Glucocorticoid-regulated localization of cell surface glycoproteins in rat hepatoma cells is mediated within the Golgi complex.

Glucocorticoid hormones regulate the post-translational maturation and sorting of cell surface and extracellular mouse mammary tumor virus (MMTV) glycoproteins in M1.54 cells, a stably infected rat hepatoma cell line. Exposure to monensin significantly reduced the proteolytic maturation and externalization of viral glycoproteins resulting in a stable cellular accumulation of a single 70,000-Mr glycosylated polyprotein (designated gp70). Cell surface- and intracellular-specific immunoprecipitations of monensin-treated cells revealed that gp70 can be localized to the cell surface only in the presence of 1 microM dexamethasone, while in uninduced cells gp70 is irreversibly sequestered in an intracellular compartment. Analysis of oligosaccharide processing kinetics demonstrated that gp70 acquired resistance to endoglycosidase H with a half-time of 65 min in the presence or absence of hormone. In contrast, gp70 was inefficiently galactosylated after a 60-min lag in uninduced cells while rapidly acquiring this carbohydrate modification in the presence of dexamethasone. Furthermore, in the absence or presence of monensin, MMTV glycoproteins failed to be galactosylated in hormone-induced CR4 cells, a complement-selected sorting variant defective in the glucocorticoid-regulated compartmentalization of viral glycoproteins to the cell surface. Since dexamethasone had no apparent global effects on organelle morphology or production of total cell surface-galactosylated species, we conclude that glucocorticoids induce the localization of cell surface MMTV glycoproteins by regulating a highly selective step within the Golgi apparatus after the acquisition of endoglycosidase H-resistant oligosaccharide side chains but before or at the site of galactose attachment

Can changes in malaria transmission intensity explain prolonged protection and contribute to high protective efficacy of intermittent preventive treatment for malaria in infants?

BACKGROUND: Intermittent preventive (or presumptive) treatment of infants (IPTi), the administration of a curative anti-malarial dose to infants whether or not they are known to be infected, is being considered as a new strategy for malaria control. Five of the six trials using sulphadoxine-pyrimethamine (SP) for IPTi showed protective efficacies (PEs) against clinical malaria ranging from 20.1 - 33.3% whilst one, the Ifakara study, showed a protective efficacy of 58.6%. MATERIALS AND METHODS: The possible mechanisms that could explain the differences in the reported PE of IPTi were examined by comparing output from a mathematical model to data from the six published IPTi trials. RESULTS: Under stable transmission, the PE of IPTi predicted by the model was comparable with the observed PEs in all but the Ifakara study (ratio of the mean predicted PE to that observed was 1.02, range 0.39 - 1.59). When a reduction in the incidence of infection during the study was included in the model, the predicted PE of IPTi increased and extended into the second year of life, as observed in the Ifakara study. CONCLUSION: A decrease in malaria transmission during the study period may explain part of the difference in observed PEs of IPTi between sites and the extended period of protection into the second year of life observed in the Ifakara study. This finding of continued benefit of interventions in settings of decreasing transmission may explain why rebound of clinical malaria was absent in the large scale trials of insecticide-treated bed nets

Safety and Immunogenicity of a Recombinant Plasmodium falciparum AMA1 Malaria Vaccine Adjuvanted with Alhydrogel™, Montanide ISA 720 or AS02

Contains fulltext : 71100.pdf (publisher's version ) (Open Access)BACKGROUND: Plasmodium falciparum Apical Membrane Antigen 1 (PfAMA1) is a candidate vaccine antigen expressed by merozoites and sporozoites. It plays a key role in red blood cell and hepatocyte invasion that can be blocked by antibodies. METHODOLOGY/PRINCIPAL FINDINGS: We assessed the safety and immunogenicity of recombinant PfAMA1 in a dose-escalating, phase Ia trial. PfAMA1 FVO strain, produced in Pichia pastoris, was reconstituted at 10 microg and 50 microg doses with three different adjuvants, Alhydrogel, Montanide ISA720 and AS02 Adjuvant System. Six randomised groups of healthy male volunteers, 8-10 volunteers each, were scheduled to receive three immunisations at 4-week intervals. Safety and immunogenicity data were collected over one year. Transient pain was the predominant injection site reaction (80-100%). Induration occurred in the Montanide 50 microg group, resulting in a sterile abscess in two volunteers. Systemic adverse events occurred mainly in the AS02 groups lasting for 1-2 days. Erythema was observed in 22% of Montanide and 59% of AS02 group volunteers. After the second dose, six volunteers in the AS02 group and one in the Montanide group who reported grade 3 erythema (>50 mm) were withdrawn as they met the stopping criteria. All adverse events resolved. There were no vaccine-related serious adverse events. Humoral responses were highest in the AS02 groups. Antibodies showed activity in an in vitro growth inhibition assay up to 80%. Upon stimulation with the vaccine, peripheral mononuclear cells from all groups proliferated and secreted IFNgamma and IL-5 cytokines. CONCLUSIONS/SIGNIFICANCE: All formulations showed distinct reactogenicity profiles. All formulations with PfAMA1 were immunogenic and induced functional antibodies. TRIAL REGISTRATION: (Clinicaltrials.gov) NCT00730782

Leptin Resistance in Vagal Afferent Neurons Inhibits Cholecystokinin Signaling and Satiation in Diet Induced Obese Rats

Background and Aims: The gastrointestinal hormone cholecystokinin (CCK) plays an important role in regulating meal size and duration by activating CCK1 receptors on vagal afferent neurons (VAN). Leptin enhances CCK signaling in VAN via an early growth response 1 (EGR1) dependent pathway thereby increasing their sensitivity to CCK. In response to a chronic ingestion of a high fat diet, VAN develop leptin resistance and the satiating effects of CCK are reduced. We tested the hypothesis that leptin resistance in VAN is responsible for reducing CCK signaling and satiation. Results: Lean Zucker rats sensitive to leptin signaling, significantly reduced their food intake following administration of CCK8S (0.22 nmol/kg, i.p.), while obese Zucker rats, insensitive to leptin, did not. CCK signaling in VAN of obese Zucker rats was reduced, preventing CCK-induced up-regulation of Y2 receptor and down-regulation of melanin concentrating hormone 1 receptor (MCH1R) and cannabinoid receptor (CB1). In VAN from diet-induced obese (DIO) Sprague Dawley rats, previously shown to become leptin resistant, we demonstrated that the reduction in EGR1 expression resulted in decreased sensitivity of VAN to CCK and reduced CCK-induced inhibition of food intake. The lowered sensitivity of VAN to CCK in DIO rats resulted in a decrease in Y2 expression and increased CB1 and MCH1R expression. These effects coincided with the onset of hyperphagia in DIO rats. Conclusions: Leptin signaling in VAN is required for appropriate CCK signaling and satiation. In response to high fat feeding

Feeding-dependent activation of enteric cells and sensory neurons by lymphatic fluid: evidence for a neurolymphocrine system

Fulltext embargoed for: 12 months post date of publicationLymphatic fluid is a plasma filtrate that can be viewed as having biological activity through the passive accumulation of molecules from the interstitial fluid. The possibility that lymphatic fluid is part of an active self-contained signaling process that parallels the endocrine system, through the activation of G-protein coupled receptors (GPCR), has remained unexplored. We show that the GPCR lysophosphatidic acid 5 (LPA5) is found in sensory nerve fibers expressing calcitonin gene-related peptide (CGRP) that innervate the lumen of lymphatic lacteals and enteric nerves. Using LPA5 as a model for nutrient-responsive GPCRs present on sensory nerves, we demonstrate that dietary protein hydrolysate (peptone) can induce c-Fos expression in enterocytes and nerves that express LPA5. Mesenteric lymphatic fluid (MLF) mobilizes intracellular calcium in cell models expressing LPA5 upon feeding in a time- and dose-dependent manner. Primary cultured neurons of the dorsal root ganglia expressing CGRP are activated by MLF, which is enhanced upon LPA5 overexpression. Activation is independent of the known LPA5 agonists, lysophosphatidic acid and farnesyl pyrophosphate. These data bring forth a pathway for the direct stimulation of sensory nerves by luminal contents and interstitial fluid. Thus, by activating LPA5 on sensory nerves, MLF provides a means for known and yet to be identified constituents of the interstitial fluid to act as signals to comprise a "neurolymphocrine" system

Therapeutic potential of neuropeptide Y (NPY) receptor ligands

Neuropeptide Y (NPY) is widely distributed in the human body and contributes to a vast number of physiological processes. Since its discovery, NPY has been implicated in metabolic regulation and, although interest in its role in central mechanisms related to food intake and obesity has somewhat diminished, the topic remains a strong focus of research concerning NPY signalling. In addition, a number of other uses for modulators of NPY receptors have been implied in a range of diseases, although the development of NPY receptor ligands has been slow, with no clinically approved receptor therapeutics currently available. Nevertheless, several interesting small molecule compounds, notably Y2 receptor antagonists, have been published recently, fueling optimism in the field. Herein we review the role of NPY in the pathophysiology of a number of diseases and highlight instances where NPY receptor signalling systems are attractive therapeutic targets

Inositol hexakisphosphate increases the size of platelet aggregates

The inositol phosphates, InsP5 and InsP6, have recently been identified as binding partners of fibrinogen, which is critically involved in hemostasis by crosslinking activated platelets at sites of vascular injury. Here, we investigated the putative physiological role of this interaction and found that platelets increase their InsP6 concentration upon stimulation with the PLC-activating agonists thrombin, collagen I and ADP and present a fraction of it at the outer plasma membrane. Cone and plate analysis in whole blood revealed that InsP6 specifically increases platelet aggregate size. This effect is fibrinogen-dependent, since it is inhibited by an antibody that blocks fibrinogen binding to platelets. Furthermore, InsP6 has only an effect on aggregate size of washed platelets when fibrinogen is present, while it has no influence in presence of von Willebrand factor or collagen. By employing blind docking studies we predicted the binding site for InsP6 at the bundle between the γ and β helical subunit of fibrinogen. Since InsP6 is unable to directly activate platelets and it did not exhibit an effect on thrombin formation or fibrin structure, our data indicate that InsP6 might be a hemostatic agent that is produced by platelets upon stimulation with PLC-activating agonists to promote platelet aggregation by supporting crosslinking of fibrinogen and activated platelets

Inositol hexakisphosphate increases the size of platelet aggregates

The inositol phosphates, InsP5 and InsP6, have recently been identified as binding partners of fibrinogen, which is critically involved in hemostasis by crosslinking activated platelets at sites of vascular injury. Here, we investigated the putative physiological role of this interaction and found that platelets increase their InsP6 concentration upon stimulation with the PLC-activating agonists thrombin, collagen I and ADP and present a fraction of it at the outer plasma membrane. Cone and plate analysis in whole blood revealed that InsP6 specifically increases platelet aggregate size. This effect is fibrinogen-dependent, since it is inhibited by an antibody that blocks fibrinogen binding to platelets. Furthermore, InsP6 has only an effect on aggregate size of washed platelets when fibrinogen is present, while it has no influence in presence of von Willebrand factor or collagen. By employing blind docking studies we predicted the binding site for InsP6 at the bundle between the γ and β helical subunit of fibrinogen. Since InsP6 is unable to directly activate platelets and it did not exhibit an effect on thrombin formation or fibrin structure, our data indicate that InsP6 might be a hemostatic agent that is produced by platelets upon stimulation with PLC-activating agonists to promote platelet aggregation by supporting crosslinking of fibrinogen and activated platelets