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Biomedical subjects

R A Clayton

Publications and source records attributed to R A Clayton.

At least 19 recordsLinked to original sources
PubMed2001

The effect of acute alteration in oxygen tension on the bronchodilator response to salbutamol in vitro and in vivo in man.

These studies examine the effect of acute hypoxia on airway smooth muscle relaxation in response to salbutamol in vitro in human isolated bronchi from non-asthmatics and in vivo in-patients with asthma. Isometric responses were measured from rings of human bronchi pre-constricted with methacholine under oxygen tensions of 95% (hyperoxia), 20% (normoxia) and 4% (hypoxia). Once contractions had plateaued, concentration - response curves were conducted to salbutamol (10(-9)-10(-4)m). Twelve stable asthmatic patients were studied in a randomised double blind fashion. On two study days following baseline measurements, patients were randomised to receive either oxygen (FiO(2)1.0) or a hypoxic gas mixture (FiO(2)0.15) followed by three incremental doses of nebulised salbutamol at 15 min intervals. On two further study days nebulised saline was administered instead of salbutamol. In isolated bronchi, salbutamol-induced relaxations were significantly (P< 0.001) greater in hyperoxia and normoxia (P< 0.01) when compared to hypoxia. Among patients with asthma no significant differences were found in the mean maximum % change in forced expiratory volume (FEV(1)) from baseline between the hypoxic and hyperoxic study days on which nebulised salbutamol was administered. We conclude that acute hypoxia attenuates airway smooth muscle relaxation in response to salbutamol in vitro but has no effect on salbutamol-induced bronchodilation in in-patients with asthma.

Administration, Inhalation↗
PubMed2001

Screening for herpesvirus genomes in common acute lymphoblastic leukemia.

There is epidemiological evidence that infection may play a role in the etiology of childhood leukemia in particular common B cell precursor acute lymphoblastic leukemia. A panel of 20 leukemic samples (panel 1) was examined for the presence of four lymphotropic herpesviruses using conventional molecular techniques. A second independent panel of 27 leukemic samples (panel 2), along with 28 control peripheral blood samples from children with other forms of cancer, was tested for the presence of the same four viruses using sensitive real-time quantitative PCR. While herpesvirus genomes were detected, they were present at very low levels; detection rates and levels were similar in the leukemic and control panels. In addition we surveyed 18 leukemic samples (five from panel 1, six from panel 2 and a further seven samples not previously analyzed) using a degenerate PCR assay capable of detecting the genomes of known herpesviruses plus putative new members of the family. No novel herpesvirus genomes were detected suggesting that a herpesvirus is unlikely to be etiologically involved as a transforming agent in common acute lymphoblastic leukemia.

Adolescent↗
PubMed2000

DNA sequence of both chromosomes of the cholera pathogen Vibrio cholerae.

Here we determine the complete genomic sequence of the gram negative, gamma-Proteobacterium Vibrio cholerae El Tor N16961 to be 4,033,460 base pairs (bp). The genome consists of two circular chromosomes of 2,961,146 bp and 1,072,314 bp that together encode 3,885 open reading frames. The vast majority of recognizable genes for essential cell functions (such as DNA replication, transcription, translation and cell-wall biosynthesis) and pathogenicity (for example, toxins, surface antigens and adhesins) are located on the large chromosome. In contrast, the small chromosome contains a larger fraction (59%) of hypothetical genes compared with the large chromosome (42%), and also contains many more genes that appear to have origins other than the gamma-Proteobacteria. The small chromosome also carries a gene capture system (the integron island) and host 'addiction' genes that are typically found on plasmids; thus, the small chromosome may have originally been a megaplasmid that was captured by an ancestral Vibrio species. The V. cholerae genomic sequence provides a starting point for understanding how a free-living, environmental organism emerged to become a significant human bacterial pathogen.

Base Sequence↗
PubMed1999

Chronic exposure to hypoxia attenuates contractile responses in rat airways in vitro: a possible role for nitric oxide.

We investigated the effect of chronic hypoxia (10% O(2) for 14 days) on airway responsiveness in rats. Chronic hypoxia significantly (P<0. 05, P<0.01, P<0.01, respectively) attenuated contractions evoked by methacholine (10(-9)-3x10(-4) M), endothelin-1 (10(-10)-3x10(-7) M) and potassium chloride (10(-3)-7x10(-2) M) in rat isolated trachea. To investigate this attenuation, we studied the effect of epithelial removal, indomethacin (3x10(-6) M), and L-nitro arginine methyl ester (L-NAME, 10(-4) M), on contractile responses in control and chronically hypoxic rat trachea. Indomethacin did not alter contractions evoked by methacholine or endothelin-1 in control or hypoxic rats. In contrast, epithelial removal and L-NAME both significantly potentiated responses to methacholine and endothelin-1 in trachea from control and chronically hypoxic rats. In separate experiments, tracheal rings were first contracted with methacholine (10(-6) M) and then relaxed, either by the nitric oxide donor sodium nitroprusside or by the beta(2)-adrenoceptor agonist, salbutamol. Sodium nitroprusside was significantly (P<0.001) more effective at reversing induced tone in tracheal rings from chronically hypoxic than control rats. Salbutamol, however, was equally effective in chronically hypoxic and control rats. These results suggest that, in trachea from both control and chronically hypoxic rats, contractile responses to methacholine and endothelin-1 are inhibited by nitric oxide, probably released from the epithelium. The attenuation of contractile responses in airways from chronically hypoxic rats may be due to an enhanced guanylyl cyclase activity and hence, an increased response to nitric oxide.

Albuterol↗
PubMed1999

Evidence for lateral gene transfer between Archaea and bacteria from genome sequence of Thermotoga maritima.

The 1,860,725-base-pair genome of Thermotoga maritima MSB8 contains 1,877 predicted coding regions, 1,014 (54%) of which have functional assignments and 863 (46%) of which are of unknown function. Genome analysis reveals numerous pathways involved in degradation of sugars and plant polysaccharides, and 108 genes that have orthologues only in the genomes of other thermophilic Eubacteria and Archaea. Of the Eubacteria sequenced to date, T. maritima has the highest percentage (24%) of genes that are most similar to archaeal genes. Eighty-one archaeal-like genes are clustered in 15 regions of the T. maritima genome that range in size from 4 to 20 kilobases. Conservation of gene order between T. maritima and Archaea in many of the clustered regions suggests that lateral gene transfer may have occurred between thermophilic Eubacteria and Archaea.

Archaea↗
PubMed1999

The effect of chronic hypoxia on endothelin receptor subtype-mediated responses in rat isolated airways.

Contractile responses to endothelin-1 (ET-1) were investigated in isolated trachea from rats previously exposed to chronic hypoxia (10% O(2)) or room air for 14 days. Concentration-response curves were constructed to ET-1 (10(-11)-3x10(-7)m ) in the presence and absence of the ET(A)receptor antagonist FR 139317 (10(-8), 10(-7)and 10(-6)m ), the ET(B)receptor antagonist BQ 788 (10(-6)and 3x10(-6)m ), the non-selective ET receptor antagonist SB 209670 (10(-7)and 10(-6)m ) and a combination of FR 139317 (10(-6)m ) and BQ 788 (10(-6)m ). Concentration-response curves were also conducted to the ET(B)receptor agonist sarafotoxin S6c (10(-11)-3x10(-7)m ). In addition, responses to ET-1 (10(-11)-3x10(-7)m ) were examined in the presence and absence of the nitric oxide synthase inhibitor, L-NAME. In control rat trachea, both FR 139317 and BQ 788 failed to inhibit ET-1-induced contractions and, indeed, FR 139317 (10(-8)m ) and BQ 788 actually potentiated responses. In trachea from chronic hypoxic rats, FR 139317 did not alter ET-1 responses whereas BQ 788 again potentiated ET-1-induced contractions. The non-selective ET receptor antagonist SB 209670 attenuated ET-1-evoked contractions in trachea from control and chronically hypoxic rats. A combination of FR 139317 (10(-6)m ) and BQ 788 (10(-6)m ) also attenuated ET-1 responses in control rat trachea, but not trachea from chronically hypoxic rats. In trachea from both control and chronically hypoxic rats, L-NAME significantly potentiated responses to ET-1. To investigate ET receptor-mediated relaxation, tissues were preconstricted with methacholine and concentration-response curves were conducted to ET-1 (10(-13)-10(-8)m ) in the presence and absence of BQ 788 (10(-6)m ) and to the ET(B)receptor agonist sarafotoxin S6c (10(-13)-10(-8)m ). In trachea from control and chronic hypoxic rats, ET-1 and sarafotoxin S6c evoked only very small, non-reproducible relaxatory responses.

Animals↗
PubMed1998

Findings emerging from complete microbial genome sequences.

Sixteen microorganisms, including one eukaryote, four archaeons, and 11 eubacteria, have been completely sequenced and published. More than 50 genomes are scheduled to be completed by the year 2000. This explosive growth of information is forcing change in many scientific disciplines (e.g. bioinformatics and molecular genetics), spawning new fields, and even changing the way scientific information is used and shared. Novel, global genome sequence comparisons seem slow to appear but the infrastructure for these projects is being built, and we expect exciting developments in the near future.

Databases, Factual↗
PubMed1997

The complete genome sequence of the hyperthermophilic, sulphate-reducing archaeon Archaeoglobus fulgidus.

Archaeoglobus fulgidus is the first sulphur-metabolizing organism to have its genome sequence determined. Its genome of 2,178,400 base pairs contains 2,436 open reading frames (ORFs). The information processing systems and the biosynthetic pathways for essential components (nucleotides, amino acids and cofactors) have extensive correlation with their counterparts in the archaeon Methanococcus jannaschii. The genomes of these two Archaea indicate dramatic differences in the way these organisms sense their environment, perform regulatory and transport functions, and gain energy. In contrast to M. jannaschii, A. fulgidus has fewer restriction-modification systems, and none of its genes appears to contain inteins. A quarter (651 ORFs) of the A. fulgidus genome encodes functionally uncharacterized yet conserved proteins, two-thirds of which are shared with M. jannaschii (428 ORFs). Another quarter of the genome encodes new proteins indicating substantial archaeal gene diversity.

Archaeoglobus fulgidus↗
PubMed1997

The complete genome sequence of the gastric pathogen Helicobacter pylori.

Helicobacter pylori, strain 26695, has a circular genome of 1,667,867 base pairs and 1,590 predicted coding sequences. Sequence analysis indicates that H. pylori has well-developed systems for motility, for scavenging iron, and for DNA restriction and modification. Many putative adhesins, lipoproteins and other outer membrane proteins were identified, underscoring the potential complexity of host-pathogen interaction. Based on the large number of sequence-related genes encoding outer membrane proteins and the presence of homopolymeric tracts and dinucleotide repeats in coding sequences, H. pylori, like several other mucosal pathogens, probably uses recombination and slipped-strand mispairing within repeats as mechanisms for antigenic variation and adaptive evolution. Consistent with its restricted niche, H. pylori has a few regulatory networks, and a limited metabolic repertoire and biosynthetic capacity. Its survival in acid conditions depends, in part, on its ability to establish a positive inside-membrane potential in low pH.

Antigenic Variation↗
PubMed1997

Changing the oxygen tension alters the ability of bronchodilators to protect against methacholine-induced challenge in bovine isolated bronchial rings.

The ability of atrial natriuretic peptide, salbutamol, sodium nitroprusside and isosorbide dinitrate to protect against challenge with methacholine in bovine isolated bronchi was compared in different O2 tensions. Perfusing the Krebs-Henseleit solution with gas mixtures containing 95% O2 (hyperoxia), 20% O2 (approximately normoxia) and 0% O2 (hypoxia) produced O2 tensions in the organ-baths of 524, 147 and 26 mm Hg, respectively. In hyperoxia, pre-incubation of atrial natriuretic peptide at concentrations of 3 x 10(-7) M and 10(-6) M significantly attenuated responses to methacholine, whereas in normoxia, these concentrations of atrial natriuretic peptide had no effect. Furthermore, in hypoxia, 3 x 10(-7) M and 10(-6) M atrial natriuretic peptide significantly enhanced responses to methacholine. Salbutamol, at concentrations of 3 x 10(-7) M and 10(-6) M significantly attenuated responses to methacholine in hyperoxia, whereas in normoxia and hypoxia, pre-incubation of salbutamol did not alter the methacholine response. Pre-incubation of 10(-5) M sodium nitroprusside significantly attenuated methacholine-induced contractions in hyperoxia and when the oxygen tension in the gas mixture was lowered to 20% or 0%, the ability of sodium nitroprusside to protect against methacholine challenge was enhanced. In hyperoxia, isosorbide dinitrate, at the 10(-4) M level, evoked a rightward shift of the methacholine response curve. Lowering the oxygen tension to either 20% or 0% enhanced the protectant effect of isosorbide dinitrate, with the effect being greater in 20% O2. Thus, the effect of these bronchodilators on methacholine-induced challenge in hyperoxia O2 differed from those in normoxia and hypoxia, although the direction of the changes varied among the agents used. This suggests that the responses evoked by bronchodilators in 95% O2 may not necessarily predict those in the physiological range of oxygen tensions and that the relative effectiveness of bronchodilators may vary between normoxic and hypoxic conditions.

Albuterol↗
PubMed1997

Large-scale production and characterization of Bacillus thuringiensis subsp. tenebrionis insecticidal protein from Escherichia coli.

Bacillus thuringiensis subsp. tenebrionis insecticidal protein was produced in recombinant Escherichia coli and purified to near homogeneity to provide quantities of protein for safety-assessment studies associated with the registration of transgenic potato plants. The 68-kDa protein is produced naturally by Bacillus thuringiensis subsp. tenebrionis by translation initiation at an internal initiation site in the native DNA sequence. The gene sequence specific for this truncated protein was expressed in E. coli strain JM 101 and fermented at the 1000-1 scale. The protein accumulated as insoluble inclusion bodies, and was purified by extraction at pH 10.8 with carbonate buffer, selective precipitation at pH 9.0, and differential centrifugation. No chromatography steps were required to produce over 50 g purified protein as a lyophilized powder with a purity greater than 95% and demonstrating full insecticidal activity against Colorado potato beetle larvae. The protein was further characterized to assure identity and suitability for use in safety-assessment studies.

Animals↗
PubMed1997

Effect of angiotensin II on histamine-induced bronchoconstriction in the human airway both in vitro and in vivo.

The renin-angiotensin system is activated in acute severe asthma. Angiotensin II causes bronchoconstriction in mild asthmatics and potentiates methacholine-evoked bronchoconstriction both in vitro and in vivo. To evaluate the effect of angiotensin II on histamine-induced bronchoconstriction, human bronchial rings (n = 6) were obtained from lung tissue at thoracotomy and were prepared in organ baths. Contractions were measured isometrically and cumulative concentration-response curves obtained to angiotensin II alone and to histamine in the presence and absence of threshold concentrations of angiotensin II. Eight asthmatic patients with bronchial hyper-reactivity to histamine were challenged with histamine during intravenous infusion of placebo, angiotensin II 1 ng kg-1 min-1 and angiotensin 2 ng kg-1 min-1 administered in a randomized, double-blind fashion, FEV1 was measured prior to, during the infusion and during the histamine challenge. Angiotensin II (3 x 10(-7)M and 10(-6)M) alone evoked small contractions (< 0.25 g) of human bronchi in vitro, but pre-incubation with threshold concentrations of angiotensin II (10(-7)M, 3 x 10(-7)M and 10(-6)M) had no effect on histamine-evoked contractions. In asthmatic patients, angiotensin II alone had no effect on baseline FEV1 at the low levels infused and did not affect the response to nebulized histamine as measured by the PC20 histamine: Geometric mean (range) PC20 histamine (mg ml-1) screening day 3.58 (1.26-7.75), placebo infusion 2.67 (0.89-9.57), angiotensin II 1 ng kg-1 min-1 2.45 (0.42-6.97) and angiotensin II 2 ng kg-1 3.09 (0.8-10.78). It is concluded that, in contrast to its potentiating effect on methacholine-induced bronchoconstriction, angiotensin II has no effect on histamine-evoked bronchoconstriction in human bronchi in vitro or in vivo.

Adult↗
PubMed1997

Interactions between endothelin-I-induced contractions and bronchodilators in human isolated bronchi.

1. The aim of this study was to compare the ability of four different bronchodilators (atrial natriuretic peptide, salbutamol, sodium nitroprusside and isosorbide dinitrate) to reverse and also to protect against contractions evoked by the spasmogens endothelin-1 and methacholine in human isolated bronchial rings. 2. Contractions evoked by either endothelin-1 or methacholine were reversed by atrial natriuretic peptide (10(-9)-10(-6) mol/l), salbutamol (10(-9)-10(-5) mol/l), sodium nitroprusside (10(-9)-10(-5) mol/l) and isosorbide dinitrate (10(-7)-10(-4) mol/l). 3. Sodium nitroprusside produced a significantly (P < 0.05 for data points) greater mean maximal inhibition of endothelin-1-induced tone than methacholine-induced tone; however, the other dilators were equally effective at reversing either endothelin-1- or methacholine-induced contractions. Comparing sodium nitroprusside and salbutamol, sodium nitroprusside was significantly (P < 0.05 for data set) less effective than salbutamol at reversing either endothelin-1- or methacholine-induced contractions. 4. To compare the ability of these bronchodilator substances to protect against spasmogen challenge, cumulative concentration-response curves to either endothelin-1 (10(-10)-3 x 10(-7) mol/l) or methacholine (10(-9)-3 x 10(-4) mol/l) were constructed in the presence and absence of each bronchodilator. Atrial natriuretic peptide, at a concentration of 10(-6) mol/l, significantly attenuated contractions evoked by methacholine. In contrast, contractions evoked by endothelin-1 were enhanced by atrial natriuretic peptide at concentrations of 3 x 10(-7) and 10(-6) mol/l. Preincubation of salbutamol at a concentration of 10(-6) mol/l significantly attenuated methacholine-induced contractions, but responses to endothelin-1 were not altered by preincubation of salbutamol at concentrations of 3 x 10(-7), 10(-6) and 3 x 10(-6) mol/l. Sodium nitroprusside (10(-6) mol/l) and isosorbide dinitrate (3 x 10(-5) mol/l) did not alter responses evoked by subsequent addition of either endothelin-1 or methacholine. At a concentration of 10(-4) mol/l, however, isosorbide dinitrate significantly attenuated endothelin-1-evoked contractions. 5. These results show that drugs which reverse agonist-induced tone in isolated bronchial rings may not necessarily protect against subsequent challenge with this agonist. This suggests that the pharmacology of relaxation may be dissimilar to that of protection.

Albuterol↗
PubMed1997

Effect of acute alterations in inspired oxygen tension on methacholine induced bronchoconstriction in patients with asthma.

BACKGROUND: Recent in vitro and in vivo studies in animals have suggested that ambient oxygen tension may influence airway responsiveness to bronchoconstrictor stimuli. These observations may have relevance to the management of acute exacerbations of asthma. The present studies were designed to examine the influence of inspired oxygen tension (Fio2 1.0, 0.21, 0.15) on methacholine-induced broncho-constriction in patients with asthma. METHODS: In a dual study two groups of asthmatic patients performed methacholine inhalation challenges breathing either air (Fio2 0.21) or a hypoxic gas mixture (Fio2 0.15) in study 1 and air (Fio2 0.21) or hyperoxia (Fio2 1.0) in study 2. The gases were administered through a closed breathing circuit in a randomised double blind fashion. The PC20 values (dose of methacholine causing a 20% fall in forced expiratory volume in one second (FEV1) were calculated after each methacholine challenge by linear interpolation from the logarithmic dose response curve. Plasma catecholamine levels were measured before and after methacholine challenges as well as heart rate, oxygen saturation, and percentage end tidal carbon dioxide levels. RESULTS: The geometric mean PC20 value for methacholine was significantly lower on the hypoxic study day than on the normoxic day in study 1 (mean difference in PC20 values 2.88 mg/ml (95% CI 1.4 to 5.3); p < 0.05), but there was no significant difference in the geometric mean PC20 value for methacholine between the hyperoxic and normoxic study days in study 2 (mean difference in PC20 values 1.45 mg/ ml (95% CI 0.83 to 2.51)). CONCLUSIONS: Acute hypoxia potentiates methacholine induced bronchoconstriction and acute hyperoxia has no effect in mild to moderate patients with stable asthma.

Administration, Inhalation↗
PubMed1996

Complete genome sequence of the methanogenic archaeon, Methanococcus jannaschii.

The complete 1.66-megabase pair genome sequence of an autotrophic archaeon, Methanococcus jannaschii, and its 58- and 16-kilobase pair extrachromosomal elements have been determined by whole-genome random sequencing. A total of 1738 predicted protein-coding genes were identified; however, only a minority of these (38 percent) could be assigned a putative cellular role with high confidence. Although the majority of genes related to energy production, cell division, and metabolism in M. jannaschii are most similar to those found in Bacteria, most of the genes involved in transcription, translation, and replication in M. jannaschii are more similar to those found in Eukaryotes.

Amino Acid Sequence↗
PubMed1996

The effect of oxygen tension on responses evoked by methacholine and bronchodilators in bovine isolated bronchial rings.

This study examined the effect of acute changes in oxygen tension on responses evoked by the bronchoconstrictor methacholine and the dilators salbutamol, atrial natriuretic peptide and isosorbide dinitrate in isolated bovine bronchi. Cumulative concentration-response curves to methacholine (10(-9)-3 x 10(-4) M) were constructed in three oxygen tensions; hyperoxia (95%), normoxia (20%) and hypoxia (4% O2). Oxygen tensions of 20% and 4% each significantly enhanced contractions to methacholine compared to those in 95% O2. There was no significant difference, however, between responses in 20 and 4%. The ability of salbutamol, atrial natriuretic peptide and isosorbide dinitrate to reverse methacholine induced tone was also compared in the three oxygen tensions (95%, 20% and 4%). Lowering the oxygen tension from 95% enhanced the ability of each of the drugs to reverse contraction, however the pattern differed between drugs; salbutamol was more effective in 20% O2 than 4%, atrial natriuretic peptide was more effective in 4% than either 95 or 20% O2 and isosorbide dinitrate was more effective in both 4 and 20% than 95% O2. In conclusion, both bronchoconstrictor and bronchodilator responses in 95% O2 (hyperoxia) differed from those in 20% O2 (normoxia) and further changes occurred on moving to 4% (hypoxia), although the direction of the changes varied among the dilators. This suggests that the responses evoked by bronchodilators in 95% O2 may not necessarily predict those in the physiological range of oxygen tensions and that the relative effectiveness of bronchodilators may vary between normoxic and hypoxic conditions.

Adrenergic beta-Agonists↗
PubMed1996

Microbial sterility testing of oil-formulated bovine somatotropin using Tween 80 dispersion.

The Food and Drug Administration requires that sterile pharmaceutical products be free of viable microorganisms. Sterility testing of pharmaceutical products provides added assurance that the product is sterile. Sterility testing is typically done by inoculating the drug product into microbial growth media followed by visual inspection for growth during incubation for a specified time period. A lack of visual growth indicates that the drug product samples tested were sterile. Formulated Posilac bovine somatotropin consists of protein particles suspended in an oil-based excipient. The product formulation is immiscible in aqueous media due to the excipient's water insolubility and the insolubility of the protein particles at near neutral pH values. Because the formulation is packaged and sold as a sterile product, it is critical that a sensitive microbial sterility test method be used for this key quality test. A sterility test method was developed for Posilac that utilized Tween 80 (i.e. polysorbate 80) as a dispersant. Dispersion of the product using Tween 80 produced a homogeneous suspension of bovine somatotropin particles and oil droplets in the micron size range. The suspension did not appreciably settle out with time, attesting to the homogeneous nature of the mixture. This method was found to be compatible with survival, recovery, and growth from low numbers of the test organisms required by the U.S. Pharmacopeia XXIII as well as from two additional test cultures.

Animals↗