RESUMO
BACKGROUND: The epithelial growth factor receptor family of tyrosine kinases modulates embryonic formation of semilunar valves. We hypothesized that mice heterozygous for a dominant loss-of-function mutation in epithelial growth factor receptor, which are EgfrVel/+ mice, would develop anomalous aortic valves, valve dysfunction, and valvular cardiomyopathy. METHODS AND RESULTS: Aortic valves from EgfrVel/+ mice and control mice were examined by light microscopy at 2.5 to 4 months of age. Additional EgfrVel/+ and control mice underwent echocardiography at 2.5, 4.5, 8, and 12 months of age, followed by histologic examination. In young mice, microscopy revealed anatomic anomalies in 79% of EgfrVel/+ aortic valves, which resembled human unicuspid aortic valves. Anomalies were not observed in control mice. At 12 months of age, histologic architecture was grossly distorted in EgfrVel/+ aortic valves. Echocardiography detected moderate or severe aortic regurgitation, or aortic stenosis was present in 38% of EgfrVel/+ mice at 2.5 months of age (N=24) and in 74% by 8 months of age. Left ventricular enlargement, hypertrophy, and reversion to a fetal myocardial gene expression program occurred in EgfrVel/+ mice with aortic valve dysfunction, but not in EgfrVel/+ mice with near-normal aortic valve function. Myocardial fibrosis was minimal or absent in all groups. CONCLUSIONS: A new mouse model uniquely recapitulates salient functional, structural, and histologic features of human unicuspid aortic valve disease, which are phenotypically distinct from other forms of congenital aortic valve disease. The new model may be useful for elucidating mechanisms by which congenitally anomalous aortic valves become critically dysfunctional.
Assuntos
Valva Aórtica/anormalidades , Receptores ErbB/genética , Cardiopatias Congênitas/genética , Doenças das Valvas Cardíacas/genética , Mutação com Perda de Função , Animais , Valva Aórtica/diagnóstico por imagem , Valva Aórtica/fisiopatologia , Insuficiência da Valva Aórtica/genética , Insuficiência da Valva Aórtica/fisiopatologia , Estenose da Valva Aórtica/genética , Estenose da Valva Aórtica/fisiopatologia , Modelos Animais de Doenças , Progressão da Doença , Feminino , Regulação da Expressão Gênica , Predisposição Genética para Doença , Cardiopatias Congênitas/diagnóstico por imagem , Cardiopatias Congênitas/patologia , Cardiopatias Congênitas/fisiopatologia , Doenças das Valvas Cardíacas/diagnóstico por imagem , Doenças das Valvas Cardíacas/patologia , Doenças das Valvas Cardíacas/fisiopatologia , Hemodinâmica , Humanos , Hipertrofia Ventricular Esquerda/genética , Hipertrofia Ventricular Esquerda/fisiopatologia , Masculino , Camundongos Endogâmicos C57BL , Camundongos Mutantes , Fenótipo , Fatores de Tempo , Função Ventricular EsquerdaRESUMO
OBJECTIVE: Hypercholesterolemia and hypertension are associated with aortic valve stenosis (AVS) in humans. We have examined aortic valve function, structure, and gene expression in hypercholesterolemic/hypertensive mice. APPROACH AND RESULTS: Control, hypertensive, hypercholesterolemic (Apoe(-/-)), and hypercholesterolemic/hypertensive mice were studied. Severe aortic stenosis (echocardiography) occurred only in hypercholesterolemic/hypertensive mice. There was minimal calcification of the aortic valve. Several structural changes were identified at the base of the valve. The intercusp raphe (or seam between leaflets) was longer in hypercholesterolemic/hypertensive mice than in other mice, and collagen fibers at the base of the leaflets were reoriented to form a mesh. In hypercholesterolemic/hypertensive mice, the cusps were asymmetrical, which may contribute to changes that produce AVS. RNA sequencing was used to identify molecular targets during the developmental phase of stenosis. Genes related to the structure of the valve were identified, which differentially expressed before fibrotic AVS developed. Both RNA and protein of a profibrotic molecule, plasminogen activator inhibitor 1, were increased greatly in hypercholesterolemic/hypertensive mice. CONCLUSIONS: Hypercholesterolemic/hypertensive mice are the first model of fibrotic AVS. Hypercholesterolemic/hypertensive mice develop severe AVS in the absence of significant calcification, a feature that resembles AVS in children and some adults. Structural changes at the base of the valve leaflets include lengthening of the raphe, remodeling of collagen, and asymmetry of the leaflets. Genes were identified that may contribute to the development of fibrotic AVS.
Assuntos
Estenose da Valva Aórtica/etiologia , Valva Aórtica/patologia , Hipercolesterolemia/complicações , Hipertensão/complicações , Angiotensinogênio/genética , Angiotensinogênio/metabolismo , Animais , Valva Aórtica/metabolismo , Valva Aórtica/fisiopatologia , Estenose da Valva Aórtica/metabolismo , Estenose da Valva Aórtica/patologia , Estenose da Valva Aórtica/fisiopatologia , Apolipoproteínas E/deficiência , Apolipoproteínas E/genética , Modelos Animais de Doenças , Feminino , Fibrose , Regulação da Expressão Gênica , Hipercolesterolemia/genética , Hipercolesterolemia/metabolismo , Hipertensão/genética , Hipertensão/metabolismo , Masculino , Camundongos Endogâmicos C57BL , Camundongos Knockout , Inibidor 1 de Ativador de Plasminogênio/genética , Inibidor 1 de Ativador de Plasminogênio/metabolismo , Renina/genética , Renina/metabolismo , Índice de Gravidade de DoençaRESUMO
OBJECTIVE: We studied the mechanistic links between fibrocalcific changes in the aortic valve and aortic valve function in mice homozygous for a hypomorphic epidermal growth factor receptor mutation (Wave mice). We also studied myocardial responses to aortic valve dysfunction in Wave mice. APPROACH AND RESULTS: At 1.5 months of age, before development of valve fibrosis and calcification, aortic regurgitation, but not aortic stenosis, was common in Wave mice. Aortic valve fibrosis, profibrotic signaling, calcification, osteogenic markers, lipid deposition, and apoptosis increased dramatically by 6 and 12 months of age in Wave mice. Aortic regurgitation remained prevalent, however, and aortic stenosis was rare, at all ages. Proteoglycan content was abnormally increased in aortic valves of Wave mice at all ages. Treatment with pioglitazone prevented abnormal valve calcification, but did not protect valve function. There was significant left ventricular volume overload, hypertrophy, and fetal gene expression, at all ages in Wave mice with aortic regurgitation. Left ventricular systolic function was normal until 6 months of age in Wave mice, but became impaired by 12 months of age. Myocardial transverse tubules were normal in the presence of left ventricular hypertrophy at 1.5 and 3 months of age, but became disrupted by 12 months of age. CONCLUSIONS: We present the first comprehensive phenotypic and molecular characterization of spontaneous aortic regurgitation and volume-overload cardiomyopathy in an experimental model. In Wave mice, fibrocalcific changes are not linked to valve dysfunction and are epiphenomena arising from structurally incompetent myxomatous valves.
Assuntos
Insuficiência da Valva Aórtica/patologia , Insuficiência da Valva Aórtica/fisiopatologia , Doenças das Valvas Cardíacas/patologia , Doenças das Valvas Cardíacas/fisiopatologia , Actinas/metabolismo , Animais , Valva Aórtica/efeitos dos fármacos , Valva Aórtica/patologia , Valva Aórtica/fisiopatologia , Calcinose/patologia , Calcinose/prevenção & controle , Morte Celular , Progressão da Doença , Fibrose , Expressão Gênica , Metabolismo dos Lipídeos , Camundongos , Camundongos Mutantes , Osteocalcina/metabolismo , Pioglitazona , Proteoglicanas/metabolismo , Fator de Transcrição Sp7 , Sístole , Tiazolidinedionas/farmacologia , Fatores de Transcrição/metabolismo , Fator de Crescimento Transformador beta/metabolismoRESUMO
Risk factors for fibrocalcific aortic valve disease (FCAVD) are associated with systemic decreases in bioavailability of endothelium-derived nitric oxide (EDNO). In patients with bicuspid aortic valve (BAV), vascular expression of endothelial nitric oxide synthase (eNOS) is decreased, and eNOS(-/-) mice have increased prevalence of BAV. The goal of this study was to test the hypotheses that EDNO attenuates profibrotic actions of valve interstitial cells (VICs) in vitro and that EDNO deficiency accelerates development of FCAVD in vivo. As a result of the study, coculture of VICs with aortic valve endothelial cells (vlvECs) significantly decreased VIC activation, a critical early phase of FCAVD. Inhibition of VIC activation by vlvECs was attenuated by N(G)-nitro-l-arginine methyl ester or indomethacin. Coculture with vlvECs attenuated VIC expression of matrix metalloproteinase-9, which depended on stiffness of the culture matrix. Coculture with vlvECs preferentially inhibited collagen-3, compared with collagen-1, gene expression. BAV occurred in 30% of eNOS(-/-) mice. At age 6 mo, collagen was increased in both bicuspid and trileaflet eNOS(-/-) aortic valves, compared with wild-type valves. At 18 mo, total collagen was similar in eNOS(-/-) and wild-type mice, but collagen-3 was preferentially increased in eNOS(-/-) mice. Calcification and apoptosis were significantly increased in BAV of eNOS(-/-) mice at ages 6 and 18 mo. Remarkably, these histological changes were not accompanied by physiologically significant valve stenosis or regurgitation. In conclusion, coculture with vlvECs inhibits specific profibrotic VIC processes. In vivo, eNOS deficiency produces fibrosis in both trileaflet and BAVs but produces calcification only in BAVs.
Assuntos
Valva Aórtica/patologia , Calcinose/metabolismo , Cardiopatias Congênitas/metabolismo , Doenças das Valvas Cardíacas/metabolismo , Óxido Nítrico Sintase Tipo III/metabolismo , Animais , Valva Aórtica/metabolismo , Valva Aórtica/fisiopatologia , Apoptose , Doença da Válvula Aórtica Bicúspide , Calcinose/patologia , Calcinose/fisiopatologia , Células Cultivadas , Colágeno Tipo I/genética , Colágeno Tipo I/metabolismo , Colágeno Tipo III/genética , Colágeno Tipo III/metabolismo , Cardiopatias Congênitas/patologia , Cardiopatias Congênitas/fisiopatologia , Doenças das Valvas Cardíacas/patologia , Doenças das Valvas Cardíacas/fisiopatologia , Células Intersticiais de Cajal/metabolismo , Células Intersticiais de Cajal/patologia , Metaloproteinase 9 da Matriz/metabolismo , Camundongos , Óxido Nítrico Sintase Tipo III/deficiência , Óxido Nítrico Sintase Tipo III/genética , Esclerose/metabolismo , Esclerose/patologia , Esclerose/fisiopatologia , SuínosRESUMO
BACKGROUND: There are no rigorously confirmed effective medical therapies for calcific aortic stenosis. Hypercholesterolemic Ldlr (-/-) Apob (100/100) mice develop calcific aortic stenosis and valvular cardiomyopathy in old age. Osteoprotegerin (OPG) modulates calcification in bone and blood vessels, but its effect on valve calcification and valve function is not known. OBJECTIVES: To determine the impact of pharmacologic treatment with OPG upon aortic valve calcification and valve function in aortic stenosis-prone hypercholesterolemic Ldlr (-/-) Apob (100/100) mice. METHODS: Young Ldlr (-/-) Apob (100/100) mice (age 2 months) were fed a Western diet and received exogenous OPG or vehicle (Nâ=â12 each) 3 times per week, until age 8 months. After echocardiographic evaluation of valve function, the aortic valve was evaluated histologically. Older Ldlr (-/-) Apob (100/100) mice were fed a Western diet beginning at age 2 months. OPG or vehicle (Nâ=â12 each) was administered from 6 to 12 months of age, followed by echocardiographic evaluation of valve function, followed by histologic evaluation. RESULTS: In Young Ldlr (-/-) Apob (100/100) mice, OPG significantly attenuated osteogenic transformation in the aortic valve, but did not affect lipid accumulation. In Older Ldlr (-/-) Apob (100/100) mice, OPG attenuated accumulation of the osteoblast-specific matrix protein osteocalcin by â¼80%, and attenuated aortic valve calcification by â¼ 70%. OPG also attenuated impairment of aortic valve function. CONCLUSIONS: OPG attenuates pro-calcific processes in the aortic valve, and protects against impairment of aortic valve function in hypercholesterolemic aortic stenosis-prone Ldlr (-/-) Apob (100/100) mice.
Assuntos
Estenose da Valva Aórtica/tratamento farmacológico , Valva Aórtica/efeitos dos fármacos , Valva Aórtica/patologia , Calcinose/tratamento farmacológico , Hipercolesterolemia/tratamento farmacológico , Osteoprotegerina/farmacologia , Fatores Etários , Animais , Valva Aórtica/diagnóstico por imagem , Valva Aórtica/metabolismo , Estenose da Valva Aórtica/diagnóstico por imagem , Estenose da Valva Aórtica/metabolismo , Estenose da Valva Aórtica/patologia , Apolipoproteína B-100/genética , Calcinose/diagnóstico por imagem , Calcinose/metabolismo , Calcinose/patologia , Modelos Animais de Doenças , Feminino , Hipercolesterolemia/diagnóstico por imagem , Hipercolesterolemia/metabolismo , Hipercolesterolemia/patologia , Injeções , Masculino , Camundongos , Camundongos Transgênicos , Osteogênese/efeitos dos fármacos , Receptores de LDL/deficiência , Receptores de LDL/genética , UltrassonografiaRESUMO
OBJECTIVE: Development of calcific aortic valve stenosis involves multiple signaling pathways, which may be modulated by peroxisome proliferator-activated receptor-γ). This study tested the hypothesis that pioglitazone (Pio), a ligand for peroxisome proliferator-activated receptor-γ, inhibits calcification of the aortic valve in hypercholesteremic mice. METHODS AND RESULTS: Low density lipoprotein receptor(-/-)/apolipoprotein B(100/100) mice were fed a Western-type diet with or without Pio (20 mg/kg per day) for 6 months. Pio attenuated lipid deposition and calcification in the aortic valve, but not aorta. In the aortic valve, Pio reduced levels of active caspase-3 and terminal deoxynucleotidyl transferase dUTP nick end labeling staining. Valve function (echocardiography) was significantly improved by Pio. To determine whether changes in gene expression are associated with differential effects of Pio on aortic valves versus aorta, Reversa mice were fed Western diet with or without Pio for 2 months. Several procalcific genes were increased by Western diet, and the increase was attenuated by Pio, in aortic valve, but not aorta. CONCLUSIONS: Pio attenuates lipid deposition, calcification, and apoptosis in aortic valves of hypercholesterolemic mice, improves aortic valve function, and exhibits preferential effects on aortic valves versus aorta. We suggest that Pio protects against calcific aortic valve stenosis, and Pio or other peroxisome proliferator-activated receptor-γ ligands may be useful for early intervention to prevent or slow stenosis of aortic valves.
Assuntos
Estenose da Valva Aórtica/prevenção & controle , Valva Aórtica/efeitos dos fármacos , Calcinose/prevenção & controle , Hipercolesterolemia/tratamento farmacológico , Tiazolidinedionas/farmacologia , Adiponectina/sangue , Animais , Aorta/efeitos dos fármacos , Aorta/metabolismo , Valva Aórtica/diagnóstico por imagem , Valva Aórtica/metabolismo , Valva Aórtica/patologia , Valva Aórtica/fisiopatologia , Estenose da Valva Aórtica/diagnóstico , Estenose da Valva Aórtica/genética , Estenose da Valva Aórtica/metabolismo , Estenose da Valva Aórtica/fisiopatologia , Apolipoproteína B-100/deficiência , Apolipoproteína B-100/genética , Apoptose/efeitos dos fármacos , Biomarcadores/sangue , Glicemia/metabolismo , Calcinose/diagnóstico , Calcinose/genética , Calcinose/metabolismo , Calcinose/fisiopatologia , Caspase 3/metabolismo , Colesterol/sangue , Modelos Animais de Doenças , Ativação Enzimática , Feminino , Regulação da Expressão Gênica , Hipercolesterolemia/diagnóstico , Hipercolesterolemia/genética , Hipercolesterolemia/metabolismo , Marcação In Situ das Extremidades Cortadas , Camundongos , Camundongos Knockout , Osteogênese/efeitos dos fármacos , Osteogênese/genética , PPAR gama/agonistas , PPAR gama/metabolismo , Pioglitazona , Receptores de LDL/deficiência , Receptores de LDL/genética , Proteína Amiloide A Sérica/metabolismo , Fatores de Tempo , UltrassonografiaRESUMO
Endothelial vasomotor function decreases with increasing age. Extracellular superoxide dismutase (ecSOD) protects against vascular dysfunction in several disease states. The purpose of this study was to determine whether endogenous ecSOD protects against endothelial dysfunction in old mice. Vasomotor function of the aorta was studied ex vivo in wild-type (ecSOD(+/+)) and ecSOD-deficient (ecSOD(-/-)) mice at 11 (adult) and 29 (old) mo of age. Maximal relaxation to acetylcholine (10(-4) M) was impaired in vessels from adult ecSOD(-/-) mice [75 +/- 3% (mean +/- SE)] compared with wild-type mice (89 +/- 2%, P < 0.05). Maximal relaxation to acetylcholine (10(-4) M) was profoundly impaired in aorta from old ecSOD(-/-) mice (45 +/- 5%) compared with wild-type mice (75 +/- 4%, P < 0.05). There was a significant correlation between expression of ecSOD and maximal relaxation to acetylcholine in adult and old mice. Tempol (1 mM), a scavenger of superoxide, improved relaxation in response to acetylcholine (63 +/- 8%) in old ecSOD(-/-) mice (P < 0.05), but not wild-type mice (75 +/- 4%). Maximal relaxation to sodium nitroprusside was similar in aorta from adult and old wild-type and ecSOD(-/-) mice. Quantitative RT-PCR showed a decrease in mRNA levels of ecSOD and catalase in aorta of old mice and an increase in levels of TNFalpha and Nox-4 in aorta of old mice compared with adult mice. The findings support the hypothesis that impaired antioxidant mechanisms may contribute to cumulative increases in oxidative stress and impaired endothelial function in old mice. In conclusion, endogenous ecSOD plays an important role in protection against endothelial dysfunction during aging.
Assuntos
Envelhecimento/metabolismo , Endotélio Vascular/metabolismo , Estresse Oxidativo/fisiologia , Superóxido Dismutase/genética , Superóxido Dismutase/metabolismo , Acetilcolina/farmacologia , Animais , Antioxidantes/metabolismo , Endotélio Vascular/efeitos dos fármacos , Espaço Extracelular/enzimologia , Feminino , Regulação Enzimológica da Expressão Gênica/fisiologia , Camundongos , Camundongos Mutantes , Óxido Nítrico/metabolismo , RNA Mensageiro/metabolismo , Superóxidos/metabolismo , Vasodilatação/efeitos dos fármacos , Vasodilatação/fisiologia , Vasodilatadores/farmacologiaRESUMO
UNLABELLED: Our previous studies have shown vascular dysfunction in small coronary and mesenteric arteries in Zucker obese rats, a model of the metabolic syndrome, and Zucker Diabetic Fatty (ZDF) rats, a model of type 2 diabetes. Because of their lipid lowering action and antioxidant activity, we predicted that treatment with Rosuvastatin, an HMG-CoA reductase inhibitor (statin) or Enalapril, an angiotensin converting enzyme (ACE) inhibitor would improve vascular dysfunction associated with the metabolic syndrome and type 2 diabetes. METHODS: 20-week-old Zucker obese and 16-week-old ZDF rats were treated with Rosuvastatin (25 mg/kg/day) or Enalapril (20 mg/kg/day) for 12 weeks. We examined metabolic parameters, indices of oxidative stress and vascular dysfunction in ventricular and mesenteric small arteries (75-175 microm intraluminal diameter) from lean, Zucker obese and ZDF rats (untreated and treated). RESULTS: Endothelial dependent responses were attenuated in coronary vessels from Zucker obese and ZDF rats compared to responses from lean rats. Both drugs improved metabolic parameters, oxidative stress, and vascular dysfunction in Zucker obese rats, however, only partial improvement was observed in ZDF rats, suggesting more aggressive treatment is needed when hyperglycemia is involved. CONCLUSION: Vascular dysfunction is improved when Zucker obese and, to a lesser degree, when ZDF rats were treated with Rosuvastatin or Enalapril.
Assuntos
Doenças Cardiovasculares/tratamento farmacológico , Diabetes Mellitus Tipo 2/tratamento farmacológico , Enalapril/farmacologia , Fluorbenzenos/farmacologia , Síndrome Metabólica/tratamento farmacológico , Pirimidinas/farmacologia , Sulfonamidas/farmacologia , Fatores Etários , Inibidores da Enzima Conversora de Angiotensina/farmacologia , Inibidores da Enzima Conversora de Angiotensina/uso terapêutico , Animais , Glicemia/metabolismo , Pressão Sanguínea/efeitos dos fármacos , Peso Corporal/efeitos dos fármacos , Doenças Cardiovasculares/complicações , Doenças Cardiovasculares/fisiopatologia , Colesterol/sangue , Vasos Coronários/efeitos dos fármacos , Vasos Coronários/metabolismo , Vasos Coronários/fisiopatologia , Diabetes Mellitus Tipo 2/complicações , Diabetes Mellitus Tipo 2/fisiopatologia , Enalapril/uso terapêutico , Endotélio Vascular/efeitos dos fármacos , Endotélio Vascular/metabolismo , Endotélio Vascular/fisiopatologia , Ácidos Graxos/sangue , Fluorbenzenos/uso terapêutico , Inibidores de Hidroximetilglutaril-CoA Redutases/farmacologia , Inibidores de Hidroximetilglutaril-CoA Redutases/uso terapêutico , Masculino , Artérias Mesentéricas/efeitos dos fármacos , Artérias Mesentéricas/fisiopatologia , Síndrome Metabólica/complicações , Síndrome Metabólica/fisiopatologia , Estresse Oxidativo/efeitos dos fármacos , Pirimidinas/uso terapêutico , Ratos , Ratos Zucker , Rosuvastatina Cálcica , Sulfonamidas/uso terapêutico , Triglicerídeos/sangueRESUMO
OBJECTIVE: Obese Zucker rats, animal model for the metabolic syndrome, develop a diabetes-like neuropathy that is independent of hyperglycemia. The purpose of this study was to determine whether drugs used to treat cardiovascular dysfunction in metabolic syndrome also protect nerve function. METHODS AND PROCEDURES: Obese Zucker rats at 20 weeks of age were treated for 12 weeks with enalapril or rosuvastatin. Lean rats were used as controls. Vasodilation in epineurial arterioles was measured by videomicroscopy. Endoneurial blood flow (EBF) was measured by hydrogen clearance and nerve conduction velocity was measured following electrical stimulation of motor or sensory nerves. RESULTS: Enalapril treatment decreased serum angiotensin-converting enzyme (ACE) activity and both drugs reduced serum cholesterol levels. In obese Zucker rats at 32 weeks of age superoxide levels were elevated in the aortas and epineurial arterioles, which were reduced by treatment with either drug. Nitrotyrosine levels were increased in epineurial arterioles and reduced with enalapril treatment. EBF was decreased and corrected by treatment with either drug. Motor nerve conduction velocity was decreased and significantly improved with enalapril treatment. Obese Zucker rats were hypoalgesic in response to a thermal stimulus and this was significantly improved with either treatment. Treatment with either enalapril or rosuvastatin significantly reversed the decrease in acetylcholine-mediated vascular relaxation of epineurial arterioles in obese Zucker rats. DISCUSSION: Even though obese Zucker rats have normal glycemia vascular and neural dysfunctions develop with age and can be improved by treatment with either enalapril or rosuvastatin.
Assuntos
Inibidores da Enzima Conversora de Angiotensina/farmacologia , Sistema Cardiovascular/fisiopatologia , Enalapril/farmacologia , Fluorbenzenos/farmacologia , Inibidores de Hidroximetilglutaril-CoA Redutases/farmacologia , Obesidade/fisiopatologia , Nervos Periféricos/fisiopatologia , Pirimidinas/farmacologia , Sulfonamidas/farmacologia , Acetilcolina/farmacologia , Animais , Arteríolas/efeitos dos fármacos , Arteríolas/fisiologia , Sistema Cardiovascular/efeitos dos fármacos , Modelos Animais de Doenças , Masculino , Síndrome Metabólica/fisiopatologia , Neurônios Motores/efeitos dos fármacos , Neurônios Motores/fisiologia , Condução Nervosa/efeitos dos fármacos , Condução Nervosa/fisiologia , Neurônios Aferentes/efeitos dos fármacos , Neurônios Aferentes/fisiologia , Nociceptores/efeitos dos fármacos , Nociceptores/fisiologia , Nervos Periféricos/efeitos dos fármacos , Ratos , Ratos Zucker , Rosuvastatina Cálcica , Nervo Isquiático/irrigação sanguínea , Nervo Isquiático/fisiologia , Superóxidos/metabolismo , Tirosina/análogos & derivados , Tirosina/metabolismo , Vasodilatadores/farmacologiaRESUMO
Endotoxin [or lipopolysaccharide (LPS)] increases levels of superoxide in blood vessels and impairs vasomotor function. Angiotensin II plays an important role in the generation of superoxide in several disease states, including hypertension and heart failure. The goal of this study was to determine whether the activation of the renin-angiotensin system contributes to oxidative stress and endothelial dysfunction after endotoxin. We examined the effects of enalapril (an angiotensin-converting enzyme inhibitor) or L-158809 (an angiotensin receptor blocker) on increases of superoxide and vasomotor dysfunction in mice treated with LPS. C57BL/6 mice were treated with either enalapril (60 mg.kg(-1).day(-1)) or L-158809 (30 mg.kg(-1).day(-1)) for 4 days. After the third day, LPS (10-20 mg/kg) or vehicle was injected intraperitoneally, and one day later, vasomotor function of the aorta was examined in vitro. After precontraction with PGF(2alpha), the maximal responses to sodium nitroprusside were similar in the aorta from normal and LPS-treated mice. In contrast, the relaxation to acetylcholine was impaired after LPS (54 +/- 5% at 10(-5), mean +/- SE) compared with vessels treated with vehicle (88 +/- 1%; P < 0.05). Enalapril improved (P < 0.05) relaxation in response to acetylcholine to 81 +/- 6% after LPS. L-158809 also improved relaxation in response to acetylcholine to 77 +/- 4% after LPS. Superoxide (measured with lucigenin and hydroethidine) was increased (P < 0.05) in aorta after LPS, and levels were reduced (P < 0.05) following enalapril and L-158809. Thus, after LPS, enalapril and L-158809 reduce superoxide levels and improve relaxation to acetylcholine in the aorta. The findings suggest that activation of the renin-angiotensin system contributes importantly to oxidative stress and endothelial dysfunction after endotoxin.
Assuntos
Bloqueadores do Receptor Tipo 1 de Angiotensina II/farmacologia , Angiotensina II/metabolismo , Inibidores da Enzima Conversora de Angiotensina/farmacologia , Enalapril/farmacologia , Endotélio Vascular/efeitos dos fármacos , Endotoxemia/metabolismo , Imidazóis/farmacologia , Sistema Renina-Angiotensina/efeitos dos fármacos , Tetrazóis/farmacologia , Vasodilatação/efeitos dos fármacos , Acetilcolina/farmacologia , Animais , Modelos Animais de Doenças , Relação Dose-Resposta a Droga , Endotélio Vascular/metabolismo , Endotélio Vascular/fisiopatologia , Endotoxemia/induzido quimicamente , Endotoxemia/fisiopatologia , Lipopolissacarídeos , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Nitroprussiato/farmacologia , Estresse Oxidativo/efeitos dos fármacos , Superóxidos/metabolismo , Vasodilatadores/farmacologiaRESUMO
A common gene variant in the heparin-binding domain (HBD) of extracellular superoxide dismutase (ECSOD) may predispose human carriers to ischaemic heart disease. We have demonstrated that the HBD of ECSOD is important for ECSOD to restore vascular dysfunction produced by endotoxin. The purpose of this study was to determine whether the gene variant in the HBD of ECSOD (ECSOD(R213G)) protects against endothelial dysfunction in a model of inflammation. We constructed a recombinant adenovirus that expresses ECSOD(R213G). Adenoviral vectors expressing ECSOD, ECSOD(R213G) or beta-galactosidase (LacZ, a control) were injected i.v. in mice. After 3 days, at which time the plasma SOD activity is maximal, vehicle or endotoxin (lipopolysaccharide or LPS, 40 mg kg(-1)) was injected i.p. Vasomotor function of aorta in vitro was examined 1 day later. Maximal relaxation to sodium nitroprusside was similar in aorta from normal and LPS-treated mice. Maximal relaxation to acetylcholine (10(-5)) was impaired after LPS and LacZ (63 +/- 3%, mean +/- s.e.m.) compared to normal vessels (83 +/- 3%) (P < 0.05). Gene transfer of ECSOD improved (P < 0.05) relaxation in response to acetylcholine (76 +/- 5%) after LPS, whereas gene transfer of ECSOD(R213G) had no effect (65 +/- 4%). Superoxide was increased in aorta (measured using lucigenin and hydroethidine) after LPS, and levels of superoxide were significantly reduced following ECSOD but not ECSOD(R213G). Thus, ECSOD reduces superoxide and improves relaxation to acetylcholine in the aorta after LPS, while the ECSOD variant R213G had minimal effect. These findings suggest that, in contrast to ECSOD, the common human gene variant of ECSOD fails to protect against endothelial dysfunction produced by an inflammatory stimulus.
Assuntos
Endotélio Vascular/fisiopatologia , Inflamação/fisiopatologia , Polimorfismo Genético , Superóxido Dismutase/metabolismo , Vasoconstrição , Vasodilatação , Acetilcolina/farmacologia , Adenoviridae/genética , Animais , Aorta/enzimologia , Aorta/fisiopatologia , Dinoprosta/farmacologia , Modelos Animais de Doenças , Relação Dose-Resposta a Droga , Endotélio Vascular/efeitos dos fármacos , Endotélio Vascular/enzimologia , Genes Reporter , Vetores Genéticos , Humanos , Inflamação/enzimologia , Óperon Lac , Lipopolissacarídeos , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Transgênicos , Nitroprussiato/farmacologia , Superóxido Dismutase/genética , Superóxidos/metabolismo , Vasoconstrição/efeitos dos fármacos , Vasoconstritores/farmacologia , Vasodilatação/efeitos dos fármacos , Vasodilatadores/farmacologia , beta-GalactosidaseRESUMO
In epineurial arterioles, acetylcholine-mediated vascular relaxation is mediated by nitric oxide and endothelium-derived hyperpolarizing factor (EDHF), and both mechanisms are impaired by diabetes. The mediator responsible for the effect of EDHF is unknown. In epineurial arterioles, C-type natriuretic peptide (CNP) has properties consistent with EDHF-like activity. Epineurial arterioles express CNP, and exogenous CNP causes a concentration-dependent vascular relaxation. In streptozotocin-induced diabetic rats, CNP-mediated vascular relaxation in epineurial arterioles is decreased. Since CNP may be a regulator of vascular function, a vasopeptidase inhibitor may be an effective treatment for diabetes-induced vascular and neural disease. Vasopeptidase inhibitors inhibit ACE activity and neutral endopeptidase, which degrades natriuretic peptides. Streptozotocin-induced diabetic rats were treated with AVE7688 (450 mg/kg in the diet), a vasopeptidase inhibitor, for 8-10 weeks after 4 weeks of untreated diabetes. Treatment of diabetic rats corrected the diabetes-induced decrease in endoneurial blood flow, significantly improved motor and sensory nerve conduction velocity, prevented the development of hypoalgesia in the hind paw, and reduced superoxide and nitrotyrosine levels in epineurial arterioles. The diabetes-induced decrease in acetylcholine-mediated vascular relaxation by epineurial arterioles was significantly improved with treatment. These studies suggest that vasopeptidase inhibitors may be an effective approach for the treatment of diabetic vascular and neural dysfunction.
Assuntos
Angiopatias Diabéticas/tratamento farmacológico , Neuropatias Diabéticas/tratamento farmacológico , Compostos Heterocíclicos com 3 Anéis/uso terapêutico , Peptídeo Natriurético Tipo C/metabolismo , Nervos Periféricos/irrigação sanguínea , Inibidores de Proteases/uso terapêutico , Animais , Arteríolas/metabolismo , Glicemia , Peso Corporal , Diabetes Mellitus Experimental/tratamento farmacológico , Diabetes Mellitus Experimental/fisiopatologia , Compostos Heterocíclicos com 3 Anéis/farmacologia , Masculino , Neprilisina/efeitos dos fármacos , Condução Nervosa/efeitos dos fármacos , Nervos Periféricos/metabolismo , Pró-Fármacos/farmacologia , Pró-Fármacos/uso terapêutico , Inibidores de Proteases/farmacologia , Ratos , Ratos Sprague-Dawley , Estreptozocina/farmacologia , Vasodilatação/efeitos dos fármacosRESUMO
We investigated the progression of vascular dysfunction associated with the metabolic syndrome with and without hyperglycemia in lean, Zucker obese, and Zucker diabetic fatty (ZDF) rats. Responses of aorta and small coronary and mesenteric arteries were measured to endothelium-dependent and -independent vasodilators. Indices of oxidative stress were increased in serum from ZDF rats throughout the study, whereas values were increased in Zucker obese rats later in the study [thiobarbituric acid reactive substances: 0.45 +/- 0.02, 0.59 +/- 0.03 (P < 0.05), and 0.58 +/- 0.03 (P < 0.05) mug/ml in serum from 28- to 40-wk-old lean, Zucker obese, and ZDF rats, respectively]. Acetylcholine (ACh)-induced relaxation was not altered in vessels from lean animals from 8-40 wk. ACh-induced relaxation was nearly abolished in coronary arteries from 28- to 36-wk-old Zucker obese rats and by 16-36 wk in ZDF rats and was attenuated in aorta and mesenteric vessels from ZDF rats [%relaxation to 10 muM ACh: 72.2 +/- 7.1, 17.9 +/- 5.9 (P < 0.05), and 23.0 +/- 4.5 (P < 0.05) in coronary vessels; and 67.9 +/- 9.2, 50.1 +/- 5.5, and 42.3 +/- 4.7 (P < 0.05) in mesenteric vessels from 28- to 40-wk-old lean, Zucker obese, and ZDF rats, respectively]. The attenuated ACh-induced relaxation was improved when vessels were incubated with tiron, suggesting superoxide as a mechanism of endothelial dysfunction. Sodium nitroprusside-induced relaxation was not altered in aorta or coronary arteries and was potentiated in mesenteric arteries from Zucker obese rats. Our data suggest that diabetes enhances the progression of vascular dysfunction. Increases in indices of oxidative stress precede the development of dysfunction and may serve as a marker of endothelial damage.
Assuntos
Doenças Cardiovasculares/fisiopatologia , Vasos Coronários/fisiopatologia , Diabetes Mellitus Experimental/fisiopatologia , Artérias Mesentéricas/fisiopatologia , Síndrome Metabólica/complicações , Obesidade/fisiopatologia , Animais , Vasos Coronários/efeitos dos fármacos , Diabetes Mellitus Experimental/sangue , Progressão da Doença , Endotélio Vascular/patologia , Hiperglicemia/complicações , Insulina/sangue , Estudos Longitudinais , Artérias Mesentéricas/efeitos dos fármacos , Síndrome Metabólica/sangue , Síndrome Metabólica/fisiopatologia , Nitroprussiato/farmacologia , Obesidade/sangue , Estresse Oxidativo/fisiologia , Ratos , Ratos Zucker , Superóxidos/metabolismo , Substâncias Reativas com Ácido Tiobarbitúrico/análise , Vasodilatadores/farmacologiaRESUMO
Aging is an independent risk factor for cardiovascular disease, but mechanisms leading to vascular dysfunction have not been fully elucidated. Recent studies suggest that oxidative stress may increase in blood vessels during aging. Levels of superoxide are influenced by the activity of SODs. The goal of this study was to examine the effect of extracellular superoxide dismutase (ECSOD) on superoxide levels and vascular function in an animal model of aging. Aortas from young (4-8 mo old) and old (29-31 mo old) Fischer 344 rats were examined in vitro. Relaxation of aorta to ACh was impaired in old rats compared with young rats; e.g., 3 muM ACh produced 57 +/- 4% (mean +/- SE) and 84 +/- 2% relaxation in old and young rats, respectively (P < 0.0001). Three days after gene transfer of adenovirus expressing human ECSOD (AdECSOD), the response to ACh was not affected in young rats but was improved in old rats. There was no difference in relaxation to the endothelium-independent dilator sodium nitroprusside between young, aged, and AdECSOD-treated old rats. Superoxide levels (lucigenin-enhanced chemiluminescence) were significantly increased in aged rats compared with young rats. After gene transfer of ECSOD to aged rats, superoxide levels in aorta were similar in old and young rats. Gene transfer of an ECSOD with the heparin-binding domain deleted had no effect on vascular function or superoxide levels in old rats. These results suggest that 1) vascular dysfunction associated with aging is mediated in part by increased levels of superoxide, 2) gene transfer of ECSOD reduces vascular superoxide and dysfunction in old rats, and 3) beneficial effects of ECSOD in old rats require the heparin-binding domain of ECSOD.
Assuntos
Envelhecimento/fisiologia , Superóxido Dismutase/genética , Doenças Vasculares/prevenção & controle , Adenoviridae/genética , Animais , Aorta Torácica/fisiologia , Glicemia/metabolismo , Western Blotting , Peso Corporal/fisiologia , Colesterol/sangue , Eletroforese em Gel de Poliacrilamida , Espaço Extracelular/enzimologia , Regulação Enzimológica da Expressão Gênica/fisiologia , Técnicas de Transferência de Genes , Heparina/metabolismo , Masculino , Músculo Liso Vascular/fisiologia , Ratos , Ratos Endogâmicos F344 , Superóxidos/metabolismoRESUMO
ACE inhibition and/or blocking of the angiotensin II receptor are recognized as first-line treatment for nephropathy and cardiovascular disease in diabetic patients. However, little information is available about the potential benefits of these drugs on diabetic neuropathy. We examined vascular and neural activity in streptozotocin-induced diabetic rats that were treated for 12 weeks with enalapril, an ACE inhibitor, or L-158809, an angiotensin II receptor blocker. A prevention protocol (group 1) as well as three intervention protocols (treatment was initiated after 4, 8, or 12 weeks of diabetes [groups 2, 3, and 4, respectively]) were used. Endoneurial blood flow and motor nerve conduction velocity (MNCV) were impaired in all groups of untreated diabetic rats. In group 1, treatment of diabetic rats with enalapril or L-158809 partially prevented the diabetes-induced decrease in endoneurial blood flow and MNCV. In groups 2-4, intervention with enalapril was more effective in reversing the diabetes-induced impairment in endoneurial blood flow and MNCV than L-158809. The superoxide level in the aorta and epineurial arterioles of diabetic rats was increased. Treatment of diabetic rats with enalapril or L-158809 reduced the superoxide level in the aorta in all groups but was less effective in epineurial arterioles. Acetylcholine and calcitonin gene-related peptide (CGRP) cause vasodilation in epineurial arterioles of the sciatic nerve, which was impaired by diabetes. Treatment of diabetic rats (all groups) with enalapril or L-158809 completely prevented/reversed the diabetes-induced impairment in CGRP-mediated vascular relaxation. Treatment with enalapril or L-158809 was also effective in improving impaired acetylcholine-mediated vasodilation, but the efficacy was diminished from groups 1 to 4. These studies suggest that ACE inhibitors and/or angiotensin II receptor blockers may be effective treatments for diabetes and vascular and neural dysfunction. However, the efficacy of these treatments may be dependent on when the treatment is initiated.
Assuntos
Bloqueadores do Receptor Tipo 1 de Angiotensina II/uso terapêutico , Inibidores da Enzima Conversora de Angiotensina/uso terapêutico , Neuropatias Diabéticas/tratamento farmacológico , Neuropatias Diabéticas/prevenção & controle , Enalapril/uso terapêutico , Imidazóis/uso terapêutico , Tetrazóis/uso terapêutico , Animais , Artérias/efeitos dos fármacos , Artérias/metabolismo , Diabetes Mellitus Experimental/induzido quimicamente , Diabetes Mellitus Experimental/complicações , Neuropatias Diabéticas/induzido quimicamente , Neuropatias Diabéticas/complicações , Quimioterapia Combinada , Masculino , Relaxamento Muscular/efeitos dos fármacos , Músculo Liso Vascular/efeitos dos fármacos , Músculo Liso Vascular/metabolismo , Estresse Oxidativo , Ratos , Ratos Sprague-DawleyRESUMO
BACKGROUND: Extracellular superoxide dismutase (ECSOD) is a major extracellular antioxidant enzyme. We have demonstrated that vascular effects of ECSOD require an intact heparin-binding domain. A common genetic variant with a substitution in the heparin-binding domain (ECSOD(R213G)) was reported recently to be associated with ischemic heart disease. The goal of this study was to examine vascular effects of ECSOD(R213G). METHODS AND RESULTS: A recombinant adenovirus (Ad) that expresses ECSOD(R213G) was constructed. ECSOD(R213G) and ECSOD proteins bound to collagen type I in vitro, but binding to aorta ex vivo was 10-fold greater with ECSOD than ECSOD(R213G). Three days after intravenous injection of AdECSOD(R213G) or AdECSOD in spontaneously hypertensive rats (SHR), immunostaining demonstrated binding of ECSOD to carotid arteries and kidneys but minimal binding of ECSOD(R213G). Binding to aorta and carotid artery was 2.5- to 3-fold greater with ECSOD than ECSOD(R213G) by immunoblotting. Arterial pressure was significantly reduced by AdECSOD but not by AdECSOD(R213G). Responses to acetylcholine and basal levels of nitric oxide in carotid arteries were impaired in SHR compared with normotensive Wistar-Kyoto rats and were improved after AdECSOD but not AdECSOD(R213G). Levels of superoxide and nitrotyrosine in aorta were higher in SHR than Wistar-Kyoto rats and were greatly reduced after AdECSOD but not AdECSOD(R213G). CONCLUSIONS: In contrast to ECSOD, ECSOD(R213G) has no significant protective effect on arterial pressure, vascular function, or vascular levels of oxidative stress in SHR. These findings may provide a mechanistic basis for association studies that suggest that human beings carrying ECSOD(R213G) are predisposed to vascular diseases.
Assuntos
Polimorfismo de Nucleotídeo Único , Superóxido Dismutase/genética , Substituição de Aminoácidos , Animais , Arginina , Nefropatias Diabéticas/genética , Modelos Animais de Doenças , Técnicas de Transferência de Genes , Glicina , Humanos , Falência Renal Crônica/genética , Falência Renal Crônica/mortalidade , Falência Renal Crônica/terapia , Masculino , Isquemia Miocárdica/genética , Ratos , Ratos Endogâmicos SHR , Ratos Endogâmicos WKY , Proteínas Recombinantes/farmacologiaRESUMO
We tested the hypotheses that 1) systemic IL-10, after adenoviral gene transfer, protects arteries from impaired relaxation produced by LPS; 2) local expression of IL-10 within the arterial wall protects against vasomotor dysfunction after LPS; and 3) IL-10 protects against vascular dysfunction mediated by inducible NO synthase (iNOS) after LPS. In IL-10-deficient (IL-10-/-) and wild-type (WT, IL-10+/+) mice, LPS in vivo impaired relaxation of arteries to acetylcholine and gene transfer of IL-10 improved responses to acetylcholine. Superoxide levels were elevated in arteries after LPS, and increased levels of superoxide were prevented by gene transfer of IL-10. In arteries incubated with a low concentration of LPS in vitro to eliminate systemic effects of LPS and IL-10 from nonvascular sources, responses to acetylcholine were impaired in IL-10-deficient mice and impairment was largely prevented by gene transfer in vitro of IL-10. In arteries from WT mice in vitro, the low concentration of LPS did not impair responses to acetylcholine. Thus IL-10 within the vessel wall protects against LPS-induced dysfunction. In IL-10-deficient mice, aminoguanidine, which inhibits iNOS, protected against vasomotor dysfunction after LPS. In arteries from iNOS-deficient mice, LPS did not impair responses to acetylcholine. These findings suggest that both systemic and local effects of IL-10 provide important protection of arteries against an inflammatory stimulus and that IL-10 decreases iNOS-mediated impairment of vasorelaxation after LPS.
Assuntos
Artérias Carótidas/efeitos dos fármacos , Interleucina-10/farmacologia , Lipopolissacarídeos/farmacologia , Doenças Vasculares/prevenção & controle , Vasodilatação/efeitos dos fármacos , Acetilcolina/farmacologia , Animais , Adesão Celular , Endotélio Vascular , Técnicas de Transferência de Genes , Humanos , Técnicas In Vitro , Interleucina-10/deficiência , Interleucina-10/genética , Leucócitos/fisiologia , Masculino , Camundongos , Camundongos Knockout , Óxido Nítrico Sintase/deficiência , Óxido Nítrico Sintase/metabolismo , Óxido Nítrico Sintase Tipo II , Doenças Vasculares/induzido quimicamente , Vasodilatadores/farmacologiaRESUMO
We have examined the progression of vascular and neural deficits in Zucker rats, Zucker diabetic fatty (ZDF) diabetic rats, and age-matched lean ZDF rats from 8 to 40 wk of age. Both the ZDF diabetic and Zucker rats were glucose intolerant at 8 wk of age. The Zucker rats did not become hyperglycemic but were hyperinsulinemic through 32 wk of age. All ZDF diabetic rats became hyperglycemic by 8 wk of age. Through their life span, serum free fatty acids and triglycerides levels were significantly higher in Zucker and ZDF diabetic rats compared with age-matched lean ZDF rats. After 24 and 28 wk of age, endoneurial blood flow was significantly decreased in ZDF diabetic and Zucker rats. Motor nerve conduction velocity was significantly decreased after 12-14 wk of age in ZDF diabetic rats and at 32 wk of age in Zucker rats. ACh-mediated vascular relaxation of epineurial arterioles of the sciatic nerve was impaired after 8-10 wk of age in ZDF diabetic rats and after approximately 16 wk of age in Zucker rats. In contrast, vascular relaxation mediated by calcitonin gene-related peptide was impaired significantly after 28 wk of age in ZDF diabetic rats but not impaired in Zucker rats up to 40 wk of age. Markers of oxidative stress were differentially elevated in ZDF diabetic rats and Zucker rats. These data indicate that vascular and neural dysfunction develops in both Zucker and ZDF diabetic rats but at different rates, which may be the result of hyperglycemia.
Assuntos
Diabetes Mellitus Tipo 2/fisiopatologia , Angiopatias Diabéticas/fisiopatologia , Neuropatias Diabéticas/fisiopatologia , Síndrome Metabólica/fisiopatologia , Obesidade/fisiopatologia , Neuropatia Ciática/fisiopatologia , Animais , Glicemia/análise , Peso Corporal , Peptídeo Relacionado com Gene de Calcitonina/metabolismo , Diabetes Mellitus Tipo 2/complicações , Angiopatias Diabéticas/etiologia , Neuropatias Diabéticas/etiologia , Progressão da Doença , Hiperglicemia/complicações , Hiperglicemia/fisiopatologia , Hiperlipidemias/complicações , Hiperlipidemias/fisiopatologia , Insulina/metabolismo , Masculino , Síndrome Metabólica/complicações , Condução Nervosa , Obesidade/complicações , Ratos , Ratos Zucker , Neuropatia Ciática/etiologiaRESUMO
Lipopolysaccharide (LPS) impairs vascular function, in part by generation of reactive oxygen species. One goal of this study was to determine whether gene transfer of extracellular SOD (ECSOD) improves vascular responsiveness in LPS-treated rats. A second goal was to determine whether effects of ECSOD are dependent on the heparin-binding domain of the enzyme, which facilitates binding of ECSOD to the outside of cells. Adenoviruses containing ECSOD (AdECSOD), ECSOD with deletion of its heparin-binding domain (AdECSOD-HBD), or a control virus (AdLacZ) were injected intravenously into rats. Three days later, vehicle or LPS (10 mg/kg ip) was injected. After 24 h, vascular reactivity was examined in aortic rings in vitro. Maximum relaxation to acetylcholine was 95 +/- 1% (means +/- SE) after AdlacZ plus vehicle and 77 +/- 3% after AdlacZ plus LPS (P < 0.05). Responses to calcium ionophore A-23187 and submaximal concentrations of nitroprusside also were impaired by LPS. Gene transfer of ECSOD, but not AdECSOD-HBD, improved (P < 0.05) relaxation to acetylcholine and A-23187 after LPS. Maximum relaxation to acetylcholine was 88 +/- 3% after LPS plus AdECSOD. Superoxide was increased in aorta after LPS, and the levels were reduced after AdECSOD but not AdECSOD-HBD. LPS-induced adhesion of leukocytes to aortic endothelium was reduced by AdECSOD but not by AdECSOD-HBD. We conclude that after gene transfer in vivo, binding of ECSOD to arteries effectively decreases the numbers of adherent leukocytes and levels of superoxide and improves impaired endothelium-dependent relaxation produced by LPS.