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1.
Am J Physiol Heart Circ Physiol ; 302(1): H159-66, 2012 Jan 01.
Artigo em Inglês | MEDLINE | ID: mdl-22058149

RESUMO

Arginase can cause vascular dysfunction by competing with nitric oxide synthase for l-arginine and by increasing cell proliferation and collagen formation, which promote vascular fibrosis/stiffening. We have shown that increased arginase expression/activity contribute to vascular endothelial cell (EC) dysfunction. Here, we examined the roles of the two arginase isoforms, arginase I and II (AI and AII, respectively), in this process. Experiments were performed using streptozotocin-induced diabetic mice: wild-type (WT) mice and knockout mice lacking the AII isoform alone (AI(+/+)AII(-/-)) or in combination with partial deletion of AI (AI(+/-)AII (-/-)). EC-dependent vasorelaxation of aortic rings and arterial fibrosis and stiffness were assessed in relation to arginase activity and expression. Diabetes reduced mean EC-dependent vasorelaxation markedly in diabetic WT and AI(+/+)AII(-/-) aortas (53% and 44% vs. controls, respectively) compared with a 27% decrease in AI(+/-)AII (-/-) vessels. Coronary fibrosis was also increased in diabetic WT and AI(+/+)AII(-/-) mice (1.9- and 1.7-fold vs. controls, respectively) but was not altered in AI(+/-)AII (-/-) diabetic mice. Carotid stiffness was increased by 142% in WT diabetic mice compared with 51% in AI(+/+)AII(-/-) mice and 19% in AI(+/-)AII (-/-) mice. In diabetic WT and AI(+/+)AII(-/-) mice, aortic arginase activity and AI expression were significantly increased compared with control mice, but neither parameter was altered in AI(+/-)AII (-/-) mice. In summary, AI(+/-)AII (-/-) mice exhibit better EC-dependent vasodilation and less vascular stiffness and coronary fibrosis compared with diabetic WT and AI(+/+)AII(-/-) mice. These data indicate a major involvement of AI in diabetes-induced vascular dysfunction.


Assuntos
Arginase/metabolismo , Artérias/enzimologia , Diabetes Mellitus Experimental/complicações , Angiopatias Diabéticas/etiologia , Vasodilatação , Animais , Aorta/enzimologia , Aorta/fisiopatologia , Arginase/genética , Artérias/efeitos dos fármacos , Artérias/patologia , Artérias/fisiopatologia , Artérias Carótidas/enzimologia , Artérias Carótidas/fisiopatologia , Complacência (Medida de Distensibilidade) , Vasos Coronários/enzimologia , Vasos Coronários/fisiopatologia , Diabetes Mellitus Experimental/enzimologia , Diabetes Mellitus Experimental/genética , Diabetes Mellitus Experimental/patologia , Diabetes Mellitus Experimental/fisiopatologia , Angiopatias Diabéticas/enzimologia , Angiopatias Diabéticas/genética , Angiopatias Diabéticas/patologia , Angiopatias Diabéticas/fisiopatologia , Relação Dose-Resposta a Droga , Fibrose , Peróxido de Hidrogênio/metabolismo , Hidroxiprolina/metabolismo , Peroxidação de Lipídeos/efeitos dos fármacos , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Superóxidos/metabolismo , Vasoconstrição , Vasoconstritores/farmacologia , Vasodilatadores/farmacologia
2.
Circ Res ; 102(1): 95-102, 2008 Jan 04.
Artigo em Inglês | MEDLINE | ID: mdl-17967788

RESUMO

Increases in arginase activity have been reported in a variety of disease conditions characterized by vascular dysfunction. Arginase competes with NO synthase for their common substrate arginine, suggesting a cause and effect relationship. We tested this concept by experiments with streptozotocin diabetic rats and high glucose (HG)-treated bovine coronary endothelial cells (BCECs). Our studies showed that diabetes-induced impairment of vasorelaxation to acetylcholine was correlated with increases in reactive oxygen species and arginase activity and arginase I expression in aorta and liver. Treatment of diabetic rats with simvastatin (5 mg/kg per day, subcutaneously) or L-citrulline (50 mg/kg per day, orally) blunted these effects. Acute treatment of diabetic coronary arteries with arginase inhibitors also reversed the impaired vasodilation to acetylcholine. Treatment of BCECs with HG (25 mmol/L, 24 hours) also increased arginase activity. This effect was blocked by treatment with simvastatin (0.1 micromol/L), the Rho kinase inhibitor Y-27632 (10 micromol/L), or L-citrulline (1 mmol/L). Superoxide and active RhoA levels also were elevated in HG-treated BCECs. Furthermore, HG significantly diminished NO production in BCECs. Transfection of BCECs with arginase I small interfering RNA prevented the rise in arginase activity in HG-treated cells and normalized NO production, suggesting a role for arginase I in reduced NO production with HG. These results indicate that increased arginase activity in diabetes contributes to vascular endothelial dysfunction by decreasing L-arginine availability to NO synthase.


Assuntos
Arginase/metabolismo , Doença das Coronárias/enzimologia , Doença das Coronárias/etiologia , Complicações do Diabetes , Animais , Arginina/sangue , Arginina/metabolismo , Ligação Competitiva , Bovinos , Vasos Coronários/fisiopatologia , Diabetes Mellitus Experimental/induzido quimicamente , Diabetes Mellitus Experimental/complicações , Endotélio Vascular/fisiopatologia , Óxido Nítrico Sintase Tipo III/metabolismo , Ratos
3.
Cardiovasc Drug Rev ; 24(3-4): 275-90, 2006.
Artigo em Inglês | MEDLINE | ID: mdl-17214603

RESUMO

L-citrulline is the natural precursor of L-arginine, substrate for nitric oxide synthase (NOS) in the production of NO. Supplemental administration L-arginine has been shown to be effective in improving NO production and cardiovascular function in cardiovascular diseases associated with endothelial dysfunction, such as hypertension, heart failure, atherosclerosis, diabetic vascular disease and ischemia-reperfusion injury, but the beneficial actions do not endure with chronic therapy. Substantial intestinal and hepatic metabolism of L-arginine to ornithine and urea by arginase makes oral delivery very ineffective. Additionally, all of these disease states as well as supplemental L-arginine enhance arginase expression and activity, thus reducing the effectiveness of L-arginine therapy. In contrast, L-citrulline is not metabolized in the intestine or liver and does not induce tissue arginase, but rather inhibits its activity. L-citrulline entering the kidney, vascular endothelium and other tissues can be readily converted to L-arginine, thus raising plasma and tissue levels of L-arginine and enhancing NO production. Supplemental L-citrulline has promise as a therapeutic adjunct in disease states associated with L-arginine deficiencies.


Assuntos
Arginina/metabolismo , Doenças Cardiovasculares/tratamento farmacológico , Citrulina/uso terapêutico , Animais , Citrulina/metabolismo , Citrulina/farmacologia , Células Endoteliais/metabolismo , Humanos , Mucosa Intestinal/metabolismo , Rim/metabolismo , Miócitos de Músculo Liso/metabolismo
4.
Free Radic Biol Med ; 39(10): 1353-61, 2005 Nov 15.
Artigo em Inglês | MEDLINE | ID: mdl-16257644

RESUMO

Increased expression of vascular endothelial growth factor (VEGF) has been correlated with increased oxidative stress and formation of peroxynitrite in numerous disease conditions, including diabetic microangiopathy, tumor angiogenesis, and atherosclerosis. In this study we tested the hypothesis that peroxynitrite stimulates VEGF expression. Treatment of microvascular endothelial cells with exogenous peroxynitrite induced a time- and dose-dependent increase in VEGF mRNA, which peaked within 1 h of treatment at a concentration of 100 muM. The increase in VEGF mRNA was followed by a significant increase in VEGF protein. To define the molecular mechanisms involved, the effect of peroxynitrite was determined on the activation of two transcription factors known to regulate VEGF expression during hypoxia and tumor angiogenesis-signal transducer and activator of transcription 3 (STAT3) and hypoxia-inducible factor-1 (HIF-1). Peroxynitrite caused activation and nuclear translocation of STAT3, but not HIF-1. Moreover, transduction of endothelial cells with dominant-negative STAT3 abrogated the peroxynitrite-induced increase in VEGF mRNA. The increase in VEGF mRNA was also blocked by inhibitors of transcription and was unaffected by the inhibition of protein synthesis. These results indicate that peroxynitrite causes increased expression of VEGF in vascular endothelial cells by a process that requires the activation of STAT3.


Assuntos
Endotélio Vascular/citologia , Ácido Peroxinitroso/farmacologia , Fator de Transcrição STAT3/metabolismo , Fator A de Crescimento do Endotélio Vascular/biossíntese , Transporte Ativo do Núcleo Celular , Adenoviridae/genética , Animais , Western Blotting , Bovinos , Movimento Celular , Núcleo Celular/metabolismo , Células Cultivadas , Relação Dose-Resposta a Droga , Endotélio Vascular/metabolismo , Ensaio de Imunoadsorção Enzimática , Radicais Livres , Regulação da Expressão Gênica , Genes Dominantes , Vetores Genéticos , Proteínas de Fluorescência Verde/metabolismo , Fator 1 Induzível por Hipóxia/metabolismo , Immunoblotting , Microcirculação , Microscopia de Fluorescência , Músculo Liso/citologia , Estresse Oxidativo , Ácido Peroxinitroso/química , Ácido Peroxinitroso/metabolismo , RNA Mensageiro/metabolismo , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Fatores de Tempo , Transcrição Gênica , Fator A de Crescimento do Endotélio Vascular/genética , Fator A de Crescimento do Endotélio Vascular/metabolismo
5.
Am J Pathol ; 167(2): 599-607, 2005 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-16049343

RESUMO

Because oxidative stress has been strongly implicated in up-regulation of vascular endothelial growth factor (VEGF) expression in ischemic retinopathy, we evaluated the role of NAD(P)H oxidase in causing VEGF overexpression and retinal neovascularization. Dihydroethidium imaging analyses showed increased superoxide formation in areas of retinal neovascularization associated with relative retinal hypoxia in a mouse model for oxygen-induced retinopathy. The effect of hypoxia in stimulating superoxide formation in retinal vascular endothelial cells was confirmed by in vitro chemiluminescence assays. The superoxide formation was blocked by specific inhibitors of NAD(P)H oxidase activity (apocynin, gp91ds-tat) indicating that NAD(P)H oxidase is a major source of superoxide formation. Western blot and immunolocalization analyses showed that retinal ischemia increased expression of the NAD(P)H oxidase catalytic subunit gp91phox, which localized primarily within vascular endothelial cells. Treatment of mice with apocynin blocked ischemia-induced increases in oxidative stress, normalized VEGF expression, and prevented retinal neovascularization. Apocynin and gp91ds-tat also blocked the action of hypoxia in causing increased VEGF expression in vitro, confirming the specific role of NAD(P)H oxidase in hypoxia-induced increases in VEGF expression. In conclusion, NAD(P)H oxidase activity is required for hypoxia-stimulated increases in VEGF expression and retinal neovascularization. Inhibition of NAD(P)H oxidase offers a new therapeutic target for the treatment of retinopathy.


Assuntos
Isquemia , NADPH Oxidases/antagonistas & inibidores , Doenças Retinianas/patologia , Neovascularização Retiniana/prevenção & controle , Vasos Retinianos/patologia , Fator A de Crescimento do Endotélio Vascular/metabolismo , Acetofenonas/farmacologia , Animais , Bovinos , Células Cultivadas , Endotélio Vascular/metabolismo , Endotélio Vascular/patologia , Inibidores Enzimáticos/farmacologia , Hipóxia , Glicoproteínas de Membrana/metabolismo , Camundongos , Camundongos Endogâmicos C57BL , NADPH Oxidase 2 , NADPH Oxidases/metabolismo , Oxigênio , Retina/citologia , Retina/metabolismo , Retina/patologia , Doenças Retinianas/etiologia , Doenças Retinianas/prevenção & controle , Superóxidos/metabolismo , Regulação para Cima
6.
Curr Drug Targets ; 6(4): 511-24, 2005 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-16026270

RESUMO

Retinal neovascularization and macular edema are central features of diabetic retinopathy, a major cause of blindness in working age adults. The currently established treatment for diabetic retinopathy targets the vascular pathology by laser photocoagulation. This approach is associated with significant adverse effects due the destruction of neural tissue and is not always effective. Characterization of the molecular and cellular processes involved in vascular growth and hyperpermeability has led to the recognition that the angiogenic growth factor and vascular permeability factor VEGF (vascular endothelial growth factor) play a pivotal role in the retinal microvascular complications of diabetes. Thus, VEGF represents an important target for therapeutic intervention in diabetic retinopathy. Agents that directly inhibit the actions of VEGF and its receptors show considerable promise, but have not proven to be completely effective in blocking pathological angiogenesis. Therefore, a better understanding of the molecular events that control VEGF expression and mediate its downstream actions is important to define more precise therapeutic targets for intervention in diabetic retinopathy. This review highlights the current understanding of the process by which VEGF gene expression is regulated and how VEGF's biological effects are altered during diabetes. In particular, cellular and molecular alterations seen in diabetic models are considered in the context of high glucose-mediated oxidative stress effects on VEGF expression and action. Potential therapeutic strategies for preventing VEGF overexpression or blocking its pathological actions in the diabetic retina are considered.


Assuntos
Retinopatia Diabética/etiologia , Estresse Oxidativo , Fator A de Crescimento do Endotélio Vascular/fisiologia , Corticosteroides/uso terapêutico , Animais , Antioxidantes/uso terapêutico , Canabinoides/uso terapêutico , Permeabilidade Capilar , Sobrevivência Celular , Retinopatia Diabética/tratamento farmacológico , Retinopatia Diabética/metabolismo , Proteínas do Olho/fisiologia , Proteínas do Olho/uso terapêutico , Humanos , Inibidores de Hidroximetilglutaril-CoA Redutases/uso terapêutico , Neovascularização Fisiológica , Fatores de Crescimento Neural/fisiologia , Fatores de Crescimento Neural/uso terapêutico , Receptores de Fatores de Crescimento do Endotélio Vascular/fisiologia , Serpinas/fisiologia , Serpinas/uso terapêutico , Fator A de Crescimento do Endotélio Vascular/antagonistas & inibidores
7.
Diabetes Metab Res Rev ; 19(6): 442-55, 2003.
Artigo em Inglês | MEDLINE | ID: mdl-14648803

RESUMO

Retinal neovascularization and macular edema are central features of diabetic retinopathy, the major cause of blindness in the developed world. Current treatments are limited in their efficacy and are associated with significant adverse effects. Characterization of the molecular and cellular processes involved in vascular growth and permeability has led to the recognition that the angiogenic growth factor and vascular permeability factor vascular endothelial growth factor (VEGF) plays a pivotal role in the retinal microvascular complications of diabetes. Therefore, VEGF represents an exciting target for therapeutic intervention in diabetic retinopathy. This review highlights the current understanding of the mechanisms that regulate VEGF gene expression and mediate its biological effects and how these processes may become altered during diabetes. The cellular and molecular alterations that characterize experimental models of diabetes are considered in relation to the influence of high glucose-mediated oxidative stress on VEGF expression and on the mechanisms of VEGF's actions under hyperglycemic induction. Finally, potential therapeutic strategies for preventing VEGF overexpression or blocking its pathological effects in the diabetic retina are considered.


Assuntos
Retinopatia Diabética/fisiopatologia , Fator A de Crescimento do Endotélio Vascular/fisiologia , Retinopatia Diabética/etiologia , Retinopatia Diabética/terapia , Glicosilação , Humanos , Degeneração Macular/etiologia , Neovascularização Patológica/etiologia , Espécies Reativas de Oxigênio/efeitos adversos , Receptores de Fatores de Crescimento do Endotélio Vascular/fisiologia
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