PubMed HealthSearch

Biomedical subjects

D S Munro

Publications and source records attributed to D S Munro.

At least 19 recordsLinked to original sources

Human infection in Tayside, Scotland due to Salmonella serotype Livingstone.

Livingstone was the third most common salmonella serotype isolated from cases of human salmonellosis in the Tayside region of Scotland in 1989-1991; latterly, it spread to Grampian region. The significant upsurge of Livingstone in these two Scottish regions was not matched by similar increases in its frequency of isolation from human cases of salmonellosis in other regions of Scotland or elsewhere in the UK. Although Salmonella Livingstone is usually associated in the UK with incidents of infection among poultry flocks, our detailed investigations found no clear evidence that poultry, eggs or poultry-related products were responsible for this outbreak. Most cases occurred in the summer months from July to September and many of the patients required hospital treatment. Other than one outbreak among geriatric patients in a long-stay hospital in north Tayside, most of the cases were sporadic. The extent of the outbreak, covering 3 years, was recognised mainly because Livingstone was previously an uncommon serotype in Tayside. There were few Livingstone isolations from non-human sources in Scotland in these same years. Possible sources of infection and predisposing factors among patients are discussed. Livingstone was not isolated in Scotland in 1992.

Adolescent

Measurement of thyroid stimulating immunoglobulins in a new cell line transfected with a functional human TSH receptor (JPO9 cells), compared with an assay using FRTL-5 cells.

OBJECTIVE: Most bioassays used to measure thyroid stimulating antibodies (TSAb) rely on the ability of patient sera to generate cAMP in cultured cells of non-human origin. The cell line most commonly used has been the rat thyroid cell line, FRTL-5. Recently, a new cell line (JPO9) possessing a transfected human TSH receptor has been developed. The aim of this study was to evaluate the JPO9 cells by comparing them with the widely used FRTL-5 cells, using the method of 3H-adenine incorporation for the direct measurement of intracellular cAMP. DESIGN: The ability of patient sera to generate cAMP production in JPO9 and FRTL-5 cells was used as the index of thyroid stimulation. The specificity of the assay was determined using sera from patients with autoimmune thyroid and non-thyroid autoimmune diseases. PATIENTS: We studied sera from 28 patients with newly diagnosed Graves' disease, 72 patients with Graves' disease at various stages of treatment, 8 patients with Hashimoto's thyroiditis, 40 patients with a variety of non-thyroid autoimmune disease and 42 control subjects. MEASUREMENTS: The intracellular 3H-cAMP generated in JPO9 cells following preincubation with 3H-adenine and subsequent incubation with patient or control sera, was used as a measure for TSAb. The results obtained with these cells were compared to those obtained with the widely used FRTL-5 cells and they were also correlated with the measurement of TSH binding inhibitory immunoglobulins by a conventional radioreceptor assay. RESULTS: JPO9 cells were more sensitive and had a wider range of response to bovine TSH (bTSH) than had FRTL-5 cells (10(-7)-10(-2) U/ml compared to 10(-5)-10(-3) U/ml respectively). Unlike FRTL-5 cells, JPO9 cells respond well to unmodified serum and grow constitutively in the absence of TSH, thereby not requiring TSH deprivation prior to assay. The TSAb values obtained with both cell lines correlated well (r = 0.87). The JPO9 cells responded specifically to Graves' sera (23 out of 28 newly diagnosed patients were positive), whereas no patients with Hashimoto's thyroiditis and only 4 of 40 patients with non-thyroid autoimmune diseases gave low positive results for the presence of TSAb. CONCLUSIONS: The JPO9 cells provide similar diagnostic information to FRTL-5 cells in patients with autoimmune thyroid disease. However, because they are more sensitive, grow faster, have less fastidious growth requirements and respond to unextracted sera, compared to FRTL-5 cells, we conclude that the JPO9 cells are preferable for the measurement of TSAb.

Animals

An investigation of the ability of TSH and Graves' immunoglobulin G to increase intracellular calcium in human thyroid cells, rat FRTL-5 thyroid cells and eukaryotic cells transfected with the human TSH receptor.

The purpose of this study was to determine if immunoglobulin G preparations (IgGs) from patients with Graves' disease can increase intracellular calcium in thyroid cells, as has been reported for TSH. Both TSH and Graves' IgGs (prepared by protein G affinity chromatography) increased calcium in a range of thyroid cells; however, the response seen, using Fura-2-loaded coverslips of cell monolayers, varied considerably. Chinese hamster ovary (CHO/JPO9) cells transfected with a high number of human TSH receptors showed the greatest response: TSH (10 mU/ml) increased calcium in 46% of experiments and 18 out of 25 (72%) Graves' IgGs increased calcium at 0.1 mg/ml (significantly greater, P < 0.001, than for control IgGs where cells responded to 2 out of 13 preparations). Rat FRTL-5 cells only responded to TSH in 22% of experiments and to 2 out of 8 (25%) of Graves' IgGs. Similarly, human thyroid cells responded to TSH in 22% of experiments and to 2 out of 9 (22%) of Graves' IgGs. (When studying cyclic AMP responses in JPO9 cells, much higher concentrations of Graves' IgGs were required (1-3 mg/ml). However, higher concentrations (0.3 mg/ml) of both Graves' IgGs, and to a lesser extent of control IgGs, were capable of increasing calcium in cells both with and without TSH receptors (control CHO cells and normal human dermal fibroblasts). We conclude that relatively low concentrations of patient IgGs can be distinguished from control IgGs in JPO9 cells on the basis of their ability to increase calcium, but that additionally all IgG preparations possibly contain another factor which can increase calcium in a range of cells independent of the presence of the TSH receptor.

Adolescent

Thyroid stimulating immunoglobulins, like thyrotropin activate both the cyclic AMP and the PIP2 cascades in CHO cells expressing the TSH receptor.

In human thyrocytes and in a permanent CHO cell line expressing the human thyroid stimulating hormone (TSH) receptor cDNA (JP09 cells), TSH activates both the cyclic AMP and the phosphatidylinositol 4,5-bisphosphate (PIP2) cascade, although the latter effect requires higher TSH concentrations. Thyroid stimulating autoantibodies (TSAb) activate also the human thyroid leading to the hyperthyroidism of Graves' disease. They bind to the TSH receptor and mimic the TSH stimulation of the gland by increasing intracellular cyclic AMP, but they do not enhance PIP2 hydrolysis in human thyroid slices. We show in this study that TSAb are able to activate the PIP2 cascade in JP09 cells, a cell line expressing high levels of TSH receptor. This suggests that the mechanism of action of TSAb on the TSH receptor is qualitatively similar to that of TSH.

Animals

The epidemiological relationship between Salmonella isolated from poultry meat and sewage effluents at a long-stay hospital.

Between February 1988 and March 1989 chicken carcases delivered to the kitchen of a long stay psycho-geriatric hospital were screened every week for salmonella contamination. While 214 of 477 (45%) individual carcases carried one or more salmonella types, every single consignment examined contained affected carcases. Simultaneously, sewers draining the residential accommodation and excluding kitchen effluent, were also monitored. Thirty out of 79 (38%) of Moore's swabs were positive for salmonella. There was a statistically significant association between the salmonella types isolated from chicken and those isolated from sewers the following week. Following a change in kitchen policy to order only cooked chicken there was a significant reduction in the isolation of salmonella from the sewers.

Animals

Clinical outcome and costs of care in radioiodine treatment of hyperthyroidism.

The long-term clinical outcome and costs of treatment of hyperthyroidism with radioiodine have been examined in two cohorts of patients from Sheffield and Scotland. The majority of patients in both series were considered to have Graves' disease. The Sheffield patients (660) were included in a trial of three radioiodine dose regimens of 3,500 (312), 7,000 (323) and 14,000 (25) rad determined using a formula for accurate dosimetry. The Scottish patients (3,920) drawn from five centres in Aberdeen, Dundee, Edinburgh, Glasgow and Inverness were treated using an arbitrary scale, for the activity of radioiodine administered, related to goitre size. Their results are grouped into five MBq 'dose' bands: 37-185, 186-370, 371-555, 556-740 and 741+. The proportion of patients with persistent hyperthyroidism was higher in both cohorts for low-dose radioiodine regimens, but 15-25% of patients who received high doses showed persistent hyperthyroidism. Early and late onset hypothyroidism was lower after low doses but differences between the treatment groups were small in terms of clinical benefit. Total morbidity at 10 years follow-up, in terms of hyperthyroidism, and hypothyroidism, was highest after low-dose therapy. There was little variation in total costs, but patient costs were lowest for the Scottish regimen and highest for low-dose therapy. A dose of at least 370-555 MBq which will ensure early elimination of hyperthyroidism will also limit the medical workload and total costs.

Adult

A role for insulin-like growth factor-I in the regulation of human thyroid cell growth by thyrotrophin.

Human thyroid epithelial cells in monolayer culture were found to release radioimmunoassayable insulin-like growth factor-I (IGF-I) over a 48-h culture period in serum-free medium. In the presence of supraphysiological concentrations of TSH (1-100 mU/ml) known to be inhibitory to DNA synthesis by human thyroid cells, the release of IGF-I was found to be inhibited in six thyroid cultures studied. In only one out of the six was IGF-I release increased in the presence of physiological mitogenic concentrations of TSH (0.1-100 microU/ml). Human thyroid fibroblasts, established by long-term culture of thyroid epithelial cells under fibroblast-selective conditions, also secreted IGF-I which was unaffected by the presence of TSH at both low and high concentrations. Using a monoclonal antibody against human IGF-I, monolayer cultures of both human thyroid epithelial cells and human thyroid fibroblasts showed positive immunocytochemical staining for IGF peptide. However, fixed sections of intact thyroid tissue only showed positive staining for IGF peptide associated with the fibrous layers surrounding the thyroid follicle, with no staining of the follicular epithelial cells. The growth of human thyroid epithelial cells was also found to be increased by IGF-I (25-100 ng/ml) added in medium plus 1% fetal calf serum as assessed by the incorporation of [3H]thymidine into DNA. In the presence of a monoclonal antibody to IGF-I the increase in [3H]thymidine uptake in response to IGF-I was abolished as was that seen in response to TSH.(ABSTRACT TRUNCATED AT 250 WORDS)

Cell Division

Extracellular calmodulin and its association with epidermal growth factor in normal human body fluids.

In this study we describe the occurrence of a calmodulin-like protein in normal human biological fluids. Extraction of the calmodulin-like protein from breast milk, saliva, serum and urine provided an extract with enhanced calmodulin immunoreactivity which, in the case of milk and saliva, showed a protein band comigrating with authentic calmodulin (Mr 17,000) on sodium dodecylsulphate-polyacrylamide gel electrophoresis. However, in milk, saliva and serum a major protein band of Mr 14,000-15,000 was always observed, which we speculate may be related to calmodulin, possibly as a partially degraded form. Estimates of biologically active calmodulin in most normal extracellular fluids were of the order which we have found will stimulate cell division when added to the extracellular medium of cells in culture. Levels ranged from 0.03 nmol/l in urine to 18.6 nmol/l in breast milk, and exhibited a quantitative relationship (r = 0.79, P less than 0.01) to epidermal growth factor (EGF) levels in fluids. Where EGF concentrations varied from normal (increased in saliva 24 h after oral surgery and reduced in the urine of patients with renal failure) calmodulin concentrations were similarly affected. The presence of calmodulin in serum may in part be attributable to its release from platelets which are particularly rich in calmodulin. Release of calmodulin from the platelet was associated with that of EGF and other platelet products.

Blood Platelets

Old and new techniques together resolve a problem of infection by Salmonella typhimurium.

Isolates of Salmonella typhimurium, recovered over a 9-month period from a child with gastroenteritis, were characterized by biotyping, phage-typing and plasmid-profile analysis. Because the data from the different methods were discrepant, it was difficult to establish whether her infection was due to a single strain that had changed character in vivo or was due to recurrent infections with different, unrelated strains. Restriction-enzyme fingerprinting of the plasmids from the different isolates provided an explanation for the initial discrepancy and high-lighted a source of potential confusion in epidemiological studies.

Bacteriophage Typing

A comparative study of the anti-proliferative effects of calmodulin antagonists in cultured cells--W7 derivatives of improved cytostatic potential.

We have compared the effects of the calmodulin antagonist N-(6-aminohexyl)-5-chloro-1-naphthalene-sulphonamide (W7) and relatives of the parent compound, modified by substituting the 5-chloro- for an iodo-residue and increasing stepwise the length of the carbon chain from 6 to 12, for their ability to inhibit cell proliferation in tissue culture. These species showed improved specificity and potency, as determined against a calcium calmodulin-dependent beef heart phosphodiesterase in vitro, exhibited time courses of inhibition similar to the parent compound W7, but were more potent at inhibiting DNA synthesis in K562 human leukaemic cells cultured in serum-free medium. The elevation observed in the percentage of cells in G1 of the cell cycle following drug exposure indicated that these drugs, like W7, arrested growth by inhibiting entry of cells into and through S phase. Higher doses of all drugs were irreversibly cytotoxic as determined by the inability of the cells to recover DNA synthetic capacity and to form colonies in soft agar or to recover their normal cell cycle distribution. We also discuss the possible implications of extracellular calmodulin and antagonism thereof on cell proliferation.

Animals

Recognition of the cryptic plasmid, pSLT, by restriction fingerprinting and a study of its incidence in Scottish Salmonella isolates.

The plasmid pSLT is a cryptic plasmid of 60 megadaltons (Md) present in Salmonella typhimurium LT2. We present evidence that it has a characteristic fingerprint when digested with the restriction enzymes PstI and SmaI. Among a representative collection of S. typhimurium isolates it was present in 67% of strains and was widely distributed amongst different phage types (DT) with the exception of DT10 and U285. Furthermore, its prevalence among veterinary isolates was significantly higher than among human isolates. It was not found among any of the 96 strains representative of other salmonella serotypes currently prevalent and thus appears to be serotype-specific.

Chromosome Mapping

The distribution of plasmids among a representative collection of Scottish strains of Salmonellae.

The distribution of plasmids was studied in a representative collection of salmonella strains which comprised 98 Salmonella typhimurium and 96 other serotypes. Plasmids were detected in 72% of strains (mean 1.3 plasmids/strain) and individual strains harboured between 0 and 7 plasmids. They were more common among S. typhimurium than other serotypes (incidence 92 and 53%; mean 1.9 and 0.8 plasmids/strain respectively). Although a higher proportion of S. typhimurium (33%) were antibiotic-resistant compared to other serotypes (14%) the evidence presented indicated that R-plasmids were not responsible for the difference observed in the number and distribution of plasmids in these strains. These results were discussed in comparison with similar studies of Escherichia coli and other enteric genera.

DNA, Bacterial

Relationship between growth and function of human thyroid cells in culture.

Subconfluent human thyroid cells in monolayer, isolated from thyrotoxic tissue or non-toxic goitres obtained at surgery, responded to the addition of epidermal growth factor (EGF) with an increase in cell growth as measured by increased incorporation of [3H]thymidine into trichloroacetic acid-precipitable material. The growth response to EGF was concentration-dependent and the characteristics of the responses were the same using EGF from murine or human sources. With concentrations which stimulated growth, EGF was found to inhibit human thyroid cell function as measured by the release of radioimmunoassayable tri-iodothyronine into the incubation medium. Thyrotrophin (TSH) was also found to stimulate human thyroid cell growth but at concentrations far lower than those used to stimulate thyroid cell function in this system. The effect of EGF on the differentiating action of TSH on human thyroid cells in culture was also investigated; the association of thyroid cells into two-dimensional follicular structures produced by the incubation of thyroid cells at a high cell density with TSH was found to be inhibited by the addition of EGF.

Cells, Cultured

Calcium and calmodulin in the regulation of human thyroid adenylate cyclase activity.

TSH (thyrotropin)-stimulated human thyroid adenylate cyclase has a biphasic response to Ca2+, being activated by submicromolar Ca2+ (optimum 22nM), with inhibition at higher concentrations. Calmodulin antagonists caused an inhibition of TSH-stimulated adenylate cyclase in a dose-dependent manner. Inhibition of TSH-and TSIg-(thyroid-stimulating immunoglobulins)-stimulated activity was more marked than that of basal, NaF- or forskolin-stimulated activity. This inhibition was not due to a decreased binding of TSH to its receptor. Addition of pure calmodulin to particulate preparations of human non-toxic goitre which had not been calmodulin-depleted had no effect on adenylate cyclase activity. EGTA was ineffective in removing calmodulin from particulate preparations, but treatment with the tervalent metal ion La3+ resulted in a loss of up to 98% of calmodulin activity from these preparations. Addition of La3+ directly to the adenylate cyclase assay resulted in a partial inhibition of TSH- and NaF-stimulated activity, with 50% inhibition produced by 5.1 microM and 4.0 microM-La3+ respectively. Particulate preparations with La3+ showed a decrease of TSH- and NaF-stimulated adenylate cyclase activity (approx. 40-60%). In La3+-treated preparations there was a decrease in sensitivity of TSH-stimulated adenylate cyclase to Ca2+ over a wide range of Ca2+ concentrations, but most markedly in the region of the optimal stimulatory Ca2+ concentration. In particulate preparations from which endogenous calmodulin had been removed by La3+ treatment, the addition of pure calmodulin caused an increase (73 +/- 22%; mean +/- S.E.M., n = 8) in TSH-stimulated thyroid adenylate cyclase activity. This was seen in 8 out of 13 experiments.

Adenylyl Cyclase Inhibitors

The pathogenesis of Graves' disease.

The abnormally increased thyroid activity that is characteristic of Graves' disease is caused by immunoglobulins which specifically interact with the thyroid cell and stimulate it. Increases and decreases in thyroid activity in Graves' disease can be clearly related to rise and fall of these immunoglobulin-mediated activities. The level of immunoglobulin stimulatory activity can be used for prediction of the likelihood of neonatal Graves' disease and of recurrence of disease after cessation of treatment with antithyroid drugs. Investigation of patients with Graves' disease and their families has led to identification of particular human leukocyte antigens and genetically linked markers on immunoglobulins which both appear to incur increased susceptibility to certain autoimmune diseases. Differences in immune function, when compared with control populations, have been found in patients with these genetically linked markers. Protection against autoimmune disease is maintained by purposeful inhibition of any self-directed activity within each function of the immune system and by the controlling interaction of other immune functions. No single deficiency of immune function can be selected as giving the major risk of autoimmune disease, but rather a sum of relative defects resulting in an increased risk. In some patients with Graves' disease the self-protection mechanisms regain sufficient control of the immune functions to reduce the activity of the autoimmune disease, and the patient may achieve clinical remission. Often, however, there is evidence that abnormal immune activity directed against thyroid tissue has persisted with liability to recurrence of the Graves' disease.

Autoantibodies

An epidemiological study of Salmonella montevideo by biotyping.

Among 622 cultures of Salmonella montevideo, 27 biotypes belonging to two biogroups were recognized. One biogroup (10di) was predominant in all animals in Scotland but only in sheep in England and Wales. The other (biogroup 2d) was responsible for almost all human, cattle and poultry infection in England and Wales, but only 24% of human infection in Scotland.

Animals

Adrenal scintigraphy with 75Se selenonorcholestenol: a review.

We have reviewed 58 patients on whom adrenal scintigraphy has been performed using 75Se selenonorcholestenol. For 15 patients whose adrenal function was biochemically normal, the upper limit of normal of the 7 day adrenal uptake test was 0.45%, considerably higher than the generally accepted value of 0.3%. There is evidence from this group of patients that stress and obesity might account for uptakes in the range 0.3-0.45%. The sensitivity of the uptake test is poor, with 7 out of 23 patients with Cushing's syndrome having uptakes within the normal range. Scintigraphy of such patients may still be useful in differentiating between unilateral and bilateral adrenal involvement.

Adrenal Cortex