bluuebunny/crossref_metadata_2025_split huggingface dataset:
Expression of the genes for α inhibin, βA inhibin and follistatin in the ovaries of Booroola ewes which were homozygotes or non-carriers of the fecundity gene FecB
Authors: Fleming J. S. , Tisdall D. J. , Greenwood P. J. , Hudson N. L. , Heath D. A. , McNatty K. P. 
Published: December 2008

ABSTRACT
Ovine cDNA probes for the α and βA inhibin subunits and for follistatin were used to investigate the mRNA species for these hormones in ovaries obtained during the luteal phase of the oestrous cycle, from Booroola ewes which were homozygous carriers (BB) or non-carriers (++) of the FecB gene. BB ewes had significantly higher concentrations of peripheral FSH and LH immunoreactivity than ++ ewes, but the peripheral inhibin immunoreactivity and ovarian inhibin and progesterone secretion rates were not significantly different between genotypes. No gene-specific differences in the number or size of mRNA transcripts detected by Northern blotting were noted for any of these genes. A single α inhibin mRNA species at 1.5 kb was observed in the follicle RNA from ++ and BB ovaries. Low amounts of α inhibin hybridization were discerned occasionally in + + and BB stroma and also in BB, but not in ++, corpora lutea. The βA inhibin gene was expressed only in the follicles from both ++ and BB ovaries. At least three βA inhibin transcripts were observed; one at 7.5kb and at least two between 1.4 and 5.0kb. The follistatin cDNA probe detected two major transcripts at 2.7 and 1.5 kb and a minor band at 0.5 kb in both follicle and corpora lutea RNA. Densitometry of the Northern blots revealed no significant gene-specific differences in the levels of α inhibin and follistatin gene mRNA transcripts. However, significantly greater amounts of total βA inhibin hybridization were detected in follicle RNA from BB compared with ++ ovaries (P<0.001) and this FecB-specific difference appeared to be associated with the 7.5 kb transcript. We conclude that the Booroola FecB gene does not influence the synthesis of the α inhibin subunit or follistatin during the luteal phase of the oestrous cycle, but may affect inhibin or activin synthesis in the ovaries of FecB carriers, by increasing the transcription or stability of the βA inhibin mRNA species.
Expression of the clusterin gene in the tissues of Booroola sheep which were homozygotes or non-carriers of the fecundity gene FecB
Authors: Fleming J. S. , Greenwood P. J. , Chen C.-L. C. 
Published: December 2008

ABSTRACT
Clusterin or sulphated glycoprotein-2 is a major component of the rete testis fluid, synthesized by the rete testis epithelial cells and Sertoli cells. Differences in the two-dimensional polyacrylamide gel electrophoresis pattern of clusterin-like proteins have been reported in rete testis fluid from Booroola rams carrying the fecundity gene FecB, when compared with that from non-carrier rams. In order to determine whether the FecB gene influences the expression of the clusterin gene, we used a rat clusterin cRNA probe to investigate mRNA species in the tissues of homozygous (BB) or non-carrier (+ +) Booroola sheep. Northern blots of polyadenylated RNA showed hybridization to the cRNA probe in the testis, ovarian follicles, corpora lutea and stroma, pituitary and liver. A major mRNA transcript was observed at 2·3 kb and a minor transcript in some tissues at 0·8 kb. Densitometry of the autoradiographs revealed no FecB-specific differences in the densities of the hybridization signals from + + and BB testis or ovarian follicle, corpora lutea or stromal RNA. We conclude that the gene for ovine clusterin is expressed widely in the tissues of sheep and that its expression is not affected by the presence of the FecB gene.
Effects of dexamethasone on G protein levels and adenylyl cyclase activity in rat vascular smooth muscle cells
Authors: McLellan A. R. , Tawil S. , Lyall F. , Milligan G. , Connell J. M. C. , Kenyon C. J. 
Published: December 2008

ABSTRACT
Dexamethasone administration in vitro has been shown to increase adenylyl cyclase activity in vascular smooth muscle cells (VSMC) from renal arteries and in non-vascular cell lines. To investigate whether G proteins are involved in this response, cultured VSMC from mesenteric arteries of Sprague—Dawley rats were incubated in the presence and absence of 10 nm dexamethasone for 24 and 48 h. Basal and stimulated adenylyl cyclase activities were increased by approximately 50% after treatment with dexamethasone. The changes were neither specifically associated with ligands which stimulate adenylyl cyclase catalytic unit via Gs (isoproterenol and prostaglandin E1) nor with guanylylimidodiphosphate (0·1 nm), which inhibits the catalytic unit via Gi. This suggests that dexamethasone enhances adenylyl cyclase activity through changes at the level of the catalytic unit, rather than through the G proteins which modulate its activity. No differences were seen in immunoblotting studies of the levels of Giα2, Gsα, Giα3 and β subunits. Similarly, dexamethasone had no effect on the expression of mRNA for Giα2 and Gsα.
The results indicate that glucocorticoid-induced increases of adenylyl cyclase activity are due to changes at the level of the adenylyl cyclase catalytic unit rather than alteration of the levels or turnover of Gsα, Giα2, Giα3 and β subunits in the membranes of VSMC.
An association between a BclI restriction fragment length polymorphism of the glucocorticoid receptor locus and hyperinsulinaemia in obese women
Authors: Weaver J. U. , Hitman G. A. , Kopelman P. G. 
Published: December 2008

ABSTRACT
Obesity is likely to be a multifactorial disease with an important genetic component. Animal models of genetic and experimentally induced obesity suggest that glucocorticoid receptor (GR) activity plays a role in the aetiology and maintenance of the obese state. Glucocorticoid activity appears to be essential for the development of hyperinsulinaemia and subsequent fat deposition. In humans, glucocorticoid excess is associated with central fat distribution. We have therefore investigated the restriction fragment length polymorphisms of the human GR gene locus (GRL) and have sought associations of specific alleles with anthropometric measurements and indices of insulin secretion and resistance in obesity.
Fifty-six extremely obese, unrelated, nondiabetic premenopausal British Caucasian females and 43 age-matched, normal weight controls were studied. The obese subjects were characterized by fat distribution (waist to hip ratio), insulin secretion and insulin resistance (fasting insulin (FI)), an index of insulin resistance (HOMA), stimulated insulin secretion during an oral glucose tolerance test and insulin-mediated glucose disposal, steady-state plasma glucose). A BclI polymorphism (fragments of 4·5 and 2·3 kb) demonstrated significant association with indices of glucose metabolism in obesity; those subjects homozygous for the 4·5 kb fragment had elevated FI (Pc=0·012) and HOMA (Pc=0·012) values. The genotypic and allelic frequencies of the GRL BclI polymorphism were otherwise similar in obese and normal weight subjects. We postulate that the GRL BclI polymorphism may directly affect GR gene expression, or be in linkage disequilibrium with a possible mutation within one of three exons of the GR gene, and thereby modulate GR transcriptional activity on target genes involved in glucose and insulin homeostasis.
Identification of exchange mechanisms for the regulation of intracellular pH in rat thyroid FRTL-5 cells
Authors: Wood A. M. , Warhurst G. , Bidey S. P. , Soden J. , Taylor R. , Robertson W. R. 
Published: December 2008

ABSTRACT
pH is maintained in cells by plasma membrane exchange mechanisms. In the absence of HCO3− ions, FRTL-5 cells regulate intracellular pH (pHi) by an Na+/H+ antiport but HCO3−-dependent exchangers cannot operate. We have investigated pHi regulation (by microfluorimetry and the pH sensitive dye 2′,7′-bis(2-carboxyethyl)-5(6′)-carboxy-fluorescein) in small groups (five to six cells) of FRTL-5 thyroid cell monolayers held in kREBS—Ringer buffer (pH 7·4) with or without HCO3− ions. The exchangers were investigated with inhibitors (amiloride or its derivative dimethylamiloride for the Na+/H+ antiporter and the stilbene derivative disodium 4,4′-diisothiocyanatostilbene-2,2′-disulphonic acid (DIDS) for HCO3 −-dependent mechanisms), ionic substitution and by NH4+/NH3 (10mm) acid loading. Basal pHi was lower in the presence (7·3±0·058, mean±s.d., n= 14) than in the absence (7·59±0·078, n=10) of HCO3 ions. In HCO3 −-free media, cells recovered from acid load by 0·34±0·04 pH units in the first 2 min and finally reached a pHi of 7·35±0·06. This recovery was Na+-dependent and blocked by dimethylamiloride during the 15 min following intracellular acidification. In HCO3−-containing media, cells recovered from an acid load at a similar rate, but reached 99 ± 10% (n = 9) of the baseline pH; this recovery was also dependent on Na+ ions. Moreover, although dimethylamiloride and DIDS reduced the rate of recovery to 0·06±0·02 and 0·18±0·04 pH units respectively during the 2-min period, the cells returned to the basal pHi within 15 min. Removal of Na+ from HCO3−-containing media acidified the cells (ΔpH=–0·82±0·05, n=10) within 40 min; this acidification was partially blocked by either amiloride or DIDS. Removal of Cl− alkalinized the cells (ΔpH=+0·51 ± 0·06, n=10) after 40 min, and this alkalinization was totally prevented by DIDS. Furthermore, in the absence of Na+ and presence of amiloride, alkalinization was still seen on the removal of Cl−, albeit at a diminished rate (i.e. ΔpH = +0·25±0·05, n=8) after 40 min. In conclusion, FRTL-5 cells maintain pHi by two Na+-dependent exchangers, one sensitive to amiloride, the other to DIDS, and a Na+-independent, Cl−/HCO3− mechanism.
The mouse relaxin gene: nucleotide sequence and expression
Authors: Evans B A , John M , Fowler K J , Summers R J , Cronk M , Shine J , Tregear G W 
Published: December 2008

ABSTRACT
Relaxin is a polypeptide hormone that has a variety of physiological effects both on remodelling of collagen and on uterine contractility. These are most apparent during pregnancy. The sequences of relaxin cDNAs derived from ovaries of late-pregnant random-bred Swiss mice have been established. Multiple subclones obtained from three independent polymerase chain reaction experiments were found to encode relaxins which were identical except at position 11 in the A chain (Ile or Val). All mouse relaxin cDNAs expressed in the ovary during pregnancy had an extra tyrosine inserted prior to the final A chain cysteine residue, a result confirmed by direct sequencing of relaxin peptides. Whilst this tyrosine insertion must have local effects on the folding of the A chain, structure—activity studies will clarify whether it perturbs functional interaction with the relaxin receptor. We have shown that there is a single relaxin gene in the mouse genome, and that expression during pregnancy occurs in the ovary but is not detectable in the placenta, uterus or fetus.
Glucose-regulated protein-78 expression in the rat adrenal cortex
Authors: Gregoire J , Mertz L M , Pedersen R C 
Published: December 2008

ABSTRACT
We have postulated that steroidogenesis activator polypeptide (SAP) is a product of glucose-regulated protein-78 (grp78) proteolysis on the basis of a number of considerations, including a striking sequence similarity between the carboxyl-terminal region of grp78 and SAP. Since ACTH stimulates the rapid intracellular accumulation of SAP, experiments were conducted to determine whether ACTH might also regulate levels of grp78 mRNA and/or protein. Using a grp78 cDNA probe, Northern analysis of total RNA isolated from hypophysectomized or dexamethasone-suppressed rats revealed that neither treatment had a measurable influence on steady-state levels of grp78 mRNA over a 4-day period. Moreover, immunoblotting with an antiserum directed against a shared grp78/SAP sequence failed to detect a significant change in the grp78 content of adrenal homogenates from dexamethasone-suppressed rats as compared with untreated controls. On the other hand, grp78 in cultured rat adrenocortical cells fell to 50% of that in time-zero controls after 72 h in the absence of ACTH, whereas inclusion of ACTH in the medium blocked this decline. We conclude that while adrenocortical grp78 may be under some measure of trophic ACTH control, the rapid fluctuations reported for SAP are not likely to be driven by large changes in the size of the grp78 pool.
Localization of progesterone-associated endometrial protein mRNA by in-situ hybridization in human pregnancy decidua, endometriosis and borderline endometrioid adenoma
Authors: Kämäräinen M , Leivo I , Julkunen M , Seppälä M 
Published: December 2008

ABSTRACT
Progesterone-associated endometrial protein (PAEP) has been isolated from human decidualized endometrium. In-situ hybridization histochemistry was employed to determine the cellular localization of PAEP mRNA in decidua during pregnancy. PAEP mRNA was found to be expressed in the glandular epithelium of decidua spongiosa throughout pregnancy. Substantial variations in the amount of PAEP mRNA during the course of pregnancy were observed, and it was most abundant at the end of the first trimester. We also found that the PAEP gene was expressed in endometriosis and in a borderline endometrioid adenoma. As in decidual tissues, PAEP mRNA in endometriosis was abundant in the glandular compartment.
Specific oxytocin agonist stimulates prolactin release but has no effect on inositol phosphate accumulation in isolated rat anterior pituitary cells
Authors: Chadio S E , Antoni F A 
Published: December 2008

ABSTRACT
We have previously characterized specific oxytocin receptors in the rat anterior pituitary gland, using a highly selective oxytocin receptor antagonist as radio-ligand. The aim of the present study was to examine whether occupation of these receptors by oxytocin produces a stimulation of prolactin release and a rise in the accumulation of total inositol phosphates in the rat adenohypophysis. Anterior pituitary cells harvested from randomly cycling and diethylstilboestrol (100 μg s.c.)-treated rats were perifused with Dulbecco's minimal essential medium at a rate of 0·3 ml/min. Oxytocin and the specific oxytocin agonist [Thr4-Gly7]-oxytocin (TG-OT) both stimulated a significant prolactin release at concentrations of 10-6 and 10-7 m. Oestrogen treatment did not affect the response to oxytocin, indicating that there is no straightforward correlation between receptor number and prolactin secretory response in the rat pituitary gland.
The involvement of phosphoinositide hydrolysis was investigated in dispersed anterior pituitary cells and uterine tissue from randomly cycling rats. Oxytocin and arginine-vasopressin stimulated a significant (P<0·05) and dose-related increase in total inositol phosphates, vasopressin being more potent. The specific oxytocin agonist TG-OT had no effect on total inositol phosphate production in pituitary cells, but when tested in uterine tissue it significantly (P< 0.05) stimulated the accumulation of total inositol phosphate at all concentrations tested (10-5 to 10-9 m). In conclusion, the data show that oxytocin has prolactin-releasing activity, acting on specific receptors in the anterior pituitary gland. Furthermore, although oxytocin receptors in the rat uterus are coupled to the inositol phospholipid cycle, it would appear that this is not a prerequisite for the stimulation of prolactin secretion when specific oxytocin receptors in the rat adenohypophysis are activated.
Up-regulation of insulin-like growth factor-I (IGF-I) receptor gene expression in patients with reduced serum IGF-I levels
Authors: Eshet R , Werner H , Klinger B , Silbergeld A , Laron Z , LeRoith D , Roberts C T 
Published: December 2008

ABSTRACT
We have analysed the expression of the IGF-I receptor gene in lymphocytes of patients with low levels of circulating IGF-I (four patients with isolated GH deficiency (IGHD) and one Laron-type dwarf (LTD)) in comparison with a control group exhibiting normal serum IGF-I levels and endocrine profiles. 125I-Labelled IGF-I binding assays were performed on erythrocytes to determine the number of IGF-I binding sites per cell and their dissociation constants. Erythrocytes from patients with IGHD or LTD contained significantly (P=0·002) more receptors per cell (10·9±3·1 binding sites/cell), with a reduced affinity (Kd = 0·49±0·05 nm), than erythrocytes from controls (2·0±0·4 sites/cell; Kd = 0·14 nm). The levels of IGF-I receptor mRNA in circulating lymphocytes were determined by an RNA template-specific reverse transcription/polymerase chain reaction method. There was a statistically significant increase in IGF-I receptor mRNA levels in lymphocytes from patients with LTD or IGHD when compared with controls (3108·1±775·9 vs 576·0±465·7 arbitrary units, P=0·006). The increased level of IGF-I binding due to increased IGF-I receptor gene expression may represent a compensatory up-regulation process activated in response to the low levels of IGF-I in the circulation of patients with LTD or IGHD.
The human insulin gene-linked polymorphic region adopts a G-quartet structure in chromatin assembled in vitro
Authors: Hammond-Kosack M C U , Kilpatrick M W , Docherty K 
Published: December 2008

ABSTRACT
The insulin gene-linked polymorphic region (ILPR), located 363 bp upstream of the human insulin gene, is composed of tandem repeats of the consensus sequence ACAGGGGT(G/C)(T/C)GGGG. It has previously been shown that an insulin gene fragment containing the ILPR adopts an altered DNA structure in vitro. Furthermore, oligonucleotides containing the consensus repeat sequence exhibit multiple quadriplex DNA structures. The present study was undertaken to determine whether such altered DNA structures existed within the ILPR when the insulin gene was assembled into chromatin in vitro. Chromatin assembly was achieved using histones and an extract from unfertilized eggs from Xenopus laevis. The presence of altered DNA conformations within the 5′ region of the human insulin gene was investigated using the structural probe nuclease P1. Nuclease P1 recognized multiple distinct sites in the 5′ flanking region of the human insulin gene in naked DNA. Most of these sites disappeared when the recombinant plasmid DNA was treated with histones and unfertilized egg extract. In the assembled DNA, the ILPR appeared as the major site of nuclease P1 hypersensitivity. Fine-mapping of the multiple reactive sites within the ILPR showed a pattern characteristic of G-quartet foldback structures similar to those that have been observed for telomeric DNA.
Changes in progesterone and oestrogen receptor mRNA and protein and oxytocin receptors in endometrium of ewes after intrauterine injection of ovine trophoblast interferon
Authors: Mirando M A , Harney J P , Zhou Y , Ogle T F , Ott T L , Moffatt R J , Bazer F W 
Published: December 2008

ABSTRACT
This study determined whether twice-daily intrauterine injections of ovine conceptus secretory proteins (oCSP) containing type-I trophoblast interferon (25 μg/uterine horn) from day 11 to day 15 post-oestrus (oestrus = day 0) could alter the binding capacities of endometrial receptors for oxytocin, progesterone and oestrogen in cyclic ewes when compared with control ewes receiving serum protein (SP) injections. Injections of oCSP on days 11–15 post-oestrus decreased concentrations of oestrogen receptors (P<0·06), oestrogen receptor mRNA (P<0·05) and progesterone receptors (P<0·08) in endometrium on day 16 when compared with SP-infused control ewes, which were undergoing corpus luteum regression on days 14–16. Injection of oCSP also decreased the number (P<0·10) and affinity (P<0·06) of oxytocin receptors. Inositol phosphate formation induced in the endometrium on day 16 by 100 nm oxytocin in vitro was highly correlated with the concentration (r≥0·93, P<0·001) and Kd (r= –0·91, P<0·01) of oxytocin receptors in SP-infused ewes, but was not as highly correlated with concentration (r≤0·83, P<0·06) and Kd (r≤ –0·40, P>0·40) of oxytocin receptors in oCSP-infused ewes. This indicates that oCSP disrupted the relationship between oxytocin receptor binding and oxytocin-induced activation of its second messenger system. These results indicate that antiluteolytic type-I trophoblast interferon may prevent oxytocininduced luteolytic pulsatile secretion of prostaglandin F2α during maternal recognition of pregnancy in sheep, by reducing the synthesis and affinity of endometrial oxytocin receptors. Inhibition of other components of the oxytocin receptor—phospholipase C system by ovine trophoblast interferon may also be involved in reducing endometrial responsiveness to oxytocin. Ovine trophoblast interferon may inhibit the synthesis of endometrial oestrogen receptors to inhibit responsiveness to oxytocin during early pregnancy in ewes.
Expression of the β3-adrenergic receptor in human white adipose tissue
Authors: Revelli J-P , Muzzin P , Paoloni A , Moinat M , Giacobino J-P 
Published: December 2008

ABSTRACT
The results of this study suggest that the atypical β-adrenergic receptor (β3 subtype) is expressed in human white adipose tissue. A β3-adrenergic receptor mRNA could be detected in human omental fat poly(A)+ RNA by polymerase chain reaction amplification with appropriate primers. The expression of the β3-adrenergic receptor in human fat was confirmed by Northern blot analysis; however, the level of its mRNA was lower than those of the β1- and β2-adrenergic receptors. Two populations of ((−)-4-(3-t-butylamino-2-hydroxypropoxy)-[5,7-3H]benz-imidazole-2-one) ([3H]CGP 12177)-binding sites were identified in human omental fat membranes, one with a high affinity (Kd= 1·7 nm) and the other with a low affinity (Kd = 22 nm). The low affinity binding site population, which should represent the β3-adrenergic receptor, was predominant (75% of the total binding sites).
Effects of free fatty acids on somatostatin secretion, content and mRNA levels in cortical and hypothalamic fetal rat neurones in monolayer culture
Authors: Señarís R M , Lewis M D , Lago F , Domínguez F , Scanlon M F , Diéguez C 
Published: December 2008

ABSTRACT
The effects of free fatty acids on somatostatin secretion, content and mRNA levels in fetal rat hypothalamic and cortical cell cultures were investigated. Somatostatin secretion and content were quantified by radioimmunoassay. Somatostatin mRNA levels were measured by Northern blot hybridization using a cRNA probe. Treatment with either caprylic acid (5×10-3m) or oleic acid (5× 10-5 m) for 90 min inhibited basal somatostatin secretion in both hypothalamic and cortical cell cultures. In addition, the increase in somatostatin secretion induced by incubation with veratridine (10-4m) or carbachol (10-4m) for 90 min was significantly reduced by the addition of caprylic acid, but somatostatin release stimulated by 5·6×10-2m KCl was not affected. Furthermore, treatment with these free fatty acids for 90 min markedly decreased somatostatin mRNA levels in both types of neurone culture. These inhibitory effects were transient, being observed after 90 min, but not after 5 h. These results support the probability that there is a role for free fatty acids in the regulation of somatostatin mRNA levels and somatostatin secretion in both hypothalamic and cortical cell cultures.