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Biomedical subjects

C J Evans

Publications and source records attributed to C J Evans.

At least 91 records · Page 5Linked to original sources
PubMed1994

Preferential localization of a vesicular monoamine transporter to dense core vesicles in PC12 cells.

Neurons and endocrine cells have two types of secretory vesicle that undergo regulated exocytosis. Large dense core vesicles (LDCVs) store neural peptides whereas small clear synaptic vesicles store classical neurotransmitters such as acetylcholine, gamma-aminobutyric acid (GABA), glycine, and glutamate. However, monoamines differ from other classical transmitters and have been reported to appear in both LDCVs and smaller vesicles. To localize the transporter that packages monoamines into secretory vesicles, we have raised antibodies to a COOH-terminal sequence from the vesicular amine transporter expressed in the adrenal gland (VMAT1). Like synaptic vesicle proteins, the transporter occurs in endosomes of transfected CHO cells, accounting for the observed vesicular transport activity. In rat pheochromocytoma PC12 cells, the transporter occurs principally in LDCVs by both immunofluorescence and density gradient centrifugation. Synaptic-like microvesicles in PC12 cells contain relatively little VMAT1. The results appear to account for the storage of monoamines by LDCVs in the adrenal medulla and indicate that VMAT1 provides a novel membrane protein marker unique to LDCVs.

Adrenal Medulla↗
PubMed1994

Absolute measurements of total-body calcium using prompt gamma neutron analysis and internal chlorine standardization: results in renal patients.

Measurement of total-body calcium (TBCa) is important in the study of several medical conditions. Neutron activation analysis (NAA) is sufficiently precise for serial measurements, but in order to provide single, absolute determinations of TBCa, it requires a correction for body habitus. A method is described for the absolute determination of TBCa by comparing the total-body chlorine, obtained by NAA, with that obtained by stable bromine dilution. The method is applied to a group of thirty patients who had been treated for chronic renal failure, and significant differences are found between measured values of TBCa and those predicted for 'normal' healthy subjects. The overall experimental error in the absolute determination of TBCa is estimated to be 8%, for a dose equivalent of 3.1 mSv (based on skin dose, and a quality factor of 10 for neutrons).

Body Composition↗
PubMed1994

Epidemiology of varicose veins. A review.

Disease of the venous system is a major problem affecting western societies, resulting in considerable morbidity in the population and cost to the health service. In many countries "varicose veins are probably the commonest disorder presenting to general surgeons" and an average of 30% of district nursing time is estimated to be spent caring for patients with venous ulcers. For the patient with varicose veins or leg ulceration, there is often persistent discomfort and disability extending over long periods of time. Despite this, little epidemiological research has been carried out on venous disease, perhaps partly because of society's perception that venous disease is not a major problem and it is not normally a cause death. More recently however, efforts have been made to conduct structured epidemiological studies to identify risk factors and to clarify the geographical variations suggested in the past by anecdotal evidence. This article reviews recent epidemiological studies, discusses the prevalence of varicose veins and presents evidence for and against the differing theories of causation.

Adolescent↗
PubMed1993

The measurement of silicon in a lung phantom--a comparison of two nuclear reactions for in vivo activation analysis.

The amount of silica in the human lung may be estimated by measurement of silicon using in vivo neutron activation analysis. A pulsed, fast neutron beam, produced with a 2 MV Van de Graaff generator using the 2H + 2H reaction, was used to irradiate a Si-doped chest phantom in order to determine minimum detection limits (MDL). Two 'in-beam' nuclear reactions on Si were studied; prompt fast neutron inelastic scatter 28Si (n,n' gamma)28Si reaction was measured during the beam burst and the slow neutron prompt capture reaction was measured between the fast neutron bursts. Although the latter reaction appeared less favourable due to neutron cross section and measurement efficiency considerations, it yielded an MDL of 1.8 g compared with 2.3 g for the 28Si(n,n' gamma)28Si reaction. A comparison was made with a 252Cf neutron irradiation system where a Si MDL of 6.3 g was obtained using the slow neutron capture reaction. The Van de Graaff system permits 'exposed' Si lung burdens to be measured but not normal levels. Improved measurement sensitivity may be achieved by reduction of high counting-rate losses and high background radiation.

Humans↗
PubMed1993

A five-compartment model of body composition of healthy subjects assessed using in vivo neutron activation analysis.

A body composition study of 31 healthy subjects covering a wide range of age (23.5-72.0 years) and weight (44.5-104.2 kg) has been undertaken. Subjects were assessed by in vivo neutron activation and tritiated water analysis and values of total body nitrogen, hydrogen and fat obtained by utilization of a five-compartment model of body composition comprising protein, water, fat, minerals and glycogen. The protein (as 6.25 x nitrogen) and water compartments were measured but the smaller compartments of minerals and glycogen were calculated as fixed fractions of the fat-free mass estimated from the water space. Fat was calculated as the body mass less the sum of the four other compartments. Mean values (+/- SEM), expressed as a percentage of body mass, for nitrogen, hydrogen and fat were 2.56 (+/- 0.07)%, 10.07 (+/- 0.04)%, and 21.9 (+/- 1.7)% respectively for men and 2.14 (+/- 0.07)%, 10.40 (+/- 0.04)%, and 35.5 (+/- 1.7)% respectively for women. The accuracy of the nitrogen measurements was evaluated by comparison with calculated values from two prediction equations; correlation coefficients, the mean bias (estimated from the mean differences between the measured and predicted nitrogen), the confidence interval for the bias, and limits of agreement were calculated. The correlation coefficients were high (r > 0.93) and the mean bias indicative of agreement. The ratio of nitrogen to the fat-free mass (derived from the body composition model) was also calculated and mean values (+/- SEM) of 32.7 (+/- 0.4) and 33.1 (+/- 0.4) g/kg for men and women, respectively were obtained. The hydration of the fat-free mass was determined to be 0.725 (+/- 0.002) and 0.722 (+/- 0.002) kg/kg for men and women respectively. The accuracy of the body fat estimate was evaluated by comparison with skinfold-thickness-derived values and computation from tritiated water space. The ratio of the body composition model to skinfold-thickness-derived fat was significantly (P < 0.005) greater than unity. The mean bias between the body composition model and tritiated-water-derived fat was -0.6 percentage points of fat (95% confidence interval from -0.3 to -0.9 percentage points of fat). Finally a prediction equation (r2 = 0.908, SEE = 108 g) for body nitrogen in healthy subjects based on weight, age and sex was calculated.

Adipose Tissue↗
PubMed1993

Agonist-induced state of the delta-opioid receptor that discriminates between opioid peptides and opiate alkaloids.

We report a novel agonist-induced change in delta-opioid receptor binding properties in NG108-15 cells. Pretreatment of these cells with opioid agonists substantially diminishes the binding of peptide agonists and a peptide antagonist to opioid receptors in intact cells or membrane preparations. However, similar agonist-induced changes in the binding of opiate alkaloid agonists and antagonists were not detected. The change in opioid peptide binding occurs rapidly at 37 degrees (t1/2 approximately 10 min) but is not induced by agonist treatment at 4 degrees. Because of its lability at 37 degrees, the binding change is only detected when equilibrium binding assays are performed at 4 degrees. Both alkaloid and peptide agonists induce the binding change in a dose-dependent manner, with an ED50 for etorphine of approximately 10 nM. The induction of the binding change is completely blocked by the opiate antagonist naloxone. Stimulation of muscarinic receptors (which, like opioid receptors, inhibit adenylate cyclase in these cells) does not induce or block the binding change. These data reveal the operation of a homologous regulation mechanism that rapidly diminishes the interaction of delta-subtype opioid receptors with peptide ligands but does not detectably change the interaction of receptors with alkaloid ligands.

Alkaloids↗
PubMed1992

Cloning of a delta opioid receptor by functional expression.

Opiate drugs have potent analgesic and addictive properties. These drugs interact with receptors that also mediate the response to endogenous opioid peptide ligands. However, the receptors for opioids have eluded definitive molecular characterization. By transient expression in COS cells and screening with an iodinated analog of the opioid peptide enkephalin, a complementary DNA clone encoding a functional delta opioid receptor has been identified. The sequence shows homology to G protein-coupled receptors, in particular the receptors for somatostatin, angiotensin, and interleukin-8.

Amino Acid Sequence↗
PubMed1992

Multicenter evaluation of the novel ABN Western blot (immunoblot) system in comparison with an enzyme-linked immunosorbent assay and a different Western blot.

A new, modular Western blot (immunoblot) system for human immunodeficiency virus (HIV) antibodies (ABN WesPage; Wellcome) was compared with enzyme immunoassays (Wellcome, Behringwerke, and Abbott) and with a U.S. Food and Drug Administration (FDA)-licensed Western blot (DuPont) in a multicenter study. A total of 649 serum samples from HIV patients at different stages of the disease, as well as from high-risk patients, from patients with conditions unrelated to AIDS, and from healthy blood donors, were used in the evaluation along with nine seroconversion panels. For evaluation of Western blot reactivity, both Centers for Disease Control (CDC) and FDA criteria were used. With the DuPont Western blot as the reference assay, the overall sensitivity and specificity of the ABN WesPage were 100 and 99.1%, respectively, when indeterminate results were not taken into account and when both tests were interpreted in accordance with CDC criteria. The DuPont Western blot detected significantly more antibodies to pol and gag gene products than the ABN WesPage. The ABN WesPage showed a higher positive rate of detection of viral envelope band gp160. When both Western blots were interpreted in accordance with CDC criteria, the ABN WesPage and the DuPont Western blot yielded 9.3 and 10.4% indeterminate results, respectively. When the DuPont Western blot was interpreted in accordance with the manufacturer's instructions (FDA criteria), 25.7% of the samples tested were regarded as indeterminate. The choice of interpretation criteria is of paramount importance for the evaluation of HIV Western blot patterns.

AIDS Serodiagnosis↗
PubMed1991

Dual determination of extracellular cholecystokinin and neurotensin fragments in rat forebrain: microdialysis combined with a sequential multiple antigen radioimmunoassay.

Microdialysis was combined with a highly sensitive sequential multiple antigen radioimmunoassay to simultaneously measure extracellular cholecystokinin and neurotensin fragments from discrete regions of the rat brain in vivo. The assay was conducted in 96-well plates and provided a limit of detection for both peptides of 0.1 fmol. Dialysis membranes composed of polyacrylonitrile, Cuprophan and polycarbonate were evaluated in vitro using both radiolabelled peptides and radioimmunoassay. Polycarbonate probes were implanted in the posterior medial nucleus accumbens-septum, medial caudate nucleus or medial prefrontal cortex of halothane-N2O-anaesthetized rats. Cholecystokinin immunoreactivity levels were generally above the assay detection limits (0.1-0.7 fmol) in 30-min samples from all three regions under basal conditions. Recovered basal amounts of neurotensin immunoreactivity were detectable in the nucleus accumbens-septum in approximately 50% of experiments (0.1-0.2 fmol) but were not measured in the caudate nucleus or prefrontal cortex. In the nucleus accumbens-septum, a 10-min pulse of 200 mM K(+)-containing artificial cerebrospinal fluid in the perfusion medium during a 30-min sampling period increased the recovered cholecystokinin and neurotensin immunoreactivity to 9.7 fmol +/- 1.9 S.E.M. and 5.8 +/- 1.6 S.E.M., respectively. A second stimulation following a 2.5-h interval produced similar elevations with S2:S1 ratios of 0.62 +/- 0.07 and 0.68 +/- 0.07 for cholecystokinin and neurotensin, respectively. In a separate series of experiments the second stimulation of both peptides was prevented by perfusion of a 10 mM EGTA-containing medium. Similar results were obtained in the caudate nucleus for cholecystokinin, but K(+)-induced elevations in neurotensin immunoreactivity were much smaller (0.5 fmol) in this brain region and calcium dependency was not established. Sequential K+ stimulations at 50, 100 and 200 mM produced progressively greater increases in recovered cholecystokinin and neurotensin immunoreactivity from the nucleus accumbens-septum and of cholecystokinin immunoreactivity from the prefrontal cortex. No neurotensin immunoreactivity was detected in the prefrontal cortex following K+ stimulation. Large post mortem increases in the recovered amounts of cholecystokinin and neurotensin immunoreactivity were observed. This effect was significantly attenuated by EGTA although there was a large calcium-independent component of the cholecystokinin immunoreactivity. On reverse-phase high-performance liquid chromatography the major cholecystokinin-immunoreactive peak co-eluted with sulphated cholecystokinin octapeptide. Neurotensin-immunoreactive material co-eluted with neurotensin (1-13), neurotensin (1-12), neurotensin (1-11), neurotensin (1-10) and neurotensin (1-8). These results further demonstrate the potential of microdialysis for studying neuropeptide release and metabolism in vivo when combined with sufficiently sensitive assay procedures.

Animals↗
PubMed1991

Focal stimulation of the mossy fibers releases endogenous dynorphins that bind kappa 1-opioid receptors in guinea pig hippocampus.

Physiological release of endogenous opioids in guinea pig hippocampal slices was detected in an in vitro competition binding assay using [3H]U69,593, a kappa 1-selective radioligand. Veratridine-induced opioid release caused a decrease in [3H]U69,593 binding that was blocked by either tetrodotoxin addition or the removal of calcium from the incubation buffer. Focal electrical stimulation of opioid peptide-containing afferent pathways resulted in a decrease in [3H]U69,593 binding, whereas stimulation of a major afferent lacking endogenous opioid immunoreactivity had no effect. The addition of 6-cyano-7-nitroquinoxaline-2,3-dione blocked the reduction in [3H]U69,593 binding caused by perforant path stimulation, but not the reduction caused by mossy fiber stimulation, suggesting that the primary source of endogenous kappa ligands was likely to be the dentate granule cells. Antisera against dynorphin A(1-8) or dynorphin B peptides inhibited the effects of mossy fiber stimulation in the [3H]U69,593 displacement assay. Antisera against other prodynorphin- and proenkephalin-derived opioid peptides had no effect. As shown by receptor autoradiography, the distribution of kappa 1 binding sites was limited to the molecular layer of the dentate gyrus and the presubiculum region of temporal hippocampal slices. These results indicate that prodynorphin-derived opioids released under physiological conditions from the mossy fibers act at kappa 1 receptors in the guinea pig dentate gyrus.

6-Cyano-7-nitroquinoxaline-2,3-dione↗
PubMed1991

Isolation and characterization of opioid peptides from rabbit cerebellum.

The rabbit cerebellum has been shown to contain significant quantities of opioid receptors consisting of both mu- and kappa-subtypes. To determine the nature of the endogenous opioid ligands in this tissue, extracts from rabbit cerebellum were separated by various chromatography techniques and fractions were assayed initially for opioid peptides with a radioimmunoassay capable of detecting all peptides with an amino-terminal Tyr-Gly-Gly-Phe sequence. This sequence is common to all mammalian opioid peptides and is critical for recognition by all known opioid receptors. Each of the three immunoreactive opioid peptide peaks detected was purified to homogeneity and subjected to amino acid composition and sequence analysis. One peak was analyzed further by mass spectrometry. This identified the major opioid peptides in the cerebellum as [Met5]enkephalin, [Leu5]enkephalin, and heptapeptide [Met5]enkephalyl-Arg6-Phe7. The comprehensiveness of this initial detection scheme in identifying biologically active opioid peptides was substantiated through subsequent analysis. Using specific radioimmunoassays for representative opioid peptides of the three opioid systems currently known, no other peptides of either the proenkephalin, proopiomelanocortin, or prodynorphin series were detected in any appreciable amounts. Collectively, these results are consistent with the position that rabbit cerebellar opioids are derived from proenkephalin. However, given that no appreciable quantities of either [Met5]enkephalyl-Arg6-Arg7-Val8-NH2 (metorphamide) or [Met5]enkephalyl-Arg6-Gly7-Leu8 were detected suggests that rabbit proenkephalin may have a slightly altered sequence and/or is differentially processed relative to other mammalian species studied.

Amino Acid Sequence↗