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Biomedical subjects

D Slater

Publications and source records attributed to D Slater.

At least 55 records · Page 3Linked to original sources
PubMed1993

Production of biologically active light chain of tetanus toxin in Escherichia coli. Evidence for the importance of the C-terminal 16 amino acids for full biological activity.

The activity of the light (L) chain of tetanus toxin, and of mutants constructed by site-directed mutagenesis, was studied by expression and purification of the proteins from E. coli. Wild-type recombinant L chain (pTet87) was active in the inhibition of exocytosis from cultured bovine adrenal chromaffin cells, although at a level 5-15% of that of L chain purified from tetanus toxin. L chain mutants which terminated at Leu-438 (pTet89), or which contained a Cys-to-Ser mutation at residue 439 (pTet88) were equally as active as the full-length recombinant protein. The reduced activity of pTet87 L chain correlated with C-terminal proteolysis of the protein upon purification. A tryptic fragment derived from native light chain and which terminated at Leu-434 also showed reduced activity in the exocytosis assay, consistent with a requirement of the C-terminal region of the L chain for maximal activity. pTet87 L chain, but neither of the mutants, could be associated with purified H (heavy) chain to form a covalent dimer which induced the symptoms of tetanus in mice. The ability to form biologically active toxin using recombinant L chain will be of great value in structure-function studies of tetanus toxin.

Adrenal Medulla↗
PubMed1993

Effective control of hypercholesterolemia in children with dietary interventions based in pediatric practice.

BACKGROUND: Based on recent recommendations, the number of hypercholesterolemic children who would require dietary therapy could overwhelm current preventive pediatric cardiology resources. No previous studies have established the efficacy of such therapy in the pediatrician's office. The purpose of this study was to evaluate two programs of office-based therapy. METHODS: We randomly assigned 295 children with hypercholesterolemia (> 185 mg/dl) two interventions: one single or four multiple 90-min sessions of family-oriented nutritional education, based in pediatric practices. We examined total cholesterol, 3-day food records, height and weight, and in the multisession group high-density lipoprotein cholesterol and triglycerides at the beginning and at intervals of 8.5-9 and 21 to 33 weeks (single-session and multisession groups, respectively). RESULTS: Total cholesterol was lowered equally in both treatment groups over the course of the study. This was accompanied by dietary changes: a decrease in calories derived from total and saturated fats, and increased intake of fiber, protein, and carbohydrate. However, more single-session patients withdrew from the program during the study. CONCLUSIONS: The two interventions were equally effective in lowering total and low-density lipoprotein cholesterol and in reducing intake of total and saturated fat. However, the higher completion rate of the multisession group suggests that this approach may be the more effective.

Adolescent↗
PubMed1993

Expression of a common cellular phospholipase A2 by human intrauterine tissues.

A subclone of the three prime (3') end of a human extrapancreatic phospholipase cDNA has been generated using the polymerase chain reaction. This has been used to detect mRNA transcripts in RNA extracted from human amnion, chorion-decidua, placenta and myometrium. The carboxyl-terminus of phospholipases A2 is poorly conserved but hybridisation remained stable under conditions of high stringency. This supports the hypothesis that these tissues express a common cellular phospholipase A2 which is identical to that expressed in platelets.

Amnion↗
PubMed1993

Changes in amniotic arachidonic acid metabolism associated with increased cyclo-oxygenase gene expression.

OBJECTIVES: To study the differences in the metabolism of arachidonic acid, to prostaglandins and other eicosanoids, between amnion cells before and after labour. To study the changes in the expression of the type 1 cyclo-oxygenase gene associated with the changes in arachidonic acid metabolism. DESIGN: Amnion cells collected before labour, at elective caesarean section, and following spontaneous labour and delivery were established in mono-layer culture. Intracellular arachidonic acid pools were radio-labelled and the metabolic fate of endogenous archidonic acid was then studied using high performance liquid chromatography. RNA was extracted from the cell cultures and expression of the cyclooxygenase gene was studied using northern hybridisation to a sheep cyclo-oxygenase cDNA. RESULTS: Metabolism of endogenous arachidonic acid in amnion cells established in culture prior to labour is principally via lipoxygenase and epoxygenase enzyme pathways to di- and mono-HETEs and EET with little cyclo-oxygenase metabolism to prostaglandins. Following labour there is a large increase in arachidonic acid metabolism with a change in the cyclo-oxygenase: lipoxygenase enzyme pathway ratio in favour synthesis of prostaglandin E2. Whilst synthesis of lipoxygenase metabolites doubles, the increase in prostaglandin E2 synthesis is ten fold. Expression of the cyclo-oxygenase gene is significantly greater in cell cultures established following spontaneous labour and delivery than in those established following elective caesarean section prior to labour. CONCLUSIONS: In association with labour there is an increase in arachidonic acid metabolism in amnion cells and a change in the ratio of metabolism via cyclo-oxygenase and lipoxygenase enzyme pathways to increase synthesis of prostaglandin E2. This change is associated with increased expression of the cyclooxygenase gene.

Amnion↗
PubMed1993

Particulate air pollution and hospital emergency room visits for asthma in Seattle.

Recent studies have associated short-term exposure to respirable particulate matter (PM10) exposure with peak flow decrements, increased symptoms of respiratory irritation, increased use of asthma medications, and increased hospitalization for asthma. Increased mortality from chronic respiratory disease has also been reported. To help confirm whether PM10 exposure is a risk factor for the exacerbation of asthma, we compiled daily records of asthma emergency room visits from eight hospitals in the Seattle area. In Poisson regressions controlling for weather, season, time trends, age, hospital, and day of the week, the daily counts of emergency room visits for persons under age 65 were significantly associated with PM10 exposure on the previous day. The mean of the previous 4 days' PM10 was a better predictor (p < 0.005). The relative risk for a 30 micrograms/m3 increase in PM10 was 1.12 (95% confidence interval 1.20 to 1.04). Daily PM10 concentrations never exceeded 70% of the current ambient air quality standards during the period. The consistency of investigations of the health effects of PM10 suggest that increased attention should be given to the control of particulate matter air pollution.

Adolescent↗
PubMed1993

Multianalyte assay system developed for drugs of abuse.

A simple, 10-min immunoassay system has been developed that simultaneously screens for five different classes of drugs of abuse in a urine sample. This system tests for amphetamines, cannabinoids, cocaine metabolites, opiates, and phencyclidine, and each assay has a specific preset cutoff concentration. Accuracy is > 99% for reporting positive or negative results for samples with 200% or 50%, respectively, of the cutoff concentrations of the drugs. Tests of a panel of 96 compounds yielded only three cases of nonspecific reactivity (at a drug concentration of 100 mg/L). Another panel of 12 compounds that could normally be found in urine samples was also evaluated and no interferences were observed. Concordance was > 95% between this system and the comparable automated immunoassays for detecting drugs of abuse. Greater than 98% of GC/MS-confirmed positive samples gave positive results with this assay system.

Autoanalysis↗
PubMed1992

Thermogenic actions of tryptophan in the rat are mediated independently of 5-HT.

Serotonin (5-HT) has been implicated in the central control of energy balance, via inhibition of food intake and stimulation of thermogenesis. Its rate of synthesis in brain is dependent on the availability of its precursor amino acid, tryptophan. The objective of the present study was therefore to investigate the thermogenic actions of tryptophan and to determine whether these actions are mediated by 5-HT. Central or peripheral injections of 5-HT (i.c.v.; 0.5-40 micrograms), 5-hydroxytryptophan (5-HTP) (i.c.v.; 20 micrograms) or tryptophan (i.p.; 20 mg/kg, i.c.v.; 12-60 micrograms) significantly increased resting oxygen consumption (VO2 by approximately 15-20%) in conscious rats, without apparent effects on physical activity. Small increases (5-7%) in VO2 were also observed following peripheral injections of aspartate or glycine (20 mg/kg) but not taurine, whilst central injections of tyrosine or leucine (15-18 micrograms) significantly increased VO2 by 15%. We have previously reported that the thermogenic and anorexic actions of 5-HT are mediated by corticotropin-releasing factor (CRF). In the present study, the thermogenic actions of 5-HTP, like those of 5-HT, were significantly reduced by pretreatment (5 min before) with the CRF antagonist alpha-helical CRF9-41 (25 micrograms, i.c.v.) or a polyclonal antibody to CRF. However, the thermogenic actions of tryptophan were not significantly modified by pretreatment with either the 5-HT antagonist, methysergide (20 micrograms, i.c.v.) or with the CRF antagonist or antibody and thus appear to act through different mechanisms to 5-HT.(ABSTRACT TRUNCATED AT 250 WORDS)

5-Hydroxytryptophan↗
PubMed1992

Use of the nirB promoter to direct the stable expression of heterologous antigens in Salmonella oral vaccine strains: development of a single-dose oral tetanus vaccine.

Plasmid pTETnir15, which directs the expression of the non-toxic immunogenic fragment C of tetanus toxin from the anaerobically inducible nirB promoter, was introduced into the Salmonella typhimurium aroA aroD live oral vaccine strain BRD509. The resulting strain, designated BRD847, was used to vaccinate orally BALB/c mice and was tested for plasmid stability and its ability to protect against a lethal tetanus toxin challenge. pTETnir15 was stably inherited by bacteria growing or persisting in the tissues of immunized mice whereas another BRD509 derivative, designated BRD753, harboring plasmid pTET85 which directs fragment C expression from the tac promoter, was highly unstable. Mice immunized with a single oral dose of BRD847 developed high levels of circulating anti-fragment C antibodies and were solidly protected against tetanus toxin challenge. Mice immunized with a single oral dose of BRD753 developed no detectable anti-fragment C antibodies. After boosting, antibodies were detected, but the mice were only partially protected against tetanus toxin challenge. Thus the use of an in vivo inducible promoter such as nirB may be a generally applicable approach to obtaining the stable in vivo expression of heterologous antigens in Salmonella vaccine strains.

Anaerobiosis↗
PubMed1992

Influence of cardiopulmonary bypass on water balance hormones in children.

OBJECTIVE: To determine the changes in the endocrine mechanisms of fluid balance after cardiopulmonary bypass in children. DESIGN: Prospective study; analysis of numbered plasma samples performed blind with respect to clinical data. SETTING: Regional paediatric cardiothoracic unit. PATIENTS: Nine patients, median age 4, range 2 to 9 years, five males. Patients under the age of 1 year were excluded because of the frequent blood sampling involved. MAIN OUTCOME MEASURES: Plasma concentrations of atrial natriuretic peptide (ANP), arginine vasopressin, plasma renin activity, aldosterone, noradrenaline and adrenaline, and urinary concentrations of cyclic guanosine monophosphate (cGMP) as measured by radioimmunoassay. RESULTS: After 30 minutes of cardiopulmonary bypass plasma atrial natriuretic peptide (ANP) decreased from (mean (SEM)) 151 (71) pg/ml to 52 (44) pg/ml (NS), and urinary production of its second messenger cyclic guanosine monophosphate (cGMP) decreased from 1286 (600) pmol/ml to 151 (414) pmol (p < 0.05). Other plasma concentrations of hormones studied did not change significantly although arginine vasopressin, adrenaline, and noradrenaline increased whereas aldosterone and plasma renin activity decreased. After cardiopulmonary bypass stopped there was an immediate and significant rise in plasma ANP, but within the next 24 hours plasma ANP declined significantly (p < 0.05), decreasing from 294 (49) pg/ml to 64 (29) pg/ml at 22 hours. In the postoperative period there was a significant correlation between plasma ANP and both mean fluid balance (r = 0.96, p < 0.001) and mean urine output (r = 0.97, p < 0.001). Plasma aldosterone peaked (p < 0.05) at 22 hours after operation, and argine vasopressin peaked (p < 0.05) at two hours and then declined (p < 0.05) to a trough at 24 hours. Plasma renin activity, adrenaline, noradrenaline, and urinary cGMP concentrations, and mean central venous pressure did not change significantly in the postoperative period. CONCLUSION: The changes documented show the differing pattern of release of water balance hormones invoked by cardiopulmonary bypass. The central role of ANP is shown by its strong correlation with urinary output and its similarly strong relation to fluid balance.

Aldosterone↗
PubMed1991

Ultrastructure of melatonin-responsive cells in the ovine pars tuberalis.

Functional receptors for melatonin have been localized and characterized on the pars tuberalis (PT) of a number of mammalian species, but the cell-type responsive to melatonin is unknown. The ultrastructure of the ovine pars tuberalis has been examined and these findings correlated with the functional response of the gland to melatonin. This study revealed that two secretory cell types predominate in the ovine PT, which differ in the abundance of dense-core granules. The most abundant of the cells are either agranular or very sparsely granulated and represent 90% of the total population, with the remaining 10% being composed of cells with abundant dense-core vesicles. Few follicular cells were observed. This ratio of secretory cell-types persisted in primary culture, with the two types non-separable by Percoll gradient centrifugation. Using forskolin, as a non-specific stimulant of adenylate cyclase, melatonin was shown to inhibit the formation of cyclic AMP by 80-90% in cells both before and after Percoll centrifugation. The results demonstrate that the agranular secretory cells of the ovine pars tuberalis are the melatonin responsive cell-type of this gland.

Animals↗