RESUMEN

The transcription factor YBX1 can act as a mediator of signals transmitted via the EGFR-RAS-MAPK axis. YBX1 expression has been associated with tumor progression and prognosis in multiple types of cancer. Immunohistochemical studies have revealed dependency between YBX1 expression and individual EGFR family members. We analyzed YBX1 and EGFR family proteins in a colorectal cancer (CRC) cohort and provide functional analyses of YBX1 in the context of EGFR-RAS-MAPK signaling. Immunohistochemistry for YBX1 and EGFR family receptors with two antibodies for YBX1 and EGFR were performed and related to clinicopathological data. We employed Caco2 cells expressing an inducible KRASV12 gene to determine effects on localization and levels of YBX1. Mouse xenografts of Caco2-KRASV12 cells were used to determine YBX1 dynamics in a tissue context. The two different antibodies against YBX1 showed discordant immunohistochemical stainings in cell culture and clinical specimens. Expression of YBX1 and EGFR family members were not correlated in CRC. Analysis of Caco2 xenografts displayed again heterogeneity of YBX1 staining with both antibodies. Our results suggest that YBX1 is controlled via complex regulatory mechanisms involving tumor stroma interaction and signal transduction processes. Our study highlights that YBX1 antibodies have different specificities, advocating their use in a combined manner.

RESUMEN

Eight cases of psittacosis due to Chlamydia psittaci were identified in May 2013 among 15 individuals involved in chicken gutting activities on a mixed poultry farm in France. All cases were women between 42 and 67 years-old. Cases were diagnosed by serology and PCR of respiratory samples. Appropriate treatment was immediately administered to the eight hospitalised individuals after exposure to birds had been discovered. In the chicken flocks, mainly C. gallinacea was detected, a new member of the family Chlamydiaceae, whereas the ducks were found to harbour predominantly C. psittaci, the classical agent of psittacosis. In addition, C. psittaci was found in the same flock as the chickens that the patients had slaughtered. Both human and C. psittaci-positive avian samples carried the same ompA genotype E/B of C. psittaci, which is widespread among French duck flocks. Repeated grassland rotations between duck and chicken flocks on the farm may explain the presence of C. psittaci in the chickens. Inspection by the veterinary service led to temporary closure of the farm. All birds had to be euthanised on site as no slaughterhouses accepted processing them. Farm buildings and grasslands were cleaned and/or disinfected before the introduction of new poultry birds.

Asunto(s)

Chlamydophila psittaci/aislamiento & purificación , Brotes de Enfermedades , Exposición Profesional , Enfermedades de las Aves de Corral/microbiología , Psitacosis/diagnóstico , Psitacosis/epidemiología , Adulto , Animales , Pollos/microbiología , Chlamydophila psittaci/genética , Femenino , Francia/epidemiología , Genotipo , Humanos , Persona de Mediana Edad , Aves de Corral , Enfermedades de las Aves de Corral/epidemiología , Enfermedades de las Aves de Corral/transmisión , Reacción en Cadena en Tiempo Real de la Polimerasa

RESUMEN

The human Y-box binding protein, YB-1, is a multifunctional protein that regulates gene expression. Nuclear expression of YB-1 has been associated with chemoresistance and poor prognosis of tumour patients. Representative samples from autopsied material of primary tumours from 77 patients with NSCLC were investigated by immunohistochemistry for subcellular distribution of YB-1 and p53, in order to evaluate the prognostic role of nuclear expression of YB-1. Cytoplasmic YB-1 expression was found in all tumour samples, whereas nuclear expression was only observed in 48%. There was no correlation with histological classification, clinical parameters or tumour size, stage and metastasis status. However, patients with positive nuclear YB-1 expression in tumours showed reduced survival times when compared with patients without nuclear expression. Including information about the histology and mutational status for p53 increased the prognostic value of nuclear YB-1. Patients with nuclear YB-1 expression and p53 mutations had the worst prognosis (median survival 3 months), while best outcome was found in patients with no nuclear YB-1 and wildtype p53 (median survival 15 months). This suggests that the combined analysis of both markers allows a better identification of subgroups with varying prognosis. Nuclear expression of Y-box binding protien seems to be an independent prognostic marker.

Asunto(s)

Biomarcadores de Tumor/análisis , Proteínas Potenciadoras de Unión a CCAAT/análisis , Carcinoma de Pulmón de Células no Pequeñas/mortalidad , Núcleo Celular/química , Proteínas de Unión al ADN , Neoplasias Pulmonares/mortalidad , Factores de Transcripción , Anciano , Anciano de 80 o más Años , Carcinoma de Pulmón de Células no Pequeñas/química , Citoplasma/química , Femenino , Estudios de Seguimiento , Humanos , Inmunohistoquímica , Neoplasias Pulmonares/química , Masculino , Persona de Mediana Edad , Mutación , Factores de Transcripción NFI , Proteínas Nucleares , Pronóstico , Proteína p53 Supresora de Tumor/análisis , Proteína 1 de Unión a la Caja Y

RESUMEN

Genotoxic stress leads to nuclear translocation of the Y-box transcription factor YB-1 and enhanced expression of the multidrug resistance gene MDR1. Because hyperthermia is used for the treatment of colon cancer in combination with chemoradiotherapy, we investigated the influence of hyperthermia on YB-1 activity and the expression of multidrug resistance-related genes. Here we report that hyperthermia causes YB-1 translocation from the cytoplasm into the nucleus of human colon carcinoma cells HCT15 and HCT116. Nuclear translocation of YB-1 was associated with increased MDR1 and MRP1 gene activity, which is reflected in strong efflux pump activity. However, a combination of hyperthermia and drug treatment effectively reduced cell survival of the HCT15 and HCT116 cells. These results demonstrate that activation of MDR1 and MRP1 gene expression and increased efflux pump activity after hyperthermia were insufficient to cause an increase in drug resistance in colon cancer cell lines. The ability of hyperthermia to abrogate drug resistance in the presence of an increase in functional MDR proteins may provide an explanation for the efficacious results seen in the clinic in colon cancer patients treated with a combination of hyperthermia and chemotherapy.

Asunto(s)

Miembro 1 de la Subfamilia B de Casetes de Unión a ATP/biosíntesis , Transporte Activo de Núcleo Celular , Proteínas Potenciadoras de Unión a CCAAT/metabolismo , Proteínas de Unión al ADN , Fiebre , Factores de Transcripción , Transportadoras de Casetes de Unión a ATP/biosíntesis , Antineoplásicos/farmacología , Línea Celular , Núcleo Celular/metabolismo , Supervivencia Celular/efectos de los fármacos , Cloranfenicol O-Acetiltransferasa/metabolismo , Neoplasias del Colon/tratamiento farmacológico , Neoplasias del Colon/metabolismo , Citoplasma/metabolismo , ADN Complementario/metabolismo , Doxorrubicina/farmacología , Citometría de Flujo , Técnica del Anticuerpo Fluorescente Indirecta , Genes Reporteros , Humanos , Microscopía Confocal , Microscopía Fluorescente , Proteínas Asociadas a Resistencia a Múltiples Medicamentos , Factores de Transcripción NFI , Proteínas Nucleares , Plásmidos/metabolismo , ARN/metabolismo , Reacción en Cadena de la Polimerasa de Transcriptasa Inversa , Rodaminas/metabolismo , Temperatura , Factores de Tiempo , Transfección , Células Tumorales Cultivadas , Proteína 1 de Unión a la Caja Y

RESUMEN

Eight dogs with generalized demodicosis and five with sarcoptic manage were treated with 1.25% amitraz solution applied weekly and associated with an antidote treatment (atipamezol, 0.1 mg kg-1 i.m. once: and yohimbine 0.1 mg kg-1 once daily for 3 days, orally). Results of skin scrapings were used to determine whether therapy should be continued or stopped. The median number of treatments for demodicosis and sarcoptic mange was three (range 2-5) and two (range 1-3), respectively. Some side-effects were observed but all were stopped with antidote treatment; no failure or relapses occurred at 6-36 months after treatment.

Asunto(s)

Agonistas alfa-Adrenérgicos/uso terapéutico , Enfermedades de los Perros/tratamiento farmacológico , Insecticidas/uso terapéutico , Enfermedades Cutáneas Parasitarias/veterinaria , Toluidinas/uso terapéutico , Administración Oral , Administración Tópica , Agonistas alfa-Adrenérgicos/administración & dosificación , Antagonistas Adrenérgicos alfa/administración & dosificación , Antagonistas Adrenérgicos alfa/uso terapéutico , Animales , Antiparasitarios , Perros , Femenino , Imidazoles/administración & dosificación , Imidazoles/uso terapéutico , Inyecciones Intramusculares/veterinaria , Insecticidas/administración & dosificación , Masculino , Escabiosis/tratamiento farmacológico , Escabiosis/veterinaria , Toluidinas/administración & dosificación , Resultado del Tratamiento , Yohimbina/administración & dosificación , Yohimbina/uso terapéutico

Asunto(s)

Ciclo Celular/fisiología , Premio Nobel , Historia del Siglo XXI , Reino Unido , Estados Unidos

RESUMEN

Regulated mRNA turnover is a highly important process, but its mechanism is poorly understood. Using interleukin-2 (IL-2) mRNA as a model, we described a role for the JNK-signaling pathway in stabilization of IL-2 mRNA during T-cell activation, acting via a JNK response element (JRE) in the 5' untranslated region (UTR). We have now identified two major RNA-binding proteins, nucleolin and YB-1, that specifically bind to the JRE. Binding of both proteins is required for IL-2 mRNA stabilization induced by T-cell activation signals and for JNK-induced stabilization in a cell-free system that duplicates essential features of regulated mRNA decay. Nucleolin and YB-1 are required for formation of an IL-2 mRNP complex that responds to specific mRNA stabilizing signals.

Asunto(s)

Proteínas Potenciadoras de Unión a CCAAT , Proteínas de Unión al ADN/metabolismo , Interleucina-2/genética , Activación de Linfocitos , Proteínas Quinasas Activadas por Mitógenos/metabolismo , Fosfoproteínas/metabolismo , Estabilidad del ARN , Proteínas de Unión al ARN/metabolismo , Linfocitos T/metabolismo , Factores de Transcripción , Secuencia de Aminoácidos , Secuencia de Bases , Citoplasma/química , Proteínas de Unión al ADN/química , Proteínas de Unión al ADN/genética , Proteínas de Unión al ADN/aislamiento & purificación , Activación Enzimática , Regulación de la Expresión Génica/genética , Semivida , Humanos , Proteínas Quinasas JNK Activadas por Mitógenos , Células Jurkat , Sistema de Señalización de MAP Quinasas , Quinasas de Proteína Quinasa Activadas por Mitógenos/metabolismo , Proteínas Quinasas Activadas por Mitógenos/genética , Datos de Secuencia Molecular , Mutación/genética , Factores de Transcripción NFI , Proteínas Nucleares , Fosfoproteínas/química , Fosfoproteínas/genética , Fosfoproteínas/aislamiento & purificación , Pruebas de Precipitina , Unión Proteica , ARN Mensajero/genética , ARN Mensajero/metabolismo , Proteínas de Unión al ARN/química , Proteínas de Unión al ARN/genética , Proteínas de Unión al ARN/aislamiento & purificación , Elementos de Respuesta/genética , Linfocitos T/citología , Linfocitos T/enzimología , Linfocitos T/inmunología , Proteína 1 de Unión a la Caja Y , Nucleolina

RESUMEN

STUDY OBJECTIVES: There is accumulating evidence that the common cold produces impairments in psychomotor vigilance. This has led some investigators to hypothesize that such illnesses may also have disruptive effects on sleep. While several self-report studies suggest that viral illness may influence sleep parameters, no studies have assessed polysomnographically recorded sleep following viral infections. DESIGN: Parallel control group comparison. SETTING: Sleep laboratory in a large urban medical center. PARTICIPANTS: Twenty-one men and women with susceptibility to the rhinovirus type 23. INTERVENTIONS: Nasal inoculation with rhinovirus type 23. MEASUREMENTS: Polysomnographically recorded sleep for five nights (2300-0700 h) post-viral inoculation. Twice daily (1030 and 1430 h) performance assessment during each experimental day using auditory vigilance and divided attention tasks. A multiple sleep latency test (MSLT) was performed daily for the duration of the study. RESULTS: In symptomatic individuals, total sleep time decreased an average of 23 min, consolidated sleep decreased an average of 36 min, and sleep efficiency was reduced by an average of 5% during the active viral period (experimental days/nights 3-5) compared with the incubation period. Psychomotor performance was impaired. These changes were significantly greater than those observed in asymptomatic individuals. CONCLUSIONS: The common cold can have detrimental effects on sleep and psychomotor performance in symptomatic individuals during the initial active phase of the illness.

Asunto(s)

Atención/fisiología , Resfriado Común/psicología , Desempeño Psicomotor/fisiología , Rhinovirus , Sueño/fisiología , Adolescente , Adulto , Afecto/fisiología , Femenino , Humanos , Masculino , Persona de Mediana Edad , Polisomnografía

Asunto(s)

Vacunas contra el Cólera , Programas de Inmunización , Refugiados , Factores de Riesgo , Estudios de Factibilidad , Uganda , Sudán

RESUMEN

In refugee settings, the use of cholera vaccines is controversial since a mass vaccination campaign might disrupt other priority interventions. We therefore conducted a study to assess the feasibility of such a campaign using a two-dose oral cholera vaccine in a refugee camp. The campaign, using killed whole-cell/recombinant B-subunit cholera vaccine, was carried out in October 1997 among 44,000 south Sudanese refugees in Uganda. Outcome variables included the number of doses administered, the drop-out rate between the two rounds, the proportion of vaccine wasted, the speed of administration, the cost of the campaign, and the vaccine coverage. Overall, 63,220 doses of vaccine were administered. At best, 200 vaccine doses were administered per vaccination site and per hour. The direct cost of the campaign amounted to US$ 14,655, not including the vaccine itself. Vaccine coverage, based on vaccination cards, was 83.0% and 75.9% for the first and second rounds, respectively. Mass vaccination of a large refugee population with an oral cholera vaccine therefore proved to be feasible. A pre-emptive vaccination strategy could be considered in stable refugee settings and in urban slums in high-risk areas. However, the potential cost of the vaccine and the absence of quickly accessible stockpiles are major drawbacks for its large-scale use.

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PIP: This study was undertaken to assess the feasibility of mass vaccination using a two-dose oral cholera vaccine in a refugee setting in Uganda. A total of 44,000 south Sudanese refugees were involved in the study. The campaign was conducted using killed whole-cell/recombinant B-subunit cholera vaccine. Measured outcomes include the total number of doses administered, the dropout rate between the two rounds, the amount of vaccine wasted, the cost of the campaign, and the vaccine coverage. Given the results of the study, the mass vaccination of a refugee population with a two-dose oral cholera vaccine proved to be feasible. A total of 63,220 vaccines were administered, with 200 vaccine doses given per vaccination area per hour. The campaign cost was US$14,655, excluding the cost of the vaccine. Vaccine coverage was 83% for the first round and 75.9% for the second round. A presumptive vaccination strategy could be taken into account in stable refugee settings and in urban slums in high-risk sites. However, the potential amount of the vaccine and the absence of immediately accessible stockpiles are major constraints for its large-scale implementation.

Asunto(s)

Vacunas contra el Cólera/administración & dosificación , Cólera/prevención & control , Refugiados , Vacunación/métodos , Administración Oral , Adolescente , Niño , Preescolar , Vacunas contra el Cólera/economía , Vacunas contra el Cólera/inmunología , Costos Directos de Servicios/estadística & datos numéricos , Estudios de Factibilidad , Femenino , Humanos , Lactante , Masculino , Aceptación de la Atención de Salud/psicología , Aceptación de la Atención de Salud/estadística & datos numéricos , Evaluación de Programas y Proyectos de Salud , Refugiados/psicología , Sudán/etnología , Uganda , Vacunación/economía , Vacunación/psicología , Vacunación/estadística & datos numéricos

RESUMEN

Hodgkin's disease is histopathologically characterized by the relative scarcity of neoplastic Hodgkin and Reed-Sternberg cells and for yet unknown reasons by an abundant reactive background of T lymphocytes and often eosinophils. Eotaxin is a CC-chemokine attracting eosinophils and T helper 2 (Th2) cells in allergic inflammation. We now report that eotaxin is strongly expressed in fibroblasts of Hodgkin's disease tissues, whereas Hodgkin/Reed-Sternberg cells do not express this chemokine. In tissue culture, Hodgkin's disease tumor cells induce eotaxin expression in cocultured dermal fibroblasts in a concentration leading to a specific chemotactic response of a Th2 cell clone. Production of tumor necrosis factor-alpha (TNF-alpha) by Hodgkin/Reed-Sternberg cells appears to be responsible for this induction, because blocking of TNF-alpha by neutralizing antibodies prevented fibroblast eotaxin expression. Our data suggest that eotaxin is involved in the pathobiology of Hodgkin's disease by contributing to eosinophil and T-lymphocyte recruitment.

Asunto(s)

Quimiocinas CC , Quimiotaxis de Leucocito , Citocinas/genética , Enfermedad de Hodgkin/patología , Células de Reed-Sternberg/patología , Piel/citología , Transcripción Genética , Células Cultivadas , Quimiocina CCL11 , Factores Quimiotácticos Eosinófilos/genética , Citocinas/metabolismo , Ensayo de Inmunoadsorción Enzimática , Fibroblastos/citología , Fibroblastos/patología , Fibroblastos/fisiología , Regulación de la Expresión Génica , Células HeLa , Humanos , Hibridación in Situ , Ganglios Linfáticos/patología , ARN Mensajero/genética , Células de Reed-Sternberg/fisiología , Células Th2/fisiología , Células Tumorales Cultivadas

RESUMEN

David P. Phillips has reported evidence that famous people are often able to postpone their deaths until after a birthday. A reexamination of Phillips' data shows some aspects of his analysis to be questionable, including the lumping together of deaths that occur during the birthmonth, which does not distinguish deaths that occurred before the birthday from those that occurred afterward. A reanalysis of his data shows that there were actually a relatively large number of deaths in the month preceding and the months following the birthday. One explanation is that the anxiety associated with this milestone and the excesses associated with its celebration are sometimes fatal. Another explanation is that Phillips' results were a fluke created by a selective use of data.

Asunto(s)

Ansiedad/psicología , Muerte , Personajes , Trabajo de Parto , Interpretación Estadística de Datos , Femenino , Humanos , Embarazo , Reproducibilidad de los Resultados

RESUMEN

Breast cancers are either primarily resistant to chemotherapy (intrinsic resistance), or respond to chemotherapy but later recur with a multidrug-resistant phenotype because of overexpression of the multidrug transporter P-glycoprotein. The MDR1 gene encoding P-glycoprotein may be transcriptionally regulated by a Y-box transcription factor. We now report that, in multidrug-resistant MCF-7 breast cancer cells, nuclear localization of YB-1 is associated with MDR-1 gene expression. In drug-sensitive MCF-7 cells, however, YB-1 was localized to the cytoplasm. Regulated overexpression of YB-1 in drug-sensitive diploid breast epithelial cells induced MDR-1 gene expression and multidrug resistance. In 27 out of 27 untreated primary breast cancers, YB-1 protein was expressed in the cytoplasm although it was undetectable in normal breast tissue of these patients. In a subgroup of tumors (9/27), however, YB-1 was also localized to the nucleus and, in these cases, high levels of P-glycoprotein were present. These results show that in a subset of untreated primary breast cancers, nuclear localization of YB-1 protein is associated with intrinsic multidrug resistance. Our data show that YB-1 has an important role in controlling MDR1 gene transcription and this finding provides a basis for the analysis of molecular mechanisms responsible for intrinsic multidrug resistance in human breast cancer.

Asunto(s)

Miembro 1 de la Subfamilia B de Casetes de Unión a ATP/metabolismo , Neoplasias de la Mama/genética , Proteínas Potenciadoras de Unión a CCAAT , Proteínas de Unión al ADN/metabolismo , Resistencia a Múltiples Medicamentos/genética , Regulación Neoplásica de la Expresión Génica , Factores de Transcripción/metabolismo , Adulto , Anciano , Neoplasias de la Mama/patología , Compartimento Celular , Núcleo Celular/metabolismo , Femenino , Humanos , Persona de Mediana Edad , Factores de Transcripción NFI , Proteínas Nucleares , Transcripción Genética , Proteína 1 de Unión a la Caja Y

RESUMEN

The pathogenesis and etiology of Hodgkin's disease, a common human malignant lymphoma, is still unresolved. As a unique characteristic, we have identified constitutive activation of the transcription factor nuclear factor (NF)-kappaB p50-RelA in Hodgkin/Reed-Sternberg (H/RS) cells, which discriminates these neoplastic cells from most cell types studied to date. In contrast to other lymphoid and nonlymphoid cell lines tested, proliferation of H/RS cells depended on activated NF-kappaB. Furthermore, constitutive NF-kappaB p50-RelA prevented Hodgkin's lymphoma cells from undergoing apoptosis under stress conditions. Consistent with this dual function, Hodgkin's lymphoma cells depleted of constitutive nuclear NF-kappaB revealed strongly impaired tumor growth in severe combined immunodeficient mice. Our findings identify NF-kappaB as an important component for understanding the pathogenesis of Hodgkin's disease and for developing new therapeutic strategies against it.

Asunto(s)

Enfermedad de Hodgkin/patología , FN-kappa B/fisiología , Animales , Apoptosis , División Celular , Línea Celular , Supervivencia Celular , Enfermedad de Hodgkin/metabolismo , Humanos , Ratones , Ratones SCID , Subunidad p50 de NF-kappa B , Factor de Transcripción ReIA , Células Tumorales Cultivadas

RESUMEN

Resistance to apoptosis plays an important role in malignancies that are refractory to chemotherapy treatment. Recently we have shown that the expression of bax-alpha, a death-promoting member of the bcl-2 family, is down-regulated in breast cancer and have provided evidence that low bax expression might contribute to the pathogenesis of breast cancer. In this study we were able to demonstrate the role of this gene in chemotherapy-induced apoptosis. We transfected bax-alpha into the breast-cancer cell lines R30C and MCF-7 under the control of an inducible tetracycline-dependent expression system. Induction of bax-alpha expression did not affect viability by itself but strongly increased chemosensitivity to epirubicin. We were able to demonstrate that this sensitization is due to apoptosis. These data might explain the recently published observation that reduced expression of bax is associated with poor response rates to chemotherapy in breast cancer.

Asunto(s)

Apoptosis/efectos de los fármacos , Neoplasias de la Mama/patología , Proteínas Proto-Oncogénicas/genética , Proto-Oncogenes , Neoplasias de la Mama/genética , Epirrubicina/farmacología , Femenino , Humanos , Proteínas Proto-Oncogénicas/análisis , Proteínas Proto-Oncogénicas c-bcl-2 , Células Tumorales Cultivadas , Proteína X Asociada a bcl-2

RESUMEN

We have studied the expression of members of the bcl-2 family in human breast cancer. The expression pattern of these genes in breast cancer tissue samples was compared with the expression pattern in normal breast epithelium. No marked difference with regard to bcl-2 and bcl-xL expression was observed between normal breast epithelium and cancer tissue. In contrast, bax-alpha, a splice variant of bax, which promotes apoptosis, is expressed in high amounts in normal breast epithelium, whereas only weak or no expression could be detected in 39 out of 40 cancer tissue samples examined so far. Of interest, downregulation of bax-alpha was found in different histological subtypes. Furthermore, we transfected bax-alpha into breast cancer cell lines under the control of a tetracycline-dependent expression system. We were able to demonstrate for the first time that induction of bax expression in breast cancer cell lines restores sensitivity towards both serum starvation and APO-I/Fas-triggered apoptosis and significantly reduces tumor growth in SCID mice. Therefore, we propose that dysregulation of apoptosis might contribute to the pathogenesis of breast cancer at least in part due to an imbalance between members of the bcl-2 gene family.

Asunto(s)

Apoptosis , Neoplasias de la Mama/genética , Neoplasias de la Mama/patología , Regulación Neoplásica de la Expresión Génica , Proteínas Proto-Oncogénicas/biosíntesis , Animales , Secuencia de Bases , Mama/metabolismo , División Celular , Línea Celular , Cartilla de ADN , ADN de Neoplasias/biosíntesis , Epitelio/metabolismo , Femenino , Humanos , Ratones , Ratones SCID , Datos de Secuencia Molecular , Reacción en Cadena de la Polimerasa , Proteínas Proto-Oncogénicas c-bcl-2 , ARN Mensajero/análisis , ARN Mensajero/biosíntesis , Proteínas Recombinantes/biosíntesis , Valores de Referencia , Transcripción Genética , Transfección , Células Tumorales Cultivadas , Proteína X Asociada a bcl-2 , Receptor fas/fisiología

RESUMEN

There is a considerable lack of understanding about the common molecular defects that form the basis for the occurrence of Hodgkin's lymphoma. Despite a number of molecular tools used thus far in immunophenotypic and karyotypic studies, it has not been possible to establish a single common trait among various Hodgkin (H)-cell lines or primary tumor cells that would allow classification into a particular hematopoietic lineage. With this study, we demonstrate that a characteristic expression pattern of transcription factors provides a unifying principle. Seven different cell lines derived from patients with Hodgkin's disease (HD), as well as primary H/Reed-Sternberg (RS) cells isolated from the pericardial fluid of a patient with HD, were compared with a number of hematopoietic and nonhematopoietic cell lines for the expression of Oct-2, a tissue-specific transcription factor normally restricted to B cells, and nuclear factor kappa B (NF-kappa B), an inducible transcription factor. Regardless of the heterogeneous phenotypes and genotypes of the H cell lines, which varied inconsistently between B-cell-, T-cell-, or monocyte-like properties, all H cells tested displayed expression of Oct-2 protein at levels comparable to those seen in B cells. Furthermore, all cell lines showed an abundant constitutive nuclear NF-kappa B activity. Interestingly, anaplastic large-cell lymphoma (ALCL) cell lines, which have many features in common with H/RS cells, were devoid constitutive nuclear NF-kappa B activity. Unlike the constitutive NF-kappa B activity known for B cells, which mainly consists of the p50 and c-Rel or RelB subunits, we demonstrate by antibody supershifting experiments that H cells contain constitutive nuclear p50 and p65, the dimeric NF-kappa B normally observed only for limited time intervals after stimulation with diverse inducers. Additionally, some H-cell lines also displayed nuclear c-Rel activity, whereas RelB or p52 were not detected as part of the constitutive activity. The expression pattern of Oct-2 and NF-kappa B appears to be a unifying and characteristic property of H cells and might explain the deregulated expression of various cytokines leading to the clinical and pathologic manifestations of HD.

Asunto(s)

Proteínas de Unión al ADN/biosíntesis , Regulación Neoplásica de la Expresión Génica , Enfermedad de Hodgkin/patología , FN-kappa B/biosíntesis , Proteínas de Neoplasias/biosíntesis , Células de Reed-Sternberg/metabolismo , Factores de Transcripción , Linfocitos B/patología , Secuencia de Bases , Neoplasias de la Mama/patología , Proteínas de Unión al ADN/genética , Femenino , Enfermedad de Hodgkin/genética , Enfermedad de Hodgkin/metabolismo , Humanos , Inmunofenotipificación , Leucemia/patología , Persona de Mediana Edad , Datos de Secuencia Molecular , Familia de Multigenes , FN-kappa B/genética , Proteínas de Neoplasias/genética , Factor 2 de Transcripción de Unión a Octámeros , Plasmacitoma/patología , Proteínas Proto-Oncogénicas/química , Proteínas Proto-Oncogénicas/genética , Proteínas Proto-Oncogénicas c-rel , Linfocitos T/patología , Células Tumorales Cultivadas

RESUMEN

The nuclear factor p92, originally discovered by its interaction with the human papillomavirus type 18 enhancer, is a cellular protein whose activity is restricted to S phase in human primary fibroblasts. The human papillomavirus type 18 p92 binding sequence confers enhancer activity on a heterologous promoter, suggesting that p92 acts as a transcription factor. We have identified a class of nuclear inhibitory proteins, I-92s, which noncovalently associate with p92 but not with other transcription factors such as AP1, E2F, or NF-kappaB. Different I-92s occur in G1, G2, and G0, while no I-92 is detectable in S phase. Phase-specific inhibitors, therefore, are responsible for the cell cycle dependence of p92 activity and provide a novel mechanism linking transcription factor regulation with the cell cycle.

Asunto(s)

Ciclo Celular , Núcleo Celular/metabolismo , Proteínas de Unión al ADN/metabolismo , Proteínas Nucleares/metabolismo , Factores de Transcripción/metabolismo , Secuencia de Bases , Sitios de Unión , Células Cultivadas , Elementos de Facilitación Genéticos , Fibroblastos , Citometría de Flujo , Células HeLa , Humanos , Datos de Secuencia Molecular , Proteínas Nucleares/aislamiento & purificación , Oligodesoxirribonucleótidos , Papillomaviridae/genética , Regiones Promotoras Genéticas , Proteínas Recombinantes/metabolismo , Fase S , Virus 40 de los Simios/genética , Transfección

RESUMEN

The transcription factor E2F is regulated during the cell cycle through interactions with the product of the retinoblastoma susceptibility gene and related proteins. It is thought that E2F-mediated gene regulation at the G1/S boundary and during S phase may be one of the rate-limiting steps in cell proliferation. It was reported that in vivo overexpression of E2F-1 in fibroblasts induces S phase entry and leads to apoptosis. This observation suggests that E2F plays a role in both cell cycle regulation and apoptosis. To further understand the role of E2F in cell cycle progression, cell death, and tumor development, we have blocked endogenous E2F activity in HBL-100 cells, derived from nonmalignant human breast epithelium, using dominant-negative mutants under the control of a tetracycline-dependent expression system. We have shown here that induction of dominant-negative mutants led to strong downregulation of transiently transfected E2F-dependent chloramphenicol acetyl transferase reporter constructs and of endogenous c-myc, which has been described as a target gene of the transcription factor E2F/DP. In addition, we have shown that blocking of E2F could efficiently protect from apoptosis induced by serum starvation within a period of 10 d, whereas control cells started to die after 24 h. Surprisingly, blocking of E2F did not alter the rate of proliferation or of DNA synthesis of these cells; this finding indicates that cell-cycle progression could be driven in an E2F-independent manner. In addition, we have been able to show that blocking of endogenous E2F in HBL-100 cells led to rapid induction of tumor growth in severe combined immunodeficiency mice. No tumor growth could be observed in mice that received mock-transfected clones or tetracycline to block expression of the E2F mutant constructs in vivo. Thus, it appears that E2F has a potential tumor-suppressive function under certain circumstances. Furthermore, we provide evidence that dysregulation of apoptosis may be an important step in tumorigenesis.

Asunto(s)

Neoplasias de la Mama/genética , Neoplasias de la Mama/patología , Proteínas Portadoras , Proteínas de Ciclo Celular , Ciclo Celular , Proteínas de Unión al ADN , Regulación de la Expresión Génica , Genes myc , Factores de Transcripción/metabolismo , Animales , Apoptosis , Secuencia de Bases , División Celular , Línea Celular , Cloranfenicol O-Acetiltransferasa/biosíntesis , Cartilla de ADN , Replicación del ADN , Factores de Transcripción E2F , Factor de Transcripción E2F1 , Femenino , Fibroblastos , Genes Dominantes , Humanos , Cinética , Ratones , Datos de Secuencia Molecular , Trasplante de Neoplasias , Reacción en Cadena de la Polimerasa , Regiones Promotoras Genéticas , Proteínas Proto-Oncogénicas c-myc/biosíntesis , ARN Mensajero/análisis , ARN Mensajero/biosíntesis , Proteínas Recombinantes/biosíntesis , Proteína 1 de Unión a Retinoblastoma , Factor de Transcripción DP1 , Factores de Transcripción/antagonistas & inhibidores , Factores de Transcripción/biosíntesis , Transfección