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Biomedical subjects

A E Jones

Publications and source records attributed to A E Jones.

At least 37 records · Page 2Linked to original sources
PubMed1998

Cell-surface biotinylation of GLUT4 using bis-mannose photolabels.

New cell-impermeant bis-mannose photolabels have been developed with biotinyl groups attached to 4-(1-azi-2,2,2-trifluoroethyl)-benzoyl-1, 3-bis(d-mannos-4-yloxy)-2-propylamine (ATB-BMPA) by either a polyethoxy spacer (Bio-ATB-BMPA) or an additional hexanoic acid spacer (Bio-LC-ATB-BMPA). The half-maximal inhibition constants, Ki values, for inhibition of glucose transport activity in insulin-stimulated rat adipocytes were determined to be 359+/-10 and 273+/-28 microM for Bio-ATB-BMPA and Bio-LC-ATB-BMPA, respectively. These values are similar to those previously reported for the non-biotinylated compound ATB-BMPA. Following UV-irradiation-induced cross-linking of the biotinylated photolabels to rat adipocytes, the biotinylated glucose transporter isoform 4 (GLUT4) could be detected by non-radioactive and radioactive methods that utilized the interaction with streptavidin. Biotinylated GLUT4 from 1-2 microg of adipose cell membranes, precipitated onto magnetic streptavidin beads, could be sensitively and quantitatively detected using an electrochemiluminescent assay method. This utilized a ruthenium-tagged anti-GLUT4 antibody that on excitation at an electrode generated an electrochemiluminescent signal in an ORIGEN analyser. Alternatively, surface-biotinylated GLUT4 could be easily, but less sensitively, detected in streptavidin agarose precipitates which were analysed by conventional GLUT4 Western blotting. Data obtained using the non-radioactive methods compared favourably with those using tritiated versions of the biotinylated probes. Insulin treatment of adipocytes increased the levels of signals from surface biotinylated GLUT4 by approximately 10-fold or approximately 20-fold, respectively, when the electrochemiluminescent or the Western blot detection methods were used and these signals were blocked by cytochalasin B.

Adipocytes↗
PubMed1998

Determination of the enantiomers of salbutamol and its 4-O-sulphate metabolites in biological matrices by chiral liquid chromatography tandem mass spectrometry.

Sensitive, mass spectrometry based bioanalytical methods are described for the determination of the R- and S-enantiomers of the beta-agonist salbutamol (albuterol) and its 4-O-sulphate metabolite in human plasma and urine. In both methods samples are prepared by 96 well format solid phase extraction using a custom built robotic system. Extracts are then analysed by liquid chromatography tandem mass spectrometry (LC-MS/MS) using a teicoplanin-based chiral stationary phase and selected reaction monitoring. The methods are accurate (bias < +/- 10%), precise (%CV < 11%) and sensitive, providing lower limits of quantitation (LLoQ) in plasma of 100 pg/mL and 5 ng/mL for the enantiomers of salbutamol and its 4-O-sulphate metabolite, respectively. By restricting the chiral method for plasma to the enantiomers of salbutamol only, it was possible to revalidate at an improved LLoQ of 25 pg/mL. A high throughout LC-MS/MS method has also been developed for racemic salbutamol only, which uses a similar extraction procedure but a conventional C8 column. The method has a reduced analysis time of three minutes per sample and using a high sensitivity, triple quadrupole mass spectrometer provides an LLoQ of 5 pg/mL based on extraction of 0.5 mL of plasma.

Adrenergic beta-Agonists↗
PubMed1998

The effect of age and gender on the metabolic disposal of [1-13C] palmitic acid.

OBJECTIVE: To examine the effect of age and gender on the metabolic disposal of [1-13C]palmitic acid. DESIGN: Cross-sectional. SETTING: Clinical Nutrition and Metabolism Unit at Southampton General Hospital, Institute of Human Nutrition, University of Southampton. SUBJECTS AND MEASUREMENTS: Twelve children (5 boys and 7 girls; aged 5-10 y) and six men (BMI 23.3 +/- 2.6 kg/m2; aged 20-30 y) were recruited. Following oral administration of a bolus dose of [1-13C]palmitic acid (10 mg/kg body weight) consumed with a test meal (1667 kJ) the excretion of 13C-label was measured on breath as 13CO2 over 24 h and in stool over 5 d to account for differences in absorption of [1-13C]palmitic acid. The 13C-enrichment of samples was determined by continuous flow-isotope ratio mass spectrometry. Net substrate oxidation was estimated from gaseous exchange measurements in the postabsorptive state and over 6 h postprandially. RESULTS: The excretion of 13CO2 on breath varied between subjects both in the pattern and amount excreted over 24 h. Breath 13CO2 was not different between boys (61.0 +/- 22.4% of absorbed dose) and girls (54.2 +/- 17.9% of absorbed dose). The excretion of breath 13CO2 was less in the men (35.1 +/- 9.3% of absorbed dose; P = 0.005) and that observed previously by our group in women (30.7 +/- 6.7% of absorbed dose; P = 0.005) than in the children. Net fat oxidation was greater in the children in both the postabsorptive (2.43 +/- 0.78 g/h) and postprandial (11.89 +/- 3.13 g/6 h) states than in the men (0.93 g/h +/- 1.50; P = 0.016; 9.86 +/- 10.53 g/6 h; NS) and women studied previously (0.53 +/- 0.68 g/h; P = 0.003; 0.03 +/- 3.21 g/6 h; P = 0.001). CONCLUSIONS: Our observations that children oxidised nearly twice the amount of [1-13C]palmitic acid than adults in conjunction with greater net fat oxidation in children than adults in both the postabsorptive and postprandial states should be considered before current UK dietary recommendations for fat and saturated fats, developed for adults, are applied to growing children. For dietary recommendations to be developed further more information is required, particularly in groups of infants and the elderly, about the factors that influence the postprandial handling of dietary fat.

Absorption↗
PubMed1998

Human sperm function in co-culture with human, macaque or bovine oviduct epithelial cell monolayers.

Human sperm function was compared in co-culture with monolayers of oviduct epithelial cells (OEC) from three species, human, macaque and bovine. For all species, freeze-thawed and passaged OEC from females in the periovulatory phase were used. OEC cultured on an extracellular matrix (Matrigel) formed a monolayer which supported human sperm attachment to OEC from all three species. Spermatozoa in co-culture with OEC from all three species showed prolonged survival and improved motility characteristics over those cultured in medium alone. This paper describes an efficient, repeatable co-culture system for human spermatozoa which supports sperm attachment to OEC and subsequently improves sperm function over that seen in control medium cultures. Because the improved sperm function in co-culture did not differ significantly between human and bovine OEC for those attributes studied, it is proposed that bovine OEC could be used as an alternative to human OEC in certain human sperm coculture studies. Follicular phase bovine OEC from reproductively normal donors are far more accessible than their human counterparts, thus making this co-culture system more widely available for the study of human spermatozoa-female tract interactions.

Animals↗
PubMed1998

Metabolic handling of 13C labelled tripalmitin in healthy controls and patients with cystic fibrosis.

AIM: To examine the gastrointestinal handling and metabolic disposal of emulsified [1-13C]palmitic acid esterified into a triglyceride in nine healthy children and seven patients with cystic fibrosis on enzyme replacement treatment. METHODS: After an overnight fast, each child was given 10 mg/kg body weight [1,1,1-13C]tripalmitin with a standardised test meal of low natural 13C abundance. The total enrichment of 13C was measured using isotope ratio mass spectrometry in stool collected for a period of up to five days and in breath samples collected over a 24 hour period. RESULTS: The mean proportion of administered 13C label excreted in stool was 6% (range, 1-12.7%) in healthy children and 24.6% (range, 0-64%) in patients with cystic fibrosis. Healthy children excreted 31.3% of the administered label on their breath (range, 14.2-42.9%). Correcting the excretion of administered 13C label on the breath for differences in digestion and absorption in patients with cystic fibrosis increased the difference between individuals from 0-31.3% of administered dose (mean, 17.9%) to 0-49.1% of absorbed dose (mean, 23.2%) and was poorly related to the amount of 13C label in stool. CONCLUSION: Measurements of breath 13CO2 do not consistently reflect the gastrointestinal handling of emulsified 13C labelled tripalmitin because of differences in digestion and absorption in cystic fibrosis. Further studies need to examine whether "breath tests" alone can predict with confidence the gastrointestinal handling of other 13C labelled triglycerides and fatty acids.

Breath Tests↗
PubMed1998

Bronchopleural fistula resulting from the use of a thoracic vent: a case report and review.

Pneumothorax is defined as the presence of gas or air within the pleural space. Standard treatment is usually based on the evacuation of the gas by various methods. The thoracic vent is a relatively new device used in the treatment of pneumothorax. This report focuses on the first major complication, as far as is known, associated with the use of a thoracic vent.

Adult↗
PubMed1997

Stable-isotope method for determining the gastrointestinal handling of [1-13C]palmitic acid.

The 13C enrichment in individual fatty acids extracted from human feces following the oral administration of [1-13C]palmitic acid has been determined using a novel approach based upon gas chromatography-isotope ratio mass spectrometry. The method was established and tested for precision and repeatability. Analytical precision was determined from 10 repeated injections of a sample containing 16:0 and 18:0 with levels of delta 13C abundance measured at -34.01 +/- 0.60 and -23.62 +/- 0.95 delta per mil (parts per thousand) (/1000), respectively (mean +/- SD). For the repeatability study, measurement of enrichment of the same mixture of unlabeled fatty acid methyl ester (FAME) standards (13:0, 14:0, 16:0, and 18:0) was found to have standard deviations (0.45, 0.56, 1.46 and 1.54/1000, respectively). When labeled [1-13C]palmitic acid was serially diluted with naturally enriched palmitic acid, a linear relationship was obtained to a dilution of 10% enriched compound (530/1000). FAME were prepared from two fecal samples from a normal healthy adult; the first, a baseline specimen, containing no added label and the second, followed a single oral dose of [1-13C]palmitic acid and was enriched. Enrichment in 13C was confined to the solvent-soluble fraction following lipid extraction, and was only identified with prior acidification. The enrichments were measured in triplicate, baseline sample -32.66 +/- 0.5/1000, enriched sample +268.61 +/- 8.0/1000. Enrichment was restricted to the labeled species consumed, 16:0. The methodology described here allows for the separation of compounds prior to the determination of enrichment and can be utilized to contribute to a more complete description of the gastrointestinal handling of labeled substrates than previously obtained.

Carbon Isotopes↗
PubMed1997

Gastrointestinal handling of [1-13C]palmitic acid in healthy controls and patients with cystic fibrosis.

AIM: To examine the gastrointestinal handling of [1-13C]palmitic acid given as the free acid by measuring the excretion of 13C label in stool in 16 healthy children and 11 patients with cystic fibrosis on their habitual enzyme replacement treatment. METHODS: After an overnight fast, each child ingested 10 mg/kg body weight [1-13C]palmitic acid with a standardised test meal of low natural 13C abundance. A stool sample was collected before the test and all stools were collected thereafter for a period of up to five days. The total enrichment of 13C in stool and the species bearing the 13C label was measured using isotope ratio mass spectrometry. RESULTS: The proportion of administered 13C label excreted in stool was 24.0% (range 10.7-64.9%) in healthy children and only 4.4% (range 1.2-11.6%) in cystic fibrosis patients. The enrichment of 13C in stool was primarily restricted to the species consumed by the subjects (that is as palmitic acid). CONCLUSIONS: There does not appear to be a specific defect in the absorption of [1-13C]palmitic acid in patients with cystic fibrosis. The reasons why cystic fibrosis patients appear to absorb more of this saturated fatty acid than healthy children is not clear and requires further investigation.

Adolescent↗
PubMed1997

Reduced cardiac contractile responsiveness to isoproterenol in obese rabbits.

Although obesity is characterized by increased sympathetic nervous system activity, there is often a paradoxical reduction in cardiovascular end-organ response to sympathetic stimulation. Mechanisms involved in reduced sympathetic responsiveness in obesity have not been well characterized. Therefore, we determined cardiac contractile responsiveness to beta-stimulation in the obese rabbit model using both isolated heart (IH) and isolated papillary muscle (IPM) preparations. Female New Zealand White rabbits were fed control (IH: n=9; IPM: n=6) or 10% fat diets (IH: n=9; IPM: n=7) for 12 weeks. Contractile responsiveness in the IH was determined using a modified Langendorff preparation to evaluate the dose-response relationship between isoproterenol and 1) peak developed pressure/g of left ventricular wet weight and 2) maximal rate of pressure development (+dP/dt/P). Contractile responsiveness in the IPM was determined using right ventricular papillary muscles to evaluate the dose-response relationship between isoproterenol and (1) peak developed tension (T)/mm2 cross-sectional area (CSA) and (2) maximal rate of tension development (dT/dt/CSA). In the IH, baseline and maximum developed pressure/g were reduced in obese rabbits by 37% and 31%, respectively (P< or =.05). In the IPM, baseline and maximum T/CSA responses were reduced in obese rabbits by 59% and 33%, respectively (P< or =.05). Potency of isoproterenol as reflected by the EC50 did not differ between lean and obese animals in either preparation. These results demonstrate that left ventricular contractility in obesity is reduced at baseline and in response to stimulation with isoproterenol and suggest that decreased responsiveness to beta-stimulation may be a factor in the obesity-related systolic dysfunction.

Animals↗
PubMed1996

Determination of the potent antiprotozoal compound atovaquone in plasma using liquid-liquid extraction followed by reversed-phase high-performance liquid chromatography with ultraviolet detection.

A specific and robust method is presented for the determination of atovaquone in plasma. Atovaquone is a potent antiprotozoal compound for use in immunocompromised patients who are intolerant of conventional therapies. The method involves a liquid-liquid extraction of the compound into hexane modified with 2% (v/v) isoamyl alcohol. The processed extracts are analysed by reversed-phase high-performance liquid chromatography with ultraviolet detection at 254 nm. The assay has a limit of quantification of 0.1 microgram/ml and is linear between 0.1 and 50 micrograms/ml. The method has been applied to many clinical studies and has been demonstrated to be precise and accurate with high sample throughput. Atovaquone is not significantly metabolised in humans.

Antiprotozoal Agents↗
PubMed1996

Sedimentation analyses of the salt- and divalent metal ion-induced oligomerization of nucleolar protein B23.

Protein B23 is a major nonribosomal nucleolar protein and putative ribosome assembly factor that has been demonstrated to form oligomers. Sedimentation velocity and equilibrium analyses were used to examine the oligomerization properties of recombinant proteins B23.1 and B23.2. Under low ionic strength conditions protein B23.1 was predominantly a 2.1S monomer with small amounts of a 7.1S oligomer. At NaCl concentrations of 0.1 M and above the protein was almost exclusively the 7.1S oligomer. The oligomer remained the predominant species in NaCl concentrations as high as 1 M, suggesting that oligomers are not stabilized by electrostatic interactions. Low concentrations of divalent metal ions (0.1 - 1mM Ca2+ or Mg2+) also promoted oligomerization. Reducing agents had no effect on oligomerization, indicating that disulfide bridges are not important in oligomer formation. Protein B23.2, the carboxyl-terminal truncated isoform, had sedimentation characteristics similar to that of protein B23.1, suggesting that the carboxyl-terminal end of protein B23.1 is not essential for oligomerization. Protein B23.1 was previously shown to bind nucleic acids [Wang, D., Baumann, A., Szebeni, A., & Olson, M. O. J.(1995) J. Biol. Chem. 269, 30994-30998]. The effect of protein B23.1 oligomerization on its interaction with a 230 base pair DNA fragment was examined by sedimentation analyses. Under conditions where significant amounts of monomer were present, protein B23.1 was capable of binding DNA, whereas conditions that strongly favored oligomerization caused a nearly complete abolition of DNA binding activity. These studies suggest that protein B23 exists in an equilibrium between monomer and oligomer and that the quaternary structure of the protein may regulate its DNA binding properties.

Animals↗
PubMed1995

Identification of urinary and biliary conjugated metabolites of the neuromuscular blocker 51W89 by liquid chromatography/mass spectrometry.

Cisatracurium, (1R, 1'R, 2R, 2'R)-2,2-[1,5-pentanediylbis-[oxy(3-oxo- 3,1-propanediyl]]bis[1-[(3,4-dimethoxyphenyl)-methyl]-1,2,3,4- tetrahydro-6,7-dimethoxy-2-methylisoquinolinium] dibenzenesulphonate (51W89), is an intermediate-acting neuromuscular blocking agent. 51W89 is one of ten isomers contained in Tracrium (atracurium besylate) and represents approximately 15 percent of the atracurium mixture. Clinical studies have indicated that 51W89 is more potent and is significantly weaker as a histamine releaser than atracurium. In vitro studies in human plasma have shown that, like atracurium, 51W89 spontaneously degrades at physiological pH by Hoffmann elimination to form laudanosine and the quaternary monoacrylate. Subsequent ester hydrolysis of the monoacrylate generates the monoquaternary alcohol. In rat plasma, 51W89 is also metabolized by non-specific carboxylesterases to the monoquaternary alcohol and the monoquaternary acid, the former being rapidly hydrolysed further to the more stable acid. It has been reported that laudanosine can be further metabolized via N-dimethylation to yield tetrahydropapaverine. The rate-limiting step in the degradation of 51W89 in human plasma is Hofmann elimination, whilst in rat plasma, the action of non-specific carboxylesterases is rate limiting. As part of the development of 51W89, the disposition of 14C-51W89 following a single intravenous bolus dose was studied in various animal species and humans. In the present work, we describe the identification of 51W89 metabolites in urine and bile from these studies by high performance liquid chromatography/mass spectrometry using pneumatically-assisted electrospray ionization coupled to an on-line radioactivity monitor.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗
PubMed1995

Isolation of serotype 1 Marek's disease viruses from vaccinated Australian flocks.

Field samples were received for Marek's disease virus (MDV) isolation from clinically affected flocks from several regions of Eastern Australia. Lymphocytes were fractionated in Ficoll-Paque and passaged once or twice in chicken embryo kidney cultures. Serotype-specific virus was detected in infected cultures by indirect immunofluorescence with monoclonal antibodies. Serotype 1 MDV was isolated from 10 flocks. In samples from 5 of these flocks, serotype 2 and 3 vaccine viruses were isolated from the same specimen. In a parallel study, plasmas obtained during lymphocyte isolation were tested for antibodies to MDVs by agar gel precipitin (AGP) tests using serotype 1 and 3 antigen extracts. No correlation was observed between the rate of virus isolation and AGP positivity. The AGP test was incapable of discriminating between the different MDV serotypes.

Animals↗
PubMed1995

Practical evolution and application of direct intracytoplasmic sperm injection for male factor and idiopathic fertilization failure infertilities.

OBJECTIVE: To analyze the introduction of a new assisted fertilization technique for the treatment of severe male factor and idiopathic fertilization failure infertilities. DESIGN: Retrospective analysis of 16-month clinical application of IVF-ET where insemination was performed solely by direct intracytoplasmic sperm injection. SETTING: Clinical IVF-ET program. PATIENTS: Ninety-two couples undergoing 105 cycles of sperm injection. RESULTS: One hundred embryo transfers yielded 28 viable pregnancies (28%) from which eight normal deliveries have occurred to date. Complete cleavage arrest or fertilization failure occurred in four cycles, and one couple had all embryos cryopreserved. One thousand one hundred forty-three eggs were injected of which 173 (15%) degenerated. Four hundred seventy-nine of the surviving 970 eggs became normally fertilized (49%), and 381 of these zygotes (79.5%) developed suitably for cryopreservation or for transfer. Thirty-four of 310 embryos transferred implanted, yielding an implantation rate of 11%. Both testicular and epididymal sperm were used successfully to achieve fertilization and pregnancies, as was sperm retrieved by electroejaculation. Older women and couples suffering from prior idiopathic fertilization failure had a markedly poorer outcome. CONCLUSIONS: These results confirm that the intracytoplasmic sperm injection technique is a successful form of assisted fertilization that can be applied to a wide range of couples at significant risk from fertilization failure.

Adult↗
PubMed1995

Factors affecting success with intracytoplasmic sperm injection.

In this study, 141 couples underwent 163 cycles of in vitro fertilization (IVF) and embryo transfer in which the eggs were inseminated by intracytoplasmic sperm injection (ICSI). Overall, 41% of the injected eggs were normally fertilized and 81% of the resulting embryos were suitable for cryopreservation (91 embryos) or uterine transfer. From 153 fresh embryo transfers, 45 ongoing or delivered pregnancies (27.6% per cycle) were achieved, and of the 507 embryos transferred, 54 successfully implanted giving an implantation rate per embryo of 10.7%. Five additional pregnancies did not yield a viable fetus or underwent a spontaneous abortion, giving a miscarriage rate of 10% (5/50). Increased maternal age or a prior diagnosis of failed fertilization after conventional IVF had a significantly negative impact on success. Sperm from the testis and epididymis, those retrieved by electro-ejaculation, and completely immotile ejaculated sperm all gave rise to pregnancies. ICSI reinsemination was used with limited success to rescue failed fertilization cycles, although the implantation rate per embryo was poor (5%). ICSI has greatly improved the ability to use IVF for treating couples with a poor fertilization potential.

Adult↗
PubMed1994

Ten different dietary fibers have significantly different effects on serum and liver lipids of cholesterol-fed rats.

The comparative effects of 10 different dietary fibers on serum and liver lipids were investigated by feeding male Sprague-Dawley rats diets containing 10 g cholesterol + 2 g cholic acid/kg diet, with 60 g fiber/kg diet. Diets were fed for 3 wk; cellulose was the control fiber. Rats fed psyllium (rich in soluble fiber) had the lowest serum and liver cholesterol concentrations. Rats fed other soluble fiber-rich fibers (oat gum, guar gum and pectin) also had significantly lower serum and liver cholesterol concentrations than rats fed cellulose. Although feeding diets containing both soluble and insoluble fibers (soybean fiber and oat bran) did not significantly alter serum cholesterol, liver cholesterol values were significantly lower than those of cellulose-fed rats. Rats fed rice bran, predominantly an insoluble fiber source, had significantly higher liver cholesterol and significantly lower body weight gains and serum triglyceride concentrations than cellulose-fed rats. Values for serum and liver cholesterol were similar for rats were fed insoluble-rich fibers (corn bran, cellulose and wheat bran). These observations indicate that feeding dietary fibers rich in soluble fiber produces lower serum and liver cholesterol concentrations than does feeding commonly available sources of water-insoluble fiber.

Analysis of Variance↗