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Biomedical subjects

D B Ramsden

Publications and source records attributed to D B Ramsden.

At least 73 records · Page 4Linked to original sources
PubMed1988

Ioxynil and 3,5,3'-triiodothyronine: comparison of binding to human plasma proteins.

Ioxynil, an iodine-containing compound with structural similarity to T3 affects the thyroids of experimental animals. Binding to human plasma proteins was compared with that of T3, to gain insight into possible effects in humans. After incubating human serum with [125I]ioxynil, 2DIEP against anti-whole human serum and rocket immunoelectrophoresis against anti-TBPA, albumin or TBG were carried out. Ioxynil bound to TBPA but not to TBG nor albumin. The Ka calculated for the TBPA-ioxynil complex was 2.6 x 10(6) l.mol-1. Ioxynil competed for the iodothyronine-binding sites of TBPA.

Autoradiography↗
PubMed1988

The enhancement of radiographic images from a multiwire camera using a maximum entropy algorithm.

The multiwire camera (MWC) produces high speed, quantitative autoradiography of radiolabelled substances in two-dimensional systems. While greatly superior to film-based systems in respect of speed and quantitativity the MWC has significantly poorer spatial resolution (particularly for high energy beta-emitting radiolabels) and the performance is ultimately limited by the noise induced in the images by Poisson statistics and counter background. Processing the MWC images with a maximum entropy algorithm significantly improves the performance of the system in these respects. The algorithm has been tested using one-dimensional data taken from images of known tritium, 14C and 125I distributions. Processed images are visually more acceptable with improved quantitative accuracy and spatial resolution. Quantitative accuracy, calculated as the root mean square deviation between an image and the known sample activities, is 10-40% lower for processed images compared with original camera images. Spatial resolution, calculated from slopes in the images representing edges of activity in the sources, is improved by 20-40% for the processed images. The algorithm is used to improve a two-dimensional image from a biological study. The source distribution consisted of a set of circular dots of varying activity. The dots with lowest activity were barely discernible in the raw MWC image but are clearly resolved after processing. The algorithm used is simple and effective and executes acceptably quickly on a personal computer. It should prove useful in any context where the imaging performance of a system is limited by Poisson statistics.

Algorithms↗
PubMed1988

Synthesis and nuclear magnetic resonance spectroscopic, mass spectroscopic, and plasma amine oxidase inhibitory properties of analogues of 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine.

Seventeen analogues of 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine were synthesized using three reaction pathways: condensation of phenols with 1-methyl-4-piperidone, reaction of Grignard reagents with 1-methyl-4-piperidone followed by dehydration of the product, and aminomethylation of olefins. The identity of the products of synthesis was established by nuclear magnetic resonance spectroscopy, mass spectroscopy, and elemental analysis. Thirteen analogues were shown to inhibit the oxidation of benzylamine by bovine plasma amine oxidase. Increasing the length of the aliphatic chain of N-substituted analogues resulted in increased inhibition. In 4-phenyl-substituted analogues, both the position and electronic character of the substituent group affected the degree of inhibition.

1-Methyl-4-phenyl-1,2,3,6-tetrahydropyridine↗
PubMed1987

A comparison of the effects of bromocriptine and somatostatin on growth hormone gene expression in the rat anterior pituitary gland in vitro.

The effects of the dopamine agonist bromocriptine (BCR) have been compared with those of somatostatin (SS) on growth hormone (GH) synthesis and secretion by rat anterior pituitary cells in vitro. Both BCR and SS produced a dose-related reduction in GH release. Cytoplasmic GH mRNA levels were unchanged by BCR treatment and this finding was associated with an increase in total intracellular GH content. The reduction in GH release seen following SS treatment was accompanied by a fall in cytoplasmic GH mRNA levels and no significant change in intracellular GH content. These results suggest that the effects of BCR are predominantly on GH release mechanisms, whereas SS appears not only to regulate hormone release but also to regulate GH gene expression at a pre-translational level.

Animals↗
PubMed1987

Quantitation of images from a multiwire camera for autoradiography of tritium-labelled substances.

We have shown that tritium-labelled substances in two-dimensional systems can be quantitated using a multiwire camera. Its accuracy has now been improved by correcting results for non-uniformity of response over the detection area. Uniformity was assessed by imaging plates of nominally uniform activity. The results were then used to correct images from plates containing tritium-labelled proteins using a computer program. Errors were reduced from 11.3 (+/- 6.1) to 7.7 (+/- 2.8)% for standard sources and from 6.2 (+/- 1.8) to 1.9 (+/- 0.6)% for a plate containing the labelled proteins. The conducting carbon layer covering the plate absorbed 36 (+/- 3)% of the tritium beta radiation and was estimated to be 85 nm in thickness. Quantitation of the labelled proteins by the camera gave a good correlation with protein content (chi-squared: 30-40%). The activities of the protein samples were measured to an accuracy of 10% by comparison with standard sources. These results indicate useful quantitation of tritiated compounds in two-dimensional media using the multiwire camera.

Autoradiography↗
PubMed1987

Effect of hypothyroidism and thyroid hormone replacement in vivo on pituitary cytoplasmic concentrations of thyrotropin-beta and alpha-subunit messenger ribonucleic acids.

We have studied the influence of hypothyroidism and thyroid hormone replacement in vivo on rat pituitary cytoplasmic concentrations of TSH beta and alpha-subunit mRNA, measured by cytoplasmic dot hybridization, as well as circulating TSH and pituitary TSH content. Cytoplasmic concentrations of GH, PRL, and LH beta-subunit mRNA were measured in parallel. Marked increases in serum TSH and TSH beta and alpha mRNA were found in hypothyroidism; these changes reversed 72 h after beginning T3 replacement. A more marked effect of hypothyroidism on TSH beta mRNA than on alpha mRNA was evident, suggesting differential control of regulation of the TSH genes. In contrast to a rapid fall in serum TSH after T3 administration, an increase in TSH beta and alpha mRNA was evident at 1 and 6 h, followed by a later fall in TSH mRNA to euthyroid values. Pituitary cytoplasmic concentrations of GH and PRL mRNA were reduced in hypothyroidism and increased after administration of T3; LH beta mRNA was unaffected by hypothyroidism or T3 replacement. The differential effects of thyroid status on TSH, GH, PRL, and LH mRNA indicate specificity of regulation of these anterior pituitary hormone genes.

Animals↗
PubMed1987

The effect of thyroid hormones on growth hormone gene expression in vivo in rats.

Thyroid hormones are important regulators of GH synthesis and secretion. In this study we have made a detailed examination of the time-course of the effects of hypothyroidism and tri-iodothyronine (T3) replacement in the intact rat on GH gene expression in the anterior pituitary gland. Changes in pituitary cytoplasmic GH messenger (m)RNA levels were compared with total pituitary GH content and serum GH concentration during the development of hypothyroidism and following short-term T3 replacement in vivo. Hypothyroidism was associated with a fall in pituitary GH mRNA levels. Treatment of hypothyroid animals with T3 rapidly stimulated GH mRNA levels to values above those seen in euthyroid controls. The reduction in GH mRNA levels seen during the development of hypothyroidism was accompanied by a fall in serum GH and pituitary GH content, both of which were partially restored by T3 replacement. Thus thyroid hormone replacement in hypothyroidism rapidly stimulates GH mRNA synthesis, which is followed by the gradual restoration of pituitary GH stores and serum GH concentration.

Animals↗
PubMed1987

Modulation by oestrogen of thyroid hormone effects on thyrotrophin gene expression.

The role of oestrogen in the regulation of TSH gene expression is unclear. We have examined the effect of administration of oestrogen in the rat on serum TSH, pituitary TSH content and pituitary cytoplasmic concentrations of mRNA encoding the TSH beta and alpha subunits, thus deriving measures of hormone release and synthesis. In addition, we have examined the effect of oestrogen on the binding of tri-iodothyronine (T3) to nuclear receptors in the anterior pituitary. Administration of oestrogen did not affect serum concentrations of TSH in euthyroid or untreated hypothyroid rats, but did augment the effects of T3 (1 and 2 micrograms) on serum TSH in hypothyroid animals 6 h after injection of T3. No influence of oestrogen or of thyroid status on pituitary content of TSH was seen. A marked increase in the concentrations of TSH beta and alpha mRNA in pituitary cytoplasm was found in hypothyroidism, compared with those in the euthyroid state. No effect of oestrogen on TSH mRNA was seen in euthyroid animals but concentrations of TSH beta and alpha mRNA were lower in hypothyroid animals than in vehicle-treated controls. A stimulatory influence of T3 on TSH mRNA was seen 6 h after injection of T3; this stimulation was absent in oestrogen-treated rats. No effect of oestrogen on the action of T3 was evident 72 h after beginning treatment with T3. In addition to effects on serum TSH and TSH mRNA, an increase in the number of pituitary nuclear receptors for T3 was seen after oestrogen treatment.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗
PubMed1987

Thyroid status affects the regulation of prolactin mRNA accumulation by tri-iodothyronine and thyrotrophin-releasing hormone in cultured rat anterior pituitary cells.

The effects of tri-iodothyronine (T3) and TRH on prolactin mRNA accumulation in monolayer pituitary cell cultures prepared from both euthyroid and hypothyroid rats were investigated. Basal prolactin mRNA concentrations and prolactin release into culture medium were increased in hypothyroid cultures, the increase being related to the duration of hypothyroidism in vivo. The inhibitory effects of T3 seen in euthyroid cells were preserved in cells derived from hypothyroid animals, and the degree of inhibition was greater in cells from the most severely hypothyroid rats. However, the stimulation of prolactin synthesis and secretion induced by TRH in euthyroid cultures was not found in the hypothyroid cells. Hypothalamic and anterior pituitary TRH content were measured in similarly hypothyroid and euthyroid rats. A large hypothalamic pool of TRH was found, which was unchanged in hypothyroidism, whereas anterior pituitary TRH content was increased in the hypothyroid rats. The consequent down-regulation of anterior pituitary TRH receptors may explain the poor response of prolactin to TRH seen in vitro.

Animals↗
PubMed1987

Hormonal regulation of thyrotrophin synthesis and secretion.

We have examined the effects of triiodothyronine treatment on serum TSH and pituitary cytoplasmic TSH alpha and beta mRNA levels in the hypothyroid rat. Serum TSH and the cytoplasmic levels of the TSH alpha and beta subunit mRNAs rose concomitantly in hypothyroidism. Treatment of hypothyroid animals with triiodothyronine resulted in a fall in serum TSH but an initial increase in subunit mRNAs, followed by a reduction to euthyroid levels. We have also studied the modulation of these responses by oestrogen and testosterone administration. Treatment with oestradiol during the development of hypothyroidism gave rise to lower levels in alpha and beta subunit mRNAs compared with hypothyroid animals not receiving the oestrogen. Oestrogen also abolished the early rise in subunit mRNA levels seen following triiodothyronine replacement. Testosterone administration during the development of hypothyroidism had a similar effect to that of oestrogen in that subunit mRNA levels were lower in the androgen-treated hypothyroid animals than in hypothyroid animals receiving no exogenous androgen. These data suggest that gonadal steroids render the pituitary thyrotroph cell more sensitive to thyroid hormone.

Animals↗
PubMed1987

The use of a multiwire camera for the detection of 3H labelled cDNA/mRNA hybrids.

We have investigated the use of a multiwire camera for the rapid direct quantitation of 3H labelled cDNA/mRNA hybrids. Growth hormone (GH) complementary DNA (cDNA) probes were labelled with either 3H or 32P and used in dot-blot hybridization assays of pituitary cytoplasmic samples. The results were assessed either by counting in the multiwire camera (3H) or by autoradiography and scanning densitometry (32P). Results obtained by both methods were comparable over a similar range of pituitary cytoplasmic dilutions. The use of a multiwire camera may have several advantages over conventional methods for the detection of radioisotopically labelled cDNA/mRNA hybrids and has great potential for use in this field of endocrine research.

Animals↗
PubMed1986

Glycosylation of human thyroglobulin and characterization by lectin affinity electrophoresis.

Thyroglobulin, a 660 kDa glycoprotein, is the major product of protein synthesis in the thyroid gland. It has been suggested that modifications of thyroglobulin glycosylation occur in various thyroid disorders. In order to study possible changes in glycosylation of tissue thyroglobulin associated with thyroid disease, we have developed a lectin affinity electrophoresis system which allows characterization of small (less than 1 microgram) quantities of thyroglobulin. Human thyroglobulin was extracted and purified. Agarose gels were cast containing concanavalin A, Ricinus communis agglutinin, L-phytohaemagglutinin and pokeweed mitogen at various concentrations. Purified human thyroglobulin was serially diluted, loaded onto lectin gels and electrophoresed. Concanavalin A, R. communis agglutinin and phytohaemagglutinin all bound thyroglobulin in a concentration-dependent manner. Pokeweed mitogen did not bind thyroglobulin. Purified thyroglobulin was treated with neuraminidase and endoglycosidase H. Two-dimensional immunoelectrophoresis revealed the migration of thyroglobulin to be modified by neuraminidase but not by endoglycosidase H. Lectin affinity electrophoresis of purified human thyroglobulin with and without enzyme treatment indicated the presence of: oligomannose structures as shown by concanavalin A reactivity and modification by endoglycosidase H, and complex oligosaccharides as shown by affinity for R. communis agglutinin and modification by neuraminidase. These structures are in keeping with the proposed patterns of glycosylation of human thyroglobulin and indicate suitability of the method for characterizing the glycosylation of small quantities of thyroglobulin.

Acetylglucosaminidase↗
PubMed1986

Preliminary assessment of a multiwire camera for quantitative autoradiography of tritium-labelled substances.

We have assessed a multiwire camera for the high speed, quantitative autoradiography of tritium-labelled substances in two-dimensional systems. Exposure times for 3H were typically 1000 times less than film-based methods, with a detection efficiency of 19 +/- 3% for those beta particles predicted to escape into the sensitive region from uniformly 3H-labelled plastic segments of 30 micron thickness. The spatial resolving power was calculated to be 0.4 mm full width half maximum (FWHM), the background count rate was 1 CPM cm-2 and the lowest activity monitored for a biochemical study was 0.15 Bq cm-2. The camera is expected to have wide applications for detecting and quantifying biological substances in a wide range of separation media.

Autoradiography↗
PubMed1986

3,5-Diiodotyrosine and thyronine in the urine of patients with chronic renal disease.

Urinary 3,5-diiodotyrosine (DIT) and thyronine (T0) excretion was investigated in 18 patients with chronic renal disease. In accord with previous findings serum T4 and thyroid hormone binding proteins measured in 17 patients were in the low or normal range. Urinary albumin excretion was elevated in all 18 and T4 binding prealbumin (TBPA) in 15 of the 18. Urinary T0 excretion measured in 12 patients was also significantly lower than normal (mean +/- SD 4.4 +/- 2.6 vs 15.8 +/- 5.8 nmol/24 h renal vs normal 2 P less than 0.001). In contrast urinary DIT excretion was significantly elevated in renal patients compared with normal subjects (2.0 +/- 1.5 vs 0.75 +/- 0.41 nmol/24 h, respectively). Possible sources of the increased DIT are discussed.

Adult↗
PubMed1986

Serum concentrations of 3,5,3',5'-tetraiodothyroacetate (T4A) in subjects with hypo-, hyper- and euthyroidism.

Some factors influencing serum 3,5,3',5'-tetradiodothyroacetate (T4A) concentrations were examined in hypothyroid, euthyroid and hyperthyroid human subjects. Serum T4A concentration was shown to be correlated with parameters such as serum, total and free thyroxine (T4) concentrations and thyroxine: thyroxine binding globulin ratio, which are indicative of the availability of T4 for peripheral metabolism. The relationship between T4A and these parameters was not a simple linear one; serum levels of T4A tended to show less variation from the normal range than did serum total T4 in hyperthyroid subjects.

Humans↗
PubMed1986

Effect of hypothyroidism on pituitary cytoplasmic concentrations of messenger RNA encoding thyrotrophin beta and alpha subunits, prolactin and growth hormone.

Thyroid hormones may directly regulate gene expression in the anterior pituitary. In order to examine this possibility we have studied the effect of hypothyroidism in the rat on pituitary cytoplasmic concentrations of messenger RNA (mRNA) encoding thyrotrophin (TSH) beta and alpha subunits, prolactin and GH. We demonstrated a marked increase in TSH beta and alpha subunit mRNA, accompanied by a decrease in GH mRNA, in the hypothyroid state, changes largely reversed by thyroid hormone replacement. We have thus shown a direct influence of thyroid status on the pretranslational events occurring in pituitary hormone synthesis. The simultaneous rise in cytoplasmic TSH beta and alpha mRNA levels and fall in GH mRNA in hypothyroidism suggests that thyroid status exerts a differential effect on the expression of these genes.

Animals↗
PubMed1986

Demonstration of thyrotrophin beta-subunit messenger RNA in rat pituitary cells in primary culture--evidence for regulation by thyrotrophin-releasing hormone and forskolin.

We have reported previously the effect of thyroid status in vivo on pituitary cytoplasmic concentrations of messenger RNA (mRNA) encoding the thyrotrophin (TSH) beta-subunit (Franklyn, Lynam, Docherty et al, 1985). Studies in vitro of the regulation of TSH beta gene transcription have been confined to thyrotrophic tumour cells. We now report the demonstration of TSH beta-subunit mRNA in non-tumorous rat pituitary cells in primary culture. Treatment of cells with thyrotrophin-releasing hormone (TRH) and with forskolin resulted in a marked increase in cellular concentration of TSH beta-mRNA. These results suggest that TRH exerts a direct effect on the pretranslational events involved in TSH synthesis and further that the adenylate cyclase system may be involved in the regulation of synthesis. We have thus described a novel system for the study of TSH beta-subunit gene expression in normal rat pituitary cells in vitro.

Animals↗