RESUMO
Endothelial cells injury and pro-inflammation cytokines release are the initial steps of hyperhomocysteinemia (HHcy)-associated vascular inflammation. Pyroptosis is a newly identified pro-inflammation form of programmed cell death, causing cell lysis and IL-1ß release, and characterized by the caspases-induced cleavage of its effector molecule gasdermins (GSDMs). However, the effect of homocysteine (Hcy) on endothelial cells pyroptosis and the underlying mechanisms have not been fully defined. We have previously reported that Hcy induces vascular endothelial inflammation accompanied by the increase of high mobility group box-1 protein (HMGB1) and lysosomal cysteine protease cathepsin V in endothelial cells, and other studies have shown that HMGB1 or cathepsins are involved in activation of NLRP3 inflammasome and caspase-1. Here, we investigated the role of HMGB1 and cathepsin V in the process of Hcy-induced pyroptosis. We observed an increase in plasma IL-1ß levels in HHcy patients and mice models, cathepsin V inhibitor reduced the plasma IL-1ß levels and cleavage of GSDMD full-length into GSDMD N-terminal in the thoracic aorta of hyperhomocysteinemia mice. Using cultured HUVECs, we observed that Hcy promoted GSDMD N-terminal expression, silencing GSDMD or HMGB1 rescued Hcy-induced pyroptosis. HMGB1 also increased GSDMD N-terminal expression, and silencing cathepsin V reversed HMGB1-induced pyroptosis. HMGB1 could increase lysosome permeability, and silencing cathepsin V attenuated HMGB1-induced activation of caspase-1. In conclusion, this study has delineated a novel mechanism that HMGB1 mediated Hcy-induced endothelial cells pyroptosis partly via cathepsin V-dependent pathway.
Assuntos
Catepsinas/fisiologia , Cisteína Endopeptidases/fisiologia , Endotélio Vascular/citologia , Proteína HMGB1/fisiologia , Homocisteína/fisiologia , Piroptose , Idoso , Animais , Caspase 1/metabolismo , Linhagem Celular , Feminino , Células Endoteliais da Veia Umbilical Humana/citologia , Células Endoteliais da Veia Umbilical Humana/metabolismo , Humanos , Hiper-Homocisteinemia/sangue , Hiper-Homocisteinemia/metabolismo , Interleucina-1beta/sangue , Peptídeos e Proteínas de Sinalização Intracelular/metabolismo , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Pessoa de Meia-Idade , Proteínas de Ligação a Fosfato/metabolismo , Artérias Torácicas/metabolismoRESUMO
Despite commonly used for coronary artery bypass surgery, saphenous vein (SV) grafts have significantly lower patency rates in comparison to internal thoracic artery (ITA) grafts, which might be due to the structural characteristics of the vessel wall but also due to differences in oxidative stress adaptation and molecular signaling and regulation. This human post mortem study included a total of 150 human bypass grafts (75 SV grafts and 75 ITA grafts) obtained from 60 patients divided into five groups due to the time period of implantation: group 1: baseline group without grafting; group 2: 1 day; group 3: > 1 day-1 week; group 4: > 1 week-1 month; group 5: > 1 month-1 year. Pieces of 3 mm length were fixed with formaldehyde, dehydrated, wax embedded, cut into sections of 3 µm thickness, and histologically and immunohistochemically examined. Over the whole time period, we observed a lower neointima formation and a better preserved media in ITA grafts with a higher percentage of TNF-α, PDGFR-α, and VEGF-A in nearly all vessel wall layers, a higher amount of MMP-7, MMP-9, EGFR, and bFGF positive cells in SV grafts and a timely different peak not only between ITA and SV grafts but also within the various vessel wall layers of both graft types. Since most of the examined growth factors, growth factor receptors and cytokines are regulated by MAPKs, our results suggest an activation of different pathways in both vessel graft types immediately after bypass grafting.
Assuntos
Ponte de Artéria Coronária , Citocinas/análise , Peptídeos e Proteínas de Sinalização Intercelular/análise , Receptores de Fatores de Crescimento/análise , Veia Safena/metabolismo , Artérias Torácicas/metabolismo , Citocinas/metabolismo , Feminino , Humanos , Peptídeos e Proteínas de Sinalização Intercelular/metabolismo , Masculino , Receptores de Fatores de Crescimento/metabolismo , Veia Safena/cirurgia , Artérias Torácicas/cirurgia , Fatores de TempoRESUMO
AIMS: Chemerin has been recently identified as a vasoactive adipokine implicated in blood pressure regulation. In this context, we evaluated whether chemerin could influence pulmonary vasoreactive response. MATERIALS AND METHODS: Vascular reactivity to chemerin and to phenylephrine, serotonin and endothelin-1 after chemerin pretreatment was evaluated in rat isolated pulmonary artery versus thoracic aorta with and without endothelium. Vasoreactivity to acetylcholine in presence of nitric oxide (NO)-synthase inhibitor (L-NAME) and to NO donor sodium nitroprusside (SNP) was evaluated in chemerin-pretreated pulmonary artery versus thoracic aorta with endothelium. Pretreatment with ODQ, a soluble guanylate cyclase inhibitor and apocynin, a ROS production inhibitor, were also tested. Arteries and lung tissue were harvested for pathobiological evaluation. KEY FINDINGS: Chemerin contracted endothelium-denuded pulmonary artery, while no response was observed in arteries with endothelium. Chemerin potentiated phenylephrine-, endothelin-1- and serotonin-induced vasoconstriction, which was further enhanced by endothelium removal. Chemerin decreased acetylcholine-induced vasorelaxation in arteries with endothelium, while it did not affect SNP-induced relaxation. In presence of L-NAME, there remained a vasorelaxation in chemerin-pretreated arteries. Chemerin or ODQ alone partly decreased acetylcholine-induced vasorelaxation in pulmonary artery and thoracic aorta, while combined chemerin and ODQ incubation abolished it. Treatment with apocynin partly or totally reversed chemerin effects. In both types of arteries, chemerin reduced acetylcholine-induced NO production, as well as endothelial and inducible NO-synthase expression. SIGNIFICANCE: Chemerin potentiates vascular responses to vasoconstrictors in pulmonary artery and thoracic aorta and, impairs acetylcholine-induced pulmonary artery vasodilatation, by mechanisms involving at least partly NO signaling and oxidative stress.
Assuntos
Quimiocinas/fisiologia , Peptídeos e Proteínas de Sinalização Intercelular/fisiologia , Artéria Pulmonar/efeitos dos fármacos , Vasoconstrição/efeitos dos fármacos , Acetilcolina/farmacologia , Adipocinas/metabolismo , Animais , Quimiocinas/metabolismo , Endotelina-1/metabolismo , Endotelinas/efeitos dos fármacos , Endotélio Vascular/efeitos dos fármacos , Peptídeos e Proteínas de Sinalização Intercelular/metabolismo , Masculino , NG-Nitroarginina Metil Éster/farmacologia , Óxido Nítrico/metabolismo , Nitroprussiato/farmacologia , Estresse Oxidativo/efeitos dos fármacos , Fenilefrina/farmacologia , Artéria Pulmonar/metabolismo , Ratos , Ratos Wistar , Serotonina/metabolismo , Artérias Torácicas/efeitos dos fármacos , Artérias Torácicas/metabolismo , Vasoconstritores/farmacologia , Vasodilatação/efeitos dos fármacosRESUMO
BACKGROUND: An imbalance between the activity of matrix metalloproteinases (MMPs), particularly gelatinases, and tissue inhibitors of metalloproteinases (TIMPs) is considered as one of the mechanisms leading to aortocoronary graft failure. AIM: We aimed to assess the variability in gelatinase expression in the walls of aortocoronary conduits and to evaluate its impact on coronary artery bypass grafting (CABG) outcomes. METHODS: The study included 101 consecutive patients (61 men and 40 women) who underwent CABG. An immunohisto-chemical analysis of MMP-2, MMP-9, TIMP-1, and TIMP-2 expression was performed on the cross-sections of the internal thoracic artery (ITA), radial artery (RA), and saphenous vein (SV). The histological findings were compared between patients with SV graft disease (SVGD[+] group) and those without occlusions in the SV (SVGD[-] group). RESULTS: The median MMP and TIMP expression was the weakest in the ITA wall. MMP expression was comparable between the RA and SV cross-sections, whereas TIMP expression was stronger in the RA than in the SV wall (p < 0.05). In most SV segments, but not in the arteries, immunostaining intensity for MMP was comparable to or stronger than for TIMPs. In the veins harvested from the SVGD(+) group, MMP-2 and MMP-9 tissue expression was more pronounced than in the SVGD(-) group. TIMP levels were comparable between groups. CONCLUSIONS: Imbalance in the metalloproteinase-to-inhibitor tissue expression in the vessel wall might predispose to graft failure. A stronger expression of TIMPs than MMPs in the arterial grafts might explain favourable long-term outcomes.
Assuntos
Vasos Sanguíneos/enzimologia , Ponte de Artéria Coronária , Doença das Coronárias/enzimologia , Gelatinases/genética , Inibidores Teciduais de Metaloproteinases/genética , Adulto , Idoso , Idoso de 80 Anos ou mais , Vasos Sanguíneos/metabolismo , Doença das Coronárias/genética , Doença das Coronárias/metabolismo , Doença das Coronárias/cirurgia , Feminino , Regulação da Expressão Gênica , Humanos , Masculino , Pessoa de Meia-Idade , Artéria Radial/enzimologia , Artéria Radial/metabolismo , Veia Safena/enzimologia , Veia Safena/metabolismo , Artérias Torácicas/enzimologia , Artérias Torácicas/metabolismo , Inibidor Tecidual de Metaloproteinase-1/genética , Inibidor Tecidual de Metaloproteinase-2/genética , Resultado do TratamentoRESUMO
We investigated the potential effects of monosodium glutamate (MSG)-induced obesity with regards to nitric oxide and prostanoid production, as well as potassium channel function, in rat thoracic arteries. Newborn male Wistar rats were injected intraperitoneally with typically reported MSG (4.0â¯mg/g) once daily for 4 consecutive days. At 90â¯days postnatal, the rats were sacrificed and the thoracic aortas were evaluated for vascular responses and for prostanoid production. Nitric oxide was studied with calcium ionophore (A23187), acetylcholine (ACh) and sodium nitroprusside (SNP). The release of prostanoids was measured under basal and ACh-stimulated conditions, and the vasomotor effect of exogenous thromboxane A2 mimetic, U46619 was assessed. Potassium channel activities were analyzed using an NS1619 opener for BKCa channels and pinacidil for KATP channels. Arteries from MSG-obese rats exhibited a reduced maximal contraction to potassium chloride and hyper-responsiveness to U46619, suggesting that MSG also alters the responsiveness of vascular smooth muscles. The endothelium-dependent relaxation to ACh and A23817 was attenuated, suggesting low nitric oxide bioavailability. The hypersensitivity of arteries to an exogenous nitric oxide donor, SNP, occurred. The secondary contraction to A23817 was augmented, suggesting increased activation of the prostanoid receptor. The prostanoid release was increased in both basal- and acetylcholine-stimulated rings. In addition, down-regulation of KATP and BKCa channels influenced hyperpolarizing mechanisms. Our findings suggest that increased prostanoid production and hypersensitivity to thromboxane A2 together with down-regulation of potassium channels and low nitric oxide bioavailability may contribute to the increase in blood pressure found in adult MSG-obese male rats.
Assuntos
Aditivos Alimentares/toxicidade , Óxido Nítrico/metabolismo , Obesidade/induzido quimicamente , Obesidade/patologia , Canais de Potássio/efeitos dos fármacos , Prostaglandinas/metabolismo , Glutamato de Sódio/toxicidade , Artérias Torácicas/efeitos dos fármacos , Artérias Torácicas/patologia , Ácido 15-Hidroxi-11 alfa,9 alfa-(epoximetano)prosta-5,13-dienoico/farmacologia , Acetilcolina/farmacologia , Animais , Aorta Torácica/patologia , Regulação para Baixo/efeitos dos fármacos , Injeções Intraperitoneais , Masculino , Contração Muscular/efeitos dos fármacos , Músculo Liso Vascular/efeitos dos fármacos , Ratos , Ratos Wistar , Artérias Torácicas/metabolismo , Tromboxano A2/farmacologia , Vasoconstritores/farmacologiaRESUMO
BACKGROUND: Vascular defects in the mechanical properties of aorta and muscular arteries have been previously reported in animals with copper-deficient feed. However, the interaction between copper nanoparticles (CuNPs) and mechanical properties of arteries has not been reported. Hence, the present study was aimed to evaluate the effect of copper nanoparticles on the vasoreactivity of rat isolated thoracic arteries. METHODS: In this study, 5 week old male Wistar rats were fed a copper-adequate diet (CuA, 6.5mg copper/kg diet), copper-deficient diet (CuD) and copper-modified diets, enriched with copper as a salt (CuS) and as copper nanoparticles (CuNPs) of 40-60nm in diameter. RESULTS: There was a strong relationship between CuNPs and CuS administration in the tensile strength of the thoracic aorta subjected to phenylephrine treatment in the concentration range of 10-7-10-5M. This was also seen between CuNPs and the control diet in the same concentration ranges. In addition vasodilation induced by acetylcholine at the concentration range of 10-7-10-5M was significantly reduced in CuD and NPs feed animals. In CuNPs fed rats, activities of Cu,Zn-SOD, CAT and copper concentration in cardiomyocytes were not influenced when compared with CuS control. In contrast, in CuS-low diet the activities of studied enzymes and copper concentration were pointing towards copper deficiency. CONCLUSIONS: Our results demonstrate for the first time that the observed effects of copper administration in the form of NPs are attributed mainly to the NPs rather than copper itself. Thus another mechanism not related with Cu,Zn-SOD and CAT seems to be involved.
Assuntos
Cobre/farmacologia , Nanopartículas Metálicas , Superóxido Dismutase/metabolismo , Artérias Torácicas/efeitos dos fármacos , Acetilcolina/farmacologia , Animais , Cobre/administração & dosagem , Cobre/química , Dieta , Masculino , Miócitos Cardíacos/efeitos dos fármacos , Miócitos Cardíacos/metabolismo , Tamanho da Partícula , Fenilefrina/farmacologia , Ratos , Ratos Wistar , Sais , Artérias Torácicas/metabolismo , Vasodilatação/efeitos dos fármacosRESUMO
INTRODUCTION: The role of Pulmonary and Activation-Regulated Chemokine (PARC) in the physiopathology of Chronic Obstructive Pulmonary Disease (COPD) is not fully understood. The aim of the present study is to analyze the expression of PARC in lung tissue and its relationship with the vascular remodeling of the systemic and pulmonary arteries of COPD subjects. METHODS: To achieve this objective, protein and gene expression experiments, together with ELISA assays, were performed on the lung tissue, intercostal arteries and serum samples from COPD patients, non-obstructed smokers (NOS) and never-smokers (NS). RESULTS: A total of 57 subjects were included in the analysis (23 COPD, 18 NOS and 16 NS). In the comparisons between groups, a significantly increased lung protein expression of PARC was observed in the COPD group compared to the NOS group (1.96±0.22 vs. 1.29±0.27, P-adjusted = 0.038). PARC was located predominantly in the smooth muscle cells of the remodeled pulmonary muscular arteries and the macrophage-rich area of the alveolar parenchyma. No differences were detected in PARC gene expression analyses. The protein content of PARC in the intercostal arteries were similar between groups, though little remodeling was observed in these arteries. Circulating levels of PARC were numerically higher in patients with COPD compared to NOS and NS. CONCLUSION: The results of the present study suggest an increased lung protein expression of PARC in COPD subjects. This protein was mainly localized in the smooth muscle cells of the pulmonary muscular arteries and was associated with the severity of intimal thickening, indicating its possible role in this remodeling process.
Assuntos
Quimiocinas CC/metabolismo , Artéria Pulmonar/metabolismo , Doença Pulmonar Obstrutiva Crônica/metabolismo , Idoso , Quimiocinas CC/genética , Ensaio de Imunoadsorção Enzimática , Feminino , Expressão Gênica , Humanos , Pulmão/metabolismo , Pulmão/patologia , Masculino , Pessoa de Meia-Idade , Artéria Pulmonar/patologia , Doença Pulmonar Obstrutiva Crônica/patologia , Fumar/metabolismo , Artérias Torácicas/metabolismo , Artérias Torácicas/fisiopatologia , Remodelação VascularRESUMO
UNLABELLED: Essentials Patients with unprovoked pulmonary embolism (PE) are at increased risk of arterial thromboembolism. Coronary and thoracic aorta calcium were evaluated in patients with and without (unprovoked) PE. No association was found between (unprovoked) PE and coronary or aortic calcification. Assessment of both calcium scores on computed tomography pulmonary angiography was highly reproducible. SUMMARY: Objective To evaluate the potential association between (unprovoked) pulmonary embolism (PE) and the presence and extent of coronary artery calcium (CAC) and thoracic aorta calcium (TAC). Methods CAC and TAC derived from computed tomography pulmonary angiography of 100 patients with PE were compared to that of 100 patients in whom PE was ruled out. Results Intraobserver and interobserver agreements for both TAC and CAC were excellent (intraclass correlation > 0.95 for both). In patients with PE vs. patients without PE, no significant differences were found in the presence of CAC or TAC (CAC 64% vs. 67%, odds ratio [OR] 1.0, 95% confidence interval [CI] 0.67-1.6; TAC 46% vs. 59%, OR 1.2, 95% CI 0.80-2.1). Mean CAC and TAC scores were significantly lower in patients with PE than in patients without PE (CAC 3.4 vs. 4.9, absolute difference 1.5, 95% CI 0.2-2.8; TAC 1.1 vs. 1.8, absolute difference 0.9, 95% CI 0.2-1.2). No significant differences were found in the presence of CAC or TAC or in mean CAC and TAC scores between patients with unprovoked PE vs. patients with provoked PE, or between patients with unprovoked PE vs. no PE. Conclusion No significant differences were found between the extent of CAC and TAC in patients with unprovoked PE compared to those with provoked PE or without PE. The observed difference in the extend of CAC and TAC between patients with and without PE was dependent on prevalent cardiovascular risk factors.
Assuntos
Aorta Torácica/fisiopatologia , Cálcio/metabolismo , Vasos Coronários/fisiopatologia , Embolia Pulmonar/metabolismo , Artérias Torácicas/fisiopatologia , Adulto , Idoso , Aorta/fisiopatologia , Aorta Torácica/metabolismo , Calcinose , Estudos de Casos e Controles , Vasos Coronários/metabolismo , Feminino , Humanos , Masculino , Pessoa de Meia-Idade , Variações Dependentes do Observador , Estudos Prospectivos , Fatores de Risco , Artérias Torácicas/metabolismo , Tomografia Computadorizada por Raios X , Resultado do TratamentoRESUMO
BACKGROUND: The goal of this study was to investigate the modulation of the contraction-relaxation effects in isolated human thoracic artery samples of three calcium-channel antagonists, amlodipine (CAS [88150-42-9]), cerebrocrast (CAS [118790-71-9]) and diltiazem (CAS [42399-41-7]), and two calcium-channel agonists, CGP 28392 (CAS [89289-93-0]) and benzimidazole derivative. To estimate the endothelial function of the artery samples, carbachol, an agonist of muscarinic receptors, was used. METHODS: The experiments were conducted on isolated human thoracic artery samples, and their isometric contractions were recorded using an i-FOT10 force transducer. Cumulative concentration-contraction curves of the tested agents (10(-7) to 10(-4) M) were established. RESULTS: Carbachol at concentrations of 10(-7) to 10(-6) M did not cause any relaxation of artery rings precontracted by 10(-4) M phenylephrine, and at concentrations of 10(-5) and 10(-4) M, isometric contractions increased by 7% and 20%, respectively. In response to amlodipine, cerebrocrast and CGP28392, the contraction of artery samples increased significantly, whereas diltiazem and benzimidazole derivative caused their relaxation by ≈55%. CONCLUSION: The obtained data indicate that the endothelium of the thoracic artery, which is used for coronary artery bypass grafting, is damaged by and may have some influence on the inadequate response of 1,4-dihydropyrine type calcium-channel antagonists.
Assuntos
Agonistas dos Canais de Cálcio/farmacologia , Bloqueadores dos Canais de Cálcio/farmacologia , Endotélio Vascular/efeitos dos fármacos , Artérias Torácicas/efeitos dos fármacos , Adulto , Idoso , Idoso de 80 Anos ou mais , Anlodipino/administração & dosagem , Anlodipino/farmacologia , Benzimidazóis/administração & dosagem , Benzimidazóis/farmacologia , Agonistas dos Canais de Cálcio/administração & dosagem , Bloqueadores dos Canais de Cálcio/administração & dosagem , Carbacol/administração & dosagem , Carbacol/farmacologia , Ponte de Artéria Coronária/métodos , Di-Hidropiridinas/administração & dosagem , Di-Hidropiridinas/farmacologia , Diltiazem/administração & dosagem , Diltiazem/farmacologia , Relação Dose-Resposta a Droga , Endotélio Vascular/metabolismo , Feminino , Humanos , Técnicas In Vitro , Masculino , Pessoa de Meia-Idade , Contração Muscular/efeitos dos fármacos , Relaxamento Muscular/efeitos dos fármacos , Piridinas/administração & dosagem , Piridinas/farmacologia , Artérias Torácicas/metabolismoRESUMO
Arterial stiffness is a key marker in metabolic diseases and can be evaluated by arterial compliance. Most compliance measurements are performed in large conduit arteries in advanced stage of metabolic diseases, which may not provide information on mechanisms associated with the initiation of the pathology. For this reason, we compared arterial compliance of two different size arteries: carotid and a smaller artery (thoracodorsal artery, TDA). The arterial compliance was compared between control and high fat-fed mice for 6 weeks. We show that the compliance of the TDAs was dramatically reduced in high fat-fed mice whereas the compliance of the carotids remained unchanged. An abundance of collagen deposition in the media/adventitia of the carotids and TDAs was observed in high fat-fed mice. These results demonstrate that the structural and mechanical properties of small arteries are rapidly altered even after only 6 weeks of high fat feeding.
Assuntos
Artérias Carótidas/patologia , Dieta Hiperlipídica , Obesidade/patologia , Artérias Torácicas/patologia , Doenças Vasculares/patologia , Animais , Artérias Carótidas/metabolismo , Artérias Carótidas/fisiopatologia , Colágeno/metabolismo , Complacência (Medida de Distensibilidade) , Modelos Animais de Doenças , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Obesidade/etiologia , Obesidade/metabolismo , Obesidade/fisiopatologia , Artérias Torácicas/metabolismo , Artérias Torácicas/fisiopatologia , Fatores de Tempo , Doenças Vasculares/etiologia , Doenças Vasculares/metabolismo , Doenças Vasculares/fisiopatologiaRESUMO
OBJECTIVES: The internal thoracic artery (ITA) is the preferred conduit for coronary artery bypass graft (CABG) surgery. The authors investigated whether thoracic epidural anesthesia (TEA) as an adjunct to general anesthesia (GA) can increase the blood flow of the ITA. DESIGN: A prospective randomized study. SETTING: A university hospital. PARTICIPANTS: Patients with ischemic heart disease. INTERVENTIONS: Thirty patients scheduled for elective CABG surgery were randomized to receive either GA (n = 15) or GA + TEA (n = 15) after receiving institutional review board approval. Demographics showed similarity between the groups. The epidural catheter was inserted in the thoracic region between T1 and T5 levels. In the GA + TEA group, the patients received a 20-mg bolus of 0.25% bupivacaine through epidural catheters 1 hour before surgery, and this was followed by the infusion (20 mg/h) of 0.25% bupivacaine. In all patients, ITA free blood flow was measured before cardiopulmonary bypass and without the administration of any vasodilatory agent. A short segment of ITA was excised for histologic examination; immunocytochemistry analysis was performed using antirabbit polyclonal VEGF antibody, rabbit polyclonal inducible nitric oxide synthase (i-NOS) antibody, and adenosine anti-A2B receptor antibody. The immunoreactivity rates then were evaluated. MAIN RESULTS: The mean ITA free flow in the GA + TEA group was significantly higher than in the GA group (56.0 ± 9.0 mL/min v 39.6 ± 14 mL/min, p = 0.001). Immunostaining intensity in the sections after incubation with each primary antibody increased in the GA + TEA group compared with the GA group. CONCLUSIONS: The results of this study indicated that TEA increased ITA free blood flow significantly via increased VEGF, i-NOS, and adenosine-A2B receptor expressions. Therefore, the use of TEA as an adjunct to GA might be considered as an alternative to vasoactive agents for increasing ITA flow in CABG surgery.
Assuntos
Anestesia Epidural/efeitos adversos , Ponte de Artéria Coronária/métodos , Endotélio Vascular/fisiologia , Artérias Torácicas/fisiologia , Vértebras Torácicas , Adenosina/metabolismo , Idoso , Anestesia Geral , Anestésicos Locais/administração & dosagem , Bupivacaína/administração & dosagem , Endotélio Vascular/metabolismo , Feminino , Hemodinâmica/fisiologia , Humanos , Imuno-Histoquímica , Masculino , Pessoa de Meia-Idade , Óxido Nítrico Sintase Tipo II/biossíntese , Receptor A2B de Adenosina/biossíntese , Fluxo Sanguíneo Regional/fisiologia , Tamanho da Amostra , Volume Sistólico/fisiologia , Artérias Torácicas/metabolismo , Fator A de Crescimento do Endotélio Vascular/biossínteseRESUMO
Protein kinase CK1alpha regulates several fundamental cellular processes including proliferation and differentiation. Up to four forms of this kinase are expressed in vertebrates resulting from alternative splicing of exons; these exons encode either the L-insert located within the catalytic domain or the S-insert located at the C terminus of the protein. Whereas the L-insert is known to target the kinase to the nucleus, the functional significance of nuclear CK1alphaLS has been unclear. Here we demonstrate that selective L-insert-targeted short hairpin small interfering RNA-mediated knockdown of CK1alphaLS in human vascular endothelial cells and vascular smooth muscle cells impairs proliferation and abolishes hydrogen peroxide-stimulated proliferation of vascular smooth muscle cells, with the cells accumulating in G(0)/G(1). In addition, selective knockdown of CK1alphaLS in cultured human arteries inhibits vascular activation, preventing smooth muscle cell proliferation, intimal hyperplasia, and proteoglycan deposition. Knockdown of CK1alphaLS results in the harmonious down-regulation of its target substrate heterogeneous nuclear ribonucleoprotein C and results in the altered expression or alternative splicing of key genes involved in cellular activation including CXCR4, MMP3, CSF2, and SMURF1. Our results indicate that the nuclear form of CK1alpha in humans, CK1alphaLS, plays a critical role in vascular cell proliferation, cellular activation, and hydrogen peroxide-mediated mitogenic signal transduction.
Assuntos
Caseína Quinase Ialfa/metabolismo , Proliferação de Células , Células Endoteliais/metabolismo , Endotélio Vascular/metabolismo , Túnica Íntima/metabolismo , Túnica Íntima/patologia , Adulto , Idoso , Idoso de 80 Anos ou mais , Western Blotting , Contagem de Células , Células Cultivadas , Células Endoteliais/patologia , Endotélio Vascular/patologia , Feminino , Citometria de Fluxo , Humanos , Hiperplasia/metabolismo , Hiperplasia/patologia , Masculino , Pessoa de Meia-Idade , Músculo Liso Vascular/metabolismo , Músculo Liso Vascular/patologia , Miócitos de Músculo Liso/metabolismo , Miócitos de Músculo Liso/patologia , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Transdução de Sinais/fisiologia , Artérias Torácicas/metabolismo , Artérias Torácicas/patologiaRESUMO
OBJECTIVE: Alpha2-antiplasmin (alpha2-AP) is the major circulating inhibitor of plasmin, which plays a determining role in the regulation of intravascular fibrinolysis. We investigated the role of alpha(2)-AP on vascular remodeling in response to angiotensin II (Ang II). METHODS AND RESULTS: alpha2-AP-deficient mice were performed. Ang II and N(omega)-nitro- L-arginine methyl ester (L-NAME)-induced perivascular fibrosis was significantly decreased in alpha2-AP-/- mice compared with wild-type mice. In situ gelatinolytic activity analysis shows that perivascular gelatinolytic activity was increased in alpha2-AP-/- mice, which was responsible for decreased perivascular fibrosis in response to Ang II and L-NAME. Ang II-induced arterial wall thickening, vascular cell proliferation, apoptosis, c-Myc, and collagen I expression were significantly decreased in alpha2-AP-/- mice compared with wild-type mice. Further analysis shows that increased p53 and p21 expression were responsible for inhibition of Ang II-induced vascular remodeling in alpha2-AP-/- mice. CONCLUSIONS: The results show that alpha2-AP is a critical regulator for vascular remodeling by inhibiting p53/p21 pathway, suggesting that alpha2-AP is proposed to be a potential therapeutic target for vascular remodeling.
Assuntos
Cardiomegalia/etiologia , Proteínas do Olho/metabolismo , Hipertensão/metabolismo , Fatores de Crescimento Neural/metabolismo , Serpinas/metabolismo , Artérias Torácicas/metabolismo , Angiotensina II , Animais , Apoptose , Cardiomegalia/metabolismo , Cardiomegalia/patologia , Proliferação de Células , Colágeno Tipo I/metabolismo , Inibidor de Quinase Dependente de Ciclina p21/metabolismo , Modelos Animais de Doenças , Proteínas do Olho/genética , Fibrose , Gelatinases/metabolismo , Hipertensão/induzido quimicamente , Hipertensão/complicações , Hipertensão/patologia , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Miocárdio/metabolismo , Miocárdio/patologia , NG-Nitroarginina Metil Éster , Fatores de Crescimento Neural/deficiência , Fatores de Crescimento Neural/genética , Proteínas Proto-Oncogênicas/metabolismo , Proteínas Proto-Oncogênicas c-bcl-2 , Proteínas Proto-Oncogênicas c-myc/metabolismo , Serpinas/deficiência , Serpinas/genética , Artérias Torácicas/enzimologia , Artérias Torácicas/patologia , Proteína Supressora de Tumor p53/metabolismoRESUMO
Although probucol is known to prevent restenosis by regulating vascular remodeling after percutaneous transluminal coronary angioplasty, the mechanisms remain unclear. The present study was designed to investigate whether probucol mediates vascular remodeling via the extracellular signal-regulated kinase 1/2 (ERK1/2) signalling pathway. A rabbit restenosis model was used, in which the New Zealand white rabbits received angioplasty with a 3.5 F angioplasty balloon catheter and the proliferation and migration of smooth muscle cells (SMCs) was induced by oxidized low-density lipoprotein (ox-LDL). We evaluated several vascular remodeling parameters and found that probucol prevented lumen restenosis and mediated expansive remodeling with a remodeling index greater than 1 and that the proliferation and migration of SMCs was inhibited. Based on Western blot analyses, probucol decreased the expression of phospho-mitogen-activated protein kinase kinases 1 (p-MEK1) and phospho-ERK1/2 and enhanced the expression of mitogen-activated protein (MAP) kinase phosphatase-1 (MKP-1) and caveolin-1. Cells treated with the MEK1 inhibitor PD98059 demonstrated a remarkable suppression of the effects of probucol. Furthermore, immunofluorescence analysis showed that probucol inhibited the activation of ERK1/2 by preventing its translocation to the nucleus. It was also found that c-myc expression in aortic tissue after angioplasty and the activator protein 1 (AP1) activity in SMCs induced by ox-LDL were decreased with probucol treatment. In conclusion, probucol mediated vascular remodeling to prevent restenosis after angioplasty by down-regulating the ERK1/2 signaling pathway.
Assuntos
Angioplastia com Balão , Antioxidantes/farmacologia , Miócitos de Músculo Liso/efeitos dos fármacos , Probucol/farmacologia , Artérias Torácicas/efeitos dos fármacos , Túnica Íntima/efeitos dos fármacos , Animais , Aterosclerose/metabolismo , Aterosclerose/patologia , Aterosclerose/terapia , Caveolina 1/metabolismo , Movimento Celular/efeitos dos fármacos , Proliferação de Células/efeitos dos fármacos , Sobrevivência Celular/efeitos dos fármacos , Células Cultivadas , Reestenose Coronária/prevenção & controle , Regulação para Baixo , Fosfatase 1 de Especificidade Dupla/metabolismo , Genes myc/fisiologia , Humanos , Hiperplasia/tratamento farmacológico , Hiperplasia/metabolismo , MAP Quinase Quinase 1/metabolismo , Masculino , Proteína Quinase 1 Ativada por Mitógeno/metabolismo , Proteína Quinase 3 Ativada por Mitógeno/metabolismo , Músculo Liso Vascular/citologia , Miócitos de Músculo Liso/citologia , Miócitos de Músculo Liso/metabolismo , Coelhos , Artérias Torácicas/metabolismo , Artérias Torácicas/patologia , Fator de Transcrição AP-1/metabolismo , Túnica Íntima/patologiaRESUMO
INTRODUCTION: We investigated both the effect and the role(s) of potassium channels, nitric oxide (NO) and cyclooxygenase (COX) products in the effect of hydrogen peroxide (H(2)O(2)) in human internal thoracic artery (ITA) rings. MATERIALS AND METHODS: Samples of redundant ITA obtained from patients undergoing a coronary artery bypass graft surgery were cut into 3 mm wide rings and suspended in 20 ml organ baths. Isometric tension was continuously measured with an isometric force transducer connected to a computer-based data acquisition system. RESULTS: H(2)O(2) (10(-7)-10(-4) M) produced concentration-dependent relaxation responses in human ITA precontracted by phenylephrine. The relaxant responses to H(2)O(2) did not differ significantly between endothelium-intact and endothelium-denuded preparations. Incubation of human ITA rings with superoxide dismutase (50 U/ml) did not affect the relaxant responses to H(2)O(2), while 1,000 U/ml catalase caused a significant decrease. Incubation of endothelium-intact or endothelium-denuded human ITA rings with voltage-dependent potassium channel blocker 4-aminopyridine (5 mM) significantly inhibited the relaxant responses to H(2)O(2). COX inhibitor indomethacin (10(-5) M) also caused a significant inhibition. Incubation with ATP-dependent potassium channel blocker glibenclamide (10(-6) M) or Ca(2+)-activated potassium channel blocker iberiotoxin (10(-7) M) or NO synthase (NOS) blocker N(omega)-nitro-L: -arginine methyl ester (10(-4) M) did not alter relaxant responses of ITA rings to H(2)O(2). CONCLUSION: The findings of the present study suggested that H(2)O(2)-induced relaxation responses in human ITA were neither dependant on the endothelium nor blocked by NOS inhibition but they rather seem to depend on the activation of voltage-dependent potassium channels and COX.
Assuntos
Peróxido de Hidrogênio/farmacologia , Óxido Nítrico/metabolismo , Oxidantes/farmacologia , Prostaglandina-Endoperóxido Sintases/metabolismo , Relação Dose-Resposta a Droga , Endotélio Vascular/efeitos dos fármacos , Humanos , Peróxido de Hidrogênio/administração & dosagem , Técnicas In Vitro , Contração Isométrica/efeitos dos fármacos , Masculino , Pessoa de Meia-Idade , Músculo Liso Vascular/efeitos dos fármacos , Óxido Nítrico Sintase/antagonistas & inibidores , Óxido Nítrico Sintase/metabolismo , Oxidantes/administração & dosagem , Canais de Potássio de Abertura Dependente da Tensão da Membrana , Prostaglandina-Endoperóxido Sintases/efeitos dos fármacos , Artérias Torácicas/metabolismoRESUMO
The aim of our study was to evaluate the relevance of tissue- and species-specific endothelial cells (EC) to study EC-dependent mechanisms in inflammatory-mediated tissue injury. We established an isolation protocol for highly purified EC (pEC) preparations of different origin and compared EC-specific inflammatory responses. Fluorescence-activated cell separation was used to obtain pEC cultures from different human arterial (coronary artery, internal thoracic artery) and venous (umbilical vein, saphenous vein) vessels. All pEC were analyzed for growth kinetics, morphology, release of cytokines/chemokines, and expression of E-selectin. For all different EC cultures, purities of >or=99% were reproducibly achieved. The EC isolation did not affect EC growth, morphology, and function. However, characterization of pEC from different vessel materials revealed an intrinsic, tissue-specific functional heterogeneity of EC cultures. Despite an arterial and venous difference in the secretion of IL-8 and monocyte chemoattractant protein-1, especially EC from coronary arteries produced significantly more IL-6 compared with other EC types, independent of age, gender, and disease of the cell donors. In contrast, the expression of E-selectin was not affected. We conclude that the proposed isolation protocol allows the generation of a pEC bank, enabling us to study tissue-specific aspects at the level of the endothelium.
Assuntos
Vasos Coronários/citologia , Vasos Coronários/metabolismo , Endotélio Vascular/citologia , Endotélio Vascular/metabolismo , Interleucina-6/metabolismo , Idoso , Biomarcadores/metabolismo , Técnicas de Cultura de Células/métodos , Células Cultivadas , Quimiocina CCL2/metabolismo , Feminino , Humanos , Interleucina-8/metabolismo , Masculino , Pessoa de Meia-Idade , Veia Safena/citologia , Veia Safena/metabolismo , Artérias Torácicas/citologia , Artérias Torácicas/metabolismo , Veias Umbilicais/citologia , Veias Umbilicais/metabolismoRESUMO
Here, we report the existence of endothelial precursor (EPC) and stem cells in a distinct zone of the vascular wall that are capable to differentiate into mature endothelial cells, hematopoietic and local immune cells, such as macrophages. This zone has been identified to be localized between smooth muscle and adventitial layer of human adult vascular wall. It predominantly contains CD34-positive (+) but CD31-negative (-) cells, which also express VEGFR2 and TIE2. Only few cells in this zone of the vascular wall are positive for CD45. In a ring assay using the fragments of human internal thoracic artery (HITA), we show here that the CD34+ cells of the HITA-wall form capillary sprouts ex vivo and are apparently recruited for capillary formation by tumor cells. New vessels formed by these vascular wall resident EPCs express markers for angiogenically activated endothelial cells, such as CEACAM1, and also for mature endothelial cells, such as VE-cadherin or occludin. Vascular wall areas containing EPCs are found in large and middle sized arteries and veins of all organs studied here. These data suggest the existence of a ;vasculogenic zone' in the wall of adult human blood vessels, which may serve as a source for progenitor cells for postnatal vasculogenesis, contributing to tumor vascularization and local immune response.
Assuntos
Movimento Celular/fisiologia , Endotélio Vascular/citologia , Endotélio Vascular/crescimento & desenvolvimento , Neovascularização Fisiológica/fisiologia , Células-Tronco/citologia , Células-Tronco/fisiologia , Adulto , Animais , Antígenos CD34/metabolismo , Diferenciação Celular/fisiologia , Células Cultivadas , Células Endoteliais/citologia , Células Endoteliais/metabolismo , Células Endoteliais/fisiologia , Endotélio Vascular/metabolismo , Humanos , Imuno-Histoquímica , Macrófagos/fisiologia , Ratos , Receptores de Fatores de Crescimento do Endotélio Vascular/fisiologia , Artérias Torácicas/citologia , Artérias Torácicas/metabolismo , Artérias Torácicas/fisiologiaRESUMO
OBJECTIVE: The aim of this study was to evaluate the role of glycoxidation in the calcification of the internal thoracic artery (ITA) in diabetes mellitus (DM). METHODS: ITA samples were obtained from 17 patients with type 2 DM (age 62.9 +/- 10.5 years) and 12 age-matched, nondiabetic patients (age 62.5 +/- 10.2 years) who underwent coronary artery bypass grafting. These samples were analyzed histopathologically and assessed for calcification by von Kossa staining and for glycoxidation by immunohistochemistry using anti-N(epsilon)-(carboxymethyl)lysine (CML) antibody. Morphometric evaluation of calcification of the medial layer, intimal thickness and intima-to-media ratio was performed using NIH image software. To evaluate the mechanism of the interaction between calcification and glycoxidation, we developed an in vitro model of calcification of collagen that was chemically modified by glucose, glutaraldehyde or epoxy compound. The calcium-binding activity of these collagens was determined in hydrolysates using atomic absorption spectrophotometry. RESULTS: ITAs of both diabetic and nondiabetic patients were free of atherosclerosis, and no differences were found between the two groups with regard to intimal thickness and intima-to-media ratio. Areas of calcification were noticed in both groups in the tunica media, but not in the tunica intima. Calcium deposits were localized within the extracellular matrix, which was immunohistochemically positive for CML. The extent of medial layer calcification was significantly greater in diabetic patients than nondiabetic subjects, but was independent of known risk factors such as hypertension, hyperlipidemia, obesity and history of old myocardial infarction. The binding activity of collagen was time-dependently increased with in vitro incubation of glucose. A significant increase in the calcium-binding ability was observed in glucose- and glutaraldehyde-modified collagens, but not in epoxy compound-modified collagen. CONCLUSION: Our results suggest that glycoxidative modification of the extracellular matrix, in particular collagen, of the vascular wall may enhance the development of ITA calcification in diabetic patients.
Assuntos
Calcinose/metabolismo , Diabetes Mellitus Tipo 2/metabolismo , Diabetes Mellitus Tipo 2/patologia , Artérias Torácicas/metabolismo , Adulto , Idoso , Idoso de 80 Anos ou mais , Calcinose/epidemiologia , Calcinose/patologia , Cálcio/metabolismo , Colágeno/metabolismo , Diabetes Mellitus Tipo 2/epidemiologia , Proteínas da Matriz Extracelular/metabolismo , Feminino , Glicoproteínas/metabolismo , Humanos , Imuno-Histoquímica , Masculino , Pessoa de Meia-Idade , Oxirredução , Fatores de Risco , Artérias Torácicas/patologiaRESUMO
The effect of peroxynitrite on the development of atherosclerosis is one of the major foci of recent studies. Here, the cytotoxic effect of peroxynitrite was investigated by quantitatively measuring nitrated tyrosine, 3-nitrotyrosine (3-NT) levels in atherosclerotic blood vessels. Atherosclerotic vessels were obtained from the patients who underwent either coronary artery or peripheric artery bypass surgery. Internal thoracic arteries of the patients were treated as non-atherosclerotic control vessels. 3-NT was measured by reverse-phase HPLC and plasma nitrite-nitrate levels were measured by spectrophotometry. 3-NT levels were significantly elevated in atherosclerotic vessels (46.6 +/- 23.3 nmol/mg protein, n = 15; p < 0.001) in comparison to control vessels (15.8 +/- 2.5 nmol/mg protein, n = 10). Vessel 3-NT correlated weakly with plasma nitrate levels (r = 0.38). Thus, atherosclerotic arteries have higher 3-NT levels than non-atherosclerotic blood vessels.
Assuntos
Arteriosclerose/metabolismo , Vasos Sanguíneos/metabolismo , Artérias Torácicas/metabolismo , Tirosina/análogos & derivados , Tirosina/metabolismo , Arteriosclerose/patologia , Estudos de Casos e Controles , Humanos , Técnicas de Diagnóstico Molecular , Nitratos/análise , Nitratos/sangue , Nitritos/análise , Nitritos/sangue , Espectrofotometria , Artérias Torácicas/patologia , Regulação para CimaRESUMO
OBJECTIVE: Endothelial nitric oxide inhibits smooth muscle cell proliferation, reducing the chance of vascular intimal thickening. In this study we investigated whether the superior long-term patency of the internal thoracic artery in human coronary bypass grafting compared with that of the saphenous vein could be explained by different levels of nitric oxide production. METHODS: The baseline endogenous nitric oxide production appeared to be 50% higher in the internal thoracic artery than in the saphenous vein. Previously, it was shown that vascular endothelial growth factor and the vascular endothelial growth factor receptors KDR (Flk-1) and Flt-1 are expressed in both internal thoracic arteries and saphenous veins and that vascular endothelial growth factor receptor density was higher in internal thoracic arteries than in saphenous veins. Therefore, we also investigated the influence of vascular endothelial growth factor on nitric oxide release in both the internal thoracic artery and the saphenous vein. RESULTS: Vascular endothelial growth factor augmented nitric oxide production by approximately 50% in the saphenous vein and 100% in the internal thoracic artery. As shown by means of immunohistochemistry, expression of endothelial constitutive nitric oxide synthase was similar in the internal thoracic artery and the saphenous vein, and no inducible nitric oxide synthase was expressed in any of the vascular segments. CONCLUSION: Vascular endothelial growth factor augments endothelial constitutive nitric oxide synthase-dependent nitric oxide release to a greater extent in the internal thoracic artery than in the saphenous vein. These findings may help to explain the long-term superiority of the internal thoracic artery versus the saphenous vein as a conduit for coronary artery bypass.