RESUMO
The objective was to assess the number of patients with acute oesophageal bolus obstruction that resolves spontaneously and to aid the identification of the best practice. This prospective and retrospective case series study at a teaching hospital and a district general hospital in Scotland, UK, involved 37 patients with acute oesophageal obstruction from a food bolus who were observed for 24 h from the beginning of symptoms. The bolus passed spontaneously in 54% of the patients during the observational period. A short observational period following the admission of patients with acute food bolus obstruction is reasonable as it may reduce exposure to surgical morbidity and decrease inpatient stay.
Assuntos
Esôfago , Corpos Estranhos/terapia , Doença Aguda , Adolescente , Adulto , Idoso , Idoso de 80 Anos ou mais , Humanos , Pessoa de Meia-IdadeRESUMO
Retropharyngeal space swelling is a rare occurrence following minor head and neck trauma. Upper airway obstruction is a potentially life-threatening sequela. The authors present a case of retropharyngeal space haematoma following minor blunt head and neck trauma. Management was conservative with gradual spontaneous resolution of the haematoma. The literature is reviewed and the management and treatment principles presented.
Assuntos
Traumatismos Craniocerebrais/complicações , Hematoma/etiologia , Lesões do Pescoço/complicações , Doenças Faríngeas/etiologia , Acidentes por Quedas , Traumatismos Craniocerebrais/diagnóstico por imagem , Traumatismos Craniocerebrais/terapia , Hematoma/diagnóstico por imagem , Hematoma/terapia , Humanos , Masculino , Pessoa de Meia-Idade , Lesões do Pescoço/diagnóstico por imagem , Lesões do Pescoço/terapia , Doenças Faríngeas/diagnóstico por imagem , Doenças Faríngeas/terapia , Radiografia , Remissão Espontânea , Resultado do TratamentoRESUMO
Epstein Barr Virus (EBV) associated lymphoproliferative disorders (LPD) express EBV latent antigens that are also expressed on normal B-cells transformed with EBV. This could potentially be exploited to develop immunotherapeutic strategies for LPD and other EBV associated malignancies. To this end we investigated the capacity of human monocyte derived dendritic cells (DC) pulsed with lysate from autologous EBV transformed B-cell lymphoblastoid cell (BCL) lysate to elicit an in vitro antitumor response. BCL lysate pulsed DC generate BCL specific cytotoxic lymphocytes, as lymphocytes primed with such DCs induce cytolysis of autologous (>60%) but not allogeneic BCL (<5%). In addition, lymphocytes primed with BCL lysate pulsed DC secrete gamma-IFN (3176 pg/ml). Whereas gamma-IFN production was markedly reduced (>99%) when BCL specific T-cells were stimulated by BCL lysate pulsed DC in the presence of blocking antibodies to HLA-DR, DP and DQ, use of antibodies to MHC class-I resulted in only a minimal reduction in gamma-IFN production (17%). These studies demonstrate that BCL lysate pulsed DC elicit a predominantly BCL specific, MHC class-Il restricted T cell response. This suggests that vaccination with autologous BCL lysate pulsed DC may represent a viable immunotherapeutic approach for the treatment of LPD.
Assuntos
Linfócitos B/imunologia , Transformação Celular Viral/imunologia , Células Dendríticas/imunologia , Herpesvirus Humano 4/fisiologia , Antígenos de Histocompatibilidade Classe II/imunologia , Linfócitos T Citotóxicos/imunologia , Células Cultivadas , Citocinas/biossíntese , Humanos , Interferon gama/biossíntese , Ativação Linfocitária , Monócitos/imunologiaRESUMO
Dietary fats have been shown by many investigators to affect immune responses in vitro and in vivo. However, the exact mechanism(s) by which fats or their metabolic derivatives affect immune function is still unknown. In this report we have investigated whether short-term in vitro exposure to fatty acids alters the adhesion of lymphocytes to extracellular matrix proteins. We found remarkably heterogeneous effects with these agents on lymphocyte adhesion; increases and decreases in adhesion were observed depending upon the fatty acid, cell type and extracellular matrix protein used. Alterations in the adhesion potential of lymphocytes could serve as a mechanism for the reported effects of fatty acids on immune function since lymphocytes are dependent upon rapid and reversible adherence events for most of their effector activities.
Assuntos
Linfócitos B/fisiologia , Adesão Celular/fisiologia , Gorduras Insaturadas na Dieta/farmacologia , Gorduras na Dieta , Proteínas da Matriz Extracelular/fisiologia , Ácidos Graxos não Esterificados/farmacologia , Fluidez de Membrana/efeitos dos fármacos , Linfócitos T/fisiologia , Animais , Linfócitos B/efeitos dos fármacos , Adesão Celular/efeitos dos fármacos , Células Cultivadas , Colágeno/fisiologia , Feminino , Fibronectinas/fisiologia , Laminina/fisiologia , Camundongos , Camundongos Endogâmicos BALB C , Baço/imunologia , Linfócitos T/efeitos dos fármacosRESUMO
Transcription initiation by RNA polymerase II is effected by an ordered series of general factor interactions with core promoter elements (leading to basal activity) and further regulated by gene-specific factors acting from distal elements. Both the general factor TFIID (refs 2,3), including the constituent TBP (TATA-binding polypeptide) and associated factors, and the interacting factor TFIIB (refs 9-11) have been implicated as targets for various activators. Towards an understanding of the basis for activator function, including the multiplicity of TBP interactions, we have now identified mutations in yeast TBP that selectively block activator (GAL4-VP16)-dependent but not basal transcription. We further show an effect of GAL4-VP16 on TFIIB recruitment to early preinitiation complexes, and that recruitment is disrupted by TBP mutations that impair its interactions with VP16 (L114K), TFIIB (L189K) or an unidentified component (K211L). Thus, GAL4-VP16 function seems to involve both direct interactions with TBP and a corresponding induction (or stabilization) of an activation-specific TBP-TFIIB-promoter complex.
Assuntos
Proteínas de Ligação a DNA/metabolismo , Saccharomyces cerevisiae/genética , Fatores de Transcrição/metabolismo , Transcrição Gênica/fisiologia , Arabidopsis/química , Cristalografia por Raios X , Proteínas de Ligação a DNA/química , Proteínas de Ligação a DNA/genética , Proteínas Fúngicas/metabolismo , Proteínas de Plantas/química , Mutação Puntual , Testes de Precipitina , Regiões Promotoras Genéticas , Ligação Proteica , Conformação Proteica , RNA Polimerase II/metabolismo , Proteínas Recombinantes/metabolismo , Saccharomyces cerevisiae/metabolismo , Proteína de Ligação a TATA-Box , Transativadores/metabolismo , Fator de Transcrição TFIIA , Fator de Transcrição TFIIB , Fatores de Transcrição/química , Fatores de Transcrição/genéticaRESUMO
Resolution of whole cell extract through two chromatographic steps yields a single protein fraction requiring only the addition of TFIID for the initiation of transcription at RNA polymerase II promoters. This approach allows the convenient generation of RNA polymerase II transcription systems from Saccharomyces cerevisiae, human lymphocytes, and Schizosaccharomyces pombe. TFIIDs from all three organisms are interchangeable among all three systems. The S. cerevisiae and Sch. pombe systems support effects of acidic activator proteins, provided a further protein fraction from S. cerevisiae is supplied. This further fraction is distinct from the mediator of transcriptional activation described previously and represents a second component in addition to general initiation factors that may facilitate a response to acidic activators.
Assuntos
Regulação Fúngica da Expressão Gênica , Regiões Promotoras Genéticas , RNA Polimerase II/genética , Schizosaccharomyces/genética , Schizosaccharomyces/metabolismo , Fatores de Transcrição/metabolismo , Transcrição Gênica , Linhagem Celular , Regulação Enzimológica da Expressão Gênica , Humanos , Cinética , Linfócitos , Saccharomyces cerevisiae/genética , Saccharomyces cerevisiae/metabolismo , Fator de Transcrição TFIID , Fatores de Transcrição/isolamento & purificaçãoRESUMO
Previous work showed that human TFIID fails to support yeast cell growth, although it is nearly identical to yeast TFIID in a carboxy-terminal region of the molecule that suffices for basal, TATA-element-dependent transcription in vitro. These and other findings raised the possibility that TFIID participates in species-specific interactions, possibly with mediator factors, required for activated transcription. Here, we report that human TFIID and amino-terminally truncated derivatives of yeast TFIID are fully functional in support of both basal transcription and the response to acidic activator proteins in a yeast in vitro transcription system. Conversely, and in contrast to previously published results, yeast TFIID supports both basal and activated transcription in reactions reconstituted with human components. This functional interchangeability of yeast and human TFIIDs argues strongly against species specificity with regard to TFIID function in basal transcription and the response to acidic activator proteins. In addition, our results suggest that any intermediary factors between acidic activators and TFIID are conserved from yeast to man.
Assuntos
Proteínas de Ligação a DNA , Proteínas Quinases , Proteínas de Saccharomyces cerevisiae , Saccharomyces cerevisiae/genética , Fatores de Transcrição/metabolismo , Transcrição Gênica/fisiologia , Northern Blotting , Proteína Receptora de AMP Cíclico/metabolismo , Proteínas Fúngicas/genética , Humanos , Fragmentos de Peptídeos/metabolismo , RNA Polimerase II/metabolismo , Transativadores/genética , Fator de Transcrição TFIID , Fatores de Transcrição/genéticaRESUMO
Activator proteins bind to enhancer DNA elements and stimulate the initiation of transcription. It has been proposed that activators contact general initiation factors at a promoter, and evidence for such direct interaction has been obtained. Studies of transcription in vitro, however, have suggested that activators might function through an intermediary molecule(s) distinct from the general factors. In the first of these studies, we exploited the finding that one activator could inhibit transcription stimulated by a second activator (activator interference or 'squelching'). This inhibition, which is attributed to competition between the activators for a common target factor, could not be relieved by addition of a large excess of general initiation factors, suggesting that the target for which activators compete is distinct from these factors. Similar conclusions came from the observation that TFIID's expressed from cloned genes fail to replace partially purified 'natural' TFIID fractions in supporting activation, evidently because they lacked some component present in the impure fractions. While these lines of evidence for a novel 'mediator' of activation were negative, we also showed that a partially purified fraction from yeast would reverse activator interference. This positive effect of a presumptive mediator provided an assay for its activity, but its role in activation was still only inferred. We now present direct evidence for a mediator which is required for stimulation of transcription in vitro by the activators GAL4-VP16 and GCN4, but which has no effect on transcription in the absence of activator protein.
Assuntos
Proteínas Nucleares/fisiologia , RNA Polimerase II/metabolismo , Proteínas de Saccharomyces cerevisiae , Transcrição Gênica , Proteínas de Ligação a DNA/metabolismo , Ativação Enzimática , Proteínas Fúngicas/metabolismo , Técnicas In Vitro , Proteínas Recombinantes/metabolismo , Saccharomyces cerevisiae , Fator de Transcrição TFIID , Fatores de Transcrição/metabolismoAssuntos
Núcleo Celular/metabolismo , RNA Polimerase II/metabolismo , Saccharomyces cerevisiae/genética , Transcrição Gênica , Fracionamento Celular/métodos , Núcleo Celular/ultraestrutura , Sistema Livre de Células , Regiões Promotoras Genéticas , Saccharomyces cerevisiae/metabolismo , Esferoplastos/metabolismo , Esferoplastos/ultraestruturaRESUMO
Fractionation of a yeast nuclear extract reveals at least four factors required in addition to RNA polymerase II for accurate initiation of transcription. One of these factors can be replaced by HeLa transcription factor IID or by its yeast counterpart expressed in Escherichia coli. Each of the remaining three factors can be replaced by a fraction from yeast whole cell extract, facilitating further purification of the factors.
Assuntos
RNA Polimerase II/metabolismo , Saccharomyces cerevisiae/enzimologia , Fatores de Transcrição/farmacologia , Transcrição Gênica , Núcleo Celular/enzimologia , Escherichia coli/metabolismo , Células HeLa/análise , Saccharomyces cerevisiae/ultraestrutura , Fator de Transcrição TFIIA , Fator de Transcrição TFIID , Fatores de Transcrição/isolamento & purificaçãoRESUMO
One gene activator protein may interfere with the effects of another in eukaryotic cells. We report here that a hybrid yeast-herpes gene activator protein inhibits transcriptional activation by a thymidine-rich DNA element in yeast. This example of activator interference can be faithfully reproduced in vitro. Interference is reversed by a partially purified yeast component, but not by RNA polymerase II or various polymerase II transcription factors. We conclude that the partially purified yeast component is a novel factor, and we suggest this factor mediates the transcriptional activation process.
Assuntos
Proteínas Fúngicas/metabolismo , Fosfoproteínas/metabolismo , RNA Polimerase II/metabolismo , Proteínas de Saccharomyces cerevisiae , Saccharomyces cerevisiae/genética , Transativadores/metabolismo , Fatores de Transcrição/metabolismo , Transcrição Gênica , Núcleo Celular/metabolismo , DNA Fúngico/genética , Proteínas de Ligação a DNA , Proteínas Fúngicas/genética , Regulação Fúngica da Expressão Gênica , Cinética , Fosfoproteínas/genética , Plasmídeos , Proteínas Recombinantes de Fusão/metabolismo , Simplexvirus/genética , Transativadores/genética , beta-Galactosidase/genética , beta-Galactosidase/metabolismoRESUMO
A 53-year-old female patient who presented with recurrent meningitis as a result of a meningioma, is reported. The meningioma was found to be wholly contained within the left mastoid antrium. To our knowledge this is the first reported case of a meningioma localised to the mastoid antrium. The patient had been assessed in the ENT department on two separate occasions, 17 years and 19 years previously for nebulous symptoms related to the left ear which had cleared spontaneously. Following a second episode of pyogenic meningitis, both of which were associated with aural symptoms, radiological examination suggested an intramastoid pathology which prompted mastoid exploration. Histological examination of the mass confined to the mastoid antum provided the diagnosis of meningioma. There was no clinical or radiological evidence of extratemporal spread of tumour.
Assuntos
Processo Mastoide , Meningioma/complicações , Meningite/etiologia , Neoplasias Cranianas/complicações , Feminino , Humanos , Pessoa de Meia-Idade , RecidivaRESUMO
Conformational transitions in topologically constrained duplex DNA necessarily affect and are affected by other transitional processes throughout the entire molecule. This conformational interdependence of discrete sequences within a given superhelical domain arises through a requisite competition for the free energy of supercoiling. Here we present a generalized statistical mechanical analysis of multiple, competing conformational equilibria in superhelical DNA. This model has been applied, using experimentally determined parameters, to the energetic coupling of two independent B-Z transitions. Specifically, we have monitored the extent of B-Z transition, as a function of negative superhelicity, in topoisomers of a plasmid containing two identical d(C-G)n inserts using two-dimensional gel electrophoresis. The theoretical results were found to be in good agreement with the experimental data, and we have used this model to predict the competitive behavior of B-Z transitions within sequences differing in length and sequence composition. This competition is shown to have a profound effect upon the B-Z equilibria of those sequences analyzed, resulting in a complex modulation in the extent of Z-DNA formation as a function of negative superhelicity. These theoretical and experimental results show that DNA sequences separated by large distances are capable of communicating structural information.
Assuntos
DNA Super-Helicoidal , DNA , Conformação de Ácido Nucleico , Modelos Teóricos , Plasmídeos , Polidesoxirribonucleotídeos , TermodinâmicaRESUMO
The ability of a stretch of alternating dA-dT to adopt the left-handed Z form has been assessed by examining the behavior of the sequence d(CG)6(TA)4(CG)6 contained in the plasmid pBR322. The structural transition occurring within this sequence as a function of negative superhelicity was analyzed by several methods, including (1) the supercoiling-dependent unwinding of the insert as determined by two-dimensional gel electrophoresis, (2) the binding of anti-Z-DNA antibodies to the insert, (3) the sensitivity of the sequence to a single strand specific endonuclease, and (4) the sensitivity of the insert to digestion by a restriction endonuclease that cuts within the d(CG)6 segments when in the right-handed form. These studies have shown that in negatively supercoiled DNA the two d(CG)6 portions of the insert adopt the Z form, while the central d(TA)4 segment forms an underwound structure with a helical repeat that is best approximated as being intermediate between the B form and the Z form. A statistical mechanical treatment of the unwinding of the insert as a function of negative superhelicity provides an estimate of the minimum free energy required to convert an A-T bp from the B form to the Z form, as well as the free energy associated with the conversion of an A-T bp from the B form to the unwound form. These results strongly indicate that Z DNA is an unfavored structural alternative for stretches of d(AT)n in negatively supercoiled DNA.
Assuntos
DNA Super-Helicoidal , DNA , Conformação de Ácido Nucleico , Plasmídeos , Poli dA-dT , Polidesoxirribonucleotídeos , Sequência de Bases , TermodinâmicaRESUMO
The likelihood that a given DNA sequence will adopt the Z conformation in negatively supercoiled DNA depends on the energy difference between the B form and the Z form for that sequence relative to other sequences in the same molecule. This energy can be viewed simply as a sum of energies for the nearest-neighbor interactions within the sequence plus the energy required to stabilize the B-Z boundaries. Knowledge of these energetic terms would be of value in predicting when sequences become left-handed in response to negative superhelicity. Here we present an approach that can be used to determine the free-energy changes associated with all the nearest-neighbor interactions that can occur in Z-DNA. Synthetic stretches of d(C-G)n containing one or two transversions were cloned into plasmids, and the extent of the B-Z transition as a function of negative superhelicity was determined for each insert by two-dimensional agarose gel electrophoresis. By subjecting the data to statistical mechanical analysis, it was possible to evaluate the energetic penalty resulting from each base-pair (bp) substitution. Guanine to cytosine transversions cost 2.4 kcal (1 cal = 4.18 J)/(mol X bp), whereas guanine to thymine transversions cost 3.4 kcal/(mol X bp), to stabilize in the Z conformation. We have used these numbers, along with energetic values determined by others for the B-Z transition, to predict that certain strictly nonalternating purine and pyrimidine sequences may adopt the Z form readily.
Assuntos
DNA Super-Helicoidal , Conformação de Ácido Nucleico , Sequência de Bases , Plasmídeos , Relação Estrutura-Atividade , TermodinâmicaRESUMO
The in vivo production of RNA that is complementary to the mRNA of a particular target gene (anti-sense RNA) appears to be an effective tool for the regulation of genes in Escherichia coli (Coleman, J., Green, P.J., and Inouye, M. (1984) Cell 37, 429-436). These investigators demonstrated that short anti-sense transcripts which are complementary to the ribosome binding site of the target mRNA are overwhelmingly the most effective in blocking protein synthesis. We have constructed plasmids which produce thermally regulated anti-sense transcripts of three regions of the E. coli lac Z gene coding sequence, and have examined the relative effects of these constructs on the synthesis of the lac Z gene product, beta-galactosidase. We conclude that there is a strong correlation between the length of RNA complementarity and the suppression of beta-galactosidase synthesis. Furthermore, a significant inhibition of translation can be obtained when the anti-sense transcript lacks complementarity to the 5' noncoding region of the mRNA, provided that the extent of complementarity with the coding sequence is considerable.
Assuntos
Escherichia coli/enzimologia , Escherichia coli/genética , Galactosidases/genética , Genes Bacterianos , Genes Reguladores , Genes , RNA Bacteriano/genética , RNA Mensageiro/genética , beta-Galactosidase/genética , Enzimas de Restrição do DNA , Cinética , Plasmídeos , Temperatura , Transcrição Gênica , beta-Galactosidase/biossínteseRESUMO
Some general features of dexamethasone resistance in five murine lymphoid cell lines were investigated. To obtain large numbers of dexamethasone-resistant (Dexr) variants, a technique was developed by which mouse lymphoid cell lines can be grown with high efficiency on the surface of agar plates without a feeder layer. A total of 271 Dexr variants were investigated, and 90% of them turned out to lack detectable steroid receptor whereas 10% have receptor with, in most cases, a normal affinity for the steroid hormone. Most of this latter class of variants, however, have reduced amounts of receptor and the receptor of all of them displayed altered nuclear binding characteristics. None of the five investigated lymphoid cell lines yielded a Dexr variant with a normal receptor. These results confirm the idea that the high incidence of receptor variant may be due, at least in part, to the haploid state of a gene coding for the receptors. In cell fusion experiments it could be shown that Dexs is dominant over Dexr, but that a Dexr a-lele in a tetraploid cell can lead to an increased frequency of steroid resistance.
Assuntos
Dexametasona/metabolismo , Variação Genética , Linfócitos/metabolismo , Animais , Fusão Celular , Linhagem Celular , Núcleo Celular/metabolismo , Dexametasona/farmacologia , Resistência a Medicamentos , Linfócitos/efeitos dos fármacos , Linfoma , Camundongos , Receptores de Glucocorticoides/metabolismoRESUMO
Gene dosage and escape synthesis experiments support the hypothesis that diaminopimelate decarboxylase repression by lysine involves a repressor molecule in a negative control system.
Assuntos
Carboxiliases/biossíntese , Escherichia coli/enzimologia , Genes Reguladores , Repressão Enzimática , Escherichia coli/metabolismo , Lisina/biossíntese , Lisina/farmacologia , Mutação , Temperatura , Transdução GenéticaRESUMO
Hybrid clones arising after inactivated Sendai virus mediated cell fusion between a bromodeoxyuridine-resistant derivative of the BALB/c mouse lymphoma cell line, S49, and a thioguanine-resistant derivative (A9) of the C3H mouse fibroblast cell line, L929, were selected in HAT medium. The responses of the clones to growth inhibitors as well as their chromosome numbers were consistent with properties expected of hybrids. Hybrid clones expressed the major histocompatibility (H-2) surface antigens of both parental types, i.e., H-2d of BALB/c and H-2k of C3H. The Thy-1.2 antigen, expressed on the surface of the lymphoma parent but not the fibroblast parent, was not detected on the hybrids.
Assuntos
Antígenos HLA/análise , Antígenos de Histocompatibilidade/análise , Células Híbridas/imunologia , Células L/imunologia , Linfoma/imunologia , Animais , Fusão Celular , Membrana Celular/imunologia , Cromossomos , Técnicas In Vitro , Linfoma/patologia , Camundongos , Fenótipo , Linfócitos T/imunologiaRESUMO
Biochemical and genetic data were obtained from a series of 51 Pyr- strains of Bacillus subtilis. The observed enzymatic deficiencies allowed the mutants to be placed into 12 clases, some of which represent defects in more than one of the six known pyrimidine biosynthetic enzymes. Mapping analysis by transformation has shown that all the Pyr- mutations are located in a single small area of the B. subtilis genome. A correlation of the biochemical defects and the genetic data has been made. Those mutations conferring similar enzymatic deficiencies were found to be contiguous on the B. subtilis map. Regulatory aspects of the pyrimidine pathway have also been investigated and are compared to previously reported results from other organisms. Evidence is presented which bears upon the possible physical association of the first three enzymes and the association of at least some of the enzymes of this pathway with particulate elements of the cell. A model for the organization of the enzymes is presented with dihydroorotate dehydrogenase as the central enzyme in a proposed aggregate.