PubMed Health⌕ Search

Biomedical subjects

D B Ramsden

Publications and source records attributed to D B Ramsden.

At least 55 records · Page 3Linked to original sources
PubMed1994

Retinoic acid inhibition of thyroxine binding to human transthyretin.

All-trans retinoic acid is a potent inhibitor of [125I]-thyroxine (T4) binding to human erythrocyte membranes and can block the activation by thyroid hormone of erythrocyte Ca(2+)-ATPase [J. Biol. Chem. (1989) 264, 687-689]. In the present studies, retinoic acid was examined for its ability to displace thyroxine from binding sites on human transthyretin (TTR). Scatchard analysis of [125I]T4 binding to purified TTR, determined by equilibrium dialysis, revealed two classes of binding sites with association constants of 3.2 x 10(9) M-1 and 8.1 x 10(6) M-1. All-trans retinoic acid also displaced [125I]T4; 40% of the specifically bound [125I]T4 was displaced at a retinoic acid concentration of 2 x 10(-5) M. Analysis of the high affinity T4 binding site suggests that the Ka for retinoic acid to that site is approx. 10(7) M-1. 8-Anilinonaphthalene-1-sulfonate (ANS), a strongly fluorescing dye, binds to the thyroxine binding sites on TTR. T4 and 3,5,3'-L-triiodothyronine (T3) shifted the fluorescence emission maximum and intensity of an ANS-TTR solution toward the spectrum obtained from uncomplexed ANS. All-trans retinoic acid caused a similar shift in the emission spectrum of ANS, but was less potent than T4. Retinol failed to quench the emission intensity of the ANS-TTR complex, while 13-cis-retinoic acid was less effective than all-trans retinoic acid.

Anilino Naphthalenesulfonates↗
PubMed1993

The structure of the human thyroxine binding globulin (TBG) gene.

A portion of the human X-chromosome (> 5 kb) encoding the translated portion of the thyroxine-binding globulin (TBG) gene was sequenced. The primary templates for sequencing were isolated from a human X-chromosome library (two positive plaques from 400,000 screened initially with a TBG cDNA probe) or were produced by PCR amplification using leucocyte genomic DNA as the amplification template. Potential hormone response elements (HREs) were identified at either end of the gene. These HREs have sequences based on the consensus half-site of thyroid hormone response elements, although it is unclear whether the structures are functional HREs. Other potential regulatory elements also were identified towards the 3' end of the gene.

Animals↗
PubMed1992

Activation of bovine plasma benzylamine oxidase (BzAO) by 1-methyl-4-(2-methylphenyl)-1,2,3,6-tetrahydropyridine.

We have shown previously that certain analogues of 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP) are potent inhibitors of human and bovine plasma benzylamine oxidase (BzAO: EC 1.4.3.6). Inhibition was competitive, reversible and allosteric. Under certain conditions competitive inhibitors of allosteric enzymes can act as allosteric activators. In the present work, 1-methyl-4-(2-methylphenyl)-1,2,3,6-tetrahydropyridine (2'-CH3MPTP) was found to activate bovine plasma BzAO at low substrate and 2'-CH3MPTP concentrations. At higher 2'-CH3MPTP concentrations, the activation was negated.

1-Methyl-4-phenyl-1,2,3,6-tetrahydropyridine↗
PubMed1992

Severity of alcoholic liver disease and markers of thyroid and steroid status.

Alcoholic liver disease is associated with abnormalities in circulating levels of thyroid, adrenal and gonadal steroid hormones. The relative importance of ethanol consumption and severity of liver disease in the aetiology of these changes and their relationship to clinical abnormalities are unclear. We studied 31 subjects with alcohol-induced liver disease divided into three groups according to the severity of histological features: fatty change, hepatitis and cirrhosis. Circulating concentrations of thyroid, adrenal and gonadal steroid hormones, together with their major binding proteins, were measured in all subjects, and changes related to histology and tests of liver function, as well as clinical endocrine status. A reduction in circulating free tri-iodothyronine (fT3) was seen in subjects with alcoholic hepatitis and cirrhosis, in association with normal or reduced levels of thyrotrophin (TSH). The absence of abnormalities in subjects with fatty change despite similar ethanol intake to the other groups, and correlations between fT3 and liver function tests, suggest that changes in fT3 reflect the severity of underlying liver disease. Similarly, marked increases in circulating cortisol in the hepatitis and cirrhosis groups, and correlations between cortisol and liver function, suggest that changes largely reflect hepatic disease. The absence of clinical features of hypothyroidism or Cushing's syndrome in these groups, despite abnormalities of fT3 and cortisol, suggest an altered tissue sensitivity to hormone effects. In contrast, increases in circulating oestradiol and reductions in testosterone were found in all three groups in males. These findings suggest that both direct effects of ethanol and hepatic dysfunction determine changes in gonadal steroids in males.

Adrenal Cortex Hormones↗
PubMed1991

An inherited non-amyloidogenic transthyretin variant, [Ser6]-TTR, with increased thyroxine-binding affinity, characterized by DNA sequencing.

Amplification and sequencing of the four transthyretin (TTR) exons of a subject with a variant TTR with four-fold increased affinity for thyroxine revealed a heterozygous G to A point mutation at base 7 of exon 2. This results in a serine for glycine change at residue 6 of the mature TTR monomer. No other mutations were found in any exon. Amplification and MspI digestion of TTR exon 2 from the leucocyte DNA of eight members of the subject's family revealed that all but one member were also heterozygous for [Ser6]-TTR.

DNA↗
PubMed1991

Familial nephropathic non-neuropathic amyloidosis: clinical features, immunohistochemistry and chemistry.

Classification of familial amyloidosis by the chemical nature of the fibrillar protein has become possible. Most such amyloidogenic proteins so far recognized are variant transthyretins, but two kindreds with the same apolipoprotein AI modification have been reported. We describe the clinical features of another such family in whom petechial skin rash appeared to be a marker for the disease, which was non-neuropathic and of the Ostertag-type. Immunohistochemistry showed the protein to be apolipoprotein AI, but allele-specific DNA amplification indicated that it was not the Arg26 variant previously identified.

Amyloidosis↗
PubMed1990

Inhibition of human benzylamine oxidase (BzAO) by analogues of 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP).

A sensitive assay for human plasma BzAO, involving the conversion of 14C-benzylamine to 14C-benzaldehyde, was developed. MPTP and several of its analogues were found to be competitive inhibitors of the enzyme. Ki values for the MPTP analogues in the presence of human plasma BzAO were determined. The analogues had a different rank order of inhibition of human plasma BzAO compared with the rank order of inhibition of bovine plasma BzAO found previously. MPTP and 1-methyl-4-(2-methylphenyl)-1,2,3,6-tetrahydropyridine (2'-CH3-MPTP), which are potent nigrostriatal toxins, were weak inhibitors of human plasma BzAO.

1-Methyl-4-phenyl-1,2,3,6-tetrahydropyridine↗
PubMed1990

Thyroxine-binding prealbumin gene polymorphism: a population study.

The human thyroxine-binding prealbumin (TBPA) gene was examined for restriction fragment length polymorphism (RFLP) in normal subjects and a subject with euthyroid hyperthyroxinaemia, due to increased thyroxine binding by TBPA, using 16 restriction enzymes. Only Taq I and Msp I were shown to detect RFLPs. In a male of the normal population and one of his daughters, an additional Taq I site was found in the 3'-flanking region of the TBPA gene. The RFLP in a subject with euthyroid hyperthyroxinaemia was due to the deletion of a MspI site. All three subjects with RFLPs were heterozygous.

Adult↗
PubMed1990

Regulation of alpha and thyrotrophin-beta subunit mRNA levels by androgens in the female rat.

Thyroid and steroid hormones act by similar mechanisms to influence gene expression in the anterior pituitary gland. The genes encoding the common alpha and TSH-beta glycoprotein subunits are known to be regulated by thyroid hormones; we report here the effects of androgen administration on levels of alpha and TSH-beta mRNA in pituitary cytoplasm in the euthyroid and hypothyroid female rat. Dihydrotestosterone (DHT) suppressed both alpha and TSH-beta mRNAs to levels lower than those found in untreated animals; a similar reduction was seen in hypothyroid animals treated with DHT. A biphasic response of TSH-beta mRNA was seen following administration of tri-iodothyronine (T3) to hypothyroid rats, with early stimulation followed by later inhibition; these changes were also evident after administration of T3 to androgen-treated animals, although mRNA levels were again suppressed. The effects of testosterone were similar to those of DHT. In contrast to the changes in mRNA levels, androgen administration did not lead to significant alterations in serum TSH concentrations or pituitary TSH content. These results indicate that, like thyroid hormones, androgens suppress both alpha and TSH-beta subunit mRNA levels in the female rat. Androgens, however, exert differential effects on TSH synthesis and release which contrast with those of thyroid hormones.

Analysis of Variance↗
PubMed1990

Theoretical considerations for improving tumour targeting.

To determine the relative importance of factors influencing tumour uptake of antibodies, we used a mathematical model to simulate intravenous injection of substances of varying molecular sizes and tumour-binding affinities at several dose levels. The FACSIMILE program was used to simulate the time course of tumour uptake of the tumour-binding substance by calculating the instantaneous tumour content (TC) and tumour:background uptake ratios (UR). Relative total doses to tumour and normal tissue were calculated by integration of TC/time curves. The model was used to make theoretical predictions on the effects of altering different parameters. The size of the injected dose in relation to the number of tumour receptors was crucial:if too low, uptake could not be improved by manipulating other variables, and if too high, the UR for large binding molecules was reduced. Using the standard scanning dose of labelled antibody, absolute numbers of labelled molecules binding to tumour could be increased by injection of a large excess of unlabelled molecules. Given an adequate dose, peak tumour content increased with increasing affinity up to receptor saturation. The peak uptake ratio rose progressively with affinity for a small ligand, but reached a relatively low plateau for antibody due to constant high background levels. At low doses such as those currently administered for diagnostic scanning with antibody, no effect of increasing affinity was predicted.

Antibodies, Monoclonal↗
PubMed1989

Thyroid hormone response elements in pituitary genes.

Thyroid hormones regulate the transcription of a number of genes in the anterior pituitary gland. A thyroid hormone response element in the regulatory region of the rat growth hormone gene has previously been shown to mediate the effects of thyroid hormones on growth hormone gene transcription. The 5' flanking regions of the thyrotrophin alpha and beta subunit and prolactin genes have now been examined for the presence of sequences similar to this response element. Southern blotting reveals that no such sequences are present in the regions of the thyrotrophin subunit and prolactin genes examined, suggesting that the thyroid response elements of these genes differ from that of the rat growth hormone gene.

Animals↗
PubMed1989

Effect of dose, molecular size, affinity, and protein binding on tumor uptake of antibody or ligand: a biomathematical model.

A mathematical model has been developed to determine the best approach to improving tumor targeting with antibody. The amount of antibody in the tumor (tumor content) and the tumor:normal tissue antibody concentration ratio (uptake ratio) were calculated over 12 days from injection, using the computer program FACSIMILE to solve the stiff nonlinear differential equations describing the system. Results indicate that success requires an optimal combination of dose, size, and binding affinity of antibody. Increasing the dose to 100 times that presently used for scanning increased both the percentage of injected antibody in the tumor and the uptake ratio by up to 2 orders of magnitude to maximal values determined by affinity. This result could be achieved by coinjecting unlabeled antibody. Increasing affinity from Keq = 10(9) to 10(13)M-1 increased the uptake ratio from 5 to 100 for whole antibody and to 550 for a small ligand, at the calculated optimal dose, but had no effect at the current scanning dose. With decreasing molecular size at average affinity, the same maximum tumor content and uptake ratio were achieved but progressively earlier. At high affinity there was a substantial advantage for a small ligand compared with whole antibody in terms of uptake ratio (550 versus 100) and tumor:normal tissue integral dose ratio (330 versus 60). The uptake of a small ligand was not increased by binding to plasma protein but with increasing time the tumor content was higher than without protein binding.

Animals↗
PubMed1989

The influence of dexamethasone on serum thyrotrophin and thyrotrophin synthesis in the rat.

Glucocorticoid hormones suppress circulating concentrations of thyrotrophin (TSH), but their effect on synthesis of TSH in the pituitary gland is unclear. We have examined the influence of the glucocorticoid dexamethasone on serum TSH, pituitary TSH content and TSH beta- and alpha-subunit mRNA concentrations in pituitary cytoplasm in both the euthyroid and hypothyroid rat, and following triiodothyronine (T3) treatment in the hypothyroid rat. The rise in serum TSH in hypothyroidism was attenuated in animals treated with dexamethasone; in addition the suppression of serum TSH 6 h after T3 administration to hypothyroid rats was enhanced by dexamethasone. In contrast to the changes in serum TSH, pituitary TSH content was unaffected by dexamethasone. Furthermore dexamethasone had no significant effect on changes in pituitary cytoplasmic TSH beta- and alpha-subunit mRNA levels with thyroid status. These findings demonstrate that dexamethasone exerts differential effects on serum TSH levels and TSH biosynthesis which contrast with those of thyroid hormones.

Animals↗
PubMed1989

Is oleic acid the thyroxine binding inhibitor in the serum of ill patients?

The possibility that oleic acid is the thyroxine binding inhibitor in the serum of seriously ill patients was investigated. 3H-Oleic acid was shown to bind directly to human thyroxine-binding globulin (TBG) by the techniques of one and two-dimensional immunoelectrophoresis in combination with autoradiography. However, no correlation was seen between serum thyroxine concentration and oleic acid concentration in two groups of patients, one of which underwent routine cholecystectomy, whilst the other group was admitted to an intensive therapy unit (mortality 75%). No correlation was seen between serum total thyroxine concentration and either stearic, palmitic, linoleic or arachidonic acid concentrations in these groups. Therefore, it was concluded that oleic acid was unlikely to be the circulating inhibitor of thyroxine binding.

Blood Proteins↗
PubMed1989

Nuclear proteins from the rat pituitary gland bind to regulatory sequences of the thyrotrophin-beta gene.

Interactions between DNA sequences and nuclear proteins are important in the regulation of gene expression. We have applied a gel retardation method to examine the binding of nuclear proteins from the rat pituitary gland to regulatory sequences of the rat TSH beta-subunit gene. Binding between nuclear protein and DNA is demonstrated by an alteration in electrophoretic mobility of the DNA. A 0.4 M NaCl fraction of pituitary nuclear proteins bound to a fragment of DNA containing the promoter region of the TSH-beta gene, but not to plasmid DNA or insulin cDNA of comparable size. This nuclear protein fraction also bound to fragments of DNA containing sequences from the 5'-flanking region of the gene; binding of nuclear proteins was evident as far as 1 kb 5' to the structural gene. Each of these regions of the TSH-beta gene formed a number of distinct complexes with the nuclear protein extract, distinguished by their differing binding affinities in the presence of poly(dI-dC), and by their electrophoretic mobility. These findings suggest that transcriptional regulation of the rat TSH-beta gene may be mediated by interaction of a number of nuclear proteins with regulatory sequences of DNA up to 1 kb upstream of the transcriptional start site of the gene.

Animals↗
PubMed1989

Subunit assembly and secretion of transthyretin: studies in a cell-free translation system and in microinjected Xenopus oocytes.

Transthyretin (TTR), or thyroid-binding prealbumin, is a protein of 55 kDa, composed of four identical subunits, which is synthesized by the liver and choroid plexus epithelium. In order to study the subunit assembly and secretion of TTR, cRNA encoding TTR was translated in a rabbit reticulocyte lysate or microinjected into Xenopus oocytes, and radiolabelled biosynthetic products were immunoprecipitated with an antibody against TTR and analysed by sodium dodecyl sulphate-polyacrylamide gel electrophoresis (SDS-PAGE) and fluorography. In the cell-free translation system in the absence of dog pancreatic microsomes, a single protein of Mr 17,000 was synthesized. In the presence of dog pancreatic microsomes, two proteins of Mr 15,000 and 37,000 were observed. The Mr 17,000 protein was identified as pre-TTR and the Mr 15,000 and 37,000 proteins as monomeric and dimeric forms of TTR. When the mRNA was microinjected into Xenopus oocytes both Mr 15,000 and 37,000 proteins were secreted into the media. It was shown that, under the SDS-PAGE conditions used in this study, the TTR tetramer dissociated to the dimeric form (Mr 37,000), but that there was no, or at least very little, further breakdown to the monomer. Therefore, to determine whether tetrameric or dimeric forms of TTR were secreted from the oocytes, the media from microinjected oocytes were subjected to gel permeation chromatography under non-dissociating conditions, and the eluted fractions analysed by SDS-PAGE. TTR eluted from the column as a dimer; there was no tetramer or monomer. The dimer, however, was completely dissociated to the monomer when analysed by SDS-PAGE, which suggested that incomplete or incorrect subunit assembly had occurred.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗
PubMed1989

Kinetic analysis of the uptake of thyroid hormones by the human erythrocyte.

The uptake of 3,5,3'-tri-iodothyronine (T3) by human erythrocytes was shown to occur by two saturable processes. System I (Michaelis constant (Km) = 109 +/- 40 (mean +/- S.D.) nmol/l; maximal velocity (Vmax) = 43 +/- 16 pmol/min per 10(9) cells) was observed over the range 1-50 nmol T3/l whilst system II (Km = 8.8 +/- 8.0 pmol/l; Vmax = 29 +/- 6 fmol/min per 10(9) cells) was observed over the range 1-500 pmol T3/l. At low concentrations (1-500 pmol/l) uptake processes equivalent to system II could be demonstrated for thyroxine and 3,3',5'-tri-iodothyronine also. System II had the properties of a carrier-mediated process.

Erythrocytes↗