bluuebunny/crossref_metadata_2025 huggingface dataset:
Dexamethasone blocks antioestrogen- and oxidant-induced death of pituitary tumour cells
Authors: Newton CJ , Bilko D , Pappa S , Atkin SL
Published: December 2004

The oestrogen receptor is fundamental to the growth and survival of the rat pituitary tumour cell line, GH(3). Our previous studies have shown that antioestrogens such as RU 58668 and ZM 182780 will reduce the rate of cell division and also induce cell death. Death of these cells in response to antioestrogen treatment appears to be due to a heightened sensitivity to reactive oxygen species (ROS). As part of a study to determine the cross-talk between steroid receptor systems in these cells, we have observed that the glucocorticoid, dexamethasone (Dex), inhibits antioestrogen-induced cell death. Cell death induced by H(2)O(2) is enhanced by ZM 182780 and this effect is also blocked by Dex. As apoptotic cell death in a number of systems involves an early loss of mitochondrial membrane potential (DeltaPsi(m)), we have performed detailed studies on the time-course of DeltaPsi(m) loss in relation to the loss in cell membrane function. These studies have indicated that a loss of DeltaPsi(m) parallels a loss of cell membrane function - this is more characteristic of necrosis than of apoptosis. From microscopic observations of these cells in response to H(2)O(2), it has been noted that early cell membrane blebbing, induced by H(2)O(2), is blocked in the presence of ZM 182780. Cell membrane blebbing can precede necrosis as well as apoptosis and it is thought to involve cytoskeletal changes, for which localised glycolytic reactions provide ATP. These observations, together with those showing that removal of glucose, but not inhibition of mitochondrial function, enhances ROS-induced cell death, prompted studies on the glycolytic pathway. As a strong candidate mechanism, it would appear that, via an effect on one of the rate-limiting glycolytic enzymes, glyceraldehyde-3-phosphate dehydrogenase, Dex is able to overcome the antioestrogen-enhanced loss of glycolytic function following exposure of cells to ROS. This report contributes to the growing body of evidence showing that glucocorticoids provide a survival advantage to both normal and tumour cell types.
Anterior pituitary trophic responses to dexamethasone withdrawal and repeated dexamethasone exposures
Authors: Nolan LA , Levy A
Published: December 2004

Glucocorticoid withdrawal, depending on the dose and duration of treatment, results in a transient but sometimes prolonged reduction in hypothalamo-pituitary-adrenal (HPA) axis secretory responsiveness. As the anatomic basis of HPA axis suppression remains uncertain, we have directly examined changes in trophic activity within the rat anterior pituitary gland following dexamethasone withdrawal and re-treatment. Treatment of adrenalectomised, male Wistar rats with dexamethasone results in a discrete, highly significant burst of apoptosis in the anterior pituitary with concurrent suppression of mitosis. Despite a surge in mitotic activity immediately after dexamethasone withdrawal, calculated total anterior pituitary cell populations remain below that seen in untreated adrenalectomised controls. Repeated exposures to dexamethasone show that the dexamethasone-sensitive cell population that is deleted by apoptosis is partially but not completely restored. As the amplitude of apoptotic bursts induced by second and third dexamethasone exposures are similar but smaller than that induced by initial exposure, it appears that the very first exposure to dexamethasone deletes a subset of anterior pituitary cells that are either not restored at all, or are only replaced very slowly. The reduced proportion of corticotrophs contributing to the increase in mitotic index after dexamethasone withdrawal corroborates this. Although continued cell turnover within the pituitary predicts that the absolute cellular deficit would diminish with time, the effects seen may contribute to the delayed recovery of pituitary axis function following cessation of glucocorticoid treatment.
Progesterone and testosterone in combination act in the hypothalamus of castrated rams to regulate the secretion of LH
Authors: Turner AI , Tilbrook AJ , Clarke IJ , Scott CJ
Published: December 2004

We tested the hypotheses that progesterone enhances the negative feedback actions of testosterone in rams and that this occurs through actions at the hypothalamus. In the first part of this study, blood samples were collected every 10 min for 12 h before and after 7 days of treatment (i.m.) of castrated Romney Marsh rams (n=5 per group) with vehicle, progesterone (4 mg/12 h), testosterone (4 mg/12 h) or a combination of progesterone (4 mg/12 h) and testosterone (4 mg/12 h). In the second part of this study the brains of four gonad-intact Romney Marsh rams were collected, the hypothalamus was sectioned and in situ hybridisation of mRNA for progesterone receptors conducted. After 7 days of treatment with vehicle or progesterone or testosterone alone, there were no changes in the secretion of LH. In contrast, treatment with a combination of progesterone and testosterone resulted in a significant (P<0.01, repeated measures ANOVA) decrease in mean plasma concentrations of LH, the number of LH pulses per hour and the pre-LH pulse nadir and a significant (P<0.01) increase in the inter-LH pulse interval. We found cells containing mRNA for progesterone receptors throughout the hypothalamus, including the preoptic area (where most GnRH neurons are located in sheep), the periventricular, ventromedial and arcuate nuclei and the bed nucleus of the stria terminalis. This study shows that progesterone is capable of acting centrally with testosterone to suppress the secretion of LH in castrated rams and that cells containing mRNA for progesterone receptors are located in the hypothalamus of rams in the vicinity of GnRH neurons.
Contribution of progesterone, follicle stimulating hormone and glucocorticoids in survival of serum-free cultured granulosa cell explants
Authors: Mussche S , D'Herde K
Published: December 2004

To investigate the role of progesterone (P4) as a survival factor in quail granulosa cell explants, P4 content was determined under various conditions and correlated with apoptotic indexes (AIs) evaluated by 2',6'-diamidino-2-phenylindole (DAPI)-staining. Analysis of serum-free cultures from 24 to 96 h shows decreased P4 levels in the medium paralleled by increasing AI. Inhibiting apoptosis by gonadotropic support (FSH, 100 ng/ml) stimulates a 3-fold increase of the P4 level in the medium (83.49+/-8.69 vs 26.31+/-1.61 ng/ml in serum-free controls) together with a significant decrease in AI from 8.81+/-1.06% in serum-free controls to 3.50+/-0.72%. Substantial evidence for P4 as an autocrine/paracrine survival factor can be inferred from experiments with aminoglutethimide (AG, 1 mM) and RU486 (20 microM). Blocking P4 synthesis by AG causes a 2-fold increase in apoptosis from 6.08+/-0.67% in serum-free controls to 12.53+/-1.60%. Blocking P4 receptors by RU486 causes a similar increase in AI (3.02+/-0.98% in serum-free controls to 17.07+/-3.20%) and about a 50% decrease in P4. The effect of RU486 could be attenuated by exogenous P4 but not by dexamethasone indicating selective binding of P4 to the progesterone receptor. Dexamethasone treatment promotes survival without affecting P4 levels. In further support of an autocrine/paracrine action for P4 in the granulosa cells, both the A and B form of the avian P4 receptor (PR) are identified in vivo and in vitro by Western blotting. Exogenous administration of P4 only affects survival when endogenous P4 synthesis is blocked or after 48 h of serum-free culture when endogenous P4 production is very low. Because FSH also affects survival when its stimulatory effect on P4 synthesis is blocked by AG (AI decrease from 6.08+/-0.67% in serum-free controls to 1.64+/-0.71% in FSH+AG treated) it is proposed that (1) P4 is an autocrine/paracrine survival factor in the preovulatory granulosa and (2) FSH mediates both P4-dependent and P4-independent survival pathways.
Autoregulation of the gonadotropin-releasing hormone (GnRH) system during puberty: effects of antagonistic versus agonistic GnRH analogs in a female rat model
Authors: Roth C , Schricker M , Lakomek M , Strege A , Heiden I , Luft H , Munzel U , Wuttke W , Jarry H
Published: December 2004

To address whether gonadotropin-releasing hormone (GnRH) regulates its own expression and the expression of its receptor in the hypothalamus and ovary, we treated five groups of prepubertal/peripubertal female rats from postnatal days 25-36 with either the GnRH agonist triptorelin (TRIP) or the GnRH antagonist cetrorelix (CET), each 10 or 100 microgram/day, or a placebo. We compared their effects regarding pubertal development, serum gonadotropins and the expression of GnRH and GnRH-receptor in the hypothalamus, pituitary, ovary and uterus. Onset of puberty was determined by vaginal opening, and expression levels of GnRH and GnRH-receptor were determined using either quantitative real-time PCR or competitive RT-PCR. Onset of puberty was retarded by both analogs but CET (100 microgram/day) inhibited while TRIP (10 and 100 microgram/day) stimulated serum gonadotropins (P<0.05). The expression of GnRH in the preoptic area did not show significant differences among the treatment groups but ovarian GnRH mRNA levels were significantly stimulated by CET (100 microgram/day). GnRH mRNA could not be detected in the uterus by either real-time PCR or competetive RT-PCR. The GnRH-receptor expression in the hypothalamus (preoptic area and mediobasal hypothalamus) did not vary among any of the groups, whereas in the pituitary GnRH-receptor mRNA levels were stimulated by TRIP (10 microgram/day) but inhibited by CET (100 microgram/day). In contrast, in the ovary GnRH-receptor mRNA levels were inhibited by both TRIP (100 microgram/day) and CET (100 microgram/day). Interestingly, the GnRH-receptor was even expressed in the uterus where it was strongly stimulated by both CET and TRIP in a dose-related manner. This shows that in addition to their different pituitary effects, the GnRH analogs cetrorelix and triptorelin exert different actions at the hypothalamic, ovarian and uterine level. This study also demonstrates an organ-specific regulation of GnRH and GnRH-receptor gene expression which is likely part of a local autoregulatory system. We conclude that the ovarian and uterine effects of GnRH analogs must be considered in addition to their known pituitary effects when deciding which GnRH analog is most suitable for treating precocious puberty.
Differential expression of suppressors of cytokine signalling genes in response to nutrition and growth hormone in the septic rat
Authors: Johnson TS , O'Leary M , Justice SK , Maamra M , Zarkesh-Esfahani SH , Furlanetto R , Preedy VR , Hinds CJ , El Nahas AM , Ross RJ
Published: December 2004

GH treatment during critical illness and sepsis may increase mortality. A family of negative regulators of cytokine signalling, the suppressors of cytokine signalling (SOCS), have been characterised. SOCS provide a mechanism for cross-talk between the cytokine receptors, including GH. Here, we have investigated the impact of nutrition and GH treatment on GH receptor, SOCS1, SOCS-2, SOCS-3 and cytokine-inducible SH2-containing protein (CIS) hepatic mRNA expression in a rat model of sepsis, caecal ligation and puncture (CLP). Four groups of rats were studied: control (food given ad libitum, n=7), CLP only (n=8), CLP and total parenteral nutrition (TPN) (n=9), and CLP, TPN and GH (n=10). CLP rats underwent surgery and 18 h later received saline or TPN or TPN+GH for 6 h before they were killed. Serum IGF-I levels were lower in all CLP groups (P<0.001). The combination of TPN and GH treatment increased IGF-I levels compared with the saline-treated CLP rats (P<0.01), but IGF-I levels remained lower than control animals (P<0.001). GH receptor and GH-binding protein expression in liver was reduced in animals subjected to CLP and was unaffected by nutrition or GH treatment. Hepatic SOCS-1 was detectable in normal rats, induced in all CLP animals but was unaffected by nutrition and GH. Hepatic SOCS-2 expression was difficult to detect in normal and CLP rats but was greatly induced in CLP rats treated with GH. Hepatic SOCS-3 expression was only just detectable in the control group but was elevated in all CLP groups and unaffected by nutrition and GH. Hepatic CIS expression was difficult to detect in normal rats, was not induced by CLP but was induced by both nutrition and GH. In conclusion, CLP induced low IGF-I levels associated with increased expression of SOCS-1 and SOCS-3, both of which are known to inhibit GH receptor signalling. GH induced SOCS-2 and CIS in the CLP rat despite resistance with respect to IGF-I generation, and parenteral feeding induced CIS in the CLP rat. Thus, there is potential for a complex interaction between GH and cytokine signalling at the level of SOCS expression whereby the inflammatory response may alter GH signalling and GH may influence the inflammatory response.
Effects of the gp130 cytokines ciliary neurotropic factor (CNTF) and interleukin-11 on pituitary cells: CNTF receptors on human pituitary adenomas and stimulation of prolactin and GH secretion in normal rat anterior pituitary aggregate cultures
Authors: Perez Castro C , Carbia Nagashima A , Paez Pereda M , Goldberg V , Chervin A , Carrizo G , Molina H , Renner U , Stalla GK , Arzt E
Published: December 2004

Two of the most potent cytokines regulating anterior pituitary cell function are leukemia inhibitory factor (LIF) and interleukin (IL)-6, which belong to the cytokine family using the common gp130 signal transducer. Recently, the expression and action of two other members of this family, IL-11 and ciliary neurotrophic factor (CNTF), on different cell lines has also been demonstrated. We studied the expression of the specific receptor subunits for CNTF in mammotropic, non-functioning and somatotropic tumors and the action of CNTF and IL-11 in the regulation of hormone secretion in these and normal pituitary cells. The mRNA for the alpha chain specific for the CNTF receptor was detected by Northern blot in tumors secreting prolactin (PRL) and GH and in non-functioning tumors. We found that both IL-11 and CNTF exerted a similar stimulatory effect on GH mRNA expression in somatotropic monolayer cell cultures from acromegalic tumors, but these cytokines had no significant influence on GH secretion. CNTF stimulates prolactin secretion in lactotropic monolayer cell cultures from patients with prolactinoma. In monolayer cell cultures from normal rat anterior pituitary, IL-11 and CNTF had no significant effect on the release of either GH or PRL, or on GH mRNA. However, when the cells were cultured in aggregate cultures, in which the three-dimensional structure of the cells is reconstituted, both cytokines, in doses at which they had no effect on monolayer cultures, significantly stimulated both PRL and GH secretion. These data show that IL-11 and CNTF may act as regulatory factors in anterior pituitary cells, in which the three-dimensional structure of the gland is of critical importance.
Mitogenic activity of high molecular weight forms of insulin-like growth factor-II in amniotic fluid
Authors: Blahovec J , Kostecka Z , Lacroix MC , Cabanie L , Godeau F , Mester J , Cavaille F
Published: December 2004

Amniotic fluid (AF) collected from ewes and goats at mid gestation displayed mitogenic activity in mouse fibroblasts. Upon fractionation of this material by size exclusion chromatography, the mitogenic activity was resolved into two peaks, whose activity was inhibited by an anti-IGF type 1 receptor blocking antibody. One of the peaks contained IGF-I and IGF-II (mature form), whereas the other contained high M(r) precursor forms of IGF-II. The presence in this latter fraction of IGF-binding proteins (IGFBP) suggests that the AF IGFBPs do not efficiently inhibit the mitogenic activity of the high M(r) forms of IGF-II. In agreement with this conclusion, exogenous IGFBP-1 failed to affect this activity. Analysis of IGF-II in sheep AF showed that the AF concentrations of both forms of IGF-II increased dramatically from mid pregnancy until 106-120 days of gestation, and fell thereafter. The amniotic IGFBPs followed a similar evolution. High M(r) forms of IGF-II were also found in human AF, with a pattern of electrophoretic migration different from that of sheep. We suggest that the precursor forms of IGF-II may play an important role in foetal development.
Extracts from tumors causing oncogenic osteomalacia inhibit phosphate uptake in opossum kidney cells
Authors: Jonsson KB , Mannstadt M , Miyauchi A , Yang IM , Stein G , Ljunggren O , Juppner H
Published: December 2004

In oncogenic osteomalacia (OOM), a tumor produces an unknown substance that inhibits phosphate reabsorption in the proximal tubules. This causes urinary phosphate wasting and, as a consequence, hypophosphatemic osteomalacia. To characterize this poorly understood biological tumor activity we generated aqueous extracts from several OOM tumors. Extracts from three of four tumors inhibited, dose- and time-dependently, (32)P-orthophosphate uptake by opossum kidney (OK) cells; maximum inhibition was about 45% of untreated control. Further characterization revealed that the factor is resistant to heat and several proteases, and that it has a low molecular weight. The tumor extracts also stimulated cAMP accumulation in OK cells, but not in osteoblastic ROS 17/2.8 and UMR106 cells, or in LLC-PK1 kidney cells expressing the parathyroid hormone (PTH)/PTH-related peptide receptor or the PTH-2 receptor. HPLC separation of low molecular weight fractions of the tumor extracts revealed that the flow-through of all three positive tumor extracts inhibited (32)P uptake and stimulated cAMP accumulation in OK cells. Additionally, a second peak with inhibitory activity on phosphate transport, but without cAMP stimulatory activity, was identified in the most potent tumor extract. We have concluded that several low molecular weight molecules with the ability to inhibit phosphate transport in OK cells can be found in extracts from OOM tumors. It remains uncertain, however, whether these are related to the long-sought phosphaturic factor responsible for the phosphate wasting seen in OOM patients.
The effects of intense exercise on the female reproductive system
Authors: Warren MP , Perlroth NE
Published: December 2004

Women have become increasingly physically active in recent decades. While exercise provides substantial health benefits, intensive exercise is also associated with a unique set of risks for the female athlete. Hypothalamic dysfunction associated with strenuous exercise, and the resulting disturbance of GnRH pulsatility, can result in delayed menarche and disruption of menstrual cyclicity. Specific mechanisms triggering reproductive dysfunction may vary across athletic disciplines. An energy drain incurred by women whose energy expenditure exceeds dietary energy intake appears to be the primary factor effecting GnRH suppression in athletes engaged in sports emphasizing leanness; nutritional restriction may be an important causal factor in the hypoestrogenism observed in these athletes. A distinct hormonal profile characterized by hyperandrogenism rather than hypoestrogenism is associated with athletes engaged in sports emphasizing strength over leanness. Complications associated with suppression of GnRH include infertility and compromised bone density. Failure to attain peak bone mass and bone loss predispose hypoestrogenic athletes to osteopenia and osteoporosis. Metabolic aberrations associated with nutritional insult may be the primary factors effecting low bone density in hypoestrogenic athletes, thus diagnosis should include careful screening for abnormal eating behavior. Increasing caloric intake to offset high energy demand may be sufficient to reverse menstrual dysfunction and stimulate bone accretion. Treatment with exogenous estrogen may help to curb further bone loss in the hypoestrogenic amenorrheic athlete, but may not be sufficient to stimulate bone growth. Treatment aimed at correcting metabolic abnormalities may in fact prove more effective than that aimed at correcting estrogen deficiencies.
Ethical aspects and the prevalence of hormone abuse in sport
Authors: Verroken M
Published: December 2004

Abuse of hormones by sportsmen or sportswomen might lead to some serious health consequences for the individual user. Such abuse also damages the very spirit of sport, cheating fellow athletes, the officials and spectators. Where hormone findings cause most controversy is with endogenously produced substances. Some hormone findings might be indicators of a medical condition, and it requires sensitive handling to discover the facts. Examining the prevalence of hormone abuse using a theoretical perspective on ethics permits a philosophical study of the dilemmas facing sport, and a clearer identification of the issues that science can help sport to resolve. This paper looks at the way in which rules on doping have evolved in an attempt to set out the ethical standards that should apply and to discuss how some sportsmen and sportswomen have worked around the rules, challenging them to the extent that the anti-doping system itself is questioned. The data emerging from the testing programmes give one guide to the actual prevalence of hormone findings. However, as not all findings may constitute doping offences this cannot be said to be the definitive guide to the extent of hormone abuse. Use of hormone medications with or without a therapeutical indication further complicates the disciplinary process. Sensitive management of hormone findings is absolutely necessary to avoid accusations of doping or embarrassing breaches of confidentiality when the origin may be a serious medical condition. Because hormone findings require careful consideration, the door is open for the anti-doping system to be exploited by unscrupulous scientists, raising challenges that test the limits of credibility. Sex, alcohol and decomposition are arguments that have been put forward to explain findings. A close partnership between scientists and sport is called for to avoid the athlete becoming a victim of the rules that are intended to protect sport.
Drugs in sport - the role of the physician
Authors: Dawson RT
Published: December 2004

Sportsmen have used anabolic steroids since the 1950s and yet it was not until the 1980s that we, as physicians, admitted that they could improve performance. We now find ourselves in the insidious position of being unable to predict convincingly either safety or major health risks with performance-enhancing drug use. The use of performance-enhancing drugs is no longer limited to the elite athlete. In 1993 the Canadian Center for Drug-free Sport estimated that 83 000 children between the ages of 11 and 18 had used anabolic steroids in the previous 12 months. Recent evidence suggests anabolic steroids are now the third most commonly offered drugs to children in the UK, behind cannabis and amphetamines. The role of the physician of today is to regain our position of impartiality and objectivity within both the sporting and general community. Only then will we be able to pursue a harm minimisation strategy designed to convince the public that it is better to be the best you can be naturally. For the majority, the improvement through the use of performance-enhancing drugs can equally be achieved through dietary and training advice. For the elite athlete, what price a gold medal that is tarnished by deceit? Its value then can only lie with the sponsors and politicians, for they can no longer claim to be sportsmen, only entertainers.
Desensitization of beta2-adrenoceptor function in non-pregnant rat myometrium is modulated by sex steroids
Authors: Engstrom T , Vilhardt H , Bratholm P , Christensen NJ
Published: December 2004

The effects of in vivo treatment with estrogen and progesterone on isoproterenol-induced uterine relaxation and beta(2)-adrenoceptor (beta(2)AR) mRNA production in non-pregnant rat myometrium were investigated. Whether homologous myometrial desensitization of beta(2)AR function was dependent on or modulated by the two steroids was also examined. Estrogen treatment alone or in combination with progesterone reduced maximal relaxation (E(max)) of isolated uterine strips subsequently challenged with isoproterenol whereas progesterone alone had no effect on this parameter. The reduction was accompanied by an enhanced beta(2)AR mRNA concentration. The concentration of isoproterenol giving half-maximal relaxing response (EC(50)) increased following estrogen treatment and this effect was curbed by progesterone. Isoproterenol had no effect on beta(2)AR transcription irrespective of the steroid regimes employed. E(max) of isolated uterine strips was reduced following prolonged in vivo treatment with isoproterenol but the effect was found only when estrogen alone was administered concomitantly. Finally, in vivo treatment with isoproterenol increased EC(50) of uterine strips subsequently stimulated with isoproterenol in vitro. This effect was independent of steroid treatment. We conclude that homologous desensitization of beta(2)AR function in non-pregnant rat myometrium in terms of sensitivity (EC(50)) is independent of sex steroids but in terms of maximal response (E(max)) occurs only in the presence of estrogen. We speculate whether progesterone withdrawal in connection with the well-known estrogen dominance at rat parturition may strengthen the desensitization induced by beta(2)AR activation and thus contribute to the transformation of the uterus from a quiescent to a highly contractile organ.
Osteoclast differentiation antigen, distinct from receptor activator of nuclear factor kappa B, is involved in osteoclastogenesis under calcitonin-regulated conditions
Authors: Kukita T , Kukita A , Watanabe T , Iijima T
Published: December 2004

Although calcitonin has been clinically utilized as a primary treatment for several metabolic bone diseases, its inhibitory effects against osteoclastic function diminish after several days owing to the calcitonin 'escape phenomenon'. We have previously found a unique cell-surface antigen (Kat1-antigen) expressed on rat osteoclasts. Here we show evidence that, in the presence of calcitonin, the Kat1-antigen is involved in osteoclastogenesis. Treatment of bone marrow cultures for forming osteoclast-like cells with anti-Kat1-antigen monoclonal antibody (mAb Kat1) provoked a marked stimulation of osteoclast-like cell formation only in the presence of calcitonin but not in its absence. Osteoclastogenesis stimulated by the receptor activator of nuclear factor kappa B (NF-kappaB) ligand/osteoclast differentiation factor was further augmented by mAb Kat1 in the presence of calcitonin. Furthermore, even in the presence of the osteoprotegerin/osteoclast inhibitory factor, mAb Kat1 induced osteoclast-like cell formation. Our current data suggest that the Kat1-antigen is a molecule that is distinct from receptor activator of NF-kappaB. The presence of the unique Kat1-antigen on cells in the osteoclast lineage appears to contribute to the fine regulation of osteoclastogenesis in vivo. Expression of this cell-surface molecule in cells in the osteoclast lineage may partly explain the mechanism responsible for the escape phenomenon.
In vitro free radical scavenging capacity of thyroid hormones and structural analogues
Authors: Oziol L , Faure P , Vergely C , Rochette L , Artur Y , Chomard P , Chomard P
Published: December 2004

It was reported that thyroid hormones decreased Cu(2+)-induced low-density lipoprotein (LDL) oxidation in vitro. Here, we investigated free radical scavenging capacities of thyroid hormones (3,5,3'-tri-iodo-L-thyronine (T(3)), thyroxine (T(4)) and 3,3',5'-tri-iodo-L-thyronine (rT(3))) and structural analogues (L-thyronine (T(0)), 3,5,3'tri-iodothyroacetic acid (TA(3)) and 3,5,3',5'-tetra-iodothyroacetic acid (TA(4))), using three different models of free radical generation. T(0), T(3) and TA(3) slowed down production of conjugated diene and thiobarbituric acid-reactive substances during LDL oxidation by 2,2'-azobis-[2-amidinopropane] (water-soluble), whereas rT(3), T(4) and TA(4) had practically no effect. In this system, T(0) was the more active compound. Using a 1,1-diphenyl-2-picrylhydrazyl (lipid-soluble) test, all compounds also revealed free radical scavenging capacities, but rT(3), T(4) and TA(4) were more active than T(0), T(3) and TA(3). T(3) was able to scavenge superoxide anion and hydroxyl radicals generated in an aqueous phase by a xanthine-xanthine oxidase system, as measured by electron paramagnetic resonance spectroscopy. It may be concluded that: (1) thyroid hormones and analogues with a 4'-hydroxy diphenylether structure have free radical scavenging capacities, (2) this property is influenced by the number of iodines on the phenolic ring, and (3) thyroid hormone scavenging capacity should not be the only mechanism explaining their protective effect on Cu(2+)-induced LDL oxidation. The physiological significance of the findings is discussed.
Long-term thyroxine administration increases heat stress protein-70 mRNA expression and attenuates p38 MAP kinase activity in response to ischaemia
Authors: Pantos CI , Malliopoulou VA , Mourouzis IS , Karamanoli EP , Tzeis SM , Carageorgiou HC , Varonos DD , Cokkinos DV
Published: December 2004

The present study was undertaken to investigate heat stress protein (HSP)-70 mRNA induction and p38 MAP kinase (MAPK) activity in response to ischaemic stress in the hyperthyroid rat heart. L-Thyroxine (T(4)) (25 microg/100 g body weight) was administered to Wistar rats for 2 days (THYRacute) or 14 days (THYR), while animals treated similarly with normal saline served as controls (NORMacute and NORM). In addition, abdominal aortic banding was performed in another group of rats to produce constriction-induced hypertrophy (HYP), while sham-operated (SOP) animals served as controls. Isolated rat hearts were perfused in a Langendorff mode. Hearts from NORMacute (n=6), THYRacute animals (n=8), NORM (n=6), THYR (n=6), SOP (n=5) and HYP (n=7) animals were subjected to 20 min of zero-flow global ischaemia followed by 45 min of reperfusion. HSP70 mRNA expression and phosphorylated p38 MAPK protein expression were detected in response to ischaemia and protein kinase C-epsilon (PKCepsilon) protein expression was detected at baseline. Thyroid hormones were measured in plasma. Long-term T(4) administration and aortic constriction resulted in the development of cardiac hypertrophy. Thyroid hormones were increased in both THYR and THYRacute as compared with normal groups (P<0.05). HSP70 mRNA induction was increased 2.3-fold in THYR as compared with NORM hearts (P<0.05), whereas there was not any difference between THYRacute and NORMacute hearts (P>0.05). Phosphorylated p38 MAPK protein expression was 2.2-fold more in NORM than in THYR hearts (P<0.05), but it was not different between NORMacute and THYRacute hearts (P>0.05). HSP70 mRNA induction was 1.8-fold greater in HYP than in SOP hearts (P<0.05), whereas phosphorylated p38 MAPK protein expression was similar between the two groups (P>0.05). PKCepsilon protein expression at baseline was 1.7-fold more in NORM than in THYR hearts (P<0.05), and not different between NORMacute and THYRacute hearts (P>0.05) as well as HYP and SOP hearts (P>0.05). This study shows that HSP70 mRNA expression is increased, whereas p38 MAPK activation is attenuated in response to ischaemia in long-term T(4)-treated rat hearts as compared with normal and acute hyperthyroid hearts.
Growth hormone endocrinology of Atlantic salmon (Salmo salar): pituitary gene expression, hormone storage, secretion and plasma levels during parr-smolt transformation
Authors: Agustsson T , Sundell K , Sakamoto T , Johansson V , Ando M , Th Bjornsson B
Published: December 2004

A number of studies on the Atlantic salmon (Salmo salar), have reported changes in plasma GH during parr-smolt transformation, but there is a lack of information about the endocrinology of the GH system during this process. In order to elucidate the mechanisms underlying these changes in plasma GH levels during the parr-smolt transformation of Atlantic salmon, GH mRNA expression in the pituitary was studied together with total pituitary GH content, in vitro GH secretion rate and plasma GH and IGF-I levels. Atlantic salmon were kept in outside tanks, under natural condition from early February until late June. Approximately three times a month fish were killed and pituitaries and blood were sampled for investigation. Further, pituitaries were moved to the laboratory for in vitro GH secretion studies. The results show that the GH system is first activated by an increase in GH secretion rate, which leads to an increase in plasma GH levels and causes a drop in the total GH content of the pituitary. This drop in pituitary GH content is later reversed by an increased GH synthesis seen as an increase in GH mRNA expression. Maximal activation of the GH system is seen to occur in early May, when plasma IGF-I levels reach highest levels, after which a certain deactivation of the GH system takes place. The data show that plasma levels of GH are to a large extent regulated by the secretion rate from the pituitary, although changes in the GH clearance rate are also likely to take place and influence the plasma GH levels. The study further underlines the significant role that the GH-IGF-I axis plays in the parr-smolt transformation of the Atlantic salmon.
Autocrine role of adrenomedullin in the human adrenal cortex
Authors: Thomson LM , Kapas S , Carroll M , Hinson JP
Published: December 2004

Previous studies from our laboratory have reported that adrenomedullin is synthesised in rat zona glomerulosa cells. In the present studies, it was found that the human adrenocortical cell line H295R expresses the gene encoding adrenomedullin, and that immunoreactive adrenomedullin is released into the culture medium. Furthermore, it was found that secretion of adrenomedullin is regulated by angiotensin II and forskolin. Studies on the actions of adrenomedullin and calcitonin gene-related peptide (CGRP) revealed a stimulatory effect of adrenomedullin, but not of CGRP, on aldosterone and cortisol secretion. These data suggest that adrenomedullin is not acting by a CGRP receptor-mediated mechanism in the H295R cell line. Adrenomedullin was also found to increase cAMP production, suggesting that in the adrenal, as in other cell types, cAMP is a second messenger for adrenomedullin action. However, the effects of adrenomedullin were not fully mimicked by forskolin, possibly suggesting a role for an additional second messenger. The presence of mRNA encoding both the putative adrenomedullin receptors, L1 and calcitonin receptorlike receptor/receptor-associated modulatory protein 2 (CRLR/RAMP-2), was demonstrated in H295R cells, but RAMP-1 was not detected, suggesting that these cells do not express the CGRPI receptor CRLR/RAMP-1. Taken together, these data have demonstrated that adrenomedullin is synthesised and secreted by H295R cells. The observed rate of adrenomedullin synthesis suggests that this peptide exerts a paracrine/autocrine effect in this adrenocortical cell line, probably acting through a specific adrenomedullin receptor, to stimulate steroidogenesis and increase aldosterone synthase expression.
Uroguanylin and guanylate cyclase C in the human pancreas: expression and mutuality of ligand/receptor localization as indicators of intercellular paracrine signaling pathways
Authors: Kulaksiz H , Cetin Y
Published: December 2004

The intestinal peptide hormone uroguanylin regulates electrolyte/fluid transport in the gastrointestinal epithelium by binding to its receptor, guanylate cyclase C (GC-C), and thus specifically coupling to activation of cystic fibrosis transmembrane conductance regulator (CFTR). Since CFTR is crucially involved in pancreatic electrolyte secretion, we investigated the human pancreas for expression and cell-specific localization of uroguanylin and guanylate cyclase C as potential regulatory components of pancreatic electrolyte secretion. RT-PCR analyses with specific primers revealed that uroguanylin and GC-C are expressed in the human pancreas (and in the duodenum, used as positive control); at the translational level, western blotting analyses with peptide- and region-specific antibodies identified the presence of 12.5 kDa uroguanylin and 130 kDa GC-C in both human pancreatic and intestinal extracts. At the cellular level, uroguanylin and GC-C immunoreactivities were absent from the islets of Langerhans but were exclusively confined to the exocrine parenchyma. Hence, uroguanylin was localized to the centroacinar cells typical of the pancreas, and also to epithelial cells of the intercalated, intralobular and interlobular ducts where the peptide was primarily concentrated adluminally to the apical portion of the respective cells. Coincidently, correlative studies localized the GC-C receptor to the epithelial cells of the ductal network, where it was confined exclusively to the apical cell membrane that evidently represents the functionally relevant target membrane domain for the regulatory peptide. In view of the fact that CFTR is highly expressed in pancreatic ductal cells where uroguanylin and its receptor are also localized, we assume that uroguanylin, an intrinsic pancreatic peptide, is involved in the regulation of electrolyte/water secretion in the ductal system via GC-C and CFTR. The particular cellular expression of uroguanylin in duct cells and the localization of GC-C to the duct cell apical membrane domain predict a novel route of intercellular signaling and luminal activation of GC-C via the pancreatic juice.
Daily patterns of plasma leptin in sheep: effects of photoperiod and food intake
Authors: Marie M , Findlay PA , Thomas L , Adam CL
Published: December 2004

Circulating concentrations of leptin in sheep correlate with body fatness and are affected by level of food intake and photoperiod. The present objective was to elucidate the short-term dynamics of leptin secretion. Frequent blood samples were taken over 48 h from 12 Soay rams after 16 weeks in short-day photoperiod (SD, 16 h darkness:8 h light) with freely available food, and then after 16 weeks in long days (16 h light:8 h darkness) with food freely available (LD) or restricted to 90% maintenance (LDR) (n=6/group). During the second 24 h of sampling, half were food deprived (n=6, SD and LD) and half had their meal times shifted (n=6, SD and LDR). A homologous RIA was developed, using antibodies raised in chicken against recombinant ovine leptin, to measure plasma concentrations. Simultaneous 24 h profiles of plasma insulin, glucose and non-esterified fatty acids (NEFA) were measured. Plasma leptin was higher in LD than SD, and in LD than LDR, associated with higher food intake, liveweight and body condition score (adiposity), but tended to be lower in LDR than SD, associated with lower food intake, liveweight and body condition score. There was no evidence for a circadian rhythm of plasma leptin, but clear evidence for post-prandial peaks of low amplitude (15-36%) 2-8 h after meals given at normal and shifted times. Complete food deprivation caused a dramatic fall in plasma leptin to basal levels within 24 h. There was a positive association of plasma leptin with plasma insulin, and negative association with NEFA, both between meals and during fasting. Thus, plasma leptin concentrations in sheep are sensitive to short-term changes in energy balance, as well as to long-term photoperiod-driven changes in food intake and adiposity.
Up to date with human thyroglobulin
Authors: van de Graaf SA , Ris-Stalpers C , Pauws E , Mendive FM , Targovnik HM , de Vijlder JJ
Published: December 2004

The coding region of the human thyroglobulin (TG) mRNA has been resequenced, and comparison with the TG sequence originally published in 1987 showed many variations. All of the variations were validated in 20--40 other alleles, and this resulted in the revision of 41 nucleotide positions. This review presents the revised wild-type human TG sequence, including all known exon/exon boundaries and additional data on the TG mRNA population, concerning alternative splicing and variability of the polyadenylation cleavage site. The amino acid sequence derived shows one additional, 12 changed, and 10 polymorphic residues. Protein characteristics, such as acceptor and donor tyrosine residues, N-glycosylation sites, cysteine-rich repeats, the proposed receptor domain, and antigenic epitopes, are included, and their relationship to the revised sequence is discussed. Furthermore, all reported TG mutations causing dyshormonogenesis in humans and animals are designated in the nucleotide and amino acid sequences. This up-to-date profile of the human TG molecule presents the features of importance for its complex role in thyroid hormonogenesis, and is the basis for future studies on the structure--function relationship.
The role of the IGF axis in IGFBP-1 and IGF-I induced renal enlargement in Snell dwarf mice
Authors: van Kleffens M , Lindenbergh-Kortleve DJ , Koster JG , van Neck JW , Flyvbjerg A , Rasch R , Drop SL , van Buul-Offers SC
Published: December 2004

Insulin-like growth factor (IGF) binding protein-1 (IGFBP-1) is generally believed to inhibit IGF action in the circulation. In contrast, IGFBP-1 has been reported to interact with cell surfaces and enhance IGF-I action locally in some tissues. Renal IGFBP-1 levels are found elevated in various conditions characterized by renal growth (e.g. diabetes mellitus, hypokalemia). To test whether IGFBP-1 is a renotropic factor, IGFBP-1 was administered alone or in combination with IGF-I to Snell dwarf mice, an in vivo model without compensatory feedback effects on growth hormone (GH) secretion. In three control groups of Snell dwarf mice, placebo, GH or IGF-I was administered. Compared with placebo, kidney weight increased in all treated groups, however, with different effects on kidney morphology. Administration of IGF-I, alone or in combination with IGFBP-1, tended to increase glomerular volume, while no changes were seen in the other groups. Administration of IGFBP-1 or IGFBP-1+IGF-I both caused dilatation of the thin limbs of Henle's loop, while GH or IGF-I administration had no visible effect. Furthermore, IGF-I administration resulted in an increased mean number of nuclei per cortical area and renal weight, whereas GH, IGF-I+IGFBP-1 or IGFBP-1 caused a decreased renal nuclei number. In situ hybridization and immunohistochemistry showed specific changes of the renal IGF system expression patterns in the different groups. Particularly, IGFBP-1 administration resulted in extensive changes in the mRNA expression of the renal IGF system, whereas the other administration regimen resulted in less prominent modifications. In contrast, administration of IGFBP-1 and IGFBP-1+IGF-I resulted in identical changes in the protein expression of the renal IGF system. Our results indicate that IGFBP-1, alone or in combination with IGF-I, demonstrated effects on the renal tubular system that differ from the effects of IGF-I.
Retinoic acid and vitamin D(3) powerfully inhibit in vitro leptin secretion by human adipose tissue
Authors: Menendez C , Lage M , Peino R , Baldelli R , Concheiro P , Dieguez C , Casanueva FF
Published: December 2004

Leptin, the product of the ob gene, is secreted into the circulation by white adipose tissue; its major role being to participate in the regulation of energy homeostasis. Plasma leptin levels are mainly determined by the relative adiposity of the subject; however, the great dispersion of values for any given body mass index and the noteworthy gender-based differences indicate that other factors are operating. Steroid hormones actively participate in the regulation of leptin secretion; however, non-steroid nuclear hormones have either not been studied or have provided contradictory results. In order to understand the role of hormones of the non-steroid superfamily such as 3,5,3'-tri-iodothyronine (T(3)), vitamin D(3) and retinoic acid (RA) in the control of leptin secretion, in the present work doses of 10(-9), 10(-8) and 10(-7) M of these compounds have been studied on in vitro leptin secretion. The organ culture was performed with omental adipose tissue samples from healthy donors (n=28). T(3) was devoid of effect at any dose studied, while an inhibition of leptin secretion was observed with 9-cis-RA (slight) and all-trans-RA (potent). Interestingly, vitamin D(3) exerted a powerfully inhibitory role at the doses studied, and its action was synergistic with all-trans-RA. In conclusion, in vitro leptin secretion by human adipose tissue is negatively controlled by either RA or vitamin D(3). The clinical significance of leptin regulation by this superfamily of nuclear receptors remains to be ascertained.
Nitric oxide as a second messenger in parathyroid hormone-related protein signaling
Authors: Kalinowski L , Dobrucki LW , Malinski T
Published: December 2004

Parathyroid hormone (PTH)-related protein (PTHrP) is produced in smooth muscles and endothelial cells and is believed to participate in the local regulation of vascular tone. No direct evidence for the activation of endothelium-derived nitric oxide (NO) signaling pathway by PTHrP has been found despite attempts to identify it. Based on direct in situ measurements, it is reported here for the first time that the human PTH/PTHrP receptor analogs, hPTH(1--34) and hPTHrP(1--34), stimulate NO release from a single endothelial cell. A highly sensitive porphyrinic microsensor with a response time of 0.1 ms and a detection limit of 1 nmol/l was used for the measurement of NO. Both hPTH(1--34) and hPTHrP(1--34) stimulated NO release at nanomolar concentrations. The peak concentration of 0.1 micromol/l hPTH(1--34)- and 0.1 micromol/l hPTHrP(1--34)-stimulated NO release was 175+/-9 and 248+/-13 nmol/l respectively. This represents about 30%--40% of maximum NO concentration recorded in the presence of (0.1 micromol/l) calcium ionophore. Two competitive PTH/PTHrP receptor antagonists, 10 micromol/l [Leu(11),d -Trp(12)]-hPTHrP(7--34)amide and 10 micromol/l [Nle(8,18),Tyr(34)]-bPTH(3--34)amide, were equipotent in antagonizing hPTH(1--34)-stimulated NO release; [Leu(11),d -Trp(12)]-hPTHrP(7--34)amide was more potent than [Nle(8,18),Tyr(34)]-bPTH(3--34)amide in inhibiting hPTHrP(1--34)-stimulated NO release. The PKC inhibitor, H-7 (50 micromol/l), did not change hPTH(1--34)- and hPTHrP(1--34)-stimulated NO release, whereas the combined effect of 10 micromol/l of the cAMP antagonist, Rp-cAMPS, and 50 micromol/l of the calmodulin inhibitor, W-7, was additive. The present studies show that both hPTH(1--34) and hPTHrP(1--34) activate NO production in endothelial cells. The activation of NO release is through PTH/PTHrP receptors and is mediated via the calcium/calmodulin pathway.