RESUMEN
The mechanisms of ectopic bone formation in arteries are poorly understood. Osteoblasts might originate either from stem cells that penetrate atherosclerotic plaques from the blood stream or from pluripotent mesenchymal cells that have remained in the arterial wall from embryonic stages of the development. We have examined the frequency of the expression and spatial distribution of osteoblast-specific factor-2/core binding factor-1 (Osf2/Cbfa1) in carotid and coronary arteries. Cbfa1-expressing cells were rarely observed but were found in all tissue specimens in the deep portions of atherosclerotic plaques under the necrotic cores. The deep portions of atherosclerotic plaques under the necrotic cores were characterized by the lack of capillaries of neovascularization. In contrast, plaque shoulders, which were enriched by plexuses of neovascularization, lacked Cbfa1-expressing cells. No bone formation was found in any of the 21 carotid plaques examined and ectopic bone was observed in only two of 12 coronary plaques. We speculate that the sparse invasion of sprouts of neovascularization into areas underlying the necrotic cores, where Cbfa1-expressing cells reside, might explain the rarity of events of ectopic bone formation in the arterial wall. This study has also revealed that Cbfa1-expressing cells contain alpha-smooth muscle actin and myofilaments, indicating their relationship with arterial smooth muscle cells.
Asunto(s)
Arterias , Aterosclerosis/patología , Subunidad alfa 1 del Factor de Unión al Sitio Principal/metabolismo , Osificación Heterotópica , Osteoblastos/fisiología , Osteogénesis/fisiología , Anciano , Arterias/metabolismo , Arterias/patología , Subunidad alfa 1 del Factor de Unión al Sitio Principal/genética , Humanos , Masculino , Persona de Mediana Edad , Osteoblastos/citologíaRESUMEN
Mishandled concerns about clinical standards resulted in whistleblowing in four Australian hospitals. Official inquiries followed with recommendations to improve patient safety. In the aftermath of the inquiries, common themes included loss of trust in management and among clinical colleagues, and loss of trust from patients and the community. Without first rebuilding trust, staff will not report mistakes or other concerns about safety. Successful implementation of patient safety procedures requires policies to stress the professional duty of staff to report concerns about colleagues when they believe there is a risk to patients.
Asunto(s)
Actitud del Personal de Salud , Errores Médicos/prevención & control , Personal de Hospital/psicología , Administración de la Seguridad , Denuncia de Irregularidades , Australia , Administración Hospitalaria/métodos , Hospitales/normas , Humanos , Relaciones Interprofesionales , ConfianzaRESUMEN
Over recent years, the role of matrix vesicles in the initial stages of arterial calcification has been recognized. Matrix calcifying vesicles have been isolated from atherosclerotic arteries and the biochemical composition of calcified vesicles has been studied. No studies have yet been carried out to examine the fine structure of matrix vesicles in order to visualize the features of the consequent stages of their calcification in arteries. In the present work, a high resolution ultrastructural analysis has been employed and the study revealed that matrix vesicles in human atherosclerotic lesions are heterogeneous with two main types which we classified. Type I calcified vesicles were presented by vesicles surrounded by two electron-dense layers and these vesicles were found to be resistant to the calcification process in atherosclerotic lesions in situ. Type II matrix vesicles were presented by vesicles surrounded by several electron-dense layers and these vesicles were found to represent calcifying vesicles in atherosclerotic lesions. To test the hypothesis that calcification of matrix vesicles surrounded by multilayer sheets may occur simply as a physicochemical process, independently from the cell regulation, we produced multilamellar liposomes and induced their calcification in vitro in a manner similar to that occurring in matrix vesicles in atherosclerotic lesions in situ.
Asunto(s)
Aterosclerosis/patología , Calcinosis/patología , Arterias Carótidas/ultraestructura , Matriz Extracelular/ultraestructura , Anciano , Femenino , Humanos , Masculino , Microscopía Electrónica de Transmisión , Persona de Mediana EdadRESUMEN
We have previously demonstrated that amounts of ganglioside GM3 are markedly higher in human atherosclerotic lesions compared to that in non-diseased arterial tissue. Because the fatty acid composition of GM3 in blood plasma low density lipoproteins (LDL) and the fatty acid composition of GM3 in atherosclerotic lesions differed, we hypothesized that, in addition to GM3 originating from LDL infiltrating the arterial wall from the blood, excessive GM3 may be synthesized locally in atherosclerotic lesions. In the present work, using an anti-GM3 antibody developed by us, we showed that the levels of GM3 synthase in membrane fractions isolated from the atherosclerotic intima were higher compared to those in non-diseased arterial tissue. Using an immunohistochemical approach, we examined the expression of GM3 synthase in sections of atherosclerotic plaques and non-diseased arterial wall. GM3 synthase immunopositivity was found to be low in non-diseased arterial intima but large numbers of GM3 synthase-immunopositive cells were observed in atherosclerotic plaques. GM3 synthase was overexpressed by macrophages and dendritic cells and double immunostaining demonstrated cellular co-localization of GM3 synthase and GM3. Further in vitro experiments showed that both monocyte-derived dendritic cells and macrophages expressed high levels of GM3 synthase. The findings of the present study indicate that, at least partially, excessive amounts of GM3 in atherosclerotic lesions can be synthesized by macrophages and dendritic cells directly within the arterial wall.
Asunto(s)
Aterosclerosis/enzimología , Sialiltransferasas/biosíntesis , Túnica Íntima/enzimología , Adulto , Anciano , Aorta/enzimología , Aorta/patología , Aterosclerosis/patología , Western Blotting , Arterias Carótidas/enzimología , Arterias Carótidas/patología , Células Cultivadas , Células Dendríticas/enzimología , Citometría de Flujo , Humanos , Inmunohistoquímica , Técnicas In Vitro , Macrófagos/enzimología , Persona de Mediana Edad , Túnica Íntima/patologíaRESUMEN
Only a few previous works investigated the involvement of Chlamydia pneumoniae (Chlamydophila pneumoniae) in arterial calcification. The present study investigated a possible association between C. pneumoniae and medial calcification. Carotid artery segments obtained by endarterectomy from 60 patients were examined by PCR and immunohistochemistry to identify the presence of C. pneumoniae. Arterial specimens showing double-positive (n = 17), double-negative (n = 22), and single-positive results (n = 21) were further analyzed by a combination of histology, immunohistochemistry, and electron microscopy. Medial calcification occurred in 10 of 17 (58.8%) C. pneumoniae double-positive arterial specimens, but no medial calcification was observed in any of 22 C. pneumoniae double-negative arterial specimens. Electron microscopy indicated C. pneumoniae in smooth muscle cells (SMCs) in foci of medial calcification. Medial SMCs showing damage to the cytoplasm and basement membrane contained the structures with the appearance of elementary, reticulate, and aberrant bodies of C. pneumoniae. The presence of C. pneumoniae in SMCs was confirmed by electron-microscopic immunocytochemistry. In the extracellular matrix, calcification was observed in C. pneumoniae aberrant bodies that exited the SMCs. The findings offer a new hypothesis of arterial calcification: they suggest that C. pneumoniae infection of medial SMCs may be associated with the pathophysiological events of arteriosclerotic calcification of the tunica media.
Asunto(s)
Aterosclerosis/microbiología , Aterosclerosis/patología , Calcinosis/microbiología , Calcinosis/patología , Chlamydophila pneumoniae/aislamiento & purificación , Túnica Media/microbiología , Túnica Media/patología , Anciano , Humanos , Hallazgos Incidentales , Masculino , Persona de Mediana Edad , Estadística como AsuntoRESUMEN
Atherogenesis is a complex process involving inflammation. S100A8 and S100A9, the Ca2+-binding neutrophil cytosolic proteins, are associated with innate immunity and regulate processes leading to leukocyte adhesion and transmigration. In neutrophils and monocytes the S100A8-S100A9 complex regulates phosphorylation, NADPH-oxidase activity, and fatty acid transport. The proteins have anti-microbial properties, and S100A8 may play a role in oxidant defense in inflammation. Murine S100A8 is regulated by inflammatory mediators and recruits macrophages with a proatherogenic phenotype. S100A9 but not S100A8 was found in macrophages in ApoE-/- murine atherosclerotic lesions, whereas both proteins are expressed in human giant cell arteritis. Here we demonstrate S100A8 and S100A9 protein and mRNA in macrophages, foam cells, and neovessels in human atheroma. Monomeric and complexed forms were detected in plaque extracts. S100A9 was strongly expressed in calcifying areas and the surrounding extracellular matrix. Vascular matrix vesicles contain high levels of Ca2+-binding proteins and phospholipids that regulate calcification. Matrix vesicles characterized by electron microscopy, x-ray microanalysis, nucleoside triphosphate pyrophosphohydrolase assay and cholesterol/phospholipid analysis contained predominantly S100A9. We propose that S100A9 associated with lipid structures in matrix vesicles may influence phospholipid-Ca2+ binding properties to promote dystrophic calcification. S100A8 and S100A9 were more sensitive to hypochlorite oxidation than albumin or low density lipoprotein and immunoaffinity confirmed S100A8-S100A9 complexes; some were resistant to reduction, suggesting that hypochlorite may contribute to protein cross-linking. S100A8 and S100A9 in atherosclerotic plaque and calcifying matrix vesicles may significantly influence redox- and Ca2+-dependent processes during atherogenesis and its chronic complications, particularly dystrophic calcification.
Asunto(s)
Arterias/patología , Calgranulina A/fisiología , Calgranulina B/fisiología , Adulto , Anciano , Anciano de 80 o más Años , Antiinfecciosos/farmacología , Aorta/patología , Apolipoproteínas E/genética , Arterias/ultraestructura , Aterosclerosis/metabolismo , Western Blotting , Calcio/metabolismo , Calgranulina A/metabolismo , Calgranulina B/metabolismo , Arterias Carótidas/patología , Adhesión Celular , Movimiento Celular , Colesterol/química , Reactivos de Enlaces Cruzados/farmacología , Citosol/metabolismo , Relación Dosis-Respuesta a Droga , Endotelio Vascular/metabolismo , Femenino , Células Espumosas/metabolismo , Humanos , Ácido Hipocloroso/farmacología , Inmunohistoquímica , Hibridación in Situ , Inflamación , Lípidos/química , Macrófagos/metabolismo , Masculino , Microscopía Electrónica , Microscopía Electrónica de Transmisión , Persona de Mediana Edad , Monocitos/metabolismo , Neutrófilos/metabolismo , Oxidantes/química , Oxígeno/química , Fenotipo , Fosfolípidos/metabolismo , Fosforilación , ARN Mensajero/metabolismo , Proteínas Recombinantes/químicaRESUMEN
We previously reported that CD1d, a molecule responsible for the presentation of lipid antigens, is expressed in atherosclerotic lesions and that its expression is restricted to dendritic cells. Recent studies demonstrating that CD1d-restricted natural killer T (NKT) cells are involved in atherogenesis prompted the present study investigating whether NKT cells are present in human atherosclerotic lesions and, if so, whether there is an association between NKT cells and dendritic cells. We found that NKT cells do accumulate in rupture-prone shoulders of atherosclerotic plaques and observed direct contacts of dendritic cells with NKT cells in rupture-prone regions of plaque.
Asunto(s)
Arteriosclerosis/inmunología , Arteriosclerosis/patología , Células Dendríticas/patología , Células Asesinas Naturales/patología , Arterias Carótidas/patología , Humanos , InmunohistoquímicaRESUMEN
BACKGROUND: Since graduated compression stockings (GCS) reduce the risk of deep venous thrombosis (DVT) in both hospital and ambulant patients, we checked the compressive efficiency of 20-30 mmHg GCS in the standing position. METHODS: In 30 volunteers (17 normal legs, 13 varicose legs), duplex ultrasound was used to measure the internal diameters of the long saphenous vein, posterior tibial veins, peroneal veins, and soleal veins in the lying and standing position and with and without 20-30 mmHg GCS. RESULTS: Graduated compression stockings effectively compressed both superficial and deep veins in supine individuals but not the superficial or the deep veins when standing. In the varicose leg, the stockings did not compress the long saphenous vein at the mid-calf level even when supine. In the varicose leg the long saphenous vein was constricted at the upper band of the stocking, which might explain why superficial venous thrombosis is more common when compression stockings are worn. CONCLUSIONS: In the standing position, GCS did not compress the deep or superficial veins of the calf.
Asunto(s)
Vendajes , Postura , Várices/terapia , Femenino , Humanos , Pierna/irrigación sanguínea , Masculino , Persona de Mediana Edad , Fenómenos Físicos , FísicaRESUMEN
Dendritic cells (DCs) populate atherosclerotic lesions and might be involved in the regulation of immune reactions in atherosclerosis. The present work was undertaken to examine a possible association of DCs with Chlamydophila pneumoniae in human atherosclerotic plaques obtained by endarterectomy. C. pneumoniae was identified in 17 of 60 (28%) atherosclerotic plaques by a combination of immunohistochemistry and polymerase chain reaction (PCR). Double immunohistochemistry identified the presence of C. pneumoniae within S100(+) DCs that were localised predominantly in the deep layer of the intima under the necrotic core. Quantitative analysis showed that there were no differences in the numbers of DCs between C. pneumoniae(+) and C. pneumoniae(-) groups of atherosclerotic specimens. There were also no differences in the expression of Lag-antigen and HLA-DR by DCs between the groups of specimens. Markers of DC activation CD80 and CD86 were absent from both groups of specimens. Flow cytometry analysis of the effects of C. pneumoniae infection on immature monocyte-derived DCs in vitro showed no changes in the expression of CD1a, MHC class II, CD80 and CD86. The results of this study demonstrate that C. pneumoniae might infect DCs within the atherosclerotic intima but whether the presence of C. pneumoniae in DCs affects the intensity of immune reactions in atherosclerosis needs further clarification.
Asunto(s)
Arteriosclerosis/microbiología , Arteriosclerosis/patología , Infecciones por Chlamydia/diagnóstico , Chlamydophila pneumoniae/aislamiento & purificación , Anciano , Biopsia con Aguja , Estudios de Casos y Controles , Células Cultivadas , Células Dendríticas/microbiología , Células Dendríticas/patología , Femenino , Citometría de Flujo , Técnica del Anticuerpo Fluorescente Indirecta , Humanos , Inmunohistoquímica , Masculino , Persona de Mediana Edad , Probabilidad , Valores de Referencia , Medición de Riesgo , Muestreo , Sensibilidad y Especificidad , Túnica Íntima/microbiología , Túnica Íntima/patologíaRESUMEN
OBJECTIVE: Dendritic cells (DCs) accumulate in atherosclerotic lesions but their characteristics and their role in atherogenesis are poorly understood. C1q, an element of the first component of complement, is expressed by interdigitating dendritic cells and follicular dendritic cells in the spleen. It has been suggested that C1q is involved in capturing immune complexes in the lymphoid tissue. Immune complexes are also detected in atherosclerotic lesions. The present study investigated whether C1q is expressed by DCs in the arterial wall. Because DCs accumulating within atherosclerotic lesions might originate from monocytes that infiltrate the intima from very early stages of atherosclerosis, C1q expression was also examined in monocyte-derived DCs in vitro. METHODS: Specimens of the aorta, carotid, mammary, popliteal and tibial arteries were obtained during operation. Expression of C1q in the arterial wall was studied by immunohistochemistry. The nature of cells expressing C1q was studied in sections double stained with antibodies to C1q and cell type specific markers including CD1a and S-100 (for identification of DCs), CD68 (macrophages), CD3 (T-cells), von Willebrand factor (endothelial cells), and smooth muscle alpha-actin (smooth muscle cells). In vitro, DCs were differentiated from human peripheral blood monocytes using GM-CSF and IL-4. Peripheral blood monocytes were differentiated to macrophages using M-CSF. The expression of C1q in monocytes and in vitro monocyte-derived DCs and macrophages was determined by RT-PCR, Western blotting, immunofluorescence microscopy and flow cytometry. RESULTS: In all the arterial specimens studied, DCs expressing C1q were detected. C1q was also found in macrophages, macrophage foam cells and in neovascular endothelial cells in atherosclerotic lesions, but no C1q expression was detected in T-cells and smooth muscle cells. In vitro analysis demonstrated that monocyte-derived DCs and macrophages express C1q but no C1q was detected in monocytes. CONCLUSION: C1q is expressed by DCs residing in the arterial wall as well as by monocyte-derived DCs in vitro. Expression of C1q occurs during differentiation of monocytes to DCs and macrophages and might be important in binding and trapping immune complexes in atherosclerotic lesions.
Asunto(s)
Complejo Antígeno-Anticuerpo , Arteriosclerosis/inmunología , Complemento C1q/análisis , Células Dendríticas/inmunología , Túnica Íntima/inmunología , Actinas/análisis , Aorta , Arterias Carótidas , Células Cultivadas , Citometría de Flujo , Células Espumosas/inmunología , Humanos , Inmunohistoquímica/métodos , Macrófagos/inmunología , Arterias Mamarias , Microscopía Confocal , Arteria Poplítea , Reacción en Cadena de la Polimerasa de Transcriptasa Inversa , Arterias TibialesRESUMEN
Regulation of alphavbeta3 and alpha5beta1 integrin function plays a crucial role in atherosclerosis. Possible regulators of integrin-matrix interactions are integrin-binding ADAMs (proteins with a disintegrin- and metalloproteinase-domain), like ADAM-15 and ADAM-9. Molecular interactions between ADAM-15, alpha5beta1, and alphavbeta3 have been demonstrated. ADAM-9 and ADAM-15 were found to be interdependently regulated. This study, therefore, investigated whether the upregulation of integrins alpha5beta1 and alphavbeta3 was correlated with the expression of integrin-binding ADAMs in atherosclerotic processes. Human arterial and venous vascular smooth muscle cells (VSMCs) were incubated with PDGF over different time intervals up to a 3-day culture period. mRNA concentrations, quantified by real-time RT-PCR and normalized to PBGD, of integrins alphavbeta3 and alpha5beta1 were strongly increased after a 12-h PDGF-incubation in arterial and venous VSMC. ADAM-15 and ADAM-9 mRNA production showed a corresponding increase following integrin upregulation after a 24-h incubation period. Western blot anaylsis revealed an increased protein expression of integrins and ADAMs in PDGF-stimulated VSMC. Additionally, mRNA concentrations of atherosclerotic and normal human specimens were quantified by real-time RT-PCR. mRNA of ADAMs and integrins was significantly increased in atherosclerotic arteries compared to normal arteries. Immunohistochemistry of these specimens showed an increased expression and codistribution of both ADAMs and integrins in atherosclerosis. In conclusion, upregulation of ADAM-15 and ADAM-9 in atherosclerosis appears to follow an increase in alpha5beta1 and alphavbeta3 integrins. Since alpha5beta1 and alphavbeta3 are known to promote smooth muscle cell migration and proliferation, upregulation of ADAM-15 and ADAM-9 could balance integrin-matrix interactions and cell migration, thus modulating neointima progression.
Asunto(s)
Arteriosclerosis/metabolismo , Desintegrinas/biosíntesis , Desintegrinas/metabolismo , Integrina alfa5beta1/biosíntesis , Integrina alfaVbeta3/biosíntesis , Proteínas de la Membrana/biosíntesis , Metaloendopeptidasas/biosíntesis , Proteínas ADAM , Anciano , Arterias/citología , Arterias/metabolismo , Arterias/patología , Arteriosclerosis/genética , Arteriosclerosis/patología , Cartilla de ADN/genética , Femenino , Humanos , Integrina alfa5beta1/genética , Integrina alfa5beta1/metabolismo , Integrina alfaVbeta3/genética , Integrina alfaVbeta3/metabolismo , Masculino , Proteínas de la Membrana/metabolismo , Metaloendopeptidasas/metabolismo , Persona de Mediana Edad , Músculo Liso Vascular/metabolismo , Músculo Liso Vascular/patología , Factor de Crecimiento Derivado de Plaquetas/farmacología , ARN Mensajero/biosíntesis , ARN Mensajero/genética , Reacción en Cadena de la Polimerasa de Transcriptasa Inversa/métodos , Regulación hacia Arriba , Venas/citología , Venas/metabolismo , Venas/patologíaRESUMEN
Plasminogen activator inhibitor-1 (PAI-1) and two-chain high molecular weight kininogen (HKa) exert anti-adhesive properties in vitronectin-dependent cell adhesion. Here, the hypothesis was tested that these anti-adhesive components promote apoptosis in vascular cells. PAI-1 or HKa induced a 2- to 3-fold increase in apoptosis of human umbilical-vein endothelial cells (HUVEC) and vascular smooth muscle cells (VSMC) adherent to vitronectin, as determined by annexin V-FACS assay, similar to alphav-integrin inhibitor cyclo-(Arg-Gly-Asp-D-Phe-Val)-peptide (cRGDfV). Apoptosis occurred after 12 h incubation and was attributable to caspase 3 activation that in turn induced DNA fragmentation. Induction of apoptosis strongly correlated with the anti-adhesive effect of PAI-1 and HKa on these cells. In contrast, PAI-1 and HKa did not affect fibronectin-dependent adhesion or cell survival. uPA did not influence apoptosis in vitronectin- or fibronectin-adherent cells. In atherosclerotic vessel sections, congruent distribution of vitronectin, PAI-1, HK, and of components of the urokinase plasminogen activator/receptor system with apoptotic cells lining foam cell lesions was demonstrated by immunostaining. These results indicate that inhibition of vitronectin-dependent cell adhesion through PAI-1 and HKa correlates with apoptosis induction in vascular cells mediated through the caspase 3 pathway. Co-distribution of apoptosis with plasminogen activation system components in atherosclerosis exemplifies the significance of anti-adhesive mechanisms and apoptosis for tissue remodeling, such as in neointima development.
Asunto(s)
Apoptosis/efectos de los fármacos , Endotelio Vascular/citología , Quininógenos/farmacología , Músculo Liso Vascular/citología , Inhibidor 1 de Activador Plasminogénico/farmacología , Apoptosis/fisiología , Arteriosclerosis/fisiopatología , Adhesión Celular/efectos de los fármacos , Células Cultivadas , Fibronectinas/farmacología , Humanos , Peso Molecular , Vitronectina/farmacologíaAsunto(s)
Amputación Quirúrgica , Isquemia/terapia , Pierna/irrigación sanguínea , Enfermedad Crónica , Humanos , Isquemia/complicaciones , Resistencia a la Meticilina , Infecciones Estafilocócicas/complicaciones , Infecciones Estafilocócicas/tratamiento farmacológico , Staphylococcus aureus/efectos de los fármacos , Insuficiencia del Tratamiento , Resistencia a la VancomicinaRESUMEN
OBJECTIVE: Overexpression of heat shock proteins (HSPs) is an important means of cell protection during physiologic stress such as occurs during atherogenesis. Immune responses are early events in atherosclerosis, with recent studies indicating that both humoral and cellular autoimmune processes in atherogenesis are directed toward HSPs. Dendritic cells are the key cells in the initiation and regulation of immune responses. This study examined whether HSP70 is overexpressed by dendritic cells in atherosclerotic lesions. METHODS: Twenty-six carotid artery and 16 aortic specimens obtained at endarterectomy and aortic reconstruction were examined with immunohistochemical techniques. The nature of cells that overexpressed HSP70 was studied in consecutive sections that were double stained with antibodies to HSP70 and cell type-specific markers, including CD1a and fascin (to identify dendritic cells), CD14 (monocytes), CD68 (macrophages), CD3 (T cells), CD15 (mast cells), von Willibrand factor (endothelial cells), and alpha-smooth muscle actin (smooth muscle cells). Staining with HLA-DR and CD1d was used to identify cells involved in antigen presentation. RESULTS: In advanced atherosclerotic lesions, several cell types, including monocytes, macrophages, dendritic cells, and smooth muscle cells, overexpressed HSP70. In contrast, in early atherosclerotic lesions, only dendritic cells overexpressed HSP70. Dendritic cells that overexpressed HSP70 frequently contacted T cells and also expressed HLA-DR. Furthermore, dendritic cells that clustered with T cells expressed CD1d, a unique molecule responsible for presenting lipid antigens. CONCLUSION: The results suggest that direct contacts between activated dendritic cells that overexpress HSP70 and T cells might be responsible for T cell activation and might facilitate the presentation of lipid antigens to T cells directly within the arterial wall. In early intimal lesions, HSP70 is overexpressed exclusively by dendritic cells, which suggests that dendritic cells might be involved in the early phases of atherogenesis.