Photoaffinity labeling of aldehyde dehydrogenase from horse liver by P1-N6-(4-azidophenylethyl)adenosine-P2-[4-(3-azidopyridinio)butyl] diphosphate.
Explore the source record for details and available documents.
Affinity Labels↗
Biomedical subjects
S Becker
Publications and source records attributed to S Becker.
Correlation of the expression of double-stranded RNA-dependent protein kinase (p68) with differentiation in head and neck squamous cell carcinoma.
p68 is an inducible protein kinase which is believed to be an important factor in the regulation of both viral and cellular protein synthesis. We have produced a monoclonal antibody (TJ4C4) which specifically detects p68, and which can be used to detect this antigen in formalin-fixed, paraffin-embedded tissues. Because p68 plays an important role in cellular protein synthesis, we hypothesized that it may correlate with normal and neoplastic cellular differentiation. One hundred and seventy-seven head and neck squamous cell carcinoma specimens, representing 82 patients, were studied. The relative amount, frequency, and distribution of p68 expression were determined by microscopic evaluation of ABC immunoperoxidase-stained specimens. A spectrum of immunoreactivity was detected in 156 of 177 tumors, as well as within the normal squamous epithelium. Normal, actively proliferating cells, such as the basal layer of squamous epithelium, expressed comparatively little p68. Increased p68 expression was noted to parallel the morphologic features of cellular differentiation. In neoplastic tissue, p68 expression also increased with the degree of cellular differentiation. These data demonstrate that the expression of p68 parallels the degree of cellular differentiation in squamous cell carcinoma of the head and neck region, as well as within normal squamous mucosa. Therefore, p68 may provide an objective biologic measure of cellular differentiation which does not depend on morphologic features.
Aged↗
dcp gene of Escherichia coli: cloning, sequencing, transcript mapping, and characterization of the gene product.
Dipeptidyl carboxypeptidase is a C-terminal exopeptidase of Escherichia coli. We have isolated the respective gene, dcp, from a low-copy-number plasmid library by its ability to complement a dcp mutation preventing the utilization of the unique substrate N-benzoyl-L-glycyl-L-histidyl-L-leucine. Sequence analysis of a 2.9-kb DNA fragment revealed an open reading frame of 2,043 nucleotides which was assigned to the dcp gene by N-terminal amino acid sequencing and electrophoretic molecular mass determination of the purified dcp product. Transcript mapping by primer extension and S1 protection experiments verified the physiological significance of potential initiation and termination signals for dcp transcription and allowed the identification of a single species of monocistronic dcp mRNA. The codon usage pattern and the effects of elevated gene copy number indicated a relatively low level of dcp expression. The predicted amino acid sequence of dipeptidyl carboxypeptidase, containing a potential zinc-binding site, is highly homologous (78.8%) to the corresponding enzyme from Salmonella typhimurium. It also displays significant homology to the products of the S. typhimurium opdA and the E. coli prlC genes and to some metalloproteases from rats and Saccharomyces cerevisiae. No potential export signals could be inferred from the amino acid sequence. Dipeptidyl carboxypeptidase was enriched 80-fold from crude extracts of E. coli and used to investigate some of its biochemical and biophysical properties.
Amino Acid Sequence↗
Respiratory syncytial virus-induced cytokine production by a human bronchial epithelial cell line.
Respiratory syncytial virus (RSV) is the most common cause of lower respiratory infection in infants and young children, but the pathogenesis of RSV-induced inflammation is not well defined. We hypothesized that in vitro infection of a human bronchial epithelial cell line (BEAS) would induce production of proinflammatory cytokines. BEAS cells were infected with RSV, and cells and supernatants were assayed for cytokine mRNA and protein changes at several time points after infection. Cytokine mRNA in BEAS cells was measured by polymerase chain reaction of reverse-transcribed RNA from whole cell lysates; cytokine levels in supernatants were measured by bioassay or immunoassay. Our results indicated that interleukin-5ay or immunoassay. Our results indicated that interleukin-8 (IL-8) was induced at 4 h after infection (during the eclipse phase of RSV infection) with accumulation of IL-8 in supernatants by 24 h after infection. Increased levels of IL-6 and granulocyte macrophage colony-stimulating factor in supernatants were only detected by 96 h after infection, during the RSV replicative phase. Interferon-alpha and -gamma transcripts were not detectable at any time point. We conclude that the effects of RSV on airway inflammation may be at least partly mediated by sequential production of proinflammatory cytokines in infected airway epithelium.
Base Sequence↗
Effect of storage temperature on the activity of superoxide dismutase, catalase, glutathione peroxidase, glutathione reductase and glutathione S-transferase in rat liver and kidney homogenates.
The behavior of the catalytic activities of the enzymes superoxide dismutase, catalase, glutathione peroxidase, glutathione S-transferase and glutathione reductase was tested in rat liver and kidney homogenates stored at 4, -20 and -70 degrees C and in corresponding tissue samples stored at -70 degrees C. The stabilities of enzymes were different for various enzymes and were dependent on the organ (liver, kidney) and the storage temperature. The storage temperature of -70 degrees C guaranteed the best stability and the five enzymes investigated were sufficiently stable in preserved tissue samples or in homogenates prepared with conventional mannit/sucrose homogenization solution. Under such conditions, these enzymes were stable over at least 4 or 1 weeks, respectively.
Animals↗
Interleukin-8 expression in normal nasal epithelium and its modulation by infection with respiratory syncytial virus and cytokines tumor necrosis factor, interleukin-1, and interleukin-6.
Inflammation in nasal and airway tissue caused by allergens, microbial infection, and air pollution are likely to be regulated by inflammatory mediators produced by airway epithelial cells. We have therefore investigated the baseline expression of a number of cytokine genes known to be important inducers and modulators of inflammation, in freshly isolated human nasal epithelium. Cells were obtained by superficial scraping of turbinate tissue, and cDNA for polymerase chain reaction (PCR) amplification was reverse-transcribed directly from lysates of 3 x 10(3) to 5 x 10(3) epithelial cells using random hexamers. Constitutive expression of relatively high levels of interleukin-8 (IL-8) mRNA but undetectable levels (< 1 mRNA copy/cell) of granulocyte/macrophage colony-stimulating factor (GM-CSF), IL-6, IL-1, or tumor necrosis factor (TNF) mRNA were found after PCR amplification of the cDNA. IL-8 protein, but not IL-6, was identified in the nasal epithelial cells by immunocytochemistry. Infection with respiratory syncytial virus (RSV) or stimulation of nasal epithelium for 4 h with TNF or IL-1 in vitro resulted in a 4- to 10-fold increase in IL-8 mRNA expression but not in the expression of detectable levels of mRNA for the other cytokines. IL-8 was secreted by RSV-, IL-1-, and TNF-stimulated as well as unstimulated nasal epithelial cells after 6 to 20 h of culture. Neither IL-6, GM-CSF, nor TNF activity/immunoreactivity was detectable in the culture supernatants. Thus, it appears that IL-8 is a major cytokine of human nasal epithelium, constitutively expressed and readily secreted upon virus infection or stimulation with IL-1 and TNF.
Adult↗
Chemical nature and immunotoxicological properties of arachidonic acid degradation products formed by exposure to ozone.
Ozone (O3) exposure in vivo has been reported to degrade arachidonic acid (AA) in the lungs of rodents. The O3-degraded AA products may play a role in the responses to this toxicant. To study the chemical nature and biological activity of O3-exposed AA, we exposed AA in a cell-free, aqueous environment to air, 0.1 ppm O3, or 1.0 ppm O3 for 30-120 min. AA exposed to air was not degraded. All O3 exposures degraded > 98% of the AA to more polar products, which were predominantly aldehydic substances (as determined by reactivity with 2,4-dinitrophenylhydrazine and subsequent separation by HPLC) and hydrogen peroxide. The type and amount of aldehydic substances formed depended on the O3 concentration and exposure duration. A human bronchial epithelial cell line (BEAS-2B, S6 subclone) exposed in vitro to either 0.1 ppm or 1.0 ppm O3 for 1 hr produced AA-derived aldehydic substances, some of which eluted with similar retention times as the aldehydic substances derived from O3 degradation of AA in the cell-free system. In vitro, O3-degraded AA induced an increase in human peripheral blood polymorphonuclear leukocyte (PMN) polarization, decreased human peripheral blood T-lymphocyte proliferation in response to mitogens, and decreased human peripheral blood natural killer cell lysis of K562 target cells. The aldehydic substances, but not hydrogen peroxide, appeared to be the principal active agents responsible for the observed effects. O3-degraded AA may play a role in the PMN influx into lungs and in decreased T-lymphocyte mitogenesis and natural killer cell activity observed in humans and rodents exposed to O3.
Aldehydes↗
Rapid screening of low molecular mass proteinuria: evaluation of the first immunochemical test strip for the detection of alpha 1-microglobulin in urine.
A new semiquantitative immunochemical test strip for urinary alpha 1-microglobulin, a marker protein for tubular proteinuria, was assessed. This test strip has four colour zones, reflecting alpha 1-microglobulin concentrations of ca. 10, 25, 50, and 80 mg/l. alpha 1-Microglobulin concentrations were measured by means of the test strip and an immunonephelometric method in 330 samples collected as the second voided morning urine. The reading time of the test strip must be strictly observed. Reading one minute earlier or later than the 5 min stated in the instructions led to misclassification of over 70% of the results. Correlation between both methods was highly significant, with a Spearman rank correlation coefficient of rs = 0.84 (P < 0.001). There was a partial overlap of the test strip results in different concentration ranges. An elevation of alpha 1-microglobulin was defined as > 25 mg/l, calculated as the upper limit of the central 95% interval of alpha 1-microglobulin concentration in urine samples measured in a previous study of 304 healthy adults. Using this definition of alpha 1-microglobulin elevation, a sensitivity of 97.5%, specificity of 73.6%, a false-positive rate of 16.6%, and a false-negative rate of 0.9% of the test strip results were obtained. A fraction of 82.4% of the 330 samples investigated was correctly classified as having increased alpha 1-microglobulin concentration or not. Methodical improvements of the test strip are necessary to reduce overlapping results, in order to make the test suitable for screening purposes.
Humans↗
Action of derivatives of mu-conotoxin GIIIA on sodium channels. Single amino acid substitutions in the toxin separately affect association and dissociation rates.
We have studied binding and block of sodium channels by 12 derivatives of the 22-residue peptide mu-conotoxin GIIIA (mu-CTX) in which single amino acids were substituted as follows: Arg or Lys by Gln, Gln-18 by Lys, Asp by Asn, and HO-Pro by Pro. Derivatives were synthesized as described by Becker et al. [(1989) Eur. J. Biochem. 185, 79]. Binding was measured by displacement of labeled saxitoxin from eel electroplax membranes (100 mM choline chloride, 10 mM HEPES-NaOH, pH 7.4). Blocking kinetics were evaluated from steady-state, single-channel recordings from rat skeletal muscle sodium channels incorporated into planar, neutral phospholipid/decane bilayers (200 mM NaCl, 10 mM HEPES-NaOH, pH 7.0). Blocking events generally appeared as periods of seconds to minutes in which current through the single channel was completely eliminated. A notable exception was seen for the substitution Arg-13-Gln for which the "blocked" events showed measurable conductances of about 20-40% of the open state. The substitution of Arg-13 reduced binding to electroplax membranes to undetectable levels and increased the apparent dissociation constant determined for skeletal muscle channels by greater than 80-fold compared with the native peptide. Other substitutions caused smaller decreases in affinity. The decreased potency of the toxin derivatives resulted both from increases in the rates of dissociation from the channel, and from decreases in association rates. Our data support the suggestion by Sato et al. [(1991) J. Biol. Chem. 265, 16989] that Arg-13 associates intimately with the binding site on the channel. In addition, our results suggest that certain residues affect almost exclusively the approach and docking of the toxin with its binding site, others appear to be important only to the strength of the association once binding has taken place, and yet others affect both.
Amino Acid Sequence↗
Self-organizing neural network that discovers surfaces in random-dot stereograms.
The standard form of back-propagation learning is implausible as a model of perceptual learning because it requires an external teacher to specify the desired output of the network. We show how the external teacher can be replaced by internally derived teaching signals. These signals are generated by using the assumption that different parts of the perceptual input have common causes in the external world. Small modules that look at separate but related parts of the perceptual input discover these common causes by striving to produce outputs that agree with each other. The modules may look at different modalities (such as vision and touch), or the same modality at different times (for example, the consecutive two-dimensional views of a rotating three-dimensional object), or even spatially adjacent parts of the same image. Our simulations show that when our learning procedure is applied to adjacent patches of two-dimensional images, it allows a neural network that has no prior knowledge of the third dimension to discovery depth in random dot stereograms of curved surfaces.
Artificial Intelligence↗
Differences in arachidonic acid metabolism by human myelomonocytic cell lines.
The production of arachidonic acid metabolites by the HL60, ML3, and U937 human phagocyte cell lines was determined after incubation with interferon-gamma (IFN-gamma, 500 U/ml) or vehicle for 4 days. Cells were prelabeled with tritiated arachidonic acid, [3H]AA, for 4 h, and media supernatants were analyzed by high-performance liquid chromatography. None of the cell lines produced [3H]AA metabolites in large amounts during an unstimulated, basal release period (30 or 60 min). In response to 10 microM calcium ionophore A23187 incubation (30 min), undifferentiated and IFN-gamma-differentiated HL60 cells formed both cyclooxygenase products (thromboxane and prostaglandins) and lipoxygenase products (leukotrienes and hydroxyeicosatetraenoic acids). In contrast to the HL60 cells, IFN-gamma-differentiated U937 cells formed primarily cyclooxygenase products and undifferentiated and IFN-gamma-differentiated ML3 cells did not form any [3H]AA metabolites in response to A23187. These results indicate the need to be careful in selecting a cell line for use in a phagocyte assay system when cyclooxygenase and/or lipoxygenase products could influence the assay results.
Antigens, CD↗
Evidence for occurrence of filovirus antibodies in humans and imported monkeys: do subclinical filovirus infections occur worldwide?
In the present serological study 120 monkey sera from different species originating from the Philippines, China, Uganda and undetermined sources and several groups of human sera comprising a total of 1288 specimens from people living in Germany were examined for the presence of antibodies directed against filoviruses (Marburg virus, strain Musoke/Ebola virus, subtype Zaire, strain Mayinga/Reston virus). Sera were screened using a filovirus-specific enzyme-linked immunosorbent assay (ELISA). ELISA-positive sera were then confirmed by the indirect immunofluorescence technique, Western blot technique, and a blocking assay, and declared positive when at least one confirmation test was reactive. Altogether 43.3% of the monkey sera and 6.9% of the human sera reacted positively with at least one of the three different filovirus antigens. The blocking assays show that antibodies, detected in the sera, are directed to specific filovirus antigens and not caused by antigenic cross-reactivity with hitherto unknown agents. Data presented in this report suggest that subclinical filovirus infections may also occur in humans and in subhuman primates. They further suggest that filoviruses are not restricted to the African continent.
Animals↗
Radiotherapy of the rhabdomyosarcoma R1H of the rat: recovery from radiation injury in tumour and skin.
The response of the rhabdomyosarcoma R1H of the rat to fractionated irradiation has been reported to be almost independent of the dose per fraction in the range from 0.43 to 10 Gy. To elucidate whether recovery from radiation injury is impaired in fractionated irradiation or not, tumours were exposed on 5 days per week over 6 weeks either to a single daily dose or to two daily fractions separated by an interfraction interval of 2 h. Tumour response was assessed by net growth delay. Skin damage was scored for comparison. A significant reduction in tumour response (p less than 0.009) and skin damage (p less than 0.001) was observed when the daily dose was administered in a twice daily irradiation regimen, indicating that recovery from radiation injury does occur in the course of fractionated irradiation in our tumour system. A tumour response was almost independent of the dose per fraction for time intervals between fractions of at least 8 h, a possible explanation might be that recovery is compensated by improved reoxygenation.
Animals↗
Localization of endoderm-specific mRNAs in differentiating F9 embryoid bodies.
Primitive endoderm in the peri-implantation mouse embryo differentiates into two separate lineages, visceral and parietal endoderm (VE and PE). F9 teratocarcinoma cells, when grown in suspension in the presence of retinoic acid (RA), differentiate into embryoid bodies (EBs) which can be used as a model system to study the spatially appropriate induction of VE- and PE-specific gene expression. We have used a whole mount in situ hybridization technique to follow the localization of VE-specific AFP and PE-specific tPA mRNAs during EB differentiation. We show that the putative endoderm-specific markers are localized to the endoderm in mature EBs, and that AFP mRNA is localized to the apical edge of the VE in RA EBs. Surprisingly, prior to localization of endoderm-specific markers at the periphery of EBs, these genes are expressed at low to moderate levels in all cells of the EB. Our data suggest that the establishment of endoderm in EBs is position dependent and not the result of sorting out of predetermined, randomly distributed cells. Our observations also suggest a two-step process for establishing endoderm-specific gene expression, involving up-regulation of transcripts throughout the EB prior to the restriction of their expression to the outer differentiating cell layer.
Animals↗
[Heroin abuse and methadone substitution in pregnancy].
The pre- and postnatal data of 20 pregnant heroin addicts and their neonates are described. 16 women were treated with methadone to prevent withdrawal symptoms. A relatively stabile prenatal condition with a decrease of complications was achieved. On the other hand, the neonates suffered from severe withdrawal symptoms including seizures in spite of intensive paediatric care and prophylactic treatment with barbiturates. After a mean follow-up of one to two years a relatively good neurological development of the children was observed.
Asphyxia Neonatorum↗
Diet-induced dermatitis response of hairless rats to systemic treatment with cyclosporin A (Sandimmun), cyclosporin H and FK506.
Weaned hairless rats were fed a diet deficient in fat, magnesium and folacin. After approximately 1 week, an erythematous dermatitis developed which was associated with extreme generalized pruritus. Scratching led to excoriations and hemorrhagic crusting. The acute stage (pruritic rash) resolved after several days and was followed by sporadic non-itching relapses. Subsequent to the onset of symptoms, rats were treated orally, once daily for 3 days with CyA, CyH or FK506. The immunosuppressants CyA and FK506 caused a dose-dependent inhibition of symptoms in contrast to CyH. The immediate clinical response was associated with changes in blood histamine, white blood cell counts and histological parameters. Since CyH is known to lack immunosuppressive activity, these results may indicate that the cutaneous changes induced by the nutritional deficiency are associated with immunological abnormalities. The results may also indicate mechanisms influenced by CyA and FK506 but not by CyH; for example, release of chemical mediators from inflammatory cells.
Animals↗
Location of the dcp gene on the physical map of Escherichia coli.
Explore the source record for details and available documents.
Chromosomes, Bacterial↗