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AIMS: Here, we aimed to determine the therapeutic effect of longevity-associated variant (LAV)-BPIFB4 gene therapy on atherosclerosis. METHODS AND RESULTS: ApoE knockout mice (ApoE-/-) fed a high-fat diet were randomly allocated to receive LAV-BPIFB4, wild-type (WT)-BPIFB4, or empty vector via adeno-associated viral vector injection. The primary endpoints of the study were to assess (i) vascular reactivity and (ii) atherosclerotic disease severity, by Echo-Doppler imaging, histology and ultrastructural analysis. Moreover, we assessed the capacity of the LAV-BPIFB4 protein to shift monocyte-derived macrophages of atherosclerotic mice and patients towards an anti-inflammatory phenotype. LAV-BPIFB4 gene therapy rescued endothelial function of mesenteric and femoral arteries from ApoE-/- mice; this effect was blunted by AMD3100, a CXC chemokine receptor type 4 (CXCR4) inhibitor. LAV-BPIFB4-treated mice showed a CXCR4-mediated shift in the balance between Ly6Chigh/Ly6Clow monocytes and M2/M1 macrophages, along with decreased T cell proliferation and elevated circulating levels of interleukins IL-23 and IL-27. In vitro conditioning with LAV-BPIFB4 protein of macrophages from atherosclerotic patients resulted in a CXCR4-dependent M2 polarization phenotype. Furthermore, LAV-BPIFB4 treatment of arteries explanted from atherosclerotic patients increased the release of atheroprotective IL-33, while inhibiting the release of pro-inflammatory IL-1ß, inducing endothelial nitric oxide synthase phosphorylation and restoring endothelial function. Finally, significantly lower plasma BPIFB4 was detected in patients with pathological carotid stenosis (>25%) and intima media thickness >2 mm. CONCLUSION: Transfer of the LAV of BPIFB4 reduces the atherogenic process and skews macrophages towards an M2-resolving phenotype through modulation of CXCR4, thus opening up novel therapeutic possibilities in cardiovascular disease.
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Aterosclerose , Placa Aterosclerótica , Idoso , Animais , Apolipoproteínas E , Aterosclerose/genética , Espessura Intima-Media Carotídea , Feminino , Humanos , Inflamação , Peptídeos e Proteínas de Sinalização Intercelular , Longevidade , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Camundongos Knockout para ApoE , Pessoa de Meia-Idade , Fosfoproteínas , Receptores CXCR4RESUMO
Diabetes mellitus is a common disease that affects 3â»5% of the general population in Italy. In some countries of northern Europe or in North America, it can even affect 6â»8% of the population. Of great concern is that the number of cases of diabetes is constantly increasing, probably due to the increase in obesity and the sedentary nature of the population. According to the World Health Organization, in the year 2030 there will be 360 million people with diabetes, compared to 170 million in 2000. This has important repercussions on the lives of patients and their families, and on health systems that offer assistance to patients. In this review, we try to describe in an organized way the pathophysiological continuity between diabetes mellitus, endothelial dysfunction, and platelet hyperaggregation, highlighting the main molecular mechanisms involved and the interconnections.
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Diabetes Mellitus/fisiopatologia , Endotélio Vascular/fisiopatologia , Doenças Vasculares/fisiopatologia , Animais , Humanos , Modelos Biológicos , Agregação PlaquetáriaRESUMO
A growing number of evidences report that aging represents the major risk factor for the development of cardio and cerebrovascular diseases. Understanding Aging from a genetic, biochemical and physiological point of view could be helpful to design a better medical approach and to elaborate the best therapeutic strategy to adopt, without neglecting all the risk factors associated with advanced age. Of course, the better way should always be understanding risk-to-benefit ratio, maintenance of independence and reduction of symptoms. Although improvements in treatment of cardiovascular diseases in the elderly population have increased the survival rate, several studies are needed to understand the best management option to improve therapeutic outcomes. The aim of this review is to give a 360° panorama on what goes on in the fragile ecosystem of elderly, why it happens and what we can do, right now, with the tools at our disposal to slow down aging, until new discoveries on aging, cardio and cerebrovascular diseases are at hand.
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Envelhecimento , Doenças Cardiovasculares/epidemiologia , Doenças Cardiovasculares/etiologia , Transtornos Cerebrovasculares/epidemiologia , Transtornos Cerebrovasculares/etiologia , Fatores Etários , Animais , Vasos Sanguíneos/fisiologia , Vasos Sanguíneos/fisiopatologia , Suscetibilidade a Doenças , Endotélio/metabolismo , Endotélio/fisiopatologia , Predisposição Genética para Doença , Coração/fisiologia , Coração/fisiopatologia , Humanos , Estresse Oxidativo , Risco , VasodilataçãoRESUMO
RATIONALE: Long living individuals show delay of aging, which is characterized by the progressive loss of cardiovascular homeostasis, along with reduced endothelial nitric oxide synthase activity, endothelial dysfunction, and impairment of tissue repair after ischemic injury. OBJECTIVE: Exploit genetic analysis of long living individuals to reveal master molecular regulators of physiological aging and new targets for treatment of cardiovascular disease. METHODS AND RESULTS: We show that the polymorphic variant rs2070325 (Ile229Val) in bactericidal/permeability-increasing fold-containing-family-B-member-4 (BPIFB4) associates with exceptional longevity, under a recessive genetic model, in 3 independent populations. Moreover, the expression of BPIFB4 is instrumental to maintenance of cellular and vascular homeostasis through regulation of protein synthesis. BPIFB4 phosphorylation/activation by protein-kinase-R-like endoplasmic reticulum kinase induces its complexing with 14-3-3 and heat shock protein 90, which is facilitated by the longevity-associated variant. In isolated vessels, BPIFB4 is upregulated by mechanical stress, and its knock-down inhibits endothelium-dependent vasorelaxation. In hypertensive rats and old mice, gene transfer of longevity-associated variant-BPIFB4 restores endothelial nitric oxide synthase signaling, rescues endothelial dysfunction, and reduces blood pressure levels. Furthermore, BPIFB4 is implicated in vascular repair. BPIFB4 is abundantly expressed in circulating CD34(+) cells of long living individuals, and its knock-down in endothelial progenitor cells precludes their capacity to migrate toward the chemoattractant SDF-1. In a murine model of peripheral ischemia, systemic gene therapy with longevity-associated variant-BPIFB4 promotes the recruitment of hematopoietic stem cells, reparative vascularization, and reperfusion of the ischemic muscle. CONCLUSIONS: Longevity-associated variant-BPIFB4 may represent a novel therapeutic tool to fight endothelial dysfunction and promote vascular reparative processes.
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Células Progenitoras Endoteliais/metabolismo , Células Endoteliais da Veia Umbilical Humana/metabolismo , Longevidade/genética , Músculo Esquelético/irrigação sanguínea , Neovascularização Fisiológica , Fosfoproteínas/genética , Fosfoproteínas/metabolismo , Proteínas 14-3-3/metabolismo , Fatores Etários , Idoso , Idoso de 80 Anos ou mais , Animais , Pressão Sanguínea , Movimento Celular , Modelos Animais de Doenças , Europa (Continente) , Feminino , Estudos de Associação Genética , Terapia Genética , Genótipo , Células HEK293 , Proteínas de Choque Térmico HSP90/metabolismo , Membro Posterior , Humanos , Hipertensão/genética , Hipertensão/metabolismo , Hipertensão/fisiopatologia , Hipertensão/terapia , Peptídeos e Proteínas de Sinalização Intercelular , Isquemia/genética , Isquemia/metabolismo , Isquemia/fisiopatologia , Isquemia/terapia , Masculino , Camundongos Endogâmicos C57BL , Pessoa de Meia-Idade , Óxido Nítrico Sintase Tipo III/metabolismo , Fenótipo , Fosforilação , Interferência de RNA , Ratos Endogâmicos SHR , Transdução de Sinais , Estresse Mecânico , Transfecção , Estados Unidos , Vasodilatação , eIF-2 Quinase/metabolismoRESUMO
After publication of the original article [1], it was noticed that one of the affiliations of author Carmine Vecchione was missing.
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BACKGROUND: NAFLD is associated to Insulin Resistance (IR). IR is responsible for Endothelial Dysfunction (ED) through the impairment of eNOS function. Although eNOS derangement has been demonstrated in experimental models, no studies have directly shown that eNOS dysfunction is associated with NAFLD in humans. The aim of this study is to investigate eNOS function in NAFLD patients. METHODS: Fifty-four NAFLD patients were consecutively enrolled. All patients underwent clinical and laboratory evaluation and liver biopsy. Patients were divided into two groups by the presence of NAFL or NASH. We measured vascular reactivity induced by patients' platelets on isolated mice aorta rings. Immunoblot assays for platelet-derived phosphorylated-eNOS (p-eNOS) and immunohistochemistry for hepatic p-eNOS have been performed to evaluate eNOS function in platelets and liver specimens. Flow-mediated-dilation (FMD) was also performed. Data were compared with healthy controls. RESULTS: Twenty-one (38, 8%) patients had NAFL and 33 (61, 7%) NASH. No differences were found between groups and controls except for HOMA and insulin (p < 0.0001). Vascular reactivity demonstrated a reduced function induced from NAFLD platelets as compared with controls (p < 0.001), associated with an impaired p-eNOS in both platelets and liver (p < 0.001). NAFL showed a higher impairment of eNOS phosphorylation in comparison to NASH (p < 0.01). In contrast with what observed in vitro, the vascular response by FMD was worse in NASH as compared with NAFL. CONCLUSIONS: Our data showed, for the first time in humans, that NAFLD patients show a marked eNOS dysfunction, which may contribute to a higher CV risk. eNOS dysfunction observed in platelets and liver tissue didn't match with FMD.
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Fígado/irrigação sanguínea , Óxido Nítrico Sintase Tipo III/metabolismo , Hepatopatia Gordurosa não Alcoólica/enzimologia , Hepatopatia Gordurosa não Alcoólica/fisiopatologia , Vasodilatação , Adulto , Animais , Aorta/metabolismo , Biópsia , Plaquetas/metabolismo , Estudos de Casos e Controles , Fígado Gorduroso/patologia , Fígado Gorduroso/fisiopatologia , Feminino , Humanos , Resistência à Insulina , Fígado/patologia , Masculino , Camundongos , Pessoa de Meia-Idade , Hepatopatia Gordurosa não Alcoólica/patologia , Fosforilação , Estudos ProspectivosRESUMO
BACKGROUND: Pentraxin 3 (PTX3), the prototype of long pentraxins, has been described to be associated with endothelial dysfunction in different cardiovascular disorders. No study has yet evaluated the possible direct effect of PTX3 on vascular function. METHODS AND RESULTS: Through in vitro experiments of vascular reactivity and ultrastructural analyses, we demonstrate that PTX3 induces dysfunction and morphological changes in the endothelial layer through a P-selectin/matrix metalloproteinase-1 pathway. The latter hampered the detachment of endothelial nitric oxide synthase from caveolin-1, leading to an impairment of nitric oxide signaling. In vivo studies showed that administering PTX3 to wild-type mice induced endothelial dysfunction and increased blood pressure, an effect absent in P-selectin-deficient mice. In isolated human umbilical vein endothelial cells, PTX3 significantly blunted nitric oxide production through the matrix metalloproteinase-1 pathway. Finally, using ELISA, we found that hypertensive patients (n=31) have higher plasma levels of PTX3 and its mediators P-selectin and matrix metalloproteinase-1 than normotensive subjects (n=21). CONCLUSIONS: Our data show for the first time a direct role of PTX3 on vascular function and blood pressure homeostasis, identifying the molecular mechanisms involved. The findings in humans suggest that PTX3, P-selectin, and matrix metalloproteinase-1 may be novel biomarkers that predict the onset of vascular dysfunction in hypertensive patients.
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Proteína C-Reativa/fisiologia , Endotélio Vascular/fisiopatologia , Hipertensão/fisiopatologia , Metaloproteinase 1 da Matriz/fisiologia , Selectina-P/fisiologia , Componente Amiloide P Sérico/fisiologia , Animais , Pressão Sanguínea , Caveolina 1/metabolismo , Membrana Celular/metabolismo , Células Cultivadas , Células Endoteliais da Veia Umbilical Humana , Humanos , Metaloproteinase 1 da Matriz/deficiência , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Proteínas do Tecido Nervoso/fisiologia , Óxido Nítrico/metabolismo , Receptores de IgG/deficiência , Transdução de Sinais/fisiologia , VasodilataçãoRESUMO
The acute phase protein Pentraxin 3 (PTX3) plays a non-redundant role as a soluble pattern recognition receptor for selected pathogens and it represents a rapid biomarker for primary local activation of innate immunity and inflammation. Recent evidence indicates that PTX3 exerts an important role in modulating the cardiovascular system in humans and experimental models. In particular, there are conflicting points concerning the effects of PTX3 in cardiovascular diseases (CVD) since several observations indicate a cardiovascular protective effect of PTX3 while others speculate that the increased plasma levels of PTX3 in subjects with CVD correlate with disease severity and with poor prognosis in elderly patients. In the present review, we discuss the multifaceted effects of PTX3 on the cardiovascular system focusing on its involvement in atherosclerosis, endothelial function, hypertension, myocardial infarction and angiogenesis. This may help to explain how the specific modulation of PTX3 such as the use of different dosing, time, and target organs could help to contain different vascular diseases. These opposite actions of PTX3 will be emphasized concerning the modulation of cardiovascular system where potential therapeutic implications of PTX3 in humans are discussed.
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Considered as a superfood of the future, Spirulina platensis matrix has been extensively used because of its beneficial effect on the management of cardiovascular diseases. However, its nutraceutical properties, bioactive compounds, and molecular mechanisms are unknown. Here, we demonstrate that S platensis matrix processed in vitro by simulated gastrointestinal digestion induces direct endothelial nitric oxide (NO)-mediated vasorelaxation of resistance vessels in mice. To gain insight into the bioactive compounds responsible for this effect, we used a complex multistep peptidomic approach to fractionate the crude digest: of the 5 peptide fractions identified (A-E), only fraction E evoked vasorelaxation. High-resolution mass spectrometry-based screening revealed in E the presence of 4 main peptides (SP3-SP6 [spirulina peptides]), of which only SP6 (GIVAGDVTPI) exerted direct endothelium-dependent vasodilation of ex vivo vessels, an effect occurring via a PI3K (phosphoinositide-3-kinase)/AKT (serine/threonine kinase Akt) pathway converging on NO release. In vivo, administration of SP6 evoked a significant hemodynamic effect, reducing blood pressure, an action absent in eNOS (endothelial NO synthase)-deficient mice. Of note, although lower doses of SP6 had no hemodynamic effects, it still enhanced endothelial NO vasorelaxation. Finally, in an experimental model of arterial hypertension, SP6 exerted an antihypertensive effect, improving endothelial vasorelaxation associated with enhanced serum nitrite levels. Based on our results, this novel decameric peptide may extend the possible fields of application for spirulina-derived peptides and could be developed into a promising nonpharmacological approach for the containment of pathologies associated with vascular NO misregulation.
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Proteínas de Bactérias/farmacologia , Pressão Sanguínea/efeitos dos fármacos , Óxido Nítrico Sintase Tipo III/fisiologia , Peptídeos/farmacologia , Fosfatidilinositol 3-Quinases/fisiologia , Proteínas Proto-Oncogênicas c-akt/fisiologia , Spirulina/química , Animais , Endotélio Vascular/efeitos dos fármacos , Endotélio Vascular/fisiologia , Camundongos , Óxido Nítrico/fisiologia , Vasodilatação/efeitos dos fármacosRESUMO
Peptides derived from buffalo dairy products possess multiple healthy properties that cannot be exerted as long as they are encrypted in parent proteins. To evaluate the biological activities of encrypted peptide sequences from buffalo ricotta cheese, we performed a simulated gastrointestinal (GI) digestion. Chemical and pharmacological characterization of the digest led to the identification of a novel peptide endowed with antioxidant and antihypertensive action. The GI digest was fractionated by Semiprep-HPLC, and fractions were tested against reactive oxygen species (ROS) release in an H2O2-treated intestinal epithelial cell line. UHPLC-PDA-MS/MS analysis revealed the presence of an abundant ß-lactoglobulin peptide (BRP2) in the most active fraction. Pharmacological characterization of BRP2 highlighted its antioxidant activity, involving ROS reduction, nuclear factor erythroid 2-related factor 2 (Nrf2) activation, and cytoprotective enzyme expression. The bioavailability of BRP2 was evaluated in intestinal transport studies through a Caco-2 cell monolayer. Equal bidirectional transport and linear permeability indicate that BRP2 was absorbed mainly through passive diffusion. In addition to its local effects, the BRP2 administration on mouse mesenteric arteries was able to reduce the angiotensin II-induced vasoconstriction by the Nrf2 nuclear translocation, the reduction of the active form of Ras-related C3 botulinum toxin substrate 1 (Rac1), and the NADPH oxidase activity. These data further highlight the role of buffalo ricotta cheese-derived peptides against oxidative stress-related diseases and suggest their health-promoting potential.
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Angiotensina II/metabolismo , Células Epiteliais/metabolismo , Lactoglobulinas/farmacologia , Artérias Mesentéricas/metabolismo , Fator 2 Relacionado a NF-E2/metabolismo , Estresse Oxidativo/efeitos dos fármacos , Fragmentos de Peptídeos/farmacologia , Animais , Antioxidantes/farmacologia , Células CACO-2 , Células Epiteliais/efeitos dos fármacos , Humanos , Mucosa Intestinal/efeitos dos fármacos , Mucosa Intestinal/metabolismo , Camundongos , Ratos , Espécies Reativas de Oxigênio/metabolismo , Vasoconstrição/efeitos dos fármacosRESUMO
A correction to this article has been published and is linked from the HTML and PDF versions of this paper. The error has been fixed in the paper.
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BACKGROUND: Vascular complications and abnormal platelet function contribute to morbidity and mortality in diabetes mellitus. We hypothesized that the Rho-related GTPase protein, Rac1, can influence both endothelial and platelet function and might represent a potential novel therapeutic target in diabetes mellitus. METHODS AND RESULTS: We used both in vitro and ex vivo approaches to test the effects of pharmacological inhibition of Rac1 during hyperglycemic condition. We evaluated the effect of NSC23766, a pharmacological inhibitor of Rac1, on vascular function in diabetic mice and platelet aggregation in diabetic subjects. We demonstrated that the administration of NSC23766 protects from hyperglycemia-induced endothelial dysfunction, restoring NO levels, and reduces oxidative stress generated by nicotinamide adenine dinucleotide phosphate oxidase. Mechanistically, we identified Rho-associated coiled-coil serine/threonine kinase-1 as a downstream target of Rac1. Moreover, we reported that during hyperglycemic conditions, human platelets showed hyperactivation of Rac1 and impaired NO release, which were both partially restored after NSC23766 treatment. Finally, we characterized the antiplatelet effect of NSC23766 during hyperglycemic conditions, demonstrating the additional role of Rac1 inhibition in reducing platelet aggregation in diabetic patients treated with common antiplatelet drugs. CONCLUSIONS: Our data suggest that the pharmacological inhibition of Rac1 could represent a novel therapeutic strategy to reduce endothelial dysfunction and platelet hyperaggregation in diabetes mellitus.
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Plaquetas/metabolismo , Diabetes Mellitus Experimental/sangue , Endotélio Vascular/fisiopatologia , Agregação Plaquetária/fisiologia , Vasodilatação/fisiologia , Proteínas rac1 de Ligação ao GTP/metabolismo , Aminoquinolinas/farmacologia , Animais , Glicemia/metabolismo , Diabetes Mellitus Experimental/tratamento farmacológico , Diabetes Mellitus Experimental/fisiopatologia , Feminino , Humanos , Masculino , Camundongos , Pessoa de Meia-Idade , Ativação Plaquetária , Agregação Plaquetária/efeitos dos fármacos , Pirimidinas/farmacologia , Proteínas rac1 de Ligação ao GTP/efeitos dos fármacosRESUMO
MitoAKAPs (mitochondrial A kinase anchoring proteins), encoded by the Akap1 gene, regulate multiple cellular processes governing mitochondrial homeostasis and cell viability. Although mitochondrial alterations have been associated to endothelial dysfunction, the role of mitoAKAPs in the vasculature is currently unknown. To test this, postischemic neovascularization, vascular function, and arterial blood pressure were analyzed in Akap1 knockout mice (Akap1-/- ) and their wild-type (wt) littermates. Primary cultures of aortic endothelial cells (ECs) were also obtained from Akap1-/- and wt mice, and ECs migration, proliferation, survival, and capillary-like network formation were analyzed under different experimental conditions. After femoral artery ligation, Akap1-/- mice displayed impaired blood flow and functional recovery, reduced skeletal muscle capillary density, and Akt phosphorylation compared with wt mice. In Akap1-/- ECs, a significant enhancement of hypoxia-induced mitophagy, mitochondrial dysfunction, reactive oxygen species production, and apoptosis were observed. Consistently, capillary-like network formation, migration, proliferation, and AKT phosphorylation were reduced in Akap1-/- ECs. Alterations in Akap1-/- ECs behavior were also confirmed in Akap1-/- mice, which exhibited a selective reduction in acetylcholine-induced vasorelaxation in mesenteric arteries and a mild but significant increase in arterial blood pressure levels compared with wt. Finally, overexpression of a constitutively active Akt mutant restored vascular reactivity and ECs function in Akap1-/- conditions. These results demonstrate the important role of mitoAKAPs in the modulation of multiple ECs functions in vivo and in vitro, suggesting that mitochondria-dependent regulation of ECs might represent a novel therapeutic approach in cardiovascular diseases characterized by endothelial dysfunction.
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Proteínas de Ancoragem à Quinase A/metabolismo , Células Endoteliais/patologia , Mitocôndrias/patologia , Neovascularização Patológica/patologia , Espécies Reativas de Oxigênio/metabolismo , Análise de Variância , Animais , Movimento Celular/fisiologia , Células Cultivadas , Modelos Animais de Doenças , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Mitocôndrias/metabolismo , Óxido Nítrico/metabolismo , Fosforilação , Distribuição Aleatória , Valores de Referência , Fatores de Risco , Estatísticas não Paramétricas , Doenças Vasculares/patologia , Doenças Vasculares/fisiopatologiaRESUMO
The kynurenine pathway of tryptophan metabolism is activated by pro-inflammatory cytokines. L-kynurenine, an upstream metabolite of the pathway, acts as a putative endothelium-derived relaxing factor, and has been hypothesized to play a causative role in the pathophysiology of inflammation-induced hypotension. Here, we show that xanthurenic acid (XA), the transamination product of 3-hydroxykynurenine, is more efficacious than L-kynurenine in causing relaxation of a resistance artery, but fails to relax pre-contracted aortic rings. In the mesenteric artery, XA enhanced activating phosphorylation of endothelial nitric oxide synthase (NOS), and the relaxing action of XA was abrogated by pharmacological inhibition of NOS and endothelial-derived hyperpolarizing factor. Systemic injection of XA reduced blood pressure in mice, and serum levels of XA increased by several fold in response to a pulse with the endotoxin, lipopolysaccharide (LPS). LPS-induced hypotension in mice was prevented by pre-treatment with the kynurenine monooxygenase (KMO) inhibitor, Ro-618048, which lowered serum levels of XA but enhanced serum levels of L-kynurenine. UPF 648, another KMO inhibitor, could also abrogate LPS-induced hypotension. Our data identify XA as a novel vasoactive compound and suggest that formation of XA is a key event in the pathophysiology of inflammation-induced hypotension.
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AIMS: Ageing is associated with impairment of endothelial nitric oxide synthase (eNOS) and progressive reduction in endothelial function. A genetic study on long-living individuals-who are characterized by delays in ageing and in the onset of cardiovascular disease-previously revealed I229V (rs2070325) in bactericidal/permeability-increasing fold-containing-family-B-member-4 (BPIFB4) as a longevity-associated variant (LAV); the LAV protein enhanced endothelial NO production and vasorelaxation through a protein kinase R-like endoplasmic reticulum kinase/14-3-3/heat shock protein 90 signal. Here, we further characterize the molecular mechanisms underlying LAV-BPIFB4-dependent enhancement of vascular function. METHODS AND RESULTS: LAV-BPIFB4 upregulated eNOS function via mobilization of Ca2+ and activation of protein kinase C alpha (PKCα). Indeed, the overexpression of LAV-BPIFB4 in human endothelial cells enhanced ATP-induced Ca2+ mobilization and the translocation of PKCα to the plasma membrane. Coherently, pharmacological inhibition of PKCα blunted the positive effect of LAV-BPIFB4 on eNOS and endothelial function. In addition, although LAV-BPIFB4 lost the ability to activate PKCα and eNOS in ex vivo vessels studied in an external Ca2+-free medium and in vessels from eNOS-/- mice, it still potentiated endothelial activity, recruiting an alternative mechanism dependent upon endothelium-derived hyperpolarizing factor (EDHF). CONCLUSIONS: We have identified novel molecular determinants of the beneficial effects of LAV-BPIFB4 on endothelial function, showing the roles of Ca2+ mobilization and PKCα in eNOS activation and of EDHF when eNOS is inhibited. These results highlight the role LAV-BPIFB4 can have in restoring signals that are lost during ageing.
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Sinalização do Cálcio , Células Endoteliais/enzimologia , Artérias Mesentéricas/enzimologia , Óxido Nítrico Sintase Tipo III/metabolismo , Fosfoproteínas/metabolismo , Proteína Quinase C-alfa/metabolismo , Vasodilatação , Animais , Sinalização do Cálcio/efeitos dos fármacos , Células Cultivadas , Relação Dose-Resposta a Droga , Células Endoteliais/efeitos dos fármacos , Ativação Enzimática , Humanos , Peptídeos e Proteínas de Sinalização Intercelular , Potenciais da Membrana , Artérias Mesentéricas/efeitos dos fármacos , Camundongos Knockout , Óxido Nítrico Sintase Tipo III/deficiência , Óxido Nítrico Sintase Tipo III/genética , Fosfoproteínas/genética , Fosforilação , Isoformas de Proteínas , Proteína Quinase C-alfa/antagonistas & inibidores , Proteína Quinase C-alfa/genética , Inibidores de Proteínas Quinases/farmacologia , Transporte Proteico , Transfecção , Regulação para Cima , Vasodilatação/efeitos dos fármacos , Vasodilatadores/farmacologiaRESUMO
BPIFB4 is associated with exceptional longevity: four single-nucleotide polymorphisms distinguish the wild-type form from a longevity-associated variant conferring positive effects on blood pressure. The effect of a rare variant (RV; allele frequency, 4%) on blood pressure is unknown. Here, we show that overexpression of RV-BPIFB4 in ex-vivo mouse vessels impairs phosphorylation of endothelial nitric oxide synthase (eNOS), blunting acetylcholine-evoked vasorelaxation; in vivo, virally mediated overexpression of RV-BPIFB4 increases blood pressure, an action absent in eNOS-deficient mice. In humans, we found RV carriers to have increased diastolic blood pressure, a finding that was more marked in subjects on anti-hypertensive medication; moreover, recombinant RV-BPIFB4 protein impaired eNOS function in ex-vivo human vessels. Thus, RV-BPIFB4 acts directly on blood pressure homeostasis and may represent a novel biomarker of vascular dysfunction and hypertension.
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Pressão Sanguínea/genética , Predisposição Genética para Doença , Variação Genética , Hipertensão/genética , Hipertensão/metabolismo , Óxido Nítrico/metabolismo , Fosfoproteínas/genética , Transdução de Sinais , Idoso , Alelos , Animais , Biomarcadores , Vasos Sanguíneos/efeitos dos fármacos , Feminino , Frequência do Gene , Estudos de Associação Genética , Haplótipos , Humanos , Hipertensão/fisiopatologia , Peptídeos e Proteínas de Sinalização Intercelular , Masculino , Camundongos , Pessoa de Meia-Idade , Fosfoproteínas/farmacologia , Polimorfismo de Nucleotídeo Único , Proteínas Recombinantes/farmacologiaRESUMO
BACKGROUND: Endothelial dysfunction contributes significantly to the development of vascular diseases. However, a therapy able to reduce this derangement still needs to be identified. We evaluated the effects of pharmacological inhibition of Rac1, a small GTPase protein promoting oxidative stress, in human endothelial dysfunction. METHODS AND RESULTS: We performed vascular reactivity studies to test the effects of NSC23766, a Rac1 inhibitor, on endothelium-dependent vasorelaxation of saphenous vein segments collected from 85 subjects who had undergone surgery for venous insufficiency and from 11 patients who had undergone peripheral vascular surgery. The endothelium-dependent vasorelaxation of the varicose segments of saphenous veins collected from patients with venous insufficiency was markedly impaired and was also significantly lower than that observed in control nonvaricose vein tracts from the same veins. Rac1 activity, reactive oxygen species levels, and reduced nicotine adenine dinucleotide phosphate (NADPH) oxidase activity were significantly increased in varicose veins, and NSC23766 was able to significantly improve endothelium-dependent vasorelaxation of dysfunctional saphenous vein portions in a nitric oxide-dependent manner. These effects were paralleled by a significant reduction of NADPH oxidase activity and activation of endothelial nitric oxide synthase. Finally, we further corroborated this data by demonstrating that Rac1 inhibition significantly improves venous endothelial function and reduces NADPH oxidase activity in saphenous vein grafts harvested from patients with vascular diseases undergoing peripheral bypass surgery. CONCLUSIONS: Rac1 pharmacological inhibition rescues endothelial function and reduces oxidative stress in dysfunctional veins. Rac1 inhibition may represent a potential therapeutic intervention to reduce human endothelial dysfunction and subsequently vascular diseases in various clinical settings.
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Aminoquinolinas/farmacologia , Endotélio Vascular/fisiopatologia , Pirimidinas/farmacologia , Veia Safena/fisiopatologia , Vasodilatação/efeitos dos fármacos , Insuficiência Venosa/fisiopatologia , Proteínas rac1 de Ligação ao GTP/biossíntese , Adolescente , Adulto , Idoso , Idoso de 80 Anos ou mais , Doença Crônica , Endotélio Vascular/efeitos dos fármacos , Endotélio Vascular/metabolismo , Feminino , Humanos , Immunoblotting , Masculino , Pessoa de Meia-Idade , NADP/metabolismo , Óxido Nítrico Sintase Tipo III/metabolismo , Estresse Oxidativo/efeitos dos fármacos , Espécies Reativas de Oxigênio/metabolismo , Veia Safena/efeitos dos fármacos , Veia Safena/metabolismo , Insuficiência Venosa/metabolismo , Adulto Jovem , Proteínas rac1 de Ligação ao GTP/antagonistas & inibidoresRESUMO
Bcl2-associated athanogene 3 (BAG3), is constitutively expressed in a few normal cell types, including myocytes, peripheral nerves and in the brain, and is also expressed in certain tumors. To date, the main studies about the role of BAG3 are focused on its pro-survival effect in tumors through various mechanisms that vary according to cellular type. Recently, elevated concentrations of a soluble form of BAG3 were described in patients affected by advanced stage of heart failure (HF), identifying BAG3 as a potentially useful biomarker in monitoring HF progression. Despite the finding of high levels of BAG3 in the sera of HF patients, there are no data on its possible role on the modulation of vascular tone and blood pressure levels. The aim of this study was to investigate the possible hemodynamic effects of BAG3 performing both in vitro and in vivo experiments. Through vascular reactivity studies, we demonstrate that BAG3 is capable of evoking dose-dependent vasorelaxation. Of note, BAG3 exerts its vasorelaxant effect on resistance vessels, typically involved in the blood pressure regulation. Our data further show that the molecular mechanism through which BAG3 exerts this effect is the activation of the PI3K/Akt signalling pathway leading to nitric oxide release by endothelial cells. Finally, we show that in vivo BAG3 administration is capable of regulating blood pressure and that this is dependent on eNOS regulation since this ability is lost in eNOS KO animals.
Assuntos
Proteínas Adaptadoras de Transdução de Sinal/metabolismo , Proteínas Reguladoras de Apoptose/metabolismo , Vasos Sanguíneos/fisiologia , Homeostase , Animais , Pressão Sanguínea/efeitos dos fármacos , Vasos Sanguíneos/efeitos dos fármacos , Sobrevivência Celular/efeitos dos fármacos , Endotélio Vascular/efeitos dos fármacos , Homeostase/efeitos dos fármacos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Contração Muscular/efeitos dos fármacos , Óxido Nítrico Sintase Tipo III/metabolismo , Fenilefrina/farmacologia , Fosfatidilinositol 3-Quinases/metabolismo , Proteínas Proto-Oncogênicas c-akt/metabolismo , Transdução de Sinais/efeitos dos fármacos , Sístole/efeitos dos fármacos , Vasodilatação/efeitos dos fármacosRESUMO
SCOPE: Morus alba is a promising phytomedicine cultivated in oriental countries that is extensively used to prevent and treat various cardiovascular problems. To date, despite its beneficial effects, the molecular mechanisms involved remain unclear. Thus, we investigate the vascular and haemodynamic effects of Morus alba extract in an experimental model focusing our attention on the molecular mechanisms involved. METHODS AND RESULTS: Through vascular reactivity studies, we demonstrate that Morus alba extract evokes endothelial vasorelaxation through a nitric oxide-dependent pathway. Our molecular analysis highlights an increase in endothelial nitric oxide synthase (eNOS) phosphorylation. In vivo administration of Morus alba extract reduces blood pressure levels exclusively in wild-type mice, whereas it fails to evoke any haemodynamic effects in eNOS-deficient mice. Molecular analyses revealed that its beneficial action on vasculature is mediated by the activation of two important proteins that act as stress sensors and chaperones: PERK and heat shock protein 90. Finally, Morus alba extract exerts antihypertensive action in an experimental model of arterial hypertension. CONCLUSION: Through its action on eNOS signaling, Morus alba extract could act as a food supplement for the regulation of cardiovascular system, mainly in clinical conditions characterized by eNOS dysfunction, such as arterial hypertension.
Assuntos
Pressão Sanguínea/efeitos dos fármacos , Morus/química , Óxido Nítrico Sintase Tipo III/metabolismo , Extratos Vegetais/farmacologia , Administração Oral , Animais , Cromatografia Líquida de Alta Pressão , Relação Dose-Resposta a Droga , Proteínas de Choque Térmico HSP90/metabolismo , Artérias Mesentéricas/efeitos dos fármacos , Artérias Mesentéricas/fisiologia , Camundongos Endogâmicos C57BL , Camundongos Knockout , Óxido Nítrico/metabolismo , Óxido Nítrico Sintase Tipo III/genética , Fosforilação/efeitos dos fármacos , Extratos Vegetais/administração & dosagem , Extratos Vegetais/química , Ratos Endogâmicos SHR , Transdução de Sinais/efeitos dos fármacos , Vasodilatação/efeitos dos fármacos , eIF-2 Quinase/metabolismoRESUMO
Within the family of endogenous gasotransmitters, nitric oxide (NO) is the smallest gaseous intercellular messenger involved in the modulation of several processes, such as blood flow and platelet aggregation control, essential to maintain vascular homeostasis. NO is produced by nitric oxide synthases (NOS) and its effects are mediated by cGMP-dependent or cGMP-independent mechanisms. Growing evidence suggests a crosstalk between the NO signaling and the occurrence of oxidative stress in the onset and progression of vascular diseases, such as hypertension, heart failure, ischemia, and stroke. For these reasons, NO is considered as an emerging molecular target for developing therapeutic strategies for cardio- and cerebrovascular pathologies. Several natural derived compounds, such as polyphenols, are now proposed as modulators of NO-mediated pathways. The aim of this review is to highlight the experimental evidence on the involvement of nitric oxide in vascular homeostasis focusing on the therapeutic potential of targeting NO with some natural compounds in patients with vascular diseases.