RESUMO
We report the cloning and subcellular localization of a novel Schwann cell-specific protein of 147 kd that we have named periaxin. Periaxin has a remarkable domain of repetitive pentameric units in the primary sequence. It is expressed in the first uncompacted whorls of membrane that ensheathe the axon, and further synthesis of the protein in the rat sciatic nerve parallels the deposition of myelin. In mature myelin, periaxin colocalizes with the myelin-associated glycoprotein in the cytoplasm-filled periaxonal regions of the sheath but is excluded from compact myelin. We propose that periaxin has a role in axon-glial interactions, possibly by interacting with the cytoplasmic domains of integral membrane proteins such as myelin-associated glycoprotein in the periaxonal regions of the Schwann cell plasma membrane.
Assuntos
Axônios/fisiologia , Proteínas de Membrana/fisiologia , Bainha de Mielina/fisiologia , Células de Schwann/fisiologia , Sequência de Aminoácidos , Animais , Sequência de Bases , Clonagem Molecular , Proteínas de Membrana/genética , Proteínas de Membrana/metabolismo , Microscopia Imunoeletrônica , Sondas Moleculares/genética , Dados de Sequência Molecular , Bainha de Mielina/metabolismo , Nervos Periféricos/metabolismo , Ratos , Nervo Isquiático/crescimento & desenvolvimento , Nervo Isquiático/metabolismo , Nervo Isquiático/ultraestrutura , Frações Subcelulares/metabolismoRESUMO
BACKGROUND: The use of the combination of fluorouracil (5-FU) and levamisole has been shown to improve the survival of patients with resected Dukes' stage C colon carcinoma. 5-FU is incorporated into RNA, which results in aberrant processing and turnover of RNA. Neither the mechanism of synergy between the two drugs nor the precise molecular mechanism of action of levamisole is known. Each drug has previously been shown to alter the expression of class I human leukocyte antigens (HLA class I) in colorectal cancer cell lines. PURPOSE: The purpose of this study was to explore the mechanism of interaction between 5-FU and levamisole by investigating the effect of this combination on HLA class I gene expression in the colorectal cancer cell line WiDr. METHODS: WiDr cells were treated either with 5-FU alone or with 5-FU and levamisole. Expression of HLA class I antigens was analyzed by flow cytometry using the monoclonal antibody W6/32. Specific DNA probes for HLA class I, beta 2-microglobulin, beta-actin, HLA class II, and p53 (also known as TP53) were used in Northern blot analysis of the steady-state level of messenger RNAs (mRNAs) and for "run-on" transcription analysis. RESULTS: 5-FU alone produced more than 50% increases in the expression of the HLA class I antigens, and levamisole caused a further 8%-18% increase. 5-FU caused the steady-state level of HLA class I mRNAs to increase by about 80%, and levamisole enhanced this effect of 5-FU by a further 70%. 5-FU did not increase the other mRNAs. In vitro run-on transcription revealed that 5-FU caused a 20%-57% reduction in RNA synthesis, while levamisole caused a 30%-190% increase in RNA synthesis. Levamisole therefore reversed the inhibition of RNA synthesis caused by 5-FU. Both drugs had a general effect on RNA synthesis that was not restricted to HLA class I transcription. CONCLUSIONS: The apparent synergy between levamisole and 5-FU is a result of the incorporation of 5-FU, which may stabilize HLA class I mRNAs, leading to their accumulation, while levamisole augments the accumulation of these stable mRNAs by increasing the rate of transcription. IMPLICATIONS: Levamisole reduces the toxicity of 5-FU caused by generalized inhibition of RNA synthesis, and at the same time augments the effects of 5-FU, which may be due to selective stabilization of certain mRNAs.
Assuntos
Neoplasias Colorretais/tratamento farmacológico , Neoplasias Colorretais/genética , Fluoruracila/farmacologia , Genes MHC Classe I/efeitos dos fármacos , Levamisol/farmacologia , Northern Blotting , Neoplasias Colorretais/imunologia , Sondas de DNA , Sinergismo Farmacológico , Quimioterapia Combinada , Citometria de Fluxo , Genes MHC Classe I/genética , Humanos , RNA Mensageiro/efeitos dos fármacos , RNA Neoplásico/efeitos dos fármacos , Células Tumorais Cultivadas , Microglobulina beta-2/efeitos dos fármacosRESUMO
Highly oncogenic adenovirus type 12 (Ad12) transformed rat cells contain decreased levels of cytoplasmic and nuclear major histocompatibility (MHC) class I transcripts, a process dependent on the presence of the Ad12 E1A oncogene. Here we show that the reduction in MHC expression is due at least in part to a decreased rate of initiation of transcription of MHC class I genes. Gamma-interferon treatment of Ad12 transformed cells causes an elevation in steady-state levels of cytoplasmic MHC class I transcripts.
Assuntos
Adenovírus Humanos/genética , Transformação Celular Neoplásica , Regulação da Expressão Gênica , Genes MHC Classe I , Transcrição Gênica , Animais , Northern Blotting , Linhagem Celular , Interferon gama/farmacologia , Poli A/genética , Poli A/isolamento & purificação , RNA/genética , RNA/isolamento & purificação , RNA Mensageiro , Ratos , Transcrição Gênica/efeitos dos fármacosRESUMO
The intracellular distribution of the transformation-associated cellular protein p53 was studied by indirect immunofluorescence in a series of adenovirus-transformed rodent cells. In most adenovirus 2 or 5 (group C) transformed cell lines p53 was detected in discrete areas in nuclei and in all cell lines p53 was also present in a perinuclear structure. The adenovirus 2 or 5 E1B-58 kD protein, previously found to form molecular complexes with p53 in group C transformed cells, colocalized with p53 in both intracellular locations. Further studies on the region of the cell corresponding to the perinuclear body containing p53 showed that it frequently included the centrosomes of the transformed cell. The intranuclear p53 was released by mild DNAase I digestion.
Assuntos
Transformação Celular Viral , Proteínas de Neoplasias/metabolismo , Fosfoproteínas/metabolismo , Animais , Compartimento Celular , Núcleo Celular/metabolismo , Citoplasma/ultraestrutura , Desoxirribonucleases/metabolismo , Imunofluorescência , Mitose , Proteínas Nucleares/metabolismo , Tubulina (Proteína)/metabolismo , Proteína Supressora de Tumor p53RESUMO
We have examined the possibility that the E7 proteins of the high-risk human papillomavirus (HPV) type 16 and 18 and the oncogenic adenovirus (Ad) type 12 E1A protein share the ability to down-regulate the expression of components of the antigen processing and presentation pathway, as a common strategy in the evasion of immune surveillance during the induction of cell transformation. Expression of the HPV 18 E7 oncoprotein, like Ad 12 E1A, resulted in repression of the major histocompatibility complex (MHC) class I heavy chain promoter, as well as repression of a bidirectional promoter that regulates expression of the genes encoding the transporter associated with antigen processing subunit 1 (TAP1) and a proteasome subunit, low molecular weight protein 2 (LMP2). HPV 16 E7 also caused a reduction in class I heavy chain promoter activity, however it did not have any significant effect on the activity of the bidirectional promoter. Interestingly, expression of the low-risk HPV 6b E7 protein resulted in an increase in MHC class I heavy chain promoter activity, while repressing the TAP1/LMP2 promoter. Interference with the class I pathway could also explain the ability of low-risk HPVs in inducing benign lesions.
Assuntos
Transportadores de Cassetes de Ligação de ATP/genética , Proteínas de Ligação a DNA , Regulação Viral da Expressão Gênica , Antígenos H-2/genética , Complexo Principal de Histocompatibilidade/genética , Proteínas Oncogênicas Virais/fisiologia , Papillomaviridae/fisiologia , Transcrição Gênica , Proteínas da Matriz Viral/genética , Membro 2 da Subfamília B de Transportadores de Cassetes de Ligação de ATP , Transportadores de Cassetes de Ligação de ATP/biossíntese , Proteínas E1A de Adenovirus/fisiologia , Animais , Fibroblastos , Genes MHC Classe I , Antígenos H-2/biossíntese , Humanos , Camundongos , Proteínas Oncogênicas Virais/genética , Papillomaviridae/classificação , Papillomaviridae/genética , Proteínas E7 de Papillomavirus , Regiões Promotoras Genéticas , Ratos , Proteínas Recombinantes de Fusão/fisiologia , Transfecção , Proteínas da Matriz Viral/biossínteseRESUMO
We show using mild extraction procedures that the p53 proto-oncogene forms a complex with adenovirus 5 E1b-58 kD during infection. These complexes are detected as coimmunoprecipitates from radiolabeled extracts of adenovirus infected cells on SDS-PAGE. Furthermore, adenovirus mutants with defects in E1b-58 kD fail to form complexes, whereas mutants in other early region genes still show evidence of complex. Using a panel of monoclonal antibodies to mouse p53, we show that antibodies reacting with N-terminal epitopes on p53, displace E1b-58 kD. This result suggests that E1b-58 kD binds to an N-terminal region of mouse p53. In addition, in a transient transfection assay in monkey COS cells, we show that an N-terminal deletion mutant of mouse p53 does not bind to E1b-58 kD but wild-type mouse p53 does bind. This result again suggests that E1b-58 kD binds an N-terminal determinant on p53.
Assuntos
Adenoviridae/genética , Proteínas Oncogênicas Virais/metabolismo , Proteína Supressora de Tumor p53/metabolismo , Proteínas Precoces de Adenovirus , Animais , Anticorpos Monoclonais , Sítios de Ligação , Linhagem Celular , Deleção Cromossômica , Células HeLa/fisiologia , Humanos , Peso Molecular , Proteínas Oncogênicas Virais/genética , Ligação Proteica , Proto-Oncogene Mas , Transfecção , Proteína Supressora de Tumor p53/genéticaRESUMO
Membrane dipeptidase (MDP) is a zinc metalloenzyme located in the lungs and on the brush border membranes of the kidney and intestine. The gene for MDP (also termed DPEP1) is both frequently lost in Wilm's tumours and is located on human chromosome 16q24.3, a region of the genome known to contain a tumour suppressor gene(s). We now report on the regulation of MDP gene expression in normal and transformed cells. MDP enzyme activity and mRNA was detected in primary baby rat kidney (BRK) cells maintained in culture for up to 4 weeks. In contrast all stable transformed cell lines that were tested, derived either by transformation with the DNA tumour viruses SV40 or adenovirus, or in human tumour cell lines, contained very low levels of or no detectable MDP mRNA or enzyme activity. In BRK cells transformed by the temperature-sensitive tsA58 mutant of SV40 T antigen, MDP activity was not detectable, in cell lines grown at the permissive temperature (33 degrees C) but after 5-14 days of incubation at the non-permissive temperature (39.5 degrees C), MDP protein and enzyme activity could be readily detected. Taken together, these results indicate that MDP expression is characteristic of differentiated kidney epithelial cells and is down-regulated in proliferating, transformed cells.
Assuntos
Dipeptidases/metabolismo , Rim/enzimologia , Animais , Antígenos Transformantes de Poliomavirus/genética , Membrana Celular/enzimologia , Transformação Celular Viral , Células Cultivadas , Cromossomos Humanos Par 16 , Proteínas Ligadas por GPI , Regulação Enzimológica da Expressão Gênica , Regulação Neoplásica da Expressão Gênica , Genes Supressores de Tumor , Humanos , Metaloproteínas/metabolismo , RNA Mensageiro/genética , RNA Neoplásico/genética , Ratos , Vírus 40 dos Símios , Fatores de TempoRESUMO
Cell-type specific transcription of the myelin basic protein (MBP) gene in primary oligodendrocytes (OL) is regulated by cis-acting regulatory elements located at both upstream and downstream of the TATA-box region of the MBP promoter. To identify cell-type specific factors that bind to the downstream regulatory elements, we utilised DNase I footprinting analysis and gel retardation assays with nuclear extracts from myelin-forming OL as well as a non-myelin forming cell line, C6 glioma (C6) cells. Several regions of DNA were protected from DNAse I digestion by nuclear extracts of both cell types. However, two regions, from -17 to +17 and from +47 to +58 were protected specifically in OL, while three regions, from + 17 to + 22, from +43 to +49 and from +58 to +64 were protected only with C6 nuclear extracts. Inspection of the protected regions for homology with known transcription factor binding sites revealed that sequences at from +47 to +58 and from +56 to +68 showed extensive homology to the negative regulatory element (NRE1), of the mouse renin gene and to the interferon (IFN) consensus sequence of major histocompatibility complex class I genes (MHC I-ICS), respectively. Gel retardation assays using a MHC I-ICS oligonucleotide and transient transfection assays using MBP-CAT constructs were used to study the effect of IFNs on MBP promoter activity in OL and C6 cells. In OL, IFN-alpha/beta caused little induction of CAT activity, but IFN-gamma resulted in a 2-3.5-fold decrease in CAT activity. In contrast, in C6 cells both IFN-alpha/beta and IFN-gamma induced a 1.5-2.5-fold increase in CAT activity. The cooperative effects of factors binding to NREs and ICS may be responsible for the cell-type specific regulation of MBP gene transcription.
Assuntos
Proteínas de Ligação a DNA/metabolismo , Proteína Básica da Mielina/genética , Regiões Promotoras Genéticas , Sequências Reguladoras de Ácido Nucleico , TATA Box , Animais , Sítios de Ligação , Linhagem Celular , Humanos , Interferon-alfa/farmacologia , Interferon beta/farmacologia , Interferon gama/farmacologia , Camundongos , Ratos , Células Tumorais CultivadasRESUMO
Human adenoviruses (Ads) have the ability to transform primary cells, and certain Ads, the subgenus A adenoviruses such as Ad12, induce tumours in immunocompetent rodents. The oncogenic phenotype of the subgenus A adenoviruses is determined by the viral E1A oncogene. In order to generate tumours, Ad12-transformed cells must evade the cellular immune system of the host. Ad12 E1A gene products mediate transcriptional repression of several genes in the major histocompatibility complex (MHC) involved in antigen processing and presentation, resulting in evasion of cytotoxic T lymphocyte (CTL) killing of transformed cells. In this review, the molecular mechanisms of E1A-mediated transcriptional repression of MHC gene expression are described. In addition, evasion of natural killer (NK) cell killing by Ad-transformed cells is also considered.
Assuntos
Adenoviridae/imunologia , Antígenos de Histocompatibilidade Classe I/imunologia , Sistema Imunitário/efeitos dos fármacos , Proteínas Oncogênicas Virais/farmacologia , Adenoviridae/genética , Adenoviridae/metabolismo , Adenoviridae/fisiologia , Animais , Regulação para Baixo , Inativação Gênica/efeitos dos fármacos , Antígenos de Histocompatibilidade Classe I/biossíntese , Antígenos de Histocompatibilidade Classe I/genética , Humanos , Proteínas Oncogênicas Virais/antagonistas & inibidoresRESUMO
Hepatocellular carcinoma (HCC) develops on the background of chronic hepatitis. Leukocytes found within the HCC microenvironment are implicated as regulators of tumour growth. We show that diethylnitrosamine (DEN)-induced murine HCC is attenuated by antibody-mediated depletion of hepatic neutrophils, the latter stimulating hepatocellular ROS and telomere DNA damage. We additionally report a previously unappreciated tumour suppressor function for hepatocellular nfkb1 operating via p50:p50 dimers and the co-repressor HDAC1. These anti-inflammatory proteins combine to transcriptionally repress hepatic expression of a S100A8/9, CXCL1 and CXCL2 neutrophil chemokine network. Loss of nfkb1 promotes ageing-associated chronic liver disease (CLD), characterized by steatosis, neutrophillia, fibrosis, hepatocyte telomere damage and HCC. Nfkb1(S340A/S340A)mice carrying a mutation designed to selectively disrupt p50:p50:HDAC1 complexes are more susceptible to HCC; by contrast, mice lacking S100A9 express reduced neutrophil chemokines and are protected from HCC. Inhibiting neutrophil accumulation in CLD or targeting their tumour-promoting activities may offer therapeutic opportunities in HCC.
Assuntos
Carcinoma Hepatocelular/genética , Neoplasias Hepáticas Experimentais/genética , Subunidade p50 de NF-kappa B/genética , Neutrófilos/imunologia , Alquilantes/toxicidade , Animais , Calgranulina A/genética , Calgranulina B/genética , Carcinoma Hepatocelular/induzido quimicamente , Carcinoma Hepatocelular/imunologia , Quimiocina CXCL1/genética , Quimiocina CXCL2/genética , Dietilnitrosamina/toxicidade , Hepatopatias/genética , Hepatopatias/imunologia , Neoplasias Hepáticas Experimentais/induzido quimicamente , Neoplasias Hepáticas Experimentais/imunologia , Camundongos , Camundongos Knockout , MutaçãoRESUMO
The myelin (My) basic protein-encoding gene (MBP) is specifically expressed by oligodendrocytes (OL) in the central nervous system (CNS) and by Schwann cells in the peripheral nervous system (PNS). To define cell-type-specific regulatory elements, a series of chimaeric constructs containing varying lengths of the 5'-flanking region and exon 1 of MBP linked to the cat reporter gene was transfected into several cell types, including primary cultures of differentiated rat OL, Schwann cells and kidney cells, as well as neuronal and non-neuronal cell lines. All the constructs generated variable levels of chloramphenicol acetyltransferase (CAT) activity in all cell types, except in primary OL and Schwann cells, where distinct positive (PRE) and negative regulatory elements (NRE) were found to be involved in regulating cat expression. The nucleotide (nt) -53/+70 construct gave maximal activity in all cell types, except OL and Schwann cells, where sequences upstream from nt -53 were necessary for maximal promoter activity. The sequences located at nt -655 to -397 and nt -394 to -54 showed enhancer and repressor effects, respectively, in OL. In Schwann cells, sequences from -394 to -253 showed positive regulatory effects, while those between -655 to -397 and -253 to -54 showed negative regulatory effects. In the downstream region of the promoter, sequences from +20 to +70 and +70 to +200 showed strong silencer and enhancer activities specifically in OL. In gel-retardation assays using nuclear extracts prepared from several cell types and the -253 to -53 repressor sequences, specific DNA-protein complexes unique to OL were identified.(ABSTRACT TRUNCATED AT 250 WORDS)
Assuntos
Regulação da Expressão Gênica , Proteína Básica da Mielina/biossíntese , Oligodendroglia/metabolismo , Sequência de Aminoácidos , Animais , Sequência de Bases , Linhagem Celular , Células Cultivadas , Sistema Nervoso Central/metabolismo , Cloranfenicol O-Acetiltransferase/biossíntese , Sequência Conservada , Éxons , Humanos , Camundongos , Dados de Sequência Molecular , Proteína Básica da Mielina/genética , Oligodesoxirribonucleotídeos , Regiões Promotoras Genéticas , Splicing de RNA , Ratos , Proteínas Recombinantes de Fusão/biossíntese , Sequências Reguladoras de Ácido Nucleico , Sequências Repetitivas de Ácido Nucleico , Mapeamento por Restrição , Células de Schwann/metabolismo , Homologia de Sequência do Ácido Nucleico , Suínos , Células Tumorais CultivadasRESUMO
Cells transformed by human adenovirus 12 (Ad12) exhibit extremely low surface levels of MHC class I molecules and contain reduced levels of class I heavy chain mRNAs. We report that levels of MHC-encoded TAP1 and LMP2 mRNAs are also down-regulated in Ad12-transformed rat cells, and that transcription of rat TAP1 and LMP2 transcripts is directed from a 564 bp intergenic region which is significantly less active in Ad12-transformed cells compared to those transformed with Ad5. Our results suggest that, in common with MHC class I gene expression, TAP1 and LMP2 gene expression is reduced mainly at the level of transcription in Ad12-transformed cells.
Assuntos
Transportadores de Cassetes de Ligação de ATP/genética , Adenovírus Humanos/genética , Cisteína Endopeptidases , Regulação para Baixo , Regulação da Expressão Gênica , Complexo Principal de Histocompatibilidade , Regiões Promotoras Genéticas , Proteínas/genética , Membro 2 da Subfamília B de Transportadores de Cassetes de Ligação de ATP , Animais , Sequência de Bases , Linhagem Celular Transformada , Clonagem Molecular , DNA Complementar , Expressão Gênica , Humanos , Dados de Sequência Molecular , Oncogenes , RNA Mensageiro , Ratos , Ratos WistarRESUMO
The potential for transfer of antibiotic resistance genes from genetically modified (GM) plant material to microbes through genetic recombination in the human or animal gut is a consideration that has engendered caution in the use of GM foods. This study was aimed at defining the optimal physical and chemical conditions necessary to ensure sufficient fragmentation of DNA in plant tissues to a size where it would be unlikely to be stably transferred to bacterial gut microflora. The ribulose 1,5-bisphosphate carboxylase/oxygenase small subunit (Rubisco SS) genes are of similar size (approximately 1.4 kb) to transgenes present in GM plants. DNA analysis and PCR amplification of Rubisco SS genes showed that fresh maize and maize silage contained high molecular weight DNA and intact Rubisco SS genes. Relatively high temperatures and pressurised steam were necessary to degrade fully genomic DNA and Rubisco SS genes in maize and wheat grains, the source of most animal feedstuffs. Furthermore, chemical expulsion and extrusion of oilseeds resulted in residues with completely degraded genomic DNA. These results imply that stringent conditions are needed in the processing of GM plant tissues for feedstuffs to eliminate the possibility of transmission of transgenes.
Assuntos
DNA Recombinante/análise , Técnicas de Transferência de Genes , Plantas Comestíveis/genética , Plantas Geneticamente Modificadas/genética , Transgenes/genética , Fragmentação do DNA , DNA Bacteriano/análise , DNA Bacteriano/química , DNA Bacteriano/genética , DNA de Plantas/análise , DNA de Plantas/química , DNA de Plantas/genética , DNA de Plantas/metabolismo , DNA Recombinante/química , DNA Recombinante/genética , DNA Recombinante/metabolismo , Resistência Microbiana a Medicamentos , Manipulação de Alimentos , Genes de Plantas/genética , Engenharia Genética , Temperatura Alta , Pressão Hidrostática , Peso Molecular , Reação em Cadeia da Polimerase , Recombinação Genética/genética , Ribulose-Bifosfato Carboxilase/genética , Segurança , Transformação Genética/genética , Triticum/genética , Zea mays/genéticaRESUMO
Infection of mouse BALB/c 3T3 cells by adenovirus 5 resulted in at least 1000-fold lowered yields of virus compared to human cells. The molecular basis of this restriction was analysed at the level of viral gene expression. Steady-state levels of viral DNA and RNA were greatly reduced in infected mouse, compared to human cells. Both early region 1A (E1A) and E1B mRNAs were decreased in mouse cells and their protein products were barely detectable by metabolic labelling of infected cells. The E2A-72 kDa protein and the hexon protein were detected by metabolic labelling, and immunocytochemical analysis showed that they were correctly located in nuclei of infected mouse cells. Only a minor proportion of infected mouse 3T3 cells expressed the E2A-72 kDa or hexon proteins. Low yields of virus were obtained by infection of SV40 transformed BALB/c 3T3 cells showing that SV40 does not provide a helper function for adenovirus 5 growth in this cell system.
Assuntos
Adenovírus Humanos/fisiologia , Adenovírus Humanos/classificação , Animais , Linhagem Celular , DNA Viral/metabolismo , Vírus Auxiliares/fisiologia , Humanos , Camundongos , RNA Viral/metabolismo , Vírus 40 dos Símios/fisiologia , Especificidade da Espécie , Proteínas Virais/metabolismo , Replicação ViralRESUMO
Gene therapy strategies based on modifying tumour cells using high efficiency adenoviral vectors have shown promise in the clinic. Recently the Coxsackie and adenovirus receptor (CAR) has been shown to mediate adenoviral entry into tumour cells, although previous studies also suggested a role for MHC class I heavy chain. Detailed evaluation of the expression of both CAR and MHC class I in prostate cancer cell lines would have important implications for therapeutic strategies. We have found that, unlike cell lines derived from other malignancies, in human and murine prostate cancer loss of CAR expression appears to be relatively infrequent and does not correlate with loss of MHC class I expression. These findings, together with the demonstration of appreciable levels of cell-surface expression of integrins, suggest that cancer vaccine strategies based on modifying whole prostate cancer cells should be feasible using the current generation of recombinant adenoviral vectors, without deleterious effects on either the virus vector or the target cell.
Assuntos
Genes MHC Classe I/fisiologia , Terapia Genética , Integrinas/biossíntese , Neoplasias da Próstata/metabolismo , Receptores Virais/biossíntese , Adenoviridae/genética , Animais , Proteína de Membrana Semelhante a Receptor de Coxsackie e Adenovirus , Citometria de Fluxo , Terapia Genética/métodos , Vetores Genéticos , Proteínas de Fluorescência Verde , Humanos , Imuno-Histoquímica , Integrinas/genética , Proteínas Luminescentes/biossíntese , Proteínas Luminescentes/genética , Masculino , Neoplasias da Próstata/terapia , Ratos , Receptores Virais/genética , Transdução Genética , Células Tumorais CultivadasRESUMO
A simple polymerase chain reaction (PCR) assay for detecting enteric adenoviruses (Ads 40 and 41) in faecal extracts is described. A pair of PCR primers designed to hybridise to the EIB genes of Ad40 and 41 was found to amplify only Ad40 and 41 DNA but not EIB genes or viral DNA from representative numbers of the other human adenovirus subgenera. The PCR assay was tested on a panel of 10 faecal extracts, all of which contained adenovirus particles (as judged by electron microscopy) but only four of which proved amenable to serotyping. Extracts in which enteric adenoviruses had been detected serologically yielded positive results in the PCR assay. These results suggest that this PCR assay may be useful for detecting enteric adenoviruses in clinical samples.
Assuntos
Infecções por Adenovirus Humanos/diagnóstico , Adenovírus Humanos/isolamento & purificação , Fezes/microbiologia , Enteropatias/diagnóstico , Reação em Cadeia da Polimerase/métodos , Adenovírus Humanos/genética , Sequência de Bases , Primers do DNA , DNA Viral/análise , DNA Viral/isolamento & purificação , Humanos , Dados de Sequência MolecularRESUMO
This study was done to determine whether Elizur's anxiety scoring (AL) for Rorschach content was correlated to scores on Spielberger's State-Trait Anxiety Inventory (STAI). The definition of anxiety presented by Elizur implies that his technique measures anxiety as a long-term, relatively stable personality characteristic rather than a transitory emotional state, but no research has shown whether AL was correlated with state and/or trait anxiety as defined and measured by Spielberger. The State-Trait Anxiety Inventory was administered in small groups to 40 college students with a repeated measure of STAI State-anxiety and the Rorschach given individually following a delay of at least five days. Analysis indicated that the STAI Trait-anxiety measure correlated significantly with AL. Test-retest correlations for STAI State-anxiety measures and STAI State- and Trait-anxiety measures obtained in the same testing session were significant. State-anxiety scores obtained just prior to Rorschach testing were related to STAI Trait-anxiety scores and initial STAI State-anxiety scores correlated with AL.
Assuntos
Ansiedade/psicologia , Inventário de Personalidade , Teste de Rorschach , Adolescente , Adulto , Feminino , Humanos , Masculino , Pessoa de Meia-Idade , PsicometriaRESUMO
The subjects, 60 undergraduate students, were administered the Test of Nonverbal Intelligence (TONI) individually. The Shipley Institute of Living Scale was administered in small groups. A Pearson correlation of .56 was obtained for TONI Quotients, Forms A and B. TONI Quotients, Forms A and B, correlated with Shipley estimated WAIS--R IQ .50 and .46, respectively, and correlated to .71 and .64, with Shipley Total T scores, .52 and .44, respectively (corrected to .71 and .61), with Shipley Abstraction T scores, .51 and .42, respectively (corrected, .63 and .52), and with Shipley Vocabulary T scores .26 and .32, respectively (corrected to .63 and .52). TONI scores seem more closely related to Shipley Total and Abstraction scores than to Shipley Vocabulary.
Assuntos
Logro , Gestos , Testes de Inteligência , Cinésica , Comportamento Verbal , Adolescente , Adulto , Feminino , Humanos , Masculino , Pessoa de Meia-Idade , Resolução de Problemas , PsicometriaRESUMO
Transduction of 11 pancreatic cancer cell lines with a replication-deficient adenovirus 5 expressing enhanced green fluorescent protein (Ad5EGFP) was analyzed and variable EGFP levels were observed, ranging from <1% to â¼40% of cells transduced, depending on the cell line. Efficient Ad5EGFP transduction was associated mainly with higher levels of cell surface Coxsackie and adenovirus receptor (CAR) but not with expression of α(v)ß(3) and α(v)ß(5) integrins and was fiber dependent. Reduction of CAR by RNA interference resulted in a corresponding decrease in Ad5EGFP transduction. Pre-treatment of Ad5EGFP with blood coagulation Factor X increased virus entry even in the presence of low CAR levels generated by RNA interference, suggesting a potential alternative route of Ad5 entry into pancreatic cancer cells. Immunohistochemistry carried out on 188 pancreatic ductal adenocarcinomas and 68 matched controls showed that CAR was absent in 102 (54%) of adenocarcinomas, whereas moderate and strong staining was observed in 58 (31%) and 28 (15%) cases, respectively. Weak or absent CAR immunolabeling correlated with poor histological differentiation of pancreatic cancer. In normal tissue, strong immunolabeling was detected in islet cells and in the majority of inter- and intralobular pancreatic ducts.