RESUMEN
Sialic acids (Sias), 9-carbon-backbone sugars, are among the most complex and versatile molecules of life. As terminal residues of glycans on proteins and lipids, Sias are key elements of glycotopes of both cellular and microbial lectins and thus act as important molecular tags in cell recognition and signaling events. Their functions in such interactions can be regulated by post-synthetic modifications, the most common of which is differential Sia-O-acetylation (O-Ac-Sias). The biology of O-Ac-Sias remains mostly unexplored, largely because of limitations associated with their specific in situ detection. Here, we show that dual-function hemagglutinin-esterase envelope proteins of nidoviruses distinguish between a variety of closely related O-Ac-Sias. By using soluble forms of hemagglutinin-esterases as lectins and sialate-O-acetylesterases, we demonstrate differential expression of distinct O-Ac-sialoglycan populations in an organ-, tissue- and cell-specific fashion. Our findings indicate that programmed Sia-O-acetylation/de-O-acetylation may be critical to key aspects of cell development, homeostasis, and/or function.
Asunto(s)
Acetilesterasa/biosíntesis , Hemaglutininas Virales/genética , Ácido N-Acetilneuramínico/genética , Ácidos Siálicos/genética , Proteínas Virales de Fusión/genética , Acetilación , Acetilesterasa/genética , Animales , Regulación de la Expresión Génica , Genoma , Hemaglutininas Virales/química , Hemaglutininas Virales/metabolismo , Humanos , Lípidos/química , Lípidos/genética , Mamíferos , Ácido N-Acetilneuramínico/química , Ácido N-Acetilneuramínico/metabolismo , Nidovirales/química , Proteínas/química , Proteínas/genética , Ácidos Siálicos/química , Especificidad de la Especie , Proteínas Virales de Fusión/química , Proteínas Virales de Fusión/metabolismoRESUMEN
Infection is a major cause of failure of inserted or implanted biomedical devices (biomaterials). During surgery, bacteria may adhere to the implant, initiating biofilm formation. Bacteria are also observed in and recultured from the tissue surrounding implants, and may even reside inside host cells. Whether these bacteria originate from biofilms is not known. Therefore, we investigated the fate of Staphylococcus epidermidis inoculated on the surface of implants as adherent planktonic cells or as a biofilm in mouse experimental biomaterial-associated infection. In order to discriminate the challenge strain from potential contaminating mouse microflora, we constructed a fully virulent green fluorescent S. epidermidis strain. S. epidermidis injected along subcutaneous titanium implants, pre-seeded on the implants or pre-grown as biofilm, were retrieved from the implants as well as the surrounding tissue in all cases after 4days, and in histology bacteria were observed in the tissue co-localizing with macrophages. Thus, bacteria adherent to or in a biofilm on the implant are a potential source of infection of the surrounding tissue, and antimicrobial strategies should prevent both biofilm formation and tissue colonization.
Asunto(s)
Biopelículas/crecimiento & desarrollo , Infecciones Relacionadas con Prótesis/inmunología , Infecciones Relacionadas con Prótesis/microbiología , Infecciones Estafilocócicas/inmunología , Infecciones Estafilocócicas/microbiología , Staphylococcus epidermidis/inmunología , Titanio , Animales , Adhesión Bacteriana , Femenino , Ratones , Ratones Endogámicos C57BL , Infecciones Estafilocócicas/patologíaRESUMEN
AIMS: Genetic factors explain a proportion of the inter-individual variation in the risk for atherosclerotic events, but the genetic basis of atherosclerosis and atherothrombosis in families with Mendelian forms of premature atherosclerosis is incompletely understood. We set out to unravel the molecular pathology in a large kindred with an autosomal dominant inherited form of premature atherosclerosis. METHODS AND RESULTS: Parametric linkage analysis was performed in a pedigree comprising 4 generations, of which a total of 11 members suffered from premature vascular events. A parametric LOD-score of 3.31 was observed for a 4.4 Mb interval on chromosome 12. Upon sequencing, a non-synonymous variant in KERA (c.920C>G; p.Ser307Cys) was identified. The variant was absent from nearly 28,000 individuals, including 2,571 patients with premature atherosclerosis. KERA, a proteoglycan protein, was expressed in lipid-rich areas of human atherosclerotic lesions, but not in healthy arterial specimens. Moreover, KERA expression in plaques was significantly associated with plaque size in a carotid-collar Apoe-/- mice (r2â=â0.69; p<0.0001). CONCLUSION: A rare variant in KERA was identified in a large kindred with premature atherosclerosis. The identification of KERA in atherosclerotic plaque specimen in humans and mice lends support to its potential role in atherosclerosis.
Asunto(s)
Aterosclerosis/genética , Análisis Mutacional de ADN , Ligamiento Genético , Mutación , Linaje , Proteoglicanos/genética , Anciano , Animales , Apolipoproteínas E/deficiencia , Aterosclerosis/patología , Espacio Extracelular/metabolismo , Femenino , Humanos , Masculino , Ratones , Persona de Mediana Edad , Simulación de Dinámica Molecular , Conformación Proteica , Proteoglicanos/químicaRESUMEN
As biomarker discovery takes centre-stage, the role of immunohistochemistry within that process is increasing. At the same time, the number of antibodies being produced for "research use" continues to rise and it is important that antibodies to be used as biomarkers are validated for specificity and sensitivity before use. This guideline seeks to provide a stepwise approach for the validation of an antibody for immunohistochemical assays, reflecting the views of a consortium of academic and pharmaceutical based histopathology researchers. We propose that antibodies are placed into a tier system, level 1-3, based on evidence of their usage in immunohistochemistry, and that the degree of validation required is proportionate to their place on that tier.
Asunto(s)
Anticuerpos/química , Biomarcadores/metabolismo , Inmunohistoquímica/métodos , Proteínas/química , Animales , Biomarcadores/química , Biomarcadores de Tumor/metabolismo , Investigación Biomédica/métodos , Línea Celular , Química Farmacéutica/métodos , Epítopos/química , Regulación Neoplásica de la Expresión Génica , Humanos , Neoplasias/inmunología , Neoplasias/metabolismo , Fenotipo , ARN Mensajero/metabolismo , Reproducibilidad de los ResultadosRESUMEN
Tuberculosis (TB), caused by Mycobacterium (M.) tuberculosis, is a devastating infectious disease causing many deaths world-wide. Thrombomodulin (TM) is a multidomain glycoprotein expressed on all vascular endothelial cells. We here studied the role of the lectin-like domain of TM, responsible for a variety of anti-inflammatory properties of TM, during TB. We compared the extent of TM-expression in human lung tissue of TB and control patients. The, the role of the lectin-like domain of TM was investigated by comparing mice lacking this domain (TMLeD/LeD mice) with wild-type (WT) mice during experimental lung TB induced by infection with M. tuberculosis via the airways. Lungs were harvested for analyses at two, six and 29 weeks after infection. Lung TM-expression was downregulated in TB patients, which was not related to changes in the amount of endothelium in infected lungs. TMLeD/LeD mice showed unaltered mycobacterial loads in lungs, liver and spleen during experimental TB. Additionally, lung histopathology and cytokine concentrations were largely similar in TMLeD/LeD and WT mice, while total leukocyte counts were increased in lungs of TMLeD/LeD mice after 29 weeks of infection. Mortality did not occur in either group. The lectin-like domain of TM does not play an important role in the host response to M. tuberculosis infection in mice.
Asunto(s)
Pulmón/metabolismo , Mycobacterium tuberculosis/patogenicidad , Trombomodulina/metabolismo , Tuberculosis Pulmonar/metabolismo , Adolescente , Adulto , Anciano , Anciano de 80 o más Años , Animales , Estudios de Casos y Controles , Células Cultivadas , Citocinas/metabolismo , Modelos Animales de Enfermedad , Femenino , Interacciones Huésped-Patógeno , Humanos , Mediadores de Inflamación/metabolismo , Pulmón/microbiología , Pulmón/patología , Masculino , Ratones , Ratones Endogámicos C57BL , Ratones Mutantes , Persona de Mediana Edad , Pronóstico , Estructura Terciaria de Proteína , Bazo/inmunología , Bazo/metabolismo , Bazo/microbiología , Trombomodulina/genética , Factores de Tiempo , Tuberculosis Pulmonar/genética , Tuberculosis Pulmonar/microbiología , Tuberculosis Pulmonar/patología , Adulto JovenRESUMEN
OBJECTIVE: Angiotensin-converting enzyme and its effector peptide angiotensin II have been implicated in the pathogenesis of acute respiratory distress syndrome. Recently, angiotensin-converting enzyme 2 was identified as the counter-regulatory enzyme of angiotensin-converting enzyme that converts angiotensin II into angiotensin-(1-7). The aim of this study was to determine pulmonary angiotensin-converting enzyme and angiotensin-converting enzyme 2 activity in patients with acute respiratory distress syndrome. DESIGN: Prospective observational pilot study. SETTING: A PICU of a university hospital. PATIENTS: Fourteen patients admitted, requiring mechanical ventilation for respiratory syncytial virus lower respiratory tract infection. INTERVENTIONS: None. MEASUREMENTS AND MAIN RESULTS: Two groups of patients were distinguished at admission: a group fulfilling the criteria for acute respiratory distress syndrome and a non-acute respiratory distress syndrome group. Angiotensin-converting enzyme and angiotensin-converting enzyme 2 activity were measured in bronchoalveolar lavage fluid. Patients with acute respiratory distress syndrome had increased angiotensin-converting enzyme activity and decreased angiotensin-converting enzyme 2 activity (p < 0.001) compared with the control group. CONCLUSION: It is shown for the first time that in acute respiratory distress syndrome, enhanced angiotensin-converting enzyme activity is paralleled by a reduced angiotensin-converting enzyme 2 activity, similar to that found in an experimental rat model of acute respiratory distress syndrome. The reduced angiotensin-converting enzyme 2 activity may be counteracted by restoring angiotensin-(1-7) level, thereby offering a novel treatment modality for this syndrome.
Asunto(s)
Peptidil-Dipeptidasa A/metabolismo , Síndrome de Dificultad Respiratoria del Recién Nacido/enzimología , Enzima Convertidora de Angiotensina 2 , Líquido del Lavado Bronquioalveolar/química , Estudios de Casos y Controles , Femenino , Humanos , Lactante , Recién Nacido , Pulmón/enzimología , Masculino , Peptidil-Dipeptidasa A/análisis , Estudios ProspectivosRESUMEN
BACKGROUND: Preoperative portal vein embolization (PVE) is used to increase future remnant liver volume through induction of hepatocellular regeneration. This event, however, potentially enhances tumor growth. The aim of our study was to assess tumor growth and liver regeneration after PVE in a rabbit hepatic tumor model. The VX2 carcinoma is derived from a virus-induced papilloma tumor in rabbits. The tumor grows rapidly, and its blood supply is similar to that of human hepatocellular carcinoma. MATERIALS AND METHODS: Two weeks after subcapsular implantation of a VX2 carcinoma in the cranial liver lobe, New Zealand White rabbits were allocated to a control or PVE group (n = 5 per group). In the PVE group, the portal vein branch to the cranial liver lobes (80%) was embolized using particles and coils, leaving the caudal liver lobe (20%) free. In the tumor control group, the liver was mobilized. Computed tomography volumetry was performed on days 3, 7, 10, and 14. Tumor growth rate (TGR), hepatocellular proliferation rate, and liver damage parameters were assessed before PVE and on days 1, 3, 7, 10, and 14. RESULTS: Portography confirmed complete occlusion of the portal vein branch to the cranial liver lobes in all PVE rabbits. The hypertrophy response and proliferation rate in the nonembolized liver lobes were significantly higher in the PVE group, which was confirmed by liver-to-body weight index assessment. TGR was increased in both groups, with a significantly larger increase in the PVE group over time (day 14: mean, 34.4 ± 4.3 mL/d versus control: mean, 24.1 ± 7.2 mL/d; P < 0.05). CONCLUSIONS: TGR was significantly increased after PVE in the rabbit tumor model. This finding supports the notion that PVE potentially enhances tumor growth, along with the regeneration of the nonembolized liver lobe.
Asunto(s)
Embolización Terapéutica/efectos adversos , Neoplasias Hepáticas Experimentales/patología , Vena Porta , Animales , Proliferación Celular , Modelos Animales de Enfermedad , Femenino , Hipertrofia , Regeneración Hepática , Conejos , Tomografía Computarizada por Rayos XRESUMEN
Determination of hepatocyte proliferation activity is hampered by the presence of Ki67-positive non-parenchymal cells. We validated a multicolor immunohistochemical (IHC) approach using multispectral tissue and cell segmentation software. Portal vein branches to the cranial liver lobes of 10 rabbits were embolized, leading to atrophy of the cranial lobes and hyperplasia of the caudal lobes. Slides from cranial and caudal lobes (n=20) were double-stained (CK8+18 and Ki67) and triple-stained (CK8+18, Ki67, and CD31). The Ki67 proliferation index was calculated using automated tissue and cell segmentation software and compared with manual counting by two independent observers. A substantial variation was seen in the number of Ki67-positive hepatocytes in the different specimens in both double and triple staining (range, 0-50). Correlation coefficients between manual counting and the digital analysis were 0.76 for observer 1 (p<0.001) and 0.78 for observer 2 (p<0.001) with double staining and R(2) = 0.91 for observer 1 and R(2) = 0.89 for observer 2, p<0.001 with triple staining. In conclusion, in rabbit, the hepatocellular proliferation index can be reliably determined using automated tissue and cell segmentation software in combination with IHC multiple staining. Our findings may be useful in clinical practice when Ki67 proliferation index yields prognostic significance.
Asunto(s)
Hepatocitos/citología , Antígeno Ki-67/metabolismo , Hígado/citología , Animales , Compartimento Celular , Proliferación Celular , Hepatocitos/metabolismo , Humanos , Inmunohistoquímica , Queratina-18/metabolismo , Queratina-8/metabolismo , Hígado/metabolismo , Molécula-1 de Adhesión Celular Endotelial de Plaqueta/metabolismo , Conejos , Programas Informáticos , Coloración y EtiquetadoRESUMEN
BACKGROUND: Episodes of microvascular proliferation associated with volume expansion have been observed in arteriovenous malformations (AVMs) of skin and soft tissue. OBJECTIVE: We sought to investigate the relationship between a microvascular proliferative response and flow velocity in AVMs. METHODS: Resection specimens of 80 AVMs were clinically categorized as either high- or low-flow lesions, and histopathologically screened for the presence of microvessels, inflammation, thrombosis, or a combination of these. Immunohistochemistry was performed with endoglin (CD105), von Willebrand factor, and fibrinogen antibodies. RESULTS: Clinically, 37 AVMs were classified as high-flow lesions and 43 as low-flow lesions. In 81% of high-flow lesions microvascular proliferations were seen versus in 14% of low-flow lesions (P < .005). In high-flow lesions, which were embolized before surgery (30% of all), 88% showed microvascular proliferation, 88% inflammation, and 33% thrombosis. However, similar vasoproliferative responses were also observed in nonembolized AVM (69% high-flow and 14% low-flow lesions). Endoglin was more frequently expressed in high-flow lesions. Extracellular von Willebrand factor staining was found in most lesions, irrespective of flow type or presence of microvascular proliferations. LIMITATIONS: The study was carried out at a single tertiary referral center. CONCLUSIONS: Microvascular proliferative masses in AVMs appear to be strongly associated with high-flow characteristics. This could be explained to some extent by previous therapeutic embolization and/or inflammation in the lesion. However, occurrence of similar microvascular responses in AVM that were not embolized before surgery suggests that the biomechanical effects of high flow in these lesions may also have an angiogenic effect.
Asunto(s)
Malformaciones Arteriovenosas/patología , Malformaciones Arteriovenosas/fisiopatología , Embolización Terapéutica/efectos adversos , Inflamación/complicaciones , Microvasos/patología , Adolescente , Adulto , Anciano , Velocidad del Flujo Sanguíneo , Niño , Preescolar , Femenino , Humanos , Masculino , Persona de Mediana Edad , Estudios Retrospectivos , Adulto JovenRESUMEN
Neutrophils are important cellular sources of interleukin (IL) 17A and -F. Moreover, upon activation neutrophils are able to excrete chromatin embedded with components from their cytoplasmic granules to form 'neutrophil extracellular traps' (NETs). Recent studies suggested that NETs contribute to thrombosis by promoting fibrin deposition and platelet aggregation. IL17A may also promote thrombosis by enhancing platelet aggregation. In the present study we investigated the presence of neutrophils, NETs and IL17A and -F in coronary thrombosuction specimens obtained from patients after acute myocardial infarction. Neutrophils and NETs were identified using histochemical (HE, Feulgen procedure) and immunohistochemical stainings (Histone H1, myeloperoxidase, neutrophil elastase) in 15 fresh, 15 lytic and 15 organised thrombi. The presence and distribution of IL17A and -F was studied using (immuno)histochemical double staining and spectral image analysis, rtPCR and Western blot. High numbers of neutrophils are present (10-30% of the thrombus mass) in fresh and lytic, but not in organized thrombus. NETs were frequently observed in fresh (4/15) and lytic (12/15), but never in organised thrombus specimens. Double staining combining the Feulgen reaction with Histone-H1, MPO or neutrophil elastase confirmed colocalisation with DNA. Cytoplasmatic IL17A/F staining was found in the majority of the neutrophils, extracellularly and in NETs. Western blotting confirmed the presence of IL17A and IL17F in thrombus specimens. In conclusion, a large burden of neutrophils, neutrophil extracellular traps and IL17A and -F are important constituents of fresh and lytic thrombus after acute myocardial infarction. The specific colocalisation of these indicates a role during thrombus stabilisation and growth.
Asunto(s)
Interleucina-17/análisis , Infarto del Miocardio/inmunología , Activación Neutrófila , Neutrófilos/inmunología , Trombosis/inmunología , Biomarcadores/análisis , Western Blotting , Histonas/análisis , Humanos , Inmunohistoquímica , Interleucina-17/genética , Elastasa de Leucocito/análisis , Infarto del Miocardio/genética , Neutrófilos/enzimología , Peroxidasa/análisis , ARN Mensajero/análisis , Reacción en Cadena de la Polimerasa de Transcriptasa Inversa , Colorantes de Rosanilina , Trombectomía , Trombosis/genética , Trombosis/cirugíaRESUMEN
OBJECTIVE: Macrophages are decisive in the chronic inflammatory processes that drive atherogenesis. The purpose of this study was to explore the presence and spatial distribution of polarized macrophage populations in human atherosclerosis. METHODS & RESULTS: We used transcriptomics and immunohistochemistry to analyze macrophage subset dynamics in successive stages of atherogenesis. Developing lesions progressively accumulated both M1 and M2 cells, as was signified by the enhanced expression of associated markers at the transcriptional and protein level. Histologically, these markers were confined to overlapping, but spatially distinct CD68(+) areas of the intima. We subsequently quantified the presence of these markers in relation to morphological determinants of plaque stability. In line with their pro-inflammatory characteristics, M1 macrophages dominated the rupture-prone shoulder regions of the plaque over M2 polarized cells, while the fibrous caps of lesions showed no significant differences between subsets. In contrast, vascular adventitial tissue displayed a pronounced M2 activation profile. As expected, areas of intraplaque hemorrhage clearly associated with CD163 staining. Rather than being limited to complicated lesions, this M2 marker was also readily detectable in stable plaques. Finally, foamy macrophages displayed an ambiguous repertoire that incorporates individual M1 and M2 markers. CONCLUSION: M1 and M2 macrophage populations are present throughout atherogenesis. These subsets display disparity when it comes to their prevalence in morphological compartments of the vessel wall. Our current findings warrant continued investigation into the functional implications of polarized macrophage populations in human atherosclerosis.
Asunto(s)
Arterias Carótidas/inmunología , Enfermedades de las Arterias Carótidas/inmunología , Mediadores de Inflamación/análisis , Macrófagos/inmunología , Adventicia/inmunología , Adventicia/patología , Anciano , Anciano de 80 o más Años , Biomarcadores/análisis , Arterias Carótidas/patología , Enfermedades de las Arterias Carótidas/genética , Enfermedades de las Arterias Carótidas/patología , Progresión de la Enfermedad , Femenino , Fibrosis , Perfilación de la Expresión Génica , Regulación de la Expresión Génica , Humanos , Inmunohistoquímica , Macrófagos/clasificación , Macrófagos/patología , Masculino , Placa Aterosclerótica , ARN Mensajero/análisis , Rotura Espontánea , Índice de Severidad de la Enfermedad , TranscriptomaRESUMEN
Despite potential clinical importance, target cells for mother-to-child transmission of HIV-1 have not yet been identified. Cord blood-derived CD4(+) T cells are largely naive and do not express CCR5, the mandatory coreceptor for transmitted HIV-1 R5 strains in infants. In the present study, we demonstrate that in the human fetal and infant gut mucosa, there is already a large subset of mucosal memory CD4(+)CCR5(+) T cells with predominantly a Th1 and Th17 phenotype. Using next-generation sequencing of the TCRß chain, clonally expanded T cells as a hallmark for memory development predominated in the gut mucosa (30%), whereas few were found in the lymph nodes (1%) and none in cord blood (0%). The gut mucosal fetal and infant CD4(+) T cells were highly susceptible to HIV-1 without any prestimulation; pol proviral DNA levels were similar to infected phytohemagglutinin-stimulated adult PBMCs. In conclusion, in the present study, we show that extensive adaptive immunity is present before birth and the gut mucosa is the preferential site for memory CD4(+) T cells. These CD4(+)CCR5(+) T cells in the infant mucosa provide a large pool of susceptible cells for ingested HIV-1 at birth and during breastfeeding, indicating a mucosal route of mother-to-child transmission that can be targeted in prevention strategies.
Asunto(s)
Linfocitos T CD4-Positivos/citología , Infecciones por VIH/transmisión , Memoria Inmunológica , Transmisión Vertical de Enfermedad Infecciosa , Intestinos/inmunología , Receptores CCR5/metabolismo , Adulto , Recuento de Linfocito CD4 , Linfocitos T CD4-Positivos/inmunología , Linfocitos T CD4-Positivos/metabolismo , Células Cultivadas , Femenino , Infecciones por VIH/inmunología , VIH-1/inmunología , Humanos , Memoria Inmunológica/inmunología , Memoria Inmunológica/fisiología , Recién Nacido , Mucosa Intestinal/citología , Mucosa Intestinal/inmunología , Mucosa Intestinal/metabolismo , Intestinos/citología , Intestinos/virología , Masculino , Relaciones Madre-Hijo , Embarazo , Complicaciones Infecciosas del Embarazo/inmunologíaRESUMEN
BACKGROUND: Areas of microvascular proliferation have been observed in a subpopulation of symptomatic congenital vascular malformations later in life. We investigated whether this angiogenic response is followed by a stage of maturation. METHODS: Resections of vascular malformations (n = 15), infantile hemangiomas (IHs) (n = 8) and pyogenic granulomas (PGs) (n = 5) were studied. Histopathologically, all lesions were screened for presence of foci of immature and/or mature microvessels. These areas were further studied immunohistochemically for differential expression of several angiogenic factors, cell cycle-dependent proteins, p53 and active caspase3. Immunostains were scored semiquantitatively. RESULTS: Immature microvessel areas were present in five vascular malformations (all of the arteriovenous type), five IHs and five PGs; these lesions also contained transitions between immature and mature microvessels. Conglomerates of mature microvessels were found in 19 cases (6 vascular malformations, 5 PGs and 8 IHs). Expression of vascular endothelial growth factor-A, angiopoietin-1, Ki-67, p16 and p21/27 ratios were overall significantly lower in mature areas than in immature areas including those in vascular malformations. P53 and caspase3 expression was scarce in all lesions. CONCLUSIONS: Microvascular areas in vascular malformations appear to follow the same pattern of vascular proliferation and maturation as seen in other microvascular lesions of skin and soft tissue.
Asunto(s)
Angiopoyetina 1/biosíntesis , Malformaciones Arteriovenosas , Proteínas de Ciclo Celular/biosíntesis , Ciclo Celular , Microvasos , Neovascularización Patológica , Piel , Factor A de Crecimiento Endotelial Vascular/biosíntesis , Malformaciones Arteriovenosas/metabolismo , Malformaciones Arteriovenosas/patología , Caspasa 3/metabolismo , Femenino , Granuloma Piogénico/metabolismo , Granuloma Piogénico/patología , Hemangioma/metabolismo , Hemangioma/patología , Humanos , Inmunohistoquímica , Masculino , Microvasos/metabolismo , Microvasos/patología , Neovascularización Patológica/metabolismo , Neovascularización Patológica/patología , Piel/irrigación sanguínea , Piel/metabolismo , Piel/patología , Proteína p53 Supresora de Tumor/metabolismoRESUMEN
BACKGROUND: Cerebral vascular malformations were investigated for the presence of the glucose transporter protein GLUT1, which is normally expressed in endothelial cells of the pre-existing microvasculature of the brain and absent in the vasculature of the choroid plexus and extracranial vasculature without a barrier function. Extracranial arteriovenous malformations (AVM) are known to show an absence of GLUT1 expression which distinguishes them from infantile hemangioma of skin and soft tissue. The expression of GLUT1 in cerebrovascular malformations is not systematically investigated. METHODS: Paraffin-embedded sections of cerebral AVM (4), including one choroid plexus AVM, cerebral cavernous malformations (CCM, 3) and extracranial (facial) AVM (3) were immunostained with anti-CD31 and GLUT1 in doublestaining procedure which was further analyzed with the use of spectral analysis software. RESULTS: All 7 cases of cerebral vascular malformations showed colocalization of GLUT1/CD31 of endothelial cells of the vessels within the malformation. Only in the extracranial AVM expression of GLUT1 was completely absent. CONCLUSION: Cerebral AVM differ from extracranial AVM by their endothelial immunoexpression of GLUT1, indicating that the vessels of these malformations retain the endothelial phenotype of the local vascular beds from which they are derived during embryogenesis.
Asunto(s)
Malformaciones Arteriovenosas/metabolismo , Transportador de Glucosa de Tipo 1/biosíntesis , Malformaciones Arteriovenosas Intracraneales/metabolismo , Adolescente , Adulto , Malformaciones Arteriovenosas/patología , Niño , Femenino , Hemangioma Cavernoso del Sistema Nervioso Central/metabolismo , Hemangioma Cavernoso del Sistema Nervioso Central/patología , Humanos , Inmunohistoquímica , Malformaciones Arteriovenosas Intracraneales/patología , MasculinoRESUMEN
AIMS: Vulnerable atherosclerotic plaques are lesions with a high propensity to develop plaque disruption and superimposed thrombosis. No systematic studies have been carried out on tissue markers for plaque vulnerability throughout the entire coronary artery system at the end stages of coronary atherosclerosis. METHODS AND RESULTS: Nine autopsied patients (mean age 77 years) with angiographically severe trivascular coronary atherosclerosis were selected for this study. All visible lesions in postmortem coronary angiograms (n = 125) were histologically and immunohistochemically screened for the presence of intraplaque haemorrhages (activated) microvessels and inflammatory infiltrates. Intraplaque haemorrhages were observed in 76/125 plaques (61%). Chronic inflammation was found superficially in 59/125 plaques (47%) and deeply inside the plaque tissue in 103/125 plaques (83%). Microvessels were found in 100/125 lesions (80%), of which 58% showed endothelial expression of the vascular activation marker CD105. Moreover, microvascular CD105 positivity correlated positively with plaque haemorrhage and deeply seated plaque inflammation. CONCLUSIONS: Plaque inflammation and haemorrhages can be found at a high frequency throughout the coronary artery system of elderly patients with multivessel coronary atherosclerosis. Microvascular expression of endoglin (CD105), which correlates positively with both of these features of plaque vulnerability, can serve as a marker of the risk of developing coronary thrombotic complications.
Asunto(s)
Antígenos CD/metabolismo , Enfermedad de la Arteria Coronaria/metabolismo , Vasos Coronarios/metabolismo , Microvasos/metabolismo , Placa Aterosclerótica/metabolismo , Receptores de Superficie Celular/metabolismo , Anciano , Anciano de 80 o más Años , Autopsia , Biomarcadores/metabolismo , Cadáver , Enfermedad Crónica , Angiografía Coronaria , Enfermedad de la Arteria Coronaria/complicaciones , Enfermedad de la Arteria Coronaria/patología , Vasos Coronarios/patología , Endoglina , Endotelio Vascular/metabolismo , Endotelio Vascular/patología , Femenino , Hemorragia/complicaciones , Hemorragia/patología , Humanos , Inflamación/complicaciones , Inflamación/metabolismo , Inflamación/patología , Masculino , Microvasos/patología , Persona de Mediana Edad , Placa Aterosclerótica/complicaciones , Placa Aterosclerótica/patologíaRESUMEN
OBJECTIVES: Repeated exposure to stress leads to mast cell degranulation, microscopic inflammation, and subsequent visceral hypersensitivity in animal models. To what extent this pathophysiological pathway has a role in patients with the irritable bowel syndrome (IBS) has not been properly investigated. The objective of this study was to assess the relationship between visceral hypersensitivity, microscopic inflammation, and the stress response in IBS. METHODS: Microscopic inflammation of the colonic mucosa was evaluated by immunohistochemistry in 66 IBS patients and 20 healthy volunteers (HV). Rectal sensitivity was assessed by a barostat study using an intermittent pressure-controlled distension protocol. Salivary cortisol to a psychological stress was measured to assess the stress response. RESULTS: Compared with HV, mast cells, T cells, and macrophages were decreased in IBS patients. Similarly, λ-free light chain (FLC)-positive mast cells were decreased but not immunoglobulin E (IgE)- and IgG-positive mast cells. There were no differences between hypersensitive and normosensitive IBS patients. No relation was found between any of the immune cells studied and the thresholds of discomfort, urge, first sensation, or IBS symptoms (e.g., abdominal pain, stool-related complaints, bloating). Finally, stress-related symptoms and the hypothalamic-pituitary-adrenal-axis response to stress were not correlated with the number of mast cells or the presence of visceral hypersensitivity. CONCLUSIONS: Although the number of mast cells, macrophages, T cells, and λFLC-positive mast cells is decreased in IBS compared with HV, this is not associated with the presence of visceral hypersensitivity or abnormal stress response. Our data question the role of microscopic inflammation as an underlying mechanism of visceral hypersensitivity, but rather suggest dysregulation of the mucosal immune system in IBS.
Asunto(s)
Mucosa Intestinal/inmunología , Síndrome del Colon Irritable/inmunología , Síndrome del Colon Irritable/fisiopatología , Recto/fisiopatología , Adulto , Biopsia con Aguja , Recuento de Células , Colon/inmunología , Colon/patología , Colon/fisiopatología , Colonoscopía , Femenino , Humanos , Hidrocortisona/sangre , Inmunohistoquímica , Mucosa Intestinal/patología , Síndrome del Colon Irritable/patología , Síndrome del Colon Irritable/psicología , Macrófagos/inmunología , Macrófagos/patología , Masculino , Mastocitos/inmunología , Mastocitos/patología , Persona de Mediana Edad , Presión , Umbral Sensorial , Estrés Psicológico/fisiopatología , Linfocitos T/inmunología , Linfocitos T/patología , Adulto JovenRESUMEN
Children with Down syndrome (DS) are at high risk for acute lung injury (ALI). Pulmonary epithelial apoptosis is an important factor in the pathophysiology of ALI. Whether the risk of ALI in DS is associated with a high level of pulmonary epithelial apoptosis is not known. We hypothesized that the percentage of apoptotic epithelial cells is higher in DS than in control lungs. Lung tissue sections from autopsies of 21 fetuses with DS and 12 controls were stained with antibodies against the epithelial marker pan-cytokeratin (CK) and apoptosis marker activated caspase-3 (aC3). Spectral imaging software was used to quantify the mean percentage of pixels that showed colocalization of CK and aC3. Mean (standard deviation [SD]) gestational age in weeks was 18.7 (1.4) in DS and 18.9 (2.0) in controls (P â=â 0.67). The mean (SD) percentage of CK-positive pixels was 27.2% (4.7%) in DS compared to 27.1% (6.2%) in controls (P â=â 0.97). The median (interquartile range [IQR]) percentage of CK-positive pixels that showed colocalization of aC3 was 0.16% (0.18%) in DS compared to 0.27% (0.24%) in controls (P â=â 0.45). The mean (SD) number of CK-positive pixels increased from 22.5% (5.2%) to 30.4% (4.6%) with the appearance of saccular morphology in controls but not in DS (P â=â 0.01). The percentage of apoptotic epithelial cells in DS fetal lungs does not differ from that in controls. However, we did find a difference in the development of epithelial structures between DS and controls that may be associated with anomalies in alveolar development found at birth in DS.
Asunto(s)
Apoptosis , Síndrome de Down/patología , Células Epiteliales/patología , Pulmón/patología , Lesión Pulmonar Aguda/etiología , Lesión Pulmonar Aguda/fisiopatología , Autopsia , Caspasa 3/análisis , Síndrome de Down/complicaciones , Síndrome de Down/metabolismo , Células Epiteliales/metabolismo , Feto , Humanos , Inmunohistoquímica , Queratinas/análisis , Pulmón/metabolismoRESUMEN
OBJECTIVE: Whether and how B lymphocytes contribute to the pathogenesis of spondylarthritis (SpA), a seronegative arthritis associated with gut inflammation, remains unknown. Because innate-like CD5+ B lymphocytes with regulatory functions have been identified in colitis models, we undertook the present study to analyze the presence and function of CD5+ B cells in human SpA. METHODS: Peripheral blood B cells from patients with SpA, patients with rheumatoid arthritis (RA), and healthy controls were analyzed by flow cytometry. Synovial biopsy samples were evaluated by immunohistochemistry analysis. Sorted CD5+ and CD5- B cells were analyzed for somatic hypermutation, expression of costimulatory molecules, and cytokine production. RESULTS: The naive, marginal zone-like, and to a lesser extent memory B cell compartments in patients with SpA exhibited a clear and specific increase of CD5+ B cells, which was not found in patients with RA. This increase was not due to either B cell activation or preferential migration of CD5- B cells to the inflamed synovium. Consistent with their phenotype and the low-affinity polyreactive immunoglobulins produced by their murine counterpart cells, CD5+ B cells from patients with SpA showed low levels of somatic hypermutation. With regard to antigen presentation, CD5+ B cells expressed slightly increased HLA-DR levels but low CD80 and CD86 levels. In vitro activation failed to up-regulate these costimulatory molecules but induced significant production of interleukin-10 and interleukin-6 by CD5+ B cells. CONCLUSION: CD5+ B cells are specifically increased in SpA. Analysis of somatic hypermutation, expression of antigen-presenting and costimulatory molecules, and cytokine production indicates that this B cell subset has regulatory capacities. Further investigation of the potential role of CD5+ cells in SpA is warranted.
Asunto(s)
Linfocitos B Reguladores/inmunología , Antígenos CD5/metabolismo , Espondiloartritis/inmunología , Adulto , Artritis Reumatoide/inmunología , Artritis Reumatoide/metabolismo , Linfocitos B Reguladores/metabolismo , Femenino , Citometría de Flujo , Humanos , Interleucina-10/metabolismo , Interleucina-6/metabolismo , Masculino , Persona de Mediana Edad , Espondiloartritis/metabolismo , Membrana Sinovial/inmunología , Membrana Sinovial/metabolismoRESUMEN
Prolactin is best known as the polypeptide anterior pituitary hormone, which regulates the development of the mammary gland. However, it became clear over the last decade that prolactin contributes to a broad range of pathologies, including breast cancer. Prolactin is also involved in angiogenesis via the release of pro-angiogenic factors by leukocytes and epithelial cells. However, whether prolactin also influences endothelial cells, and whether there are functional consequences of prolactin-induced signalling in the perspective of angiogenesis, remains so far elusive. In the present study, we show that prolactin induces phosphorylation of ERK1/2 and STAT5 and induces tube formation of endothelial cells on Matrigel. These effects are blocked by a specific prolactin receptor antagonist, del1-9-G129R-hPRL. Moreover, in an in vivo model of the chorioallantoic membrane of the chicken embryo, prolactin enhances vessel density and the tortuosity of the vasculature and pillar formation, which are hallmarks of intussusceptive angiogenesis. Interestingly, while prolactin has only little effect on endothelial cell proliferation, it markedly stimulates endothelial cell migration. Again, migration was reverted by del1-9-G129R-hPRL, indicating a direct effect of prolactin on its receptor. Immunohistochemistry and spectral imaging revealed that the prolactin receptor is present in the microvasculature of human breast carcinoma tissue. Altogether, these results suggest that prolactin may directly stimulate angiogenesis, which could be one of the mechanisms by which prolactin contributes to breast cancer progression, thereby providing a potential tool for intervention.
Asunto(s)
Células Endoteliales/patología , Neovascularización Patológica/patología , Prolactina/efectos adversos , Transducción de Señal/efectos de los fármacos , Inductores de la Angiogénesis/efectos adversos , Animales , Neoplasias de la Mama/patología , Línea Celular , Embrión de Pollo , Colágeno/metabolismo , Combinación de Medicamentos , Células Endoteliales/metabolismo , Femenino , Inmunohistoquímica , Laminina/metabolismo , Sistema de Señalización de MAP Quinasas/efectos de los fármacos , Ratones , Fosforilación , Proteoglicanos/metabolismo , Receptores de Prolactina/antagonistas & inhibidores , Receptores de Prolactina/metabolismo , Factor de Transcripción STAT5/genética , Factor de Transcripción STAT5/metabolismoRESUMEN
Gene signatures derived from cancer stem cells (CSCs) predict tumor recurrence for many forms of cancer. Here, we derived a gene signature for colorectal CSCs defined by high Wnt signaling activity, which in agreement with previous observations predicts poor prognosis. Surprisingly, however, we found that elevated expression of Wnt targets was actually associated with good prognosis, while patient tumors with low expression of Wnt target genes segregated with immature stem cell signatures. We discovered that several Wnt target genes, including ASCL2 and LGR5, become silenced by CpG island methylation during progression of tumorigenesis, and that their re-expression was associated with reduced tumor growth. Taken together, our data show that promoter methylation of Wnt target genes is a strong predictor for recurrence of colorectal cancer, and suggest that CSC gene signatures, rather than reflecting CSC numbers, may reflect differentiation status of the malignant tissue.