etidio/farmacología , Técnicas In Vitro , Masculino , Proteína Quinasa 1 Activada por Mitógenos/metabolismo , Proteína Quinasa 3 Activada por Mitógenos/metabolismo , Contracción Miocárdica/efectos de los fármacos , Fosforilación/efectos de los fármacos , Ratas , Ratas Sprague-Dawley
7.
J Mol Cell Cardiol ; 50(4): 634-41, 2011 Apr.
Artículo en Inglés | MEDLINE | ID: mdl-21223972

RESUMEN

Parthenolide has shown promise in treatment of various cancers via inhibition of the transcription factor signal transducer and activator of transcription 3 (STAT3). Activation of STAT3 has been observed in left ventricular hypertrophy (LVH); however, its exact role is not known. The aim of the study was to examine the effects of parthenolide on pressure overload-induced LVH in rats. Pressure overload was induced by angiotensin II (Ang II) infusion (33 µg/kg/h) for 1 week in the presence or absence of parthenolide (0.5mg/kg/day, i.p.). Ang II infusion resulted in LVH associated with increased phosphorylation of STAT3 at Tyr705 and Ser727. Parthenolide treatment had no effect on ejection fraction, but abolished the activation of STAT3 and reduced the Ang II-induced LVH (LV posterior wall thickness in end-diastole: 2.28 ± 0.12 mm vs. 1.80 ± 0.06 mm, P<0.001). Importantly, parthenolide treatment had no effect on heart rate or blood pressure. Parthenolide treatment almost completely abolished the Ang II-induced increase in the number of cells positive for prolyl-4-hydroxylase, a marker for collagen-synthesizing cells, as well as Ang II-induced interstitial fibrosis in the left ventricles. This was associated with significant attenuation of Ang II-induced increase in mRNA levels of type 1 collagen and fibronectin. Moreover, parthenolide attenuated the Ang II-induced expression of interleukin-6, a potent pro-hypertrophic fibroblast-derived factor. We conclude that pharmacological inhibition of STAT3 signaling by parthenolide has favorable effects on pressure overload-induced LVH through attenuation of fibroblast activation. Therefore parthenolide may prove as a useful therapy for certain cardiovascular disease.


Asunto(s)
Angiotensina II/uso terapéutico , Fibroblastos/efectos de los fármacos , Fibroblastos/metabolismo , Hipertrofia Ventricular Izquierda/inducido químicamente , Hipertrofia Ventricular Izquierda/tratamiento farmacológico , Factor de Transcripción STAT3/metabolismo , Sesquiterpenos/farmacología , Animales , Western Blotting , Ecocardiografía , Ensayo de Cambio de Movilidad Electroforética , Hemodinámica/efectos de los fármacos , Hipertrofia Ventricular Izquierda/metabolismo , Inmunohistoquímica , Masculino , Fosforilación/efectos de los fármacos , Ratas , Ratas Sprague-Dawley , Transducción de Señal/efectos de los fármacos
8.
Analyst ; 136(4): 835-40, 2011 Feb 21.
Artículo en Inglés | MEDLINE | ID: mdl-21132163

RESUMEN

In vivo skin analysis by Desorption Electrospray Ionization was characterized on healthy human volunteers by directing pneumatically assisted electrospray directly onto their fingertips. In order to eliminate the risk of electric shock, a high ohmic resistor was built into the system. Positive ion DESI-MS analysis yields low intensity spectra, while negative ion spectra feature a number of various biogenic carboxylic acids. Compounds of external origin and excreted molecules were found to have different analysis kinetics, with the exception of highly hydrophobic species. The difference was demonstrated in the case of nicotine and cotinine. Pharmacokinetic studies were performed using a rat animal model. The kinetics of the anesthetic ketamine was followed by DESI, and results were in agreement with off-line HPLC-MS blood analysis. Using a similar approach for N,N'-dimethylthiourea (DMTU), a novel method was developed for the real-time quantification of oxidative stress. DMTU was administered to the animals, and the ratio of the molecule and its oxidized form was monitored from the skin surface. The ratio was found to be highly sensitive to experimentally induced diabetes mellitus type I and angiotensin-induced chronic oxidative stress. It was concluded that the method has a number of potential applications in the fields of forensics, pharmacology and clinical chemistry.


Asunto(s)
Piel/química , Espectrometría de Masa por Ionización de Electrospray/métodos , Animales , Humanos , Cinética , Masculino , Estrés Oxidativo , Ratas , Ratas Wistar , Piel/metabolismo , Detección de Abuso de Sustancias
9.
Blood Press ; 19(1): 54-63, 2010 Feb.
Artículo en Inglés | MEDLINE | ID: mdl-20175653

RESUMEN

Dyxin is a novel LIM domain protein acting as a transcriptional cofactor with GATA transcription factors. Here, we characterized dyxin as a p38 mitogen-activated protein kinase (MAPK) regulated gene, since combined upstream MAPK kinase 3b and wild-type p38 alpha MAPK gene transfer increased left ventricular dyxin mRNA and protein levels in vivo. We also studied cardiac dyxin expression in experimental models of pressure overload and myocardial infarction (MI) in vivo. Angiotensin II infusion increased left ventricular dyxin mRNA levels (9.4-fold, p<0.001) rapidly at 6 h followed by induction of protein levels. Furthermore, simultaneous administration of p38 MAPK inhibitor SB203580 abolished angiotensin II-induced activation of dyxin gene expression. During the post-infarction remodeling process, increased dyxin mRNA levels (7.7-fold, p<0.01) were noted at day 1 followed by the increase in proteins levels at 2 weeks after MI (1.5-fold, p<0.05). Moreover, direct wall stretch by using isolated rat heart preparation as well as direct mechanical stretch of cardiomyocytes in vitro activated dyxin gene expression within 1 h. Our results indicate that dyxin expression is rapidly upregulated in response to mechanical load, this increase being at least partly mediated by p38 MAPK. These results suggest that dyxin may play an important role in regulating hypertrophic process.


Asunto(s)
Proteínas Portadoras/metabolismo , Corazón/fisiopatología , Miocardio/metabolismo , Proteínas Quinasas p38 Activadas por Mitógenos/metabolismo , Adenoviridae/genética , Angiotensina II , Animales , Proteínas Portadoras/genética , Proteínas Co-Represoras , Técnicas de Transferencia de Gen , Vectores Genéticos , Ventrículos Cardíacos/metabolismo , Hipertensión/inducido químicamente , Hipertensión/fisiopatología , Técnicas In Vitro , Proteínas con Dominio LIM , Masculino , Infarto del Miocardio/fisiopatología , Miocitos Cardíacos/metabolismo , ARN Mensajero/metabolismo , Ratas , Ratas Sprague-Dawley , Estrés Mecánico , Vasoconstrictores , Remodelación Ventricular
10.
Regul Pept ; 159(1-3): 9-13, 2010 Jan 08.
Artículo en Inglés | MEDLINE | ID: mdl-19616582

RESUMEN

High levels of specific prolactin-releasing peptide (PrRP) binding sites have been found in the myocardium; however, the functional importance of PrRP in the regulation of cardiac function is unknown. In isolated perfused rat hearts, infusion of PrRP (1-100 nM) induced a dose-dependent positive inotropic effect. Inhibition of cAMP catabolism by IBMX, a phosphodiesterase inhibitor, failed to augment the contractile effect of PrRP. The protein phosphatase (PP1/PP2A) inhibitor calyculin A increased the inotropic response to PrRP, whereas the PP2A inhibitor okadaic acid had no effect. Ro32-0432, a protein kinase C alpha (PKC alpha) inhibitor, significantly enhanced the inotropic effect of PrRP as well as the phosphorylation of phospholamban at Ser-16. In conclusion, the present data define a hitherto unrecognized role for PrRP in the regulation of cardiovascular system by showing that PrRP exerts a direct positive inotropic effect. Moreover, our results suggest that the cAMP-independent inotropic response to PrRP is suppressed by concurrent activation of PKC alpha and PP1.


Asunto(s)
Cardiotónicos/farmacología , Contracción Miocárdica/efectos de los fármacos , Miocardio/metabolismo , Hormona Liberadora de Prolactina/farmacología , Animales , Relación Dosis-Respuesta a Droga , Inhibidores Enzimáticos/farmacología , Masculino , Proteína Quinasa C-alfa/antagonistas & inhibidores , Proteína Quinasa C-alfa/metabolismo , Proteína Fosfatasa 1/antagonistas & inhibidores , Proteína Fosfatasa 1/metabolismo , Proteína Fosfatasa 2/antagonistas & inhibidores , Proteína Fosfatasa 2/metabolismo , Ratas , Ratas Sprague-Dawley
12.
Circulation ; 118(16): 1651-8, 2008 Oct 14.
Artículo en Inglés | MEDLINE | ID: mdl-18824646

RESUMEN

BACKGROUND: Extracellular signal-regulated kinase 1/2 (ERK1/2) and p38 mitogen-activated protein kinase (p38-MAPK) have been shown to regulate various cellular processes, including cell growth, proliferation, and apoptosis in the heart. However, the function of these signaling pathways in the control of cardiac contractility is unclear. Here, we characterized the contribution of ERK1/2 and p38-MAPK to the inotropic effect of endothelin-1 (ET-1). METHODS AND RESULTS: In isolated perfused rat hearts, infusion of ET-1 (1 nmol/L) for 10 minutes increased contractility and phosphorylation of ERK1/2 and their downstream target p90 ribosomal S6 kinase (p90RSK). Suppression of ERK1/2 activation prevented p90RSK phosphorylation and attenuated the inotropic effect of ET-1. Pharmacological inhibition of epidermal growth factor receptor kinase activity abolished ET-1-induced epidermal growth factor receptor transactivation and ERK1/2 and p90RSK phosphorylation and reduced ET-1-mediated inotropic response. Moreover, inhibition of the p90RSK target Na(+)-H(+) exchanger 1 attenuated the inotropic effect of ET-1. In contrast to ERK1/2 signaling, suppression of p38-MAPK activity further augmented ET-1-enhanced contractility, which was accompanied by increased phosphorylation of phospholamban at Ser-16. CONCLUSIONS: MAPKs play opposing roles in the regulation of cardiac contractility in that the ERK1/2-mediated positive inotropic response to ET-1 is counterbalanced by simultaneous activation of p38-MAPK. Hence, selective activation of ERK1/2 signaling and inhibition of p38-MAPK signaling may represent novel means to support cardiac function in disease.


Asunto(s)
Proteína Quinasa 1 Activada por Mitógenos/metabolismo , Proteína Quinasa 3 Activada por Mitógenos/metabolismo , Contracción Miocárdica/fisiología , Miocardio/enzimología , Proteínas Quinasas p38 Activadas por Mitógenos/metabolismo , Animales , Endotelina-1/farmacología , Sistema de Señalización de MAP Quinasas/efectos de los fármacos , Sistema de Señalización de MAP Quinasas/fisiología , Masculino , Contracción Miocárdica/efectos de los fármacos , Proteína Quinasa C/metabolismo , Ratas , Ratas Sprague-Dawley , Fosfolipasas de Tipo C/metabolismo
13.
J Hypertens ; 25(9): 1927-39, 2007 Sep.
Artículo en Inglés | MEDLINE | ID: mdl-17762659

RESUMEN

OBJECTIVE: The transcription factor nuclear factor-kappaB (NF-kappaB) has been implicated in cardiomyocyte hypertrophy in vitro as well as in vivo; however, it is unknown if activation of NF-kappaB plays a mandatory role in the hypertrophic process. Here we characterize the importance of NF-kappaB signaling in moderate and severe left ventricular (LV) hypertrophy in rats with chronic pressure overload induced by angiotensin II (Ang II) infusion. METHODS AND RESULTS: Electrophoretic mobility shift assay analysis revealed that Ang II infusion (2.5 microg/kg per min) for 6 days increased LV NF-kappaB/DNA-binding activity in a biphasic manner in Sprague-Dawley rats. Pyrrolidine dithiocarbamate (PDTC) (100 mg/kg per day), an NF-kappaB inhibitor, abolished Ang II-induced NF-kappaB activation and concomitant increase in tumor necrosis factor-alpha gene expression, while activator protein-1/DNA binding was not affected. Inhibition of NF-kappaB signaling for 6 days significantly attenuated Ang II-induced increases in LV/body weight ratio, LV mean wall thickness and cardiomyocyte cross-sectional area, without compromising LV systolic function. Moreover, PDTC abolished Ang II-induced cardiomyocyte apoptosis and interstitial fibrosis, and attenuated the gene expression of type I collagen. In contrast, a moderate LV hypertrophy induced by Ang II at a lower dose (0.5 microg/kg per min) was not associated with a significant activation of NF-kappaB, and PDTC treatment had no effect on the hypertrophic indices. CONCLUSION: Our in-vivo data indicate a critical role of NF-kappaB signaling in the advanced stage of the remodeling process, whereas development of moderate LV hypertrophy is not dependent on NF-kappaB activation.


Asunto(s)
Angiotensina II/farmacología , Ventrículos Cardíacos/efectos de los fármacos , FN-kappa B/metabolismo , Transducción de Señal , Angiotensina II/administración & dosificación , Animales , Secuencia de Bases , Cartilla de ADN , Ecocardiografía , Ensayo de Cambio de Movilidad Electroforética , Masculino , Ratas , Ratas Sprague-Dawley
14.
Paediatr Anaesth ; 17(8): 782-8, 2007 Aug.
Artículo en Inglés | MEDLINE | ID: mdl-17596223

RESUMEN

BACKGROUND: Natriuretic peptide levels are associated with cardiac output and ventricular function. We hypothesized that concomitant measurement of the peptide fragments and the hemodynamic parameters could elucidate the associations of these parameters after pediatric cardiac surgery. METHODS: After approval of the institutional review board and parents' informed consent, we investigated the clinical data of eight neonates undergoing correction of transposition of the great arteries. We measured the level of N-terminal fragments of prohormones of atrial and brain natriuretic peptides (NT-proANP, NT-proBNP) preoperatively, postoperatively and 12, 24, 48, and 72 h after arrival in the intensive care unit. The hemodynamic status was assessed by transpulmonary thermodilution at the same time points. Creatinine and other laboratory values were analyzed in the first 48 h postoperatively. RESULTS: NT-proBNP levels were inversely correlated with cardiac index (CI, r = -0.47, P = 0.030), stroke volume index (r = -0.65, P = 0.005), and global end-diastolic volume index (GEDI; r = -0.63, P = 0.011). There was strong inverse correlation between the change of NT-proBNP levels and the change of CI between two consecutive measurements during the postoperative period (r = -0.79, P = 0.001). The NT-proBNP level 12 h after surgery was strongly correlated with the creatinine level of the postoperative 24th hour (r = 0.81, P = 0.014). CONCLUSIONS: NT-proBNP correlated with the hemodynamic parameters and with the severity of renal dysfunction. Therefore, NT-proBNP is a reliable indicator of the circulatory state and the severity of a low output syndrome after arterial switch operation in neonates.


Asunto(s)
Gasto Cardíaco , Péptido Natriurético Encefálico/sangre , Fragmentos de Péptidos/sangre , Transposición de los Grandes Vasos/cirugía , Humanos , Recién Nacido , Modelos Lineales , Periodo Posoperatorio , Termodilución , Transposición de los Grandes Vasos/sangre
15.
Life Sci ; 80(14): 1303-10, 2007 Mar 13.
Artículo en Inglés | MEDLINE | ID: mdl-17266992

RESUMEN

The Na(+)/K(+)-ATPase inhibitor ouabain has been shown to trigger hypertrophic growth of cultured cardiomyocytes; however, the significance of endogenous ouabain-like compound (OLC) in the hypertrophic process in vivo is unknown. Here we characterized the involvement of OLC in left ventricular (LV) hypertrophy induced by norepinephrine (NE) and angiotensin II (Ang II) infusions in rats. Administration of NE (300 microg/kg/h) via subcutanously implanted osmotic minipumps for 72 h resulted in a significant increase in left ventricular weight to body weight (LVW/BW) ratio (P<0.001) and a substantial up-regulation of atrial natriuretic peptide (ANP) gene expression (13.2-fold, P<0.001). NE infusion induced a transient increase in plasma OLC levels at 12 h (P<0.05), which returned to control levels by 72 h. Adrenalectomy markedly reduced both basal and NE-induced increase in plasma OLC levels. LVW/BW ratio was not modulated by adrenalectomy; however, ANP gene expression was blunted by 44% (P<0.01) and 47% (P<0.05) at 12 and 72 h, respectively. In agreement, adrenalectomy reduced up-regulation of ANP without affecting LV mass in rats infused with Ang II (33 microg/kg/h). Administration of exogenous ouabain (1 nM to 100 microM) for 24 h had no effect on ANP gene expression in cultured neonatal rat ventricular myocytes. However, the up-regulation of ANP mRNA levels induced by the alpha-adrenergic agonist phenylephrine (1 microM) was markedly enhanced by ouabain (100 microM) (5.6-fold vs. 9.6-fold, P<0.01). These data show that OLC as an adrenal-derived factor may be required for the induction LV ANP gene expression during the hypertrophic process.


Asunto(s)
Cardenólidos/sangre , Hipertrofia Ventricular Izquierda/sangre , Saponinas/sangre , Adrenalectomía , Angiotensina II/farmacología , Animales , Factor Natriurético Atrial/genética , Northern Blotting , Modelos Animales de Enfermedad , Relación Dosis-Respuesta a Droga , Expresión Génica/efectos de los fármacos , Hipertrofia Ventricular Izquierda/inducido químicamente , Hipertrofia Ventricular Izquierda/genética , Masculino , Miocitos Cardíacos/efectos de los fármacos , Miocitos Cardíacos/metabolismo , Norepinefrina/farmacología , Tamaño de los Órganos/efectos de los fármacos , ARN Mensajero/metabolismo , Ratas , Ratas Sprague-Dawley , Regulación hacia Arriba/efectos de los fármacos , Vasoconstrictores/farmacología
16.
Cardiovasc Res ; 73(3): 512-20, 2007 Feb 01.
Artículo en Inglés | MEDLINE | ID: mdl-17182020

RESUMEN

OBJECTIVE: In diabetes mellitus several cardiac electrophysiological parameters are known to be affected. In rodent experimental diabetes models changes in these parameters were reported, but no such data are available in other mammalian species including the dog. The present study was designed to analyse the effects of experimental type 1 diabetes on ventricular repolarization and its underlying transmembrane ionic currents and channel proteins in canine hearts. METHODS AND RESULTS: Diabetes was induced by a single injection of alloxan, a subgroup of dogs received insulin substitution. After the development of diabetes (8 weeks) electrophysiological studies were performed using conventional microelectrodes, whole cell voltage clamp, and ECG. Expression of ion channel proteins was evaluated by Western blotting. The QTc interval and the ventricular action potential duration in diabetic dogs were moderately prolonged. This was accompanied by significant reduction in the density of the transient outward K+ current (I(to)) and the slow delayed rectifier K+ current (I(Ks)), to 54.6% and 69.3% of control, respectively. No differences were observed in the density of the inward rectifier K+ current (I(K1)), rapid delayed rectifier K+ current (I(Kr)), and L-type Ca2+ current (I(Ca)). Western blot analysis revealed a reduced expression of Kv4.3 and MinK (to 25+/-21% and 48+/-15% of control, respectively) in diabetic dogs, while other channel proteins were unchanged (HERG, MiRP1, alpha(1c)) or increased (Kv1.4, KChIP2, KvLQT1). Insulin substitution fully prevented the diabetes-induced changes in I(Ks), KvLQT1 and MinK, however, the changes in I(to), Kv4.3, and Kv1.4 were only partially diminished by insulin. CONCLUSION: It is concluded that type 1 diabetes mellitus, although only moderately, lengthens ventricular repolarization, attenuates the repolarization reserve by decreasing I(to) and I(Ks) currents, and thereby may markedly enhance the risk of sudden cardiac death.


Asunto(s)
Diabetes Mellitus Tipo 1/metabolismo , Diabetes Mellitus Tipo 1/fisiopatología , Miocardio/metabolismo , Potenciales de Acción , Animales , Western Blotting/métodos , Canales de Calcio Tipo L/metabolismo , Canales de Potasio de Tipo Rectificador Tardío/metabolismo , Diabetes Mellitus Experimental , Diabetes Mellitus Tipo 1/tratamiento farmacológico , Perros , Electrocardiografía , Femenino , Insulina/uso terapéutico , Canal de Potasio KCNQ1/metabolismo , Canal de Potasio Kv1.4/metabolismo , Masculino , Técnicas de Placa-Clamp , Canales de Potasio de Rectificación Interna/metabolismo , Canales de Potasio Shal/metabolismo
17.
J Lipid Res ; 47(6): 1219-26, 2006 Jun.
Artículo en Inglés | MEDLINE | ID: mdl-16569909

RESUMEN

Long-term dietary fatty acid intake alters the development of left ventricular hypertrophy, but the linking signaling pathways are unclear. We studied the role and underlying signaling mechanisms of dietary fat intake in the early phase of the hypertrophic process. Rats assigned for 4 weeks of high-oil, high-fat, or standard diet were subjected to angiotensin II (Ang II; 33 microg/kg/h, subcutaneous) or vehicle infusion for 24 h. The Ang II-induced increase in left ventricular mRNA levels of hypertrophy-associated genes was higher in rats fed the high-oil diet compared with the standard diet. Western blotting revealed that, in parallel with changes in gene expression, the high-oil diet increased c-Jun N-terminal kinase phosphorylation (P < 0.001). Ang II increased p38 mitogen-activated protein kinase (MAPK) phosphorylation in rats fed the high-fat diet (3-fold; P < 0.01). The increase in transcription factor activator protein-1 (AP-1) DNA binding activity in response to Ang II was higher in rats fed the high-oil diet compared with those fed the standard diet (P < 0.001). Ang II downregulated inducible nitric oxide synthase mRNA levels in fatty acid-supplemented groups compared with the standard diet group. These results show that dietary fat type modulates the early activation of hypertrophic genes in pressure-overloaded myocardium involving the distinct activation of AP-1 and MAPK signal transduction pathways.


Asunto(s)
Angiotensina II/farmacología , Grasas de la Dieta/farmacología , Hipertrofia Ventricular Izquierda/genética , Animales , Northern Blotting , Western Blotting , Grasas de la Dieta/administración & dosificación , Ensayo de Cambio de Movilidad Electroforética , Expresión Génica/efectos de los fármacos , Ventrículos Cardíacos/efectos de los fármacos , Ventrículos Cardíacos/metabolismo , Ventrículos Cardíacos/patología , Hipertrofia Ventricular Izquierda/sangre , Hipertrofia Ventricular Izquierda/inducido químicamente , Proteínas Quinasas JNK Activadas por Mitógenos/metabolismo , Lípidos/sangre , Masculino , Proteínas Quinasas Activadas por Mitógenos/metabolismo , Modelos Biológicos , Óxido Nítrico Sintasa de Tipo II/genética , Óxido Nítrico Sintasa de Tipo III/genética , Óxido Nítrico Sintasa de Tipo III/metabolismo , ARN Mensajero/genética , ARN Mensajero/metabolismo , Distribución Aleatoria , Ratas , Ratas Sprague-Dawley , Receptor de Angiotensina Tipo 1/genética
18.
Clin Chem ; 51(4): 708-18, 2005 Apr.
Artículo en Inglés | MEDLINE | ID: mdl-15718490

RESUMEN

BACKGROUND: High circulating concentrations of N-terminal fragments of A- and B-type natriuretic peptides (NT-proANP and NT-proBNP) identify patients with impaired cardiac function. ProANP-derived peptides are particularly sensitive to increased preload of the heart and proBNP-derived peptides to increased afterload; therefore, combining the information from the ANP and BNP systems into a single analyte could produce an assay with increased diagnostic and prognostic power. METHODS: We prepared a hybrid peptide containing peptide sequences from both NT-proBNP and NT-proANP (referred to as NT-proXNP) by recombinant techniques and used it to develop a RIA combining weighed concentrations of NT-proANP and NT-proBNP into a new virtual analyte, NT-proXNP. We used the novel method to measure the circulating concentrations in healthy persons and in patients with cardiac disorders. We also characterized the assay by HPLC analysis of the immunoreactive molecular forms in human plasma and serum. RESULTS: The results from the novel assay correlated well with independent home-made NT-proANP and NT-proBNP assays (r2 = 0.75-0.85) as well with the arithmetic sum of NT-proANP and NT-proBNP (r2 = 0.92). Patients with valvular heart disease (VHD) and coronary artery disease (CAD) had significantly increased NT-proXNP concentrations. The areas under the curve (AUC) of the NT-proXNP assay in detecting VHD and CAD (0.961 and 0.924, respectively) were significantly larger than the AUC of either NT-proANP (0.947 and 0.872) or NT-proBNP (0.913 and 0.782) assays. HPLC analysis showed that the novel NT-proXNP assay detects two major classes of circulating immunoreactivity corresponding to peptides derived from NT-proANP and NT-proBNP. CONCLUSIONS: Our novel immunoassay mimics the physiologic signaling system working in the body by converging the information obtained from the activation of ANP and BNP into a single virtual analyte, NT-proXNP. It appears to have a diagnostic efficiency equal to or slightly better than that of individual NT-proANP or NT-proBNP assays.


Asunto(s)
Factor Natriurético Atrial/sangre , Péptido Natriurético Encefálico/sangre , Proteínas del Tejido Nervioso/sangre , Fragmentos de Péptidos/sangre , Precursores de Proteínas/sangre , Proteínas Recombinantes de Fusión/química , Adulto , Anciano , Factor Natriurético Atrial/genética , Cromatografía Líquida de Alta Presión , Enfermedad Coronaria/diagnóstico , Enfermedades de las Válvulas Cardíacas/diagnóstico , Humanos , Sueros Inmunes , Radioisótopos de Yodo , Persona de Mediana Edad , Péptido Natriurético Encefálico/genética , Proteínas del Tejido Nervioso/genética , Fragmentos de Péptidos/genética , Precursores de Proteínas/genética , Radioinmunoensayo , Proteínas Recombinantes de Fusión/inmunología , Proteínas Recombinantes de Fusión/aislamiento & purificación , Valores de Referencia
19.
J Cardiovasc Pharmacol ; 44 Suppl 1: S195-7, 2004 Nov.
Artículo en Inglés | MEDLINE | ID: mdl-15838277

RESUMEN

Diabetes mellitus is associated with endothelial and cardiac dysfunction, and endothelin has been suggested to alter cardiac function by being a positive inotropic agent, modulating the Frank-Starling response, contracting the coronary arteries and inducing tissue proliferation. We investigated endothelin levels in diabetic and in healthy dogs, 1 and 3 days after placing arteriovenous shunts (8 weeks after diabetes induction) in the femoral regions. Right and left ventricular weight/body weight ratios and Nterminal- atrial natriuretic peptide were increased in shunted animals (P < 0.05). Plasma endothelin levels were comparable in healthy and diabetic dogs. Shunted circulation did not change systemic endothelin levels in healthy dogs but reduced endothelin levels in diabetic dogs. The functional significance of altered endothelin responses to acute hemodynamic burden in experimental diabetes needs further investigation.


Asunto(s)
Cardiomegalia/metabolismo , Diabetes Mellitus Experimental/metabolismo , Endotelina-1/sangre , Animales , Derivación Arteriovenosa Quirúrgica , Factor Natriurético Atrial/sangre , Presión Sanguínea , Cardiomegalia/complicaciones , Cardiomegalia/patología , Cardiomegalia/fisiopatología , Diabetes Mellitus Experimental/complicaciones , Diabetes Mellitus Experimental/patología , Diabetes Mellitus Experimental/fisiopatología , Modelos Animales de Enfermedad , Perros , Femenino , Masculino , Precursores de Proteínas/sangre , Factores de Tiempo
20.
J Cardiovasc Pharmacol ; 44 Suppl 1: S380-2, 2004 Nov.
Artículo en Inglés | MEDLINE | ID: mdl-15838325

RESUMEN

Endothelin-1 (ET-1) is known to have a direct arrhythmogenic effect in the mammalian heart. Diabetes mellitus is accompanied by a series of endothelial and cardiac disfunctions; however, little is known about ET-1-induced direct arrhythmias in diabetes mellitus. Therefore, we infused ET-1 (33 pmol/min) into the left anterior descending coronary artery of 28 mongrel dogs, and measured basic hemodynamic parameters, coronary flow and an electrocardiogram. Diabetes mellitus was induced by alloxan (Group 4) and experiments were carried out 8 weeks later. Metabolically healthy dogs served as controls (Group 2). In a further control group, local hyperglycemia was induced by intracoronary glucose infusion (Group 3). ET-1 infusion induced prolongation of the QT-time and frequency-adjusted QT-time in all groups. Other electrophysiological parameters were comparable between the groups. This was followed by the occurence of ventricular premature beats, coupled extra-beats and later sustained ventricular tachycardia. Most of the experiments were terminated by ventricular fibrillation. The onset of arrhythmias was shorter in diabetic dogs as compared with control and locally hyperglycemic animals (18 +/- 8 minutes versus 24 +/- 8 minutes and 30 +/- 28 minutes, P < 0.05). However, there was no difference in the number of ventricular fibrillations, and the total elapsed time until the termination of the experiments. Therefore, the diabetic heart seems to be more prone to ET-1-induced arrhythmias and this is probably not a result of locally high glucose concentrations.


Asunto(s)
Diabetes Mellitus Experimental/complicaciones , Hemodinámica , Taquicardia Ventricular/etiología , Fibrilación Ventricular/etiología , Complejos Prematuros Ventriculares/etiología , Animales , Presión Sanguínea , Circulación Coronaria , Diabetes Mellitus Experimental/fisiopatología , Perros , Electrocardiografía , Endotelina-1/administración & dosificación , Glucosa/administración & dosificación , Frecuencia Cardíaca , Infusiones Parenterales , Factores de Riesgo , Taquicardia Ventricular/inducido químicamente , Taquicardia Ventricular/fisiopatología , Factores de Tiempo , Fibrilación Ventricular/inducido químicamente , Fibrilación Ventricular/fisiopatología , Complejos Prematuros Ventriculares/inducido químicamente , Complejos Prematuros Ventriculares/fisiopatología
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