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At least 1,639 records · Page 91Linked to original sources
PubMed1997

Immunolocalization of Drosophila eye-specific diacylgylcerol kinase, rdgA, which is essential for the maintenance of the photoreceptor.

The Drosophila retinal degeneration A (rdgA) mutant has photoreceptor cells that degenerate within a week after eclosion. The degeneration starts with the disruption of the subrhabdomeric cisternae (SRC), which are the organelles essential for the transport of phospholipids to the photoreceptive membranes. Our previous biochemical and molecular studies suggested that the rdgA gene encodes an eye-specific diacylglycerol kinase (DGK). In this study, we show that retinal degeneration is prevented by the introduction of the eye-DGK gene in the rdgA mutant genome, suggesting that the DGK activity is crucial for the maintenance of the photoreceptor. Furthermore, by immunohistochemical analysis, we have demonstrated that the rdgA protein is predominantly associated with the SRC, suggesting that the conversion from diacylglycerol (DG) to phosphatidic acid (PA) most actively occurs in SRC. The analysis of the eyes of mutants homozygous for rdgA and eye-protein kinase C mutations indicates that retinal degeneration is caused by the deficiency of PA rather than excessive accumulation of DG. From these data, we conclude that the production of PA in the SRC membranes is essential for the maintenance of the photoreceptor.

Animals↗
PubMed1988

Bone marrow origin of a B-cell lymphoma.

To search for precursors of the neoplastic B cells in a patient with a nodular lymphoma, we produced a monoclonal antibody to a variable region idiotope on the lymphoma IgM heavy chain. Clonal ancestors of the lymphoma cells were identified by this marker among bone marrow pre-B cells (5% to 26%). A second antiidiotype (anti-Id) antibody specific for the complete lymphoma IgM kappa recognized 10% of B cells in bone marrow and blood and greater than 95% of B cells in lymphomatous lymph nodes, including one obtained after tumor conversion to a diffuse large cell lymphoma. Immunoglobulin gene analysis surprisingly revealed expansion of multiple clones of early B lineage cells in bone marrow, including members of the neoplastic clone. The data suggest that this lymphoma arose through a progression of transformational events beginning in bone marrow: first, creation of an oligoclonal pre-neoplastic pool of pre-B cells, subsequent conversion of a single subclone into low grade neoplastic B cells that homed to the lymph node follicles, and later progression to a more invasive form of the B-cell lymphoma.

Adult↗
PubMed2002

Use of a computerized tuberculosis register for automated generation of case finding, sputum conversion, and treatment outcome reports.

SETTING: Tuberculosis (TB) rates in southern Africa have increased dramatically in recent years. Provision of accurate data for surveillance, program management, and supervision is increasingly essential. OBJECTIVE: To develop software that would provide more efficient collection, compilation, and analysis of TB data on an ongoing basis. DESIGN: The 'Electronic TB Register' is a user-friendly, Epi-Info based software program based on the WHO/IUATLD format of recording and reporting. Individual records from the TB registry are entered in a program that provides interactive support. The software provides several patient management and supervision functions, such as lists of defaulters. Finally, it generates standard quarterly and annual reports on case-finding, sputum conversion, and cohort analysis, and provides graphs of trends and maps of TB indicators. RESULTS: The 'Electronic TB Register' software has been successfully implemented in five pilot projects in southern Africa. User acceptance has been high and quality of data has improved, although timeliness remains unchanged. Factors critical for success include a functioning, paper-based system, involvement of staff from the TB program, health information systems, and health facilities, ongoing training, and backup support. CONCLUSIONS: The 'Electronic TB Register' is a potentially powerful tool for surveillance, management, and supervision for countries with well-functioning paper-based recording and reporting systems.

Confidentiality↗
PubMedDate not supplied

[Immunological and sedimentation analysis of UV-irradiated T4 phage DNA containing 5-bromouracil].

Conversions resulting from UV-irradiation of 5-bromouracil DNA of T4 phage were studied. The use of immunological and sedimentation techniques revealed that UV dose ranging from 1.4.10(2) to 1.4.10(3) erg/mm2 induced unwinding and single-stranded breaks in 5-bromouracil DNA which are not accompanied by debromination. Beginning with 4.10(3) erg/mm2 debromination but not degradation of DNA is detected. Doses higher than 4.10(3) erg/mm2 lead to extensive degradation of 5-bromouracil T4 DNA.

Bromouracil↗
PubMed2006

InSilicoSpectro: an open-source proteomics library.

We present a new proteomics open-source project, InSilicoSpectro, aimed at implementing recurrent computations that are necessary for proteomics data analysis. Illustrative examples are mass list file format conversions, protein sequence digestion, theoretical peptide and fragment mass computations, graphical display, matching with experimental data, isoelectric point estimation, and peptide retention time prediction. The project library is written in Perl, a widely used scripting language in bioinformatics, and it offers a unique framework of integrated objects to implement complex proteomics data analyses. For instance, only a few lines of code are required to digest a protein with fixed and variable modifications, label peptides with 18O, compute the fragmentation spectra and display their match with experimental spectra. We believe that InSilicoSpectro will be of great help to bioinformaticians, without detailed knowledge of proteomics specifics, and to mass spectrometrists with computer programming interest as well.

Amino Acid Sequence↗
PubMed2001

Regulation of myogenic terminal differentiation by the hairy-related transcription factor CHF2.

We have recently cloned a novel basic helix-loop-helix factor, CHF2, that functions as a transcriptional repressor. To address its role in the regulation of myogenic terminal differentiation, we analyzed its expression pattern during C2C12 mouse myotube formation. In undifferentiated myoblasts, CHF2 is expressed at high levels. After induction of myotube formation in low serum, CHF2 expression is barely detectable at 3 days after induction. Myogenin expression, in contrast, peaks at 3 days. In transiently transfected 10T1/2 embryonic fibroblasts, CHF2 inhibited MyoD-dependent activation of the myogenin promoter in a dose-dependent fashion. Electrophoretic mobility shift analysis indicated that CHF2 inhibits the binding of the MyoD.E47 heterodimer to the E-box binding site. CHF2 also inhibited myogenic conversion of 10T1/2 cells by MyoD, as measured by skeletal myosin heavy chain protein expression. Coimmunoprecipitation analysis indicated that CHF2 forms a protein complex with MyoD. Mutational analysis of CHF2 indicated that the repression activity for both transcription and myogenic conversion mapped to a hydrophobic carboxyl-terminal region and did not require either the basic helix-loop-helix or YRPW motifs. Our data indicate that CHF2 functions as a transcriptional repressor of myogenesis by formation of an inactive heterodimeric complex with MyoD and likely plays an important role in muscle development.

Animals↗
PubMed1997

Polymorphonuclear leukocyte cells and elastase in tears.

PURPOSE: To characterize the effects that mode of sampling and overnight eye closure have on the nature of caseinolytic activity recovered in tear fluid. METHODS: Reflex, open and closed (R, O and C) eye tear fluids were collected by microcapillary tubes or from the inferior formix by Schirmer strip. Microcapillary collected samples were centrifuged and recovered cells cytochemically characterized and probed by immunofluorescence microscopy, or alternatively extracted in acidic PBS. Tear supernatants, pellets and Schirmer strip extracts were subjected to casein zymography or SDS-PAGE and immunoprobed for plasmin/plasminogen. To identify caseinolytic activity, samples were immunoprecipitated with antibodies to plasmin/plasminogen or to elastase, and the immunoprecipitated materials were subjected to zymographic analysis. RESULTS: Immunoblot assays revealed R and O samples contained low levels of plasminogen (approximately 1.1 micrograms/ml) and only trace levels of plasmin (< 0.1 ng/ml). Insufficient levels of caseinolytic activity were present to allow zymographic detection. Cytochemical analysis revealed that R and O pellets consisted almost exclusively of desquamated epithelium. Immunoblot analysis revealed that C fluid was associated with an increase in plasminogen and its partial conversion to plasmin (approximately 3.2 ng/ microliter), high molecular weight covalent complexes and degradative products. Zymographic analysis disclosed much greater caseinolytic activity than could be attributed to plasmin or its cleavage products. This consisted primarily of three bands (30-26 kDa) which were identified as polymorphonuclear leukocyte (PMN) cell elastase based on size and antigenicity. This is derived from PMNs recovered from the C pellet. Elastase could also be recovered from Schirmer strips from 90% of donors, provided that the strips were extracted in sample loading buffer. The activity was restricted to the portion of the strip that had been in contact with the ocular tissue. CONCLUSIONS: The main source of caseinolytic activity in C fluid is elastase. This arises from PMNs that undergo recruitment, activation and degranulation in the C environment. In contrast, the elastase recovered in Schirmer strip extracts is derived from intact PMNs that adhere to the strip during sample collection. This would suggest that PMN cells undergo a low level of recruitment into the open eye environment.

Adolescent↗
PubMed2000

Transcutaneous electrical nerve stimulation (TENS): its short-term and long-term effects on the masticatory muscles.

In an electromyographic study on subjects with no functional disturbances of the masticatory muscles, the duration of the post-therapeutic effects of transcutaneous electrical nerve stimulation (= TENS) on the superficial masseter and anterior temporal muscle was analyzed. The myoelectric signals were registered from 20 healthy volunteers in 3 different mandibular positions. The recordings were performed before a 20-minute TENS application with the J-4 Myomonitor and continued with a sequence of follow-up registrations with increasing interval to the initial stimulation. The EMG signals underwent computer-aided analysis and were evaluated by determining the integrated values as a parameter of muscle activity, and after Fourier transformation by 7 describing parameters of the power spectrum (e.g. mean power frequency = MPF). A detailed analysis of variance of all data was used to investigate significant changes of the parameters during the observation period. Muscular response to TENS includes a decrease in muscular activity (= reduction in integrated EMG signals) and a shift in the power spectrum to higher frequencies (increase in MPF). These changes were statistically highly significant for both analyzed muscles and for all different mandibular exercises. As these reactions to TENS are contrary to muscle fatigue, the results can be interpreted as indicating that this type of therapy stimulates a change in the biochemical and physiological muscular conditions, which leads to muscle relaxation. Electromyographically, the post-therapeutic effect lasted for 2 hours in case of normal masticatory muscle activity but for more than 7 hours in case of low muscular loading. The alterations of the integrated EMG values were more persistent than those of the parameters of the power spectrum.

Adult↗
PubMedDate not supplied

Measurement of nitric oxide in murine Hepatoma Hepa1c1c7 cells by reversed phase HPLC with fluorescence detection.

PURPOSE: Nitric oxide (NO) is produced by various cell types in picomolar to nanomolar range and has important roles in a variety of biological functions. The aim of the present study was to investigate a sensitive and reproducible fluorometric HPLC method for measuring nitrite, one of the stable oxidation products of nitric oxide, in murine hepatoma Hepa 1c1c7 cells. METHODS: Hepa 1c1c7 cells were incubated with vehicle or recombinant murine tumor necrosis factor-alpha (TNF-alpha, 10 ng/ml) in Hanks' balanced salt solution (HBSS) for 10 hours. Thereafter, the HBSS medium was collected for nitrite analysis. NO production was examined by measuring the conversion of 2, 3-diaminonaphthalene (DAN) to its fluorescent product, 2, 3-naphthotriazole (NAT). NAT was analyzed after elution with 60% of 15 mM sodium phosphate buffer (pH = 7.5) and 40% methanol through a 10-microm reversed-phase C18 column (250 x 4.00 mm, I.D.) at a flow rate of 1 ml /min. Fluorescence was monitored with excitation at 375 nm and emission at 415 nm. RESULTS: NAT appeared in approximately 16 min with no interference peaks. The assay yielded linear response within the examined range of 10 - 200 pM (r(2) >0.99) with an intra and inter-day variability of <10 % and accuracy of > 90%. The detection limit for nitrite was 10 pM. NO production by TNF- alpha treated Hepa 1c1c7 cells is estimated to be approximately 2 folds more than untreated cells. CONCLUSION: This fluorometric HPLC assay offers a sensitive and reliable measurement of NO production in cell culture medium.

Analysis of Variance↗
PubMed1996

Transcriptional upregulation of gamma-globin by phenylbutyrate and analogous aromatic fatty acids.

Phenylbutyrate has been shown recently to induce fetal hemoglobin (HbF) production in patients with sickle cell anemia and beta thalassemia. We have now examined related aromatic fatty acids in order to define the range of active structures and identify plausible mechanisms of action. Structure-function analysis revealed that for effective stimulation of HbF in erythroid precursors: (1) the ideal length for the aliphatic side chain is four carbons; (2) oxygen or sulfur substitutions in the carboxylic chain are allowed, as evidenced by the equal or increased activity of phenoxypropionate, benzylthioglycolate, and benzyloxyacetate compared with phenylbutyrate; and (3) blocking the carboxylate group by conversion to the amide form greatly reduces potency. Molecular analysis indicated that the prototype agent, phenylbutyrate, increases HbF production through transcriptional activation of the gamma-globin gene. The latter contains a butyrate responsive promoter known to up-regulate transcription in the presence of short-chain fatty acids of three to five carbons. To determine whether stimulation of an element in this promoter by phenylbutyrate and its analogues might contribute to their mechanism of action, we used a transient expression system involving K562 erythroleukemia cells transfected with a luciferase reporter gene driven by the minimum gamma-globin promoter. Transcriptional activation in this experimental system correlated well with the capacity of an aromatic fatty acid to increase HbF production in erythroid precursors (r = 0.94). Our studies identify potent analogues of phenylbutyrate for the treatment of beta-chain hemoglobinopathies, and suggest that stimulation of a butyrate responsive promoter may be responsible for their activity.

Cells, Cultured↗
PubMed2000

Use of RAPD-ALF analysis for investigating the frequency of bacterial cross-transmission in an adult intensive care unit.

Bacterial cross-transmission was investigated during a 12-month period in an adult intensive care unit (ICU) by the generation of random amplified polymorphic DNA (RAPD) fingerprinting profiles, combined with automated laser fluorescence (ALF) analysis. The potential episodes of cross-transmission identified, were compared with those detected by the conventional first-line screen of antibiogram typing. Over the year, 215 primary gram-negative bacterial isolates were obtained from 160 patients. In total, 22 possible episodes of cross-transmission, involving 70 (44%) of the 160 patients, were identified by RAPD-ALF analysis, and 19 of these were substantiated with epidemiological evidence. Conversely, 31 possible episodes were identified on the basis of antibiogram data, but only three of these episodes, two involving Acinetobacter baumannii and one involving Serratia marcescens, correlated with those identified by RAPD-ALF analysis. It was concluded that analysis of antibiogram data alone is an unreliable method for assessing bacterial cross-transmission, unless the organism involved has a particularly stable or unusual resistance pattern. In contrast, the technique of RAPD-ALF analysis may provide a rapid and simple technique for obtaining an insight into the population dynamics of gram-negative bacteria in adult ICUs.

Acinetobacter↗
PubMed2001

Treatment of word retrieval in aphasia: generalisation to conversational speech.

The question of whether treatment gains in picture naming generalise to conversation remains relatively unexplored. Several difficulties surround data collection and analysis. A quantitative measure of word retrieval in aphasia is presented along with relevant details relating to the reliability of the measure, and the relationship between word retrieval in picture naming and noun retrieval in conversation. We discuss the clinical application of the measure and its applicability outside the field of aphasia.

Aphasia↗
PubMed1989

Preliminary evaluation of an intra-arterial blood gas system in dogs and humans.

The reliability and accuracy of an intra-arterial fluorescent optode system to measure continuously pHa, PaCO2, and PaO2 were evaluated in a dog model and in a group of critically ill patients. Six hundred sixty-three arterial blood gas (ABG) samples were analyzed for pHa, PaCO2, and PaO2 in the dog studies. The intra-arterial blood gas system (IBGS) indicated a steady state in 420 instances for pH, 359 instances for PaCO2, and 256 instances for PaO2. Comparison of these ABG and IBGS values by linear regression analysis revealed r = .97 for pHa, .95 for PaCO2, and .96 for PaO2. The mean +/- SD of the difference between ABG and IBGS was -0.02 +/- 0.03 for pHa, 1.05 +/- 3.8 for PaCO2, and -17 +/- 13 for PaO2. Nonsteady states were correctly identified by the IBGS in every instance. Comparisons between 79 temporally matched ABG and IBGS values, exclusive of in vivo calibration samples, in 12 critically ill patients revealed r = .97 for pHa, .96 for PaCO2, and .99 for PaO2. The difference was 0.002 +/- 0.02 for pHa, 0.44 +/- 2.97 for PaCO2, and -1.22 +/- 9.34 for PaO2. We conclude that it is possible to measure continuously pHa, PaCO2, and PaO2 with the IBGS in critically ill patients for periods from 3 to 25.5 h while maintaining the ability to monitor BP and withdraw blood samples from the arterial cannula. Agreement between the two techniques is within clinically acceptable ranges for pHa and PaCO2, whereas PaO2 measurement by the IBGS requires further refinement.(ABSTRACT TRUNCATED AT 250 WORDS)

Adolescent↗
PubMed1995

Nitric oxide synthase activity in brain regions and spinal cord of mice and rats: kinetic analysis.

Nitric oxide synthase (NOS) activity was determined by the rate of conversion of [3H]arginine to [3H]citrulline in brain regions (midbrain, hypothalamus, cerebellum, hippocampus, corpus striatum, cortex, pons-medulla and amygdala) and spinal cord of male Swiss Webster mice and male Sprague-Dawley rats. In mice, high activity of NOS was found in cerebellum, hypothalamus and midbrain; intermediate activity in pons-medulla, hippocampus, amygdala, corpus striatum and cortex; and low activity in the spinal cord. In rat, highest activity of NOS was observed in cerebellum followed in decreasing order by midbrain, hypothalamus, cortex, striatum, pons-medulla, hippocampus and spinal cord. In all tissues, NOS activity was higher in rat than in mouse. Analysis of the Eadie-Hofstee plot indicated that the Vmax and Km values of the enzyme in cortex and cerebellum of rats were higher than in mouse tissues. These studies show differential distribution of NOS activity in mouse and rat brain regions and spinal cord and higher activity of NOS in rat brain regions and spinal cord compared to mouse tissues.

Amino Acid Oxidoreductases↗
PubMed1998

Probing the mechanism of proton coupled electron transfer to dioxygen: the oxidative half-reaction of bovine serum amine oxidase.

Bovine serum amine oxidase (BSAO) catalyzes the oxidative deamination of primary amines, concomitant with the reduction of molecular oxygen to hydrogen peroxide via a ping-pong mechanism. A protocol has been developed for an analysis of chemical and kinetic mechanisms in the conversion of dioxygen to hydrogen peroxide. Steady-state kinetics show that two groups need to be deprotonated to facilitate the oxidative half-reaction. The pH dependence of Vmax/Km(O2) reveals pKa's of 6.2 +/- 0.3 and 7.0 +/- 0.2, respectively. A pKa of 7.2 +/- 0.1 has been obtained from a titration of anaerobically reduced BSAO using UV-vis spectrophotometry. The near identity of the pKa obtained from the reduced enzyme titration with the second pKa from steady-state kinetics suggests that this second pKa arises from the reduced cofactor. The assignment of pKa is supported by the observed pH dependence for formation of the cofactor semiquinone signal, detected by EPR spectroscopy under anaerobic conditions. To address the nature of rate-limiting steps in the oxidative half-reaction, the solvent isotope effect, viscosity effect, and O-18 isotope effect on Vmax/Km(O2) have been determined. The solvent isotope effect is indistinguishable from unity, ruling out a proton transfer as a rate-determining step. Use of glucose as a solvent viscosogen shows no viscosity effect, indicating that binding of oxygen is not in the rate-determining step. The O-18 kinetic isotope effect is independent of pH with an average value of 18(V/K) = 1.0097 +/- 0. 0010. This has been compared to calculated equilibrium O-18 isotope effects for various dioxygen intermediate species [Tian and Klinman (1993) J. Am. Chem. Soc. 115, 8891], leading to the conclusion that either the first electron transfer to dioxygen or the desorption of product peroxide from a Cu(II)-OOH complex could be the rate-limiting step. The distribution of steady-state enzyme species was, therefore, analyzed through a combination of stopped-flow experiments and analysis of DV and D(V/K) for benzylamine oxidation. We conclude that the major species accumulating in the steady state are the oxidized cofactor-substrate Schiff base complex and the reduced, aminoquinol form of cofactor. These data rule out a slow release of product hydroperoxide from the aminoquinone form of enzyme, leading to the conclusion that the first electron transfer from substrate-reduced cofactor to dioxygen is the rate-determining step in the oxidative half-reaction. This step is also estimated to be 40% rate-limiting in kcat. An important mechanistic conclusion from this study is that dioxygen binding is a separate step from the rate-limiting electron-transfer step to form superoxide. On the basis of a recently determined X-ray structure for the active form of a yeast amine oxidase from Hansenula polymorpha [Li et al. (1998) Structure 6, 293], a hydrophobic space has been identified near the O-2 position of reduced cofactor as the putative dioxygen binding site. Movement of superoxide from this site onto the Cu(II) at the active site may occur prior to further electron transfer from cofactor to superoxide.

Amine Oxidase (Copper-Containing)↗
PubMed2001

The patient-physician relationship. Patient-physician communication during outpatient palliative treatment visits: an observational study.

CONTEXT: Improving health-related quality of life (HRQL) is an important goal of palliative treatment, but little is known about actual patient-physician communication regarding HRQL topics during palliative treatment. OBJECTIVES: To investigate the content of routine communication regarding 4 specific HRQL issues between oncologists and their patients and to identify patient-, physician-, and visit-specific factors significantly associated with discussion of such issues. DESIGN: Observational study conducted between June 1996 and January 1998. SETTING: Outpatient palliative chemotherapy clinic of a cancer hospital in the Netherlands. PARTICIPANTS: Ten oncologists and 240 of their patients (72% female; mean age, 55 years) who had incurable cancer and were receiving outpatient palliative chemotherapy. MAIN OUTCOME MEASURES: Patient and physician questionnaires and audiotape analysis of communication regarding daily activities, emotional functioning, pain, and fatigue during an outpatient consultation using the Roter Interaction Analysis System. RESULTS: Physicians devoted 64% of their conversation to medical/technical issues and 23% to HRQL issues. Patients' communication behavior was divided more equally between medical/technical issues (41%) and HRQL topics (48%). Of the independent variables investigated, patients' self-reported HRQL was the most powerful predictor of discussing HRQL issues. Nevertheless, in 20% to 54% of the consultations in which patients were experiencing serious HRQL problems, no time was devoted to discussion of those problems. In particular, these patients' emotional functioning and fatigue were unaddressed 54% and 48% of the time, respectively. Discussion of HRQL issues was not more frequent in consultations in which tumor response was evaluated. CONCLUSION: Despite increasing recognition of the importance of maintaining patients' HRQL as a goal of palliative treatment, the amount of patient-physician communication devoted to such issues remains limited and appears to make only a modest contribution, at least in an explicit sense, to the evaluation of treatment efficacy in daily clinical practice.

Activities of Daily Living↗
PubMed1996

The selectivity for K+ versus Na+ in DNA quadruplexes is dominated by relative free energies of hydration: a thermodynamic analysis by 1H NMR.

We have studied the competition between Na+ and K+ for coordination by G quartets using the oligonucleotide d(G3T4G3) as a model system. d(G3T4G3) forms a dimeric foldback structure containing three G quartets in the presence of either NaCl or KCl. Proton chemical shifts, which are particular to the species of coordinated ion, have been used to monitor the conversion between the sodium and potassium forms under equilibrium conditions. Analysis of titration experiments indicates that at least two K+ are coordinated by the three quartets of the dimeric molecule, and perfect fits of the data are obtained for two Na+ being displaced by two K+. Our results also indicate that the conversion of [d(G3T4G3)]2 from the sodium to the potassium form is associated with a net free energy change (delta G degrees) of -1.7 +/- 0.15 kcal/mol. It has long been suggested that the greater thermal stability of DNA quadruplex structures in the presence of K+ is primarily a result of the optimal fit of this ion in the coordination sites formed by G quartets. However, a consideration of the relatively small change in free energy associated with the conversion from the sodium to the potassium form and the relatively large difference between the free energy of hydration for Na+ and K+ indicates that this cannot be correct. Rather, the preferred coordination of K+ over Na+ is actually driven by the greater energetic cost of Na+ dehydration with respect to K+ dehydration.

DNA↗