Steroids and neuroendocrine function in anorexia nervosa

Anorexia nervosa is a primarily psychiatric syndrome of self-induced weight loss due to an intense fear of becoming obese. Numerous endocrine abnormalities occur in anorexia nervosa patients, and in many respects these alterations reflects the endocrinology of reduced energy intake. However, the basic mechanisms of those alterations are far from being understood. In an attempt to understand the disrupted mechanisms of the hypogonadotropic hypogonadism of the anorectic state, we studied 10 anorectic women in the acute phase of their illness; all met the DSM III criteria. On each patient, two tests were performed with either saline as control or infusion of the opioid antagonist naloxone, and both LH and FSH levels were measured. Four mg of naloxone as bolus was used, followed by a naloxone infusion of 2 mg/h for 4 h. Compared with the pattern of normal women, naloxone did not increase in the anorectic patients either LH or FSH levels nor pulsatility. This result suggests that endogenous opioid peptides are not implicated in the low gonadotropic situation of anorexia nervosa. An alternative explanation could be that the low estrogenic "milieu" of these patients could mask the opioid action. To test this second possibility, another group of 7 anorectic women after partial weight recovery were challenged with estrogen administration. Compared with the pattern of normal women volunteers, all the anorectic patients but one presented an abnormal response in both LH and FSH levels after estrogen administration. In fact, the negative feedback and the delayed positive feedback of LH after estrogen were absent in these patients. Interestingly enough, the only patient with near-normal LH response to estrogen was considered fully recovered by the Psychiatric Unit. Several alterations in the hypothalamic-pituitary-adrenal axis has been reported in anorexia nervosa. Seven anorectic patients and 7 aged-matched women were challenged by ACTH 1-24, 250 micrograms (i.v.) and the ratio of increments in adrenal steroid products to precursors monitored. ACTH-induced increments in cortisol with respect to increments in 17-OH-progesterone was similar in anorectics and controls. On the contrary, the ratio of increments of androstenedione with respect to increments in 17-OH-progesterone were greater in anorexia nervosa than controls. These results suggest that in anorexia nervosa the 11-beta-21-alpha-hydroxylase system is normal but a deficient 17-20 desmolase system is present. Finally, the altered pattern of GH secretion in anorexia was studied using GHRH (1 microgram/kg) as stimulus of pituitary GH secretion.(ABSTRACT TRUNCATED AT 400 WORDS)     

Inositol 1,4,5-trisphosphate-independent Ca(2+) mobilization triggered by a lipid factor isolated from vitreous body.

A complex phospholipid from bovine vitreous body with a strong Ca(2+)-mobilizing activity has been recently isolated to homogeneity by our group. In this work, a sequential analysis of its transmembrane signaling pathway has been undertaken to characterize the intracellular mechanisms responsible for the Ca(2+) rise. The results show that this phospholipid induces, in a dose-dependent manner (ED(50) of around 0.25 microgram/ml), a Ca(2+) mobilization from inositol 1,4,5-trisphosphate-insensitive intracellular stores, with no participation of extracellular Ca(2+). Upon repeated administration, it shows no signs of autologous desensitization, does not induce heterologous desensitization of the L-alpha-lysophosphatidic acid (LPA) receptor but is desensitized by the previous administration of LPA. The Ca(2+)-mobilizing activity requires a membrane protein, is blocked after preincubation of the cells with pertussis toxin and phorbol esters, as well as by U73122 (an inhibitor of phospholipases C/D), R59022 (a diacylglycerol kinase inhibitor), and D609 (which inhibits phosphatidylcholine-specific phospholipase C). Upon administration of this phospholipid, the intracellular levels of phosphatidic acid (PA) rise with a time course that parallels that of the Ca(2+) mobilization, suggesting that PA could be responsible for this Ca(2+) signal. Exposure to AACOCF(3) (a specific inhibitor of phospholipase A(2)) does not modify the Ca(2+) rise, ruling out the possibility that the PA generated could be further converted to LPA by the action of phospholipase A(2). Based on the experimental data obtained, a signaling pathway involving a phosphatidylcholine-specific phospholipase C coupled to diacylglycerol kinase is proposed. This compound may represent a new class of bioactive lipids with a putative role in the physiology of the vitreous body.     

GLP-1 induces alpha cell proliferation and overrides leptin suppression induced by negative energy balance in vagotomized rats

Glucagon-like peptide-1 (GLP-1) influences energy balance by exerting effects on food intake and glucose metabolism, through mechanisms that are partially dependent on the vagal pathway. The aim of this study was to characterize the effects of chronic GLP-1 stimulation on energy homeostasis and glucose metabolism in the absence of vagal innervation Truncal vagotomized (VGX) and sham operated rats (SHAM) received an intraperitoneal GLP-1 infusion (3.5 pmol/kg/min) trough mini-osmotic pumps. To dissect the effects derived from vagal denervation on food intake, an additional group was included consisting of sham operated rats that were PAIR FED to VGX. Food intake and body weight were recorded throughout the experimental period, while the percentage of white and brown adipose tissue, fasting glucose, insulin, gastro-intestinal hormonal profile, hypothalamic, and BAT gene expression were assessed at endpoint. VGX rats had significantly lower food intake, body weight gain, and leptin levels when compared with SHAM rats. Despite having similar body weight, PAIR-FED rats had lower fasting leptin, insulin and insulin resistance, while having higher ghrelin levels than VGX. GLP-1 infusion did not influence food intake or body weight, but was associated with lower leptin levels in VGX and lower pancreatic α-cells ki-67 staining in SHAM. Concluding, this study corroborates that the vagus nerve may modulate whole body energy homeostasis by acting in peripheral signals. Our data suggest that in the absence of vagal or parasympathetic tonus, GLP-1 mediated inhibition of cell proliferation markers in α-cells is prevented, meanwhile leptin suppression, associated with a negative energy balance, is partially overridden.     

Interleukin 6 Deficiency Modulates the Hypothalamic Expression of Energy Balance Regulating Peptides during Pregnancy in Mice

Pregnancy is associated with hyperphagia, increased adiposity and multiple neuroendocrine adaptations. Maternal adipose tissue secretes rising amounts of interleukin 6 (IL6), which acts peripherally modulating metabolic function and centrally increasing energy expenditure and reducing body fat. To explore the role of IL6 in the central mechanisms governing dam''s energy homeostasis, early, mid and late pregnant (gestational days 7, 13 and 18) wild-type (WT) and Il6 knockout mice (Il6-KO) were compared with virgin controls at diestrus. Food intake, body weight and composition as well as indirect calorimetry measurements were performed in vivo. Anabolic and orexigenic peptides: neuropeptide Y (Npy) and agouti-related peptide (Agrp); and catabolic and anorectic neuropeptides: proopiomelanocortin (Pomc), corticotrophin and thyrotropin-releasing hormone (Crh and Trh) mRNA levels were determined by in situ hybridization. Real time-PCR and western-blot were used for additional tissue gene expression and protein studies. Non-pregnant Il6-KO mice were leaner than WT mice due to a decrease in fat but not in lean body mass. Pregnant Il6-KO mice had higher fat accretion despite similar body weight gain than WT controls. A decreased fat utilization in absence of Il6 might explain this effect, as shown by increased respiratory exchange ratio (RER) in virgin Il6-KO mice. Il6 mRNA levels were markedly enhanced in adipose tissue but reduced in hypothalamus of mid and late pregnant WT mice. Trh expression was also stimulated at gestational day 13 and lack of Il6 blunted this effect. Conversely, in late pregnant mice lessened hypothalamic Il6 receptor alpha (Il6ra), Pomc and Crh mRNA were observed. Il6 deficiency during this stage up-regulated Npy and Agrp expression, while restoring Pomc mRNA levels to virgin values. Together these results demonstrate that IL6/IL6Ra system modulates Npy/Agrp, Pomc and Trh expression during mouse pregnancy, supporting a role of IL6 in the central regulation of body fat in this physiological state.     

Insuficiencia suprarrenal y su tratamiento sustitutivo. Su realidad en España

Adrenal insufficiency (AI) is a rare endocrine disease, associated to increased mortality if left untreated. It can be due to a primary failure of the adrenal glands (primary AI) or malfunctioning of the hypothalamic-pituitary-adrenal axis (HPA) (secondary AI). The lack of data on incidence/prevalence of adrenal insufficiency in Spain complicates any evaluation of the magnitude of the problem in our country. Initial symptoms are non-specific, so often there is a delay in diagnosis.Current therapy with available glucocorticoids is associated with decreased quality of life in patients with treated AI, as well as with increased mortality and morbidity, probably related to both over-treatment and lack of hydrocortisone, associated with non-physiological peaks and troughs of the drug over the 24 hours. The availability of a new drug with a modified dual release (immediate and retarded), that requires one only daily dose, improves and simplifies the treatment, increases compliance as well as quality of life, morbidity and possibly mortality. This revision deals with the knowledge on the situation both globally and in Spain, prior to the availability of this new drug.     

Stimulation by ghrelin of p42/p44 mitogen-activated protein kinase through the GHS-R1a receptor: role of G-proteins and beta-arrestins

Results presented in this study indicate that in human embryonic kidney 293 cells (HEK 293), the ghrelin receptor growth hormone secretagogue receptor type 1a (GHS-R1a) activates the extracellular signal-related kinases 1 and 2 (ERK 1/2) via three pathways. One pathway is mediated by the beta-arrestins 1 and 2, and requires entry of the receptor into a multiprotein complex with the beta-arrestins, Src, Raf-1, and ERK 1/2. A second pathway is G(q/11)-dependent and involves a Ca(2+)-dependent PKC (PKCalpha/beta) and Src. A third pathway is G(i)-dependent and involves phosphoinositide 3-kinase (PI3K), PKCepsilon, and Src. Our current study reveals that G(i/o)- and G(q/11)-proteins are crucially involved in the beta-arrestin-mediated ERK 1/2 activation. These results thus support the view that the beta-arrestins act as both scaffolding proteins and signal transducers in ERK 1/2 activation, as reported for other receptors. The different pathways of ERK 1/2 activation suggest that binding to GHS-R1a activates ERK 1/2 pools at different locations within the cell, and thus probably with different physiological consequences.     

Role of obestatin on growth hormone secretion: An in vitro approach

Obestatin, the ghrelin-associated peptide, showed to activate MAPK signaling with no effect on Akt nor cell proliferating activity in rat tumor somatotroph cells (growth cells, GC). A sequential analysis of the obestatin transmembrane signaling pathway indicated a route involving the consecutive activation of G_i, PI3k, novel PKCε, and Src for ERK1/2 activation. Furthermore, obestatin treatment triggers growth hormone (GH) release in the first 30 min, being more acute at 15 min. At 1 h, obestatin treated cells showed the same levels in GH secretion than controls. Added to this functionality, obestatin was secreted by GC cells. Based on the capacity to stimulate GH release from somatotroph cells, obestatin may act directly in the pituitary through an autocrine/paracrine mechanism.     

Niche-associated activation of rac promotes the asymmetric division of Drosophila female germline stem cells

Background

Drosophila female germline stem cells (GSCs) reside adjacent to a cellular niche that secretes Bone Morphogenetic Protein (BMP) ligands and anchors the GSCs through adherens junctions. The GSCs divide asymmetrically such that one daughter remains in the niche as a GSC, while the other is born away from the niche and differentiates. However, given that the BMP signal can be diffusible, it remains unclear how a local extracellular asymmetry is sufficient to result in a robust pattern of asymmetric division.

Methods and Findings

Here we show that GSCs are polarized with respect to the cellular niche. We first use a modified biosensor to demonstrate that the small GTPase Rac is asymmetrically activated within the GSC at the niche-GSC interface. Experiments using loss-of-function and gain-of-function mutations in Rac indicate that asymmetric Rac activity both localizes the microtubule binding protein Apc2 to orient one GSC centrosome at the niche-GSC interface during interphase and activates the Jun N-terminal kinase pathway to increase the ability of the GSC to respond to BMP ligands. Other processes act in concert with each function of Rac. Specifically, we demonstrate that the GSC cell cycle arrests at prometaphase if centrosomes are misoriented.

Conclusions

Thus, the GSCs, an adult stem cell present in a cellular niche, have a niche-associated polarity that couples control of the division plane with increased response to an extracellular maintenance signal. Other processes work in parallel with the Rac-mediated polarity to ensure a robust pattern of asymmetric division. We suggest that all adult stem cells likely employ multiple, independently acting mechanisms to ensure asymmetric division to maintain tissue homeostasis.     

Leptin inhibits lysophosphatidic acid-induced intracellular calcium rise by a protein kinase C-dependent mechanism

Leptin communicates the status of body energy stores to the central nervous system, regulating appetite, metabolic rate, and neuroendocrine functions. These effects are mediated by leptin binding and activation of the cognate cell surface receptor, a member of type I cytokine receptor family, which lead to the activation of receptor-associated kinases of the Janus family. In this work, we demonstrate that leptin inhibits the l-alpha-lysophosphatidic acid (LPA)-induced intracellular calcium mobilization in a dose-dependent manner in HEK-293 cells stably expressing full-length leptin receptor (OB-Rb). This action appears to be selective, as it was not observed when other signaling families, such as VIP or EGF, were studied. Pretreatment with the phosphatidylinositol 3-kinase (PI3K) inhibitor, wortmannin, reversed the effect of leptin, pointing to PI3K as an intermediate molecule involved in this process. An unspecific protein kinase C (PKC) inhibitor, staurosporine, disrupted the inhibitory action of leptin. Furthermore, intracellular levels of phosphorylated PKCepsilon and PKCdelta rose to a maximum 5 min after leptin administration, suggesting that these atypical PKC isoforms are involved in the observed cross-desensitization. To define the regions of the OB-Rb intracellular domain required for the cross-desensitization, a series of C-terminal deletion mutants were transfected into HEK-293 cells. C-terminal truncation that removed the consensus Box 3 motif of OB-Rb prevented leptin action, indicating that heterologous desensitization over LPA was exerted at the level of this intracellular motif. Our date demonstrate that leptin plays a key role in the regulation of the earliest signaling pathways activated by growth factors, such as LPA, through a signaling pathway involving PKCdelta and PKCepsilon coupled to Box 3 motif of the OB-Rb through PI3K.     

Changes in beta-endorphin content in discrete areas of the hypothalamus throughout proestrus and diestrus of the rat

The aim of the present study is to investigate changes in beta-endorphin content in the hypothalamus during different stages of the estrous cycle. Groups of 9 to 10 Sprague-Dawley rats were sacrificed every two hours on proestrus from 8.00 to 18.00 h and groups of 7 to 8 rats were sacrificed on diestrus at 8.00, 12.00, 14.00 and 18.00 h. Preoptic suprachiasmatic region, posterior hypothalamus, arcuate nucleus and median eminence were dissected and assayed for beta-endorphin. A significant increase in beta-endorphin content was detected in the arcuate nucleus during proestrus (9.00 h: 1.76 +/- .31; 14.00 h: 4.10 +/- .85 microgram/g tissue wet weight). Levels did not change during diestrus (1.18 +/- .06 microgram/g). The increase caused significant differences in beta-endorphin values between both days at 12.00, 14.00 and 18.00 h, while the concentrations at 8.00 h were similar. The opposite pattern was observed in the median eminence with significantly higher proestrous beta-endorphin levels at 8.00 h (11.24 +/- 3.1 vs 3.52 +/- .64 microgram/g) and nonsignificant differences for the rest of the day. No significant change in beta-endorphin concentration was seen in the preoptic suprachiasmatic region over the day of proestrus (1.35 +/- .09 microgram/g). Diestrous beta-endorphin concentrations in this region were higher during the morning (2.60 +/- .65 microgram/g) and lower at 18.00 h (0.94 +/- .12 microgram/g) when compared to proestrous values. This pattern was caused by a 50% increase in beta-endorphin during the afternoon of diestrus. No changes were observed in the posterior hypothalamus on either day with comparable levels of beta-endorphin except at 18.00 h, when values were significantly higher on proestrus (1.66 +/- .30 vs 0.83 +/- .06 microgram/g).