RESUMEN
Glucagon-like peptide-1 receptor agonists (GLP-1RAs) exert potent glucose-lowering effects without increasing risks for hypoglycemia and weight gain. Preclinical studies have demonstrated direct antiatherogenic effects of GLP-1RAs in normoglycemic animal models; however, the underlying mechanisms in hyperglycemic conditions have not been fully clarified. Here we aimed to elucidate the role of AMP-activated protein kinase (AMPK) in antiatherogenic effects of GLP-1RAs in hyperglycemic mice. Streptozotocin-induced hyperglycemic apolipoprotein E-null mice were treated with vehicle, low-dose liraglutide (17 nmol·kg-1·day-1), or high-dose liraglutide (107 nmol·kg-1·day-1) in experiment 1 and the AMPK inhibitor dorsomorphin, dorsomorphin + low-dose liraglutide, or dorsomorphin + high-dose liraglutide in experiment 2. Four weeks after treatment, aortas were collected to assess atherosclerosis. In experiment 1, metabolic parameters were similar among the groups. Assessment of atherosclerosis revealed that high-dose liraglutide treatments reduced lipid deposition on the aortic surface and plaque volume and intraplaque macrophage accumulation at the aortic sinus. In experiment 2, liraglutide-induced AMPK phosphorylation in the aorta was abolished by dorsomorphin; however, the antiatherogenic effects of high-dose liraglutide were preserved. In cultured human umbilical vein endothelial cells, liraglutide suppressed tumor necrosis factor-induced expression of proatherogenic molecules; these effects were maintained under small interfering RNA-mediated knockdown of AMPKα1 and in the presence of dorsomorphin. Conversely, in human monocytic U937 cells, the anti-inflammatory effects of liraglutide were abolished by dorsomorphin. In conclusion, liraglutide exerted AMPK-independent antiatherogenic effects in hyperlipidemic mice with streptozotocin-induced hyperglycemia, with the possible involvement of AMPK-independent suppression of proatherogenic molecules in vascular endothelial cells.
Asunto(s)
Proteínas Quinasas Activadas por AMP/genética , Diabetes Mellitus Experimental/metabolismo , Hipoglucemiantes/farmacología , Liraglutida/farmacología , Placa Aterosclerótica/patología , Seno Aórtico/efectos de los fármacos , Proteínas Quinasas Activadas por AMP/metabolismo , Animales , Aorta/efectos de los fármacos , Aorta/metabolismo , Aorta/patología , Aterosclerosis/metabolismo , Aterosclerosis/patología , Técnicas de Silenciamiento del Gen , Hiperglucemia/metabolismo , Macrófagos , Ratones , Ratones Noqueados para ApoE , Placa Aterosclerótica/metabolismo , Seno Aórtico/metabolismo , Seno Aórtico/patologíaRESUMEN
BACKGROUND AND AIMS: Recent studies have suggested that heat shock protein 70 (HSP70) may play critical roles in cardiovascular disease. However, the effects of HSP70 on the development of atherosclerosis in apoE-/- mice remain largely unknown. This study was to investigate the role and potential mechanism of HSP70 in atherosclerosis. METHODS: HSP70 was overexpressed in apoE-/- mice and THP-1-derived macrophages with lentiviral vectors. Oil Red O, hematoxylin-eosin, and Masson staining were performed to evaluate atherosclerotic plaque in apoE-/- mice fed the Western type diet. Moreover, immunostaining was employed to detect the expression of relative proteins in aortic sinus. Reporter gene and chromatin immunoprecipitation were performed to analyze the effect of Elk-1 on the promoter activity of ABCA1 and ABCG1; [3H] labeled cholesterol was used to assess the capacity of cholesterol efflux and reverse cholesterol transport (RCT). RESULTS: Our results showed that HSP70 increased lipid accumulation in arteries and promoted the formation of atherosclerotic lesion. The capacity of cholesterol efflux was reduced in peritoneal macrophages isolated from HSP70-overexpressed apoE-/- mice. The levels of ABCA1 and ABCG1 expression were also reduced in the peritoneal macrophages and the aorta from apoE-/- mice in response to HSP70. The c-Jun N-terminal kinase (JNK) and ETS transcription factor (Elk-1) played a critical role in HSP70-induced downregulation ABCA1 and ABCG1. Further, HSP70 reduced RCT from macrophages to plasma, liver, and feces in apoE-/- mice. CONCLUSIONS: HSP70 promotes the progression of atherosclerosis in apoE-/- mice by suppressing the expression of ABCA1 and ABCG1 through the JNK/Elk-1 pathway.
Asunto(s)
Transportador 1 de Casete de Unión a ATP/metabolismo , Transportador de Casetes de Unión a ATP, Subfamilia G, Miembro 1/metabolismo , Aterosclerosis/patología , Proteínas HSP70 de Choque Térmico/metabolismo , Transportador 1 de Casete de Unión a ATP/genética , Transportador de Casetes de Unión a ATP, Subfamilia G, Miembro 1/genética , Animales , Aterosclerosis/etiología , Línea Celular , Colesterol/metabolismo , Dieta Occidental/efectos adversos , Modelos Animales de Enfermedad , Progresión de la Enfermedad , Regulación hacia Abajo , Humanos , Sistema de Señalización de MAP Quinasas , Macrófagos , Masculino , Ratones , Ratones Noqueados para ApoE , Placa Aterosclerótica/metabolismo , Placa Aterosclerótica/patología , Regiones Promotoras Genéticas , Seno Aórtico/metabolismo , Seno Aórtico/patología , Proteína Elk-1 con Dominio ets/metabolismoRESUMEN
BACKGROUND/AIMS: Endothelial cell dysfunction is the principal pathological process underlying atherosclerotic cardiovascular disease. G protein-coupled receptor 124 (GPR124), an orphan receptor in the adhesion GPCR subfamily, promotes angiogenesis in the brain. In the present study, we explored the role of endothelial GPR124 in the development and progression of atherosclerosis in adult mice. METHODS: Using tetracycline-inducible transgenic systems, we generated mice expressing GPR124 specifically under control of the Tie-2 promoter. The animal model of atherosclerosis was constructed by intravenously injecting AAV-PCSK9DY into tetracycline-regulated mice and feeding the mice a high-fat diet for 16 consecutive weeks. Biochemical analysis and immunohistochemistry methods were used to address the role and mechanism of GPR124 in the pathological process of atherosclerosis. RESULTS: Higher TC (total cholesterol) and LDL-C (low density lipoprotein cholesterol) levels in serum and greater lipid deposition in the aortic sinus were found in atherosclerotic mice with GPR124 overexpression, coincident with the elevated proliferation of smooth muscle cells. We observed an elevation of ONOO- in the aortic sinus in this model by using immunofluorescence, and the experiments showed that the specific overexpression of GPR124 in the endothelium induced the up-regulation of CD68, NLRP3 and caspase-1 levels in the aortic sinus. CONCLUSION: The above results indicate that manipulating GPR124 in the endothelium may contribute to delayed pathological progression of atherosclerosis.
Asunto(s)
Aterosclerosis/patología , Receptores Acoplados a Proteínas G/metabolismo , Animales , Antígenos CD/metabolismo , Antígenos de Diferenciación Mielomonocítica/metabolismo , Aterosclerosis/metabolismo , Caspasa 1/metabolismo , Colesterol/sangre , LDL-Colesterol/sangre , Dieta Alta en Grasa , Modelos Animales de Enfermedad , Humanos , Inflamación/etiología , Ratones , Ratones Endogámicos C57BL , Ratones Transgénicos , Miocitos del Músculo Liso/citología , Miocitos del Músculo Liso/efectos de los fármacos , Miocitos del Músculo Liso/metabolismo , Proteína con Dominio Pirina 3 de la Familia NLR/metabolismo , Ácido Peroxinitroso/metabolismo , Plásmidos/genética , Plásmidos/metabolismo , Receptores Acoplados a Proteínas G/genética , Seno Aórtico/metabolismo , Seno Aórtico/patologíaRESUMEN
OBJECTIVE: VWF (von Willebrand factor) is synthesized by endothelial cells and megakaryocytes and is known to contribute to atherosclerosis. In vitro studies suggest that platelet-derived VWF (Plt-VWF) is biochemically and functionally different from endothelial cell-derived VWF (EC-VWF). We determined the role of different pools of VWF in the pathophysiology of atherosclerosis. APPROACH AND RESULTS: Using bone marrow transplantation, we generated chimeric Plt-VWF, EC-VWF, and Plt-VWF mice lacking a disintegrin and metalloprotease with thrombospondin type I repeats-13 in platelets and plasma on apolipoprotein E-deficient (Apoe-/-) background. Controls were chimeric Apoe-/- mice transplanted with bone marrow from Apoe-/- mice (wild type) and Vwf-/-Apoe-/- mice transplanted with bone marrow from Vwf-/-Apoe-/- mice (VWF-knock out). Susceptibility to atherosclerosis was evaluated in whole aortae and cross-sections of the aortic sinus in female mice fed a high-fat Western diet for 14 weeks. VWF-knock out, Plt-VWF, and Plt-VWF mice lacking a disintegrin and metalloprotease with thrombospondin type I repeats-13 exhibited reduced plaque size characterized by smaller necrotic cores, reduced neutrophil and monocytes/macrophages content, decreased MMP9 (matrix metalloproteinase), MMP2, and CX3CL1 (chemokine [C-X3-C motif] ligand 1)-positive area, and abundant interstitial collagen (P<0.05 versus wild-type or EC-VWF mice). Atherosclerotic lesion size and composition were comparable between wild-type or EC-VWF mice. Together these findings suggest that EC-VWF, but not Plt-VWF, promotes atherosclerosis exacerbation. Furthermore, intravital microscopy experiments revealed that EC-VWF, but not Plt-VWF, contributes to platelet and leukocyte adhesion under inflammatory conditions at the arterial shear rate. CONCLUSIONS: EC-VWF, but not Plt-VWF, contributes to VWF-dependent atherosclerosis by promoting platelet adhesion and vascular inflammation. Plt-VWF even in the absence of a disintegrin and metalloprotease with thrombospondin type I repeats-13, both in platelet and plasma, was not sufficient to promote atherosclerosis.
Asunto(s)
Aorta/metabolismo , Enfermedades de la Aorta/metabolismo , Aterosclerosis/metabolismo , Células Endoteliales/metabolismo , Enfermedades de von Willebrand/metabolismo , Factor de von Willebrand/metabolismo , Proteína ADAMTS13/genética , Proteína ADAMTS13/metabolismo , Animales , Aorta/patología , Enfermedades de la Aorta/sangre , Enfermedades de la Aorta/genética , Enfermedades de la Aorta/patología , Aterosclerosis/sangre , Aterosclerosis/genética , Aterosclerosis/patología , Plaquetas/metabolismo , Trasplante de Médula Ósea , Adhesión Celular , Dieta Alta en Grasa , Modelos Animales de Enfermedad , Células Endoteliales/patología , Femenino , Leucocitos/metabolismo , Leucocitos/patología , Ratones Endogámicos C57BL , Ratones Noqueados para ApoE , Placa Aterosclerótica , Adhesividad Plaquetaria , Seno Aórtico/metabolismo , Seno Aórtico/patología , Enfermedades de von Willebrand/sangre , Enfermedades de von Willebrand/genética , Factor de von Willebrand/genéticaRESUMEN
OBJECTIVE: Rosuvastatin has been widely used in the primary and secondary prevention of coronary heart disease. However, its antiatherosclerotic properties have not been tested in a mouse model that could mimic human coronary heart disease. The present study was designed to test the effects of rosuvastatin on coronary artery atherosclerosis and myocardial fibrosis in SR-B1 (scavenger receptor class B type 1) and apoE (apolipoprotein E) double knockout mice. APPROACH AND RESULTS: Three-week-old SR-B1-/-/apoE-/- mice were injected daily with 10 mg/kg of rosuvastatin for 2 weeks. Compared with saline-treated mice, rosuvastatin-treated mice showed increased levels of hepatic PCSK9 (proprotein convertase subtilisin/kexin type-9) and LDLR (low-density lipoprotein receptor) message, increased plasma PCSK9 protein but decreased levels of hepatic LDLR protein and increased plasma total cholesterol associated with apoB (apolipoprotein B) 48-containing lipoproteins. In spite of this, rosuvastatin treatment was associated with decreased atherosclerosis in both the aortic sinus and coronary arteries and reduced platelet accumulation in atherosclerotic coronary arteries. Cardiac fibrosis and cardiomegaly were also attenuated in rosuvastatin-treated SR-B1-/-/apoE-/- mice. Two-week treatment with rosuvastatin resulted in significant decreases in markers of oxidized phospholipids in atherosclerotic plaques. In vitro analysis showed that incubation of bone marrow-derived macrophages with rosuvastatin substantially downregulated cluster of differentiation (CD)36 and inhibited oxidized LDL-induced foam cell formation. CONCLUSIONS: Rosuvastatin protected SR-B1-/-/apoE-/- mice against atherosclerosis and platelet accumulation in coronary arteries and attenuated myocardial fibrosis and cardiomegaly, despite increased plasma total cholesterol. The ability of rosuvastatin to reduce oxidized phospholipids in atherosclerotic plaques and inhibit macrophage foam cell formation may have contributed to this protection.
Asunto(s)
Enfermedades de la Aorta/prevención & control , Aterosclerosis/prevención & control , Colesterol/sangre , Enfermedad de la Arteria Coronaria/prevención & control , Vasos Coronarios/efectos de los fármacos , Inhibidores de Hidroximetilglutaril-CoA Reductasas/farmacología , Placa Aterosclerótica , Rosuvastatina Cálcica/farmacología , Receptores Depuradores de Clase B/deficiencia , Seno Aórtico/efectos de los fármacos , Animales , Enfermedades de la Aorta/genética , Enfermedades de la Aorta/metabolismo , Enfermedades de la Aorta/patología , Aterosclerosis/genética , Aterosclerosis/metabolismo , Aterosclerosis/patología , Biomarcadores/sangre , Células Cultivadas , Enfermedad de la Arteria Coronaria/genética , Enfermedad de la Arteria Coronaria/metabolismo , Enfermedad de la Arteria Coronaria/patología , Vasos Coronarios/metabolismo , Vasos Coronarios/patología , Modelos Animales de Enfermedad , Células Espumosas/efectos de los fármacos , Células Espumosas/metabolismo , Células Espumosas/patología , Lipoproteínas LDL/metabolismo , Hígado/efectos de los fármacos , Hígado/metabolismo , Ratones Noqueados para ApoE , Proproteína Convertasa 9/metabolismo , Receptores de LDL/metabolismo , Receptores Depuradores de Clase B/genética , Seno Aórtico/metabolismo , Seno Aórtico/patologíaRESUMEN
Endothelial cells (EC) play a key role in atherosclerosis. Although EC are in constant contact with low density lipoproteins (LDL), how EC process LDL and whether this influences atherogenesis, is unclear. Here we show that EC take up and metabolize LDL, and when overburdened with intracellular cholesterol, generate cholesterol crystals (CC). The CC are deposited on the basolateral side, and compromise endothelial function. When hyperlipidemic mice are given a high fat diet, CC appear in aortic sinus within 1 week. Treatment with cAMP-enhancing agents, forskolin/rolipram (F/R), mitigates effects of CC on endothelial function by not only improving barrier function, but also inhibiting CC formation both in vitro and in vivo. A proof of principle study using F/R incorporated into liposomes, designed to target inflamed endothelium, shows reduced atherosclerosis and CC formation in ApoE -/- mice. Our findings highlight an important mechanism by which EC contribute to atherogenesis under hyperlipidemic conditions.
Asunto(s)
Apolipoproteínas E/metabolismo , Aterosclerosis/metabolismo , Colesterol/metabolismo , Células Endoteliales/metabolismo , Hiperlipidemias/metabolismo , Animales , Aorta/metabolismo , Apolipoproteínas E/genética , Arterias Carótidas/metabolismo , Células Cultivadas , Colforsina/farmacología , Dieta Alta en Grasa , Endotelio Vascular/metabolismo , Femenino , Arteria Femoral/metabolismo , Humanos , Lipoproteínas LDL/metabolismo , Liposomas/química , Masculino , Ratones , Ratones Endogámicos C57BL , Ratones Transgénicos , Rolipram/farmacología , Seno Aórtico/metabolismoRESUMEN
Di'ao Xinxuekang (XXK) is an herbal product in China and the Netherlands that has been clinically shown to attenuate atherosclerosis; however, the underlying antiatherosclerotic mechanism remains unclear. Because of its role in cholesterol homeostasis, reverse cholesterol transport (RCT) is a potential target for these beneficial effects. This study investigated the effects of XXK on RCT and related proteins. After treating ApoE-deficient mice with XXK for 8 weeks, we observed an increase in the expression level of ATP-binding cassette transporter A1 and ATP-binding cassette transporter G1, which in turn stimulated cholesterol efflux and reduced aortic atherosclerotic lesion area. XXK also increased high-density lipoprotein (HDL) synthesis by modulating the peroxisome proliferator-activated receptor γ/liver X receptor α/ATP-binding cassette transporter A1 pathway and promoted HDL maturity by increasing serum lecithin-cholesterol acyltransferase. In addition, XXK improved the selective uptake of HDL-cholesteryl ester by increasing the expression of scavenger receptor class B type I. This is the first study to show that XXK confers a regulation of RCT, at least in part, by improving HDL synthesis, maturation, and catabolism.
Asunto(s)
Colesterol/metabolismo , Medicamentos Herbarios Chinos/farmacología , Lipoproteínas HDL/biosíntesis , Animales , Transporte Biológico/efectos de los fármacos , Transporte Biológico/fisiología , Masculino , Metabolismo/efectos de los fármacos , Metabolismo/fisiología , Ratones , Ratones Endogámicos C57BL , Ratones Noqueados , Seno Aórtico/efectos de los fármacos , Seno Aórtico/metabolismoRESUMEN
BACKGROUND AND AIMS: Chinese dragon's blood has been used to treat blood stasis for thousands of years. Its total phenolic extract (Longxuetongluo capsule, LTC) is used for the treatment of ischemic stroke; however, its protective effect against atherosclerosis remains poorly understood. This paper aims to investigate the antiatherosclerotic effect of LTC and the underlying mechanisms in high-fat diet (HFD)-induced ApoE-/- mice. METHODS: The levels of plasma lipid and areas of atherosclerotic lesions in the aortic sinus in ApoE-/- mice were evaluated. The effect of LTC on the nitric oxide (NO) production in oxidized low-density lipoprotein (ox-LDL)-stimulated human umbilical vein endothelial cells (HUVECs) was determined. The adhesion of monocytes to ox-LDL-stimulated HUVECs was further studied. RESULTS: LTC at low, medium, and high doses markedly decreased the atherosclerotic lesion areas of the aortic sinus in HFD-induced ApoE-/- mice by 26.4% (p < 0.05), 30.1% (p < 0.05), and 46.5% (p < 0.01), respectively, although it did not improve the dyslipidemia. Furthermore, LTC restored the diminished NO production of ox-LDL-stimulated HUVECs (p < 0.001) and inhibited the adhesion between monocytes and endothelial cells (p < 0.01). LTC appeared to alleviate ox-LDL-stimulated dysfunction of HUVECs, and inhibit the adhesion of monocytes to HUVECs via the MAPK/IKK/IκB/NF-κB signaling pathway, thus decrease atherosclerotic lesions in the aortic sinus in HFD-induced ApoE-/- mice. CONCLUSIONS: These findings suggest the potential of LTC for use as an effective agent against atherosclerosis.
Asunto(s)
Enfermedades de la Aorta/prevención & control , Apolipoproteínas E/deficiencia , Aterosclerosis/prevención & control , Dieta Alta en Grasa , Medicamentos Herbarios Chinos/farmacología , Endotelio Vascular/efectos de los fármacos , Seno Aórtico/efectos de los fármacos , Animales , Enfermedades de la Aorta/genética , Enfermedades de la Aorta/metabolismo , Enfermedades de la Aorta/patología , Apolipoproteínas E/genética , Aterosclerosis/genética , Aterosclerosis/metabolismo , Aterosclerosis/patología , Adhesión Celular/efectos de los fármacos , Línea Celular Transformada , Línea Celular Tumoral , Técnicas de Cocultivo , Modelos Animales de Enfermedad , Progresión de la Enfermedad , Relación Dosis-Respuesta a Droga , Endotelio Vascular/metabolismo , Endotelio Vascular/patología , Predisposición Genética a la Enfermedad , Células Endoteliales de la Vena Umbilical Humana/efectos de los fármacos , Células Endoteliales de la Vena Umbilical Humana/metabolismo , Lipoproteínas LDL/farmacología , Masculino , Ratones Noqueados , Proteínas Quinasas Activadas por Mitógenos/metabolismo , FN-kappa B/metabolismo , Óxido Nítrico/metabolismo , Óxido Nítrico Sintasa de Tipo III/metabolismo , Fenotipo , Placa Aterosclerótica , Transducción de Señal/efectos de los fármacos , Seno Aórtico/metabolismo , Seno Aórtico/patología , Factores de TiempoRESUMEN
It has been argued whether insulin accelerates or prevents atherosclerosis. Although results from in vitro studies have been conflicting, recent in vivo mice studies demonstrated antiatherogenic effects of insulin. Insulin is a known activator of endothelial nitric oxide synthase (NOS), leading to increased production of NO, which has potent antiatherogenic effects. We aimed to examine the role of NOS in the protective effects of insulin against atherosclerosis. Male apolipoprotein E-null mice (8 wk old) fed a high-cholesterol diet (1.25% cholesterol) were assigned to the following 12-wk treatments: control, insulin (0.05 U/day via subcutaneous pellet), N(ω)-nitro-l-arginine methyl ester hydrochloride (l-NAME, via drinking water at 100 mg/l), and insulin plus l-NAME. Insulin reduced atherosclerotic plaque burden in the descending aorta by 42% compared with control (plaque area/aorta lumen area: control, 16.5 ± 1.9%; insulin, 9.6 ± 1.3%, P < 0.05). Although insulin did not decrease plaque burden in the aortic sinus, macrophage accumulation in the plaque was decreased by insulin. Furthermore, insulin increased smooth muscle actin and collagen content and decreased plaque necrosis, consistent with increased plaque stability. In addition, insulin treatment increased plasma NO levels, decreased inducible NOS staining, and tended to increase phosphorylated vasodilator-stimulated phosphoprotein staining in the plaques of the aortic sinus. All these effects of insulin were abolished by coadministration of l-NAME, whereas l-NAME alone showed no effect. Insulin also tended to increase phosphorylated endothelial NOS and total neuronal NOS staining, effects not modified by l-NAME. In conclusion, we demonstrate that insulin treatment decreases atherosclerotic plaque burden and increases plaque stability through NOS-dependent mechanisms.
Asunto(s)
Aorta/efectos de los fármacos , Inhibidores Enzimáticos/farmacología , Hipoglucemiantes/farmacología , Insulina/farmacología , NG-Nitroarginina Metil Éster/farmacología , Óxido Nítrico Sintasa/efectos de los fármacos , Placa Aterosclerótica/metabolismo , Actinas/efectos de los fármacos , Actinas/metabolismo , Animales , Aorta/metabolismo , Aorta/patología , Apolipoproteínas E/genética , Colágeno/efectos de los fármacos , Colágeno/metabolismo , Macrófagos/efectos de los fármacos , Macrófagos/patología , Masculino , Ratones , Ratones Noqueados , Necrosis , Óxido Nítrico Sintasa/metabolismo , Óxido Nítrico Sintasa de Tipo I/efectos de los fármacos , Óxido Nítrico Sintasa de Tipo I/metabolismo , Óxido Nítrico Sintasa de Tipo III/efectos de los fármacos , Óxido Nítrico Sintasa de Tipo III/metabolismo , Fosfoproteínas/efectos de los fármacos , Fosfoproteínas/metabolismo , Placa Aterosclerótica/patología , Seno Aórtico/efectos de los fármacos , Seno Aórtico/metabolismo , Seno Aórtico/patologíaRESUMEN
BACKGROUND: A number of epidemiological studies demonstrated that postprandial hyperglycemia is a risk factor for cardiovascular disease in individuals with impaired glucose tolerance. Although several laboratory studies have addressed the plausible causal role of postprandial acute hyperglycemia (glucose spikes) in the development of atherosclerosis, there is little convincing evidence in vivo whether the atherosclerotic lesion formation can be accelerated solely by glucose spikes. Here, we assessed the effect of repetitive glucose spikes on atherosclerotic lesion formation in mice. METHODS: Female C57BL/6 mice were fed an atherogenic diet from 8 to 28 weeks of age. During the atherogenic diet feeding period, the mice orally received a glucose solution (50 mg glucose/mouse; G group) or water (W group) twice daily, 6 days a week. Atherosclerotic lesion formation in the aortic sinus was quantitatively analyzed in serial cross-sections by oil red O staining. RESULTS: G group mice showed transient increases in blood glucose level (~5 mmol/L above W group), and the levels returned to levels similar to those in W group mice within 60 min. No significant differences in glucose tolerance, insulin sensitivity, and plasma lipid profiles were observed after the 20-week repetitive administration between the 2 groups. G group mice showed an approximately 4-fold greater atherosclerotic lesion size in the aortic sinus than W group mice. Gene expression levels of Cd68 and Icam1 in the thoracic aorta were higher in G group mice than in W group mice. CONCLUSIONS: These results indicate that glucose spikes can accelerate atherosclerotic lesion formation, with little influence on other metabolic disorders. Repetitive glucose administration in wild-type mice may serve as a simple and useful approach to better understanding the causal role of glycemic spikes in the development of atherosclerosis.
Asunto(s)
Aterosclerosis/metabolismo , Aterosclerosis/patología , Glucemia/metabolismo , Animales , Aorta Torácica/metabolismo , Aorta Torácica/patología , Aterosclerosis/genética , Peso Corporal , Dieta Aterogénica/efectos adversos , Femenino , Regulación de la Expresión Génica , Glucosa/administración & dosificación , Prueba de Tolerancia a la Glucosa , Insulina/sangre , Resistencia a la Insulina , Lípidos/sangre , Ratones Endogámicos C57BL , Seno Aórtico/metabolismo , Seno Aórtico/patologíaRESUMEN
RATIONALE: CD4(+) natural killer T (NKT) cells augment atherosclerosis in apolipoprotein E-deficient (ApoE)(-/-) mice but their mechanisms of action are unknown. OBJECTIVES: We investigated the roles of bystander T, B, and NK cells; NKT cell-derived interferon-γ, interleukin (IL)-4, and IL-21 cytokines; and NKT cell-derived perforin and granzyme B cytotoxins in promoting CD4(+) NKT cell atherogenicity. METHODS AND RESULTS: Transfer of CD4(+) NKT cells into T- and B-cell-deficient ApoE(-/-)Rag2(-/-) mice augmented aortic root atherosclerosis by ≈75% that was ≈30% of lesions in ApoE(-/-) mice; macrophage accumulation similarly increased. Transferred NKT cells were identified in the liver and atherosclerotic lesions of recipient mice. Transfer of CD4(+) NKT cells into T-, B-cell-deficient, and NK cell-deficient ApoE(-/-)Rag2(-/-)γC(-/-) mice also augmented atherosclerosis. These data indicate that CD4(+) NKT cells can exert proatherogenic effects independent of other lymphocytes. To investigate the role of NKT cell-derived interferon-γ, IL-4, and IL-21 cytokines and perforin and granzyme B cytotoxins, CD4(+) NKT cells from mice deficient in these molecules were transferred into NKT cell-deficient ApoE(-/-)Jα18(-/-) mice. CD4(+) NKT cells deficient in IL-4, interferon-γ, or IL-21 augmented atherosclerosis in ApoE(-/-)Jα18(-/-) mice by ≈95%, ≈80%, and ≈70%, respectively. Transfer of CD4(+) NKT cells deficient in perforin or granzyme B failed to augment atherosclerosis. Apoptotic cells, necrotic cores, and proinflammatory VCAM-1 (vascular cell adhesion molecule) and MCP-1 (monocyte chemotactic protein) were reduced in mice receiving perforin-deficient NKT cells. CD4(+) NKT cells are twice as potent as CD4(+) T cells in promoting atherosclerosis. CONCLUSIONS: CD4(+) NKT cells potently promote atherosclerosis by perforin and granzyme B-dependent apoptosis that increases postapoptotic necrosis and inflammation.
Asunto(s)
Aterosclerosis/metabolismo , Linfocitos T CD4-Positivos/metabolismo , Granzimas/deficiencia , Células T Asesinas Naturales/metabolismo , Proteínas Citotóxicas Formadoras de Poros/deficiencia , Seno Aórtico/metabolismo , Traslado Adoptivo/métodos , Animales , Aterosclerosis/inmunología , Aterosclerosis/patología , Linfocitos T CD4-Positivos/inmunología , Masculino , Ratones , Ratones Noqueados , Células T Asesinas Naturales/inmunología , Seno Aórtico/inmunología , Seno Aórtico/patologíaRESUMEN
OBJECTIVES: The CC chemokine ligand-20 (CCL-20)/macrophage inflammatory protein-3α has been seen as one of the most important chemokines and played a key role in atherogenesis, but the mechanism that underlies the regulation of CCL-20 has not been established clearly yet. The aim of this study was to investigate the influence of salvianolic acid A (SAA) on the expression of CCL-20 in macrophages and ApoE-deficient (ApoE) mice. METHODS: The expression of CCL-20 was detected both at protein and messenger RNA levels in RAW264.7 cells. We validated the result in ApoE mice that were intraperitoneally injected with SAA. Phosphorylation of p38 mitogen-activated protein kinase was detected with Western blot, and inhibitor of p38 was used to investigate the mechanism of regulation of CCL-20. Hematoxylin and eosin and Oil-Red-O staining were used to evaluate the atherosclerotic lesions and lipid accumulation in ApoE mice. Immunohistochemical analysis was used to detect the expressions of CCL-20 and CCR6 in the atherosclerotic lesions. Immunofluorescent analysis was used to certify the origination of CCL-20. RESULTS: Recombinant tumor necrosis factor-α (TNF-α) upregulated CCL-20 production in dose- and time-dependent manners in RAW264.7 cells. The activity of TNF-α-induced CCL-20 production seemed to be significantly suppressed by SAA. Using p38 mitogen-activated protein kinase inhibitor, we found that p38 mediated the effects of TNF-α- and SAA-induced CCL-20 expression changes. In addition, immunohistochemical analysis of aortic root of ApoE mice also demonstrated that the expressions of CCL-20 and CCR6 were both downregulated significantly with SAA treatment. Furthermore, treatment of SAA inhibited the progression of the atherosclerotic plaques and lipid accumulation. CONCLUSIONS: These results demonstrate that TNF-α increased but SAA suppressed CCL-20 production significantly via a novel mechanism.
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Antiinflamatorios/farmacología , Aterosclerosis/tratamiento farmacológico , Ácidos Cafeicos/farmacología , Quimiocina CCL20/genética , Lactatos/farmacología , Macrófagos/efectos de los fármacos , Factor de Necrosis Tumoral alfa/farmacología , Animales , Apolipoproteínas E/deficiencia , Apolipoproteínas E/genética , Aterosclerosis/metabolismo , Aterosclerosis/patología , Western Blotting , Línea Celular , Quimiocina CCL20/inmunología , Modelos Animales de Enfermedad , Relación Dosis-Respuesta a Droga , Metabolismo de los Lípidos/efectos de los fármacos , Macrófagos/inmunología , Ratones Endogámicos C57BL , Ratones Noqueados , Reacción en Cadena en Tiempo Real de la Polimerasa , Proteínas Recombinantes , Seno Aórtico/efectos de los fármacos , Seno Aórtico/metabolismo , Seno Aórtico/patología , Factores de Tiempo , Factor de Necrosis Tumoral alfa/inmunología , Regulación hacia ArribaRESUMEN
Our laboratory has demonstrated that the cardiomotor component of aortic baroreflex is temporarily inhibited at the onset of spontaneous motor activity in decerebrate cats, without altering carotid sinus baroreflex. A reason for this dissociation may be attributed to a difference in the responses between aortic nerve activity (AoNA) and carotid sinus nerve activity (CsNA) during spontaneous motor activity. The stimulus-response curves of AoNA and CsNA against mean arterial blood pressure (MAP) were compared between the pressor interventions evoked by spontaneous motor activity and by intravenous administration of phenylephrine or norepinephrine, in which the responses in heart rate (HR) were opposite (i.e., tachycardia vs. baroreflex bradycardia), despite the identical increase in MAP of 34-40 mmHg. In parallel to the pressor response, mean AoNA and CsNA increased similarly by 78-81 and by 88 % of the baseline control, respectively, irrespective of whether the pressor response was evoked by spontaneous motor activity or by a pharmacological intervention. The slope of the stimulus-response curve of the mean AoNA became greater (P < 0.05) during spontaneous motor activity as compared to the pharmacological intervention. On the other hand, the stimulus-response curve of the mean CsNA and its slope were equal (P > 0.05) between the two pressor interventions. Furthermore, the slopes of the stimulus-response curves of both diastolic AoNA and CsNA (defined as the minimal value within a beat) exhibited a greater increase during spontaneous motor activity. All differences in the slopes of the stimulus-response curves were abolished by restraining HR at the intrinsic cardiac frequency. In conclusion, mean mass activities of both aortic and carotid sinus baroreceptors are able to encode the beat-by-beat changes in MAP not only at rest but also during spontaneous motor activity and spontaneous motor activity-related reduction of aortic baroreceptor activity is denied accordingly.
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Seno Carotídeo/efectos de los fármacos , Seno Carotídeo/fisiología , Actividad Motora/fisiología , Presorreceptores/metabolismo , Seno Aórtico/efectos de los fármacos , Seno Aórtico/fisiología , Vasoconstrictores/farmacología , Animales , Barorreflejo/efectos de los fármacos , Barorreflejo/fisiología , Presión Sanguínea/efectos de los fármacos , Presión Sanguínea/fisiología , Seno Carotídeo/metabolismo , Gatos , Corazón/efectos de los fármacos , Frecuencia Cardíaca/efectos de los fármacos , Frecuencia Cardíaca/fisiología , Actividad Motora/efectos de los fármacos , Norepinefrina/farmacología , Fenilefrina/farmacología , Seno Aórtico/metabolismoRESUMEN
BACKGROUND: The lipid milleu exacerbates the inflammatory response in atherosclerosis but its effect on T cell mediated immune response has not been fully elucidated. We hypothesized that lipid lowering would modulate T cell mediated immune function. METHODS AND RESULTS: T cells isolated from human PBMC or splenic T cells from apoE-/- mouse had higher proliferative response to T cell receptor (TCR) ligation in medium supplemented with 10% fetal bovine serum (FBS) compared to medium with 10% delipidated FBS. The differences in proliferation were associated with changes in lipid rafts, cellular cholesterol content, IL-10 secretion and subsequent activation of signaling molecule activated by TCR ligation. Immune biomarkers were also assessed in vivo using male apoE-/- mice fed atherogenic diet (AD) starting at 7 weeks of age. At 25 weeks of age, a sub-group was switched to normal diet (ND) whereas the rest remained on AD until euthanasia at 29 weeks of age. Dietary change resulted in a lower circulating level of cholesterol, reduced plaque size and inflammatory phenotype of plaques. These changes were associated with reduced intracellular IL-10 and IL-12 expression in CD4+ and CD8+ T cells. CONCLUSION: Our results show that lipid lowering reduces T cell proliferation and function, supporting the notion that lipid lowering modulates T cell function in vivo and in vitro.
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Colesterol/metabolismo , Linfocitos T/inmunología , Animales , Apolipoproteínas E/deficiencia , Apolipoproteínas E/metabolismo , Western Blotting , Proliferación Celular/efectos de los fármacos , Separación Celular , Colesterol/sangre , Medios de Cultivo/farmacología , Grasas de la Dieta/farmacología , Ensayo de Inmunoadsorción Enzimática , Esterificación , Humanos , Interleucina-10/metabolismo , Activación de Linfocitos/efectos de los fármacos , Masculino , Microdominios de Membrana/efectos de los fármacos , Microdominios de Membrana/metabolismo , Ratones Endogámicos C57BL , Fosforilación/efectos de los fármacos , Placa Aterosclerótica/patología , Seno Aórtico/efectos de los fármacos , Seno Aórtico/metabolismo , Seno Aórtico/patología , Linfocitos T/citología , Linfocitos T/efectos de los fármacos , Proteína Tirosina Quinasa ZAP-70/metabolismoRESUMEN
To investigate the pathologic mechanisms of toll-like receptor 4 (TLR4) in lung injury and atherosclerosis, ApoEâ»/â» or wild-type mice were intraperitoneally administered saline, lipopolysaccharides (LPS), or LPS plus TAK-242 (TLR4 inhibitor), respectively, twice a week for 4 weeks. Serum autoantibody of antinuclear antibody (ANA), anti-double-stranded DNA (anti-dsDNA), and cytokines of interferon-gamma (IFN-γ), tumor necrosis factor (TNF-α ), and interleukin-1 (IL-1ß) were assessed by ELISA. Hematoxylin and eosin (HE) and Perl's stains for lung pathomorphology as well as HE staining for atherosclerosis were employed. TLR4 in macrophages was detected by double immunofluorescent staining. While protein expressions of TLR4, nuclear factor-kappa B p65 (NF-κB p65), and B cell activating factor belonging to the TNF family (BAFF) were examined by immunohistochemistry. We found that serum autoantibody (ANA and anti-dsDNA), cytokines (IFN-γ, TNF-α, IL-1ß), lung inflammation, and intima-media thickness in brachiocephalic artery were obviously increased after LPS challenge in both genotypes, but to a lesser extent in wild-type strains. And those alterations were alleviated by coadministration of LPS and TAK-242. Mechanistically, upregulation of TLR4, NF-κb, and BAFF was involved. We concluded that TLR4/NF-κb/BAFF in macrophages might be a possible common autoimmune pathway that caused lung injury and atherosclerosis. TLR4 signal will be a therapeutic target in atherosclerosis and immune-mediated lung injury.
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Aterosclerosis/etiología , Lesión Pulmonar/etiología , Lupus Eritematoso Sistémico/complicaciones , Lupus Eritematoso Sistémico/metabolismo , Receptor Toll-Like 4/metabolismo , Animales , Anticuerpos Antinucleares/sangre , Anticuerpos Antinucleares/inmunología , Apolipoproteínas E/genética , Aterosclerosis/patología , Autoanticuerpos/inmunología , Citocinas/sangre , Modelos Animales de Enfermedad , Femenino , Hemorragia , Mediadores de Inflamación/sangre , Lipopolisacáridos/inmunología , Lesión Pulmonar/patología , Lupus Eritematoso Sistémico/genética , Lupus Eritematoso Sistémico/inmunología , Macrófagos/inmunología , Macrófagos/metabolismo , Ratones , Ratones Noqueados , FN-kappa B/metabolismo , Proteínas Proto-Oncogénicas B-raf/metabolismo , Seno Aórtico/inmunología , Seno Aórtico/metabolismo , Seno Aórtico/patologíaRESUMEN
Atherosclerosis is a disease of the large arteries and a major underlying cause of myocardial infarction and stroke. Several different mouse models have been developed to facilitate the study of the molecular and cellular pathophysiology of this disease. In this manuscript we describe specific techniques for the quantification and characterization of atherosclerotic lesions in the murine aortic sinus and ascending aorta. The advantage of this procedure is that it provides an accurate measurement of the cross-sectional area and total volume of the lesion, which can be used to compare atherosclerotic progression across different treatment groups. This is possible through the use of the valve leaflets as an anatomical landmark, together with careful adjustment of the sectioning angle. We also describe basic staining methods that can be used to begin to characterize atherosclerotic progression. These can be further modified to investigate antigens of specific interest to the researcher. The described techniques are generally applicable to a wide variety of existing and newly created dietary and genetically-induced models of atherogenesis.
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Aterosclerosis/patología , Seno Aórtico/patología , Animales , Aorta/metabolismo , Aorta/patología , Aterosclerosis/metabolismo , Modelos Animales de Enfermedad , Inmunohistoquímica , Ratones , Seno Aórtico/metabolismo , Coloración y Etiquetado/métodosRESUMEN
Reverse cholesterol transport (RCT) refers to the mobilization of cholesterol on HDL particles (HDL-C) from extravascular tissues to plasma, ultimately for fecal excretion. Little is known about how HDL-C leaves peripheral tissues to reach plasma. We first used 2 models of disrupted lymphatic drainage from skin--1 surgical and the other genetic--to quantitatively track RCT following injection of [3H]-cholesterol-loaded macrophages upstream of blocked or absent lymphatic vessels. Macrophage RCT was markedly impaired in both models, even at sites with a leaky vasculature. Inhibited RCT was downstream of cholesterol efflux from macrophages, since macrophage efflux of a fluorescent cholesterol analog (BODIPY-cholesterol) was not altered by impaired lymphatic drainage. We next addressed whether RCT was mediated by lymphatic vessels from the aortic wall by loading the aortae of donor atherosclerotic Apoe-deficient mice with [2H]6-labeled cholesterol and surgically transplanting these aortae into recipient Apoe-deficient mice that were treated with anti-VEGFR3 antibody to block lymphatic regrowth or with control antibody to allow such regrowth. [2H]-Cholesterol was retained in aortae of anti-VEGFR3-treated mice. Thus, the lymphatic vessel route is critical for RCT from multiple tissues, including the aortic wall. These results suggest that supporting lymphatic transport function may facilitate cholesterol clearance in therapies aimed at reversing atherosclerosis.
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Colesterol/sangre , Vasos Linfáticos/metabolismo , Macrófagos/metabolismo , Animales , Aorta Torácica/metabolismo , Aorta Torácica/trasplante , Aterosclerosis/metabolismo , Aterosclerosis/patología , Permeabilidad Capilar , Carcinoma Pulmonar de Lewis/irrigación sanguínea , Carcinoma Pulmonar de Lewis/metabolismo , Colesterol/metabolismo , Expresión Génica , Cinética , Ratones , Ratones Endogámicos C3H , Ratones Endogámicos C57BL , Ratones Transgénicos , Seno Aórtico/metabolismo , Piel/metabolismo , Receptor 3 de Factores de Crecimiento Endotelial Vascular/antagonistas & inhibidores , Receptor 3 de Factores de Crecimiento Endotelial Vascular/genética , Receptor 3 de Factores de Crecimiento Endotelial Vascular/metabolismoRESUMEN
Information on the biomechanical properties of aortic root aneurysms that would facilitate our understanding of their rupture modes is currently unavailable. In this study, whole-thickness wall specimens from aortic root aneurysms were studied in vitro so as to compare the biomechanical properties with gross histomorphology and composition, in relation to age, region, and direction. The stress-strain relationship was determined under uniaxial loading conditions and characterized by the Fung-type material model in terms of optimized material constants; failure properties were recorded. The connective tissue contents of the basic scleroproteins were also determined through computerized histology. Aging had a deleterious influence on the tensile strength of the aneurysmal sinus tissue, causing also stiffening and reduced extensibility that was consistent with the deficient elastin and collagen contents. Direction-dependent differences were demonstrated in the noncoronary sinus, with the circumferential being stiffer and stronger than the longitudinal direction, justified by the preferred collagen reinforcement along that direction there. In the left and right coronary sinus, the material constants and failure properties were essentially the same in the two directions, justified by the arbitrary orientation of medial (collagen and elastin fibers, and cellular) components relative to the circumferential-longitudinal directions. The material characterization results afforded, and the regional and age-related differences in the strength of the sinus wall, i.e. in its capacity to withstand hemodynamic stresses, are hoped to provide novel insight into the pathophysiological mechanisms responsible for the highest incidence of ruptured aortic root aneurysms in the right coronary and noncoronary sinus.
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Envejecimiento , Aneurisma de la Aorta , Rotura de la Aorta , Seno Aórtico , Estrés Fisiológico , Envejecimiento/metabolismo , Envejecimiento/patología , Aneurisma de la Aorta/metabolismo , Aneurisma de la Aorta/patología , Aneurisma de la Aorta/fisiopatología , Rotura de la Aorta/metabolismo , Rotura de la Aorta/patología , Rotura de la Aorta/fisiopatología , Colágeno/metabolismo , Elastina/metabolismo , Femenino , Humanos , Masculino , Persona de Mediana Edad , Seno Aórtico/metabolismo , Seno Aórtico/patología , Seno Aórtico/fisiopatologíaRESUMEN
Inhibition of atherosclerosis progression has long been the subject of intensive pathophysiologic investigations. The identification of a novel target molecule to redeem the cellular processes remains a major challenge in cardiology. Signal peptide CUB domain EGF-like repeat protein (SCUBE) family has been detected on human tissues and cultured cells. Two members of the SCUBE family, SCUBE1 and SCUBE3 are reported to play a role in cardiovascular diseases. The other member, SCUBE2 has been reported to mediate Hedgehog (Hh) protein signaling and is expressed in major blood vessels during mouse embryogenesis. However its involvement in cardiovascular diseases is not known yet. The aim of this study was to investigate SCUBE2 expression and localization in diffuse intimal thickening (DIT), an early event of atherosclerosis and in advanced lesion of atherosclerotic plaque. Carotid artery ligation in C57BL/6J mice was performed to induce intimal thickening, mimicking DIT in human. After 2 weeks of ligation, mRNA level of SCUBE2 increased significantly, while in LDLr-/- mice fed with high fat diet, a human atherosclerosis model, mRNA level of SCUBE2 expression markedly increased 8 weeks after start of the high fat diet. Our findings were confirmed by the observation of SCUBE2 expression in human coronary artery with DIT and advanced lesion of atherosclerotic plaque. Previous investigations described that SCUBE2 mediates Hh signaling pathway. We have observed that SCUBE2 expression is associated with Sonic Hedgehog (Shh) and its receptor, Patched (Ptc), both in DIT and advanced plaque lesion. Our results suggested that SCUBE2 is a new target molecule in atherosclerosis and might play an important role in atherosclerotic plaque progression via Hh signal transduction.
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Arterias/metabolismo , Aterosclerosis/metabolismo , Péptidos y Proteínas de Señalización Intercelular/metabolismo , Proteínas de la Membrana/metabolismo , Proteínas Adaptadoras Transductoras de Señales , Adolescente , Adulto , Anciano , Anciano de 80 o más Años , Animales , Apolipoproteínas B/metabolismo , Aterosclerosis/etiología , Proteínas de Unión al Calcio , Arterias Carótidas/metabolismo , Vasos Coronarios/metabolismo , Modelos Animales de Enfermedad , Progresión de la Enfermedad , Proteínas Hedgehog/metabolismo , Humanos , Masculino , Ratones Endogámicos C57BL , Ratones Noqueados , Persona de Mediana Edad , Receptores Patched , Receptor Patched-1 , Receptores de Superficie Celular/metabolismo , Seno Aórtico/metabolismo , Adulto JovenRESUMEN
The detection of inflammatory changes is a key aim for the early diagnosis and treatment of several autoimmune, infectious, and metastatic diseases. While surface-enhanced Raman scattering (SERS) has the capability to provide noninvasive, in vivo imaging at sufficient depth to achieve this goal, this approach has not been exploited in the study of inflammation. SERS-active nanoparticles were coded with a unique Raman signal that was protected under a wide range of conditions and stimuli. To detect early-stage inflammation, gold nanoparticle clusters containing Raman-active molecules were conjugated to intercellular adhesion molecule 1- (ICAM-1-) specific monoclonal antibodies. SERS allowed noninvasive measurement of ICAM-1 expression in vivo with twice the sensitivity of two-photon fluorescence. This is the first time SERS has been used for in vivo detection of inflammation and is a major advance in the ever-growing toolkit of approaches for use in noninvasive, next-generation in vivo imaging.