bluuebunny/crossref_metadata_2025_split huggingface dataset:
Characterization of the receptor for glucagon-like peptide-1(7–36)amide on plasma membranes from rat insulinoma-derived cells by covalent cross-linking
Authors: Göke R. , Cole T. , Conlon J. M. 
Published: December 2008

ABSTRACT
125I-Labelled glucagon-like peptide-1(7–36)amide was cross-linked to a specific binding protein in plasma membranes prepared from RINm5F rat insulinoma-derived cells using disuccinimidyl suberate. Consistent with the presence of a single class of binding site on the surface of intact cells, only a single radiolabelled band at Mr 63 000 was identified by SDS-PAGE after solubilization of the ligand—binding protein complex. The band was not observed when 10 nm glucagon-like peptide-1(7–36)amide was included in the binding assay, but 1 μm concentrations of glucagon-like peptide1(1–36)amide, glucagon-like peptide-2 and glucagon did not decrease the intensity of labelling. No change in the mobility of the band was observed under reducing conditions, suggesting that the binding protein in the receptor is not attached to other subunits via disulphide bonds. In control incubations using plasma membranes from pig intestinal epithelial cells, which do not contain specific binding sites for glucagon-like peptide-1(7–36)amide, no cross-linked ligand-binding protein complex was observed.
Immunochemical analysis of the human chorionic gonadotrophin-like material secreted by 'normal' and neoplastic urothelial cells
Authors: Iles R. K. , Chard T. 
Published: December 2008

ABSTRACT
Material with the immunochemical characteristics of human chorionic gonadotrophin (hCG) is produced by bladder tumour cells in vitro and in vivo. In order to characterize this material further, media were collected from 17 cell cultures (three choriocarcinomas, seven bladder carcinomas and seven 'normal' urothelium). The hCG-like material was compared with pregnancy hCG and purified α- and β-subunits by specific radioimmunoassays. Media were also submitted to affinity chromatography and the fractions further analysed by SDS-PAGE and Western blotting. It was shown that both the neoplastic and normal urothelium produced only free β-subunit-like material. This urothelial 'β-hCG' has the same molecular weight and electrophoretic mobility as that present in the intact hCG of pregnancy.
Amplification or rearrangement of the β-human chorionic gonadotrophin (β-hCG)—human LH gene cluster is not responsible for the ectopic production of β-hCG by bladder tumour cells
Authors: Iles R. K. , Czepulkowski B. H. , Young B. D. , Chard T. 
Published: December 2008

ABSTRACT
The β-subunit of human chorionic gonadotrophin (hCG) is coded on chromosome 19 by the β-hCG-hLH gene cluster. Genomic DNA has been isolated from bladder tumour cell lines which ectopically express β-hCG. The β-hCG—hLH gene cluster was probed for possible rearrangement or amplification and cells karyotyped for chromosome 19 abnormalities. No rearrangement or amplification of the gene cluster and no consistent abnormalities of chromosome 19 were found. The expression of β-hCG by bladder tumours is therefore likely to be the result of altered gene regulation and not a rearrangement or amplification of this gene cluster.
The cytostatic effects of α-interferon may be mediated by transforming growth factor-β
Authors: Kerr D. J. , Pragnell I. B. , Sproul A. , Cowan S. , Murray T. , George D. , Leake R. 
Published: December 2008

ABSTRACT
There is some evidence to suggest that transforming growth factor-β (TGF-β) mediates the cytostatic effects of the anti-oestrogen tamoxifen. In this study we have demonstrated that α-interferon has a significant anti-proliferative effect on the oestrogen receptor-positive human breast cancer cell line ZR-75. There is decreased phenotypic expression of the oestrogen receptors (to about 30% of control values) and increased TGF-β mRNA. Under the growth conditions used here, ZR-75 cells had approximately 5800 TGF-β binding sites per cell, with an apparent dissociation constant of 70 pm, and we have shown that the anti-proliferative effects of α-interferon can be reduced by 60% by co-treating the cells with a TGF-β polyclonal antibody. The cytostatic effects of α-interferon may therefore be mediated by TGF-β in this human breast cancer cell line.
Gastric inhibitory polypeptide: a gut hormone with anabolic functions
Authors: Beck B. 
Published: December 2008

Summary
The gastrointestinal hormone, gastric inhibitory polypeptide (GIP), has been isolated and characterized because of its enterogastrone-type effects. It is also named glucose-dependent insulinotropic polypeptide and is actually considered to be the main incretin factor of the entero-insular axis. Besides these well-described effects on gastric secretion and pancreatic β cells, it also has direct metabolic effects on other tissues and organs, such as adipose tissue, liver, muscle, gastrointestinal tract and brain. In adipose tissue it is involved in the activation and regulation of lipoprotein lipase (LPL); it also inhibits glucagon-induced lipolysis and potentiates the effect of insulin on incorporation of fatty acids into triglycerides. It may play a role in the development of obesity because of the hypersensitivity of adipose tissue of obese animals to some of these actions. In the liver it does not modify insulin extraction, and its incretin effects are due only to the stimulation of insulin secretion and synthesis. It reduces hepatic glucose output and inhibits glucagon-stimulated glycogenolysis. It might increase glucose utilization in peripheral tissues such as muscle. GIP also has an effect on the volume and/or electrolyte composition of intestinal secretion and saliva. The functional importance of its effect on the hormones of the anterior pituitary lobe remains to be established, as it has never been detected in the brain.
Its links with insulin are very close and the presence of insulin is sometimes necessary for the greater efficiency of both hormones. GIP can be considered as a true metabolic hormone, with most of its functions tending to increase anabolism.
Evidence that the bovine ovary secretes large amounts of monomeric inhibin α subunit and its isolation from bovine follicular fluid
Authors: Knight P. G. , Beard A. J. , Wrathall J. H. M. , Castillo R. J. 
Published: December 2008

ABSTRACT
Analysis of bovine follicular fluid (FF) using sodium dodecyl sulphate-polyacrylamide gel electrophoresis (SDS-PAGE) coupled with a sensitive immunoblotting procedure resolved several components that were immunoreactive with an antiserum directed against the n-terminus of the α subunit of human inhibin (hIα(1–32)). Under non-reducing conditions, three intensely stained bands having apparent Mr values of 116 000, 44 000 and 25 000 were present, whilst under reducing conditions only two intensely stained bands (Mr 43 000 and 21 000) were detected. The Mr 44 000 and 25 000 immunoreactive forms (non-reducing conditions) were also demonstrated in bovine utero-ovarian vein and peripheral venous plasma after subjecting samples (40 ml) to immunoaffinity concentration using Sepharose beads coupled to anti-hIα(1–32), SDS-PAGE and immunoblotting. The same approach revealed the presence of the smaller (Mr 25 000) form in bovine granulosa cell-conditioned culture medium (GCCM). Gel-permeation chromatography (Sephacryl S-200), immunoaffinity chromatography (Sepharose-anti-hIα(1–32)) and reversed-phase high-performance liquid chromatography (RP-HPLC; C18 and C8 columns) were employed to isolate from bFF (30 ml, 195 g protein) 750 μg protein which appeared essentially homogeneous by RP-HPLC and SDS-PAGE and had an Mr of 25 000 (non-reducing conditions)/21 000 (reducing conditions), identical to that of the immunoreactive component of lowest Mr found in bovine FF, utero-ovarian vein plasma, peripheral plasma and GCCM. The isolated material was highly immunoreactive with antisera against both hIα(1–32) and purified Mr 32 000 bovine inhibin but was devoid of biological activity when tested in a rat pituitary cell inhibin bioassay. Amino-terminal analysis revealed an amino acid sequence (residues 1–14) identical to that reported elsewhere for the α subunit (Mr 20 000/21 000) of bovine inhibin.
In conclusion, the present study has revealed that the bovine ovary secretes considerable quantities of monomeric inhibin α subunit. The unexpected presence of this material in peripheral blood is likely to hinder attempts to obtain physiologically relevant data on circulating levels of inhibin in cattle using conventional radioimmunoassays.
A monoclonal antibody to human insulin-like growth factor-I: characterization, use in radioimmunoassay and effect on the biological activities of the growth factor
Authors: Morrell D. J. , Dadi H. , More J. , Taylor A. M. , Dabestani A. , Buchanan C. R. , Holder A. T. , Preece M. A 
Published: December 2008

ABSTRACT
A monoclonal antibody (BPL-M23) to insulin-like growth factor-I (IGF-I) was obtained following immunization of BALB/c mice with human IGF-I conjugated to ovalbumin. The affinity constant of BPL-M23 for IGF-I was 10·5 litres/nmol and the cross-reactivities of IGF-II, multiplication-stimulating activity III-2 and insulin were 08, 003 and less than 0·0001 % respectively. Porcine, bovine, ovine and rabbit sera, but not rat or mouse sera, showed substantial reactivity with the antibody.
Comparison of radioimmunoassay analyses of 54 human serum samples from normal subjects and acromegalic and GH-deficient patients using BPL-M23 and a polyclonal rabbit antiserum (R557A) to human IGF-I showed a high correlation, indicating the usefulness of the monoclonal antibody in radioimmunoassay.
Monoclonal antibody BPL-M23 was capable of abolishing the sulphation, mitogenic and insulin-like activities of IGF-I in in-vitro bioassays, suggesting that these activities may rely upon the same receptor-binding site which is near to the antibody-binding site.
Thyrotrophin-releasing hormone (TRH)-induced growth hormone secretion in fowl: binding of TRH to pituitary membranes
Authors: Harvey S. , Baidwan J. S. 
Published: December 2008

ABSTRACT
Heat-labile specific binding sites for [3H]3-methylhistidine2-TRH ([3H]Me-TRH) were demonstrated on chicken adenohypophysial membranes. These binding sites were of both high affinity (dissociation constant, Kd = 15·53 nm) and low capacity (maximum binding capacity, Bmax = 8·73 pmol/g tissue) and of low affinity (Kd = 104·5 nm) and high capacity (Bmax = 32·41 pmol/g). Binding of [3H]Me-TRH to the pituitary membranes was greater at 4 °C than at 39 °C and occurred with rate constants (k1) of 1·6× 106 and 3·39×106m-1 min-1 respectively. Dissociation of [3H]Me-TRH binding at 4 °C occurred with a rate constant (k-1) of 0·125 min-1. Binding sites for [3H]Me-TRH were found in the cephalic lobe of the pituitary gland (the location of most lactotrophs and thyrotrophs) and in the caudal lobe (the location of most somatotrophs). The number of binding sites was greater in the caudal lobe than in the cephalic lobe, although the affinity of [3H]Me-TRH binding did not differ. The binding of [3H]Me-TRH to caudal lobe membranes was displaced by Me-TRH, TRH, pGlu-His-Pro-Gly-NH2 and [Glu1]-TRH, with half-maximal effective doses of 33 nm, 70·7 nm, 1·23 μm and 22 μm respectively, but not by [Phe2]-TRH, TRH free acid or His-Pro-diketopiperazine. The number of caudal lobe binding sites for [3H]Me-TRH in old birds was less than that in young ones, and the number of binding sites was increased in birds deprived of food for 48 h. TRH-induced GH secretion in birds would thus appear to be mediated by specific receptors on caudal lobe somatotrophs, and these results suggest that physiological changes in GH secretion (during growth and periods of fasting) are causally related to the abundance of TRH-binding sites.
Differential binding of thyroid hormone receptors to mouse glandular kallikrein gene promoters: evidence for multiple binding regions in the mGK-6 gene
Authors: Barlow J. W. , Raggatt L. E. , Drinkwater C. C. , Lyons I. G. , Richards R. I. 
Published: December 2008

ABSTRACT
We have used a DNA-cellulose competition assay to investigate the binding of thyroid hormone receptors to fragments of the mouse glandular kallikrein genes and the human and rat GH genes. Nuclear extracts from human lymphoblastoid IM-9 cells were incubated with [125I]tri-iodothyronine ([125I]T3) and DNA-cellulose. The ability of cloned gene fragments to compete for radiolabelled receptors bound to DNA-cellulose was compared with that of DNA from pBR322. As previously observed, a 900 bp fragment from the human GH gene showed preferential binding to the thyroid hormone receptor. High-affinity binding was observed with a synthetic fragment of the rat GH gene encompassing positions − 163 to − 192 but not with a similar fragment from positions −224 to −192. Preferential binding was also observed with fragments of the mouse glandular kallikrein gene, mGK-6. Binding to the entire gene and fragments containing 2300 and 776 bp of the promoter region was identical. Detectable but reduced binding was seen with a shorter fragment. These results suggest that the T3 receptor binds to multiple sites within the first 776 bp of the mGK-6 gene promoter. Potential thyroid hormone response elements can be identified within this region of the gene. In contrast, the kallikrein gene mGK-3, which shows a different response to thyroid hormone from that of mGK-6, showed no significant binding in the comparable promoter region.
Treatment of ovariectomized heifers with bovine follicular fluid specifically suppresses pituitary levels of FSH-β mRNA
Authors: Beard A. J. , Savva D. , Glencross R. G. , McLeod B. J. , Knight P. G. 
Published: December 2008

ABSTRACT
To investigate the inhibin-induced suppression of FSH secretion by the anterior pituitary, chronically ovariectomized heifers (three per group) were treated for 56–58 h with either steroid-free bovine follicular fluid (bFF; 8 ml i.v. every 8 h) or 0·9% (w/v) NaCl (8 ml i.v. every 8 h). Blood was withdrawn at 8-h intervals for analysis of plasma concentrations of FSH and LH by radioimmunoassay. At the end of the treatment period, heifers were slaughtered and pituitary glands recovered for determination of gonadotrophin contents and levels of mRNA encoding FSH-β, LH-β, TSH-β and common α glycoprotein hormone subunits using [32P]cDNA probes in total RNA dot and Northern blot assays. Treatment with bFF markedly suppressed plasma FSH by 85% (P<0·001 compared with pretreatment period), but did not affect plasma LH concentrations. Plasma FSH and LH concentrations did not vary significantly in the saline-injected control heifers. The level of FSH-β subunit mRNA was reduced by 60% (P<0·001) in heifers treated with bFF, whereas no significant differences between control and bFF-treated heifers were observed in the levels of mRNA encoding LH-β, TSH-β or common α subunits. Treatments with bFF, however, did not affect pituitary content of either FSH or LH.
These results support the conclusion that inhibin exerts its selective suppressive effect on the secretion of FSH by the bovine pituitary, at least in part, by directly inhibiting expression of the gene encoding the FSH-β subunit.
Lactate alters plasma membrane potential, increases the concentration of cytosolic Ca2+ and stimulates the secretion of insulin by the hamster β-cell line HIT-T15
Authors: Meats J. E. , Tuersley M. D. , Best L. , Lynch A. M. , Tomlinson S. 
Published: December 2008

ABSTRACT
We have studied the effect of lactate on a number of intracellular events which may be important in controlling the secretion of insulin by the hamster β-cell line HIT-T15. Using the fluorescent dye Oxonol V, as well as intracellular recording techniques to measure changes in membrane potential, we found that lactate, glucose, K+ and tolbutamide caused depolarization of HIT cells, while valinomycin resulted in hyperpolarization. Consistent with these findings was the observation that 10 mm lactate caused an increase of 69·0±18·4% (s.e.m., n=6) in the level of free cytosolic Ca2+ within HIT cells (assessed by fluorescence of quin 2). This was probably due to influx of Ca2+ through voltage-sensitive calcium channels, since it was dependent upon the concentration of extracellular Ca2+ and inhibited by verapamil. Lactate also caused cytosolic acidification in HIT cells and increased the secretion of insulin. These findings are consistent with the view that the electrogenic efflux of lactate could be a determinant in the activation of HIT cells by lactate and possibly by glucose.
C4A gene deletion: association with Graves' disease
Authors: Ratanachaiyavong S. , Lloyd L. , McGregor A. M. 
Published: December 2008

ABSTRACT
The association of HLA class I and class II antigens, particularly HLA-B8,DR3, with a variety of autoimmune diseases has been well documented. The C4A*Q0 (non-expressed C4A) allele which is in linkage disequilibrium with HLA-B8,DR3 has also been reported to be associated with systemic lupus erythematosus, insulin-dependent diabetes mellitus and Graves' disease. However, the number of studies has been limited by the requirement of family data for the assignment of the C4A*Q0 allele based on C4 protein typing. Recently, with the availability of a C4 cDNA probe, a C4A gene deletion associated with HLA-B8,DR3 has been reported in normal individuals. We have tried to resolve the problem of assigning the C4A*Q0 allele by using both phenotypic and genotypic approaches and have determined the significance of the C4A*Q0 allele in 80 unrelated patients with Graves' disease and in 50 normal control subjects. Our results demonstrate a strong association of the C4A*Q0 allele with Graves' disease (56 versus 26%; P< 0·002, relative risk=3·7) and in particular in association with HLA-B8 and/or DR3 (92 versus 70·6%; P<0·04) when compared with normal controls. All the C4A*Q0 alleles that were associated with HLA-B8 and/or DR3 were due to a C4A gene deletion. Of the C4A*Q0 alleles in Graves' disease, 94% (compared with 82% in the control group) could be detected by C4 DNA analysis using either TaqI or EcoRI restriction endonucleases. It is suggested that a combination of C4 protein typing with C4 DNA analysis is the best approach for the determination of the C4A*Q0 allele in unrelated individuals without access to family data.
Structure of rhesus monkey relaxin predicted by analysis of the single-copy rhesus monkey relaxin gene
Authors: Crawford R. J. , Hammond V. E. , Roche P. J. , Johnston P. D. , Tregear G. W. 
Published: December 2008

ABSTRACT
The gene encoding rhesus monkey relaxin has been investigated. A cDNA library was prepared using corpus luteal RNA from a pregnant rhesus monkey, cDNA clones encoding relaxin were isolated and the nucleotide sequence was determined. The amino acid sequence of rhesus monkey preprorelaxin, predicted from the cDNA, demonstrates that the sequence has not been strongly conserved when compared with that of man, although features characteristic of the relaxin molecule have been maintained. This structural information will allow production of rhesus monkey relaxin, leading to studies investigating the bioactivity of relaxin in a homologous primate system. Southern blot analysis indicated that there is only one relaxin gene in the rhesus monkey and baboon genomes. In this respect these primate genomes are different from the human genome which contains two relaxin genes.
Subunit assembly and secretion of transthyretin: studies in a cell-free translation system and in microinjected Xenopus oocytes
Authors: Docherty K. , Shennan K. I. J. , Marsden R. F. , Ramsden D. B. 
Published: December 2008

ABSTRACT
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 ooctyes 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. These results lead to the conclusion that liver- and choroid plexus-specific factors may be required for correct subunit assembly of TTR.
Desensitization of the β-adrenergic receptor of the S49 murine lymphoma cell: mechanism of receptor loss and recovery
Authors: Rich K. A. , Iyengar R. 
Published: December 2008

ABSTRACT
The effect of prolonged exposure of S49 murine lymphoma cells to (−) isoprenaline on the levels of β-adrenergic receptors and the stimulatory regulatory component of adenylyl cyclase (Gs) was investigated. Exposure of wild-type S49 cells (WT cells) to isoprenaline for 16 h resulted in an 84% decrease in β-adrenergic receptors and a 78% decrease in catecholamine-stimulated adenylyl cyclase activity. The desensitization of adenylyl cyclase was homologous since no change in stimulation by any other effectors was observed. Similar treatment of cyc- S49 cells, a mutant that lacks Gs, resulted in only a 10–15% receptor loss despite a 50% decrease in catecholamine stimulation of reconstituted adenylyl cyclase activity. By quantitative extraction of Gs from control and isoprenaline-treated WT cellmembranes and reconstitution into untreated cyc- cell membranes, it was demonstrated that the levels and behaviour of Gs in WT cell membranes were unchanged after isoprenaline treatment, despite a substantial loss of receptors from the cell surface. The β-adrenergic receptors lost from the cell surface could not be identified in any intracellular membrane fraction. Addition of cycloheximide to the culture medium of WT cells previously treated with isoprenaline resulted in a complete blockade of the restoration of β-adrenergic receptors which was otherwise observed within 16 h after removal of the agonist.
It is concluded that, in the S49 cell line, prolonged exposure to agonists results in a loss of β-adrenergic receptors by a process dependent on the presence of Gs. However, the loss of receptors is selective since under conditions where the majority of receptors are lost, there is no measurable change in the level of cell surface membrane-bound Gs. The lost receptors appear to be degraded, since the restoration of receptor levels after removal of the agonist is dependent on protein synthesis.