Improving the care of patients with asthma. Interview by Rhonda Siddall.
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Asthma↗
Biomedical subjects
G Barnes
Publications and source records attributed to G Barnes.
Selective lymphoscintigraphy: a necessary adjunct to dye-directed sentinel node biopsy for breast cancer?
BACKGROUND: Dye-directed sentinel node biopsy (SNB) for breast cancer provides accurate staging with low morbidity, but for tumors distant from the axilla, its use has been questioned. HYPOTHESIS: Can preoperative breast lymphoscintigraphy (BL) applied selectively to medial hemisphere tumors predict a subset of patients who may not require surgical staging of the axilla? DESIGN: Prospective cohort study. SETTING: Tertiary, multidisciplinary breast center. PATIENTS: Thirty-two women with breast tumors located in the medial hemisphere (30) or inframammary crease (2). INTERVENTION: Peritumoral injection of 500 microCi of technetium Tc 99m sulfur colloid and biplanar imaging. Nonpalpable lesions were localized with ultrasound or mammography. At the time of definitive breast surgery, isosulfan blue dye-directed SNB was performed. Axillary dissection was performed when the SN contained a tumor or could not be identified. MAIN OUTCOME MEASURES: Regional nodal basins identified by BL; success rate of SNB. RESULTS: Preoperative BL demonstrated axillary drainage in 28 patients (88%); 2 patients (6%) had isolated internal mammary radionuclide uptake, and 2 patients had no nodal uptake. Dye-directed axillary SNB succeeded in 27 (87%) of 31 patients, including both patients with failed BL. Breast lymphoscintigraphy had predicted isolated internal mammary drainage in 2 of 4 patients whose SNs could not be identified. Metastases were found in 5 patients (16%). CONCLUSIONS: Axillary radionuclide uptake predicts but does not augment dye-directed SN identification. In those few patients with isolated internal mammary drainage, BL may obviate the need for surgical staging of the axilla.
Adult↗
Functional cooperation between the microtubule and actin cytoskeletons.
In diverse cell types, microtubule (MT) and actin filament networks cooperate functionally during a wide variety of processes, including vesicle and organelle transport, cleavage furrow placement, directed cell migration, spindle rotation, and nuclear migration. The mechanisms by which MTs and actin filaments cooperate to mediate these different processes can be grouped into two broad categories: coordinated MT- and actin-based transport to move vesicles, organelles, and cell fate determinants; and targeting and capture of MT ends at cortical actin sites. Over the past several years, a growing number of cellular factors that bridge these cytoskeletal systems have been identified. These include 'hetero-motor' complexes (physically associated myosin and kinesin), myosin-CLIP170 complexes, formin homology (FH) proteins, dynein and the dynactin complex, Kar9p, coronin, Kelch repeat-containing proteins, and ERM proteins.
Actins↗
Effects of experimental glaucoma in macaques on the multifocal ERG. Multifocal ERG in laser-induced glaucoma.
Multifocal ERGs (MERGs) of 5 adult monkeys (Macaca mulatta) with inner retinal defects caused by laser-induced glaucoma were compared to MERGs from 3 monkeys with inner retinal activity suppressed pharmacologically. MERGs were recorded with DTL fiber electrodes from anesthetized monkeys. Stimuli consisted of 103 equal size hexagons within 17 degrees of the fovea. Stimuli at each location passed through a typical VERIS m-sequence of white (200 cd/m2) and black (12 cd/m2) presentations. In animals with laser-induced glaucoma, visual field sensitivity was assessed by static perimetry using the Humphrey C24-2 full-threshold program modified for animal behavior. Inner retinal (amacrine and ganglion cell) activity was suppressed by intravitreal injection of TTX (4.7-7.6 microM) and NMDA (1.6-5 mM). In normal eyes the first order response (1st order kernel) was larger and more complex, with more distinct oscillations (>60 Hz) in central than in peripheral locations. The 2nd order kernel also was dominated by oscillatory activity. There were naso-temporal variations in both kernels. Pharmacological suppression of inner retinal activity reduced or eliminated the oscillatory behavior, and naso-temporal variations. The 1st order kernel amplitude was increased most and was largest at the fovea. Removed inner retinal responses also were largest at the fovea. The 2nd order kernel was greatly reduced at all locations. In eyes with advanced glaucoma, the effects were similar to those produced by suppressing inner retinal activity, but the later portion of the 1st order kernel waveform was different, lacking a dip after the large positive wave. Visual sensitivity losses and MERG changes both increased over the timecourse of glaucoma, with changes in the MERG being more diffusely distributed across the visual field. We conclude that 1st and 2nd order responses of the primate MERG can be identified that originate from inner retina and are sensitive indicators of glaucomatous neuropathy.
Animals↗
Assessing payback from NHS reactive research programmes.
Aims to assess retrospectively the payback from NHS reactive research programmes in the Northern and Yorkshire region. A questionnaire was sent to all recipients of regional reactive research programme funding (biomedical, health services research (HSR), and primary and community care programmes) between 1 April 1991 and 31 March 1996. The sample available for analysis involved 174 respondents covering 119 projects, with a total financial value of 2.2 million Pounds. The main outcome measures used were peer-reviewed publications, changes in individual practice, changes in NHS service delivery and organisation, and impact on the careers of researchers. Overall, 119 projects produced 230 peer-reviewed publications: this was achieved at an average cost of 10,673 Pounds, 6,386 Pounds and 22,310 Pounds per publication for the biomedical, HSR, and primary and community care programmes respectively. From the qualitative data analysis, important changes in individual practice and NHS service delivery were identified by respondents. The researchers in our sample appeared to have attracted over 6 million Pounds in R&D funding related to the initial regional grant. Although based on self-report, there is evidence to suggest that the return on investment from NHS R&D can be substantial, taking a broad view of benefits to the NHS and to researchers. The findings also confirm the need for more effective dissemination and implementation of research findings.
Efficiency, Organizational↗
Multiregional evaluation of the SimPlate heterotrophic plate count method compared to the standard plate count agar pour plate method in water.
A new SimPlate heterotrophic plate count (HPC) method (IDEXX Laboratories, Westbrook, Maine) was compared with the pour plate method at 35 degrees C for 48 h. Six laboratories tested a total of 632 water samples. The SimPlate HPC method was found to be equivalent to the pour plate method by regression analysis (r = 0. 95; y = 0.99X + 0.06).
Agar↗
Annual report of the Rotavirus Surveillance Programme, 1999/2000.
The National Rotavirus Reference Centre has conducted rotavirus surveillance by means of a collaborative laboratory based initiative started in June 1999. The serotypes of rotaviruses that lead to the hospitalisation of children with acute diarrhoea were determined from June 1999 to May 2000. We examined 1126 rotavirus specimens using a combination of monoclonal antibody immunoassay, reverse transcription-polymerase chain reaction, and hybridisation. The four most common serotypes G1-G4 were represented. More than 50% of isolates tested were serotype G1, with serotype G1 being represented in most centres Australia-wide. Serotype G9 rotaviruses were identified for the first time in Australia, and were second in importance with 10% of samples tested. The significant presence of G9 viruses throughout Australia suggests the emergence of a new serotype and has implications for current rotavirus vaccine strategies that target serotypes G1-G4.
Australia↗
Coronin promotes the rapid assembly and cross-linking of actin filaments and may link the actin and microtubule cytoskeletons in yeast.
Coronin is a highly conserved actin-associated protein that until now has had unknown biochemical activities. Using microtubule affinity chromatography, we coisolated actin and a homologue of coronin, Crn1p, from Saccharomyces cerevisiae cell extracts. Crn1p is an abundant component of the cortical actin cytoskeleton and binds to F-actin with high affinity (Kd 6 x 10(-9) M). Crn1p promotes the rapid barbed-end assembly of actin filaments and cross-links filaments into bundles and more complex networks, but does not stabilize them. Genetic analyses with a crn1Delta deletion mutation also are consistent with Crn1p regulating filament assembly rather than stability. Filament cross-linking depends on the coiled coil domain of Crn1p, suggesting a requirement for Crn1p dimerization. Assembly-promoting activity is independent of cross-linking and could be due to nucleation and/or accelerated polymerization. Crn1p also binds to microtubules in vitro, and microtubule binding is enhanced by the presence of actin filaments. Microtubule binding is mediated by a region of Crn1p that contains sequences (not found in other coronins) homologous to the microtubule binding region of MAP1B. These activities, considered with microtubule defects observed in crn1Delta cells and in cells overexpressing Crn1p, suggest that Crn1p may provide a functional link between the actin and microtubule cytoskeletons in yeast.
Actin Cytoskeleton↗
Eye movements during combined pursuit, optokinetic and vestibular stimulation in macaque monkey.
During natural behaviour in a visual environment, smooth pursuit eye movements (SP) usually override the vestibular-ocular reflex (VOR) and the optokinetic reflex (OKR), which stem from head-in-space and scene-relative-to-eye motion, respectively. We investigated the interaction of SP, VOR, and OKR, which is not fully understood to date. Eye movements were recorded in two macaque monkeys while applying various combinations of smooth eye pursuit, vestibular and optokinetic stimuli (sinusoidal horizontal rotations of visual target, chair and optokinetic pattern, respectively, at 0.025, 0.05, 0.1, 0.2, 0.4, and 0.8 Hz, corresponding to peak stimulus velocities of 1.25-40 degrees/s for a standard stimulus of -/+8 degrees). Slow eye responses were analysed in terms of gain and phase. During SP at mid-frequencies, the eyes were almost perfectly on target (gain 0.98 at 0.1 Hz), independently of a concurrent vestibular or optokinetic stimulus. Pursuit gain at lower frequencies, although being almost ideal (0.98 at 0.025 Hz with pursuit-only stimulation), became modified by the optokinetic input (gain increase above unity when optokinetic stimulus had the same direction as target, decrease with opposite direction). At higher stimulus frequencies, pursuit gain decreased (down to 0.69 at 0.8 Hz), and the pursuit response became modified by vestibular input (gain increase during functionally synergistic combinations, decrease in antagonistic combinations).Thus, the pursuit system in monkey dominates during SP-OKR-VOR interaction, but it does so effectively only in the mid-frequency range. The results can be described in the form of a simple dynamic model in which it is assumed that the three systems interact by linear summation. In the model SP and OKR dominate VOR in the low- to mid-frequency/velocity range, because they represent closed loop systems with high internal gain values (>>1) at these frequencies/velocities, whereas the VOR represents an open loop system with about unity-gain (up to very high frequencies). SP dominance over OKR is obtained by allowing an 'attentional/volitional' mechanism to boost SP gain and a predictive mechanism to improve its dynamics.
Animals↗
Intussusception and rotavirus vaccine.
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Humans↗
Central venous catheter motion: a pitfall in catheter localization on pediatric chest radiography.
OBJECTIVE: We evaluated central venous catheter motion causing misinterpretation of catheter tip location in pediatric patients and in an experimental model. MATERIALS AND METHODS: After the recognition of clinically significant catheter motion in 12 randomly selected patients, we conducted a prospective 2-month study of chest radiographs in our pediatric care unit and neonatal emergency department. Serial radiographs were examined for change in catheter tip position ascribed to motion artifact. An in vitro model was developed to replicate clinical parameters. Experimentally, catheter movement and exposure time were analyzed and their impact on catheter localization was recorded. RESULTS: In the 12 randomly selected patients, radiographic assessment of central venous catheter tip localization on sequential radiographs caused minor to major misinterpretation of the position of the catheter tip and even total nonvisualization of the catheter. Ten (3.5%) of 352 prospectively interpreted pediatric emergency department radiographs showed similar findings. Catheter motion that caused blurring was reproduced in vitro using radiographic parameters typically used in a clinical setting. CONCLUSION: Catheter motion can cause problems in assessing catheter tip position on pediatric emergency department chest radiographs. We reproduced this phenomenon in an in vitro model. Catheter removal or change in position may be mimicked by this artifact, and patient management may be affected. In our study, catheter localization was affected by catheter motion and exposure time.
Adolescent↗
Saccharomyces cerevisiae Duo1p and Dam1p, novel proteins involved in mitotic spindle function.
In this paper, we describe the identification and characterization of two novel and essential mitotic spindle proteins, Duo1p and Dam1p. Duo1p was isolated because its overexpression caused defects in mitosis and a mitotic arrest. Duo1p was localized by immunofluorescence, by immunoelectron microscopy, and by tagging with green fluorescent protein (GFP), to intranuclear spindle microtubules and spindle pole bodies. Temperature-sensitive duo1 mutants arrest with short spindles. This arrest is dependent on the mitotic checkpoint. Dam1p was identified by two-hybrid analysis as a protein that binds to Duo1p. By expressing a GFP-Dam1p fusion protein in yeast, Dam1p was also shown to be associated with intranuclear spindle microtubules and spindle pole bodies in vivo. As with Duo1p, overproduction of Dam1p caused mitotic defects. Biochemical experiments demonstrated that Dam1p binds directly to microtubules with micromolar affinity. We suggest that Dam1p might localize Duo1p to intranuclear microtubules and spindle pole bodies to provide a previously unrecognized function (or functions) required for mitosis.
Antibodies, Fungal↗
Exploiting chemical libraries, structure, and genomics in the search for kinase inhibitors.
Selective protein kinase inhibitors were developed on the basis of the unexpected binding mode of 2,6,9-trisubstituted purines to the adenosine triphosphate-binding site of the human cyclin-dependent kinase 2 (CDK2). By iterating chemical library synthesis and biological screening, potent inhibitors of the human CDK2-cyclin A kinase complex and of Saccharomyces cerevisiae Cdc28p were identified. The structural basis for the binding affinity and selectivity was determined by analysis of a three-dimensional crystal structure of a CDK2-inhibitor complex. The cellular effects of these compounds were characterized in mammalian cells and yeast. In the latter case the effects were characterized on a genome-wide scale by monitoring changes in messenger RNA levels in treated cells with high-density oligonucleotide probe arrays. Purine libraries could provide useful tools for analyzing a variety of signaling and regulatory pathways and may lead to the development of new therapeutics.
Adenine↗
Extended excretion of rotavirus after severe diarrhoea in young children.
BACKGROUND: Rotaviruses are the major cause of severe childhood diarrhoea. Knowledge of the natural history of infection, including duration of intestinal virus shedding, is important in the understanding of transmission, sources of infection, and immune responses. METHODS: We carried out a study of rotavirus excretion in 37 children admitted to hospital with severe rotavirus diarrhoea. Sequential faecal specimens were collected from each child during 100 days of surveillance, and screened for rotavirus by EIA and by amplification of genome double-stranded RNA by reverse-transcription PCR. IgA coproantibody was estimated by EIA. FINDINGS: Duration of rotavirus excretion ranged from 4 to 57 days after onset of diarrhoea. Excretion ceased within 10 days in 16 (43%) children, and within 20 days in 26 (70%) children. Extended excretion was detected for 25-57 days in the remaining 11 (30%) children owing mainly to continued excretion of the primary infecting strain. Extended excretion was significantly associated with antirotavirus IgA coproantibody boosts during 100 days of surveillance (p=0.001, log-rank test), and with recurrence of mild diarrhoea symptoms during convalescence (p=0.006, Fisher's exact test). INTERPRETATION: Severe rotavirus disease in young children may be followed by extended excretion of rotavirus. The risk of transmission to others may be greater than previously believed. Extended excretion could also explain some cases of the postgastroenteritis syndrome.
Antibodies, Viral↗
Fifty glorious years.
Since the second world war, dentistry has undergone momentous changes, not only in clinical and mechanical developments but also in changes in people's perceptions of what care their dentist can offer them. Dental graduates today will be surprised at the great difference between their own education and that of their 1946 counterparts. This is one dentist's record of the last 50 years in dentistry and how certain advancements have affected his working life.
Adult↗
Direct analysis and identification of proteins in mixtures by LC/MS/MS and database searching at the low-femtomole level.
A method to directly identify proteins contained in mixtures by microcolumn reversed-phase liquid chromatography electrospray ionization tandem mass spectrometry (LC/MS/MS) is studied. In this method, the mixture of proteins is digested with a proteolytic enzyme to produce a large collection of peptides. The complex peptide mixture is then separated on-line with a tandem mass spectrometer, acquiring large numbers of tandem mass spectra. The tandem mass spectra are then used to search a protein database to identify the proteins present. Results from standard protein mixtures show that proteins present in simple mixtures can be readily identified with a 30-fold difference in molar quantity, that the identifications are reproducible, and that proteins within the mixture can be identified at low femtomole levels. Based on these studies, methodology has been developed for direct LC/MS/MS analysis of proteins enriched by immunoaffinity precipitation, specific interaction with a protein-protein fusion product, and specific interaction with a macromolecular complex. The approach described in this article provides a rapid method for the direct identification of proteins in mixtures.
Chromatography, High Pressure Liquid↗
DNA double-strand-break sensitivity, DNA replication, and cell cycle arrest phenotypes of Ku-deficient Saccharomyces cerevisiae.
In mammalian cells, the Ku heterodimer is involved in DNA double-strand-break recognition and repair. We have established in yeast a connection between Ku activity and DNA double-strand-break damage repair, and a connection between Ku activity and commitment to DNA replication. We generated double-stranded DNA breaks in yeast cells in vivo by expressing a restriction endonuclease and have shown that yeast mutants lacking Ku p70 activity died while isogenic wild-type cells did not. Moreover, we have discovered that DNA damage occurs spontaneously during normal yeast mitotic growth, and that Ku functions in repair of this damage. We also observed that mitotically growing Ku p70 mutants have an anomalously high DNA content, suggesting a role for Ku in regulation of DNA synthesis. Finally, we present evidence that Ku p70 function is conserved between yeast, Drosophila, and humans.
Animals↗