RESUMEN
Insulin resistance (IR) and systemic hypertension are independently associated with heart failure. We reported previously that nitric oxide synthase 3 (NOS3) has a beneficial effect on left ventricular (LV) remodeling and function after pressure-overload in mice. The aim of our study was to investigate the interaction of IR and NOS3 in pressure-overload-induced LV remodeling and dysfunction. Wild-type (WT) and NOS3-deficient (NOS3(-/-)) mice were fed either a standard diet (SD) or a high-fat diet (HFD) to induce IR. After 9 days of diet, mice underwent transverse aortic constriction (TAC). LV structure and function were assessed serially using echocardiography. Cardiomyocytes were isolated, and levels of oxidative stress were evaluated using 2',7'-dichlorodihydrofluorescein diacetate. Cardiac mitochondria were isolated, and mitochondrial respiration and ATP production were measured. TAC induced LV remodeling and dysfunction in all mice. The TAC-induced decrease in LV function was greater in SD-fed NOS3(-/-) mice than in SD-fed WT mice. In contrast, HFD-fed NOS3(-/-) developed less LV remodeling and dysfunction and had better survival than did HFD-fed WT mice. Seven days after TAC, oxidative stress levels were lower in cardiomyocytes from HFD-fed NOS3(-/-) than in those from HFD-fed WT. N(ω)-nitro-L-arginine methyl ester and mitochondrial inhibitors (rotenone and 2-thenoyltrifluoroacetone) decreased oxidative stress levels in cardiomyocytes from HFD-fed WT mice. Mitochondrial respiration was altered in NOS3(-/-) mice but did not worsen after HFD and TAC. In contrast with its protective role in SD, NOS3 increases LV adverse remodeling after pressure overload in HFD-fed, insulin resistant mice. Interactions between NOS3 and mitochondria may be responsible for increased oxidative stress levels in HFD-fed WT mice hearts.
Asunto(s)
Insuficiencia Cardíaca/etiología , Hipertensión/complicaciones , Resistencia a la Insulina , Miocitos Cardíacos/enzimología , Óxido Nítrico Sintasa de Tipo III/deficiencia , Disfunción Ventricular Izquierda/etiología , Función Ventricular Izquierda , Remodelación Ventricular , Adenosina Trifosfato/metabolismo , Animales , Grasas de la Dieta/administración & dosificación , Modelos Animales de Enfermedad , Metabolismo Energético , Inhibidores Enzimáticos/farmacología , Regulación Enzimológica de la Expresión Génica , Insuficiencia Cardíaca/enzimología , Insuficiencia Cardíaca/genética , Insuficiencia Cardíaca/fisiopatología , Peróxido de Hidrógeno/metabolismo , Hipertensión/diagnóstico por imagen , Hipertensión/enzimología , Hipertensión/genética , Hipertensión/fisiopatología , Resistencia a la Insulina/genética , Masculino , Ratones , Ratones Endogámicos C57BL , Ratones Noqueados , Mitocondrias Cardíacas/enzimología , Miocitos Cardíacos/efectos de los fármacos , Miocitos Cardíacos/patología , Óxido Nítrico Sintasa de Tipo III/antagonistas & inhibidores , Óxido Nítrico Sintasa de Tipo III/genética , Estrés Oxidativo , Fosforilación , Serina , Factores de Tiempo , Ultrasonografía , Desacopladores/farmacología , Disfunción Ventricular Izquierda/diagnóstico por imagen , Disfunción Ventricular Izquierda/enzimología , Disfunción Ventricular Izquierda/genética , Disfunción Ventricular Izquierda/fisiopatología , Función Ventricular Izquierda/efectos de los fármacos , Remodelación Ventricular/efectos de los fármacosRESUMEN
Non-alcoholic fatty liver disease (NAFLD) affects over 30% of adults in the United States. Bone morphogenetic protein (BMP) signaling is known to contribute to hepatic fibrosis, but the role of BMP signaling in the development of NAFLD is unclear. In this study, treatment with either of two BMP inhibitors reduced hepatic triglyceride content in diabetic (db/db) mice. BMP inhibitor-induced decrease in hepatic triglyceride levels was associated with decreased mRNA encoding Dgat2, an enzyme integral to triglyceride synthesis. Treatment of hepatoma cells with BMP2 induced DGAT2 expression and activity via intracellular SMAD signaling. In humans we identified a rare missense single nucleotide polymorphism in the BMP type 1 receptor ALK6 (rs34970181;R371Q) associated with a 2.1-fold increase in the prevalence of NAFLD. In vitro analyses revealed R371Q:ALK6 is a previously unknown constitutively active receptor. These data show that BMP signaling is an important determinant of NAFLD in a murine model and is associated with NAFLD in humans.
Asunto(s)
Proteínas Morfogenéticas Óseas/metabolismo , Enfermedad del Hígado Graso no Alcohólico/metabolismo , Enfermedad del Hígado Graso no Alcohólico/patología , Transducción de Señal , Animales , Biomarcadores/sangre , Línea Celular Tumoral , Diacilglicerol O-Acetiltransferasa/metabolismo , Regulación de la Expresión Génica/efectos de los fármacos , Metabolismo de los Lípidos/efectos de los fármacos , Ratones , Enfermedad del Hígado Graso no Alcohólico/sangre , Enfermedad del Hígado Graso no Alcohólico/tratamiento farmacológico , Pirazoles/farmacología , Pirazoles/uso terapéutico , Pirimidinas/farmacología , Pirimidinas/uso terapéutico , Transducción de Señal/efectos de los fármacos , Proteínas Smad/metabolismoRESUMEN
Cardiovascular disease is the leading cause of morbidity and mortality in the world. Atherosclerotic plaques, consisting of lipid-laden macrophages and calcification, develop in the coronary arteries, aortic valve, aorta, and peripheral conduit arteries and are the hallmark of cardiovascular disease. In humans, imaging with computed tomography allows for the quantification of vascular calcification; the presence of vascular calcification is a strong predictor of future cardiovascular events. Development of novel therapies in cardiovascular disease relies critically on improving our understanding of the underlying molecular mechanisms of atherosclerosis. Advancing our knowledge of atherosclerotic mechanisms relies on murine and cell-based models. Here, a method for imaging aortic calcification and macrophage infiltration using two spectrally distinct near-infrared fluorescent imaging probes is detailed. Near-infrared fluorescent imaging allows for the ex vivo quantification of calcification and macrophage accumulation in the entire aorta and can be used to further our understanding of the mechanistic relationship between inflammation and calcification in atherosclerosis. Additionally, a method for isolating and culturing animal aortic vascular smooth muscle cells and a protocol for inducing calcification in cultured smooth muscle cells from either murine aortas or from human coronary arteries is described. This in vitro method of modeling vascular calcification can be used to identify and characterize the signaling pathways likely important for the development of vascular disease, in the hopes of discovering novel targets for therapy.
Asunto(s)
Calcinosis/diagnóstico por imagen , Músculo Liso Vascular/diagnóstico por imagen , Animales , Enfermedades de la Aorta/diagnóstico por imagen , Enfermedades de la Aorta/etiología , Enfermedades de la Aorta/metabolismo , Aterosclerosis/diagnóstico por imagen , Aterosclerosis/etiología , Aterosclerosis/metabolismo , Humanos , Interpretación de Imagen Asistida por Computador , Inflamación , Ratones , Músculo Liso Vascular/metabolismo , Músculo Liso Vascular/patologíaRESUMEN
OBJECTIVE: Matrix Gla protein (MGP) is reported to inhibit bone morphogenetic protein (BMP) signal transduction. MGP deficiency is associated with medial calcification of the arterial wall, in a process that involves both osteogenic transdifferentiation of vascular smooth muscle cells (VSMCs) and mesenchymal transition of endothelial cells (EndMT). In this study, we investigated the contribution of BMP signal transduction to the medial calcification that develops in MGP-deficient mice. APPROACH AND RESULTS: MGP-deficient mice (MGP(-/-)) were treated with one of two BMP signaling inhibitors, LDN-193189 or ALK3-Fc, beginning one day after birth. Aortic calcification was assessed in 28-day-old mice by measuring the uptake of a fluorescent bisphosphonate probe and by staining tissue sections with Alizarin red. Aortic calcification was 80% less in MGP(-/-) mice treated with LDN-193189 or ALK3-Fc compared with vehicle-treated control animals (P<0.001 for both). LDN-193189-treated MGP(-/-) mice survived longer than vehicle-treated MGP(-/-) mice. Levels of phosphorylated Smad1/5 and Id1 mRNA (markers of BMP signaling) did not differ in the aortas from MGP(-/-) and wild-type mice. Markers of EndMT and osteogenesis were increased in MGP(-/-) aortas, an effect that was prevented by LDN-193189. Calcification of isolated VSMCs was also inhibited by LDN-193189. CONCLUSIONS: Inhibition of BMP signaling leads to reduced vascular calcification and improved survival in MGP(-/-) mice. The EndMT and osteogenic transdifferentiation associated with MGP deficiency is dependent upon BMP signaling. These results suggest that BMP signal transduction has critical roles in the development of vascular calcification in MGP-deficient mice.
Asunto(s)
Proteínas de Unión al Calcio/metabolismo , Proteínas de la Matriz Extracelular/metabolismo , Calcificación Vascular/tratamiento farmacológico , Animales , Proteínas Morfogenéticas Óseas/metabolismo , Proteínas de Unión al Calcio/genética , Proteínas de la Matriz Extracelular/genética , Técnica del Anticuerpo Fluorescente , Ratones , Ratones Noqueados , Pirazoles/farmacología , Pirazoles/uso terapéutico , Pirimidinas/farmacología , Pirimidinas/uso terapéutico , Transducción de Señal/efectos de los fármacos , Transducción de Señal/genética , Calcificación Vascular/genética , Proteína Gla de la MatrizRESUMEN
Nitric oxide (NO) plays an essential role in regulating hypertension and blood flow by inducing relaxation of vascular smooth muscle. Male mice deficient in a NO receptor component, the α1 subunit of soluble guanylate cyclase (sGCα1), are prone to hypertension in some, but not all, mouse strains, suggesting that additional genetic factors contribute to the onset of hypertension. Using linkage analyses, we discovered a quantitative trait locus (QTL) on chromosome 1 that was linked to mean arterial pressure (MAP) in the context of sGCα1 deficiency. This region is syntenic with previously identified blood pressure-related QTLs in the human and rat genome and contains the genes coding for renin. Hypertension was associated with increased activity of the renin-angiotensin-aldosterone system (RAAS). Further, we found that RAAS inhibition normalized MAP and improved endothelium-dependent vasorelaxation in sGCα1-deficient mice. These data identify the RAAS as a blood pressure-modifying mechanism in a setting of impaired NO/cGMP signaling.