bluuebunny/crossref_metadata_2025 huggingface dataset:
Inhibin A is a follicle stimulating hormone-responsive marker of granulosa cell differentiation, which has both autocrine and paracrine actions in sheep
Authors: Campbell BK , Baird DT
Published: December 2004

The aim of these studies was to examine the origin, control and local actions of inhibin A in monovular species, using the sheep as a model. Experiment 1 examined the pattern of mRNA expression for the inhibin subunits in relation to follicular size and pattern of expression to other differentiative markers in granulosa (P450 aromatase) and thecal cells (P450 17alpha-hydroxylase). Experiment 2 examined the pattern of inhibin A production, in relation to oestradiol, by granulosa cells induced to differentiate in vitro with follicle-stimulating hormone (FSH). Experiment 3 examined possible paracrine and autocrine actions of inhibin A by determining the effect of addition of human recombinant inhibin A and/or antiserum to inhibin on gonadotrophin-stimulated cellular differentiation. The results of Experiment 1 showed that expression of mRNA encoding inhibin subnuits alpha, beta(A) and beta(B) is greater (P<0.05) in large oestrogenic follicles than in small follicles but that only expression of the inhibin beta(A) subunit differs (P<0.05) between large oestrogenic and non-oestrogenic follicles. Expression of 17alpha-hydroxylase, but not of the luteinising hormone (LH) receptor, in thecal cells was related to both the size and the oestrogenicity of antral follicles, in a manner similar to that of the inhibin subunits. Experiment 2 demonstrated that the production of inhibin A by sheep granulosa cells is FSH-responsive after prolonged exposure (P<0.001) and precedes the production of oestradiol by around 48 h in the differentiative cascade induced in granulosa cells by FSH. The results of Experiment 3 showed that inhibin A can augment gonadotrophin-stimulated steroid production by both granulosa and theca cells (P<0.01), and that the addition of antiserum to inhibin can inhibit FSH-stimulated oestradiol production by granulosa cells from both small and large follicles (P<0.001). We conclude that inhibin A is an FSH-responsive marker of granulosa cell differentiation which has both autocrine and paracrine actions in sheep.
AP-1--glucocorticoid receptor crosstalk taken to a higher level
Authors: Karin M , Chang L
Published: December 2004

More than a decade ago our view of gene regulation by glucocorticoids (GC) and other steroid hormones underwent a dramatic change with the discovery of negative crosstalk (transcriptional interference) between the GC receptor (GCR) and transcription factor AP-1 (Jun:Fos). It was initially observed that induction of the collagenase type 1 gene, which is mediated through activation of AP-1 by growth factors and inflammatory cytokines, is repressed by GC. This repression was attributed to mutual negative interactions between AP-1 and GCR. Although the exact molecular mechanism underlying this particular case of transcriptional interference is yet to be determined, it has become clear that this and analogous interactions with other transcription factors (e.g. nuclear factor-kappaB) underlie the anti-inflammatory and immunosuppressive activity of GC. Recent studies conducted at the whole animal level indicate that the interactions between the AP-1 and GC signaling pathways are much more extensive. AP-1-related signaling via the Jun N-terminal kinases can lead to increased levels of circulating GC, which eventually down-modulate AP-1 activity via transcriptional interference. This negative feedback loop is likely to be of great importance for maintenance of homeostasis and regulation of stress responses, including acute and chronic inflammation.
Differential radiosensitivity of hypothalamo-pituitary function in the young adult rat
Authors: Robinson IC , Fairhall KM , Hendry JH , Shalet SM
Published: December 2004

Cranial irradiation in children and adults often results in irreversible hypopituitarism. The earliest and most common endocrine abnormality is GH deficiency, often followed by other pituitary hormone deficits. We investigated whether a similar pattern of progressive hypopituitarism could be reproduced in an animal model. Different doses of cranial irradiation were delivered to the hypothalamo-pituitary region of normal adult male rats, and the effects on their subsequent growth, pituitary weight and hormone contents were studied. Animals received cranial irradiation with 300 kV X-rays at doses of 0, 20, 22 or 24 Gy (n=15 per group) and five animals from each group were killed at 8, 14 or 20 weeks after irradiation. Their anterior pituitary glands were weighed and assayed for GH, LH, TSH, ACTH and prolactin (PRL) content. All three doses of irradiation reduced body weight compared with that in non-irradiated controls and compromised growth between 8 and 20 weeks. Pituitary weight increased between 8 and 20 weeks in control rats, whereas it decreased significantly in the irradiated animals. Irradiation induced time- and dose-dependent changes in pituitary hormone contents. GH and PRL were most sensitive and decreased by more than 90% after irradiation; TSH contents were unaffected 8 weeks after the lowest dose of irradiation, but were reduced at 14 and 20 weeks. LH and ACTH were the slowest to be affected, and only at the greater doses of radiation. Thus progressive multiple pituitary endocrine deficits can be induced differentially in rats by increasing doses of cranial irradiation. This model should prove useful for defining the sites and mechanisms by which cranial irradiation induces neuroendocrine dysfunction.
The acid-labile subunit (ALS) of the 150 kDa IGF-binding protein complex: an important but forgotten component of the circulating IGF system
Authors: Boisclair YR , Rhoads RP , Ueki I , Wang J , Ooi GT
Published: December 2004

The insulin-like growth factors-I and -II (IGFs) are involved in a wide array of cellular processes such as proliferation, prevention of apoptosis, and differentiation. Most of these effects are mediated by the IGF-I receptor, although at higher IGF concentrations the insulin receptor can also be activated. As the expression of both the IGFs and their receptors is widespread, IGFs are thought to have autocrine/paracrine modes of actions also, particularly during foetal life. The endocrine component of the IGF system is recognised to be important after birth, with IGF-I mediating many of the effects of growth hormone (GH), and linking anabolic processes to nutrient availability. Consideration of ligands and receptors, however, is insufficient to provide a complete understanding of the biology of IGF. This is because IGFs are found in binary complexes of 40-50 kDa with members of a family of IGF-binding proteins (IGFBPs-1 to -6) in all biological fluids. In addition, in postnatal serum, most IGFs are sequestered into ternary complexes of 150 kDa consisting of one molecule each of IGF, IGFBP-3 or IGFBP-5, and acid-labile subunit (ALS). Despite evidence that ALS plays an important role in the biology of circulating IGFs, it has received only limited attention relative to the other components of the IGF system. This review provides an overview on the current knowledge of ALS protein and gene structure, organisation and regulation by hormones, and insights from novel animal models such as the ALS knockout mice.
Correlation between inhibin secretion and damage of seminiferous tubules in a model of experimental autoimmune orchitis
Authors: Suescun MO , Suescun MO , Lustig L , Calandra RS , Calandra RS , Groome NP , Campo S
Published: December 2004

The aim of the present study was to evaluate inhibin secretion in rats with autoimmune orchitis. As we have previously described, experimental autoimmune orchitis (EAO) induced in rats by active immunization with testis homogenate and adjuvants is characterized by an interstitial mononuclear cell infiltrate and sloughing of the germinal epithelium. At 120 days after the first immunization 60% of the rats exhibited a severe orchitis with large areas of aspermatogenic seminiferous tubules in which only spermatogonia and Sertoli cells with cytoplasmic vacuolization remained attached to the tubular wall. None of the untreated (N) or control (C) rats revealed pathological alterations. Sixty percent decrease in testis weight was observed in rats with EAO compared with N or C groups. A 3-fold increase in serum FSH levels was observed in rats with EAO compared with N or C groups (19.8+/-3.7 vs 5.6+/-0.3 and 5.9+/-0.1 ng/ml respectively). A significant decrease in inhibin B levels was observed in rats with EAO when compared with N or C groups (40+/-4.6 vs 207+/-38.8 and 221.4+/-28.6 pg/ml respectively). An inverse correlation between inhibin B and FSH serum levels and a direct correlation between inhibin B and testis weight were found. Strong expression of the inhibin alpha-subunit in Sertoli cells of untreated and control rats was observed; this subunit was undetectable or poorly detectable in rats with orchitis. Positive staining for the inhibin alpha-subunit was also observed in Leydig cells of all groups studied. In conclusion, using a model of autoimmune orchitis our results show that circulating inhibin B levels and inhibin alpha-subunit expression in Sertoli cell cytoplasm closely correlate with the degree of damage of the germinal epithelium.
Interleukin-6 inhibits the expression of luteinizing hormone receptor mRNA during the maturation of cultured rat granulosa cells
Authors: Tamura K , Kawaguchi T , Kogo H
Published: December 2004

Interleukin-6 (IL-6) and its receptor components have been shown to be present in rat follicular granulosa cells. The present study was designed to examine the effect of this cytokine on changes in expression of the luteinizing hormone receptor (LHR) messenger RNA and of the steroidogenic enzyme, CYP11A1 (cytochrome P450 scc) in an in vitro model of granulosa cell maturation. Ovarian granulosa cells harvested from immature rats 2 days after treatment with equine chorionic gonadotropin were cultured for 48 h in media containing 10% fetal bovine serum. They were then transferred to a chemically defined serum-free medium and cultured for an additional 72 h. Within 24 h of transfer, the expressions of LHR and CYP11A1 mRNA increased significantly and remained increased for 72 h. The cells responded to exposure to FSH, but not LH, by an increase in production of cAMP before the additional 72 h of culture. The cAMP response to LH was attained within 24 h and persisted for 72 h, whereas the response to FSH decreased continuously with time. Inclusion of IL-6 in the culture medium caused a dose-dependent decrease in expression of LHR mRNA, in addition to a decrease in the cAMP response to LH. Immunoneutralization of endogenous granulosa cell IL-6 resulted in an increase in expression of LHR mRNA, but not CYP11A1 mRNA. The results are consistent with the view that IL-6 may have a physiological role in the maturation of ovarian follicles by modulating the attainment of the LHR in granulosa cells.
Urinary hormone levels during the natural menstrual cycle: the effect of age
Authors: Hall Moran V , Leathard HL , Coley J
Published: December 2004

A number of studies have identified hormonal changes in women during their reproductive lifespan, many focusing upon changes in women over the age of 40 years. The present study has determined the effect of increasing age on hormone levels over three decades. Daily early morning urine samples were assayed for estrone-3-glucuronide (E3 G), pregnanediol-3-glucuronide (P3 G), testosterone-17-glucuronide (T17 G), FSH and LH. An examination of the validity of using creatinine as a volume adjuster in urine samples formed an integral part of the analysis. Volunteers were healthy women who had regular (25-35 days) cycles, were not taking oral contraceptives, hormone therapies or any other medication. Three age groups were compared: 20-29 years (n=13), 30-39 years (n=9) and 40-49 years (n=13). Statistical analyses were carried out using two-way ANOVA and post hoc t-tests. Creatinine excretion, despite revealing no cycle-related variation in any age group, showed a decline with increasing age. Creatinine output was significantly lower in the 40-49 years age group in all phases of the cycle than in the 20-29 and 30-39 groups (P<0.0001). Uncorrected levels of E3 G were significantly higher in the 30-39 years group when compared with the 40-49 age group (P<0.0001). Uncorrected P3 G output was significantly higher in women aged 20-29 years than in women aged 40-49 years (P<0.001) and levels of uncorrected T17 G were higher in the 20-29 year age group when compared with the 30-39 or 40-49 years age group (P<0.0001). The present study is consistent with previous reports that have revealed a decline in creatinine clearance with increasing age, and therefore casts into some doubt the validity of using creatinine clearance as a procedure to correct for volume fluctuations in differing age groups of women. The study also demonstrates unequivocally that age-related variations in hormone levels are not restricted to women over 40 years of age. The novel finding of highly significant differences in mean levels of T17 G between the age groups is of considerable interest. It is presently unclear whether this resulted from specifically increased ovarian and/or adrenal secretion. The possible impacts of changes in testosterone levels during the female reproductive lifespan merits further study.
Cloning of the mouse sodium iodide symporter and its expression in the mammary gland and other tissues
Authors: Perron B , Rodriguez AM , Leblanc G , Pourcher T
Published: December 2004

Iodide concentration in milk by mammals is a necessary step for thyroid hormone synthesis by the newborn. With the purpose of using the mouse as an animal model to analyse the role of the sodium iodide symporter (NIS) in iodide transport and its regulation in the mammary gland, mouse NIS (mNIS) cDNA was isolated from lactating mice. The cloned sequence shows an open reading frame of 1854 nucleotides encoding a protein of 618 amino acids highly homologous to the rat and human NIS (95% and 81% identity respectively). Expression of mNIS in cultured mammalian cells induced cellular iodide accumulation. This iodide uptake process is sodium dependent and inhibited by thiocyanate and perchlorate. Tissue distribution analysis revealed that mNIS mRNAs are predominantly expressed in thyroid, stomach and in the lactating mammary gland and are present to a lower extent in several other tissues. Our data show for the first time that the level of mNIS mRNA is upregulated in the mammary gland during lactation.
Systemic administration of adrenomedullin(27-52) increases bone volume and strength in male mice
Authors: Cornish J , Callon KE , Bava U , Coy DH , Mulvey TB , Murray MA , Cooper GJ , Cooper GJ , Reid IR
Published: December 2004

Adrenomedullin is a 52-amino acid peptide first described in a human phaeochromocytoma but since been found to be present in many tissues, including the vascular system and bone. Because of its structural similarity to amylin and calcitonin gene-related peptide, both of which have actions on bone cells, we have previously assessed the effects of adrenomedullin on the skeleton, and found that it increases osteoblast proliferation in vitro and bone formation following local injection in vivo. The present study carries this work forward by assessing the effects on bone of the systemic administration of a fragment of this peptide lacking the structural requirements for vasodilator activity. Two groups of 20 adult male mice received 20 injections of human adrenomedullin(27-52) 8.1 microg or vehicle over a 4-week period and bone histomorphometry and strength were assessed. In the tibia, adrenomedullin(27-52) produced increases in the indices of osteoblast activity, osteoid perimeter and osteoblast perimeter (P<0.05 for both using Student's t-test). Osteoclast perimeter was not affected. There was a 21% increase in cortical width and a 45% increase in trabecular bone volume in animals treated with adrenomedullin(27-52) (P<0.002 for both). Assessment of bone strength by three-point bending of the humerus showed both the maximal force and the displacement to the point of failure were increased in the animals treated with adrenomedullin(27-52) (P<0.03 for both). There was also a significant increase in the thickness of the epiphyseal growth plate. No adverse effects of the treatment were noted. It is concluded that adrenomedullin(27-52) acts as an anabolic agent on bone. These findings may be relevant to the normal regulation of bone mass and to the design of agents for the treatment of osteoporosis.
Expression of the adrenomedullin binding protein, complement factor H, in the pancreas and its physiological impact on insulin secretion
Authors: Martinez A , Pio R , Lopez J , Cuttitta F
Published: December 2004

Adrenomedullin (AM) is a ubiquitous peptide hormone which, among other functional roles, reduces insulin secretion in the pancreas. Recently we have described the interaction between AM and the complement regulator protein factor H, which results in mutual modulation of their respective functions. Here we identify the expression of factor H in the beta cells of the rat pancreatic islets by immunohistochemistry and multiple immunofluorescence followed by confocal microscopy. In addition, double immunogold staining under the electron microscope showed coexistence of insulin and factor H immunoreactivities within the same secretory granules; interestingly, factor H staining was found in the electron-lucent haloes whereas the insulin antibody labeled preferentially the dense cores. The existence of factor H mRNA in the pancreas was confirmed by RT-PCR and in situ hybridization. The function of factor H in the pancreas was investigated with an insulin secretion assay. Addition of factor H to freshly isolated islets in the presence of AM resulted in a further reduction in insulin secretion with a concomitant elevation of cAMP, suggesting that factor H increases AM function in glucose homeostasis. The expression of factor H in the pancreas may play other important roles such as protection against complement-mediated cell lysis.
Effect of diethylstilbestrol on cell proliferation and expression of epidermal growth factor in the developing female rat reproductive tract
Authors: Okada A , Sato T , Ohta Y , Buchanan DL , Iguchi T
Published: December 2004

To evaluate mechanisms of cell proliferation in the fetal female rat reproductive tract, diethylstilbestrol (DES) effects on cell division and estrogen receptor (ER), epidermal growth factor (EGF) and EGF receptor (EGF-R) expressions were determined from gestational day (GD) 15.5 to 21.5. Reproductive tracts were evaluated within three regions along the Mullerian duct axis; these were proximal, middle and caudal, which differentiate into oviduct, uterus and upper vagina respectively. In fetuses from non-treated dams, epithelial and mesenchymal proliferation, as evaluated by 5-bromo-2'-deoxyuridine incorporation, was decreased with development in all regions of the Mullerian duct. EGF levels were determined by immunohistochemistry. Mullerian epithelial EGF immunoreactivity was intense in the proximal and middle regions on GDs 15.5 and 17.5. EGF staining remained intense only in the proximal epithelia by GD 19.5 and was weak in the caudal epithelium, but substantially reduced throughout epithelia in all regions by GD 21.5. Thus, decreased cell proliferation correlated with decreased EGF expression in the developing Mullerian duct. DES (100 microg/kg body weight) was injected from GD 15 to 19 and female fetuses were collected on GD 19.5. DES increased Mullerian duct cell proliferation in the proximal epithelium and mesenchyme but decreased it in the caudal epithelium compared with oil-treated controls. No proliferative DES effect was observed in any cell type in the middle region. Mullerian duct EGF immunoreactivity was suppressed by DES compared with oil. Competitive RT-PCR indicated DES also decreased mRNAs for EGF, ERbeta1 and ERbeta2, but not ERalpha and EGF-R. These results indicate EGF may be an important regulatory factor of Mullerian duct cell proliferation, and that DES may alter cell proliferation by disrupting normal EGF, ERbeta1 and ERbeta2 expression in the developing female rat reproductive tract.
Dietary soy-phytoestrogens decrease testosterone levels and prostate weight without altering LH, prostate 5alpha-reductase or testicular steroidogenic acute regulatory peptide levels in adult male Sprague-Dawley rats
Authors: Weber KS , Setchell KD , Stocco DM , Lephart ED
Published: December 2004

Nutritional factors, especially phytoestrogens, have been extensively studied for their potential beneficial effects against hormone-dependent and age-related diseases. The present study describes the short-term effects of dietary phytoestrogens on regulatory behaviors (food/water intake, locomotor activity and body weight), prostate weight, prostate 5alpha-reductase enzyme activity, reproductive hormone levels, and testicular steroidogenic acute regulatory peptide (StAR) levels in adult Sprague-Dawley rats. Animals were fed either a phytoestrogen-rich diet containing approximately 600 microg/g isoflavones (as determined by HPLC) or a phytoestrogen-free diet. After 5 weeks of consuming these diets, plasma phytoestrogen levels were 35 times higher in animals fed the phytoestrogen-rich vs phytoestrogen-free diets. Body and prostate weights were significantly decreased in animals fed the phytoestrogen-rich diet vs the phytoestrogen-free fed animals; however, no significant change in prostate 5alpha-reductase enzyme activity was observed between the treatment groups. Locomotor activity levels were higher in the phytoestrogen-rich vs the phytoestrogen-free animals during the course of the treatment interval. Plasma testosterone and androstenedione levels were significantly lower in the animals fed the phytoestrogen-rich diet compared with animals fed the phytoestrogen-free diet. However, there were no significant differences in plasma LH or estradiol levels between the diet groups. Testicular StAR levels were not significantly different between the phytoestrogen-rich vs the phytoestrogen-free fed animals. These results indicated that consumption of dietary phytoestrogens resulting in very high plasma isoflavone levels over a relatively short period can significantly alter body and prostate weight and plasma androgen hormone levels without affecting gonadotropin or testicular StAR levels. The findings of this study identify the biological actions of phytoestrogens on male reproductive endocrinology and provide insights into the protective effects these estrogen mimics exert in male reproductive disorders such as benign prostatic hyperplasia and prostate cancer.
Biochemical and functional analysis of a conserved IGF-binding protein isolated from rainbow trout (Oncorhynchus mykiss) hepatoma cells
Authors:   Bauchat,   Busby WH, Garmong A , Swanson P , Moore J , Lin M , Duan C
Published: December 2004

Rainbow trout (Oncorhynchus mykiss) serum contains several IGF-binding proteins (IGFBPs) that specifically bind to IGFs. The structures of these fish IGFBPs have not been determined and their physiological functions are poorly defined. In this study, we identified a 30 kDa IGFBP present in rainbow trout serum and secreted by cultured trout hepatoma cells. This IGFBP binds to IGFs but not to insulin. This IGFBP was purified to homogeneity using a three-step procedure involving Phenyl-Sepharose chromatography, IGF-I affinity chromatography and reverse-phase HPLC. Affinity cross-linking studies indicated that this IGFBP binds to IGF-I with a higher affinity than to IGF-II. N-terminal sequence analysis of the trout IGFBP suggests that it shares high sequence identity with that of human IGFBP-1 in the N-terminal region. When added to cultured fish and human cells, the trout IGFBP inhibited IGF-I-stimulated DNA synthesis and cell proliferation in a concentration-dependent manner. The inhibitory effect of the fish IGFBP was comparable to those of human IGFBP-1 and -4. These results indicate that the IGFBP molecule is structurally and functionally conserved in evolutionarily ancient vertebrate species such as bony fish.
Transforming growth factor-beta2 antibody attenuates fibrosis in the experimental diabetic rat kidney
Authors: Hill C , Flyvbjerg A , Rasch R , Bak M , Logan A
Published: December 2004

Diabetic nephropathy is characterised by an increase in glomerular and tubular fibrosis that compromises kidney function. The transforming growth factor-betas (TGF-betas) have been shown to play a major role in fibrosis and we have shown that TGF-beta2, in particular, increases co-ordinately with fibrogenesis in the diabetic kidney. The aim of this study was to investigate the changes in expression of extracellular matrix molecules in the diabetic kidney, with and without systemic administration of a recombinant human monoclonal antibody to TGF-beta2. Streptozotocin-induced diabetic rats were split into two groups. The first were treated with 5 mg/kg irrelevant control IgG4 (placebo) and the second treated with 5 mg/kg isoform-specific recombinant monoclonal anti-TGF-beta2 IgG4 (termed CAT-152) systemically every second day for 14 days. A further group of six non-diabetic rats was also used as a control. Various biological parameters were measured daily throughout the experimental period, and on termination of the experiment at 14 days Western blotting was performed on kidney cortices for procollagen-I C-propeptide, which is an indicator of the rate of collagen-I synthesis within the kidney. In the placebo-treated diabetic rats, blood glucose, food consumption, urinary albumin excretion (UAE) and kidney weights were all significantly higher than in the non-diabetic group (P<0.05, n=24, by ANOVA). In the anti-TGF-beta2-treated diabetic rats, kidney weights and UAE levels were decreased when compared with those in placebo-treated diabetics. Western blotting for the procollagen-I C-propeptide in kidney cortices showed a significant increase in levels in placebo-treated diabetic rats compared with non-diabetic controls over the 14 day diabetic period, indicating initiation of fibrogenesis. By contrast, in anti-TGF-beta2-treated diabetic rats, levels of the propeptide remained at non-diabetic levels. In summary, a significant suppression of kidney fibrosis was seen in anti-TGF-beta2-treated diabetic rats, compared with placebo-treated diabetic rats. We conclude that systemic delivery of CAT-152, a neutralising anti-TGF-beta2 antibody, during the acute stages of diabetic nephropathy reduces the rate of pathogenic fibrosis in the kidney.
Thyroid hormone stimulation of osteocalcin gene expression in ROS 17/2.8 cells is mediated by transcriptional and post-transcriptional mechanisms
Authors: Gouveia CH , Schultz JJ , Bianco AC , Brent GA
Published: December 2004

We investigated the mechanism of thyroid hormone regulation of osteocalcin (OC) gene expression in osteoblast-like cells (ROS 17/2.8). Treatment with tri-iodothyronine (T3) (10(-8) M) increased OC mRNA levels by approximately 3-fold after 24 h and reached a maximum, approximately 5.4-fold, after 48 h. The mRNA levels of other bone-specific genes, alkaline phosphatase and osteopontin, were not affected by T3 treatment. Interestingly, T3 induction of OC mRNA varied according to cell density: approximately 4-fold at approximately 1x10(5) cells/dish and 1.5-fold at 40-60x10(5) cells/dish. The magnitude of OC mRNA induction by T3 was approximately 40% lower than induction by 1,25 dihydroxyvitamin D3 (1,25D3) alone, and the combination of T3+1,25D3 did not further stimulate OC mRNA levels. T3 induction of OC mRNA was not affected by treatment with cycloheximide (10 microg/ml) for 5 h indicating that new protein synthesis is not required for the response. To study the half-life of OC mRNA, ROS 17/2.8 cells were incubated with actinomycin D. The basal half-life of OC mRNA (means+/-s.e.m.) was 6.4+/-0.2 h which was increased significantly with either T3 or 1,25D3 treatment to 10.9+/-0.6 h and 13.5+/-0.4 h respectively. T3 modestly up-regulated the rate of OC gene transcription (1.7+/-0.2-fold) as determined by run-off assay. T3 did not induce a reporter construct containing the rat OC gene (rOC) 5'-flanking region (to -1750 bp) or the previously described rOC vitamin D response element, when transfected into ROS 17/2.8 cells. In conclusion, T3 up-regulates the OC mRNA expression in ROS 17/2.8 cells in a dose-, time- and cell confluence-dependent fashion, and does so by transcriptional and post-transcriptional mechanisms. The greater T3 induction of OC expression in ROS 17/2.8 cells at low cell density is consistent with findings of thyroid hormone action on bone development.
Expression of the antiviral protein Mx in peripheral blood mononuclear cells of pregnant and bred, non-pregnant ewes
Authors: Yankey SJ , Hicks BA , Carnahan KG , Assiri AM , Sinor SJ , Kodali K , Stellflug JN , Stellflug JN , Ott TL
Published: December 2004

Interferon-tau (IFN tau) acts locally on the endometrium to suppress estrogen and oxytocin receptor expression and block luteolysis in ruminants. Systemic administration of conceptus homogenates or recombinant ovine IFN tau does not block luteolysis or enhance pregnancy rates in sheep or cattle, respectively. However, IFN tau up-regulates expression of the antiviral protein Mx throughout the entire uterine wall during early pregnancy. These studies determined if conceptus-derived IFN tau also up-regulates Mx expression in components of the circulating immune system that migrate through the endometrial wall. In experiment one, peripheral blood mononuclear cells (PBMC) were isolated from ewes at D26 post-artificial insemination (AI) and Mx mRNA levels examined by Northern and slot-blot hybridization. Pregnancy resulted in a two-fold increase in Mx mRNA levels compared to bred, non-pregnant ewes at D26. In experiment two, PBMC were isolated from ewes at AI, and every three days from D9 to D30. Results showed a four-fold increase in Mx mRNA levels in PBMC from pregnant versus bred, non-pregnant ewes at D15. Increased Mx mRNA, which remained elevated through D30, was accompanied by increased levels of Mx protein. These results show that pregnancy recognition signaling rapidly induces Mx gene expression in PBMC, and are the first to suggest that IFN tau activates gene expression in components of the circulating immune system.
Decreased leptin uptake in hypothalamic nuclei with ageing in Wistar rats
Authors: Fernandez-Galaz C , Fernandez-Agullo T , Campoy F , Arribas C , Gallardo N , Andres A , Ros M , Carrascosa JM
Published: December 2004

Leptin interacts with specific receptors in hypothalamic nuclei and modulates energy balance. Growing evidence has shown the association of obesity and hyperleptinaemia with non-insulin-dependent diabetes mellitus and insulin resistance. The aged Wistar rat shows peripheral insulin resistance in the absence of obesity and alterations of glucose homeostasis. However, it is not known whether, in these animals, the leptin action is altered. Here we studied the effect of ageing on plasma leptin concentration and the ability of hypothalamic nuclei to capture i.c.v.-injected digoxigenin-labelled leptin. Our data indicate that 24-month-old animals are hyperleptinaemic. However, daily food intake was greater in old animals, suggesting that they are leptin resistant. Leptin uptake in the hypothalamus was reduced in old rats. This uptake was a receptor-mediated process as demonstrated by displacement. Leptin accumulation in hypothalamic nuclei was partially colocalized with neuropeptide Y fibres. Immunohistochemical and western blot analyses showed a lower amount of the long form of leptin receptors in the hypothalamus of aged rats. Analysis by RT-PCR also demonstrated a decreased expression of leptin receptor mRNA in old animals. We conclude that the lower leptin uptake may be explained, at least in part, by a decreased amount of receptors in hypothalamic neurones of the aged rats.
Thyroid function and aging: gender-related differences
Authors: da Costa VM , Moreira DG , Rosenthal D
Published: December 2004

The effects of aging on human or animal thyroid function are still not well defined. We evaluated some aspects of thyroid function during aging using an animal model (young and old Dutch-Miranda rats). In old rats of both genders, serum thyroxine (T4) decreased but serum thyrotrophin (TSH) remained unaltered, suggesting a disturbance in the pituitary-thyroid feedback mechanism during aging. Serum tri-iodothyronine (T3) only decreased in old males, possibly because female rats are almost twice as efficient in hepatic T4 to T3 deiodination. Thyroidal T4-5'-deiodinase activity did not change much during aging, although it decreased slightly in males. Thyroidal iodothyronine-deiodinase type I mRNA expression but not total thyroidal enzymatic activity were higher in female than in male rats. Thus, ovarian/testicular hormones may modulate the expression and/or the activity of hepatic and thyroidal type I iodothyronine-deiodinase. Thyroperoxidase (TPO) and thyroglobulin (Tg) expression were higher in young male rats than in females. In males, TPO and Tg gene expression decreased with aging, suggesting that androgens might increase their expression. Our results showed that aging induces real changes in rat thyroid gland function and regulation, affecting at least pituitary, thyroid and liver functions. Furthermore, some of these changes were gender related, indicating that gonadal hormones may modulate thyroid gland function and regulation.
Bimodal inhibition of connexin 43 gap junctions decreases ACTH-induced steroidogenesis and increases bovine adrenal cell population growth
Authors: Shah US , Murray SA
Published: December 2004

In order to elucidate the role of gap junctions in adrenal cell responses, we measured the effect of inhibiting gap junctions with 18-alpha glycerrhetinic acid (GA; a potent inhibitor of cell-cell communication) and connexin antisense transfection on cell proliferation and adrenocorticotropin (ACTH)-stimulated steroidogenesis. In these experiments we utilized a bovine adrenocortical cell (SBAC) population, which responds to ACTH treatment with a dose-dependent increase in steroid production, an increase in connexin 43 (alpha(1)-Cx43) gap junction protein concentrations, and a decrease in cell population growth. SBAC cell populations treated with GA had increased growth rates, decreased ACTH-stimulated steroidogenesis, but no reduction in alpha(1)-Cx43 gap junction protein contents. In contrast, when SBAC cells were transfected with alpha(1)-Cx43 antisense cDNA, gap junction protein concentration was dramatically reduced as expected, unlike the GA-treated cell populations. Cell populations transfected with alpha(1)-Cx43-antisense also exhibited increased growth rates and a decreased steroidogenic response to ACTH treatment as compared with control or vector-only transfected cell populations. The decreased responsiveness and increased number of cells in the population after gap junction function was decreased by either GA treatment or antisense transfection, suggests that gap junctions may be necessary factors in ACTH-stimulated responsiveness and growth control in the adrenal gland.
Enhanced tumor necrosis factor-alpha-specific and decreased interleukin-10-specific immune responses to LPS during the third trimester of pregnancy in mice
Authors: Vizi ES , Szelenyi J , Selmeczy ZS , Papp Z , Nemeth ZH , Hasko G
Published: December 2004

It is increasingly apparent that there is a bidirectional interaction between the maternal immune system and the reproductive system during pregnancy. Pregnancy is associated with a suppression of maternal specific immune responses, which process underlies the protection of fetal tissues expressing paternally inherited alloantigens. However, recent evidence indicates that the suppression of specific, lymphocyte-mediated immune responses during pregnancy is accompanied by activation of the non-specific arm of the maternal immune response. In the present study, we have investigated the effect of pregnancy on the non-specific immune response induced by bacterial lipopolysaccharide (LPS, endotoxin) in mice. Pregnancy enhanced the LPS-induced production of proinflammatory cytokines, including tumor necrosis factor-alpha, interleukin (IL)-6, and interferon-gamma. On the other hand, LPS-induced levels of the anti-inflammatory cytokine IL-10 were suppressed in pregnant mice. These alterations in cytokine production correlated with an increased susceptibility for endotoxemic mortality in the pregnant mice. Although adrenergic receptors are important regulators of cytokine production in non-pregnant mice, the alpha(2)- and the beta-adrenoceptor-mediated modulation of cytokine production ceases to operate during pregnancy associated with severe endotoxemia. These data may explain how excessive activation of the non-specific immune responses during pregnancy can contribute to the increased severity of some maternal diseases, including septic shock, and can be an important pathophysiological factor in disseminated intravascular coagulation or preeclampsia.
Temporally regulated overexpression of parathyroid hormone-related protein in the mammary gland reveals distinct fetal and pubertal phenotypes
Authors: Dunbar ME , Dann P , Brown CW , Van Houton J , Dreyer B , Philbrick WP , Wysolmerski JJ
Published: December 2004

We have previously demonstrated that overexpression of parathyroid hormone-related protein (PTHrP) in the mammary glands of transgenic mice results in defects in ductal elongation and branching during puberty and in lobuloalveolar development during pregnancy. In addition, we have shown that PTHrP is necessary for the formation of the initial ductal tree during embryonic mammary development. In order to examine the effect of varying the timing of PTHrP overexpression on mammary development, we created tetracycline-regulated, K14-tTA/Tet(O)-PTHrP double transgenic mice. In this report, we document that this 'tet-off' system directs transgene expression to the mammary gland and that it is fully repressed in the presence of tetracycline. Using these mice, we demonstrate that transient overexpression of PTHrP before birth causes defects in ductal branching during puberty and that overexpression of PTHrP during puberty decreases the rate of ductal elongation. Furthermore, we demonstrate that if PTHrP overexpression is initiated after ductal morphogenesis is completed, lobuloalveolar development is unaffected. Finally, we demonstrate that the impairment in ductal elongation caused by PTHrP is associated with an increase in the basal rate of epithelial cell apoptosis in terminal end buds and a failure to increase end bud cell proliferation and decrease apoptosis in response to estrogen and progesterone.
Characterization of topology-, gestation- and labor-related changes of a cassette of myometrial contraction-associated protein mRNA in the pregnant baboon myometrium
Authors: Wu WX , Ma XH , Zhang Q , Nathanielsz PW
Published: December 2004

Our objective was to examine the topology-, gestation- and labor-related changes of estrogen receptor (ER)alpha, progesterone receptor (PR), oxytocin receptor (OTR) and thrombospondin-1 (TSP1) mRNA in pregnant baboon myometrium. ER alpha, PR, OTR and TSP1 mRNAs extracted from the lower uterine segment and fundal myometrium of pregnant baboons not in labor between 121 and 180 days of gestational age (n=9) and in established spontaneous labor between 164 and 193 days of gestational age (n=5) were analyzed by Northern blot. There were no topology-, gestation- or labor-related changes of ER alpha and PR mRNA in or between the lower uterine segment or/and the fundus. OTR mRNA was the same in the lower uterine segment and the fundus from baboons not in labor and non-labor fundal, but not lower uterine segment, myometrial OTR mRNA increased with gestation (R(2)=0.81, P<0.05). Fundal OTR mRNA rose significantly compared with the lower uterine segment during spontaneous labor. TSP1 mRNA increased significantly in both the fundus and lower uterine segment during labor. TSP1 mRNA in the lower uterine segment during spontaneous labor was higher than in the fundus during spontaneous labor. In conclusion, fundal and lower uterine segment ER alpha and PR mRNA remained unchanged in late gestation and spontaneous labor. The increased OTR mRNA may serve as a mechanism to increase uterine sensitivity to OT during late gestation. The higher fundal OTR mRNA compared with the lower uterine segment provides polarity which assists fetal expulsion by uterine contractions during labor. The significance of increased TSP1 mRNA during labor may relate to homeostasis and merits further study.
Postprandial hormone and metabolic responses amongst shift workers in Antarctica
Authors: Lund J , Arendt J , Hampton SM , English J , Morgan LM
Published: December 2004

The circadian rhythms of many night-shift workers are maladapted to their imposed behavioural schedule, and this factor may be implicated in the increased occurrence of cardiovascular disease (CVD) reported in shift workers. One way in which CVD risk could be mediated is through inappropriate hormonal and metabolic responses to meals. This study investigated the responses to standard meals at different circadian times in a group of night-shift workers on a British Antarctic Survey station at Halley Bay (75 degrees S) in Antarctica. Twelve healthy subjects (ten men and two women) were recruited. Their postprandial hormone and metabolic responses to an identical mixed test meal of 3330 kJ were measured on three occasions: (i) during daytime on a normal working day, (ii) during night-time at the beginning of a period of night-shift work, and (iii) during the daytime on return from night working to daytime working. Venous blood was taken for 9 h after the meal for the measurement of glucose, insulin, triacylglycerol (TAG) and non-esterified fatty acids. Urine was collected 4-hourly (longer during sleep) on each test day for assessment of the circadian phase via 6-sulphatoxymelatonin (aMT6s) assay. During normal daytime working, aMT6s acrophase was delayed (7.7+/-1.0 h (s.e.m.)) compared with that previously found in temperate zones in a comparable age-group. During the night shift a further delay was evident (11.8+/-1.9 h) and subjects' acrophases remained delayed 2 days after return to daytime working (12.4+/-1.8 h). Integrated postprandial glucose, insulin and TAG responses were significantly elevated during the night shift compared with normal daytime working. Two days after their return to daytime working, subjects' postprandial glucose and insulin responses had returned to pre-shift levels; however, integrated TAG levels remained significantly elevated. These results are very similar to those previously found in simulated night-shift conditions; it is the first time such changes have been reported in real shift workers in field conditions. They provide evidence that the abnormal metabolic responses to meals taken at night during unadapted night shifts are due, at least in part, to a relative insulin resistance, which could contribute to the documented cardiovascular morbidity associated with shift work. When applied to the 20% of the UK workforce currently employed on shift work, these findings have major significance from an occupational health perspective.
Quantification of prolactin receptor mRNA in multiple human tissues and cancer cell lines by real time RT-PCR
Authors: Peirce SK , Chen WY , Chen WY
Published: December 2004

Human prolactin (hPRL) has been reported to be involved in breast and prostate cancer development. The hPRL receptor (hPRLR) is expressed in a wide variety of tissues in at least three isoforms. In this study, a one-step real time reverse transcription PCR technique was used to determine relative expression levels of hPRLR mRNA in eleven human breast cancer cell lines, HeLa cells, three prostate cancer cell lines and nine normal human tissues. The housekeeping gene beta-actin was used for internal normalization. We demonstrate that hPRLR mRNA is up-regulated in six of the eleven breast cancer cell lines tested when compared with normal breast tissue. Of the cancer cell lines tested, we found that T-47D cells have the highest level of hPRLR mRNA, followed by MDA-MB-134, BT-483, BT-474, MCF-7 and MDA-MB-453 cells. In two breast cancer cell lines (MDA-MB-468 and BT-549), the hPRLR levels were found to be comparable to that of normal breast tissue. Three breast cancer cell lines (MDA-MB-436, MDA-MB-157 and MDA-MB-231) expressed hPRLR mRNA at levels lower than that of normal tissue. In contrast, in all three commonly used prostate cancer cell lines (LNCaP, PC-3 and DU 145), the levels of hPRLR mRNA were found to be down-regulated relative to that of normal prostate tissue. Of nine normal human tissues tested, we found that the uterus and the breast have the highest levels of hPRLR mRNA, followed by the kidney, the liver, the prostate and the ovary. The levels of hPRLR mRNA were the lowest among the trachea, the brain and the lung.
Lithium chloride inhibits the expression and secretion of insulin-like growth factor-binding protein-1
Authors: Lewitt MS , Brismar K , Ohlson J , Hartman J
Published: December 2004

Insulin-like growth factor-binding protein-1 (IGFBP-1) regulates IGF availability for glucose homeostasis. The IGFBP-1 promoter shares common regulatory response elements with phosphoenol pyruvate carboxykinase (PEPCK), the expression and activity of which is inhibited by lithium chloride, associated with an inhibition of glycogen synthase kinase (GSK)-3 activity, in the rat hepatoma cell line H4-II-E. We therefore determined the effect of lithium chloride on IGFBP-1 expression and secretion in H4-II-E cells. Lithium chloride inhibited IGFBP-1 secretion in a dose response and reversible manner by approx 80% during 5-h and 16-h incubations. An inhibitory effect on IGFBP-1 mRNA expression was observed at 2 h. The inhibitory effect of lithium and insulin were not additive when used alone, but inhibition by lithium occurred when insulin action was blocked by activating AMP-activated protein kinase with 5-aminoimidazole-4-carboxamide-riboside (AICAR). These findings suggest that GSK-3 inhibition, or another pathway activated by lithium, may be involved in a pathway controlling IGFBP-1, inhibiting synthesis when insulin activity is absent or impaired.
GH as a co-gonadotropin: the relevance of correlative changes in GH secretion and reproductive state
Authors: Hull KL , Harvey S
Published: December 2004

It is now well established that exogenous GH promotes sexual maturation and reproductive function. The possibility that this may reflect physiological actions of endogenous GH has, however, rarely been considered. Correlative changes in GH secretion and reproductive state (puberty, pregnancy, lactation, menopause and ovarian cycles) are thus the primary focus of this review. GH secretion is, for instance, elevated during major transitions in reproductive status such as puberty and pregnancy. In some cases, augmented circulating GH levels are paired with hepatic GH resistance. This interaction may permit selective activation of gonadal responses to GH without activating IGF-I-mediated systemic responses. This selective activation may also be mediated by autocrine, paracrine or intracrine GH actions, since GH is also synthesized in reproductive tissues. Correlative changes in GH secretion and reproductive state may be mediated by events at the hypothalamic, pituitary and gonadal level. In addition to direct effects on gonadal function, GH may influence reproductive activity by increasing gonadotropin secretion at the hypothalamic and pituitary level and by enhancing gonadotropin responsiveness at the gonadal level. The close association between reproductive status and the somatotrophic axis supports the physiological importance of GH in reproductive function.