bluuebunny/crossref_metadata_2025 huggingface dataset:
Thyroid and pituitary thyroxine-5'-deiodinase activity and thyrotrophin secretion in lithium-treated rats
Authors: Frankenfeld TG , Correa Da Costa VM , Nascimento-Saba CC , Ortiga-Carvalho TM , Santos RM , Lisboa PC , Carvalho DP , Rosenthal D
Published: December 2004

Some authors have reported increased serum thyrotrophin (TSH) in animals chronically treated with lithium, suggesting that lithium might decrease pituitary thyroxine (T(4))-5'-deiodinase activity. On the other hand, the effect of lithium treatment on thyroidal T(4)-5'-deiodinase activity is also unknown. The present study was undertaken to evaluate the effects of lithium treatment on pituitary and thyroid T(4)-5'-deiodinase activity. Serum and pituitary TSH levels and thyroidal and pituitary T(4)-5'-deiodinase activities were determined in 3-month-old isogenic male Dutch-Miranda rats treated with lithium for 8 weeks. Chronic lithium treatment produced a slight increase in pituitary TSH content, but no change in serum TSH, and a significant increase in the thyroidal T(4)-5'-deiodinase activity. However, the pituitary T(4)-5'-deiodinase activity was unaffected by lithium administration. As far as we know, the present data show for the first time that chronic lithium treatment can increase the thyroxine to tri-iodothyronine conversion in the murine thyroid gland, be it directly or indirectly.
Nutrient-hormone interaction in the ovine liver: methionine supply selectively modulates growth hormone-induced IGF-I gene expression
Authors: Stubbs AK , Wheelhouse NM , Lomax MA , Hazlerigg DG
Published: December 2004

This study tested the hypothesis that specific amino acids are responsible for modulating the insulin-like growth factor-I (IGF-I) response to growth hormone (GH) in ovine hepatocytes. Cells were grown in media of defined amino acid composition, based on physiological concentrations (P.C.) of amino acids in sheep plasma. Relative to culture in 5 x P.C., amino acid limitation to 0.2 x P.C. had inhibitory effects on IGF-I RNA expression, peptide release and p70 S6 kinase phosphorylation (P<0.01 in each case). Limitation of methionine levels to 0.2 x P.C. against a background of 5 x P.C. for the other amino acids blocked GH-stimulated IGF-I peptide release and RNA expression, although basal expression was unaffected. In contrast, limitation of the other amino acids present in the culture medium had no effect on basal or GH-stimulated IGF-I expression. Selective methionine limitation to 0.2xP.C. levels had no effect on cellular or secretory protein synthesis rates relative to cells grown in complete 5 x P.C. medium but did cause a partial reduction in p70 S6 kinase phosphorylation, which was also observed when medium was selectively limited for other essential amino acids. The addition of rapamycin (5 ng/ml) to cells grown in 5xP.C. media completely abolished p70 S6 kinase phosphorylation (P<0.001), implicating mTOR in the response of S6 kinase phosphorylation to changing amino acid supply. By contrast, inclusion of rapamycin (100 ng/ml) had no effect on levels of IGF-I gene expression. These results indicate that methionine is the key limiting amino acid involved in the modulation of IGF-I expression in the ovine liver. This nutrient-hormone interaction is a highly selective phenomenon, occurring against a background of modest effects on general protein synthetic control. The partial inhibitory effects of methionine on mTOR activity are not sufficient to account for this selectivity of action.
Dexamethasone-induced growth inhibition of porcine growth plate chondrocytes is accompanied by changes in levels of IGF axis components
Authors: Smink JJ , Koedam JA , Koster JG , van Buul-Offers SC
Published: December 2004

High (pharmacological) doses of glucocorticoids inhibit the proliferation of growth plate chondrocytes, which leads to one of the side-effects of these steroids, namely suppression of longitudinal growth. Growth inhibition by glucocorticoids is thought to be mediated in part by impaired action of components of the IGF axis, which are important for chondrocyte regulation and hence for longitudinal growth. The aim of the present study was to determine whether glucocorticoid-induced growth retardation involves changes in IGF axis components. Chondrocytes were isolated from epiphyseal growth plates of neonatal piglets and treated with pharmacological doses of dexamethasone (DXM) for 24 h to study glucocorticoid-induced growth retardation. Under IGF-I-supplemented (10 nM) culture conditions, IGF-binding proteins (IGFBPs)-2, -4 and -5 were secreted by the growth plate chondrocytes and IGFBP-2 protein and mRNA levels were decreased by the DXM treatment, whereas IGFBP-4 and -5 were not affected. Proliferation of the chondrocytes, as measured by [(3)H]thymidine incorporation, was 3.5-fold higher in serum-supplemented medium in contrast to IGF-I-supplemented (10 nM) medium. In the presence of serum, DNA synthesis was significantly inhibited by 50-63% when treated with 100 nM DXM, which was prevented by the glucocorticoid-receptor antagonist Org34116. mRNA levels of IGF axis components were determined using Northern blot analysis. IGFBP-2 to -6 were expressed in the chondrocytes, IGFBP-1 was absent and both IGF-I and IGF-II, and the type I and type II IGF receptors were expressed. Treatment with DXM (100 nM) resulted in a 2-fold increase in mRNA levels of both IGFBP-5 and the type I IGF receptor, whereas IGFBP-2 mRNA levels decreased by 55%, in concert with the decrease in protein level observed under IGF-I-supplemented culture conditions. The changes in mRNA levels due to the DXM treatment were prevented by the glucocorticoid receptor antagonist. Our data show that exposure to pharmacological doses of DXM results in inhibition of proliferation and changes in components of the IGF axis, IGFBP-2 and -5 and the type I IGF receptor, suggesting a role for these components in glucocorticoid-induced growth retardation at the local level of the growth plate.
Affected skeletal growth but normal bone mineralization in rat offspring after prenatal dexamethasone exposure
Authors: Swolin-Eide D , Dahlgren J , Nilsson C , Albertsson Wikland K , Holmang A , Ohlsson C
Published: December 2004

Events occurring early in life or prenatally are able to play important roles in the pathogenesis of diseases in adult life. Different sorts of stress or hormonal influences, during particular periods of pregnancy, may result in persisting or transient changes in physiology. Glucocorticoids are used for the treatment of a variety of diseases, to promote organ maturation and to prevent preterm delivery. Glucocorticoids are also known to affect skeletal growth and adult bone metabolism. The aim of the present study was to investigate whether exposure to dexamethasone (Dex) during fetal life has any effect on skeletal growth and/or bone mineral density in adult rat offspring. Pregnant rats were given injections of either Dex (100 micro g/kg) or vehicle on days 9, 11 and 13 of gestation. Dex-exposed male but not female rat offspring showed transient increases in crown-rump length and tibia and femur lengths at 3-6 weeks of age. In contrast, the cortical bone dimensions were altered in 12-week-old female but not male Dex-exposed offspring. The areal bone mineral densities of the long bones and the spine, as determined by dual X-ray absorptiometry, and trabecular as well as cortical volumetric bone mineral density, as measured using peripheral quantitative computerized tomography, were unchanged in both male and female Dex-exposed offspring. In conclusion, prenatal Dex exposure affects skeletal growth in a gender-specific manner, while the mineralization of bones is unaffected in both male and female offspring.
Clodronate inhibits adrenocortical cell proliferation and P450c21 activity
Authors: Fassnacht M , Franke A , Dettling A , Hahner S , Zink M , Wudy S , Allolio B
Published: December 2004

Bisphosphonates (BPs), specific inhibitors of osteoclasts, are well established in the management of skeletal metastases in breast cancer. Recent studies have suggested that these compounds may also directly influence tumor cell proliferation. As adrenocortical cancer frequently leads to bone metastases, we investigated the effects of clodronate (CLO) in the human adrenocortical cancer cell line NCI-H295 and in primary cultures of bovine adrenocortical cells. Both the non-amino BP CLO and the amino BP pamidronate (PAM) exhibited a dose-dependent antiproliferative effect in both cell types (cell viability in NCI-H295 cells: 100 microM CLO: 98+/-8%; 500 microM CLO 76+/-6%; 1000 microM CLO 53+/-2; 10 microM PAM 96+/-3%; 50 microM PAM 46+/-6%; 100 microM PAM 11+/-1% vs untreated control cells: 100+/-10%; P<0.01). FACS analysis in NCI-H295 cells treated with either CLO or PAM demonstrated both apoptotic and necrotic cell death. However, while during PAM treatment cell number and hormone secretion decreased in parallel, we observed specific impairment of steroidogenesis in the presence of CLO with a dose-dependent increase in the 17-OH-progesterone/cortisol ratio (100 microM CLO 134+/-30%; 500 microM CLO 284+/-10%; 1000 microM CLO 545+/-130% vs 100+/-20% in control cells; P<0.01). Further analysis in ACTH-stimulated bovine adrenal cells using stable isotope dilution/gas chromatography-mass spectrometry demonstrated CLO-induced inhibition of adrenal 21-hydroxylase (P450c21) activity leading to a dose-dependent increase in the 17-OH-progesterone/11-deoxycortisol ratio. In conclusion, we demonstrate a dose-dependent antiproliferative effect of CLO and PAM in adrenocortical cells. In addition, for the first time, we describe a suppressive effect of CLO on steroidogenesis via inhibition of adrenal 21-hydroxylase (P450c21) activity.
Differential expression of IgG Fc binding protein (FcgammaBP) in human normal thyroid tissue, thyroid adenomas and thyroid carcinomas
Authors: O'Donovan N , Fischer A , Abdo EM , Simon F , Peter HJ , Gerber H , Buergi U , Marti U
Published: December 2004

The genetic events involved in thyroid carcinogenesis are still incompletely understood. Several rearrangements and mutations of oncogenes have been implicated in the development of thyroid papillary carcinomas, follicular adenomas and carcinomas. However, none of these molecular alterations is suitable either as a general marker for the diagnosis of thyroid carcinomas or to differentiate between thyroid follicular adenomas and carcinomas. In order to identify new genes with altered expression which could serve as such markers, we analyzed RNA from thyroid tumor and normal tissue using a novel technique called restriction-mediated differential display. Several differentially expressed genes were identified, including the gene for IgG Fc binding protein (FcgammaBP). Differential expression of FcgammaBP was confirmed by quantitative real-time RT-PCR. Our experiments showed that IgG Fc binding protein (FcgammaBP) is differentially expressed in normal thyroid tissue, thyroid adenomas and thyroid carcinomas. While the FcgammaBP gene is constitutively expressed in normal thyroid tissue, its expression is significantly increased in follicular thyroid adenomas and significantly decreased in papillary and follicular thyroid carcinomas. Thus, measurement of the expression levels of FcgammaBP in thyroid biopsies might help to make the otherwise difficult distinction between a thyroid follicular adenoma and a follicular carcinoma.
Nuclear effects: unexpected intracellular actions of insulin-like growth factor binding protein-3
Authors: Lee KW , Cohen P
Published: December 2004

Insulin-like growth factor (IGF) binding protein (IGFBP)-3 has been shown to be a growth inhibitory, apoptosis-inducing molecule by virtue of its ability to bind IGFs, in addition to previously demonstrated IGF-independent effects. The recent discovery of the interaction between nuclear IGFBP-3 and 9-cis retinoic acid receptor-alpha (retinoid X receptor alpha RXRalpha), a nuclear receptor, and its involvement in the regulation of transcriptional signaling and apoptosis represents an important paradigm shift in the understanding of IGFBP function. RXRalpha is required for the apoptosis-inducing effects of IGFBP-3. IGFBP-3 and RXR ligands are additive in inducing apoptosis in cancer cells. IGFBP-3 has direct effects on gene transcription, as RXR response element reporter signaling was enhanced and the all-trans retinoic acid receptor response element reporter signaling was inhibited. Accumulating evidence further confirms IGF-independent functions of this multifunction binding protein. Other binding proteins, in addition to other members of the IGF axis, have now been described in the nucleus and are postulated to have effects on transcriptional events. Investigation into these new interactions will expose new protein partners in the interface between the nuclear receptor and growth factor pathways and reveal new targets to be exploited in the treatment of cancer and other diseases.
Evolution of the visual cycle: the role of retinoid-binding proteins
Authors: Gonzalez-Fernandez F
Published: December 2004

The trafficking of retinoids in the retina represents a model to study soluble hormone-binding proteins in a complex system subject to profound evolutionary adaptations. Although a remarkable illustration of convergent evolution, all visual systems detect light in the same way, that is through the photoisomerization of an 11-cis retinoid to a corresponding trans isomer. What is strikingly different between the systems, is the mechanism by which the 11-cis chromophore is reformed and visual pigment regenerated in a process known as the visual cycle. The variations of the cycle address a problem inherent to retinoids themselves. That is, the properties that make these molecules suited for light detection also account for their susceptibility to oxidative and isomeric degradation, and cellular toxicity. The cycle therefore provides an opportunity to examine the role of soluble hormone-binding proteins within an integrative and evolutionary context. The present review focuses on interphotoreceptor retinoid-binding protein (IRBP), a controversial glycolipoprotein that recruits a protein fold common to Cterminal-processing proteases and the crotonase family. This unorthodox retinoid-binding protein is entrapped in the subretinal compartment of those eyes that translocate visual cycle retinoids between the photoreceptors and the retinal pigment epithelium. Recent studies suggest that we should look beyond a strictly carrier function if we are to appreciate the role of IRBP in the visual cycle. Here we draw lessons from other soluble hormone-binding proteins to anticipate avenues of future research likely to provide insight into the structure and function of IRBP in vision.
IGF and IGF-binding protein expression in the growth plate of normal, dexamethasone-treated and human IGF-II transgenic mice
Authors: Smink JJ , Koster JG , Gresnigt MG , Rooman R , Koedam JA , Van Buul-Offers SC
Published: December 2004

Glucocorticoid (GC) treatment in childhood can lead to suppression of longitudinal growth as a side effect. The actions of GCs are thought to be mediated in part by impaired action of the insulin-like growth factors (IGF-I and IGF-II) and their binding proteins (IGFBP-1 to -6). We have studied the effects of GCs on IGF and IGFBP expression at the local level of the growth plate, using non-radioactive in situ hybridization. We treated 3-week-old normal mice for 4 weeks with dexamethasone (DXM). We also treated human IGF-II (hIGF-II) transgenic mice in order to investigate whether IGF-II could protect against the growth retarding effect of this GC. DXM treatment resulted in general growth retardation in both mice strains, however, only in normal mice was tibial length decreased. In both normal and hIGF-II trangenic mice, the total width of the growth plate was not affected, whereas the width of the proliferative zone decreased as a result of the DXM treatment. Additionally, only in normal mice, the width of the hypertrophic zone thickened. Only expression of IGF-I, IGF-II and IGFBP-2 could be detected in the growth plates of 7-week-old normal mice. IGFBP-1, -3, -4, -5 and -6 mRNAs were not detected. DXM treatment of normal mice induced a significant 2.4-fold increase in the number of cells expressing IGF-I mRNA, whereas IGF-II and IGFBP-2 mRNA levels were not affected. In hIGF-II transgenic mice, IGF-I mRNA levels were significantly increased, while endogenous IGF-II and IGFBP-2 mRNAs were unaffected, compared to normal animals. DXM treatment of the hIGF-II transgenic mice induced a further increase of IGF-I mRNA expression, to a similar extent as in DXM-treated normal mice. The increase of IGF-I due to DXM treatment in normal mice might be a reaction in order to minimize the GC-induced growth retardation. Another possibility could be that the increase of IGF-I would contribute to the GC-induced growth retardation by accelerating the differentiation of chondrocytes, resulting in accelerated ossification. In the growth plates of hIGF-II transgenic mice, the higher basal level of IGF-I, might be responsible for the observed partial protection against the adverse effects of GCs on bone.
Impaired endothelial dysfunction in diabetes mellitus rats was restored by oral administration of prostaglandin I2 analogue
Authors: Matsumoto K , Morishita R , Tomita N , Moriguchi A , Yamasaki K , Aoki M , Matsumoto K , Nakamura T , Higaki J , Ogihara T
Published: December 2004

We have previously reported that a decrease in hepatocyte growth factor (HGF), which has many protective functions against endothelial damage by high d-glucose, might be a trigger of endothelial injury. However, the regulation of vascular HGF in diabetes mellitus (DM) has not been clarified in vivo, although vascular disease is frequently observed in DM patients. In addition, our previous report revealed that a prostaglandin I(2) (PGI(2)) analogue prevented endothelial cell death through the induction of vascular HGF production in cultured human epithelial cells. Thus, in this study, we examined the effects of a PGI(2) analogue in the regulation of the local HGF system using DM rats. A PGI(2) analogue (beraprost sodium; 300 and 600 micro g/kg per day) or vehicle was administered to 16-week-old DM rats induced by administration of streptozotocin for 28 days. Endothelial function was evaluated by the vasodilator response to acetylcholine, and the expression of vascular HGF mRNA was measured by Northern blotting. Of importance, expression of HGF mRNA was significantly decreased in the blood vessels of DM rats as compared with non-DM (P<0.01). In addition, the in vitro vasodilator response of the abdominal aorta to acetylcholine was markedly impaired in DM rats. Importantly, the vasodilator response was restored by PGI(2) treatment in a dose-dependent manner (P<0.01), whereas N(omega)-nitro-l-arginine methyl ester inhibited the restoration of endothelial function. Of particular interest, vascular HGF mRNA and protein were significantly increased in the blood vessels of DM rats treated with PGI(2) as compared with vehicle. Similarly, an increase in HGF protein was also confirmed by immunohistochemical analysis. In addition, the specific HGF receptor, c-met, was also increased by PGI(2) treatment. Overall, this study demonstrated that treatment with a PGI(2) analogue restored endothelial dysfunction in DM rats, accompanied by the induction of vascular HGF and c-met expression. Increased local vascular HGF production by a PGI(2) analogue may prevent endothelial injury, potentially resulting in the improvement of endothelial dysfunction.
Changes of endothelin in streptozotocin-induced diabetic rats: effects of an angiotensin converting enzyme inhibitor, enalapril maleate
Authors: Itoh Y , Imamura S , Yamamoto K , Ono Y , Nagata M , Kobayashi T , Kato T , Tomita M , Nakai A , Itoh M , Nagasaka A
Published: December 2004

Endothelin-1 (ET-1) concentrations are increased in patients with diabetes mellitus, particularly those with diabetic retinopathy, or essential hypertension. We hypothesized that ET-1 might participate in the development and progression of diabetic microangiopathy. In this study, the effects of the angiotensin converting enzyme (ACE) inhibitor, enalapril maleate, on diabetic angiopathy were examined in streptozotocin (STZ)-induced diabetic (STZ-DM) rats by monitoring variations in renal function and ET-1 concentrations in blood and organ tissues. Significant increases in kidney weight and in concentrations of urinary albumin, N-acetyl-fl-d-glucosamidase (NAG) and serum ET-1 were observed in the STZ-DM rats as compared with the non-diabetic rats, and the concentration of ET-1 in the kidneys tended to be increased. Microscopic and electron microscopic analyses showed increased mesangial cell proliferation, matrix expansion and enlarged mesangial area in the kidney of the diabetic rats. After administration of the ACE inhibitor, increased concentrations of urinary albumin and NAG in the STZ-DM rats were reduced to the control values with a slight improvement in the electron microscopic changes. These data suggest that ET-1 may be involved in the development and progression of diabetic nephropathy and may explain, in part, why diabetes is liable to complicate hypertension. ACE inhibitor may help to restore diabetic nephropathy in the STZ-induced diabetic rats.
How much LH do the Leydig cells see?
Authors: Setchell BP , Pakarinen P , Huhtaniemi I
Published: December 2004

The purpose of this study was to assess the concentrations of LH that Leydig cells are exposed to upon in vivo stimulation of steroidogenesis. The concentrations of LH were measured in rats in testicular interstitial extracellular fluid, seminiferous tubular fluid and blood plasma from testicular veins from one testis before and from the other testis of the same rats after an intravenous injection of gonadotrophin-releasing hormone (GnRH) or saline, and compared with the concentrations in blood plasma from a peripheral vein. The concentrations of LH in interstitial fluid surrounding the Leydig cells before the injections were about 10% of the levels in blood plasma, and showed no significant rise at 15 min and a much smaller rise at later times in rats injected with GnRH than those seen in blood plasma from either of the two sources, which were similar. The concentrations of LH in tubular fluid were even lower and showed no change after GnRH. Testosterone concentrations in testicular cells, interstitial fluid and testicular venous blood plasma were significantly increased by 15 min after GnRH, when compared with saline-injected controls, with no change in the levels in tubular fluid. The rise in testosterone concentrations in testicular venous plasma after GnRH was smaller than those in the cells and interstitial fluid. In conclusion, the concentrations of LH reaching the testicular interstitial fluid were only about one-tenth of that measured in the circulation, presumably because the endothelial cells restrict access of the hormone to the interstitial fluid. This indicated that either the Leydig cells are extremely sensitive to LH stimulation or that testicular endothelial cells modulate the action of LH on the Leydig cells.
Seasonal and dose-dependent effects of intracerebroventricular leptin on lh secretion and appetite in sheep
Authors: Miller DW , Findlay PA , Morrison MA , Raver N , Adam CL
Published: December 2004

The role of leptin in neuroendocrine appetite and reproductive regulation remains to be fully resolved. A series of three experiments was conducted using adequately nourished oestradiol-implanted castrated male sheep. In a cross-over design (n=6), responses to a single i.c.v. (third ventricle) injection of leptin (0.5, 1.0 and 1.5 mg ovine leptin (oLEP) and 1.0 mg murine leptin (mLEP)), N-methyl-D-aspartate (NMDA, 20 micro g) or 0.9% saline (control) were measured in terms of LH secretion (4 h post-injection compared with 4 h pre-injection) and appetite (during 2 h post-injection) in autumn (Experiment 1). NMDA and 1.0 mg oLEP treatments were repeated in the same sheep in the following spring (Experiment 2). With an additional 12 sheep (n=18 in cross-over design), responses to low-dose 'physiological' i.c.v. infusion of leptin (8 ng/h for 12 h daily for 4 days), insulin (0.7 ng/h) and artificial cerebrospinal fluid were measured in the next spring (Experiment 3). LH was studied over 8 h and appetite over 1 h on days 1 and 4 of infusion. In Experiment 1 (autumn), oLEP overall increased LH pulse frequency by up to 110% (P<0.05), decreased LH pulse amplitude (P<0.05) and decreased appetite (P<0.05). mLEP reduced LH pulse amplitude (P<0.05) without significant effect on appetite, while NMDA reduced appetite (P<0.05) but had no effect on LH. In Experiment 2 (spring), LH responses were 'surge-like' with highly significant increases in the moving average LH concentration after 1.0 mg oLEP (P<0.001) and after NMDA (P<0.001). Compared with similar analysis of experiment 1 results, the LH response in spring was greater than that in autumn for both 1.0 mg oLEP (P<0.05) and NMDA (P<0.005). Conversely, unlike in autumn (Experiment 1), there was no effect of 1.0 mg oLEP or NMDA on appetite in the spring (Experiment 2). In Experiment 3 (spring), 'physiological' i.c.v. infusion of oLEP or insulin increased LH pulse frequency by up to 100% (P<0.001) compared with the control infusion on both days 1 and 4, but there were no effects on appetite. These results indicate that intracerebral leptin both stimulates reproductive neuroendocrine output and decreases appetite in adequately nourished sheep. However, the responses of these two axes were dose-dependent and differentially affected by the time of year, suggesting dissociation of the neural pathways involved.
Endocrinology trial design: adverse event reporting in randomised controlled trials of recombinant human GH in GH-deficient adults
Authors: Bryant J , Loveman E , Cave C , Chase D , Milne R
Published: December 2004

We have evaluated the reporting of withdrawals due to adverse effects and specific adverse effects in randomised controlled trials of recombinant human GH in adults. A systematic review was carried out of randomised controlled trials of the clinical effectiveness of recombinant human GH in adults with GH deficiency in relation to impact on quality of life. Trials were identified from searching electronic databases, bibliographies of related articles and consulting experts. There was reporting of withdrawals due to adverse effects and specific adverse effects. Rates of oedema and arthralgia were reported in included trials. Seventeen randomised controlled trials, published between 1990 and 1999, met the inclusion criteria for the review. Nine trials reported data on the effectiveness of GH on quality of life in adults. Only five trials (29%) reported both withdrawals from the study because of adverse events and specific adverse events with numbers per study arm and per type. Six further trials (35%) reported either withdrawal details or specific adverse event details or partial data on specific adverse events. Six trials (35%), however, did not report information on either withdrawals or specific adverse events. Ten of the 17 studies (59%) reported the number of patients who withdrew from the study due to adverse events per study arm and type of adverse event per study arm. Seven of the 17 trials (41%) reported the number of specific adverse events per study arm and six (35%) reported the type per study arm. The reporting of adverse events in randomised controlled trials of GH is variable and not consistent across trials. It is not possible to assess the impact that adverse events may have had on unblinding patients, and therefore the extent to which the effects of GH may have been overestimated. Therefore those conducting endocrinology trials in the future need to pay attention to the reporting of withdrawals due to adverse events and specific adverse events.
Influence of maternal hyperthyroidism in the rat on the expression of neuronal and astrocytic cytoskeletal proteins in fetal brain
Authors: Evans IM , Pickard MR , Sinha AK , Leonard AJ , Sampson DC , Ekins RP
Published: December 2004

Maternal hypothyroidism during pregnancy impairs brain function in human and rat offspring, but little is known regarding the influence of maternal hyperthyroidism on neurodevelopment. We have previously shown that the expression of neuronal and glial differentiation markers in fetal brain is compromised in hypothyroid rat dam pregnancies and have now therefore extended this investigation to hyperthyroid rat dams. Study groups comprised partially thyroidectomised dams, implanted with osmotic pumps infusing either vehicle (TX dams) or a supraphysiological dose of thyroxine (T4) (HYPER dams), and euthyroid dams infused with vehicle (N dams). Cytoskeletal protein abundance was determined in fetal brain at 21 days of gestation by immunoblot analysis. Relative to N dams, circulating total T4 levels were reduced to around one-third in TX dams but were doubled in HYPER dams. Fetal brain weight was increased in HYPER dams, whereas litter size and fetal body weight were reduced in TX dams. Glial fibrillary acidic protein expression was similar in HYPER and TX dams, being reduced in both cases relative to N dams. alpha-Internexin (INX) abundance was reduced in HYPER dams and increased in TX dams, whereas neurofilament 68 (NF68) exhibited increased abundance in HYPER dams. Furthermore, INX was inversely related to - and NF68 directly related to - maternal serum total T4 levels, independently of fetal brain weight. In conclusion, maternal hyperthyroidism compromises the expression of neuronal cytoskeletal proteins in late fetal brain, suggestive of a pattern of accelerated neuronal differentiation.
Distribution and characterization of plasmalemma vesicle protein-1 in rat endocrine glands
Authors: Hnasko R , McFarland M , Ben-Jonathan N
Published: December 2004

Plasmalemma vesicle protein-1 (PV-1) is an integral membrane protein associated with endothelial cell caveolae and fenestrae. Since endocrine glands are enriched with fenestrated endothelium, we examined the distribution of PV-1 mRNA and protein in endocrine glands and determined its cellular localization. A single transcript was detected by RT-PCR in all endocrine glands examined. A synthetic peptide was used to generate antibodies for Western blotting and immunohistochemistry (IHC). Western blotting of membrane fractions from lung, pituitary, adrenal, testis and PV-1-transfected Cos-1 cells revealed a major 65 kDa protein. This protein binds to heparin with high affinity. Using IHC, PV-1 was localized to both endothelial cells of the adrenal zona reticularis and chromaffin cells of the medulla. In the pancreas, PV-1 expression was restricted to a few cells in the islets of Langerhans that partially overlap with somatostatin-positive delta-cells. In both neonatal and adult pituitaries, strong PV-1 immunoreactivity was detected in neural lobe pituicytes in a pattern similar to that of glial fibrillary acidic protein (GFAP). PV-1 and GFAP expression was seen in the adult, but not neonatal, intermediate lobe. Endothelial cells throughout the neonatal anterior lobe were PV-1 positive, but PV-1 in the adult was restricted to some endothelial and endocrine cells localized near the margins of lobe. In the adult testis, strong PV-1 expression was seen in germ cells within the seminiferous tubules that varied with the stage of spermatogenesis. In contrast, PV-1 in the neonatal testis was localized to the interstitial cells but not seminiferous tubules. In the ovary, PV-1 was expressed in stromal endothelial cells as well as the thecal layer of developing follicles. Over half the corpus luteal cells were positive for PV-1. Our data have shown that PV-1 is not restricted to endothelial cells but is localized in many types of endocrine and non-endocrine cells. Furthermore, PV-1 expression in the pituitary and testis is developmentally regulated.
Glucocorticoid effects on chondrogenesis, differentiation and apoptosis in the murine ATDC5 chondrocyte cell line
Authors: Mushtaq T , Farquharson C , Seawright E , Ahmed SF
Published: December 2004

Glucocorticoids (GC) are used extensively in children and may cause growth retardation, which is in part due to the direct effects of GC on the growth plate. We characterised the ATDC5 chondrocyte cell line, which mimics the in vivo process of longitudinal bone growth, to examine the effects of dexamethasone (Dex) and prednisolone (Pred) during two key time points in the chondrocyte life cycle - chondrogenesis and terminal differentiation. Additionally, we studied the potential for recovery following Dex exposure. During chondrogenesis, Dex and Pred exposure at 10(-8) M, 10(-7) M and 10(-6) M resulted in a significant mean reduction in cell number (28% vs 20%), cell proliferation (27% vs 24%) and proteoglycan synthesis (47% vs 43%) and increased alkaline phosphatase (ALP) activity (106% vs 62%), whereas the incidence of apoptosis was unaltered. Minimal effects were noted during terminal differentiation with both GC although all concentrations of Dex lowered apoptotic cell number. To assess catch-up growth the cells were incubated for a total of 14 days which included 1, 3, 7, 10 or 14 days exposure to 10(-6) M Dex, prior to the recovery period. Recovery of proteoglycan synthesis was irreversibly impaired following just one day exposure to Dex. Although cell number showed a similar pattern, significant impairment was only achieved following 14 days exposure. Irreversible changes in ALP activity were only noticed following 10 days exposure to Dex. In conclusion, GC have maximal effects during chondrogenesis; Dex is more potent than Pred and cells exposed to Dex recover but this may be restricted due to differential effects of GC on specific chondrocyte phenotypes.
Leptin controls the fate of fatty acids in isolated rat white adipocytes
Authors:   William WN, Ceddia RB , Curi R
Published: December 2004

Leptin directly increases the rate of exogenous glucose and fatty acids oxidation in isolated adipocytes. However, the effects of leptin on fatty acid metabolism in white adipose tIssue have not been examined in detail. Here, we report that in adipocytes incubated for 6 h in the presence of leptin (10 ng/ml), the insulin-stimulated de novo fatty acid synthesis was inhibited by 36% (P<0.05), while the exogenous oxidation of acetic and oleic acids was increased by 50% and 76% respectively. Interestingly, leptin did not alter the oxidation of intracellular fatty acids. Leptin-incubated cells presented a 16-fold increase in the incorporation of oleic acid into triglyceride (TG) and a 123% increase in the intracellular TG hydrolysis (as measured by free fatty acids release). Fatty acid-TG cycling was not affected by leptin. By employing fatty acids radiolabeled with (3)H and (14)C, we could determine the concomitant influx of fatty acids (incorporation of fatty acids into TG) and efflux of fatty acids (intracellular fatty acids oxidation and free fatty acids release) in the incubated cells. Leptin increased by 30% the net efflux of fatty acids from adipocytes. We conclude that leptin directly inhibits de novo synthesis of fatty acids and increases the release and oxidation of fatty acids in isolated rat adipocytes. These direct energy-dissipating effects of leptin may play an important role in reducing accumulation of fatty acids into TG of rat adipose cells.
Tumor necrosis factor-alpha inhibits glutathione S-transferase-alpha expression in cultured porcine Sertoli cells
Authors: Benbrahim-Tallaa L , Boussouar F , Rey C , Benahmed M
Published: December 2004

Glutathione S-transferases (GSTs) are a family of soluble enzymes of detoxification that use reduced glutathione in conjugation and reduction reactions. Toxic electrophiles are substrates for the GSTs. GSTalpha is expressed at high levels in different tIssues such as the testis. Among the different GSTs present in the testis, GSTalpha is specifically expressed in Leydig and Sertoli cells known to be under the control of hormonal and local regulatory factors. The present study investigated the regulatory action of tumor necrosis factor-alpha (TNFalpha) on basal and hormone (FSH and testosterone)-stimulated GSTalpha expression in cultured Sertoli cells. Treatment with TNFalpha (0-20 ng/ml, 48 h) induced a decrease in basal GSTalpha mRNA levels in a dose-dependent manner (fivefold decrease; P<0.001). The maximal and half maximal effects were observed at 20 ng/ml and 7 ng/ml respectively. The inhibitory effect of TNFalpha was also time-dependent with a maximal inhibitory effect (threefold decrease; P<0.001) observed at 48 h. The inhibitory effect of the cytokine was also observed on basal GSTalpha protein (28 kDa) levels. TNFalpha also inhibited the hormone-stimulated GSTalpha expression in Sertoli cells. The treatment of cultured Sertoli cells with both FSH and TNFalpha (100 ng/ml and 10 ng/ml respectively, 48 h) resulted in a complete suppression of the stimulatory action of FSH on GSTalpha mRNA levels. Similarly, in Sertoli cells treated with testosterone or its non-aromatizable metabolite dihydrotestosterone (100 ng/ml, 24 h), TNFalpha reduced the hormone-stimulated GSTalpha mRNA and protein levels. TNFalpha inhibited basal GSTalpha expression without affecting mRNA stability. Indeed, the decay curves (mRNA half-life time=18 h) for the GSTalpha basal mRNA levels in Sertoli cells was similar in the absence or presence of TNFalpha (10 ng/ml, 48 h). Testosterone increased GSTalpha mRNA without affecting the enzyme mRNA stability. TNFalpha antagonized the androgen-stimulated GSTalpha mRNA levels without affecting the enzyme mRNA stability, suggesting that the interaction between the androgen and the cytokine is mostly exerted at a transcriptional level. FSH increased GSTalpha mRNA levels through an increase in mRNA stability (increased mRNA half-life times to 119 h). TNFalpha antagonized the stimulatory effect of FSH on GSTalpha mRNA levels by antagonizing the stabilizing effect exerted by the hormone on GSTalpha mRNA. Together, these results suggest that the increase in the cytokine levels within the testis would alter the detoxification processes against genotoxic products during spermatogenesis.
Plasma ghrelin concentrations in elderly subjects: comparison with anorexic and obese patients
Authors: Rigamonti AE , Pincelli AI , Corra B , Viarengo R , Bonomo SM , Galimberti D , Scacchi M , Scarpini E , Cavagnini F , Muller EE
Published: December 2004

Ghrelin, a novel endogenous ligand for the GH secretagogue receptor, has been reported to stimulate GH secretion and food intake in both humans and other animals. Interestingly, recent data indicate that ghrelin is up- and down-regulated in anorexia nervosa (AN) and obesity, which are also known to be accompanied by increased and reduced GH levels respectively. Ageing is associated with a gradual but progressive reduction in GH secretion, and by alterations in appetite and food intake. The role of ghrelin in the decline of somatotroph function and the anorexia of ageing is unknown. To investigate the influence of age on circulating levels of ghrelin, a total of 19 young and old normal weight subjects (Y-NW, n=12; O-NW, n=7), six patients with active AN (A-AN), and seven patients with morbid obesity (OB) were studied. In addition to fasting plasma ghrelin concentrations, baseline serum TSH, IGF-I and insulin levels were measured. Mean plasma ghrelin concentrations in A-AN or OB were higher and lower respectively than those present in Y-NW. Interestingly, mean plasma ghrelin concentrations in O-NW were significantly lower than those present in Y-NW and superimposable on those of OB. The mean fasting plasma ghrelin concentrations in all groups of subjects were negatively correlated with body mass index and serum insulin levels, but not with TSH and IGF-I levels. This study provides evidence of an age-related decline of plasma ghrelin concentrations, which might explain, at least partially, the somatotroph dysregulation and the anorexia of the elderly subject.
Circulating ghrelin concentrations are lowered by intravenous glucose or hyperinsulinemic euglycemic conditions in rodents
Authors: McCowen KC , Maykel JA , Bistrian BR , Ling PR
Published: December 2004

Ghrelin is a peptide secreted mainly by gastric parietal cells that may play a role in appetite regulation. Circulating ghrelin is abruptly lowered by food intake, but factors involved in ghrelin regulation remain unclear. The aim of this study was to determine whether intravenous glucose infusion lowers ghrelin, and to determine whether glucose, insulin or some measure of insulin action best predicts the effect of feeding on ghrelin. Rats were infused over 3 h with either A. saline (controls); B. dextrose to steady state blood glucose approximately 16.7 mM, or C. insulin 7.5 mU/kg x min, plus dextrose as needed to clamp to euglycemic basal concentrations. During 3 h of infusion, group B had significantly greater (P<0.01) glucose, 17.4+/-0.3 mM, than groups A (6.6+/-0.3) or C (6.1+/- 0.2). Groups B and C had hyperinsulinemia at the end of the 3 h infusion (894+/-246, 804+/-156 pM) compared with saline-infused (222+/-24 pM, P<0.01). Ghrelin concentrations were reduced (P<0.01) in both hyperinsulinemic groups (B=85+/-2; C=103+/-0.6 pM) versus controls (163+/-9). Ghrelin was strongly correlated with insulin (r=-0.68), glucose infusion rate (r=-0.75) and free fatty acids (r=0.67), when all 3 groups were combined, although only the 2 latter variables were independent predictors of ghrelin. In conclusion, neither a rise in blood glucose nor presence of nutrient in the stomach is required for the effect of feeding on ghrelin. The data suggest that whole body insulin responsiveness plays either a direct or indirect role in meal-related ghrelin inhibition.
Reductions in adipose tissue and skeletal growth in rat adult offspring after prenatal leptin exposure
Authors: Nilsson C , Swolin-Eide D , Ohlsson C , Eriksson E , Ho HP , Bjorntorp P , Holmang A
Published: December 2004

Leptin is involved in regulating food intake, energy balance and bone formation. Increasing evidence suggests that leptin is also involved in fetal growth and development. The aim of this study was to determine if increased maternal leptin is followed by changes in body composition, skeletal growth or hormonal regulation in the adult rat offspring. Pregnant rats were given injections of either human recombinant leptin (3.5 mg/kg, i.p.) or vehicle on days 8, 10 and 12 of gestation. Both genders of leptin-exposed offspring showed significantly reduced adipose tIssue weight at adult age. Skeletal growth and cortical bone dimensions were significantly reduced. Circulating testosterone levels were significantly increased in female leptin-exposed offspring, and male leptin-exposed offspring had significant testicular enlargement. No significant effects were seen on circulating leptin levels or hypothalamic protein levels of the leptin receptor. The results demonstrate that maternally administered leptin is involved in fetal growth and development, leading to lean offspring with reduced skeletal growth.
Hypothyroidism alters the expression of Bcl-2 family genes to induce enhanced apoptosis in the developing cerebellum
Authors: Singh R , Upadhyay G , Kumar S , Kapoor A , Kumar A , Tiwari M , Godbole MM
Published: December 2004

Thyroid hormone (TH) deficiency results in delayed proliferation and migration of cerebellar granule cells. Although extensive cell loss during the development of the cerebellum under hypothyroid conditions is known, its nature and its mechanism are poorly understood. Bcl-2 family gene expression is known to determine the fate of cells to undergo apoptosis. We evaluated the effect of hypothyroidism on Bcl-2 family gene expression in the developing rat cerebellum. Electrophoresis and Western blotting were used to analyze DNA fragmentation and expression of DNA fragmentation factor (DFF-45), Bcl-2, Bcl-xL and Bax genes respectively. In the hypothyroid condition, extensive DNA fragmentation and enhanced cleavage of DFF-45 were seen throughout development (postnatal day 0 to day 24) and adulthood whereas they were absent in the euthyroid state. The anti-apoptotic genes Bcl-2 and Bcl-xL were down-regulated and the pro-apoptotic gene Bax was expressed at higher levels compared with the euthyroid state. These results suggest that normal levels of TH prevent cerebellar apoptosis to a large extent, whereas hypothyroidism not only increases the extent but also the duration of apoptosis by down-regulating the anti-apoptotic genes and maintaining a high level of the pro-apoptotic gene Bax.
Improved biological activity of Gly2- and Ser2-substituted analogues of glucose-dependent insulinotrophic polypeptide
Authors: Gault VA , Flatt PR , Harriott P , Mooney MH , Bailey CJ , O'Harte FP
Published: December 2004

The therapeutic potential of glucagon-like peptide-1 (GLP-1) in improving glycaemic control in diabetes has been widely studied, but the potential beneficial effects of glucose-dependent insulinotropic polypeptide (GIP) have until recently been almost overlooked. One of the major problems, however, in exploiting either GIP or GLP-1 as potential therapeutic agents is their short duration of action, due to enzymatic degradation in vivo by dipeptidylpeptidase IV (DPP IV). Therefore, this study examined the plasma stability, biological activity and antidiabetic potential of two novel NH2-terminal Ala2-substituted analogues of GIP, containing glycine (Gly) or serine (Ser). Following incubation in plasma, (Ser2)GIP had a reduced hydrolysis rate compared with native GIP, while (Gly2)GIP was completely stable. In Chinese hamster lung fibroblasts stably transfected with the human GIP receptor, GIP, (Gly2)GIP and (Ser2)GIP stimulated cAMP production with EC(50) values of 18.2, 14.9 and 15.0 nM respectively. In the pancreatic BRIN-BD11 beta-cell line, (Gly2)GIP and (Ser2)GIP (10(-8) M) evoked significant increases (1.2- and 1.5-fold respectively; P<0.01 to P<0.001) in insulinotropic activity compared with GIP. In obese diabetic ob/ob mice, both analogues significantly lowered (P<0.001) the glycaemic excursion in response to i.p. glucose. This enhanced glucose-lowering ability was coupled to a significantly raised (P<0.01) and more protracted insulin response compared with GIP. These data indicate that substitution of the penultimate Ala2 in GIP by Gly or Ser confers resistance to plasma DPP IV degradation, resulting in enhanced biological activity, therefore raising the possibility of their use in the treatment of type 2 diabetes.
Effect of Lepidium meyenii (Maca), a root with aphrodisiac and fertility-enhancing properties, on serum reproductive hormone levels in adult healthy men
Authors: Gonzales GF , Cordova A , Vega K , Chung A , Villena A , Gonez C
Published: December 2004

Lepidium meyenii (Maca) is a Peruvian hypocotyl that grows exclusively between 4000 and 4500 m in the central Andes. Maca is traditionally employed in the Andean region for its supposed aphrodisiac and/or fertility-enhancing properties. This study was a 12-week double-blind, placebo-controlled, randomized, parallel trial in which active treatment with different doses of Maca Gelatinizada was compared with a placebo. The study aimed to test the hypothesis that Maca has no effect on serum reproductive hormone levels in apparently healthy men when administered in doses used for aphrodisiac and/or fertility-enhancing properties. Men aged between 21 and 56 Years received 1500 mg or 3000 mg Maca. Serum levels of luteinizing hormone, follicle-stimulating hormone, prolactin, 17-alpha hydroxyprogesterone, testosterone and 17-beta estradiol were measured before and at 2, 4, 8 and 12 weeks of treatment with placebo or Maca (1.5 g or 3.0 g per day). Data showed that compared with placebo Maca had no effect on any of the hormones studied nor did the hormones show any changes over time. Multiple regression analysis showed that serum testosterone levels were not affected by treatment with Maca at any of the times studied (P, not significant). In conclusion, treatment with Maca does not affect serum reproductive hormone levels.