Biomedical subjects

A M Smith

Publications and source records attributed to A M Smith.

At least 55 records · Page 3Linked to original sources

PubMed2001

Cross-reactivity studies of a new group 2 allergen from the dust mite Glycyphagus domesticus, Gly d 2, and group 2 allergens from Dermatophagoides pteronyssinus, Lepidoglyphus destructor, and Tyrophagus putrescentiae with recombinant allergens.

BACKGROUND: Dust mites are important inducers of allergic disease. Group 2 allergens are recognized as major allergens in several mite species, including Dermatophagoides pteronyssinus, Lepidoglyphus destructor, and Tyrophagus putrescentiae. No allergens have thus far been characterized on the molecular level from the dust mite Glycyphagus domesticus. OBJECTIVE: We sought to examine the cross-reactivity among group 2 allergens of G domesticus, L destructor, T putrescentiae, and D pteronyssinus. METHODS: A group 2 allergen from G domesticus, Gly d 2, was cloned and expressed as a recombinant protein. Cross-reactivity between Gly d 2 and 3 other group 2 allergens, Lep d 2, Tyr p 2, and Der p 2, was studied by using individual sera and a serum pool RAST-positive to G domesticus, L destructor, T putrescentiae, and D pteronyssinus. Recombinant allergens were used as inhibitors of IgE binding in immunoblotting experiments. Molecular modeling on the basis of the Der p 2 structure was carried out for Gly d 2, Lep d 2, and Tyr p 2. RESULTS: Two cDNAs encoding isoforms of Gly d 2 were isolated, but only the Gly d 2.02 isoform was used in this study. Sixteen of 17 subjects had IgE to Gly d 2. The protein sequence of Gly d 2 revealed 79% identity to Lep d 2 and 46% and 41% identity to Tyr p 2 and Der p 2, respectively. Extensive cross-reactivity was demonstrated among Gly d 2, Lep d 2, and Tyr p 2, but little cross-reactivity was found between these allergens and Der p 2. According to the tertiary structure of Der p 2 and 3-dimensional models of Gly d 2, Lep d 2, and Tyr p 2, differences reside mainly in surface-exposed residues. CONCLUSION: Gly d 2 showed high sequence homology to Lep d 2. Cross-reactivity was observed between Gly d 2, Lep d 2, and Tyr p 2, but only limited cross-reactivity was demonstrated between these 3 allergens and Der p 2.

Allergens↗

PubMed2001

The molecular basis of antigenic cross-reactivity between the group 2 mite allergens.

BACKGROUND: Mite group 2 allergens Der p 2, Der f 2, and Eur m 2 are 14-kDa proteins of unknown function that share 83% to 85% amino acid sequence identity. Isoforms of the allergens within each genus have been identified which differ by 3 or 4 amino acids, but little is known of the influence of group 2 polymorphisms on human IgE antibody binding. OBJECTIVE: The purpose of this study was to investigate the importance of interspecies and isoform substitutions on murine mAb and IgE antibody binding and on the molecular structure of the group 2 allergens. METHODS: Site-directed mutagenesis was used to incorporate the isoform amino acid substitutions onto the Der p 2.0101 sequence. Recombinant allergens were expressed and purified from Escherichia coli and used to evaluate antibody binding by enzyme-linked immunosorbent assay (ELISA). Molecular modeling of the tertiary structure was used to analyze structural differences between the various group 2 allergens. RESULTS: The substitution of asparagine for aspartic acid at position 114 restored mAb binding of rDer p 2.0101; the other Der p 2 isoforms and the 3 rDer f 2 isoforms also reacted in the 2-site ELISA. The correlation of IgE binding to the Der p 2 isoforms was excellent and tended to be higher in the isoforms with the asparagine 114 substitution (r (2) = 0.87 vs r (2) = 0.95). rEur m 2.0101 bound to all mAb except 7A1; when compared with rDer p 2 for IgE binding, rEur m 2.0101 gave a correlation coefficient of r (2) = 0.68. Molecular modeling revealed that Eur m 2 and the storage mite homologs Lep d 2 and Tyr p 2 retain the tertiary fold of Der p 2. Eur m 2 has a conserved surface, whereas Lep d 2 and Tyr p 2 present most of the amino acid substitutions on this surface. Lep d 2 and Tyr p 2 did not react with mAb or with sera from patients with IgE to Dermatophagoides species. CONCLUSION: The isoform substitutions of rDer p 2 can be distinguished by mAb. The allergenic cross-reactivity between Der p 2, Der f 2, and Eur m 2 is a direct result of the conserved antigenic surface, whereas the lack of cross-reactivity with Lep d 2 and Tyr p 2 is a result of the multiple substitutions across this surface.

Amino Acid Sequence↗

PubMed2001

A cytosolic ADP-glucose pyrophosphorylase is a feature of graminaceous endosperms, but not of other starch-storing organs.

The occurrence of an extra-plastidial isoform of ADP-glucose (Glc) pyrophosphorylase (AGPase) among starch-storing organs was investigated in two ways. First, the possibility that an extra-plastidial isoform arose during the domestication of cereals was studied by comparing the intracellular distribution of enzyme activity and protein in developing endosperm of noncultivated Hordeum species with that previously reported for cultivated barley (Hordeum vulgare). As in cultivated barley, the AGPase of H. vulgare subsp. spontaneum and Hordeum murinum endosperm is accounted for by a major extra-plastidial and a minor plastidial isoform. Second, the ratio of ADP-Glc to UDP-Glc was used as an indication of the intracellular location of the AGPase activity in a wide range of starch-synthesizing organs. The ratio is expected to be high in organs in which UDP-Glc and ADP-Glc are synthesized primarily in the cytosol, because the reactions catalyzed by AGPase and UDP-Glc pyrophosphorylase will be coupled and close to equilibrium. This study revealed that ADP-Glc contents and the ratio of ADP-Glc to UDP-Glc were higher in developing graminaceous endosperms than in any other starch-storing organs. Taken as a whole the results indicate that an extra-plastidial AGPase is important in ADP-Glc synthesis in graminaceous endosperms, but not in other starch-storing organs.

Cytosol↗

PubMed2001

ADP-glucose pyrophosphorylase is located in the plastid in developing tomato fruit.

The subcellular location of activity and protein of ADP-glucose pyrophosphorylase (AGPase) in developing tomato (Lycopersicon esculentum) fruit was determined following a report that the enzyme might be present inside and outside the plastids in this organ. Plastids prepared from crude homogenates of columella and pericarp, the starch-accumulating tissues of developing fruit, contained 8% to 18% of the total activity of enzymes known to be confined to plastids, and 0.2% to 0.5% of the total activity of enzymes known to be confined to the cytosol. The proportion of the total activity of AGPase in the plastids was the same as that of the enzymes known to be confined to the plastid. When samples of plastid and total homogenate fractions were subjected to immunoblotting with an antiserum raised to AGPase, most or all of the protein detected was plastidial. Taken as a whole, these data provide strong evidence that AGPase is confined to the plastids in developing tomato fruit.

Blotting, Western↗

PubMed2001

The Arabidopsis sex1 mutant is defective in the R1 protein, a general regulator of starch degradation in plants, and not in the chloroplast hexose transporter.

Starch is the major storage carbohydrate in higher plants and of considerable importance for the human diet and for numerous technical applications. In addition, starch can be accumulated transiently in chloroplasts as a temporary deposit of carbohydrates during ongoing photosynthesis. This transitory starch has to be mobilized during the subsequent dark period. Mutants defective in starch mobilization are characterized by high starch contents in leaves after prolonged periods of darkness and therefore are termed starch excess (sex) mutants. Here we describe the molecular characterization of the Arabidopsis sex1 mutant that has been proposed to be defective in the export of glucose resulting from hydrolytic starch breakdown. The mutated gene in sex1 was cloned using a map-based cloning approach. By complementation of the mutant, immunological analysis, and analysis of starch phosphorylation, we show that sex1 is defective in the Arabidopsis homolog of the R1 protein and not in the hexose transporter. We propose that the SEX1 protein (R1) functions as an overall regulator of starch mobilization by controlling the phosphate content of starch.

Amino Acid Motifs↗

PubMed2001

The virulence factors of Bordetella pertussis: a matter of control.

Bordetella pertussis is the causative agent of whooping cough, a contagious childhood respiratory disease. Increasing public concern over the safety of whole-cell vaccines led to decreased immunisation rates and a subsequent increase in the incidence of the disease. Research into the development of safer, more efficacious, less reactogenic vaccine preparations was concentrated on the production and purification of detoxified B. pertussis virulence factors. These virulence factors include adhesins such as filamentous haemagglutinin, fimbriae and pertactin, which allow B. pertussis to bind to ciliated epithelial cells in the upper respiratory tract. Once attachment is initiated, toxins produced by the bacterium enable colonisation to proceed by interfering with host clearance mechanisms. B. pertussis co-ordinately regulates the expression of virulence factors via the Bordetella virulence gene (bvg) locus, which encodes a response regulator responsible for signal-mediated activation and repression. This strict regulation mechanism allows the bacterium to express different gene subsets in different environmental niches within the host, according to the stage of disease progression.

Animals↗

PubMed2001

Alterations in MurM, a cell wall muropeptide branching enzyme, increase high-level penicillin and cephalosporin resistance in Streptococcus pneumoniae.

We report that alteration in MurM, an enzyme involved in the biosynthesis of branched-stem cell wall muropeptides, is required for maximal expression of penicillin and cefotaxime resistance in the pneumococcus. Hungarian isolate 3191 (penicillin MIC, 16 microg/ml; cefotaxime MIC, 4 microg/ml) was a source of donor DNA in transformation experiments. Penicillin-binding protein DNA was insufficient to transform recipient strain R6 to full resistance. Further transformation with altered murM DNA was required for full expression of donor penicillin and cefotaxime resistance.

Amino Acid Sequence↗

PubMed2001

Emergence of a pneumococcal clone with cephalosporin resistance and penicillin susceptibility.

We report two South African serotype 6B pneumococcal isolates with cephalosporin resistance, yet with susceptibility to penicillin. DNA fingerprinting revealed that they were clonal in origin. pbp 2X and 1A genes showed major alterations typical of cephalosporin-resistant pneumococci. The pbp 2B gene was completely unaltered, explaining the penicillin susceptibility of the isolates.

Amino Acid Sequence↗

PubMed2001

Activity in ventral and dorsal premotor cortex in response to predictable force-pulse perturbations in a precision grip task.

This study compared the responses of ventral and dorsal premotor cortex (PMv and PMd) neurons to predictable force-pulse perturbations applied during a precision grip. Three monkeys were trained to grasp an unseen instrumented object between the thumb and index finger and to lift and hold it stationary within a position window for 2-2.5 s. The grip and load forces and the object displacement were measured on each trial. Single-unit activity was recorded from the hand regions in the PMv and PMd. In some conditions a predictable perturbation was applied to the object after 1,500 ms of static holding, whereas in other conditions different random combinations of perturbed and unperturbed trials were given. In the perturbed conditions, some were randomly and intermittently presented with a warning flash, whereas some were unsignaled. The activities of 198 cells were modulated during the task performance. Of these cells, 151 were located in the PMv, and 47 were located in the PMd. Although both PMv and PMd neurons had similar discharge patterns, more PMd neurons (84 vs. 43%) showed early pregrip activity. Forty of 106 PMv and 10/30 PMd cells responded to the perturbation with reflexlike triggered reactions. The latency of this response was always <100 ms with a mean of about 55 ms in both the PMv and the PMd. In contrast, 106 PMv and 30 PMd cells tested with the perturbations, only 9 and 10%, respectively, showed significant but nonspecific adaptations to the perturbation. The warning stimulus did not increase the occurrence of specific responses to the perturbation even though 21 of 42 cells related to the grip task also responded to moving visual stimuli. The responses were retinal and frequently involved limited portions of both foveal and peripheral visual fields. When tested with a 75 x 5.5-cm dark bar on a light background, these cells were sensitive to the direction of movement. In summary, the periarcuate premotor area activity to related to predictable force-pulse perturbations seems to reflect a general increase in excitability in contrast to a more specific anticipatory activity such as recorded in the cerebellum. In spite of the strong cerebello-thalamo-cortical projections, the results of the present study suggest that the cortical premotor areas are not involved in the elaboration of adaptive internal models of hand-object dynamics.

Action Potentials↗

PubMed2001

Activity in rostral motor cortex in response to predictable force-pulse perturbations in a precision grip task.

The purpose of this investigation was to characterize the discharge of neurons in the rostral area 4 motor cortex (MI) during performance of a precision grip task. Three monkeys were trained to grasp an object between the thumb and index finger and to lift and hold it stationary for 2-2.5 s within a narrow position window. The grip and load forces and the vertical displacement of the object were recorded on each trial. On some trials a downward force-pulse perturbation generating a shear force and slip on the skin was applied to the object after 1.5 s of static holding. In total, 72 neurons were recorded near the rostral limit of the hand area of the motor cortex, located close to the premotor areas. Of these, 30 neurons were examined for receptive fields, and all 30 were found to receive proprioceptive inputs from finger muscles. Intracortical microstimulation applied to 38 recording sites evoked brief hand movements, most frequently involving the thumb and index finger with an average threshold of 12 microA. Slightly more than one-half of the neurons (38/72) demonstrated significant increases in firing rate that on average began 284 +/- 186 ms before grip onset. Of 54 neurons tested with predictable force-pulse perturbations, 29 (53.7%) responded with a reflexlike reaction at a mean latency of 54.2 +/- 16.8 ms. This latency was 16 ms longer than the mean latency of reflexlike activity evoked in neurons with proprioceptive receptive fields in the more caudal motor cortex. No neurons exhibited anticipatory activity that preceded the perturbation even when the perturbations were delivered randomly and signaled by a warning stimulus. The results indicate the presence of a strong proprioceptive input to the rostral motor cortex, but raise the possibility that the afferent pathway or intracortical processing may be different because of the slightly longer latency.

Action Potentials↗

PubMed2001

Usefulness of magnetic resonance-derived quantitative measurements of cerebral blood flow and volume in prediction of infarct growth in hyperacute stroke.

BACKGROUND AND PURPOSE: The identification of the tissue at risk for infarction remains challenging in stroke patients. In this study, we evaluated the value of quantitative cerebral blood flow (CBF) and cerebral blood volume (CBV) measurements in the prediction of infarct growth in hyperacute stroke. METHODS: Fluid-attenuated inversion recovery (FLAIR), diffusion-weighted (DW), and gradient-echo echo-planar perfusion-weighted (PW) sequences were obtained in 66 patients within 6 hours of stroke onset; ischemia was confirmed on follow-up FLAIR images. We delineated the following: (1) the initial infarct on DW images, (2) the area of hemodynamic disturbance on mean transit time (MTT) maps, and (3) the final infarct on follow-up FLAIR images. MTT, CBF, and CBV were calculated in the following areas: area of initial infarct (INF), area of infarct growth (IGR, final minus initial infarct), the hemodynamically disturbed area that remained viable (OLI, hemodynamic disturbance minus final infarct), and all contralateral mirror regions. RESULTS: Compared with mirror regions, the MTT in abnormal areas was always prolonged. The respective mean+/-SD CBF and CBV values were as follows: for INF, 28+/-16 mL/min per 100 g and 6.9+/-2.7%; for IGR, 36+/-20 mL/min per 100 g and 8.9+/-3.1%; for OLI, 50+/-17 mL/min per 100 g and 11.2+/-3%; and for mirror regions, 64+/-23 mL/min per 100 g and 8.7+/-2.5%. The CBV and CBF values were significantly different between all abnormal areas (except for the CBF between INF and IGR). In the area of DW/PW mismatch, a combined CBF or CBV threshold of 35 or 8.2, respectively, predicted evolution to infarction with a sensitivity of 81% and a specificity of 76%. CONCLUSIONS: Quantitative measurements of CBF and CBV in hyperacute stroke may help to predict infarct growth and to select the subjects who will benefit from thrombolysis.

Acute Disease↗

PubMed2001

Hepatic perfusion parameters in chronic liver disease: dynamic CT measurements correlated with disease severity.

OBJECTIVE: The aim of our study was to determine if hepatic perfusion parameters measured with CT change in relation to disease severity in patients with chronic liver disease. SUBJECTS AND METHODS: Dynamic contrast-enhanced single-section CT scans of the liver were obtained in 40 individuals who included six control subjects, 16 patients with noncirrhotic chronic liver disease, and 18 patients with cirrhosis. Hepatic, aortic, and portal venous time-density curves were fitted to a dual-input one-compartment model to calculate the liver perfusion, arterial fraction, distribution volume, and mean transit time. RESULTS: Liver perfusion decreased in patients with cirrhosis (67 +/- 23 mL. min(-1). 100 mL(-1) versus 108 +/- 34 mL. min(-1). 100 mL(-1) in control subjects [p = 0.009] and 98 +/- 36 mL. min(-1). 100 mL(-1) in patients with noncirrhotic chronic liver disease [p = 0.003]), and the arterial fraction and the mean transit time increased (41 +/- 27% and 51 +/- 79 sec versus 17 +/- 16% and 16 +/- 5 sec in control subjects, and 19 +/- 6% and 17 +/- 8 sec in patients with noncirrhotic chronic liver disease [p < 0.05]). A significant correlation was seen between these three perfusion parameters and the severity of chronic liver disease based on clinical and biologic data (p < 0.001). No significant change in distribution volume was observed. CONCLUSION: Hepatic perfusion parameters measured with CT were significantly altered in cirrhosis and correlated with the severity of chronic liver disease.

Case-Control Studies↗

PubMed2001

A preliminary analysis of psychophysiological variables and nursing performance in situations of increasing criticality.

OBJECTIVE: To examine the relationship between psychological, physiological, and performance variables in intensive care unit (ICU) nurses in situations of increasing criticality. SUBJECTS AND METHODS: Psychophysiological variables and endotracheal suctioning performance were examined in a classroom, a skills laboratory, and an ICU. Situation-specific anxiety (state anxiety) and the predisposition to view situations as threatening (trait anxiety), cognitive appraisal, and heart rate were measured and compared with self-appraisal and a nurse instructor's ratings of suctioning performance. Baseline data were obtained during class on 45 novice ICU nurses. RESULTS: Twenty-six nurses provided complete data, which included being videotaped and monitored in the classroom, in the skills laboratory performing endotracheal suctioning, and in the ICU during suctioning. High state anxiety significantly predicted poor ICU suctioning performance (P<.04). Nurses high in state and trait anxiety, worry, and heart rate performed poorly compared with less anxious nurses. Nurses in this study who performed best had a mean heart rate of 94 beats/min. CONCLUSION: Those nurses who are high state anxious, high trait anxious, and worried and who had a faster heart rate performed less well than their more relaxed peers. Nurses with high state anxiety may be at risk for attrition, burnout, medical errors, and poor performance in other ICU nursing tasks.

Anxiety↗

PubMed2001

Statins suppress THP-1 cell migration and secretion of matrix metalloproteinase 9 by inhibiting geranylgeranylation.

Macrophages secrete matrix metalloproteinase 9 (MMP-9), an enzyme that weakens the fibrous cap of atherosclerotic plaques, predisposing them to plaque rupture and subsequent ischemic events. Recent work indicates that statins strongly reduce the possibility of heart attack. Furthermore, these compounds appear to exert beneficial effects not only by lowering plasma low-density-lipoprotein cholesterol but also by directly affecting the artery wall. To evaluate whether statins influence the proinflammatory responses of monocytic cells, we studied their effects on the chemotactic migration and MMP-9 secretion of human monocytic cell line THP-1. Simvastatin dose dependently inhibited THP-1 cell migration mediated by monocyte chemoattractant protein 1, with a 50% inhibitory concentration of about 50 nM. It also inhibited bacterial lipopolysaccharide-stimulated secretion of MMP-9. The effects of simvastatin were completely reversed by mevalonate and its derivatives, farnesylpyrophosphate and geranylgeranyl pyrophosphate, but not by ubiquinone. Additional studies revealed similar but more profound inhibitory effects with L-839,867, a specific inhibitor of geranylgeranyl transferase. However, alpha-hydroxyfarnesyl phosphonic acid, an inhibitor of farnesyl transferase, had no effect. C3 exoenzyme, a specific inhibitor of the prenylated small signaling Rho proteins, mimicked the inhibitory effects of simvastatin and L-839,867. These data supported the role of geranylgeranylation in the migration and MMP-9 secretion of monocytes.

ADP Ribose Transferases↗

PubMed2001

SNAP-shots: scenes from nursing action plans--the cultivation of leadership in oncology nursing.

PURPOSE/OBJECTIVES: To describe the experiences of oncology nurses who participated in a national workshop to prepare nurses to lead the transformation of cancer care and to illustrate the range of possibilities available to nurses motivated to affect positive change. DATA SOURCES: Literature on leadership and negotiation are referenced and related to the personal experiences and the observations of participants in the workshop. DATA SYNTHESIS: Leadership is presented in historical context. A case study approach was used to organize the personal accounts of participants' plans to affect positive organizational change in oncology care settings. Scenes from action plans illustrate how the workshop's core curriculum, which focuses on leadership models, processes, and skills, was realized in practice. CONCLUSIONS: Leadership is a dynamic process that shapes and is shaped by all stakeholders during the process of planned change. Education, risk taking, and active participation are vital components of the process of leadership development. IMPLICATIONS FOR NURSING PRACTICE: The contributions of nurses may be simple of complex and affect single individuals, groups, or institutions. In all cases, oncology nursing practice and cancer care may be enhanced.

Breast Neoplasms↗

PubMed2001

Multiple, distinct isoforms of sucrose synthase in pea.

Genes encoding three isoforms of sucrose synthase (Sus1, Sus2, and Sus3) have been cloned from pea (Pisum sativum). The genes have distinct patterns of expression in different organs of the plant, and during organ development. Studies of the isoforms expressed as recombinant proteins in Escherichia coli show that they differ in kinetic properties. Although not of great magnitude, the differences in properties are consistent with some differentiation of physiological function between the isoforms. Evidence for differentiation of function in vivo comes from the phenotypes of rug4 mutants of pea, which carry mutations in the gene encoding Sus1. One mutant line (rug4-c) lacks detectable Sus1 protein in both the soluble and membrane-associated fractions of the embryo, and Sus activity in the embryo is reduced by 95%. The starch content of the embryo is reduced by 30%, but the cellulose content is unaffected. The results imply that different isoforms of Sus may channel carbon from sucrose towards different metabolic fates within the cell.

Carbon↗

PubMed2001

Motor vehicle occupant crashes among teens: impact of the graduated licensing law in San Diego.

To assess the effect of California's graduated driver's license (GDL) law in San Diego County, we compared motor vehicle crash and passenger injury rates involving 16-year-old drivers for 1999 and 2000, the first two years of the law, against 1997, using data from a statewide database of all injury crashes. While the crash rate per 1,000 drivers did not change following GDL, a significant decline in the percentage of 16-year-olds with a driver's license corresponded with large decreases in population-based crash and injury rates.

Accidents, Traffic↗

PubMed2000

Hydrogen exchange nuclear magnetic resonance spectroscopy mapping of antibody epitopes on the house dust mite allergen Der p 2.

New strategies for allergen-specific immunotherapy have focused on reducing IgE reactivity of purified recombinant allergens while maintaining T-cell epitopes. Previously, we showed that disrupting the disulfide bonds of the major house dust mite allergen Der p 2 resulted in 10-100-fold less skin test reactivity in mite-allergic subjects but did not change in vitro T-cell proliferative responses. To provide a more complete picture of the antigenic surface of Der p 2, we report here the identification of three epitopes using hydrogen protection nuclear magnetic resonance spectroscopy. The epitopes are defined by monoclonal antibodies that are able to inhibit IgE antibody binding to the allergen. Each monoclonal antibody affected the amide exchange rate of 2-3 continuous residues in different regions of Der p 2. Based on these data, a number of other residues were predicted to belong to each epitope, and this prediction was tested for monoclonal antibody 7A1 by generating alanine point mutants. The results indicate that only a small number of residues within the predicted epitope are functionally important for antibody binding. The molecular definition of these three epitopes will enable us to target limited positions for mutagenesis and to expand our studies of hypoallergenic variants for immunotherapy.

Amides↗