RESUMEN
Solid pseudopapillary tumor (SPT) of the pancreas is an uncommon neoplasm of uncertain lineage. They have been shown to express nuclear beta-catenin believed to be due to mutations of the beta-catenin gene. The aim of this study was to investigate the status of the E-cadherin/catenin complex in SPTs. We studied the expression of 4 principal members of the E-cadherin/catenin complex using immunohistochemistry and the E-cadherin gene status by screening all exons of the gene for mutations, in 6 cases of SPT. In addition to the nuclear localization of beta-catenin, we found nuclear localization of E-cadherin in all tumors with complete absence of membranous and cytoplasmic localization. Nuclear localization of E-cadherin was independent of beta-catenin. No mutations were identified in the E-cadherin gene in any of the tumors. Ten cases of pancreatic adenocarcinomas and 15 neuroendocrine tumors were studied as well for comparison. The reported changes in the expression of the principal members of the E-cadherin/catenin complex were unique to SPTs. Our study shows abnormalities in the expression of 4 principal members of the E-cadherin/catenin complex in SPTs, which may help to explain the discohesive nature of the cells and the cystic changes in these tumors, and provide additional diagnostic features.
Asunto(s)
Cadherinas/metabolismo , Carcinoma Papilar/metabolismo , Neoplasias Pancreáticas/metabolismo , beta Catenina/metabolismo , Adulto , Cadherinas/genética , Carcinoma Papilar/genética , Análisis Mutacional de ADN , Femenino , Humanos , Inmunohistoquímica , Masculino , Persona de Mediana Edad , Neoplasias Pancreáticas/genética , Reacción en Cadena de la PolimerasaRESUMEN
Matrix metalloproteinase-9 (MMP-9) activity is implicated in pathogenesis of central nervous system tuberculosis (CNS-TB). IFNgamma, a key cytokine in TB, usually inhibits MMP-9 secretion. Addition of IFNgamma to conditioned media from M. tb-infected monocytes (CoMTB) resulted in a 7-fold increase in MMP-9 activity detected by gelatin zymography (P<0.01). In contrast, tissue inhibitor of metalloproteinase (TIMP)-1 and -2 secretion, measured by ELISA, was suppressed. Dexamethasone abolished the synergistic increase in MMP-9 activity. Interleukin (IL)-1beta in CoMTB is a critical mediator of synergy with IFNgamma, and IL-1beta alone synergizes with IFNgamma to increase MMP-9 secretion from 51 +/- 31 to 762 +/- 136 U. IL-1beta activity is dependent on p38 mitogen-activated protein (MAPK) kinase, which was found to be phosphorylated in tissue specimens from patients with CNS-TB. Extracellular signal regulated kinase (Erk) and p38 MAPK activation did not affect IFNgamma signaling pathways. Inhibition of janus-activated kinase (JAK)-2 by 50 microM AG540 decreased MMP-9 secretion to 124 +/- 11.1 from 651 +/- 229 U of activity (P<0.01). However, signal transducer and activator of transcription (STAT)-3 but not STAT-1 phosphorylation was synergistically up-regulated by IFNgamma and CoMTB. In summary, synergy between IL-1beta and STAT-3 dependent IFNgamma signaling is key in control of up-regulation of MMP-9 activity in CNS-TB and may be a significant mechanism of brain tissue destruction.
Asunto(s)
Interferón gamma/farmacología , Interleucina-1beta/farmacología , Metaloproteinasa 9 de la Matriz/metabolismo , Mycobacterium tuberculosis/crecimiento & desarrollo , Western Blotting , Línea Celular Tumoral , Medios de Cultivo Condicionados/metabolismo , Dexametasona/farmacología , Sinergismo Farmacológico , Ensayo de Inmunoadsorción Enzimática , Quinasas MAP Reguladas por Señal Extracelular/metabolismo , Humanos , Inmunohistoquímica , Quinasas Janus/metabolismo , Monocitos/efectos de los fármacos , Monocitos/metabolismo , Monocitos/microbiología , Fosforilación , Inhibidor Tisular de Metaloproteinasa-1/metabolismo , Inhibidor Tisular de Metaloproteinasa-2/metabolismo , Tuberculosis del Sistema Nervioso Central/metabolismo , Regulación hacia Arriba/efectos de los fármacos , Proteínas Quinasas p38 Activadas por Mitógenos/metabolismoRESUMEN
Bcl-6 is expressed in germinal centre derived B-cell non-Hodgkin lymphomas including diffuse large B-cell lymphoma (DLBCL) and is likely to play a major role in driving proliferation of a subset of DLBCLs, especially those of germinal centre B-cell subtype, but the role of c-Myc, which is important for proliferation in various lineages is not known. We used the highly standardised staining conditions of a tissue microarray to characterise co-expression of c-Myc and Bcl-6 in DLBCL. We carried out immunohistochemistry of 73 arrayed cases. The majority (62/73) did not express c-Myc, but 11 cases (15%) showed nuclear staining. 5/53 (9%) of Bcl-6 expressing cases co-expressed c-Myc, whereas a much higher proportion, 6/20 (30%), of Bcl-6 negative cases were positive for c-Myc. Overall survival of c-Myc expressing cases was the same as those that had absent expression. There was no significant correlation between c-Myc expression and DLBCL subtype (germinal centre or non-germinal centre subtypes).
Asunto(s)
Centro Germinal/metabolismo , Linfoma de Células B/metabolismo , Linfoma de Células B Grandes Difuso/metabolismo , Proteínas Proto-Oncogénicas c-bcl-6/metabolismo , Proteínas Proto-Oncogénicas c-myc/metabolismo , Adulto , Centro Germinal/patología , Humanos , Técnicas para Inmunoenzimas , Linfoma de Células B/patología , Linfoma de Células B Grandes Difuso/patología , Análisis de Matrices TisularesAsunto(s)
Antígenos CD20/metabolismo , Enfermedad de Castleman/patología , Infecciones por Herpesviridae/patología , Herpesvirus Humano 8/aislamiento & purificación , Linfocitos/patología , Antígenos Virales/metabolismo , Enfermedad de Castleman/metabolismo , Enfermedad de Castleman/virología , Infecciones por Herpesviridae/complicaciones , Infecciones por Herpesviridae/metabolismo , Herpesvirus Humano 8/fisiología , Humanos , Ganglios Linfáticos/metabolismo , Ganglios Linfáticos/patología , Ganglios Linfáticos/virología , Linfocitos/metabolismo , Linfocitos/virología , Proteínas Nucleares/metabolismo , Fosfoproteínas/metabolismo , Bazo/metabolismo , Bazo/patología , Bazo/virologíaRESUMEN
PURPOSE: The expression of epidermal growth factor receptor (EGFR) and human epidermal growth factor receptor 2 (HER2) is associated with poor prognosis in sporadic colorectal carcinoma (CRC). EGFR inhibitors are approved for the treatment of refractory CRC. The aim of this study was to investigate the expression of EGFR and HER2 and downstream extracellular signal regulated kinase (ERK) and mitogen activated protein kinase (MAPK) in non-neoplastic colonic mucosa, adenomas and carcinomas from familial adenomatous polyposis coli (FAP) patients, exploring the expression along the adenoma-carcinoma sequence. METHODS: The expression of EGFR, HER2, phosphorylated MAPK/ERK kinase (pMEK) and phosphorylated ERK (pERK) proteins was studied by immunohistochemistry in samples of colonic non-neoplastic mucosa (n = 65), adenomas (n = 149) and adenocarcinomas (n = 16) from each of the 16 FAP patients. RESULTS: For HER2, only weak cytoplasmic expression was seen in 8% of adenomas, 6% of carcinomas and 3% of the non-neoplastic mucosa. EGFR was expressed in non-neoplastic mucosa, adenomas and carcinomas with a statistically significant increase in expression in adenomas compared with non-neoplastic mucosa (p < 0.001). There was also a statistically significant increase in nuclear staining intensity for pERK (p < 0.001) and pMEK (p < 0.001) in adenomas compared to non-neoplastic mucosa. CONCLUSIONS: This is the first study investigating the expression of these receptors in non-neoplastic mucosa, adenomas and carcinomas from FAP patients. HER2 is not upregulated in the tumours of FAP patients, while EGFR appears to be upregulated in most adenomas and carcinomas, with associated upregulation of pERK and pMEK. We conclude that EGFR and downstream members of its signalling pathway, but not HER2, may be potential therapeutic targets in FAP patients.
Asunto(s)
Poliposis Adenomatosa del Colon/metabolismo , Neoplasias del Colon/metabolismo , Receptores ErbB/metabolismo , Quinasas MAP Reguladas por Señal Extracelular/metabolismo , Proteínas Quinasas Activadas por Mitógenos/metabolismo , Receptor ErbB-2/metabolismo , Adenocarcinoma/metabolismo , Adenocarcinoma/patología , Adenoma/metabolismo , Adenoma/patología , Poliposis Adenomatosa del Colon/patología , Biomarcadores de Tumor/metabolismo , Neoplasias del Colon/patología , Humanos , Técnicas para Inmunoenzimas , Mucosa Intestinal/metabolismo , Mucosa Intestinal/patología , Clasificación del Tumor , Fosforilación , Transducción de SeñalRESUMEN
CXCL8 is a chemokine that is implicated in the formation of tuberculous (TB) granulomas and in immunity to Mycobacterium tuberculosis (Mtb). Fibroblast chemokine secretion is important for modulating inflammatory responses in chronic lung disease and inflammatory arthritis but has not been investigated in the pathophysiology of TB. In this study, we used a cellular model to examine monocyte/macrophage-dependent stimulation of fibroblasts by Mtb in the regulation of chemokine secretion, particularly that of CXCL8. Human lung fibroblasts grown in collagen were stimulated with conditioned medium from Mtb-infected monocytes (CoMTb). CoMTb-induced prolonged dose-dependent, p38-mediated expression of stable CXCL8 mRNA by fibroblasts accompanied by a >10-fold increase in CXCL8 secretion (487 +/- 88 ng/ml vs 48.6 +/- 34 ng/ml in controls) at 120 h. Fibroblasts strongly expressed CXCL8 in vivo in human TB granulomas. Inhibition of TNF-alpha or IL-1 in CoMTb abrogated the induction of CXCL8 at a pretranscriptional level. CXCL8 secretion was NF-kappaB, C/EBP, and JNK dependent. Sustained NF-kappaB activation was demonstrated beyond 24 h in response to CoMTb. Exogenous CXCL8 reduced the survival of Mtb within macrophages, and inhibition of CXCL8 was associated with intracellular mycobacterial proliferation. These data show that fibroblasts have a previously unrecognized role in modulating inflammation in TB by their CXCL8-dependent contribution to cell recruitment and mycobacterial killing within the granuloma.
Asunto(s)
Fibroblastos/metabolismo , Interleucina-8/metabolismo , Macrófagos/metabolismo , Mycobacterium tuberculosis/metabolismo , Tuberculoma/metabolismo , Tuberculosis Pulmonar/metabolismo , Proteínas Potenciadoras de Unión a CCAAT/inmunología , Proteínas Potenciadoras de Unión a CCAAT/metabolismo , Células Cultivadas , Fibroblastos/inmunología , Fibroblastos/microbiología , Humanos , Interleucina-1/inmunología , Interleucina-1/metabolismo , MAP Quinasa Quinasa 4/inmunología , MAP Quinasa Quinasa 4/metabolismo , Macrófagos/inmunología , Macrófagos/microbiología , Viabilidad Microbiana/inmunología , Mycobacterium tuberculosis/inmunología , FN-kappa B/inmunología , FN-kappa B/metabolismo , ARN Mensajero/biosíntesis , ARN Mensajero/inmunología , Tuberculoma/inmunología , Tuberculoma/fisiopatología , Tuberculosis Pulmonar/inmunología , Factor de Necrosis Tumoral alfa/inmunología , Factor de Necrosis Tumoral alfa/metabolismoRESUMEN
CNS tuberculosis (CNS-TB) is the most deadly form of tuberculous disease accounting for 10% of clinical cases. CNS-TB is characterized by extensive tissue destruction, in which matrix metalloproteinases (MMPs) may play a critical role. We investigated the hypothesis that Mycobacterium tuberculosis activates monocyte-astrocyte networks increasing the activity of key MMPs. We examined the expression of all human MMPs and the tissue inhibitors of metalloproteinases (TIMPs) in human astrocytes stimulated by conditioned medium from M. tuberculosis-infected monocytes (CoMTB). Real-time RT-PCR showed that gene expression of MMP-1, -2, -3, -7, and -9 was increased (p < 0.05). MMP-9 secretion was significantly up-regulated at 24 h and increased over 120 h (p < 0.01). MMP-1, -3, and -7 secretion was not detected. Secretion of MMP-2 was constitutive and unaffected by CoMTB. Astrocyte gene expression and secretion of TIMP-1 was not affected by CoMTB although TIMP-2 secretion increased 3-fold at 120 h. Immunohistochemical analysis of human brain biopsies confirmed that astrocyte MMP-9 secretion is a predominant feature in CNS-TB in vivo. Dexamethasone inhibited astrocyte MMP-9, but not TIMP-1/2 secretion in response to CoMTB. CoMTB stimulated the nuclear translocation of NF-kappaB, inducing a 6-fold increase in nuclear p65 and a 2-fold increase in nuclear p50. This was associated with degradation of IkappaBalpha and beta within 30 min, persisting for 24 h. In summary, networks active between monocytes and astrocytes regulate MMP-9 activity in tuberculosis and astrocytes are a major source of MMP-9 in CNS-TB. Astrocytes may contribute to a matrix degrading environment within the CNS and subsequent morbidity and mortality.
Asunto(s)
Astrocitos/enzimología , Astrocitos/inmunología , Metaloproteinasas de la Matriz/genética , Monocitos/enzimología , Monocitos/inmunología , Tuberculosis del Sistema Nervioso Central/enzimología , Tuberculosis del Sistema Nervioso Central/inmunología , Astrocitos/metabolismo , Comunicación Celular , Células Cultivadas , Dexametasona/farmacología , Expresión Génica , Regulación Enzimológica de la Expresión Génica , Granuloma/enzimología , Humanos , Metaloproteinasas de la Matriz/metabolismo , Monocitos/efectos de los fármacos , FN-kappa B/metabolismo , Inhibidores Tisulares de Metaloproteinasas/metabolismo , Tuberculosis del Sistema Nervioso Central/patología , Regulación hacia ArribaRESUMEN
An increase in leucocyte apoptosis and impaired clearance of apoptotic cells has been observed in patients with systemic lupus erythematosus (SLE). Apoptotic cells are likely to be a key source of autoantigens in SLE as they express many of the nuclear autoantigens (in surface blebs and apoptotic bodies) that are relevant to this disease. The clearance of apoptotic cells is usually a rapid process, such that few cells are usually seen in the extracellular environment in vivo. We report a case in which multiple apoptotic bodies were observed in the bone marrow of a patient with SLE that was complicated by an immune-mediated pancytopenia. We have subsequently examined the frequency of apoptotic cells, identified morphologically, and by caspase-3 staining in bone-marrow trephine samples taken from patients with SLE over a 10-year period of follow-up. A high proportion of bone marrows contained apoptotic debris. The novel demonstration of apoptotic bodies in vivo in patients with SLE is unusual and supports the notion that the marrow may be a target organ in the disease. Their abundance is also consistent with the hypothesis that normal clearance mechanisms are defective and/or overwhelmed in SLE.
Asunto(s)
Apoptosis , Autoinmunidad , Médula Ósea/patología , Leucocitos/patología , Lupus Eritematoso Sistémico/inmunología , Lupus Eritematoso Sistémico/patología , Adulto , Caspasa 3/análisis , Femenino , Factor Estimulante de Colonias de Granulocitos/uso terapéutico , Humanos , Factores Inmunológicos/uso terapéutico , Inmunosupresores/uso terapéutico , Masculino , Metilprednisolona/uso terapéutico , Persona de Mediana Edad , Ácido Micofenólico/análogos & derivados , Ácido Micofenólico/uso terapéutico , Pancitopenia/inmunología , Pancitopenia/patología , Coloración y EtiquetadoRESUMEN
The minichromosome maintenance (Mcm) and Cdc6 proteins are important regulators of eucaryotic DNA replication. In most normal tissues, a similar proportion of cells express Mcm-2 and Ki-67. The present study showed that in both normal and abnormal states, the proportion of megakaryocytes expressing Mcm-2 is roughly seven times as many as those that express Ki-67. This is likely to be related to the process of endomitosis and endoreduplication. We also demonstrated that a significantly lower proportion of megakaryocytes in myelodysplastic syndrome express Mcm-2. These findings provide new insights into megakaryocyte biology.
Asunto(s)
Proteínas de Ciclo Celular/metabolismo , Antígeno Ki-67/metabolismo , Megacariocitos/metabolismo , Síndromes Mielodisplásicos/metabolismo , Trastornos Mieloproliferativos/metabolismo , Proteínas Nucleares/metabolismo , Médula Ósea/metabolismo , Núcleo Celular/metabolismo , Citoplasma/metabolismo , Células Precursoras Eritroides/metabolismo , Humanos , Integrina beta3/metabolismo , Componente 2 del Complejo de Mantenimiento de Minicromosoma , Síndromes Mielodisplásicos/patología , Trastornos Mieloproliferativos/patologíaRESUMEN
Pulmonary cavitation is vital to the persistence and spread of Mycobacterium tuberculosis (MTb), but mechanisms underlying this lung destruction are poorly understood. Fibrillar type I collagen provides the lung's tensile strength, and only matrix metalloproteinases (MMPs) can degrade it at neutral pH. We investigated MTb-infected lung tissue and found that airway epithelial cells adjacent to tuberculosis (Tb) granulomas expressed a high level of MMP-1 (interstitial collagenase). Conditioned media from MTb-infected monocytes (CoMTb) up-regulated epithelial cell MMP-1 promoter activity, gene expression, and secretion, whereas direct MTb infection did not. CoMTb concurrently suppressed tissue inhibitor of metalloprotease-1 (TIMP-1) secretion, further promoting matrix degradation, and in Tb patients very low TIMP-1 expression was detected. MMP-1 up-regulation required synergy between TNF-alpha and G protein-coupled receptor signaling pathways. CoMTb stimulated p38 MAPK phosphorylation, and this is the point of TNF-alpha synergy with G protein-coupled receptor activation. Furthermore, p38 phosphorylation was the switch up-regulating MMP-1 activity and decreasing TIMP-1 secretion. Activated p38 localized to MMP-1-secreting airway epithelial cells in Tb patients. These data reveal a monocyte-epithelial cell network whereby MTb may drive tissue destruction, and they demonstrate that p38 phosphorylation is a key regulatory point in the generation of a matrix-degrading phenotype.
Asunto(s)
Células Epiteliales/enzimología , Metaloproteinasa 1 de la Matriz/metabolismo , Mycobacterium tuberculosis/fisiología , Mucosa Respiratoria/enzimología , Tuberculosis Pulmonar/enzimología , Regulación hacia Arriba/fisiología , Proteínas Quinasas p38 Activadas por Mitógenos/fisiología , Bronquios/enzimología , Bronquios/metabolismo , Bronquios/microbiología , Células Cultivadas , Medios de Cultivo Condicionados , Células Epiteliales/metabolismo , Células Epiteliales/microbiología , Humanos , Ligandos , Sistema de Señalización de MAP Quinasas/fisiología , Monocitos/fisiología , Fosforilación , Receptores Acoplados a Proteínas G/fisiología , Mucosa Respiratoria/metabolismo , Mucosa Respiratoria/microbiología , Inhibidor Tisular de Metaloproteinasa-1/metabolismo , Factor de Necrosis Tumoral alfa/fisiologíaRESUMEN
RATIONALE: Pulmonary cavitation is fundamental to the global success of Mycobacterium tuberculosis. However, the mechanisms of this lung destruction are poorly understood. The biochemistry of lung matrix predicts matrix metalloproteinase (MMP) involvement in immunopathology. METHODS: We investigated gene expression of all MMPs, proteins with a disintegrin and metalloproteinase domain, and tissue inhibitors of metalloproteinases in M. tuberculosis-infected human macrophages by real-time polymerase chain reaction. MMP secretion was measured by zymography and Western analysis, and expression in patients with pulmonary tuberculosis was localized by immunohistochemistry. RESULTS: MMP-1 and MMP-7 gene expression and secretion are potently upregulated by M. tuberculosis, and no increase in tissue inhibitor of metalloproteinase expression occurs to oppose their activity. Dexamethasone completely suppresses MMP-1 but not MMP-7 gene expression and secretion. In patients with active tuberculosis, macrophages express MMP-1 and MMP-7 adjacent to areas of tissue destruction. MMP-1 but not MMP-7 expression and secretion are relatively M. tuberculosis specific, are not upregulated by tuberculosis-associated cytokines, and are prostaglandin dependent. In contrast, the vaccine M. bovis bacillus Calmette-Guérin (BCG) does not stimulate MMP-1 secretion from human macrophages, although M. tuberculosis and BCG do upregulate MMP-7 equally. BCG-infected macrophages secrete reduced prostaglandin E2 concentrations compared with M. tuberculosis-infected macrophages, and prostaglandin pathway supplementation augments MMP-1 secretion from BCG-infected cells. CONCLUSIONS: M. tuberculosis specifically upregulates MMP-1 in a cellular model of human infection and in patients with tuberculosis. In contrast, vaccine BCG, which does not cause lung cavitation, does not upregulate prostaglandin E2-dependent MMP-1 secretion.