RESUMEN
BACKGROUND: Despite advances in treatment, myocardial infarction (MI) is a leading cause of heart failure and death worldwide, with both ischemia and reperfusion (I/R) causing cardiac injury. A previous study using a mouse model of nonreperfused MI showed activation of brown adipose tissue (BAT). Recent studies showed that molecules secreted by BAT target the heart. We investigated whether BAT attenuates cardiac injury in I/R and sought to identify potential cardioprotective proteins secreted by BAT. METHODS: Myocardial I/R surgery with or without BAT transplantation was performed in wild-type (WT) mice and in mice with impaired BAT function (uncoupling protein 1 [Ucp1]-deficient mice). To identify potential cardioprotective factors produced by BAT, RNA-seq (RNA sequencing) was performed in BAT from WT and Ucp1-/- mice. Subsequently, myocardial I/R surgery with or without BAT transplantation was performed in Bmp3b (bone morphogenetic protein 3b)-deficient mice, and WT mice subjected to myocardial I/R were treated using BMP3b. RESULTS: Dysfunction of BAT in mice was associated with larger MI size after I/R; conversely, augmenting BAT by transplantation decreased MI size. We identified Bmp3b as a protein secreted by BAT after I/R. Compared with WT mice, Bmp3b-deficient mice developed larger MIs. Increasing functional BAT by transplanting BAT from WT mice to Bmp3b-deficient mice reduced I/R injury whereas transplanting BAT from Bmp3b-deficient mice did not. Treatment of WT mice with BMP3b before reperfusion decreased MI size. The cardioprotective effect of BMP3b was mediated through SMAD1/5/8. In humans, the plasma level of BMP3b increased after MI and was positively correlated with the extent of cardiac injury. CONCLUSIONS: The results of this study suggest a cardioprotective role of BAT and BMP3b, a protein secreted by BAT, in a model of I/R injury. Interventions increasing BMP3b levels or targeting Smad 1/5 may represent novel therapeutic approaches to decrease myocardial damage in I/R injury.
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Enfermedad de la Arteria Coronaria , Factor 10 de Diferenciación de Crecimiento , Infarto del Miocardio , Isquemia Miocárdica , Daño por Reperfusión Miocárdica , Animales , Humanos , Ratones , Tejido Adiposo Pardo/metabolismo , Factor 10 de Diferenciación de Crecimiento/metabolismo , Ratones Endogámicos C57BL , Infarto del Miocardio/metabolismo , Daño por Reperfusión Miocárdica/genética , Daño por Reperfusión Miocárdica/prevención & control , Daño por Reperfusión Miocárdica/tratamiento farmacológico , ReperfusiónRESUMEN
Bone morphogenetic protein (BMP) receptor signaling is critical for the regulation of the endocrine system and cardiovascular structure and function. The objective of this study was to investigate whether Bmp3b, a glycoprotein synthetized and secreted by adipose tissue, is necessary to regulate glucose and lipid metabolism, adipogenesis, and cardiovascular remodeling. Over the course of 4 mo, Bmp3b-knockout (Bmp3b-/-) mice gained more weight than wild-type (WT) mice. The plasma levels of cholesterol and triglycerides were higher in Bmp3b-/- mice than in WT mice. Bmp3b-/- mice developed insulin resistance and glucose intolerance. The basal heart rate was higher in Bmp3b-/- mice than in WT mice, and echocardiography revealed eccentric remodeling in Bmp3b-/- mice. The expression of adipogenesis-related genes in white adipose tissue was higher in Bmp3b-/- mice than in WT control mice. In vitro studies showed that Bmp3b modulates the activity of the C/ebpα promoter, an effect mediated by Smad2/3. The results of this study suggest that Bmp3b is necessary for the maintenance of homeostasis in terms of age-related weight gain, glucose metabolism, and left ventricular (LV) remodeling and function. Interventions that increase the level or function of BMP3b may decrease cardiovascular risk and pathological cardiac remodeling.
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Adipogénesis/fisiología , Factor 10 de Diferenciación de Crecimiento/deficiencia , Factor 10 de Diferenciación de Crecimiento/fisiología , Síndrome Metabólico/etiología , Adipocitos/patología , Tejido Adiposo/patología , Animales , Proteína Morfogenética Ósea 3/deficiencia , Proteína Morfogenética Ósea 3/fisiología , Dislipidemias/etiología , Femenino , Intolerancia a la Glucosa/etiología , Cardiopatías/etiología , Cardiopatías/fisiopatología , Resistencia a la Insulina/fisiología , Masculino , Síndrome Metabólico/fisiopatología , Ratones , Ratones Endogámicos C57BL , Ratones Noqueados , Obesidad/etiología , Obesidad/patología , Transducción de Señal/fisiologíaRESUMEN
The most common pediatric mitochondrial disease is Leigh syndrome, an episodic, subacute neurodegeneration that can lead to death within the first few years of life, for which there are no proven general therapies. Mice lacking the complex I subunit, Ndufs4, develop a fatal progressive encephalopathy resembling Leigh syndrome and die at ≈60 d of age. We previously reported that continuously breathing normobaric 11% O2 from an early age prevents neurological disease and dramatically improves survival in these mice. Here, we report three advances. First, we report updated survival curves and organ pathology in Ndufs4 KO mice exposed to hypoxia or hyperoxia. Whereas normoxia-treated KO mice die from neurodegeneration at about 60 d, hypoxia-treated mice eventually die at about 270 d, likely from cardiac disease, and hyperoxia-treated mice die within days from acute pulmonary edema. Second, we report that more conservative hypoxia regimens, such as continuous normobaric 17% O2 or intermittent hypoxia, are ineffective in preventing neuropathology. Finally, we show that breathing normobaric 11% O2 in mice with late-stage encephalopathy reverses their established neurological disease, evidenced by improved behavior, circulating disease biomarkers, and survival rates. Importantly, the pathognomonic MRI brain lesions and neurohistopathologic findings are reversed after 4 wk of hypoxia. Upon return to normoxia, Ndufs4 KO mice die within days. Future work is required to determine if hypoxia can be used to prevent and reverse neurodegeneration in other animal models, and to determine if it can be provided in a safe and practical manner to allow in-hospital human therapeutic trials.
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Complejo I de Transporte de Electrón/genética , Hipoxia/metabolismo , Enfermedad de Leigh/patología , Enfermedad de Leigh/terapia , Mitocondrias/patología , Enfermedades Neurodegenerativas/terapia , Animales , Modelos Animales de Enfermedad , Enfermedad de Leigh/mortalidad , Ratones , Ratones Endogámicos C57BL , Ratones Noqueados , Enfermedades Neurodegenerativas/patología , Oxígeno/uso terapéutico , RespiraciónRESUMEN
INTRODUCTION: The nitric oxide (NO), soluble guanylate cyclase (sGC), and cyclic guanosine monophosphate (cGMP) pathway is the leading pathway in penile erection. AIM: To assess erectile function in a mouse model in which sGC is deficient in heme (apo-sGC) and unresponsive to NO. METHODS: Mutant mice (sGCß1ki/ki) that express an sGC enzyme that retains basal activity but fails to respond to NO because of heme deficiency (apo-sGC) were used. Isolated corpora cavernosa from sGCß1ki/ki and wild-type mice were mounted in vitro for isometric tension recordings in response to sGC-dependent and -independent vasorelaxant agents. In addition, the erectile effects of some of these agents were tested in vivo at intracavernosal injection. MAIN OUTCOME MEASURES: In vitro and in vivo recordings of erectile responses in sGCß1ki/ki and wild-type mice after stimulation with sGC-dependent and -independent vasorelaxant agents. RESULTS: NO-induced responses were abolished in sGCß1ki/ki mice in vitro and in vivo. The ability of the heme-dependent, NO-independent sGC stimulator BAY 41-2272 to relax the corpora cavernosa was markedly attenuated in sGCß1ki/ki mice. In contrast, the relaxation response to the heme- and NO-independent sGC activator BAY 58-2667 was significantly enhanced in sGCß1ki/ki mice. The relaxing effect of sGC-independent vasorelaxant agents was similar in wild-type and sGCß1ki/ki mice, illustrating that the observed alterations in vasorelaxation are limited to NO-sGC-cGMP-mediated processes. CONCLUSION: Our results suggest that sGC is the sole target of NO in erectile physiology. Furthermore, this study provides indirect evidence that, in addition to sGCα1ß1, sGCα2ß1 is important for erectile function. In addition, the significant relaxation observed in sGCß1ki/ki mice with the cumulative addition of the sGC activator BAY 58-2667 indicates that sGC activators might offer value in treating erectile dysfunction.
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GMP Cíclico/metabolismo , Disfunción Eréctil/fisiopatología , Hemo/deficiencia , Guanilil Ciclasa Soluble/metabolismo , Animales , Modelos Animales de Enfermedad , Guanilato Ciclasa/metabolismo , Humanos , Masculino , Ratones , Óxido Nítrico/metabolismo , Erección Peniana/efectos de los fármacos , Pene/fisiopatologíaRESUMEN
BACKGROUND: Volatile anesthetics increase levels of the neurotransmitter nitric oxide (NO) and the secondary messenger molecule cyclic guanosine monophosphate (cGMP) in the brain. NO activates the enzyme guanylyl cyclase (GC) to produce cGMP. We hypothesized that the NO-GC-cGMP pathway contributes to anesthesia-induced unconsciousness. METHODS: Sevoflurane-induced loss and return of righting reflex (LORR and RORR, respectively) were studied in wild-type mice (WT) and in mice congenitally deficient in the GC-1α subunit (GC-1-/- mice). Spatial distributions of GC-1α and the GC-2α subunit in the brain were visualized by in situ hybridization. Brain cGMP levels were measured in WT and GC-1-/- mice after inhaling oxygen with or without 1.2% sevoflurane for 20 min. RESULTS: Higher concentrations of sevoflurane were required to induce LORR in GC-1-/- mice than in WT mice (1.5 ± 0.1 vs. 1.1 ± 0.2%, respectively, n = 14 and 14, P < 0.0001). Similarly, RORR occurred at higher concentrations of sevoflurane in GC-1-/- mice than in WT mice (1.0 ± 0.1 vs. 0.8 ± 0.1%, respectively, n = 14 and 14, P < 0.0001). Abundant GC-1α and GC-2α mRNA expression was detected in the cerebral cortex, medial habenula, hippocampus, and cerebellum. Inhaling 1.2% sevoflurane for 20 min increased cGMP levels in the brains of WT mice from 2.6 ± 2.0 to 5.5 ± 3.7 pmol/mg protein (n = 13 and 10, respectively, P = 0.0355) but not in GC-1-/- mice. CONCLUSION: Congenital deficiency of GC-1α abolished the ability of sevoflurane anesthesia to increase cGMP levels in the whole brain, and increased the concentration of sevoflurane required to induce LORR. Impaired NO-cGMP signaling raises the threshold for producing sevoflurane-induced unconsciousness in mice.
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Anestésicos por Inhalación/farmacología , Guanilato Ciclasa/genética , Éteres Metílicos/farmacología , Animales , Encéfalo/metabolismo , Guanosina Monofosfato/metabolismo , Ratones Noqueados , Reflejo de Enderezamiento/efectos de los fármacos , SevofluranoRESUMEN
The study of brown adipose tissue (BAT) has gained significant scientific interest since the discovery of functional BAT in adult humans. The thermogenic properties of BAT are well recognized; however, data generated in the last decade in both rodents and humans reveal therapeutic potential for BAT against metabolic disorders and obesity. Here we review the current literature in light of a potential role for BAT in beneficially mediating cardiovascular health. We focus mainly on BAT's actions in obesity, vascular tone, and glucose and lipid metabolism. Furthermore, we discuss the recently discovered endocrine factors that have a potential beneficial role in cardiovascular health. These BAT-secreted factors may have a favorable effect against cardiovascular risk either through their metabolic role or by directly affecting the heart.
Asunto(s)
Tejido Adiposo Pardo/metabolismo , Vasos Sanguíneos/metabolismo , Enfermedades Cardiovasculares/prevención & control , Miocardio/metabolismo , Comunicación Paracrina , Tejido Adiposo Pardo/fisiopatología , Animales , Vasos Sanguíneos/fisiopatología , Enfermedades Cardiovasculares/etiología , Enfermedades Cardiovasculares/metabolismo , Enfermedades Cardiovasculares/fisiopatología , Dislipidemias/complicaciones , Dislipidemias/metabolismo , Dislipidemias/fisiopatología , Trastornos del Metabolismo de la Glucosa/complicaciones , Trastornos del Metabolismo de la Glucosa/metabolismo , Trastornos del Metabolismo de la Glucosa/fisiopatología , Humanos , Obesidad/complicaciones , Obesidad/metabolismo , Obesidad/fisiopatología , Factores de Riesgo , Transducción de Señal , TermogénesisRESUMEN
Bone morphogenetic protein (BMP) signaling contributes to the development of cardiac hypertrophy. However, the identity of the BMP type I receptor involved in cardiac hypertrophy and the underlying molecular mechanisms are poorly understood. By using quantitative PCR and immunoblotting, we demonstrated that BMP signaling increased during phenylephrine-induced hypertrophy in cultured neonatal rat cardiomyocytes (NRCs), as evidenced by increased phosphorylation of Smads 1 and 5 and induction of Id1 gene expression. Inhibition of BMP signaling with LDN193189 or noggin, and silencing of Smad 1 or 4 using small interfering RNA diminished the ability of phenylephrine to induce hypertrophy in NRCs. Conversely, activation of BMP signaling with BMP2 or BMP4 induced hypertrophy in NRCs. Luciferase reporter assay further showed that BMP2 or BMP4 treatment of NRCs repressed atrogin-1 gene expression concomitant with an increase in calcineurin protein levels and enhanced activity of nuclear factor of activated T cells, providing a mechanism by which BMP signaling contributes to cardiac hypertrophy. In a model of cardiac hypertrophy, C57BL/6 mice treated with angiotensin II (A2) had increased BMP signaling in the left ventricle. Treatment with LDN193189 attenuated A2-induced cardiac hypertrophy and collagen deposition in left ventricles. Cardiomyocyte-specific deletion of BMP type I receptor ALK2 (activin-like kinase 2), but not ALK1 or ALK3, inhibited BMP signaling and mitigated A2-induced cardiac hypertrophy and left ventricular fibrosis in mice. The results suggest that BMP signaling upregulates the calcineurin/nuclear factor of activated T cell pathway via BMP type I receptor ALK2, contributing to cardiac hypertrophy and fibrosis.
Asunto(s)
Receptores de Activinas Tipo I/metabolismo , Angiotensina II , Proteína Morfogenética Ósea 2/farmacología , Proteína Morfogenética Ósea 4/farmacología , Receptores de Proteínas Morfogenéticas Óseas de Tipo 1/metabolismo , Cardiomegalia/enzimología , Miocitos Cardíacos/enzimología , Receptores de Activinas Tipo I/deficiencia , Receptores de Activinas Tipo I/genética , Receptores de Activinas Tipo II , Animales , Receptores de Proteínas Morfogenéticas Óseas de Tipo 1/deficiencia , Receptores de Proteínas Morfogenéticas Óseas de Tipo 1/genética , Cardiomegalia/inducido químicamente , Cardiomegalia/genética , Cardiomegalia/patología , Cardiomegalia/prevención & control , Células Cultivadas , Modelos Animales de Enfermedad , Relación Dosis-Respuesta a Droga , Fibrosis , Proteína 1 Inhibidora de la Diferenciación/genética , Proteína 1 Inhibidora de la Diferenciación/metabolismo , Ratones Endogámicos C57BL , Ratones Noqueados , Miocitos Cardíacos/efectos de los fármacos , Miocitos Cardíacos/patología , Factores de Transcripción NFATC/metabolismo , Fenilefrina/farmacología , Fosforilación , Pirazoles/farmacología , Pirimidinas/farmacología , Interferencia de ARN , Ratas Sprague-Dawley , Transducción de Señal , Proteínas Smad/genética , Proteínas Smad/metabolismo , Factores de Tiempo , TransfecciónRESUMEN
Dysregulated nitric oxide (NO) signaling contributes to the pathogenesis of hypertension, a prevalent and often sex-specific risk factor for cardiovascular disease. We previously reported that mice deficient in the α1-subunit of the NO receptor soluble guanylate cyclase (sGCα1 (-/-) mice) display sex- and strain-specific hypertension: male but not female sGCα1 (-/-) mice are hypertensive on an 129S6 (S6) but not a C57BL6/J (B6) background. We aimed to uncover the genetic and molecular basis of the observed sex- and strain-specific blood pressure phenotype. Via linkage analysis, we identified a suggestive quantitative trait locus associated with elevated blood pressure in male sGCα1 (-/-)S6 mice. This locus encompasses Cyp4a12a, encoding the predominant murine synthase of the vasoconstrictor 20-hydroxy-5,8,11,14-eicosatetraenoic acid (20-HETE). Renal expression of Cyp4a12a in mice was associated with genetic background, sex, and testosterone levels. In addition, 20-HETE levels were higher in renal preglomerular microvessels of male sGCα1 (-/-)S6 than of male sGCα1 (-/-)B6 mice. Furthermore, treating male sGCα1 (-/-)S6 mice with the 20-HETE antagonist 20-hydroxyeicosa-6(Z),15(Z)-dienoic acid (20-HEDE) lowered blood pressure. Finally, 20-HEDE rescued the genetic background- and testosterone-dependent impairment of acetylcholine-induced relaxation in renal interlobar arteries associated with sGCα1 deficiency. Elevated Cyp4a12a expression and 20-HETE levels render mice susceptible to hypertension and vascular dysfunction in a setting of sGCα1 deficiency. Our data identify Cyp4a12a as a candidate sex-specific blood pressure-modifying gene in the context of deficient NO-sGC signaling.
Asunto(s)
Andrógenos/farmacología , Familia 4 del Citocromo P450/genética , Ácidos Hidroxieicosatetraenoicos/metabolismo , Hipertensión/metabolismo , Guanilil Ciclasa Soluble/metabolismo , Animales , Presión Sanguínea/efectos de los fármacos , Presión Sanguínea/fisiología , Femenino , Ligamiento Genético , Hipertensión/genética , Riñón/efectos de los fármacos , Riñón/metabolismo , Masculino , Ratones , Ratones Noqueados , Sitios de Carácter Cuantitativo , Factores Sexuales , Guanilil Ciclasa Soluble/genética , Testosterona/sangreRESUMEN
Human studies suggest that insulin resistance and obesity are associated with a decrease in B-type natriuretic peptide (BNP) plasma concentrations. The objective of the study was to gain insights into the mechanisms involved in the association between insulin resistance and decreased BNP plasma concentrations. Mice fed a high-fat, high-fructose (HFHF) diet for 4 weeks developed mild obesity and systemic insulin resistance. Elevated plasma concentrations of insulin, glucose and triglycerides were noted. The HFHF diet was also associated with myocardial insulin resistance, characterized by an impaired response of the phosphoinositide 3-kinase-AKT (PI3K-AKT) pathway to insulin in the left ventricle. Myocardial BNP expression and protein were decreased in HFHF-fed mice compared with control animals. Exposure of cardiomyocytes to 100 nm insulin activated PI3K-AKT signalling (15 min) and induced a 1.9 ± 0.3-fold increase in BNP gene expression (6 h). Prolonged exposure of cardiomyocytes to a high insulin concentration (100 nm) for 48 h induced insulin resistance, characterized by an impaired response of the PI3K-AKT signalling pathway and a decreased response of the BNP gene expression to insulin. The decreased response in BNP gene expression was reproduced by treating cardiomyocytes for 7 h with a PI3-kinase inhibitor (wortmannin). In conclusion, HFHF diet in vivo, prolonged exposure to an elevated concentration of insulin or inhibition of the PI3K-AKT pathway in vitro all decrease BNP mRNA levels; this decrease may in turn contribute to the decreased BNP peptide concentrations in plasma observed in insulin-resistant individuals.
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Insulina/fisiología , Péptido Natriurético Encefálico/biosíntesis , Obesidad/metabolismo , Androstadienos/farmacología , Animales , Glucemia/metabolismo , Dieta , Dieta Alta en Grasa , Inhibidores Enzimáticos/farmacología , Expresión Génica/efectos de los fármacos , Jarabe de Maíz Alto en Fructosa , Insulina/sangre , Resistencia a la Insulina , Masculino , Ratones , Ratones Endogámicos C57BL , Miocitos Cardíacos/metabolismo , Fosfatidilinositol 3-Quinasas/metabolismo , Inhibidores de las Quinasa Fosfoinosítidos-3 , Triglicéridos/sangre , WortmaninaRESUMEN
Brown adipose tissue (BAT) has well recognized thermogenic properties mediated by uncoupling protein 1 (UCP1); more recently, BAT has been demonstrated to modulate cardiovascular risk factors. To investigate whether BAT also affects myocardial injury and remodeling, UCP1-deficient (UCP1(-/-)) mice, which have dysfunctional BAT, were subjected to catecholamine-induced cardiomyopathy. At baseline, there were no differences in echocardiographic parameters, plasma cardiac troponin I (cTnI) or myocardial fibrosis between wild-type (WT) and UCP1(-/-) mice. Isoproterenol infusion increased cTnI and myocardial fibrosis and induced left ventricular (LV) hypertrophy in both WT and UCP1(-/-) mice. UCP1(-/-) mice also demonstrated exaggerated myocardial injury, fibrosis, and adverse remodeling, as well as decreased survival. Transplantation of WT BAT to UCP1(-/-) mice prevented the isoproterenol-induced cTnI increase and improved survival, whereas UCP1(-/-) BAT transplanted to either UCP1(-/-) or WT mice had no effect on cTnI release. After 3 days of isoproterenol treatment, phosphorylated AKT and ERK were lower in the LV's of UCP1(-/-) mice than in those of WT mice. Activation of BAT was also noted in a model of chronic ischemic cardiomyopathy, and was correlated to LV dysfunction. Deficiency in UCP1, and accompanying BAT dysfunction, increases cardiomyocyte injury and adverse LV remodeling, and decreases survival in a mouse model of catecholamine-induced cardiomyopathy. Myocardial injury and decreased survival are rescued by transplantation of functional BAT to UCP1(-/-) mice, suggesting a systemic cardioprotective role of functional BAT. BAT is also activated in chronic ischemic cardiomyopathy.
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Tejido Adiposo Pardo/fisiología , Cardiomiopatías/patología , Cardiomiopatías/fisiopatología , Catecolaminas/efectos adversos , Remodelación Ventricular , Tejido Adiposo Pardo/trasplante , Animales , Biomarcadores/metabolismo , Presión Sanguínea/efectos de los fármacos , Peso Corporal/efectos de los fármacos , Cardiomiopatías/inducido químicamente , Cardiomiopatías/diagnóstico por imagen , Cardiotónicos/metabolismo , Catecolaminas/administración & dosificación , Quinasas MAP Reguladas por Señal Extracelular/metabolismo , Fibrosis , Regulación de la Expresión Génica/efectos de los fármacos , Insuficiencia Cardíaca/complicaciones , Insuficiencia Cardíaca/enzimología , Insuficiencia Cardíaca/patología , Insuficiencia Cardíaca/fisiopatología , Ventrículos Cardíacos/efectos de los fármacos , Ventrículos Cardíacos/metabolismo , Canales Iónicos/deficiencia , Canales Iónicos/genética , Canales Iónicos/metabolismo , Isoproterenol/farmacología , Masculino , Ratones Endogámicos C57BL , Proteínas Mitocondriales/deficiencia , Proteínas Mitocondriales/genética , Proteínas Mitocondriales/metabolismo , Miocardio/patología , Miocitos Cardíacos , Fosforilación/efectos de los fármacos , Proteínas Proto-Oncogénicas c-akt/metabolismo , Transducción de Señal/efectos de los fármacos , Análisis de Supervivencia , Ultrasonografía , Proteína Desacopladora 1 , Remodelación Ventricular/efectos de los fármacosRESUMEN
The nitric oxide (NO)-cyclic guanosine monophosphate (cGMP) signaling system is a well-characterized modulator of cardiovascular function, in general, and blood pressure, in particular. The availability of mice mutant for key enzymes in the NO-cGMP signaling system facilitated the identification of interactions with other blood pressure modifying pathways (e.g. the renin-angiotensin-aldosterone system) and of gender-specific effects of impaired NO-cGMP signaling. In addition, recent genome-wide association studies identified blood pressure-modifying genetic variants in genes that modulate NO and cGMP levels. Together, these findings have advanced our understanding of how NO-cGMP signaling regulates blood pressure. In this review, we will summarize the results obtained in mice with disrupted NO-cGMP signaling and highlight the relevance of this pathway as a potential therapeutic target for the treatment of hypertension.
Asunto(s)
GMP Cíclico/fisiología , Hipertensión/fisiopatología , Óxido Nítrico/fisiología , Animales , Presión Sanguínea/fisiología , GMP Cíclico/metabolismo , Proteínas Quinasas Dependientes de GMP Cíclico/metabolismo , Endotelio Vascular/fisiología , Estudio de Asociación del Genoma Completo , Humanos , Ratones , Ratones Mutantes , Modelos Animales , Óxido Nítrico/metabolismo , Transducción de Señal/fisiología , Vasoconstricción/fisiología , Vasodilatación/fisiologíaRESUMEN
PURPOSE: Soluble guanylyl cyclase (sGC), which plays a pivotal role in penile erection, is a heterodimer build up by an α and a ß subunit. For both subunits two isoforms have been characterized, but only the sGCα(1)ß(1) and sGCα(2)ß(1) isoforms seem to be functionally active. To elucidate the functional role of the sGCα(1)ß(1) heterodimer in the mechanism of erection, experiments were performed in vivo and on isolated corpora cavernosa (CC) using sGCα(1)(-/-) mice. MATERIALS AND METHODS: For the in vivo study sGC-dependent and -independent vasorelaxing agents were injected intracavernosally in sGCα(1)(-/-) and sGCα(1)(+/+) mice and the rise in intracavernosal pressure was recorded. For the in vitro study, isolated CC tissues from sGCα(1)(-/-) and sGCα(1)(+/+) mice were mounted in organ baths for isometric tension recording and concentration-dependent curves were obtained for sGC-dependent and -independent vasorelaxing agents. These experiments were performed on 2 different mice strains (129SvEvS7 and C57BL6/J) to determine potential strain differences. RESULTS: The responses in sGCα(1)(-/-) after administration of the NO-donors, sodium nitroprusside (SNP) and spermine-NO, and to electrical stimulation are significantly reduced although not completely abolished. Responses to sGC-independent vasorelaxing agents are similar in sGCα(1)(-/-) and sGCα(1)(+/+) mice from both strains suggesting that the decreased potential of smooth muscle relaxation is not related to structural changes or changes in the pathway downstream sGC. CONCLUSION: This study illustrates the strain-independent importance of the sGCα(1)ß(1) heterodimer, although remaining vasorelaxing responses in the sGCα(1)(-/-) mice suggest a complementary role for the sGCα(2)ß(1) isoform or (an) sGC-independent mechanism(s).
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Guanilato Ciclasa/fisiología , Erección Peniana/fisiología , Receptores Citoplasmáticos y Nucleares/fisiología , Vasodilatación/fisiología , Animales , Colforsina/farmacología , Estimulación Eléctrica , Guanilato Ciclasa/genética , Masculino , Ratones , Ratones Endogámicos C57BL , Ratones Noqueados , Modelos Animales , Donantes de Óxido Nítrico/farmacología , Nitroprusiato/farmacología , Erección Peniana/efectos de los fármacos , Receptores Citoplasmáticos y Nucleares/genética , Guanilil Ciclasa Soluble , Espermina/análogos & derivados , Espermina/farmacologíaRESUMEN
Altered cGMP signaling has been implicated in myocardial depression, morbidity, and mortality associated with sepsis. Previous studies, using inhibitors of soluble guanylate cyclase (sGC), suggested that cGMP generated by sGC contributed to the cardiac dysfunction and mortality associated with sepsis. We used sGC(alpha)(1)-deficient (sGC(alpha)(1)(-/-)) mice to unequivocally determine the role of sGC(alpha)(1)beta(1) in the development of cardiac dysfunction and death associated with two models of inflammatory shock: endotoxin- and TNF-induced shock. At baseline, echocardiographic assessment and invasive hemodynamic measurements of left ventricular (LV) dimensions and function did not differ between wild-type (WT) mice and sGC(alpha)(1)(-/-) mice on the C57BL/6 background (sGC(alpha)(1)(-/-B6) mice). At 14 h after endotoxin challenge, cardiac dysfunction was more pronounced in sGC(alpha)(1)(-/-B6) than WT mice, as assessed using echocardiographic and hemodynamic indexes of LV function. Similarly, Ca(2+) handling and cell shortening were impaired to a greater extent in cardiomyocytes isolated from sGC(alpha)(1)(-/-B6) than WT mice after endotoxin challenge. Importantly, morbidity and mortality associated with inflammatory shock induced by endotoxin or TNF were increased in sGC(alpha)(1)(-/-B6) compared with WT mice. Together, these findings suggest that cGMP generated by sGC(alpha)(1)beta(1) protects against cardiac dysfunction and mortality in murine inflammatory shock models.
Asunto(s)
Guanilato Ciclasa/genética , Guanilato Ciclasa/metabolismo , Receptores Citoplasmáticos y Nucleares/genética , Receptores Citoplasmáticos y Nucleares/metabolismo , Choque Cardiogénico , Disfunción Ventricular Izquierda , Animales , Presión Sanguínea/fisiología , Calcio/metabolismo , Células Cultivadas , Modelos Animales de Enfermedad , Ecocardiografía , Endotoxinas/toxicidad , Activación Enzimática/fisiología , Regulación Enzimológica de la Expresión Génica/fisiología , Estimación de Kaplan-Meier , Ratones , Ratones Endogámicos C57BL , Ratones Mutantes , Contracción Miocárdica/fisiología , Miocitos Cardíacos/citología , Miocitos Cardíacos/fisiología , Óxido Nítrico/metabolismo , Sepsis/inmunología , Sepsis/metabolismo , Sepsis/mortalidad , Choque Cardiogénico/inmunología , Choque Cardiogénico/metabolismo , Choque Cardiogénico/mortalidad , Guanilil Ciclasa Soluble , Factor de Necrosis Tumoral alfa/toxicidad , Disfunción Ventricular Izquierda/diagnóstico por imagen , Disfunción Ventricular Izquierda/inmunología , Disfunción Ventricular Izquierda/mortalidad , Presión Ventricular/fisiologíaRESUMEN
Purpose: Glaucoma, a leading cause of blindness worldwide, often remains undetected until irreversible vision loss has occurred. Treatments focus on lowering intraocular pressure (IOP), the only modifiable and readily measurable risk factor. However, IOP can vary and does not always predict disease progression. MicroRNAs (miRNAs) are promising biomarkers. They are abundant and stable in biological fluids, including plasma and aqueous humor (AqH). We aimed to identify differentially expressed miRNAs in AqH and plasma from glaucoma, exfoliation syndrome (XFS), and control subjects. Methods: Plasma and AqH from two ethnic cohorts were harvested from glaucoma or XFS (often associated with glaucoma, n = 33) and control (n = 31) patients undergoing elective surgery. A custom miRNA array measured 372 miRNAs. Molecular target prediction and pathway analysis were performed with Ingenuity Pathway Analysis (IPA) and DIANA bioinformatical tools. Results: Levels of miRNAs in plasma, a readily accessible biomarker source, correlated with miRNA levels in AqH. Twenty circulating miRNAs were at least 1.5-fold higher in glaucoma or XFS patients than in controls across two ethnic cohorts: miR-4667-5p (P = 4.1 × 10-5), miR-99b-3p (P = 4.8 × 10-5), miR-637 (P = 5.1 × 10-5), miR-4490 (P = 5.7 × 10-5), miR-1253 (P = 6.0 × 10-5), miR-3190-3p (P = 3.1 × 10-4), miR-3173-3p (P = 0.001), miR-608 (P = 0.001), miR-4725-3p (P = 0.002), miR-4448 (P = 0.002), and miR-323b-5p (P = 0.002), miR-4538 (P = 0.003), miR-3913-3p (P = 0.003), miR-3159 (P = 0.003), miR-4663 (P = 0.003), miR-4767 (P = 0.003), miR-4724-5p (P = 0.003), miR-1306-5p (P = 0.003), miR-181b-3p (P = 0.004), and miR-433-3p (P = 0.004). miR-637, miR-1306-5p, and miR-3159, in combination, allowed discrimination between glaucoma patients and control subjects (AUC = 0.91 ± 0.008, sensitivity 85.0%, specificity 87.5%). Conclusions: These results identify specific miRNAs as potential biomarkers and provide insight into the molecular processes underlying glaucoma.
Asunto(s)
Humor Acuoso/metabolismo , Biomarcadores/sangre , Síndrome de Exfoliación/sangre , Glaucoma de Ángulo Abierto/sangre , MicroARNs/sangre , Anciano , Anciano de 80 o más Años , Pueblo Asiatico/etnología , Síndrome de Exfoliación/etnología , Síndrome de Exfoliación/cirugía , Femenino , Perfilación de la Expresión Génica , Glaucoma de Ángulo Abierto/etnología , Glaucoma de Ángulo Abierto/cirugía , Humanos , Presión Intraocular , Japón/epidemiología , Masculino , Persona de Mediana Edad , Reacción en Cadena de la Polimerasa , Sensibilidad y Especificidad , Estados Unidos/epidemiología , Población Blanca/etnologíaRESUMEN
Serum levels of fibroblast growth factor 23 (FGF23) markedly increase with renal impairment, with FGF23 levels correlating with the presence of left ventricular hypertrophy (LVH) and mortality in patients with chronic kidney disease (CKD). FGF23 activates calcineurin/nuclear factor of activated T cell (NFAT) signaling and induces hypertrophy in murine cardiomyocytes. X-linked hypophosphatemia (XLH) is characterized by high circulating levels of FGF23 but, in contrast to CKD, is associated with hypophosphatemia. The cardiac effects of high circulating levels of FGF23 in XLH are not well defined. Thus, studies were undertaken to define the cardiac phenotype in the mouse model of XLH (Hyp mice). Echocardiographic and histological analyses demonstrated that Hyp left ventricles (LVs) are smaller than those of wild-type mice. Messenger RNA expression of cardiac hypertrophy markers was not altered in the LV or right ventricle of Hyp mice. However, the Hyp LVs had increased expression of the NFAT target genes NFATc1 and RCAN1. To determine whether phosphate alone can induce markers of hypertrophy, differentiated C2C12 myocytes were treated with phosphate. Phosphate treatment increased expression of cardiac hypertrophy markers, supporting a primary role for phosphate in inducing LVH. Although previous studies showed that increased circulating FGF23 and phosphate levels are associated with LVH, our results demonstrated that in XLH, high circulating levels of FGF23 in the setting of hypophosphatemia do not induce cardiac hypertrophy.
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Raquitismo Hipofosfatémico Familiar/metabolismo , Factores de Crecimiento de Fibroblastos/metabolismo , Ventrículos Cardíacos/patología , Hipertrofia Ventricular Izquierda/genética , Miocardio/patología , Animales , Proteínas de Unión al Calcio , Cardiomegalia/genética , Cardiomegalia/metabolismo , Modelos Animales de Enfermedad , Ecocardiografía , Factor-23 de Crecimiento de Fibroblastos , Expresión Génica , Ventrículos Cardíacos/metabolismo , Hipertrofia Ventricular Izquierda/metabolismo , Péptidos y Proteínas de Señalización Intracelular/genética , Masculino , Ratones , Proteínas Musculares/genética , Miocardio/metabolismo , Factores de Transcripción NFATC/genética , Tamaño de los Órganos , ARN Mensajero/metabolismoRESUMEN
Brown adipose tissue (BAT) activation increases glucose and lipid consumption; as such, it is been considered as a potential therapy to decrease obesity. BAT is highly vascularized and its activation is associated with a necessary increase in blood flow. However, whether increasing BAT blood flow per se increases BAT activity is unknown. To examine this hypothesis, we investigated whether an isolated increase in BAT blood flow obtained by ß2-adrenoreceptor (ß2-AR) stimulation with salbutamol increased BAT activity. BAT blood flow was estimated in vivo in mice using contrast-enhanced ultrasound. The absence of direct effect of salbutamol on the function of isolated brown adipocytes was assessed by measuring oxygen consumption. The effect of salbutamol on BAT activity was investigated by measuring BAT glucose uptake in vivo. BAT blood flow increased by 2.3 ± 0.6-fold during ß2-AR stimulation using salbutamol infusion in mice (P= 0.003). ß2-AR gene expression was detectable in BAT but was extremely low in isolated brown adipocytes. Oxygen consumption of isolated brown adipocytes did not change with salbutamol exposure, confirming the absence of a direct effect of ß2-AR agonist on brown adipocytes. Finally, ß2-AR stimulation by salbutamol increased BAT glucose uptake in vivo (991 ± 358 vs. 135 ± 49 ng glucose/mg tissue/45 min in salbutamol vs. saline injected mice, respectively,P= 0.046). In conclusion, an increase in BAT blood flow without direct stimulation of the brown adipocytes is associated with increased BAT metabolic activity. Increasing BAT blood flow might represent a new therapeutic target in obesity.
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Tejido Adiposo Pardo/metabolismo , Tejido Adiposo Pardo/fisiología , Receptores Adrenérgicos beta 2/metabolismo , Animales , Transporte Biológico/fisiología , Glucosa/metabolismo , Masculino , Ratones , Ratones Endogámicos C57BL , Obesidad/metabolismo , Obesidad/fisiopatología , Consumo de Oxígeno/fisiología , Flujo Sanguíneo Regional/fisiologíaRESUMEN
BACKGROUND: Pharmacological activation of cGMP-dependent protein kinase G I (PKGI) has emerged as a therapeutic strategy for humans with heart failure. However, PKG-activating drugs have been limited by hypotension arising from PKG-induced vasodilation. PKGIα antiremodeling substrates specific to the myocardium might provide targets to circumvent this limitation, but currently remain poorly understood. METHODS AND RESULTS: We performed a screen for myocardial proteins interacting with the PKGIα leucine zipper (LZ)-binding domain to identify myocardial-specific PKGI antiremodeling substrates. Our screen identified cardiac myosin-binding protein-C (cMyBP-C), a cardiac myocyte-specific protein, which has been demonstrated to inhibit cardiac remodeling in the phosphorylated state, and when mutated leads to hypertrophic cardiomyopathy in humans. GST pulldowns and precipitations with cGMP-conjugated beads confirmed the PKGIα-cMyBP-C interaction in myocardial lysates. In vitro studies demonstrated that purified PKGIα phosphorylates the cMyBP-C M-domain at Ser-273, Ser-282, and Ser-302. cGMP induced cMyBP-C phosphorylation at these residues in COS cells transfected with PKGIα, but not in cells transfected with LZ mutant PKGIα, containing mutations to disrupt LZ substrate binding. In mice subjected to left ventricular pressure overload, PKGI activation with sildenafil increased cMyBP-C phosphorylation at Ser-273 compared with untreated mice. cGMP also induced cMyBP-C phosphorylation in isolated cardiac myocytes. CONCLUSIONS: Taken together, these data support that PKGIα and cMyBP-C interact in the heart and that cMyBP-C is an anti remodeling PKGIα kinase substrate. This study provides the first identification of a myocardial-specific PKGIα LZ-dependent antiremodeling substrate and supports further exploration of PKGIα myocardial LZ substrates as potential therapeutic targets for heart failure.
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Proteínas Portadoras/metabolismo , Proteína Quinasa Dependiente de GMP Cíclico Tipo I/metabolismo , Insuficiencia Cardíaca/metabolismo , Animales , GMP Cíclico/fisiología , Modelos Animales de Enfermedad , Insuficiencia Cardíaca/etiología , Ratones , Ratones Endogámicos C57BL , Miocitos Cardíacos/metabolismo , Fosforilación , Ratas , Ratas Sprague-DawleyRESUMEN
Oxidative stress, a central mediator of cardiovascular disease, results in loss of the prosthetic haem group of soluble guanylate cyclase (sGC), preventing its activation by nitric oxide (NO). Here we introduce Apo-sGC mice expressing haem-free sGC. Apo-sGC mice are viable and develop hypertension. The haemodynamic effects of NO are abolished, but those of the sGC activator cinaciguat are enhanced in apo-sGC mice, suggesting that the effects of NO on smooth muscle relaxation, blood pressure regulation and inhibition of platelet aggregation require sGC activation by NO. Tumour necrosis factor (TNF)-induced hypotension and mortality are preserved in apo-sGC mice, indicating that pathways other than sGC signalling mediate the cardiovascular collapse in shock. Apo-sGC mice allow for differentiation between sGC-dependent and -independent NO effects and between haem-dependent and -independent sGC effects. Apo-sGC mice represent a unique experimental platform to study the in vivo consequences of sGC oxidation and the therapeutic potential of sGC activators.
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Sistema Cardiovascular/metabolismo , Guanilato Ciclasa/genética , Hemo/genética , Músculo Liso Vascular/metabolismo , Óxido Nítrico/metabolismo , Receptores Citoplasmáticos y Nucleares/genética , Animales , Benzoatos/farmacología , Presión Sanguínea/efectos de los fármacos , Sistema Cardiovascular/efectos de los fármacos , Técnicas de Sustitución del Gen , Hipertensión/genética , Hipotensión/inducido químicamente , Hipotensión/genética , Ratones , Ratones Transgénicos , Músculo Liso Vascular/efectos de los fármacos , Estrés Oxidativo/efectos de los fármacos , Agregación Plaquetaria/efectos de los fármacos , Guanilil Ciclasa Soluble , Factor de Necrosis Tumoral alfa/farmacologíaRESUMEN
BACKGROUND: Cinaciguat (BAY 58-2667), an NO- and heme-independent sGC activator, was shown to be more effective when the heme-group of sGC is oxidized in vascular tissue. In apo-sGC mice (sGCß1 (His105Phe) knockin) both sGC isoforms (sGCα1 ß1 and sGCα2 ß1 ) are heme-deficient and can no longer be activated by NO; these mice, showing decreased gastrointestinal nitrergic relaxation and decreased gastric emptying, can be considered as a model to study the consequence of heme-oxidation in sGC. Our aim was to compare the influence of cinaciguat, on in vitro muscle tone of gastrointestinal tissues, and on gastric emptying in WT and apo-sGC mice. METHODS: Gastrointestinal smooth muscle strips were mounted in organ baths for isometric force recording and cGMP levels were determined by enzyme immunoassay. Protein levels of sGC subunits were assessed by immunoblotting. Gastric emptying was determined by phenol red recovery. KEY RESULTS: Although protein levels of the sGC subunits were lower in gastrointestinal tissues of apo-sGC mice, cinaciguat induced concentration-dependent relaxations and increased cGMP levels in apo-sGC fundus and colon to a similar or greater extent than in WT mice. The sGC inhibitor ODQ increased cinaciguat-induced relaxations and cGMP levels in WT fundus and colon. In apo-sGC antrum, pylorus and jejunum, cinaciguat was not able to induce relaxations. Cinaciguat did not improve delayed gastric emptying in apo-sGC mice. CONCLUSIONS & INFERENCES: Cinaciguat relaxes the fundus and colon efficiently when sGC is in the heme-free condition; the non-effect of cinaciguat in pylorus explains its inability to improve the delayed gastric emptying in apo-sGC mice.
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Benzoatos/farmacología , Motilidad Gastrointestinal/efectos de los fármacos , Motilidad Gastrointestinal/fisiología , Guanilato Ciclasa/metabolismo , Animales , Ensayo de Inmunoadsorción Enzimática , Fundus Gástrico/metabolismo , Técnicas de Sustitución del Gen , Immunoblotting , Isoenzimas , Masculino , Ratones , Relajación Muscular/efectos de los fármacos , Relajación Muscular/fisiología , Tono Muscular/efectos de los fármacos , Tono Muscular/fisiología , Músculo Liso/efectos de los fármacos , Músculo Liso/metabolismo , Técnicas de Cultivo de ÓrganosRESUMEN
Hypertensive heart disease causes significant mortality in older patients, yet there is an incomplete understanding of molecular mechanisms that regulate age-dependent hypertensive left ventricular hypertrophy (LVH). Therefore, we tested the hypothesis that the cGMP-dependent protein kinase G I alpha (PKGIα) attenuates hypertensive LVH by evaluating the cardiac phenotype in mice with selective mutations of the PKGIα leucine zipper domain. These leucine zipper mutant (LZM) mice develop basal hypertension. Compared with wild-type controls, 8-month-old adult LZM mice developed increased left ventricular end-diastolic pressure but without frank LVH. In advanced age (15 months), the LZM mice developed overt pathological LVH. These findings reveal a role of PKGIα in normally attenuating hypertensive LVH. Therefore, mutation of the PKGIα LZ domain produces a clinically relevant model for hypertensive heart disease of aging.