Divergent androgen regulation of unfolded protein response pathways drives prostate cancer

The unfolded protein response (UPR) is a homeostatic mechanism to maintain endoplasmic reticulum (ER) function. The UPR is activated by various physiological conditions as well as in disease states, such as cancer. As androgens regulate secretion and development of the normal prostate and drive prostate cancer (PCa) growth, they may affect UPR pathways. Here, we show that the canonical UPR pathways are directly and divergently regulated by androgens in PCa cells, through the androgen receptor (AR), which is critical for PCa survival. AR bound to gene regulatory sites and activated the IRE1α branch, but simultaneously inhibited PERK signaling. Inhibition of the IRE1α arm profoundly reduced PCa cell growth in vitro as well as tumor formation in preclinical models of PCa in vivo. Consistently, AR and UPR gene expression were correlated in human PCa, and spliced XBP-1 expression was significantly upregulated in cancer compared with normal prostate. These data establish a genetic switch orchestrated by AR that divergently regulates the UPR pathways and suggest that targeting IRE1α signaling may have therapeutic utility in PCa.

The association between perceived sweetness intensity and dietary intake in young adults

Individual differences in taste perception may influence dietary habits, nutritional status, and ultimately nutrition-related chronic disease risk. Individual differences in sweetness intensity perception and the relationship between perceived sweetness intensity, food behaviors, and dietary intake was investigated in 85 adults. Subjects (body mass index [BMI]= 21 ± 3, 21 ± 4 y) completed a food and diet questionnaire, food variety survey, 2 24-h food records, and a perceived sweetness intensity measurement using the general labeled magnitude scale (gLMS). There was interindividual variation in perceived sweetness intensity (0 to 34 gLMS units, mean 10 ± 7). One-way analysis of variance (ANOVA) revealed no difference between perceived sweetness intensity and degree of importance placed on not adding sugar to tea or coffee (P = 0.2) and the degree of importance placed on avoiding sugar-sweetened or fizzy drinks (P = 1.0). Independent t-test analysis revealed no significant association between perceived sweetness intensity and the food variety measure for sugar and confectionary intake (P = 0.6) and selected fruit and vegetable intake (P = 0.1 to 0.9). One-way ANOVA also demonstrated no difference between tertiles of sweetness intensity and BMI (P = 0.1), age (P = 0.3), and food variety score (P = 0.5). No correlation was observed with regards to perceived sweetness intensity and mean total energy (kJ) intake (r = 0.05, P = 0.7), percent energy from total fat, saturated fat, protein, carbohydrate, and grams of fiber (r =–0.1 to 0.1, P = 0.2 to 0.8) and also for intake of the micronutrients: folate, magnesium, calcium, iron, and zinc (r = 0.1 to 0.2, P = 0.1 to 0.4). Only modest correlations were observed between sodium (r = 0.3, P < 0.05), vitamin C (r = 0.3, P < 0.05), and potassium (r = 0.2, P < 0.0) intake and perceived sweetness intensity. Overall, perceived sweetness intensity does not appear to play a role in food behaviors relating to sugar consumption and dietary intake in adults.

Progesterone and testosterone in combination act in the hypothalamus of castrated rams to regulate the secretion of LH

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.

Lysosomal secretion of Flightless I upon injury has the potential to alter inflammation

Intracellular Flightless I (Flii), a gelsolin family member, has been found to have roles modulating actin regulation, transcriptional regulation and inflammation. In vivo Flii can regulate wound healing responses. We have recently shown that a pool of Flii is secreted by fibroblasts and macrophages, cells typically found in wounds, and its secretion can be upregulated upon wounding. We show that secreted Flii can bind to the bacterial cell wall component lipopolysaccharide and has the potential to regulate inflammation. We now show that secreted Flii is present in both acute and chronic wound fluid.

Cytokine expression and secretion by skeletal muscle cells : regulatory mechanisms and exercise effects

Cytokines are important mediators of various aspects of health and disease, including appetite, glucose and lipid metabolism, insulin sensitivity, skeletal muscle hypertrophy and atrophy. Over the past decade or so, considerable attention has focused on the potential for regular exercise to counteract a range of disease states by modulating cytokine production. Exercise stimulates moderate to large increases in the circulating concentrations of interleukin (IL)-6, IL-8, IL-10, IL-1 receptor antagonist, granulocyte-colony stimulating factor, and smaller increases in tumor necrosis factor-α, monocyte chemotactic protein-1, IL-1β, brain-derived neurotrophic factor, IL-12p35/p40 and IL-15. Although many of these cytokines are also expressed in skeletal muscle, not all are released from skeletal muscle into the circulation during exercise. Conversely, some cytokines that are present in the circulation are not expressed in skeletal muscle after exercise. The reasons for these discrepant cytokine responses to exercise are unclear. In this review, we address these uncertainties by summarizing the capacity of skeletal muscle cells to produce cytokines, analyzing other potential cellular sources of circulating cytokines during exercise, and discussing the soluble factors and intracellular signaling pathways that regulate cytokine synthesis (e.g., RNA-binding proteins, microRNAs, suppressor of cytokine signaling proteins, soluble receptors).

Secretion and processing of a novel multi-domain cystatin-like protein by intracellular stages of Trichinella spiralis

The excretory-secretory (ES) proteins of nematode parasites are of major interest as they function at the host-parasite interface and are likely to have roles crucial for successful parasitism. Furthermore, the ES proteins of intracellular nematodes such as Trichinella spiralis may also function to regulate gene expression in the host cell. In a recent proteomic analysis we identified a novel secreted cystatin-like protein from T. spiralis L1 muscle larva. Here we show that the protein, MCD-1 (multi-cystatin-like domain protein 1), contains three repeating cystatin-like domains and analysis of the mcd-1 gene structure suggests that the repeated domains arose from duplication of an ancestral cystatin gene. Cystatins are a diverse group of cysteine protease inhibitors and those secreted by parasitic nematodes are important immuno-modulatory factors. The cystatin superfamily also includes cystatin-like proteins that have no cysteine protease inhibitory activity. A recombinant MCD-1 protein expressed as a GST-fusion protein in Escherichia coli failed to inhibit papain in vitro suggesting that the T. spiralis protein is a new member of the non-inhibitory cystatin-related proteins. MCD-1 secreted from T. spiralis exists as high- and low-molecular weight isoforms and we show that a recombinant MCD-1 protein secreted by HeLa cells undergoes pH-dependent processing that may result in the release of individual cystatin-like domains. Furthermore, we found that mcd-1 gene expression is largely restricted to intracellular stages with the highest levels of expression in the adult worms. It is likely that the major role of the protein is during the intestinal stage of T. spiralis infections.

Insulin secretion is regulated by the glucose-dependent production of islet beta cell macrophage migration inhibitory factor.

Macrophage migration inhibitory factor (MIF), originally identified as a cytokine secreted by T lymphocytes, was found recently to be both a pituitary hormone and a mediator released by immune cells in response to glucocorticoid stimulation. We report here that the insulin-secreting beta cell of the islets of Langerhans expresses MIF and that its production is regulated by glucose in a time- and concentration-dependent manner. MIF and insulin colocalize by immunocytochemistry within the secretory granules of the pancreatic islet beta cells, and once released, MIF appears to regulate insulin release in an autocrine fashion. In perifusion studies performed with isolated rat islets, immunoneutralization of MIF reduced the first and second phase of the glucose-induced insulin secretion response by 39% and 31%, respectively. Conversely, exogenously added recombinant MIF was found to potentiate insulin release. Constitutive expression of MIF antisense RNA in the insulin-secreting INS-1 cell line inhibited MIF protein synthesis and decreased significantly glucose-induced insulin release. MIF is therefore a glucose-dependent, islet cell product that regulates insulin secretion in a positive manner and may play an important role in carbohydrate metabolism.

Monoacylglycerol, alpha/beta-hydrolase domain-6, and the regulation of insulin secretion and energy metabolism

Le cycle glycérolipides/acides gras libres (GL/FFA) est une voie métabolique clé qui relie le métabolisme du glucose et des acides gras et il est composé de deux processus métaboliques appelés lipogenèse et lipolyse. Le cycle GL/FFA, en particulier la lipolyse des triglycérides, génère diverses molécules de signalisation pour réguler la sécrétion d'insuline dans les cellules bêta pancréatiques et la thermogenèse non-frissonnante dans les adipocytes. Actuellement, les lipides provenant spécifiquement de la lipolyse impliqués dans ce processus sont mal connus. L’hydrolyse des triglycérides dans les cellules β est réalisée par les actions successives de la triglycéride lipase adipocytaire pour produire le diacylglycérol, ensuite par la lipase hormono-sensible pour produire le monoacylglycérol (MAG) et enfin par la MAG lipase (MAGL) qui relâche du glycerol et des acides gras. Dans les cellules bêta, la MAGL classique est très peu exprimée et cette étude a démontré que l’hydrolyse de MAG dans les cellules β est principalement réalisée par l'α/β-Hydrolase Domain-6 (ABHD6) nouvellement identifiée. L’inhibition d’ABHD6 par son inhibiteur spécifique WWL70, conduit à une accumulation des 1-MAG à longues chaines saturées à l'intérieur des cellules, accompagnée d’une augmentation de la sécrétion d'insuline stimulée par le glucose (GSIS). Baisser les niveaux de MAG en surexprimant ABHD6 dans la lignée cellulaire bêta INS832/13 réduit la GSIS, tandis qu’une augmentation des niveaux de MAG par le « knockdown » d’ABHD6 améliore la GSIS. L'exposition aiguë des monoacylglycérols exogènes stimule la sécrétion d'insuline de manière dose-dépendante et restaure la GSIS supprimée par un inhibiteur de lipases appelé orlistat. En outre, les souris avec une inactivation du gène ABHD6 dans tous les tissus (ABHD6-KO) et celles avec une inactivation du gène ABHD6 spécifiquement dans la cellule β présentent une GSIS stimulée, et leurs îlots montrent une augmentation de la production de monoacylglycérol et de la sécrétion d'insuline en réponse au glucose. L’inhibition d’ABHD6 chez les souris diabétiques (modèle induit par de faibles doses de streptozotocine) restaure la GSIS et améliore la tolérance au glucose. De plus, les résultats montrent que les MAGs non seulement améliorent la GSIS, mais potentialisent également la sécrétion d’insuline induite par les acides gras libres ainsi que la sécrétion d’insuline induite par divers agents et hormones, sans altération de l'oxydation et l'utilisation du glucose ainsi que l'oxydation des acides gras. Nous avons démontré que le MAG se lie à la protéine d’amorçage des vésicules appelée Munc13-1 et l’active, induisant ainsi l’exocytose de l'insuline. Sur la base de ces observations, nous proposons que le 1-MAG à chaines saturées agit comme facteur de couplage métabolique pour réguler la sécrétion d'insuline et que ABHD6 est un modulateur négatif de la sécrétion d'insuline. En plus de son rôle dans les cellules bêta, ABHD6 est également fortement exprimé dans les adipocytes et son niveau est augmenté avec l'obésité. Les souris dépourvues globalement d’ABHD6 et nourris avec une diète riche en gras (HFD) montrent une faible diminution de la prise alimentaire, une diminution du gain de poids corporel et de la glycémie à jeun et une amélioration de la tolérance au glucose et de la sensibilité à l'insuline et ont une activité locomotrice accrue. En outre, les souris ABHD6-KO affichent une augmentation de la dépense énergétique et de la thermogenèse induite par le froid. En conformité avec ceci, ces souris présentent des niveaux élevés d’UCP1 dans les adipocytes blancs et bruns, indiquant le brunissement des adipocytes blancs. Le phénotype de brunissement est reproduit dans les souris soit en les traitant de manière chronique avec WWL70 (inhibiteur d’ABHD6) ou des oligonucléotides anti-sense ciblant l’ABHD6. Les tissus adipeux blanc et brun isolés de souris ABHD6-KO montrent des niveaux très élevés de 1-MAG, mais pas de 2-MAG. L'augmentation des niveaux de MAG soit par administration exogène in vitro de 1-MAG ou par inhibition ou délétion génétique d’ABHD6 provoque le brunissement des adipocytes blancs. Une autre évidence indique que les 1-MAGs sont capables de transactiver PPARα et PPARγ et que l'effet de brunissement induit par WWL70 ou le MAG exogène est aboli par les antagonistes de PPARα et PPARγ. L’administration in vivo de l’antagoniste de PPARα GW6471 à des souris ABHD6-KO inverse partiellement les effets causés par l’inactivation du gène ABHD6 sur le gain de poids corporel, et abolit l’augmentation de la thermogenèse, le brunissement du tissu adipeux blanc et l'oxydation des acides gras dans le tissu adipeux brun. L’ensemble de ces observations indique que ABHD6 régule non seulement l’homéostasie de l'insuline et du glucose, mais aussi l'homéostasie énergétique et la fonction des tissus adipeux. Ainsi, 1-MAG agit non seulement comme un facteur de couplage métabolique pour réguler la sécrétion d'insuline en activant Munc13-1 dans les cellules bêta, mais régule aussi le brunissement des adipocytes blancs et améliore la fonction de la graisse brune par l'activation de PPARα et PPARγ. Ces résultats indiquent que ABHD6 est une cible prometteuse pour le développement de thérapies contre l'obésité, le diabète de type 2 et le syndrome métabolique.

ADRENERGIC RECEPTOR GENE EXPRESSION DURING PANCREATIC REGENERATION AND INSULIN SECRETION IN RATS –

In the present study, the changes in the brain EPI (Epinephrine), adrenergic receptors and the receptor gene expression were investigated during pancreatic regeneration and insulin secretion. The changes in the pancreatic islet EPI and adrenergic receptors were also studied in the pancreatectomised rats. The regulatory function of EPI in association with Epidermal growth factor (EGF) and glucose were investigated in rat islet cultures. In vitro studies were carried out using antagonists for adrenergic receptor subtypes to see their involvement in the islet DNA synthesis. The mechanism by which the peripheral EPI regulate insulin secretion was also investigated by studying the nuclear binding proteins in the pancreatic islets during pancreatic regeneration and diabetes. The study reveals that EPI can regulate the pancreatic islet cell proliferation by controlling the insulin synthesis and secretion. The brain adrenergic receptor gene expression and functional correlation regulate the pancreatic adrenergic receptors. The functional balance of α and β-adrenergic receptors controls the insulin secretion and pancreatic β-cell proliferation, which will have immense clinical significance in the treatment of Diabetes mellitus.

Increased Insulin Secretion by Muscarinic M1 and M3 Receptor Function from Rat Pancreatic Islets in vitro

Parasympathetic system plays an important role in insulin secretion from the pancreas. Cholinergic effect on pancreatic beta cells exerts primarily through muscarinic receptors. In the present study we investigated the specific role of muscarinic M1 and M3 receptors in glucose induced insulin secretion from rat pancreatic islets in vitro. The involvement of muscarinic receptors was studied using the antagonist atropine. The role of muscarinic MI and M3 receptor subtypes was studied using subtype specific antagonists. Acetylcholine agonist, carbachol, stimulated glucose induced insulin secretion at low concentrations (10-8-10-5 M) with a maximum stimulation at 10-7 M concentration. Carbachol-stimulated insulin secretion was inhibited by atropine confirming the role of muscarinic receptors in cholinergic induced insulin secretion. Both M1 and M3 receptor antagonists blocked insulin secretion induced by carbachol. The results show that M3 receptors are functionally more prominent at 20 mM glucose concentration when compared to MI receptors. Our studies suggest that muscarinic M1 and M3 receptors function differentially regulate glucose induced insulin secretion, which has clinical significance in glucose homeostasis.

Alterations in the muscarinic M I and M3 receptor gene expression in the brain stem during pancreatic regeneration and insulin secretion in weanling rats

Muscarinic M1 and M3 receptor changes in the brain stem during pancreatic regeneration were investigated. Brain stem acetylcholine esterase activity decreased at the time of regeneration . Sympathetic activity also decreased as indicated by the norepinephrine (NE) and epinephrine (EPI) content of adrenals and also in the plasma. Muscarinic Ml and M3 receptors showed reciprocal changes in the brain stem during regeneration. Muscairnic M1 receptor number decreased at time of regeneration without any change in the affinity. High affinity M3 receptors showed an increase in the number. The affinity did not show any change . The number of low affinity receptors decreased with decreased Kd at 72 hours after partial pancreatectomy. The Kd reversed to control value with a reversal of the number of receptors to near control value . Gene expression studies also showed a similar change in the mRNA level of Ml and M3 receptors . These alterations in the muscarinic receptors regulate sympathetic activity and maintain glucose level during pancreatic regeneration. Central muscarinic M1 and M3 receptor subtypes functional balance is suggested to regulate sympathetic and parasympathetic activity, which in turn control the islet cell proliferation and glucose homeostasis.

Amygdala and ventromedial prefrontal cortex are inversely coupled during regulation of negative affect and predict the diurnal pattern of cortisol secretion among older adults

Among younger adults, the ability to willfully regulate negative affect, enabling effective responses to stressful experiences, engages regions of prefrontal cortex (PFC) and the amygdala. Because regions of PFC and the amygdala are known to influence the hypothalamic-pituitary-adrenal axis, here we test whether PFC and amygdala responses during emotion regulation predict the diurnal pattern of salivary cortisol secretion. We also test whether PFC and amygdala regions are engaged during emotion regulation in older (62- to 64-year-old) rather than younger individuals. We measured brain activity using functional magnetic resonance imaging as participants regulated (increased or decreased) their affective responses or attended to negative picture stimuli. We also collected saliva samples for 1 week at home for cortisol assay. Consistent with previous work in younger samples, increasing negative affect resulted in ventral lateral, dorsolateral, and dorsomedial regions of PFC and amygdala activation. In contrast to previous work, decreasing negative affect did not produce the predicted robust pattern of higher PFC and lower amygdala activation. Individuals demonstrating the predicted effect (decrease s attend in the amygdala), however, exhibited higher signal in ventromedial prefrontal cortex (VMPFC) for the same contrast. Furthermore, participants displaying higher VMPFC and lower amygdala signal when decreasing compared with the attention control condition evidenced steeper, more normative declines in cortisol over the course of the day. Individual differences yielded the predicted link between brain function while reducing negative affect in the laboratory and diurnal regulation of endocrine activity in the home environment.

Transcriptional regulators of the GAD acid stress island are carried by effector protein-encoding prophages and indirectly control type III secretion in enterohemorrhagic Escherichia coli O157:H7

P>Type III secretion (T3S) plays a pivotal role in the colonization of ruminant hosts by Enterohemorrhagic Escherichia coli (EHEC). The T3S system translocates effector proteins into host cells to promote bacterial attachment and persistence. The repertoire and variation in prophage regions underpins differences in the pathogenesis and epidemiology of EHEC strains. In this study, we have used a collection of deletions in cryptic prophages and EHEC O157 O-islands to screen for novel regulators of T3S. Using this approach we have identified a family of homologous AraC-like regulators that indirectly repress T3S. These prophage-encoded secretion regulator genes (psr) are found exclusively on prophages and are associated with effector loci and the T3S activating Pch family of regulators. Transcriptional profiling, mutagenesis and DNA binding studies were used to show that these regulators usurp the conserved GAD acid stress resistance system to regulate T3S by increasing the expression of GadE (YhiE) and YhiF and that this regulation follows attachment to bovine epithelial cells. We further demonstrate that PsrA and effectors encoded within cryptic prophage CP933-N are required for persistence in a ruminant model of colonization.

Effects of the aromatase inhibitor Fadrozole on plasma sex steroid secretion and oocyte maturation in female coho salmon (Oncorhynchus kisutch) during vitellogenesis

17β-estradiol, 17α-20β-dihydroxy-4-pregnen-3-one (17α-20β-P), and testosterone levels were measured in plasma samples obtained from vitellogenic coho salmon (Oncorhynchus kisutch) before and 32 days after injection of the aromatase inhibitor Fadrozole (AI). Plasma 17β-estradiol levels decreased significantly 6 h after injection in all AI treated fish. The higher the dose the longer the maintenance of low plasma 17β-estradiol levels. Inversely, plasma 17α-20β-P increased significantly 6 h after injection in all AI treated fish, and the higher the dose the longer the maintenance of high plasma 17α-20β-P levels. At 48 h after injection plasma testosterone levels were significantly higher in the AI treated groups. The oocyte maturation index showed that multiple injections with AI retarded oocyte development. Besides, oocyte diameter and GSI were lower in the same group, which presented high incidence of atresia of vitellogenic oocytes. The ovarian follicles and brain of the fish which received multiple injections secreted less 17β-estradiol, in vitro. These findings suggest that aromatase inhibitors such as Fadrozole may have a potential as a tool to regulate sexual development in salmon.