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Signalling Events (signalling + event)
Selected AbstractsSignalling events involved in interferon-,-inducible macrophage nitric oxide generationIMMUNOLOGY, Issue 4 2003Julie Blanchette Summary Nitric oxide (NO) produced by macrophages (M,) in response to interferon-, (IFN-,) plays a pivotal role in the control of intracellular pathogens. Current knowledge of the specific biochemical cascades involved in this IFN-,-inducible M, function is still limited. In the present study, we evaluated the participation of various second messengers , Janus kinase 2 (JAK2), signal transducer and activator of transcription (STAT) 1,, MAP kinase kinase (MEK1/2), extracellular signal-regulated kinases 1 and 2 (Erk1/Erk2) and nuclear factor kappa B (NF-,B) , in the regulation of NO production by IFN-,-stimulated J774 murine M,. The use of specific signalling inhibitors permitted us to establish that JAK2/STAT1,- and Erk1/Erk2-dependent pathways are the main players in IFN-,-inducible M, NO generation. To determine whether the inhibitory effect was taking place at the pre- and/or post-transcriptional level, we evaluated the effect of each antagonist on inducible nitric oxide synthase (iNOS) gene and protein expression, and on the capacity of IFN-, to induce JAK2, Erk1/Erk2 and STAT1, phosphorylation. All downregulatory effects occurred at the pretranscriptional level, except for NF-,B, which seems to exert its role in NO production through an iNOS-independent event. In addition, electrophoretic mobility shift assay (EMSA) analysis revealed that STAT1, is essential for IFN-,-inducible iNOS expression and NO production, whereas the contribution of NF-,B to this cellular regulation seems to be minimal. Moreover, our data suggest that Erk1/Erk2 are responsible for STAT1, Ser727 residue phosphorylation in IFN-,-stimulated M,, thus contributing to the full activation of STAT1,. Taken together, our results indicate that JAK2, MEK1/2, Erk1/Erk2 and STAT1, are key players in the IFN-,-inducible generation of NO by M,. [source] A novel bacterial signalling system with a combination of a Ser/Thr kinase cascade and a His/Asp two-component systemMOLECULAR MICROBIOLOGY, Issue 2 2005Renate Lux Summary Prokaryotes and eukaryotes have long been thought to use very different types of kinases (the His kinases of the ,bacterial' two-component systems versus the ,eukaryotic' Ser/Thr/Tyr kinases) to carry out signal transduction. This paradigm no longer holds true, because both systems are now found together in an increasing number of prokaryotic organisms and ,two-component' His kinase are present in eukaryotes. Pioneering work on bacterial protein serine threonine kinases (PSTKs) has been performed in Myxococcus xanthus, a soil bacterium with a complex life cycle that possesses orthologues of signalling-related kinases ,typical' of both the prokaryotic and the eukaryotic kingdoms. In the work reported in this volume of Molecular Microbiology, Nariya and Inouye describe a PSTK cascade that modulates the biochemical activity of MrpC, a CRP-like transcriptional regulator for essential developmental signalling pathways in M. xanthus whose transcription is under the control of a two-component system. This is the first report of both a functional PSTK cascade in bacteria and the use of both PSTK and two-component systems to control a single complex bacterial signalling event. [source] The I-antigens of Ichthyophthirius multifiliis are GPI-Anchored ProteinsTHE JOURNAL OF EUKARYOTIC MICROBIOLOGY, Issue 3 2001THEODORE G. CLARK ABSTRACT. The parasitic ciliate Ichthyophthirius multifiliis has abundant surface membrane proteins (i-antigens) that when clustered, trigger rapid, premature exit from the host. Similar antigens are present in free-living ciliates and are GPI-anchored in both Paramecium and Tetrahymena. Although transmembrane signalling through GPI-anchored proteins has been well-documented in metazoan cells, comparable phenomena have yet to be described in protists. Since premature exit of Ichthyophthirius is likely to involve a transmembrane signalling event, we sought to determine whether i-antigens are GPI-anchored in these cells as well. Based on their solubility properties in Triton X-114, the i-antigens of Ichthyophthirius are amphiphilic in nature and partition with the detergent phase. Nevertheless, following treatment of detergent lysates with phospholipase C, the same proteins become hydrophilic. Concomitantly, they are recognized by antibodies against a cross-reacting determinant exposed on virtually all GPI-anchored proteins following cleavage with phospholipase C. Finally, when expressed in recombinant form in Tetrahymena thermophila, full-length i-antigens are restricted to the membrane, while those lacking hydrophobic C-termini are secreted from the cell. Taken together, these observations argue strongly that the i-antigens of Ichthyophthirius multifiliis are, in fact, GPI-anchored proteins. [source] A novel mutant phenotype implicates dicephalic in cyst formation in the Drosophila ovaryDEVELOPMENTAL DYNAMICS, Issue 4 2006Ruth McCaffrey Abstract The establishment of polarity in Drosophila requires the correct specification of the oocyte in early stages of oogenesis, its positioning at the posterior of the egg chamber, and signalling events between the oocyte and the adjacent posterior follicle cells. As a consequence, the anterior-posterior and the dorsal-ventral axes are fixed. The posterior localisation of the oocyte depends on cadherin-mediated adhesion between the oocyte and the follicle cells. Here we show that dicephalic mutants affect the posterior positioning of the oocyte without interfering with oocyte specification in the germarium. Unlike other mutants that also affect the posterior placement of the oocyte, dicephalic mutants affect neither gurken expression nor karyosome formation during meiosis. By analysing in detail the mutant phenotypes of dicephalic, we find that cyst formation in mutant germaria is defective and that it shares some similarities with cysts that lack DE-cadherin in the germline cells. We propose a model in which dicephalic is involved in the proper adhesion between the oocyte and the somatic follicle cells. Developmental Dynamics 235:908,917, 2006. © 2005 Wiley-Liss, Inc. [source] Role for cAMP-protein kinase A signalling in augmented neutrophil adhesion and chemotaxis in sickle cell diseaseEUROPEAN JOURNAL OF HAEMATOLOGY, Issue 4 2007Andreia A. Canalli Abstract The significance of the leukocyte in sickle cell disease (SCD) pathophysiology is becoming increasingly recognised; we sought to examine whether the chemotactic properties of neutrophils of SCD individuals may be altered and, further, to better understand the signalling events that mediate altered SCD neutrophil function. Adhesion to immobilised fibronectin (FN) and chemotaxis of control and SCD neutrophils were assessed using in vitro static adhesion assays and 96-well chemotaxis chamber assays. Adhesion assays confirmed a significantly higher basal adhesion of SCD neutrophils to FN, compared with control neutrophils. Chemotaxis assays established, for the first time, that SCD neutrophils demonstrate greater spontaneous migration and, also, augmented migration in response to IL-8, when compared with control neutrophils. Co-incubation of SCD neutrophils with KT5720 (an inhibitor of PKA) abrogated increased basal SCD neutrophil adhesion, spontaneous chemotaxis and IL-8-stimulated chemotaxis. Stimulation of SCD neutrophils with IL-8 also significantly augmented SCD neutrophil adhesion to FN with a concomitant increase in cAMP levels and this increase in adhesion was abolished by KT5720. Interestingly, the adhesive properties of neutrophils from SCD individuals on hydroxyurea therapy were not significantly altered and results indicate that a reduction in intracellular cAMP may contribute to lower the adhesive properties of these cells. Data indicate that up-regulated cAMP signalling plays a significant role in the altered adhesive and migratory properties in SCD neutrophils. Such alterations may have important implications for the pathophysiology of the disease and the cAMP-PKA pathway may represent a therapeutic target for the abrogation of altered leukocyte function. [source] Emerging topics in Reelin functionEUROPEAN JOURNAL OF NEUROSCIENCE, Issue 9 2010Eckart Förster Abstract Reelin signalling in the early developing cortex regulates radial migration of cortical neurons. Later in development, Reelin promotes maturation of dendrites and dendritic spines. Finally, in the mature brain, it is involved in modulating synaptic function. In recent years, efforts to identify downstream signalling events induced by binding of Reelin to lipoprotein receptors led to the characterization of novel components of the Reelin signalling cascade. In the present review, we first address distinct functions of the Reelin receptors Apoer2 and Vldlr in cortical layer formation, followed by a discussion on the recently identified downstream effector molecule n-cofilin, involved in regulating actin cytoskeletal dynamics required for coordinated neuronal migration. Next, we discuss possible functions of the recently identified Reelin,Notch signalling crosstalk, and new aspects of the role of Reelin in the formation of the dentate radial glial scaffold. Finally, progress in characterizing the function of Reelin in modulating synaptic function in the adult brain is summarized. The present review has been inspired by a session entitled ,Functions of Reelin in the developing and adult hippocampus', held at the Spring Hippocampal Research Conference in Verona/Italy, June 2009. [source] Death-associated protein kinase (DAPK) and signal transduction: blebbing in programmed cell deathFEBS JOURNAL, Issue 1 2010Miia Bovellan Death-associated protein kinase (DAPK) regulates many distinct signalling events, including apoptosis, autophagy and membrane blebbing. The role of DAPK in the blebbing process is only beginning to be understood and, in this review, we will first summarize what is known about the cytoskeletal proteins and signalling cascades that participate in bleb growth and retraction and then highlight how DAPK integrates with these processes. Membrane blebs are quasispherical cellular protrusions that have a lifetime of approximately 2 min. During expansion, blebs are initially devoid of actin, although actomyosin contractions provide the motive force for growth. Once growth slows, an actin cortex reforms and actin-bundling and contractile proteins are recruited. Finally, myosin contraction powers bleb retraction into the cell body. Blebbing occurs in a variety of cell types, from cancerous cells to embryonic cells, and can be seen in cellular phenomena as diverse as cell spreading, movement, cytokinesis and cell death. Although the machinery that executes this is still undefined in detail, the conservation of blebbing phenomenon suggests a fundamental role in metazoans and DAPK offers a door to further dissect this fascinating process. [source] Listeriolysin O: a key protein of Listeria monocytogenes with multiple functionsFEMS MICROBIOLOGY REVIEWS, Issue 4 2006Samer Kayal Abstract Cholesterol-dependent cytolysins (CDCs) are produced by a large number of pathogenic Gram-positive bacteria. Most of these single-chain proteins are secreted in the extracellular medium. Among the species producing CDCs, only two species belonging to the genus Listeria (Listeria monocytogenes and Listeria ivanovii) are able to multiply intracellularly and release their toxins in the phagosomal compartment of the infected host cell. This review provides an updated overview on the importance of listeriolysin O (LLO) in the pathogenicity of L. monocytogenes, focusing mainly on two aspects: (1) the structure,function relationship of LLO and (2) its role in intra- and extracellular signalling. We first examine the specific sequence determinants, or protein domains, that make this cytolysin so well adapted to the intracellular lifestyle of L. monocytogenes. The roles that LLO has in cellular signalling events in the context of relations to pathogenesis are also discussed. [source] Novel insights into the osmotic stress response of yeastFEMS YEAST RESEARCH, Issue 3 2002Willem H Mager Abstract Response to hyperosmolarity in the baker's yeast Saccharomyces cerevisiae has attracted a great deal of attention of molecular and cellular biologists in recent years, from both the fundamental scientific and applied viewpoint. Indeed the underlying molecular mechanisms form a clear demonstration of the intricate interplay of (environmental) signalling events, regulation of gene expression and control of metabolism that is pivotal to any living cell. In this article we briefly review the cellular response to conditions of hyperosmolarity, with focus on the high-osmolarity glycerol mitogen-activated protein kinase pathway as the major signalling route governing cellular adaptations. Special attention will be paid to the recent finding that in the yeast cell also major structural changes occur in order to ensure maintenance of cell integrity. The intriguing role of glycerol in growth of yeast under (osmotic) stress conditions is highlighted. [source] Direct and indirect manipulation of the MEK-ERK pathway regulates the formation of a pericellular HA-dependent matrix by chick articular surface cells without modifying CD44 expresssionINTERNATIONAL JOURNAL OF EXPERIMENTAL PATHOLOGY, Issue 4 2004Edward R. Bastow Introduction Recent evidence suggests that hyaluronan (HA) facilitates the mechano-dependent joint cavity-forming process through the elaboration and retention of a HA-rich pericellular matrix in the developing joint interzone (IZ). The presumptive joint IZ phenotype shows a capacity to bind and synthesize HA and also exhibits elevated activated ERK, prior to synovial joint cavity formation (Lamb et al. 2001; Edwards et al. 1994; Dowthwaite et al. 1998). We have found that immobilization, which induces embryonic joint fusion with loss of the joint IZ phenotype, also reduces ERK activity levels in the IZ. As the signalling events regulating the synthesis and binding of HA have yet to be determined, we hypothesize that ERK activation plays a pivotal role in determining the presumptive joint IZ phenotype through HA synthetic and binding capacity. Materials and methods Chick articular surface (AS) cells were harvested from proximal tibiotarsal joints of embryos by collagenase digestion. Pericellular coat formation was assessed using the erythrocyte exclusion assay and cell-coat area ratios determined. ERK activity was modulated by transient transfection of GFP constructs of constitutively active (CA-) or dominant negative (DN-) forms of MEK, the direct upstream regulator of ERK or by treatment with the MEK inhibitor PD98059 (50 µm). ERK activation was monitored by immunochemistry. CD44 expression and ERK activation in PD98059-treated cells were monitored by immunoblotting and medium HA concentrations by ELISA. Results AS cells form large pericellular coats that are lost following hyaluronidase treatment and thus dependent upon HA for their construction. Treatment with PD98059 significantly reduced pericellular coat formation after 6 h. In parallel, we confirmed that PD98059 diminished active ERK expression without modifying overall levels of ERK, suggesting that the elaboration of large HA-pericellular coats is dependent upon MEK's activation of ERK. Western blot analysis of PD98059-treated cells showed that loss of pericellular coats was not, however, associated with any decreased levels of the cell surface HA receptor CD44. Although treatment with PD98059 did not change medium HA concentration after short times of exposure, at times (up to 6 h) during which coat loss was evident, prolonged treatment over 24 h significantly decreased medium HA concentration. Consistent with a role for ERK in pericellular coat formation, transfection with DN-MEK diminished, while CA-MEK increased, both active ERK expression and coat formation efficiency. We also found that, commensurate with this modification in coat forming efficiency, cells expressing DN-MEK exhibited a significant reduction in labelling of free HA on the cell surface. Discussion These studies extend our recent work to indicate that: (i) direct modulation of ERK activation by transfection with its endogenous upstream regulator modifies cell surface-associated HA (ii) PD98059-induced blockade of ERK activation restricts medium HA release and (iii) ERK-mediated changes in pericellular coat elaboration are independent of changes in cellular CD44 expression. These findings suggest an intimate relationship between ERK activation and the formation/retention of HA-rich pericellular matrices in vitro and highlight the role for ERK activation in regulating joint line-related differentiation. [source] Defining boundaries during joint cavity formation: going out on a limbINTERNATIONAL JOURNAL OF EXPERIMENTAL PATHOLOGY, Issue 2 2003K. J. Lamb Summary., Whilst factors controlling the site at which joints form within the developing limb are recognised, the mechanisms by which articular element separation occurs during the formation of the joint cavity have not been determined. Herein, we review the relationships between early limb patterning, embryonic movement, extracellular matrix composition, local signalling events and the process of joint cavity formation. We speculate that a pivotal event in this process involves the demarcation of signalling boundaries, established by local mechano-dependent modifications in glycosaminoglycan synthesis. In our opinion, studies that examine early patterning and also focus on local developmental alterations in tissue architecture are required in order to help elucidate the fundamental principals regulating joint formation. [source] Spatial insulin signalling in isolated skeletal muscle preparationsJOURNAL OF CELLULAR BIOCHEMISTRY, Issue 5 2010Peter Sogaard Abstract During in vitro incubation in the absence or presence of insulin, glycogen depletion occurs in the inner core of the muscle specimen, concomitant with increased staining of hypoxia-induced-factor-1-alpha and caspase-3, markers of hypoxia and apoptosis, respectively. The aim of this study was to determine whether insulin is able to diffuse across the entire muscle specimen in sufficient amounts to activate signalling cascades to promote glucose uptake and glycogenesis within isolated mouse skeletal muscle. Phosphoprotein multiplex assay on lysates from muscle preparation was performed to detect phosphorylation of insulin-receptor on Tyr1146, Akt on Ser473 and glycogen-synthases-kinase-3 on Ser21/Ser9. To address the spatial resolution of insulin signalling, immunohistochemistry studies on cryosections were performed. Our results provide evidence to suggest that during the in vitro incubation, insulin sufficiently diffuses into the centre of tubular mouse muscles to promote phosphorylation of these signalling events. Interestingly, increased insulin signalling was observed in the core of the incubated muscle specimens, correlating with the location of oxidative fibres. In conclusion, insulin action was not restricted due to insufficient diffusion of the hormone during in vitro incubation in either extensor digitorum longus or soleus muscles from mouse under the specific experimental settings employed in this study. Hence, we suggest that the glycogen depleted core as earlier observed is not due to insufficient insulin action. J. Cell. Biochem. 109: 943,949, 2010. © 2010 Wiley-Liss, Inc. [source] von Willebrand factor activates endothelial nitric oxide synthase in blood platelets by a glycoprotein Ib-dependent mechanismJOURNAL OF THROMBOSIS AND HAEMOSTASIS, Issue 12 2006R. RIBA Summary.,Background: The molecular regulation of endothelial nitric oxide synthase (eNOS) in blood platelets and the signalling events induced by platelet-derived NO are poorly defined. In particular, the ability of von Willebrand factor (VWF) to stimulate cyclic guanosine monophosphate (cGMP) formation in platelets has produced conflicting data. Objectives: To determine the mechanisms leading to eNOS activation and clarify the downstream signaling pathways activated by platelet-derived NO in response to VWF. Methods: We used three independent markers of NO signaling, [3H] l -citrulline production, cGMP accrual and immunoblotting of vasodilator,stimulated phosphoprotein (VASP) to examine the NO signaling cascade in response to VWF. Results: VWF increased NO synthesis and bioavailability, as evidenced by increased [3H] l -citrulline production and cGMP accrual, respectively. VWF-induced eNOS activation was GPIb-IX-dependent and independent of integrin ,IIb,3. cGMP formation in response to VWF required Ca2+ mobilization, Src family kinases, phosphatidylinositol 3-kinase and phospholipase C, but not protein kinase C. This suggests that a cross-talk between the signaling mechanisms regulates platelet activation and NO synthesis. VWF-induced cGMP accrual was completely blocked by apyrase and indomethacin, demonstrating an essential role for platelet-derived ADP and thromboxane A2 (TxA2). Elevated cGMP levels led to increased VASP phosphorylation at serine239 that was both protein kinase G (PKG)- and protein kinase A (PKA)-dependent. Conclusions: We demonstrate that VWF activates eNOS through a specific Ca2+ -dependent GPIb receptor-signaling cascade that relies on the generation of platelet-derived ADP and TxA2. Furthermore, we provide the first evidence to suggest that platelet derived-NO/cGMP activates PKA in addition to PKG. [source] Disruption of a Plasmodium falciparum cyclic nucleotide phosphodiesterase gene causes aberrant gametogenesisMOLECULAR MICROBIOLOGY, Issue 1 2008Cathy J. Taylor Summary Phosphodiesterase (PDE) and guanylyl cyclase (GC) enzymes are key components of the cGMP signalling pathway and are encoded in the genome of Plasmodium falciparum. Here we investigate the role of specific GC and PDE isoforms in gamete formation , a process that is essential for malaria transmission and occurs in the Anopheles mosquito midgut following feeding on an infected individual. Details of the intracellular signalling events controlling development of the male and female gametes from their precursors (gametocytes) remain sparse in P. falciparum. Previous work involving the addition of pharmacological agents to gametocytes implicated cGMP in exflagellation , the emergence of highly motile, flagellated male gametes from the host red blood cell. In this study we show that decreased GC activity in parasites having undergone disruption of the PfGC, gene had no significant effect on gametogenesis. By contrast, decreased cGMP-PDE activity during gametocyte development owing to disruption of the PfPDE, gene, led to a severely reduced ability to undergo gametogenesis. This suggests that the concentration of cGMP must be maintained below a threshold in the developing gametocyte to allow subsequent differentiation to proceed normally. The data indicate that PfPDE, plays a crucial role in regulating cGMP levels during sexual development. [source] Early signalling events in the Avr9/Cf-9-dependent plant defence responseMOLECULAR PLANT PATHOLOGY, Issue 1 2000Tina Romeis Resistance of tomato to the leaf mould fungus Cladosporium fulvum is controlled by the interaction between a plant-encoded resistance gene (Cf-9) and pathogen-encoded avirulence (Avr9) gene. Our objective is to understand the underlying molecular mechanisms that transmit the Cf-9/Avr9-dependent pathogen perception event and activate the plant defence response. Our approach toward the understanding of Cf -function is based on the analysis of early Cf-9/Avr9-mediated responses and signalling events. Because Cf-9 transgenically expressed in tobacco retains its specificity and activity to the Avr9 elicitor, signalling experiments were conducted in the heterologous system using these transgenic lines or derived Cf9 tobacco cell cultures. Among the earliest responses to the Avr9/Cf-9 elicitation event were rapid changes in ion-fluxes, the synthesis of active oxygen species (AOS), probably catalysed by a plant NADPH-oxidase, and the transient activation of two MAP kinases. These kinases were identified as WIPK (wounding-induced protein kinase) and SIPK (salicylic-acid induced kinase) from tobacco. Studies with pharmacological inhibitors suggested that the MAP kinases are located in an independent signalling pathway from the Avr9/Cf-9-dependent synthesis of AOS. SIPK and WIPK were involved in pathogen-related elicitation processes as well as in abiotic stress responses. This indicates that the plant defence is triggered via a signalling network that shares components with pathways originating from abiotic environmental stress stimuli. [source] Nitric oxide-induced phosphatidic acid accumulation: a role for phospholipases C and D in stomatal closurePLANT CELL & ENVIRONMENT, Issue 2 2008AYELEN M. DISTÉFANO ABSTRACT Stomatal closure is regulated by a complex network of signalling events involving numerous intermediates, among them nitric oxide (NO). Little is known about the signalling events occurring downstream of NO. Previous studies have shown that NO modulates cytosolic calcium concentration and the activation of plasma membrane ion channels. Here we provide evidence that supports the involvement of the lipid second messenger phosphatidic acid (PA) in NO signalling during stomatal closure. PA levels in Vicia faba epidermal peels increased upon NO treatment to maximum levels within 30 min, subsequently decreasing to control levels at 60 min. PA can be generated via phospholipase D (PLD) or via phospholipase C (PLC) in concerted action with diacylglycerol kinase (DGK). Our results showed that NO-induced PA is produced via the activation of both pathways. NO-induced stomatal closure was blocked either when PLC or PLD activity was inhibited. We have shown that PLC- and PLD-derived PA represents a downstream component of NO signalling cascade during stomatal closure. [source] Changes in mitogen-activated protein kinase activity occur in the maize pulvinus in response to gravistimulation and are important for the bending responsePLANT CELL & ENVIRONMENT, Issue 7 2003A. M. CLORE ABSTRACT The maize (Zea mays L.) pulvinus was used as a model system to study the signalling events that lead to differential growth in response to gravistimulation in plants. The pulvinus functions to return tipped plants to vertical via differential elongation of the cells on its lower side. By performing immunokinase assays using total soluble protein extracts and an antibody against mammalian ERK1, a mitogen-activated protein kinase (MAPK)-like activity was assayed in pulvini halves harvested at various time points after tipping. We detected a reproducible alternation of higher levels of activity occurring between the upper and lower halves of the pulvinus between 75 and 180 min after tipping, with a sustained increase in the upper half occurring at the end of the time-course. This timing roughly corresponds to the presentation time for maize (i.e. the amount of time that the plant needs to be tipped before it is committed to bend), which occurs between 2 and 4 h. Treatment of maize stem explants with an inhibitor of MAPK activation, U0126, led to a reduction in the activity of this kinase, as well as an almost 65% reduction in bending as measured at 20 h. Rinsing out of the inhibitor resulted in recovery of both bending and kinase activity. It is possible that changes in MAPK activity in the gravistimulated pulvinus are part of a signalling cascade that may help to distinguish between minor perturbations in plant orientation and more significant and long-term changes, and may also help to determine the direction of bending. [source] Thymic stromal cells and positive selectionAPMIS, Issue 7-8 2001Ann R. Chidgey The intrathymic differentiation events leading to the development and export of mature T cells tolerant to self yet capable of responding to foreign peptide antigen in the context of self-MHC are clearly both dynamic and complex. The changing phenotype of the developing thymocyte as it migrates through and interacts with the heterogeneous thymic microenvironment and the intracellular signalling events associated with such interactions are being extensively studied, yet many aspects remain poorly defined, such as the precise relationship between stromal cells and thymic selection. Positive and negative selection are crucial events in the development of T cells, leading to a diverse yet non-autoreactive immune system. A breakdown in either of these processes could lead to either a reduced T cell repertoire or the escape into the periphery of autoreactive T cells , both clearly having deleterious consequences for the health of the individual. This review aims to summarise the current status of research in thymic positive selection with emphasis on the role of different cell types and peptides. [source] Reduced signal transduction by 5-HT4 receptors after long-term venlafaxine treatment in ratsBRITISH JOURNAL OF PHARMACOLOGY, Issue 3 2010R Vidal BACKGROUND AND PURPOSE The 5-HT4 receptor may be a target for antidepressant drugs. Here we have examined the effects of the dual antidepressant, venlafaxine, on 5-HT4 receptor-mediated signalling events. EXPERIMENTAL APPROACH The effects of 21 days treatment (p.o.) with high (40 mg·kg,1) and low (10 mg·kg,1) doses of venlafaxine, were evaluated at different levels of 5-HT4 receptor-mediated neurotransmission by using in situ hybridization, receptor autoradiography, adenylate cyclase assays and electrophysiological recordings in rat brain. The selective noradrenaline reuptake inhibitor, reboxetine (10 mg·kg,1, 21 days) was also evaluated on 5-HT4 receptor density. KEY RESULTS Treatment with a high dose (40 mg·kg,1) of venlafaxine did not alter 5-HT4 mRNA expression, but decreased the density of 5-HT4 receptors in caudate-putamen (% reduction = 26 ± 6), hippocampus (% reduction = 39 ± 7 and 39 ± 8 for CA1 and CA3 respectively) and substantia nigra (% reduction = 49 ± 5). Zacopride-stimulated adenylate cyclase activation was unaltered following low-dose treatment (10 mg·kg,1) while it was attenuated in rats treated with 40 mg·kg,1 of venlafaxine (% reduction = 51 ± 2). Furthermore, the amplitude of population spike in pyramidal cells of CA1 of hippocampus induced by zacopride was significantly attenuated in rats receiving either dose of venlafaxine. Chronic reboxetine did not modify 5-HT4 receptor density. CONCLUSIONS AND IMPLICATIONS Our data indicate a functional desensitization of 5-HT4 receptors after chronic venlafaxine, similar to that observed after treatment with the classical selective inhibitors of 5-HT reuptake. [source] Induction of insulin-like growth factor-I by interleukin-17F in bronchial epithelial cellsCLINICAL & EXPERIMENTAL ALLERGY, Issue 7 2010M. Kawaguchi Summary Cite this as: M. Kawaguchi, J. Fujita, F. Kokubu, G. Ohara, S-K Huang, S. Matsukura, Y. Ishii, M. Adachi, H. Satoh and N. Hizawa, Clinical & Experimental Allergy, 2010 (40) 1036,1043. Background Increased expression of IL-17F has been noted in the airway of asthmatic patients, but its role in asthma has not been fully elucidated. Insulin-like growth factor-I (IGF-I) is known to be involved in airway remodelling and inflammation, while its regulatory mechanisms remain to be defined. Objective To further clarify the biological function of IL-17F, we investigated whether IL-17F is able to regulate the expression of IGF-I in bronchial epithelial cells. Methods Bronchial epithelial cells were stimulated with IL-17F in the presence or absence of T-helper type 2 cytokines. Various kinase inhibitors were added to the culture to identify the key signalling events leading to the expression of IGF-I, in conjunction with the use of short interfering RNAs (siRNAs) targeting mitogen- and stress-activated protein kinase (MSK) 1, p90 ribosomal S6 kinase (p90RSK), and cyclic AMP response element-binding protein (CREB). Results IL-17F significantly induced IGF-I gene and protein expression, and co-stimulation with IL-4 and IL-13 augmented its production. MAP kinase kinase (MEK) inhibitors and the Raf1 kinase inhibitor significantly inhibited IGF-I production, and the combination of PD98059 and Raf1 kinase inhibitor showed further inhibition. Overexpression of Raf1 and Ras dominant-negative mutants inhibited its expression. MSK1 inhibitors significantly blocked IL17F-induced IGF-I expression. Moreover, transfection of the siRNAs targeting MSK1, p90RSK, and CREB blocked its expression. Conclusions In bronchial epithelial cells, IL-17F is able to induce the expression of IGF-I via the Raf1,MEK1/2,ERK1/2,MSK1/p90RSK,CREB pathway in vitro. [source] The effect of ageing on macrophage Toll-like receptor-mediated responses in the fight against pathogensCLINICAL & EXPERIMENTAL IMMUNOLOGY, Issue 3 2010C. R. Dunston Summary The cellular changes during ageing are incompletely understood yet immune system dysfunction is implicated in the age-related decline in health. The acquired immune system shows a functional decline in ability to respond to new pathogens whereas serum levels of cytokines are elevated with age. Despite these age-associated increases in circulating cytokines, the function of aged macrophages is decreased. Pathogen-associated molecular pattern receptors such as Toll-like receptors (TLRs) are vital in the response of macrophages to pathological stimuli. Here we review the evidence for defective TLR signalling in normal ageing. Gene transcription, protein expression and cell surface expression of members of the TLR family of receptors and co-effector molecules do not show a consistent age-dependent change across model systems. However, there is evidence for impaired downstream signalling events, including inhibition of positive and activation of negative modulators of TLR induced signalling events. In this paper we hypothesize that despite a poor inflammatory response via TLR activation, the ineffective clearance of pathogens by macrophages increases the duration of their activation and contributes to perpetuation of inflammatory responses and ageing. [source] Tumour necrosis factor-alpha (TNF- ,) enhances lymphocyte migration into rheumatoid synovial tissue transplanted into severe combined immunodeficient (SCID) miceCLINICAL & EXPERIMENTAL IMMUNOLOGY, Issue 1 2000S. Wahid Adhesion mechanisms play a major role in the recruitment of peripheral blood lymphocytes (PBL) which characteristically infiltrate rheumatoid arthritis (RA) synovium and other chronically inflamed tissues. Through a sequential series of complex integrated adhesion and signalling events, ,multistep model of migration', specific subsets of PBL are recruited into inflamed tissues. In this process both leucocyte receptors and microvascular endothelial (MVE) counter-receptors play a critical role. The MVE in particular, during an inflammatory state, is the target of various inflammatory mediators that cause the up-regulation of several cell adhesion molecules (CAM). One of the most important factors known to be a powerful inducer of MVE CAM is TNF- ,. Conversely, blocking TNF- , causes a down-modulation of CAM expression. To test directly the capacity of TNF- , to induce cell migration into RA synovium we adapted a model in which synovial grafts were implanted into SCID mice subcutaneously. Using this model we demonstrate that: (i) transplants remain viable and become vascularized and fed by mouse subdermal vessels; (ii) the mouse vasculature connects to the transplant vasculature which maintains the ability to express human CAM; (iii) intragraft injections of TNF- , up-regulate the expression of human CAM, following the down-regulation which occurred 4 weeks post-transplantation; and (iv) the up-regulation of graft CAM is associated with increased human PBL migration into the transplants. This study provides direct evidence in vivo of the capacity of TNF- , to induce cell migration. In addition, it provides the experimental background for the optimal use of this model. [source] FUNCTIONAL DIVERSITY OF MAMMALIAN TYPE 2C PROTEIN PHOSPHATASE ISOFORMS: NEW TALES FROM AN OLD FAMILYCLINICAL AND EXPERIMENTAL PHARMACOLOGY AND PHYSIOLOGY, Issue 2 2008Gang Lu SUMMARY 1The Type 2C protein phosphatases (PP2C) represent a highly conserved gene family in the mammalian genome. Recent studies have revealed that PP2C isoforms possess unique patterns of tissue and subcellular distribution associated with diverse functionalities. 2The functional importance of PP2C isoforms has been shown in a plethora of signalling networks controlling cell differentiation, proliferation, growth, survival and metabolism. However, little is known about the regulatory mechanisms of PP2C at the molecular level. It is uncertain how PP2C isoforms are recruited, activated and inactivated during signalling transduction processes. 3In the present paper, an overview of the critical functions of individual PP2C isoforms in regulating cellular signalling events will be provided, along with our perspectives on the challenging issues to be addressed. It is clear that a better understanding of the complex biological effects elicited by specific signalling pathways involving PP2C isoforms has great potential for developing novel therapies for a variety of human diseases, including cancer, diabetes and neural disorders, as well as cardiovascular diseases. [source] | ||||||||||||||||||||||||||||