RESUMO
Arteriogenesis requires growth of pre-existing arteriolar collateral networks and determines clinical outcome in arterial occlusive diseases. Factors responsible for the development of arteriolar collateral networks are poorly understood. The Notch ligand Delta-like 4 (Dll4) promotes arterial differentiation and restricts vessel branching. We hypothesized that Dll4 may act as a genetic determinant of collateral arterial networks and functional recovery in stroke and hind limb ischemia models in mice. Genetic loss- and gain-of-function approaches in mice showed that Dll4-Notch signaling restricts pial collateral artery formation by modulating arterial branching morphogenesis during embryogenesis. Adult Dll4(+/-) mice showed increased pial collateral numbers, but stroke volume upon middle cerebral artery occlusion was not reduced compared with wild-type littermates. Likewise, Dll4(+/-) mice showed reduced blood flow conductance after femoral artery occlusion, and, despite markedly increased angiogenesis, tissue ischemia was more severe. In peripheral arteries, loss of Dll4 adversely affected excitation-contraction coupling in arterial smooth muscle in response to vasopressor agents and arterial vessel wall adaption in response to increases in blood flow, collectively contributing to reduced flow reserve. We conclude that Dll4-Notch signaling modulates native collateral formation by acting on vascular branching morphogenesis during embryogenesis. Dll4 furthermore affects tissue perfusion by acting on arterial function and structure. Loss of Dll4 stimulates collateral formation and angiogenesis, but in the context of ischemic diseases such beneficial effects are overruled by adverse functional changes, demonstrating that ischemic recovery is not solely determined by collateral number but rather by vessel functionality.
Assuntos
Peptídeos e Proteínas de Sinalização Intracelular/metabolismo , Isquemia/fisiopatologia , Proteínas de Membrana/metabolismo , Microvasos/embriologia , Morfogênese/fisiologia , Neovascularização Fisiológica/fisiologia , Receptores Notch/metabolismo , Transdução de Sinais/fisiologia , Proteínas Adaptadoras de Transdução de Sinal , Análise de Variância , Animais , Proteínas de Ligação ao Cálcio , Imuno-Histoquímica , Isquemia/metabolismo , Camundongos , Microvasos/fisiologia , Reação em Cadeia da Polimerase em Tempo Real , Fluxo Sanguíneo Regional/fisiologia , Microtomografia por Raio-XRESUMO
RATIONALE: The lymphatic vasculature plays a major role in fluid homeostasis, absorption of dietary lipids, and immune surveillance. Fluid transport depends on the presence of intraluminal valves within lymphatic collectors. Defective formation of lymphatic valves leads to lymphedema, a progressive and debilitating condition for which curative treatments are currently unavailable. How lymphatic valve formation is regulated remains largely unknown. OBJECTIVE: We investigated if the repulsive axon guidance molecule Semaphorin3A (Sema3A) plays a role in lymphatic valve formation. METHODS AND RESULTS: We show that Sema3A mRNA is expressed in lymphatic vessels and that Sema3A protein binds to lymphatic valves expressing the Neuropilin-1 (Nrp1) and PlexinA1 receptors. Using mouse knockout models, we show that Sema3A is selectively required for lymphatic valve formation, via interaction with Nrp1 and PlexinA1. Sema3a(-/-) mice exhibit defects in lymphatic valve formation, which are not due to abnormal lymphatic patterning or sprouting, and mice carrying a mutation in the Sema3A binding site of Nrp1, or deficient for Plxna1, develop lymphatic valve defects similar to those seen in Sema3a(-/-) mice. CONCLUSIONS: Our data demonstrate an essential direct function of Sema3A-Nrp1-PlexinA1 signaling in lymphatic valve formation.
Assuntos
Vasos Linfáticos/metabolismo , Proteínas do Tecido Nervoso/metabolismo , Neuropilina-1/metabolismo , Receptores de Superfície Celular/metabolismo , Semaforina-3A/metabolismo , Animais , Animais Recém-Nascidos , Anticorpos Neutralizantes/administração & dosagem , Proteínas de Bactérias/genética , Proteínas de Bactérias/metabolismo , Células Cultivadas , Células Endoteliais/metabolismo , Regulação da Expressão Gênica no Desenvolvimento , Genótipo , Idade Gestacional , Humanos , Proteínas Luminescentes/genética , Proteínas Luminescentes/metabolismo , Vasos Linfáticos/embriologia , Camundongos , Camundongos Knockout , Camundongos Transgênicos , Morfogênese , Proteínas do Tecido Nervoso/deficiência , Proteínas do Tecido Nervoso/genética , Neuropilina-1/deficiência , Neuropilina-1/genética , Neuropilina-1/imunologia , Fenótipo , RNA Mensageiro/metabolismo , Receptores de Superfície Celular/deficiência , Receptores de Superfície Celular/genética , Semaforina-3A/deficiência , Semaforina-3A/genética , Receptor 3 de Fatores de Crescimento do Endotélio Vascular/genética , Receptor 3 de Fatores de Crescimento do Endotélio Vascular/metabolismoRESUMO
BACKGROUND: Arteriogenesis and collateral formation are complex processes requiring integration of multiple inputs to coordinate vessel branching, growth, maturation, and network size. Factors regulating these processes have not been determined. METHODS AND RESULTS: We used an inhibitor of NFκB activation (IκBαSR) under control of an endothelial-specific inducible promoter to selectively suppress endothelial nuclear factor-κB activation during development, in the adult vasculature, or in vitro. Inhibition of nuclear factor-κB activation resulted in formation of an excessively branched arterial network that was composed of immature vessels and provided poor distal tissue perfusion. Molecular analysis demonstrated reduced adhesion molecule expression leading to decreased monocyte influx, reduced hypoxia-inducible factor-1α levels, and a marked decrease in δ-like ligand 4 expression with a consequent decrease in Notch signaling. The latter was the principal cause of increased vascular branching as treatment with Jagged-1 peptide reduced the size of the arterial network to baseline levels. CONCLUSIONS: These findings identify nuclear factor-κB as a key regulator of adult and developmental arteriogenesis and collateral formation. Nuclear factor-κB achieves this by regulating hypoxia-inducible factor-1α-dependent expression of vascular endothelial growth factor-A and platelet-derived growth factor-BB, which are necessary for the development and maturation of the arterial collateral network, and by regulating δ-like ligand 4 expression, which in turn determines the size and complexity of the network.
Assuntos
Células Endoteliais/metabolismo , Isquemia/fisiopatologia , Subunidade p50 de NF-kappa B/metabolismo , Neovascularização Patológica/fisiopatologia , Neovascularização Fisiológica/fisiologia , Animais , Animais Recém-Nascidos , Becaplermina , Encéfalo/metabolismo , Modelos Animais de Doenças , Membro Posterior/irrigação sanguínea , Células Endoteliais da Veia Umbilical Humana , Humanos , Subunidade alfa do Fator 1 Induzível por Hipóxia/metabolismo , Isquemia/metabolismo , Camundongos , Camundongos Transgênicos , Subunidade p50 de NF-kappa B/genética , Neovascularização Patológica/metabolismo , Proteínas Proto-Oncogênicas c-sis/metabolismo , Retina/metabolismo , Fator A de Crescimento do Endotélio Vascular/metabolismoRESUMO
Inflammatory neovascularization, such as choroidal neovascularization (CNV), occur in the presence of Notch expressing macrophages. DLL4s anti-angiogenic effect on endothelial cells (EC) has been widely recognized, but its influence on Notch signaling on macrophages and its overall effect in inflammatory neovascularization is not well understood. We identified macrophages and ECs as the main Notch 1 and Notch 4 expressing cells in CNV. A soluble fraction spanning Ser28-Pro525 of the murine extracellular DLL4 domain (sDLL4/28-525) activated the Notch pathway, as it induces Notch target genes in macrophages and ECs and inhibited EC proliferation and vascular sprouting in aortic rings. In contrast, sDLL4/28-525 increased pro-angiogenic VEGF, and IL-1ß expression in macrophages responsible for increased vascular sprouting observed in aortic rings incubated in conditioned media from sDLL4/28-525 stimulated macrophages. In vivo, Dll4(+/-) mice developed significantly more CNV and sDLL4/28-525 injections inhibited CNV in Dll4(+/-) CD1 mice. Similarly, sDLL4/28-525 inhibited CNV in C57Bl6 and its effect was reversed by a γ-secretase inhibitor that blocks Notch signaling. The inhibition occurred despite increased VEGF, IL-1ß expression in infiltrating inflammatory macrophages in sDLL4/28-525 treated mice and might be due to direct inhibition of EC proliferation in laser-induced CNV as demonstrated by EdU labelling in vivo. In conclusion, Notch activation on macrophages and ECs leads to opposing effects in inflammatory neovascularization in situations such as CNV.
Assuntos
Neovascularização de Coroide/prevenção & controle , Endotélio Vascular/fisiopatologia , Peptídeos e Proteínas de Sinalização Intercelular/fisiologia , Macrófagos Peritoneais/fisiologia , Proteínas Adaptadoras de Transdução de Sinal , Animais , Sequência de Bases , Western Blotting , Proteínas de Ligação ao Cálcio , Primers do DNA , Camundongos , Camundongos Endogâmicos C57BL , Reação em Cadeia da Polimerase Via Transcriptase ReversaRESUMO
OBJECTIVE: To investigate the novel hypothesis that neurotrophin-3 (NT-3), an established neurotrophic factor that participates in embryonic heart development, promotes blood vessel growth. METHODS AND RESULTS: We evaluated the proangiogenic capacity of recombinant NT-3 in vitro and of NT-3 gene transfer in vivo (rat mesenteric angiogenesis assay and mouse normoperfused adductor muscle). Then, we studied whether either transgenic or endogenous NT-3 mediates postischemic neovascularization in a mouse model of limb ischemia. In vitro, NT-3 stimulated endothelial cell survival, proliferation, migration, and network formation on the basement membrane matrix Matrigel. In the mesenteric assay, NT-3 increased the number and size of functional vessels, including vessels covered with mural cells. Consistently, NT-3 overexpression increased muscular capillary and arteriolar densities in either the absence or the presence of ischemia and improved postischemic blood flow recovery in mouse hind limbs. NT-3-induced microvascular responses were accompanied by tropomyosin receptor kinase C (an NT-3 high-affinity receptor) phosphorylation and involved the phosphatidylinositol 3-kinase-Akt kinase-endothelial nitric oxide synthase pathway. Finally, endogenous NT-3 was shown to be essential in native postischemic neovascularization, as demonstrated by using a soluble tropomyosin receptor kinase C receptor domain that neutralizes NT-3. CONCLUSIONS: Our results provide the first insight into the proangiogenic capacity of NT-3 and propose NT-3 as a novel potential agent for the treatment of ischemic disease.
Assuntos
Proteínas Angiogênicas/metabolismo , Células Endoteliais/metabolismo , Terapia Genética , Isquemia/terapia , Mesentério/irrigação sanguínea , Músculo Esquelético/irrigação sanguínea , Neovascularização Fisiológica , Neurotrofina 3/metabolismo , Proteínas Angiogênicas/genética , Proteínas Angiogênicas/farmacologia , Animais , Movimento Celular , Proliferação de Células , Sobrevivência Celular , Células Cultivadas , Modelos Animais de Doenças , Células Endoteliais/efeitos dos fármacos , Células Endoteliais/patologia , Terapia Genética/métodos , Membro Posterior , Humanos , Isquemia/genética , Isquemia/metabolismo , Isquemia/patologia , Isquemia/fisiopatologia , Masculino , Camundongos , Neovascularização Fisiológica/efeitos dos fármacos , Fatores de Crescimento Neural/metabolismo , Neurotrofina 3/genética , Neurotrofina 3/farmacologia , Óxido Nítrico Sintase Tipo III/metabolismo , Fosfatidilinositol 3-Quinases/metabolismo , Fosforilação , Proteínas Proto-Oncogênicas c-akt/metabolismo , RNA Mensageiro/metabolismo , Ratos , Ratos Wistar , Receptor trkC/genética , Receptor trkC/metabolismo , Proteínas Recombinantes/metabolismo , Transdução de Sinais , Fatores de Tempo , TransfecçãoRESUMO
Diabetes impairs endothelial function and reparative neovascularization. The p75 receptor of neurotrophins (p75(NTR)), which is scarcely present in healthy endothelial cells (ECs), becomes strongly expressed by capillary ECs after induction of peripheral ischemia in type-1 diabetic mice. Here, we show that gene transfer-induced p75(NTR) expression impairs the survival, proliferation, migration, and adhesion capacities of cultured ECs and endothelial progenitor cells (EPCs) and inhibits angiogenesis in vitro. Moreover, intramuscular p75(NTR) gene delivery impairs neovascularization and blood flow recovery in a mouse model of limb ischemia. These disturbed functions are associated with suppression of signaling mechanisms implicated in EC survival and angiogenesis. In fact, p75(NTR) depresses the VEGF-A/Akt/eNOS/NO pathway and additionally reduces the mRNA levels of ITGB1 [beta (1) integrin], BIRC5 (survivin), PTTG1 (securin) and VEZF1. Diabetic mice, which typically show impaired postischemic muscular neovascularization and blood perfusion recovery, have these defects corrected by intramuscular gene transfer of a dominant negative mutant form of p75(NTR). Collectively, our data newly demonstrate the antiangiogenic action of p75(NTR) and open new avenues for the therapeutic use of p75(NTR) inhibition to combat diabetes-induced microvascular liabilities.
Assuntos
Apoptose/fisiologia , Angiopatias Diabéticas/metabolismo , Endotélio Vascular/fisiologia , Isquemia/metabolismo , Músculo Esquelético/irrigação sanguínea , Neovascularização Fisiológica/fisiologia , Receptor de Fator de Crescimento Neural/metabolismo , Animais , Células Cultivadas , Diabetes Mellitus Experimental/complicações , Diabetes Mellitus Experimental/metabolismo , Diabetes Mellitus Experimental/patologia , Modelos Animais de Doenças , Endotélio Vascular/patologia , Humanos , Isquemia/etiologia , Isquemia/patologia , Masculino , Camundongos , Camundongos Endogâmicos , Óxido Nítrico/metabolismo , Óxido Nítrico Sintase/metabolismo , Proteínas Proto-Oncogênicas c-akt/metabolismo , Receptor de Fator de Crescimento Neural/genética , Transdução de Sinais/fisiologia , Estreptozocina , Transfecção , Fator A de Crescimento do Endotélio Vascular/metabolismoRESUMO
An increasing number of studies about the molecular basis of angiogenesis are rapidly disclosing novel signal pathways involved in the blood vessel formation process. This review will focus on bone morphogenic proteins, Hedgehog, Notch, ephrins, neuropilins, neurotrophins and netrins. These recently discovered angiogenesis mediators are involved in vascular development during embryogenesis and, interestingly, they are shared between the nervous and vascular systems. They represent new potential targets in the vasculature and suggest novel therapeutic opportunities.
Assuntos
Inibidores da Angiogênese/uso terapêutico , Neovascularização Patológica/tratamento farmacológico , Neovascularização Fisiológica/fisiologia , Animais , Proteínas Morfogenéticas Ósseas/fisiologia , Sistemas de Liberação de Medicamentos , Efrinas/fisiologia , Proteínas Hedgehog/fisiologia , Humanos , Fatores de Crescimento Neural/fisiologia , Netrina-1 , Neuropilinas/fisiologia , Receptores da Família Eph/fisiologia , Receptores Notch/fisiologia , Proteínas Supressoras de Tumor/fisiologiaRESUMO
Most characterized angiogenic modulators are proteolytic fragments of structural plasma and/or matrix components. Herein, we have identified a novel anti-angiogenic peptide generated by the in vitro hydrolysis of the C-terminal moiety of the fibrinogen alpha chain, produced by the snake venom metalloprotease bothropasin (SVMP), a hemorrhagic proteinase in Bothrops jararaca venom. The 14-amino acids peptide (alphastatin-C) is a potent antagonist of basic fibroblast growth factor, induced endothelial cell (HUVEC-CS) proliferation, migration and capillary tube formation in matrigel. It also inhibits cell adhesion to fibronectin. The basis of the antagonism between bFGF and alphastatin-C is elucidated by the inhibition of various bFGF induced signaling pathways and their molecular components modification, whenever the combination of the stimuli is provided, in comparison to the treatment with bFGF only. To corroborate to the potential therapeutic use of alphastatin-C, we have chosen to perform in vivo assays in two distinct angiogenic settings. In chick model, alphastatin-C inhibits chorioallantoic membrane angiogenesis. In mouse, it efficiently reduces tumor number and volume in a melanoma model, due to the impairment of tumor neovascularization in treated mice. In contrast, we show that the alphastatin-C peptide induces arteriogenesis, increasing pial collateral density in neonate mice. alphastatin-C is an efficient new antiangiogenic FGF-associated agent in vitro, it is an inhibitor of embryonic and tumor vascularization in vivo while, it is an arteriogenic agent. The results also suggest that SVMPs can be used as in vitro biochemical tools to process plasma and/or matrix macromolecular components unraveling new angiostatic peptides.
RESUMO
Diabetes mellitus is associated with macro- and micro-angiopathy, leading to increased risk of peripheral ischemia. In the present study, we have characterized the microvascular phenotype at the level of limb muscles and the spontaneous angiogenesis response to surgically-induced unilateral limb ischemia in a murine model of type-2 diabetes, the obese C57BL/KsOlaHsd-Lepr(db/db) mice (Lepr(db/db)), and in non-diabetic heterozygous Lepr(db/+). Wild type C57BL mice (WT) were used as controls. The basal microvascular phenotype was determined in mice aged 3 or 5 months, while the response to limb ischemia was studied only in 5-month old mice. Moreover, in 5-month old ischemic Lepr(db/db) and Lepr(db/+), we have tested the therapeutic potential of local angiogenesis gene therapy with human tissue kallikrein (hTK) or constitutively-activated Akt kinase (Myr-Akt). We found that in the muscles of 3- or 5-month old Lepr(db/db), apoptosis of endothelial cells was enhanced and the densities of capillary and arteriole were reduced. Arterioles of Lepr(db/db) showed hypertrophic remodelling and, occasionally, lumen occlusion. Following ischemia, Lepr(db/db) showed a defective reparative angiogenesis in ischemic muscle, delayed blood flow recovery, and worsened clinical outcome as compared with controls. Five-month old Lepr(db/+) displayed an increase in endothelial cell apoptosis under basal conditions, while capillary and arteriole densities were normal. Lepr(db/+) mounted a proper reparative angiogenesis response to limb ischemia and regained blood flow to the ischemic limb, regularly. Local gene therapy with hTK or Myr-Akt induced angiogenesis in ischemic muscles of Lepr(db/+) and Lepr(db/db). However, in the Lepr(db/db) neither gene therapy approach improved the blood flow recovery and the clinical outcome from ischemia. In contrast, either hTK or Myr-Akt gene transfer improved the post-ischemic recovery of Lepr(db/+). Type-2 diabetes has a negative impact on the basal microvascular phenotype and severely impairs post-ischemic recovery of limb muscles. Gene therapy-induced stimulation of neovascularization might not suffice as a sole therapeutic strategy to combat type-2 diabetes-related vascular complications. In type-2 diabetic patients, therapeutic angiogenesis may need to be further optimized before being recommended for clinical applications.
Assuntos
Diabetes Mellitus Tipo 2/complicações , Angiopatias Diabéticas/terapia , Terapia Genética , Isquemia/terapia , Neovascularização Fisiológica , Animais , Capilares/patologia , Angiopatias Diabéticas/patologia , Angiopatias Diabéticas/fisiopatologia , Humanos , Isquemia/patologia , Isquemia/fisiopatologia , Extremidade Inferior/irrigação sanguínea , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Obesos , Músculo Esquelético/irrigação sanguínea , Músculo Esquelético/metabolismo , Óxido Nítrico Sintase Tipo III/metabolismo , Fenótipo , Proteínas Proto-Oncogênicas c-akt/genética , Receptores de Superfície Celular/genética , Receptores para Leptina , Calicreínas Teciduais/genética , Fator A de Crescimento do Endotélio Vascular/metabolismoRESUMO
Resveratrol (RES) is a natural occurring phytoalexin that has been shown to have chemopreventive activity. Resveratrol acts both by suppressing cell proliferation and inducing apoptosis in a variety of cancer cell lines. In this study, we show that RES induces apoptosis in MOLT-4 acute lymphoblastic leukaemia cells by modulating three different pathways that regulate cells survival and cell death. We show for the first time that RES inhibits the survival signalling pathways Notch and their down stream effector and modulates the operation of interacting signalling systems. It induces an increase in the levels of the pro-apoptotic proteins p53, its effector p21waf and Bax. We also show that RES inhibits the PI3K/Akt pathway and activates Gsk-3beta. The data presented here demonstrate unequivocally that RES induces apoptosis by inhibiting the Notch pathway and markedly influencing the operation of the interacting apoptosis pathways mediated by p53 and PI3K/Akt. These data support findings from other laboratories that have suggested the use of RES as a chemopreventive agent. Here, we have identified potential signalling pathways influenced by RES and this could lead to the identification of the targets of RES-induced apoptosis and growth control.
Assuntos
Apoptose/efeitos dos fármacos , Estilbenos/farmacologia , Anticarcinógenos/farmacologia , Western Blotting , Linhagem Celular Tumoral , Proliferação de Células/efeitos dos fármacos , Sobrevivência Celular/efeitos dos fármacos , Relação Dose-Resposta a Droga , Ativação Enzimática/efeitos dos fármacos , Regulação Neoplásica da Expressão Gênica/efeitos dos fármacos , Quinase 3 da Glicogênio Sintase/genética , Quinase 3 da Glicogênio Sintase/metabolismo , Glicogênio Sintase Quinase 3 beta , Humanos , Leucemia-Linfoma de Células T do Adulto/metabolismo , Leucemia-Linfoma de Células T do Adulto/patologia , Cloreto de Lítio/farmacologia , Fosfatidilinositol 3-Quinases/metabolismo , Inibidores de Fosfoinositídeo-3 Quinase , Fosforilação/efeitos dos fármacos , Proteínas Proto-Oncogênicas c-akt/genética , Proteínas Proto-Oncogênicas c-akt/metabolismo , Receptores Notch/genética , Receptores Notch/metabolismo , Resveratrol , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Sesquiterpenos , Transdução de Sinais/efeitos dos fármacos , Terpenos/farmacologia , Fatores de Tempo , Proteína Supressora de Tumor p53/genética , Proteína Supressora de Tumor p53/metabolismo , Proteína X Associada a bcl-2/genética , Proteína X Associada a bcl-2/metabolismo , FitoalexinasRESUMO
Ocular neovascular diseases are a leading cause of blindness. Vascular endothelial growth factor (VEGF) blockade improves vision, but not all individuals respond to anti-VEGF treatment, making additional means to prevent neovascularization necessary. Slit-family proteins (Slits) are ligands of Roundabout (Robo) receptors that repel developing axons in the nervous system. Robo1 expression is altered in ocular neovascular diseases, and previous in vitro studies have reported both pro- and anti-angiogenic effects of Slits. However, genetic evidence supporting a role for Slits in ocular neovascularization is lacking. Here we generated conditional knockout mice deficient in various Slit and Robo proteins and found that Slit2 potently and selectively promoted angiogenesis via Robo1 and Robo2 in mouse postnatal retina and in a model of ocular neovascular disease. Mechanistically, Slit2 acting through Robo1 and Robo2 promoted the migration of endothelial cells. These receptors are required for both Slit2- and VEGF-induced Rac1 activation and lamellipodia formation. Thus, Slit2 blockade could potentially be used therapeutically to inhibit angiogenesis in individuals with ocular neovascular disease.
Assuntos
Peptídeos e Proteínas de Sinalização Intercelular/metabolismo , Proteínas do Tecido Nervoso/metabolismo , Receptores Imunológicos/metabolismo , Neovascularização Retiniana , Animais , Apoptose , Movimento Celular/genética , Proliferação de Células , Modelos Animais de Doenças , Células Endoteliais/citologia , Regulação da Expressão Gênica no Desenvolvimento , Células Endoteliais da Veia Umbilical Humana , Humanos , Pulmão/metabolismo , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Neovascularização Patológica , RNA Mensageiro/metabolismo , Retina/embriologia , Retina/metabolismo , Transdução de Sinais/efeitos dos fármacos , Fator A de Crescimento do Endotélio Vascular/metabolismo , Proteínas RoundaboutRESUMO
Autonomic sympathetic nerves innervate peripheral resistance arteries, thereby regulating vascular tone and controlling blood supply to organs. Despite the fundamental importance of blood flow control, how sympathetic arterial innervation develops remains largely unknown. Here, we identified the axon guidance cue netrin-1 as an essential factor required for development of arterial innervation in mice. Netrin-1 was produced by arterial smooth muscle cells (SMCs) at the onset of innervation, and arterial innervation required the interaction of netrin-1 with its receptor, deleted in colorectal cancer (DCC), on sympathetic growth cones. Function-blocking approaches, including cell type-specific deletion of the genes encoding Ntn1 in SMCs and Dcc in sympathetic neurons, led to severe and selective reduction of sympathetic innervation and to defective vasoconstriction in resistance arteries. These findings indicate that netrin-1 and DCC are critical for the control of arterial innervation and blood flow regulation in peripheral organs.