RESUMO
BACKGROUND: The metabolism of tryptophan (Trp) along the kynurenine pathway has been shown to carry strong immunoregulatory properties. Several experimental studies indicate that this pathway is a major regulator of vascular inflammation and influences atherogenesis. Knowledge of the role of this pathway in human atherosclerosis remains incomplete. OBJECTIVES: In this study, we performed a multiplatform analysis of tissue samples, in vitro and in vivo functional assays to elucidate the potential role of the kynurenine pathway in human atherosclerosis. METHODS AND RESULTS: Comparison of transcriptomic data from carotid plaques and control arteries revealed an upregulation of enzymes within the quinolinic branch of the kynurenine pathway in the disease state, whilst the branch leading to the formation of kynurenic acid (KynA) was downregulated. Further analyses indicated that local inflammatory responses are closely tied to the deviation of the kynurenine pathway in the vascular wall. Analysis of cerebrovascular symptomatic and asymptomatic carotid stenosis data showed that the downregulation of KynA branch enzymes and reduced KynA production were associated with an increased probability of patients to undergo surgery due to an unstable disease. In vitro, we showed that KynA-mediated signalling through aryl hydrocarbon receptor (AhR) is a major regulator of human macrophage activation. Using a mouse model of peritoneal inflammation, we showed that KynA inhibits leukocyte recruitment. CONCLUSIONS: We have found that a deviation in the kynurenine pathway is associated with an increased probability of developing symptomatic unstable atherosclerotic disease. Our study suggests that KynA-mediated signalling through AhR is an important mechanism involved in the regulation of vascular inflammation.
Assuntos
Doenças das Artérias Carótidas/metabolismo , Cinurenina/metabolismo , Triptofano/metabolismo , Regulação para Baixo , Humanos , Inflamação/metabolismo , Ácido Cinurênico/metabolismo , Cinurenina/sangue , Macrófagos/metabolismo , Placa Aterosclerótica/metabolismo , Triptofano/sangue , Regulação para CimaRESUMO
BACKGROUND: The interplay between innate and adaptive immunity is central in life-threatening clinical complications of atherosclerosis such as myocardial infarction and stroke. The specific mechanisms involved and their protective versus detrimental effects in the disease process remain poorly understood. We have previously shown that higher levels of Toll-like receptor 7 (TLR7) expression in human atherosclerotic lesions are correlated with better patient outcome. OBJECTIVE: In this study, we explored whether TLR7 activation can ameliorate disease in experimental atherosclerosis in mice. METHODS: Apolipoprotein E deficient mice (Apoe-/- ) with established disease were injected for five weeks intraperitoneally with the TLR7 ligand R848. Local effects were evaluated by characterization of the lesion. Systemic effects of the treatment were investigated by immune composition analysis in the spleen and plasma measurements. RESULTS: The in vivo treatment arrested lesion progression in the aorta. We also detected expansion of marginal zone B cells and Treg in the spleen together with increased plasma IgM antibodies against oxidized low-density lipoprotein (oxLDL) and reduced plasma cholesterol levels. These changes were accompanied by increased accumulation of IgM antibodies, decreased necrosis and fewer apoptotic cells in atherosclerotic lesions. CONCLUSIONS: Our findings show that TLR7 stimulation could ameliorate atherosclerotic lesion burden and reduce plasma cholesterol in Apoe-/- mice. TLR7 stimulation was associated with an atheroprotective B-cell and Treg response, which may have systemic and local effects within lesions that could prevent arterial lipid accumulation and inflammation.
Assuntos
Aterosclerose/imunologia , Aterosclerose/prevenção & controle , Hipercolesterolemia/sangue , Receptor 7 Toll-Like/fisiologia , Animais , Anticorpos/sangue , Aorta/patologia , Apolipoproteínas E/deficiência , Apoptose , Aterosclerose/patologia , Linfócitos B/metabolismo , Colesterol/sangue , Modelos Animais de Doenças , Imunoglobulina M/imunologia , Lipoproteínas LDL/imunologia , Camundongos Knockout , Necrose , Baço/metabolismo , Linfócitos T Reguladores/metabolismoRESUMO
BACKGROUND AND OBJECTIVES: The T-cell response to low-density lipoprotein (LDL) in the vessel wall plays a critical role in atherosclerotic plaque formation and stability. In this study, we used a new translational approach to investigate epitopes from human apolipoprotein B100 (ApoB100), the protein component of LDL, which triggers T-cell activation. We also evaluated the potential of two selected native ApoB100 epitopes to modulate atherosclerosis in human ApoB100-transgenic Ldlr-/- (HuBL) mice. METHODS AND RESULTS: HuBL mice were immunized with human atherosclerotic plaque homogenate to boost cellular autoimmune response to tissue-derived ApoB100 epitopes. In vitro challenge of splenocytes from immunized mice with a library of overlapping native peptides covering human ApoB100 revealed several sequences eliciting T-cell proliferation. Of these sequences, peptide (P) 265 and P295 were predicted to bind several human leucocyte antigen (HLA) haplotypes and induced high levels of interferon (IFN)-γ. Vaccination of HuBL mice with these peptides mounted a strong adaptive immune response to native ApoB100, including high levels of epitope-specific plasma IgGs. Interestingly, P265 and P295 vaccines significantly decreased plaque size, reduced macrophage infiltration and increased IgG1 deposition in the plaques. Purified IgGs from vaccinated mice displayed anti-inflammatory properties against macrophages in vitro, reducing their response to LPS in a dose-dependent manner. CONCLUSION: We identified two specific epitopes from human native ApoB100 that trigger T-cell activation and protect HuBL mice against atherosclerosis when used in a vaccine. Our data suggest that vaccination-induced protective mechanisms may be mediated at least in part through specific antibody responses to LDL that inhibit macrophage activation.
Assuntos
Apolipoproteína B-100/imunologia , Aterosclerose/imunologia , Aterosclerose/prevenção & controle , Epitopos de Linfócito T/imunologia , Vacinação , Animais , Apolipoproteína B-100/metabolismo , Modelos Animais de Doenças , Antígenos HLA-D/metabolismo , Humanos , Imunoglobulina G/biossíntese , Inflamação/prevenção & controle , Macrófagos/imunologia , Camundongos , Camundongos Transgênicos , Placa Aterosclerótica/imunologiaRESUMO
BACKGROUND: Embolism from unstable atheromas in the carotid bifurcation is a major cause of stroke. Here, we analysed gene expression in endarterectomies from patients with symptomatic (S) and asymptomatic (AS) carotid stenosis to identify pathways linked to plaque instability. METHODS: Microarrays were prepared from plaques (n = 127) and peripheral blood samples (n = 96) of S and AS patients. Gene set enrichment, pathway mapping and network analyses of differentially expressed genes were performed. RESULTS: These studies revealed upregulation of haemoglobin metabolism (P = 2.20E-05) and bone resorption (P = 9.63E-04) in S patients. Analysis of subgroups of patients indicated enrichment of calcification and osteoblast differentiation in S patients on statins, as well as inflammation and apoptosis in plaques removed >1 month compared to <2 weeks after symptom. By prediction profiling, a panel of 30 genes, mostly transcription factors, discriminated between plaques from S versus AS patients with 78% accuracy. By meta-analysis, common gene networks associated with atherosclerosis mapped to hypoxia, chemokines, calcification, actin cytoskeleton and extracellular matrix. A set of dysregulated genes (LMOD1, SYNPO2, PLIN2 and PPBP) previously not described in atherosclerosis were identified from microarrays and validated by quantitative PCR and immunohistochemistry. CONCLUSIONS: Our findings confirmed a central role for inflammation and proteases in plaque instability, and highlighted haemoglobin metabolism and bone resorption as important pathways. Subgroup analysis suggested prolonged inflammation following the symptoms of plaque instability and calcification as a possible stabilizing mechanism by statins. In addition, transcriptional regulation may play an important role in the determination of plaque phenotype. The results from this study will serve as a basis for further exploration of molecular signatures in carotid atherosclerosis.
Assuntos
Doenças das Artérias Carótidas/genética , Transcriptoma , Idoso , Feminino , Redes Reguladoras de Genes , Humanos , Masculino , Transdução de SinaisRESUMO
Atherosclerosis is a maladaptive, nonresolving chronic inflammatory disease that occurs at sites of blood flow disturbance. The disease usually remains silent until a breakdown of integrity at the arterial surface triggers the formation of a thrombus. By occluding the lumen, the thrombus or emboli detaching from it elicits ischaemic symptoms that may be life-threatening. Two types of surface damage can cause atherothrombosis: plaque rupture and endothelial erosion. Plaque rupture is thought to be caused by loss of mechanical stability, often due to reduced tensile strength of the collagen cap surrounding the plaque. Therefore, plaques with reduced collagen content are thought to be more vulnerable than those with a thick collagen cap. Endothelial erosion, on the other hand, may occur after injurious insults to the endothelium instigated by metabolic disturbance or immune insults. This review discusses the molecular mechanisms involved in plaque vulnerability and the development of atherothrombosis.
Assuntos
Endotélio Vascular , Inflamação/imunologia , Placa Aterosclerótica , Animais , Cisteína Proteases/metabolismo , Endotélio Vascular/metabolismo , Endotélio Vascular/patologia , Endotélio Vascular/fisiopatologia , Humanos , Metaloproteinases da Matriz/metabolismo , Placa Aterosclerótica/complicações , Placa Aterosclerótica/patologia , Placa Aterosclerótica/fisiopatologia , Ruptura Espontânea/complicações , Ruptura Espontânea/metabolismo , Ruptura Espontânea/fisiopatologia , Tromboembolia/etiologiaRESUMO
BACKGROUND: Acute clinical complications of atherosclerosis such as myocardial infarction (MI) and ischaemic stroke are usually caused by thrombus formation on the ruptured plaque surface. Collagen, the main structural protein of the fibrous cap, provides mechanical strength to the atherosclerotic plaque. The integrity of the fibrous cap depends on collagen fibre cross-linking, a process controlled by the enzyme lysyl oxidase (LOX). METHODS AND RESULTS: We studied atherosclerotic plaques from human carotid endarterectomies. LOX was strongly expressed in atherosclerotic lesions and detected in the regions with ongoing fibrogenesis. Higher LOX levels were associated with a more stable phenotype of the plaque. In the studied population, LOX mRNA levels in carotid plaques predicted the risk for future MI. Within the lesion, LOX mRNA levels correlated positively with levels of osteoprotegerin (OPG) and negatively with markers of immune activation. The amount of LOX-mediated collagen cross-links in plaques correlated positively also with serum levels of OPG. CONCLUSIONS: Lysyl oxidase may contribute to the healing of atherosclerotic lesions and to the prevention of its lethal complications. Mediators of inflammation may control LOX expression in plaques and hence plaque stability.
Assuntos
Aterosclerose/enzimologia , Doenças das Artérias Carótidas/enzimologia , Placa Aterosclerótica/enzimologia , Proteína-Lisina 6-Oxidase/metabolismo , Idoso , Idoso de 80 Anos ou mais , Feminino , Humanos , Masculino , Pessoa de Meia-Idade , Infarto do Miocárdio/etiologia , Osteoprotegerina/sangue , Osteoprotegerina/metabolismo , RNA Mensageiro/metabolismo , Fatores de RiscoRESUMO
OBJECTIVES: Atherosclerosis is an inflammatory disease of the arterial wall that leads to myocardial infarction and stroke. Regulatory T cells (Tregs) and IL-10 exert significant anti-atherogenic effects in experimental models of atherosclerosis by modulating vascular inflammation. We have previously shown that Mycobacterium bovis BCG killed by extended freeze-drying (EFD BCG) decreases lung and colon inflammation by recruiting IL-10-producing Tregs. Therefore, the aim of this study was to investigate the effect of EFD BCG on the development of atherosclerosis. DESIGN: We used two strains of atherosclerosis-prone mice: Ldlr(-/-) (four or six EFD BCG injections) and Apoe(-/-) (six injections). RESULTS: In both models, EFD BCG significantly reduced the size of atherosclerotic lesions, increased IL-10 production and reduced the serum levels of pro-inflammatory cytokines (IL-6, IL-13, KC and tumour necrosis factor-α). Shortly after treatment with EFD BCG, the number of plasmacytoid dendritic cells (pDCs) and Foxp3(+) Tregs in the draining lymph nodes increased. EFD BCG also led to accumulation of Tregs, but not of pDCs in the spleen, and reduced activity of NF-κB and increased activity of PPAR-γ in both the spleen and vascular tissue of treated mice. CONCLUSION: EFD BCG has atheroprotective effects through IL-10 production and Treg expansion. These findings support a novel approach to the prevention and treatment of atherosclerosis.
Assuntos
Aterosclerose , Vacina BCG/farmacologia , Interleucina-10/metabolismo , Mycobacterium bovis/imunologia , Linfócitos T Reguladores , Animais , Aterosclerose/imunologia , Aterosclerose/prevenção & controle , Modelos Animais de Doenças , Fatores de Transcrição Forkhead/metabolismo , Liofilização/métodos , Fenômenos do Sistema Imunitário/efeitos dos fármacos , Camundongos , PPAR gama/metabolismo , Linfócitos T Reguladores/efeitos dos fármacos , Linfócitos T Reguladores/imunologiaRESUMO
OBJECTIVE: Strokes, a major cause of disability, are often caused by embolism from unstable carotid plaques. The aim of this study was to validate a biobank of human carotid endarterectomies as a platform for further exploration of pathways for plaque instability. For this purpose, we investigated the relationship between clinical parameters of plaque instability and expression of genes previously shown to be associated with either plaque instability or healing processes in the vessel wall. METHODS: A database of clinical information and gene-expression microarray data from 106 carotid endarterectomies were used. RESULTS: Expression of matrix metalloproteinase (MMP)-9 and MMP-7 was 100-fold higher in plaques than in normal artery. In general, genes associated with inflammation (such as RANKL and CD68) were overexpressed in symptomatic compared with asymptomatic plaques. Plaques obtained from patients undergoing surgery within 2 weeks after an embolic event showed up-regulation of genes involved in healing reactions in the vessel wall (including elastin and collagen). Statin treatment, as well as echodense lesions, were associated with a more stable phenotype. CONCLUSION: Here, we demonstrate that gene-expression profiles reflect clinical parameters. Our results suggest that microarray technology and clinical variables can be used for the future identification of central molecular pathways in plaque instability.
Assuntos
Estenose das Carótidas/genética , Perfilação da Expressão Gênica , Embolia Intracraniana/genética , Idoso , Idoso de 80 Anos ou mais , Antígenos CD/genética , Antígenos de Diferenciação Mielomonocítica/genética , Estenose das Carótidas/cirurgia , Colágeno/genética , Bases de Dados Genéticas , Elastina/genética , Endarterectomia das Carótidas , Feminino , Humanos , Masculino , Metaloproteinase 7 da Matriz/genética , Metaloproteinase 9 da Matriz/genética , Pessoa de Meia-Idade , Análise de Sequência com Séries de Oligonucleotídeos , Fenótipo , Ligante RANK/genética , Estatística como Assunto , Suécia , Regulação para Cima/genética , Cicatrização/genéticaRESUMO
Differentiation of muscle cells is characterized morphologically by the acquisition of contractile filaments and characteristic shape changes, and on the molecular level by induction of the expression of several genes, including those for the muscle-specific alpha-actin isoforms. IFN-gamma is an inhibitor of proliferation for several cells, including vascular smooth muscle, and is also an inducer of differentiated properties for several hematopoietic cells. We have therefore investigated whether IFN-gamma affects the expression of alpha-smooth muscle actin in cultured arterial smooth muscle cells. Cells exposed to IFN-gamma show a reduction of alpha-smooth muscle actin-containing stress fibers, as detected by immunofluorescence. The effect was observed in all phases of the cell cycle, and was caused by a reduction of the synthesis of alpha-smooth muscle actin protein as revealed by two-dimensional electrophoretic analysis of actin isoforms. RNA hybridization using a cRNA probe that hybridizes to all actin mRNAs showed that IFN-gamma-treated cells have a reduced content of the 1.7-kb mRNA that codes for alpha-smooth muscle actin, and to a lesser extent, also of the 2.1-kb mRNA encoding the beta and gamma-cytoplasmic actins. The reduction of alpha-smooth muscle actin mRNA was confirmed using an alpha-smooth muscle actin-specific cRNA probe. The reduction of alpha-smooth muscle actin mRNA occurs within 12 h, and is dependent on protein synthesis, since cycloheximide treatment reversed the effect. The inhibition of this mRNA species was dose dependent, and detectable by RNA hybridization at a dose of 50 U/ml IFN-gamma. These results suggest that the differentiation of arterial smooth muscle cells is not necessarily coupled to an inhibition of cellular proliferation. Instead, IFN-gamma may regulate the expression of several genes that control both proliferation and expression of differentiation markers.
Assuntos
Actinas/genética , Divisão Celular/efeitos dos fármacos , Regulação da Expressão Gênica/efeitos dos fármacos , Interferon gama/farmacologia , Músculo Liso Vascular/citologia , Citoesqueleto de Actina/ultraestrutura , Animais , Northern Blotting , Diferenciação Celular/efeitos dos fármacos , Células Cultivadas , Desmina/fisiologia , Relação Dose-Resposta a Droga , Imunofluorescência , Técnicas In Vitro , Filamentos Intermediários/ultraestrutura , Músculo Liso Vascular/fisiologia , RNA Mensageiro/genética , Ratos , Proteínas RecombinantesRESUMO
Endothelial cells regulate vascular tone by secreting paracrine mediators that control the contractility of arterial smooth muscle cells. Nitric oxide (NO) is an important vasodilating agent that is generated from L-arginine by the enzyme nitric oxide synthase (NOS), which is expressed constitutively by the endothelium. NO also inhibits platelet aggregation, contributing to the antithrombotic properties of the endothelial surface. It would therefore be expected that loss of the endothelium during arterial injury would lead to vasospasm and thrombosis but instead, the neointima formed after injury has a nonthrombogenic surface and a maintained vascular patency. We report here that arterial smooth muscle cells in the neointima formed after a deendothelializing balloon injury to the rat carotid artery express the cytokine-inducible isoform of NOS. Expression was detectable by reverse transcription-polymerase chain reaction from day 1-14 after injury and in situ hybridization showed expression of NOS mRNA by neointimal smooth muscle cells, particularly at the surface of the lesion. This was associated with systemically detectable NO production as revealed by electron paramagnetic resonance spectroscopic analysis of nitrosylated red cell hemoglobin. Local NO production by intimal smooth muscle cells after endothelial injury could represent an important mechanism for the maintenance of arterial patency and nonthrombogenicity in the injured artery.
Assuntos
Aminoácido Oxirredutases/biossíntese , Endotélio Vascular/enzimologia , Endotélio Vascular/lesões , Músculo Liso Vascular/enzimologia , Animais , Artérias Carótidas , Hibridização In Situ , Óxido Nítrico Sintase , Reação em Cadeia da Polimerase , Ratos , Ratos Sprague-DawleyAssuntos
Doenças Cardiovasculares , Inflamação/imunologia , Placa Aterosclerótica , Imunidade Adaptativa , Doenças Cardiovasculares/etiologia , Doenças Cardiovasculares/prevenção & controle , Humanos , Imunidade Celular , Metabolismo dos Lipídeos , Placa Aterosclerótica/complicações , Placa Aterosclerótica/metabolismo , Placa Aterosclerótica/fisiopatologia , Placa Aterosclerótica/terapia , Terapias em EstudoRESUMO
Atherosclerosis is a chronic inflammatory disease characterized by accumulation of oxidized lipoproteins, increased cell death and hypertrophic degeneration of the arterial intima. The disease process is associated with local formation of modified self antigens that are targeted by both innate and adaptive immune responses. Although it remains to be firmly established it is likely that these autoimmune responses initially have a beneficial effect facilitating the removal of potentially harmful rest products from oxidized LDL and dying cells. However, studies performed on hypercholesterolaemic mice deficient in different components of the immune system uniformly suggest that the net effect of immune activation is pro-atherogenic and that atherosclerosis, at least to some extent, should be regarded as an autoimmune disease. These observations point to the possibility of developing new treatments for atherosclerosis based on modulation of immune responses against plaque antigens, an approach presently tested clinically for several other chronic inflammatory diseases with autoimmune components. Pilot studies in animals have provided promising results for both parental and oral vaccines based on oxidized LDL antigens. The time when this concept is ready for clinical testing is rapidly approaching but it will be important not to underestimate the difficulties that will be encountered in transferring the promising results from experimental animals into humans.
Assuntos
Aterosclerose/imunologia , Autoimunidade , Adulto , Animais , Apoptose , Aterosclerose/terapia , Autoantígenos/imunologia , Autoantígenos/metabolismo , Criança , Células Dendríticas/imunologia , Células Dendríticas/metabolismo , Proteínas de Choque Térmico/imunologia , Proteínas de Choque Térmico/metabolismo , Humanos , Lipoproteínas LDL/imunologia , Lipoproteínas LDL/metabolismo , Camundongos , Linfócitos T/imunologia , Linfócitos T/metabolismo , Células Th1/imunologia , Células Th1/metabolismoRESUMO
The aim of the study was to investigate T cell receptor (TCR) usage at the time of diagnosis of giant cell arteritis (GCA) and to estimate the degree of clonality of T-cells infiltrating the lesion. Seven patients with biopsy-proven giant cell arteritis were included in the study. Immunocytochemistry in biopsies from the temporal arteries and flow cytometric analysis of peripheral blood lymphocytes (PBL) was performed using monoclonal antibodies specific for CD3, CD4 and CD8 and 13 TCR Valpha and Vbeta gene segment products. The CDR3 fragment length polymorphism was assessed by gel electrophoresis of PCR-amplified TCR segments. The T lymphocytes were found to be concentrated to the adventitia rather than the media or intima. Six of the seven patients with GCA had expansions of T lymphocytes, expressing selected TCR V genes in the arterial wall. None of these expansions was found in PBL. The infiltrating T-cells were poly- or oligoclonal. In conclusion, the dominating part of the inflammatory infiltrate in GCA emanates from the adventitial microvessels. There is an uneven expression of TCR V genes by T lymphocytes in the inflammatory infiltrates as compared to peripheral blood T lymphocytes at the time of diagnosis, consistent with an antigen-driven immunological reaction in the arterial wall.
Assuntos
Linfócitos T CD4-Positivos/imunologia , Linfócitos T CD8-Positivos/imunologia , Arterite de Células Gigantes/imunologia , Receptores de Antígenos de Linfócitos T alfa-beta/análise , Artérias Temporais/imunologia , Idoso , Complexo CD3/análise , Complexo CD3/genética , Feminino , Citometria de Fluxo , Regulação da Expressão Gênica , Arterite de Células Gigantes/genética , Humanos , Imuno-Histoquímica , Imunofenotipagem , Masculino , Pessoa de Meia-Idade , Receptores de Antígenos de Linfócitos T alfa-beta/sangue , Receptores de Antígenos de Linfócitos T alfa-beta/genéticaRESUMO
Decay-accelerating factor (DAF) is a constitutively expressed plasma membrane glycoprotein on blood cells and endothelium that inhibits cell surface C3/C5 convertase formation, thus inhibiting complement activation and protecting cells from lysis by the terminal complement components. Using monoclonal anti-DAF antibodies in conjunction with anti-smooth muscle cell (SMC)-specific myosin antibodies, it was found by immunohistochemistry that vascular SMC in advanced human carotid atherosclerotic lesions express DAF antigen. The percentage of DAF-positive SMC ranged from 20 to 60% between different patient samples and SMC DAF expression was limited to SMC in the lesion proper. Normal arterial wall SMC exhibited no DAF-specific immunostaining. Essentially 100% of passaged cultured vascular SMC derived from normal human uterine artery, or from umbilical vein, expressed DAF as assessed by immunocytochemistry. A 68-kD band was observed on SDS-PAGE autoradiograms of DAF-immunoprecipitated radiolabeled cultured SMC extracts. Sensitization of rabbit erythrocytes with DAF-containing SMC extracts conferred protection against complement-mediated hemolysis in normal human serum and the protective effect could be reversed by treatment with anti-DAF antibodies. We conclude that DAF is induced on vascular SMC during atherogenesis and in culture.
Assuntos
Arteriosclerose/metabolismo , Proteínas de Membrana/análise , Músculo Liso Vascular/análise , Antígenos CD55 , Células Cultivadas , Glicoproteínas/análise , Humanos , Proteínas de Membrana/imunologia , Proteínas de Membrana/fisiologiaRESUMO
The scavenger receptor (ScR) mediates uptake of chemically modified low density lipoprotein (LDL) by human monocyte-derived macrophages. It is not down-regulated by high intracellular cholesterol levels, and exposure of macrophages to acetylated or oxidized LDL therefore leads to foam cell development. The hypothesis that this represents an important mechanism for intracellular cholesterol accumulation in atherosclerosis is supported by the finding of ScR expression in foam cells of atherosclerotic plaques. T lymphocytes are also present in such plaques and it is known that T cell products regulate macrophage activation. We have therefore studied the effect of interferon-gamma (IFN gamma), a lymphokine secreted by activated T lymphocytes, on the expression of ScR in human monocyte-derived macrophages. Binding and uptake of acetylated LDL were significantly reduced in macrophages exposed to recombinant IFN gamma or IFN gamma-containing lymphocyte-conditioned media. Competition experiments showed that the IFN gamma-regulated binding and uptake of acetylated LDL was mediated via ScR. IFN gamma exerted its effect on the saturable binding of acetylated LDL by reducing the number of cell surface binding sites without significantly affecting the affinity between acetylated LDL and its receptor. Northern analysis revealed that the type I ScR mRNA was significantly reduced in IFN gamma-treated cells. Finally, IFN gamma treatment reduced intracellular cholesteryl ester accumulation and inhibited the development of foam cells in the cultures. In conclusion, our data show that IFN gamma blocks the development of macrophage-derived foam cells by inhibiting expression of ScR. This suggests that macrophage-T lymphocyte interactions may reduce intracellular cholesterol accumulation in the atherosclerotic plaque.
Assuntos
Moléculas de Adesão Celular , Células Espumosas/efeitos dos fármacos , Interferon gama/farmacologia , Macrófagos/efeitos dos fármacos , Proteínas de Membrana , Receptores Imunológicos/efeitos dos fármacos , Receptores de LDL/efeitos dos fármacos , Receptores de Lipoproteínas , Células Cultivadas , Ésteres do Colesterol/metabolismo , Citoplasma/química , Relação Dose-Resposta a Droga , Citometria de Fluxo , Humanos , Lipoproteínas LDL/análise , Lipoproteínas LDL/metabolismo , Macrófagos/metabolismo , RNA Mensageiro/análise , Receptores Imunológicos/metabolismo , Receptores de LDL/genética , Receptores de LDL/metabolismo , Receptores Depuradores , Receptores Depuradores Classe BRESUMO
Atherosclerosis is an inflammatory-fibrotic response to accumulation of cholesterol in the artery wall. In hypercholesterolemia, low density lipoproteins (LDL) accumulate and are oxidized to proinflammatory compounds in the arterial intima, leading to activation of endothelial cells, macrophages, and T lymphocytes. We have studied immune cell activation and the autoimmune response to oxidized LDL in atherosclerotic apo E-knockout mice. Autoantibodies to oxidized LDL exhibited subclass specificities indicative of T cell help, and the increase in antibody titers in peripheral blood was associated with increased numbers of cytokine-expressing T cells in the spleen. In addition to T cell-dependent antibodies, IgM antibodies to oxidized LDL were also increased in apo E-knockout mice. This suggests that both T cell-dependent and T cell-independent epitopes may be present on oxidized LDL. In moderate hypercholesterolemia, IgG antibodies were largely of the IgG2a isotype, suggesting that T cell help was provided by proinflammatory T helper (Th) 1 cells, which are prominent components of atherosclerotic lesions. In severe hypercholesterolemia induced by cholesterol feeding of apo E-knockout mice, a switch to Th2-dependent help was evident. It was associated with a loss of IFN-gamma-producing Th1 cells in the spleen, whereas IL-4-producing Th2 cells were more resistant to hypercholesterolemia. IFN-gamma but not IL-4 mRNA was detected in atherosclerotic lesions of moderately hypercholesterolemic apo E-knockout mice, but IL-4 mRNA appeared in the lesions when mice were made severely hypercholesterolemic by cholesterol feeding. These data show that IFN-gamma-producing Th1 cells infiltrate atherosclerotic lesions and provide T cell help for autoimmune responses to oxidized LDL in apo E-knockout mice. However, severe hypercholesterolemia is associated with a switch from Th1 to Th2, which results not only in the formation of IgG1 autoantibodies to oxidized LDL, but also in the appearance of Th2-type cytokines in the atherosclerotic lesions. Since the two subsets of T cells counteract each other, this switch may have important consequences for the inflammatory/immune process in atherosclerosis.
Assuntos
Apolipoproteínas E/genética , Arteriosclerose/imunologia , Autoimunidade , Hipercolesterolemia/imunologia , Células Th1/imunologia , Células Th2/imunologia , Animais , Arteriosclerose/sangue , Arteriosclerose/genética , Autoanticorpos/sangue , Sequência de Bases , Primers do DNA/genética , Epitopos de Linfócito T , Hipercolesterolemia/sangue , Hipercolesterolemia/genética , Interferon gama/biossíntese , Interferon gama/genética , Interleucina-4/biossíntese , Interleucina-4/genética , Lipoproteínas LDL/imunologia , Masculino , Camundongos , Camundongos Knockout , Reação em Cadeia da Polimerase , RNA Mensageiro/genética , RNA Mensageiro/metabolismo , Baço/imunologia , Baço/patologiaRESUMO
A large proportion of the cells of the human atherosclerotic plaque is assumed to be derived from medial smooth muscle cells. In contrast to these, the cells of the plaque have the capacity to accumulate lipid, and they also proliferate at a higher rate than medial cells. It has therefore been suggested that smooth muscle cells undergo a change of phenotype during atherogenesis, but there has been no evidence for such a change on the molecular level. We have now analyzed carotid artery plaques using a battery of antibodies against cell surface and cytoskeletal antigens, and found that most of the cells express the class II transplantation antigen (Ia antigen) HLA-DR. Also, the beta chain of HLA-DR was detected by immunoblotting of plaque extracts with the OKIa1 monoclonal antibody. HLA-DR is normally present on cells of the immune system, but only 60% of the DR-positive cells of the plaque reacted with monoclonal antibodies specific for macrophages and lymphocytes. Many of the remaining DR-positive cells contained the muscle-specific intermediate filament protein, desmin. This indicates that smooth muscle cells of atherosclerotic plaques express DR antigen. In contrast, very few DR-positive cells were found in normal human arteries. This suggests that expression of class II antigen is part of a phenotypic change in smooth muscle cells in atherosclerosis.
Assuntos
Arteriosclerose/imunologia , Antígenos de Histocompatibilidade Classe II/análise , Idoso , Arteriosclerose/patologia , Diferenciação Celular , Desmina/metabolismo , Imunofluorescência , Antígenos HLA-DR , Humanos , Pessoa de Meia-Idade , Músculo Liso Vascular/imunologia , Receptores Fc/análiseRESUMO
Atherosclerosis is associated with immune activation. T cells and macrophages infiltrate atherosclerotic plaques and disease progression is associated with formation of autoantibodies to oxidized lipoproteins. In the apo E knockout mouse, a genetic model of cholesterol-induced atherosclerosis, congenital deficiency of macrophages, lymphocytes, or interferon-gamma receptors result in reduced lesion formation. We have now evaluated whether immune modulation in the adult animal affects disease development. Injections of 7-wk-old male apo E knockout mice with polyclonal immunoglobulin preparations (ivIg) during a 5-d period reduced fatty streak formation over a 2-mo period on cholesterol diet by 35%. Fibrofatty lesions induced by diet treatment for 4 mo were reduced by 50% in mice receiving ivIg after 2 mo on the diet. ivIg treatment also reduced IgM antibodies to oxidized LDL and led to inactivation of spleen and lymph node T cells. These data indicate that ivIg inhibits atherosclerosis, that it is effective both during the fatty streak and plaque phases, and that it may act by modulating T cell activity and/or antibody production. Therefore, immunomodulation may be an effective way to prevent and/or treat atherosclerosis.
Assuntos
Adjuvantes Imunológicos/uso terapêutico , Apolipoproteínas E/deficiência , Arteriosclerose/terapia , Imunoglobulina G/farmacologia , Animais , Aorta/patologia , Arteriosclerose/imunologia , Arteriosclerose/patologia , Divisão Celular/imunologia , Dieta , Histocitoquímica , Imunoglobulina M/imunologia , Inflamação/imunologia , Lipoproteínas LDL/imunologia , Camundongos , Camundongos Knockout , Linfócitos T/metabolismoRESUMO
Atherosclerosis is a chronic disease with high morbidity and mortality around the globe. It is characterized by chronic inflammation of the vessel wall, which is perpetuated by the continuous migration of cells to and within the atherosclerotic lesion. Chemokines (CK) and chemokine receptors (CKR) together with other chemoattractants and adhesion molecules are major mediators facilitating this process. Many CK/CKR (CC, CX3C and CXC) and other chemoattractants (e.g. leukotrienes) have been implicated in atherogenesis, but only a few have been validated as pathogenic by in vitro assays, in vivo experiments using gene-targeted animal models and genetic studies. Promising attempts are currently made to inhibit CK-dependent cell recruitment to lesion by using neutralizing antibodies, mutant proteins, viral and synthetic inhibitors or receptor antagonists. Some of the therapeutics have already entered clinical trials for other conditions and are about to be tested in human atherosclerosis. However, our limited understanding of the complex CK system and the functional specialization of individual CK/CKR, translatability of animal research into human population, limitations of current imaging techniques and surrogate markers for evaluation of the benefits of potential anti-CK compounds are still hampering therapeutic exploitation of the CK system in atherosclerosis. Hopefully we will be able to solve many of these issues in the near future and use this approach to control atherosclerotic disease in man.
Assuntos
Aterosclerose/terapia , Quimiocinas/antagonistas & inibidores , Quimiocinas/fisiologia , Animais , Aterosclerose/patologia , Movimento Celular/efeitos dos fármacos , Humanos , Receptores de Quimiocinas/efeitos dos fármacos , Receptores de Quimiocinas/fisiologiaRESUMO
OBJECTIVE: Inflammatory processes play an important role in atherosclerosis, and increasing evidence implies that microbial pathogens and proinflammatory cytokines are involved in the development and activation of atherosclerotic lesions. To find new inflammatory genes, we explored the vascular transcriptional response to an activator of innate immunity bacterial lipopolysaccharides (LPSs). METHODS AND RESULTS: Gene arrays identified the cytomegalovirus-inducible gene 5 (cig5)/viperin among the genes most potently induced by LPS in human vascular biopsies. Viperin was expressed by endothelial cells in atherosclerotic arteries and significantly elevated in atherosclerotic compared with normal arteries. In culture, cytomegalovirus infection, interferon-gamma, and LPS induced viperin expression. CONCLUSIONS: Viperin is expressed in atherosclerosis and induced in vascular cells by inflammatory stimuli and cytomegalovirus infection. The putative functions of viperin in atherosclerosis may relate to disease-associated microbes.