bluuebunny/crossref_metadata_2025_split huggingface dataset:
Characterization of the ovine LHβ-subunit gene: the promoter is regulated by GnRH and gonadal steroids in transgenic mice
Authors: McNeilly J R , Brown P , Mullins J , Clark A J , McNeilly A S 
Published: December 2004

Abstract
We have previously demonstrated that 1·9 kb of ovine LHβ promoter fused to bacterial chloramphenicol transferase (CAT) coding sequence is sufficient to target expression of the transgene specifically to the gonadotroph cells of the anterior pituitary in mice with no expression being observed in other tissues. However, it is not known if this region of the ovine LHβ promoter contains the necessary elements that confer transcriptional regulation by gonadal steroids and GnRH. Following gonadectomy, both endogenous pituitary LH and CAT activity significantly (P>0·001) increased as did plasma LH. This postgonadectomy increase in CAT, pituitary and plasma LH could be suppressed in females by treatment with oestradiol alone or oestradiol and progesterone, with an additional significant (P<0·05) reduction in CAT activity being observed in one line following the combined steroid treatment. In castrated males, testosterone suppressed CAT activity in one line. Treatment of transgenic ovariectomized females with oestradiol alone significantly suppressed plasma LH (P<0·01) with no change in pituitary LH content. There was no difference in pituitary LH between oestradiol-treated ovariectomized transgenic and non-transgenic females. Treatment of intact females from both lines with either GnRH antiserum or agonist demonstrated a decrease in pituitary CAT activity whereas similar treatment in intact males had no effect. While endogenous pituitary LH concentrations were variable, plasma LH was lower in all treated animals irrespective of line, sex or expression of the transgene. In conclusion, these results indicate that (1) the presence of CAT protein is not toxic and does not compromise either endogenous LH synthesis, storage and secretion and (2) the ovine LHβ–CAT gene is regulated in a similar but more variable manner to the endogenous LHβ gene. This may relate to the use of CAT as a reporter where its release is not necessarily related to that of the endogenous hormone whose synthesis, storage and release may differ.
Journal of Endocrinology (1996) 151, 481–489
Effects of adrenal steroids on the concentration of Na+–K+ pumps in rat skeletal muscle
Authors: Dørup I , Clausen T 
Published: December 2004

Abstract
Since adrenal steroids have been shown to upregulate the concentration of Na+–K+-ATPase in cardiac muscle, similar effects could be expected in skeletal muscle. Following infusion of dexamethasone (0·02–0·1 mg/kg per day) for 7 days in 10-week-old rats, the total concentration of [3H]ouabain-binding sites rose by up to 22–42% in soleus, extensor digitorum longus, gastrocnemius and diaphragm muscle. Dexamethasone produced no or minute changes in the Na+–K+ contents of skeletal muscle. In contrast, infusion with aldosterone (0·02–0·5 mg/kg per day) for 7 days produced hypokalemia and a graded reduction in the K+ content of skeletal muscle, which was closely correlated to a downregulation of the [3 H]ouabain-binding site concentration (r= 0·65–0·70; P<0·001). The results indicate that in skeletal muscle high doses of glucocorticoids upregulate the concentration of Na+–K+ pumps whereas mineralocorticoids induce a downregulation, which is secondary to the concomitant K+ deficiency. Since adrenalectomy produced no significant change in [3 H]ouabain-binding site concentration, basal levels of endogenous adrenal steroids seem to be of minor importance for the regulation of Na+–K+ pump concentration in skeletal muscle.
Journal of Endocrinology (1997) 152, 49–57
Concentrations of GnRH receptor and GnRH receptor mRNA in pituitary tissue of orchidectomized sheep: effect of oestradiol, progesterone, and progesterone withdrawal
Authors: Sakurai H , Adams B M , Adams T E 
Published: December 2004

Abstract
The effect of progesterone (P4) and P4 withdrawal on oestradiol (E2)-induced change in gonadotrope responsiveness (GR) and concentration of GnRH receptor and GnRH receptor mRNA in pituitary tissue of orchidectomized sheep (wethers) was determined. Thirty wethers were assigned at random to one of six treatment groups (n=5 animals/group). Wethers received E2 (2 μg/h; groups 2, 4, and 6) in 10% ethanol-saline (vehicle), or vehicle alone (groups 1, 3, and 5), as a continuous infusion for 24 h beginning 7 days after insertion (s.c.) of blank (groups 1 and 2) or P4-containing (groups 3–6) implants. The effect of P4 withdrawal was assessed by removing P4-containing implants at the beginning of vehicle (group 5) or E2 (group 6) infusion. Gonadotrope responsiveness (increase in serum LH induced by 500 ng GnRH, i.v.) was assessed at the end of infusion. In a companion study, anterior pituitary tissue was collected at the end of the 24-h infusion period. Infusion of E2 increased (P<0·05) GR relative to GR noted in control wethers receiving vehicle alone. The magnitude of E2-induced augmentation of GR was not affected by concurrent administration of P4 or P4 withdrawal. Pituitary tissue concentrations of GnRH receptor and GnRH receptor mRNA were significantly reduced in wethers implanted with P4. This P4-induced decrease in tissue concentration of GnRH receptor and GnRH receptor mRNA was not reversed during the 24-h period after P4 withdrawal. Steady-state concentrations of GnRH receptor and GnRH receptor mRNA were significantly increased by E2. However, the magnitude of oestrogen-induced increase in tissue concentrations of GnRH receptor mRNA was not significantly affected by P4 or P4 withdrawal. Conversely, concurrent progestin stimulation potentiated the E2-induced augmentation of tissue concentrations of GnRH receptor. However, this P4-induced potentiation of the oestrogenic response was not evident 24 h after removal of the P4-containing implants. Steady-state concentrations of mRNA encoding the LHβ and FSHβ subunits were reduced (P<0·05) by P4. Infusion of E2 had a similar affect. These data indicate that prolonged progestin stimulation leads to a decrease in tissue concentrations of GnRH receptor and GnRH receptor mRNA. This P4-induced suppression of GnRH receptor activity is not reversed within 24 h of P4 withdrawal. In addition, the increase in steady-state concentrations of GnRH receptor and GnRH receptor mRNA induced by E2 is not compromised by concurrent progestin stimulation.
Journal of Endocrinology (1997) 152, 91–98
Effect of the protein phosphatase inhibitor okadaic acid on FSH-induced granulosa cell steroidogenesis
Authors: Reyes J González , Santana P , Robaina I González , Oliva J Cabrera , Estévez F , Hernández I , Blanco F López , Aguiar J Quintana , Fanjul L F , de Galarreta C M Ruiz 
Published: December 2004

Abstract
To address a possible role of type 1 and 2A serine/threonine protein phosphatases (PP1 and PP2A) in regulating granulosa cell hormonal responses, we investigated the effects of okadaic acid (OA) on FSH- and cAMP-induced steroidogenesis in these cells. When added alone (0·01–1 nmol/l), the cell-permeant phosphatase inhibitor did not affect progesterone and 3β-hydroxysteroid dehydrogenase/Δ5–4 isomerase (3β-HSD) enzyme activity, whereas when added with FSH it dose-dependently augmented (minimal effective dose, 0·1 nmol/l) gonadotropin-stimulated steroidogenesis in cultured granulosa cells. A similar stimulatory effect of the toxin was observed in cells cultured for 48 h with the cell-permeant analogue dibutyryl cAMP (1 mmol/l), or when granulosa cells were stimulated with the cAMP-inducing agents cholera toxin (1 μg/ml), forskolin (15 μmol/l) or 1-methyl-3-isobutyl-xanthine (0·1 mmol/l). The observed effect of OA on FSH-supported granulosa cell steroidogenesis was not a consequence of increased cAMP generation, and time course experiments also revealed that a minimal time period of 12 h was necessary for OA (0·1 and 1 nmol/l) to significantly enhance FSH-induced progesterone and 3β-HSD enzyme activity. Since OA also inhibits the dephosphorylation of protein kinase C (PKC) substrates, we also compared the effect of OA and the PKC activator 12-0-tetradecanoylphorbol-13-acetate (TPA) on FSH-induced granulosa cell steroidogenic activity. While activation of the PKC pathway with the tumor promoter TPA (10 nmol/l) inhibited progesterone and cAMP accumulation in FSH-stimulated granulosa cells, treatment with OA augmented steroidogenesis and did not affect gonadotropin-induced cAMP generation. Collectively these results suggest that PP1 and PP2A may be important in regulating the phosphorylation state of proteins implicated in the cAMP-protein kinase A-stimulated steroidogenic activity of these cells.
Journal of Endocrinology (1997) 152, 131–139
IGF-binding protein-6 is involved in growth inhibition in SH-SY5Y human neuroblastoma cells: its production is both IGF- and cell density-dependent
Authors: Babajko S , Leneuve P , Loret C , Binoux M 
Published: December 2004

Abstract
The IGF system is involved in the growth and differentiation of neuroblastoma cells, but the precise roles played by the IGF-binding proteins (IGFBPs) remain unknown. We have examined the expression and functions of IGFBPs produced by the neuroblastoma cell line, SHSY5Y, in the presence of: insulin, IGF-I, IGF-II, des(1–3)IGF-I (an IGF-I analogue with weak affinity for IGFBPs), acidic fibroblast growth factor, basic fibroblast growth factor, or nerve growth factor. Under basal conditions, SH-SY5Y cells in serum-free medium secreted IGF-II, and traces of IGF-I, IGFBP-2 and IGFBP-4. After 24 h of culture, comparative mitogenic potencies were: des(1–3)IGF-I>IGF-1>IGF-II>insulin. After 48 h, when IGFBP-2 and IGFBP-4 concentrations in the culture media had increased, des(1–3)IGF-I remained the most active, but the activity of insulin now equalled or exceeded that of IGF-I and IGF-II. This suggests a negative feedback mechanism involving partial sequestration of IGF-I and IGF-II by IGFBP-2 and IGFBP-4. At high cell density and with high concentrations of IGF-I, des(1–3)IGF-I (40 ng/ml) or IGF-II (80 ng/ml), the mitogenic activities of the IGFs diminished concomitantly with the appearance in the culture medium of an additional IGFBP identified as IGFBP-6, whose production depended on activation of the type 1 IGF receptor. These findings suggest that IGFBP-6 contributes as an autocrine inhibitor in the regulation of growth by the IGF system in these neuroblastoma cells.
Journal of Endocrinology (1997) 152, 221–227
Mitogenic actions of endothelin and other growth factors in ovine endometrium
Authors: Salamonsen L A , Young R J , Garcia S , Findlay J K 
Published: December 2004

Abstract
Endothelin-1 (ET-1) is present in ovine endometrium, primarily in epithelial cells, and increases around the time of implantation. We examined the cell type expressing ET-binding sites in vitro and whether ET-1 has mitogenic actions in the endometrium, alone or in synergy with other growth factors. Purified epithelial and stromal cells were prepared from luteal-phase endometrium. Specific receptors were demonstrated by binding of 125I-ET-1 and proliferative effects of ET-1 and/or other growth factors determined by uptake of [3H]thymidine by cells in serum-free culture. 125I-ET-1 bound to both epithelial (2516 ± 820 c.p.m./well) and stromal (6368 ± 1350 c.p.m./well) cells and was displaced by ET-1 (1 μmol l−1). There were no proliferative effects of ET on epithelial cells. ET-1 (10 nmol l−1) stimulated uptake of [3H]thymidine by stromal cells under serum-free conditions in 13/20 individual cell preparations, to 149 ± 13% of control (untreated=100%) with dose-dependence between the range of 1 to 100 nmol l−1. Stimulation by fetal calf serum was to 377 ± 126% of control. The effects on proliferation by other growth factors (dose; % of control ± s.e.m., number of positives/total number of cell preparations) were: IGF-I (13 nmol l−1; 182 ± 14, 4/4), epidermal growth factor (EGF; 4·8 nmol l−1; 132 ± 5%, 7/7), platelet-derived growth factor-BB (0·4 nmol l−1; 146 ± 3, 2/2) and leukaemia inhibitory factor (0·4 nmol l−1; 110 ± 2, 3/3). All stimulations except that of EGF were significant and dose-responsive but only insulin was additive with ET (350 ± 35, 5/5). ET-1 also stimulated expression of the the AP-1 cis element c-jun, this being maximal at 60 min of exposure to mitogen. ET-1, along with other growth factors has a likely paracrine role in cellular proliferation in the endometrium, possibly in association with blastocyst implantation.
Journal of Endocrinology (1997) 152, 283–290
Moderate food restriction reduces serum IGF-I and alters circulating IGF-binding protein profiles in lactating rats
Authors: Monaco M H , Donovan S M 
Published: December 2004

Abstract
The role of somatogenic and lactogenic hormones in the adaptative mechanisms which occur in response to nutrient restriction during lactation is unknown. To characterize the effect of food restriction during lactation on serum IGF-I, GH and prolactin concentrations and serum IGF-binding protein (IGFBP) profiles, lactating dams had free access to food (control) or were restricted to 60% of control intake during pregnancy and lactation (RPL) or only during lactation (RL). Serum, milk and mammary gland samples were collected throughout lactation. RL dams lost body weight, control dams gained weight, while RPL dams maintained body weight during lactation. By day 20, body and mammary gland weights of RL and RPL dams did not differ and were lower than control (P<0·05). Serum IGF-I concentrations in restricted groups were lower than control (P<0·05), however, hepatic expression of IGF-I mRNA did not differ between groups in early (day 1) or mid-lactation (day 8) and was increased on day 20 in RL dams compared with RPL or control. These data suggest that serum IGF-I and hepatic IGF-I mRNA expression are not co-ordinately regulated in the food-restricted lactating rat. In early lactation, serum IGFBP-3 was lower in RPL dams than control (P<0·05), whereas IGFBP-1 and -2 were increased in RL and RPL dams in late lactation compared with control. The decrease in IGFBP-3 and increase in lower molecular weight IGFBP may have contributed to the reduction in serum IGF-I by increasing IGF-I clearance from the circulation. Serum GH and prolactin were measured in samples obtained between 0900 and 1200 h. Serum GH did not differ with the exception of an increase on day 1 in control relative to RPL dams and on day 20 in RL dams relative to RPL and control. Serum prolactin was higher in the RL dams than controls on day 4. In summary, food restriction during pregnancy and lactation or solely during lactation results in similar reductions in serum IGF-I and alterations in serum IGFBP despite differences in body weight responses to food restriction during lactation.
Journal of Endocrinology (1997) 152, 303–316
Large-scale preparation and characterization of recombinant ovine placental lactogen
Authors: Sakal E , Bignon C , Grosclaude J , Kantor A , Shapira R , Leibovitch H , Helman D , Nespoulous C , Shamay A , Rowlinson S W , Djiane J , Gertler A 
Published: December 2004

Abstract
To clone ovine placental lactogen (oPL) cDNA, total RNA from sheep placental cotyledon was reverse transcribed and the single-stranded cDNA was PCR-amplified with 5′ and 3′ primers containing, respectively, Ncol and PstI sites. The oPL cDNA fragment amplified between these two primers extended from A(−1) to the natural stop codon. The PCR product was gel-purified and subcloned into a Puc vector and the insert was sequenced on both strands, revealing several differences relative to the published sequence: S19N, S69N, D129E and R165Q. We assume that these differences can be accounted for by the high level of individual polymorphism, which has been described in detail for PLs of different species. The insert was subcloned into Ncol/PstI-digested pTrc99A procaryotic expression plasmid and protein expression was induced by isopropyl-1-thio-β-d-galactopyranoside. Because of low expression, oPL's cDNA was further subcloned into pET8 procaryotic expression plasmid. Its expression in E. coli strain BL21 transformed with this vector yielded 30–40 mg/l. The expressed protein, found in the inclusion bodies, was refolded into a monomer and purified on a Q-Sepharose column to homogeneity. Structural analysis using circular dichroism revealed a spectrum similar to that of human GH (hGH) thereby indicating proper refolding. Gel filtration and binding experiments, including real-time kinetic measurements using the surface plasmon resonance method revealed that oPL forms transient homodimeric complexes with extracellular domains of prolactin receptors from rabbit, rat and bovine and with hGH receptor. The purified oPL was biologically active in an Nb2-11C cell proliferation bioassay, in its ability to stimulate β-casein synthesis in explants of ovine and rabbit mammary gland and fat synthesis in explants of bovine mammary gland, and in a proliferation assay using FDC-P1 cells transfected with rabbit or hGH receptors.
Journal of Endocrinology (1997) 152, 317–327
Correlation between impairment of glucocorticoid feedback and the size of the pituitary gland in dogs with pituitary-dependent hyperadrenocorticism
Authors: Kooistra H S , Voorhout G , Mol J A , Rijnberk A 
Published: December 2004

Under the assumption that the impaired inhibitory effect of glucocorticoids on cell division is an important determinant in the progression of corticotrophic adenomas, it is postulated that the magnitude of proliferation and the resistance to glucocorticoids are correlated. To test this hypothesis, 67 dogs with pituitary-dependent hyperadrenocorticism were studied to determine whether a correlation could be demonstrated between the effect of dexamethasone administration on the activity of the pituitary–adrenocortical axis and the size of the pituitary gland as estimated by computed tomography.
The volumes of the pituitary glands as calculated from summations of subsequent images of pituitary areas, ranged from 11·8 to 3238·6 mm3. Among the three dimensions, the height of the pituitary was the most sensitive indicator of enlargement. Calculation of the pituitary height/brain area ratio (P/B ratio) allowed correction for the size of the dog. The P/B ratio had the highest discriminatory power in distinguishing enlarged (n=41) from non-enlarged (n=26) pituitaries.
The effects of dexamethasone (0·1 mg/kg) on the plasma concentrations of cortisol and ACTH and on the urinary corticoid/creatinine (C/C) ratios were expressed as percentage changes from the initial values. For ACTH, cortisol and C/C ratios these figures for resistance to dexamethasone were significantly correlated with the dimensions of the pituitary, particularly the height, volume and P/B ratio.
It is concluded that the magnitude of the expansion of pituitary corticotrophic adenomas is dependent upon the loss of restraint by glucocorticoids, i.e. the degree of insensitivity to glucocorticoid feedback.
Journal of Endocrinology (1997) 152, 387–394
Dexamethasone and 11-dehydrodexamethasone as tools to investigate the isozymes of 11β-hydroxysteroid dehydrogenase in vitro and in vivo
Authors: Best R , Nelson S M , Walker B R 
Published: December 2004

Abstract
Dexamethasone is used in the clinic to test the sensitivity of the hypothalamic–pituitary–adrenal axis to negative feedback. It has also been proposed that metabolism of dexamethasone might differentiate between the activities of the two isozymes of 11β-hydroxysteroid dehydrogenase (11 βHSD1 and 11βHSD2). We have developed a gas chromatographic mass spectrometric assay for dexamethasone and 11-dehydrodexamethasone and have confirmed in vitro that dexamethasone is a substrate for 11 β-HSD2 but not 11 β-HSD1 (conversion to 11-dehydrodexamethasone 0·6 ± 0·3% in homogenates of rat liver with NADP+ for 11β-HSD1, and 29·4 ±10·3% and 40·0 ± 2·0% in homogenates of rat and human kidney respectively with NAD+ for 11β-HSD2). However, we have also made the novel observation that 11-dehydrodexamethasone is a substrate for both isozymes (conversion to dexamethasone 65·0±20·4% for 11βHSD1 and 53·5 ± 20·8% and 69·0 ± 4·5% for 11βHSD2, rat and human respectively). In healthy humans, the concentrations of 11-dehydrodexamethasone in plasma after an intravenous bolus of dexamethasone were less than 10% of those of dexamethasone, and 11-dehydrodexamethasone was detected (at 0·8–65·0 nm) in plasma from only 11 of 20 subjects at 0900 h on the morning after oral dexamethasone (0·1–1 mg taken at 2400 h). Concentrations of 11-dehydrodexamethasone did not correlate with the degree of suppression of plasma cortisol. Thus dexamethasone is not useful in differentiating the activities of the isozymes of 11β-HSD in vivo and variations in 11β-HSD activity do not explain the interindividual variability in suppression of plasma cortisol by low doses of dexamethasone.
Journal of Endocrinology (1997) 153, 41–48
In vitro response to interleukin-1β and streptozotocin in pancreatic islets isolated from male and female nonobese diabetic mice
Authors: Strandell E , Sandler S 
Published: December 2004

Abstract
The aim of the present study was to examine if the islet donor gender influences the β-cell sensitivity to possible mediators of β-cell destruction in insulin-dependent diabetes mellitus (IDDM). We have currently addressed this issue by comparing the action of the cytokine interleukin-1β (IL-1β) and the alkylating agent streptozotocin (STZ), on isolated pancreatic islets derived from prediabetic female and male nonobese diabetic (NOD) mice. In this mouse strain the females have a much higher incidence of spontaneous IDDM than the males. Pancreatic islets were isolated from female and male mice of the same litter, cultured for 6 days and thereafter either 5–50 U/ml IL-1β were added for 48 h or 1·8 mm STZ for 30 min. The cytokine exposure caused a strong increase in the medium insulin accumulation in the cultures of islets obtained from both males and females. The insulin and DNA contents were similar when comparing female and male islets before and after cytokine exposure. The male pancreatic islets exhibited a higher rate of islet glucose oxidation than islets obtained from females after IL-1β addition. Non-cytokine treated islets from males showed a higher insulin content compared with corresponding female islets. After IL-1β addition male and female islets had a similar inhibition of insulin secretion following stimulation by glucose. On the other hand, islets of both genders were equally sensitive to the inhibitory action of STZ, when studied 6 days after STZ incubation, as assessed by glucose oxidation and insulin release experiments. In conclusion the data of the present study did not demonstrate a clear gender difference in the islet β-cell sensitivity to immune mediated suppression by IL-1β or alkylation damage in IDDM-prone NOD mice. However, it cannot be excluded that in vivo, under the influence of sex hormones, a decisive alteration in β-cell sensitivity to damage might develop.
Journal of Endocrinology (1997) 153, 81–86