CLINICAL AND EXPERIMENTAL PHARMACOLOGY AND PHYSIOLOGY, Issue 7 2009
Hou-Qin Xiao
SUMMARY 1Accumulating evidence suggests that vitamin D and its analogues are renoprotective. However, the precise mechanisms and the molecular targets by which active vitamin D exerts its beneficial effects remain obscure. The objective of the present study was to evaluate the effect of active vitamin D on rats with puromycin aminonucleoside (PAN) nephropathy, a model that is characterized by predominant podocyte injury. 2The PAN nephropathy rats were created by a single intravenous injection of 100 mg/kg PAN. Changes in renal pathology and podocyte numbers were observed. Real-time polymerase chain reaction (PCR) was performed to examine mRNA expression of nephrin, transforming growth factor (TGF)-,1 and bone morphogenetic protein (BMP)-7. Protein expression of nephrin, TGF-,1, BMP-7 and p-Smad2/3 and p-Smad1/5/8 was examined by immunofluorescence, immunohistochemistry and western blotting, respectively. Rats were treated with 1,25(OH)2D3 by gastric gavage at a dose of 2.5 µg/kg per day, starting 2 days before PAN injection and continuing throughout the experiment. 3A single injection of PAN induced massive proteinuria and elevated serum creatinine on Day 7, both of which were significantly suppressed by 1,25-dihydroxyvitamin D3 (1,25(OH)2D3). Immunofluorescence and real-time PCR of the podocyte-associated protein nephrin revealed reduced and discontinuous staining and this change was reversed by 1,25(OH)2D3. In PAN nephropathy rats, TGF-,1 and p-Smad2/3 expression was upregulated, whereas that of BMP-7 and p-Smad1/5/8 was downregulated. Treatment with 1,25(OH)2D3 significantly restored BMP-7/Smad signalling while suppressing TGF-,1/Smad signalling. 4In conclusion, 1,25(OH)2D3 can ameliorate podocyte damage and proteinuria induced by PAN. The beneficial effects of 1,25(OH)2D3 on podocytes may be attributable, in part, to direct modulation of TGF-,1/BMP-7 signalling. [source]

IMPLICATIONS OF CROSS-TALK BETWEEN TUMOUR NECROSIS FACTOR AND INSULIN-LIKE GROWTH FACTOR-1 SIGNALLING IN SKELETAL MUSCLE

CLINICAL AND EXPERIMENTAL PHARMACOLOGY AND PHYSIOLOGY, Issue 7 2008
Miranda D Grounds
SUMMARY 1Inflammation, particularly the pro-inflammatory cytokine tumour necrosis factor (TNF), increases necrosis of skeletal muscle. Depletion of inflammatory cells, such as neutrophils, cromolyn blockade of mast cell degranulation or pharmacological blockade of TNF reduces necrosis of dystrophic myofibres in the mdx mouse model of the lethal childhood disease Duchenne muscular dystrophy (DMD). 2Insulin-like growth factor-1 (IGF-1) is a very important cytokine for maintenance of skeletal muscle mass and the transgenic overexpression of IGF-1 within muscle cells reduces necrosis of dystrophic myofibres in mdx mice. Thus, IGF-1 usually has the opposite effect to TNF. 3Activation of TNF signalling via the c-Jun N-terminal kinase (JNK) can inhibit IGF-1 signalling by phosphorylation and conformational changes in insulin receptor substrate (IRS)-1 downstream of the IGF-1 receptor. Such silencing of IGF-1 signalling in situations where inflammatory cytokines are elevated has many implications for skeletal muscle in vivo. 4The basis for these interactions between TNF and IGF-1 is discussed with specific reference to clinical consequences for myofibre necrosis in DMD and also for the wasting (atrophy) of skeletal muscles that occurs in very old people and in cachexia associated with inflammatory disorders. [source]

Signalling and pharmacological properties of the P2Y14 receptor

ACTA PHYSIOLOGICA, Issue 2 2010
T. K. Harden
Abstract The P2Y14 receptor is a relatively broadly expressed G protein-coupled receptor that is prominently associated with immune and inflammatory cells as well as with many epithelia. This receptor historically was thought to be activated selectively by UDP-glucose and other UDP-sugars. However, UDP is also a very potent agonist of this receptor, and may prove to be one of its most important cognate activators. [source]

Multiple Cues in Status Signalling: The Role of Wingbars in Aggressive Interactions of Male House Sparrows

ETHOLOGY, Issue 10 2006
Veronika Bókony
During aggressive interactions, animals may signal their competitive ability by various ornaments referred to as badges of status. The use of a single badge predicting dominance rank occurs in many vertebrate species. However, animals often display multiple ornaments that may convey information about either different or the same aspects of the signaller's quality, or alternatively, may serve as signal amplifiers. We observed the fighting behaviour of male house sparrows in two captive flocks to investigate whether they may use multiple cues in status signalling during aggressive interactions. Beside the status-signalling bib, male sparrows possess a conspicuous white wingbar that they often display upon aggressive encounters. We tested whether bib size and the wingbar's conspicuousness (i.e. its achromatic contrast with the neighbouring dark feathers) or its area predicted success in various aspects of fighting. We found that bib size strongly predicted overall fighting success (i.e. proportion of fights won) and defence success (i.e. proportion of successful defences out of all attacks received). Wingbar conspicuousness was positively related to defence success after controlling for the effect of bib size in multivariate analyses. Furthermore, displaying the wings also tended to improve the birds' success in defence but not in attack. Wingbar area was unrelated to any measured aspect of fighting ability. We suggest that bib size and wingbar conspicuousness may convey multiple messages on fighting abilities, specifically on overall aggressiveness and defending potential, respectively. Alternatively, wingbars may serve as amplifiers for the wing displays of aggressive motivation. Thus, male sparrows may use multiple cues in assessing the competitive ability of opponents during social interactions. [source]

Successful Application of Video-Projected Human Images for Signalling to Dogs

ETHOLOGY, Issue 10 2003
Péter Pongrácz
Dogs were tested (1) in a two-way choice experiment, where the experimenter indicated a baited bowl by pointing; and (2) in a task where the owner was asked to command the dog to execute simple obedience tasks. In expt 1 dogs (n = 10) were tested at first in the presence of the experimenter (three dimensional condition, 3D), that was followed by a series of pointing trials when the life-sized image of the experimenter was projected onto the wall by the means of a video-projector (two dimensional condition, 2D). Dogs performed correctly more often than expected by chance in both 3D and 2D conditions. In expt 2 the commanding owner was either present in the room (3D), or her/his image was projected on the screen (2D), or only her/his voice was audible for the dog through a speaker (0D). The performance of the dogs (n = 10) decreased to great extent comparing the 3D and 0D condition, as the number of different actions the dogs obeyed was significantly less in the 0D condition. However, there was no difference in the number of different actions obeyed in the 3D and 2D conditions. Our results show that dogs readily obey life-sized, interactive moving image in various communicative situtations. We suppose that the difference between 2D and 3D conditions can be attributed partially to the lack of some additional communicative signals as sounds (verbal cues) and odours (from the human), and to some changes in the context. [source]

Concepts of neural nitric oxide-mediated transmission

EUROPEAN JOURNAL OF NEUROSCIENCE, Issue 11 2008
John Garthwaite
Abstract As a chemical transmitter in the mammalian central nervous system, nitric oxide (NO) is still thought a bit of an oddity, yet this role extends back to the beginnings of the evolution of the nervous system, predating many of the more familiar neurotransmitters. During the 20 years since it became known, evidence has accumulated for NO subserving an increasing number of functions in the mammalian central nervous system, as anticipated from the wide distribution of its synthetic and signal transduction machinery within it. This review attempts to probe beneath those functions and consider the cellular and molecular mechanisms through which NO evokes short- and long-term modifications in neural performance. With any transmitter, understanding its receptors is vital for decoding the language of communication. The receptor proteins specialised to detect NO are coupled to cGMP formation and provide an astonishing degree of amplification of even brief, low amplitude NO signals. Emphasis is given to the diverse ways in which NO receptor activation initiates changes in neuronal excitability and synaptic strength by acting at pre- and/or postsynaptic locations. Signalling to non-neuronal cells and an unexpected line of communication between endothelial cells and brain cells are also covered. Viewed from a mechanistic perspective, NO conforms to many of the rules governing more conventional neurotransmission, particularly of the metabotropic type, but stands out as being more economical and versatile, attributes that presumably account for its spectacular evolutionary success. [source]

Signalling and regulation of collagen I synthesis by ET-1 and TGF-,1

FEBS JOURNAL, Issue 24 2005
Angelika Horstmeyer
Endothelin-1 (ET-1) plays an important role in tissue remodelling and fibrogenesis by inducing synthesis of collagen I via protein kinase C (PKC). ET-1 signals are transduced by two receptor subtypes, the ETA- and ETB-receptors which activate different G, proteins. Here, we investigated the expression of both ET-receptor subtypes in human primary dermal fibroblasts and demonstrated that the ETA-receptor is the major ET-receptor subtype expressed. To determine further signalling intermediates, we inhibited G,i and three phospholipases. Pharmacologic inhibition of G,i, phosphatidylcholine-phospholipase C (PC-PLC) and phospholipase D (PLD), but not of phospholipase C,, abolished the increase in collagen I by ET-1. Inhibition of all phospholipases revealed similar effects on TGF-,1 induced collagen I synthesis, demonstrating involvement of PC-PLC and PLD in the signalling pathways elicited by ET-1 and TGF-,1. ET-1 and TGF-,1 each stimulated collagen I production and in an additive manner. ET-1 further induced connective tissue growth factor (CTGF), as did TGF-,1, however, to lower levels. While rapid and sustained CTGF induction was seen following TGF-,1 treatment, ET-1 increased CTGF in a biphasic manner with lower induction at 3 h and a delayed and higher induction after 5 days of permanent ET-1 treatment. Coincidentally at 5 days of permanent ET-1 stimulation, a switch in ET-receptor subtype expression to the ETB-receptor was observed. We conclude that the signalling pathways induced by ET-1 and TGF-,1 leading to augmented collagen I production by fibroblasts converge on a similar signalling pathway. Thereby, long-time stimulation by ET-1 resulted in a changed ET-receptor subtype ratio and in a biphasic CTGF induction. [source]

Some Hypotheses on the Nature of Difficulty and Ease in Second Language Reading: An Application of Schema Theory

FOREIGN LANGUAGE ANNALS, Issue 6 2000
Philip C. Hauptman
A traditional view of difficulty/ease is explained as consisting of two factors: (1) Language (grammar and vocabulary) and (2) Text Length. A modern view of difficulty/ease is then proposed via four hypotheses: (1) The first Primary Ease Factor in L2 reading is background knowledge; (2) Signalling becomes the Primary Ease Factor in L2 reading when background knowledge is not useful for accessing content schemata; (3) Other factors being equal, the degree of signalling determines the degree of accessibility of a text for the L2 reader; and (4) Other factors being equal, Language, Discourse, and Length are of secondary importance , after Background Knowledge and Signalling , for accessing a text by L2 readers. [source]

The riddle of the dual expression of IgM and IgD

IMMUNOLOGY, Issue 4 2006
Roland Geisberger
Summary Signalling through the B cell antigen receptor (BCR) is required for peripheral B lymphocyte maturation, maintenance, activation and silencing. In mature B cells, the antigen receptor normally consists of two isotypes, membrane IgM and IgD (mIgM, mIgD). Although the signals initiated from both isotypes differ in kinetics and intensity, in vivo, the BCR of either isotype seems to be able to compensate for the loss of the other, reflected by the mild phenotypes of mice deficient for mIgM or mIgD. Thus, it is still unclear why mature B cells need expression of mIgD in addition to mIgM. In the current review we suggest that the view that IgD has a simpIy definable function centred around the basic signalling function should be replaced by the assumption that IgD fine tunes humoral responses, modulates B cell selection and homeostasis and thus shapes the B cell repertoire, defining IgD to be a key modulator of the humoral immune response. [source]

B-cell antigen-receptor signalling in lymphocyte development

IMMUNOLOGY, Issue 4 2003
Leo D. Wang
Summary Signalling through the B-cell antigen receptor (BCR) is required throughout B-cell development and peripheral maturation. Targeted disruption of BCR components or downstream effectors indicates that specific signalling mechanisms are preferentially required for central B-cell development, peripheral maturation and repertoire selection. Additionally, the avidity and the context in which antigen is encountered determine both cell fate and differentiation in the periphery. Although the signalling and receptor components required at each stage have been largely elucidated, the molecular mechanisms through which specific signalling are evoked at each stage are still obscure. In particular, it is not known how the pre-BCR initiates the signals required for normal development or how immature B cells regulate the signalling pathways that determine cell fate. In this review, we will summarize the recent studies that have defined the molecules required for B-cell development and maturation as well as the theories on how signals may be regulated at each stage. [source]

Pre-handover signalling for QoS aware mobility management

INTERNATIONAL JOURNAL OF NETWORK MANAGEMENT, Issue 6 2004
Hakima Chaouchi
In this paper we present a new approach to provide fast handover in Mobile IP. A new Pre-Handover Signalling (PHS) protocol is proposed to allow the network to achieve accurate handover decisions considering different constraints such as QoS, load balancing in the base stations, the user profile, the mobile node service requirements, etc. In addition we propose to minimize the time discovery of the new base station in order to minimize the handover latency. We propose to start the PHS as soon as the mobile node crosses a predefined critical zone area in its current location, this signalling will provide a list of candidate cells to the mobile node with corresponding priorities; the mobile node will select the highest priority base station as soon as the layer two handover occurs. We propose in the current work to use an extension of COPS (Common Open Policy Service) to support the proposed PHS mechanism and overcome the blind handover decisions of Mobile IP and improve the handover performance.,Copyright © 2004 John Wiley & Sons, Ltd. [source]

The Effects of Disruption of A Kinase Anchoring Protein,Protein Kinase A Association on Protein Kinase A Signalling in Neuroendocrine Melanotroph Cells of Xenopus laevis

JOURNAL OF NEUROENDOCRINOLOGY, Issue 7 2006
G. J. H. Corstens
Abstract The secretory activity of melanotroph cells from Xenopus laevis is regulated by multiple neurotransmitters that act through adenylyl cyclase. Cyclic adenosine monophosphate (cAMP), acting on protein kinase A (PKA), stimulates the frequency of intracellular Ca2+ oscillations and the secretory activity of the melanotroph cell. Anchoring of PKA near target proteins is essential for many PKA-regulated processes, and the family of A kinase anchoring proteins (AKAPs) is involved in the compartmentalisation of PKA type II (PKA II) regulatory subunits. In the present study, we determined to what degree cAMP signalling in Xenopus melanotrophs depends on compartmentalised PKA II. For this purpose, a membrane-permeable stearated form of Ht31 (St-Ht31), which dislodges PKA II from AKAP (thus disrupting PKA II signalling), was used. The effect of St-Ht31 on both secretion of radiolabelled peptides and intracellular Ca2+ signalling by superfused Xenopus melanotrophs was assessed. St-Ht31 stimulated secretion but had no effect on Ca2+ signalling. We conclude Xenopus melanotrophs possess a St-Ht31-sensitive PKA II that is associated with the exocytosis machinery and, furthermore, that Ca2+ signalling is regulated by an AKAP-independent signalling system. Moreover, our results support a recent proposal that AKAP participates in regulating PKA activity independently from cAMP. [source]

SEPH, a Cdc7p orthologue from Aspergillus nidulans, functions upstream of actin ring formation during cytokinesis

MOLECULAR MICROBIOLOGY, Issue 1 2001
Kenneth S. Bruno
In the filamentous fungus, Aspergillus nidulans, multiple rounds of nuclear division occur before cytokinesis, allowing an unambiguous identification of genes required specifically for cytokinesis. As in animal cells, both an intact microtubule cytoskeleton and progression through mitosis are required for actin ring formation and contraction. The sepH gene from A. nidulans was discovered in a screen for temperature-sensitive cytokinesis mutants. Sequence analysis showed that SEPH is 42% identical to the serine,threonine kinase Cdc7p from fission yeast. Signalling through the Septation Initiation Network (SIN), which includes Cdc7p and the GTPase Spg1p, is emerging as a primary regulatory pathway used by fission yeast to control cytokinesis. A similar group of proteins comprise the Mitotic Exit Network (MEN) in budding yeast. This is the first direct evidence for the existence of a functional SIN,MEN pathway outside budding and fission yeast. In addition to SEPH, potential homologues were also identified in other fungi and plants but not in animal cells. Deletion of sepH resulted in a viable strain that failed to septate at any temperature. Interestingly, quantitative analysis of the actin cytoskeleton revealed that sepH is required for construction of the actin ring. Therefore, SEPH is distinct from its counterpart in fission yeast, in which SIN components operate downstream of actin ring formation and are necessary for ring contraction and later events of septation. We conclude that A. nidulans has components of a SIN,MEN pathway, one of which, SEPH, is required for early events during cytokinesis. [source]

Fine-Tuning Plant Defence Signalling: Salicylate versus Jasmonate

PLANT BIOLOGY, Issue 1 2006
G. J. M. Beckers
Abstract: Plant defences against pathogens and herbivorous insects form a comprehensive network of interacting signal transduction pathways. The signalling molecules salicylic acid (SA) and jasmonic acid (JA) play important roles in this network. SA is involved in signalling processes providing systemic acquired resistance (SAR), protecting the plant from further infection after an initial pathogen attack. SAR is long-lasting and provides broad spectrum resistance to biotrophic pathogens that feed on a living host cell. The regulatory protein NPR1 is a central positive regulator of SAR. SA-activated NPR1 localizes to the nucleus where it interacts with TGA transcription factors to induce the expression of a large set of pathogenesis-related proteins that contribute to the enhanced state of resistance. In a distinct signalling process, JA protects the plant from insect infestation and necrotrophic pathogens that kill the host cell before feeding. JA activates the regulatory protein COI1 that is part of the E3 ubiquitin ligase-containing complex SCFCOI1, which is thought to derepress JA-responsive genes involved in plant defence. Both synergistic and antagonistic interactions have been observed between SA- and JA-dependent defences. NPR1 has emerged as a critical modulator of cross-talk between the SA and JA signal and is thought to aid in fine tuning defence responses specific to the encountered attacker. Here we review SA- and JA-dependent signal transduction and summarize our current understanding of the molecular mechanisms of cross-talk between these defences. [source]

Expression and promoter activity of the desiccation-specific Solanum tuberosum gene, StDS2

PLANT CELL & ENVIRONMENT, Issue 9 2002
R. Dóczi
Abstract Environmental stresses induce the expression of several plant genes via multiple and cross-talking signalling pathways. Previously it was shown that ScDS2, a gene of the wild potato species, Solanum chacoense, is highly inducible by dehydration but not by abscisic acid (ABA), the mediator of many plant stress responses. Herein it is shown that ScDS2 -related genes are present in the cultivated potato, Solanum tuberosum (StDS2) and also in the non-tuberizing Solanum species, Solanum brevidens (SbDS2). We show that expression of StDS2 is dehydration-specific, is not inducible by cold, heat, salt, hypoxia or oxidative stresses, and is independent of ABA. Signalling of StDS2 induction, however, is dependent on the synthesis of novel proteins because cycloheximide can block StDS2 expression. To analyse the promoter region of StDS2 a genomic library of Solanum tuberosum was established and 1140 and 498 bp regions of the StDS2 promoter were isolated. The promoter fragments were fused to the , -glucuronidase (GUS) reporter gene and tested in transgenic potato plants. Both promoter fragments were able to induce GUS activity in response to dehydration. This result suggests that drought-specific cis -elements are located within 498 bp upstream to the StDS2 coding sequence. [source]

Protease-activated receptors and prostaglandins in inflammatory lung disease

BRITISH JOURNAL OF PHARMACOLOGY, Issue 4 2009
Terence Peters
Protease-activated receptors (PARs) are a novel family of G protein-coupled receptors. Signalling through PARs typically involves the cleavage of an extracellular region of the receptor by endogenous or exogenous proteases, which reveals a tethered ligand sequence capable of auto-activating the receptor. A considerable body of evidence has emerged over the past 20 years supporting a prominent role for PARs in a variety of human physiological and pathophysiological processes, and thus substantial attention has been directed towards developing drug-like molecules that activate or block PARs via non-proteolytic pathways. PARs are widely expressed within the respiratory tract, and their activation appears to exert significant modulatory influences on the level of bronchomotor tone, as well as on the inflammatory processes associated with a range of respiratory tract disorders. Nevertheless, there is debate as to whether the principal response to PAR activation is an augmentation or attenuation of airways inflammation. In this context, an important action of PAR activators may be to promote the generation and release of prostanoids, such as prostglandin E2, which have well-established anti-inflammatory effects in the lung. In this review, we primarily focus on the relationship between PARs, prostaglandins and inflammatory processes in the lung, and highlight their potential role in selected respiratory tract disorders, including pulmonary fibrosis, asthma and chronic obstructive pulmonary disease. This article is part of a themed issue on Mediators and Receptors in the Resolution of Inflammation. To view this issue visit http://www3.interscience.wiley.com/journal/121548564/issueyear?year=2009 [source]

Signalling through TLR2/MyD88 induces differentiation of murine bone marrow stem and progenitor cells to functional phagocytes in response to Candida albicans

CELLULAR MICROBIOLOGY, Issue 1 2010
Alberto Yáñez
Summary We have previously demonstrated that inactivated yeasts and hyphae of Candida albicans induce in vitro the proliferation of murine haematopoietic stem and progenitor cells (HSPCs, sorted as LKS cells: Lin - c-Kit+ Sca-1+) as well as their differentiation to lineage-positive cells, through a MyD88-dependent pathway. In this work, we have found that this process is mainly mediated by TLR2, and that expanding cells express myeloid and not lymphoid markers. Incubation of long-term repopulating HSCs (Lin - CD105+ and Sca-1+) with C. albicans yeasts resulted in their proliferation and up regulation of the common myeloid progenitors (CMPs) markers, CD34 and Fc,RII/III, by a TLR2/MyD88-dependent signalling pathway. In addition, this TLR2/MyD88 signalling promotes the differentiation of CMPs and granulocyte and macrophage progenitors (GMPs) into cells with the morphology of macrophages and neutrophils, characterized by an increase in the expression of CD11b, F4/80 and Ly6G, independently of the presence of growth and differentiation factors. These differentiated cells were able to phagocytose C. albicans yeasts and to produce proinflammatory cytokines. In conclusion, C. albicans may be sensed by TLRs on haematopoietic stem and progenitor cells to promote the host capability for rapidly replenishing myeloid cells that constitute the first line of defence against C. albicans. [source]

Signalling and phagocytosis in the orchestration of host defence

CELLULAR MICROBIOLOGY, Issue 2 2007
J. Magarian Blander
Summary Dendritic cells (DCs) orchestrate either tolerance or immunity. At the heart of this function lies phagocytosis, which allows DCs to sample the tissue microenvironment and deliver both its self and non-self constituents into endocytic compartments for clearance, degradation and presentation by major histocompatibility complex (MHC) molecules. Depending on the type of signalling pathways triggered during phagocytosis, DCs deliver appropriate signals to T cells that determine either their tolerance or activation and differentiation. Here I draw attention to the ability of DCs to read the contents of their phagosomes depending on the type of compartmentalized signalling pathways engaged during internalization. Toll-like receptors (TLRs) engaged during phagocytosis of microbial pathogens, but not syngeneic apoptotic cells exert phagosome autonomous control on both the kinetics and outcome of phagosome maturation. By bearing the assembly of signalling complexes on their membranes, individual phagosomes undergo separate programmes of maturation irrespective of the activation status of the DC carrying them. Phagosomes carrying microbial cargo are favoured for MHC class II presentation precluding phagosomes carrying self from contributing to the first signal delivered to T cells , the peptide,MHC complex. This mechanism prevents the potential presentation of peptides derived from self within the context of TLR-induced co-stimulatory signals. [source]

Inhibition of p38 MAP kinase during cellular activation results in IFN-,-dependent augmentation of IL-12 production by human monocytes/macrophages

CLINICAL & EXPERIMENTAL IMMUNOLOGY, Issue 1 2001
J. B. Marriott
Interleukin-12 (IL-12) is a key immunomodulatory cytokine produced by antigen-presenting cells that promotes cellular immunity and enables the generation of protective immunity against intracellular pathogens and tumours. Therefore, modulation of IL-12 activity is a primary immunotherapeutic goal. However, little is known about its regulation. Signalling via p38 MAPK has been implicated in the control of inflammatory responses and is therefore a potential therapeutic target. We have used the highly selective p38 MAPK inhibitor (SB203580) to examine the effect of this pathway on the production of IL-12. Surprisingly, we found that SB203580 strongly up-regulated LPS induced IL-12p40 at the protein (intracellular and secreted) and mRNA levels in PBMC cultures. The effect on IL-12 was apparent using both T cell-independent and T cell-dependent stimuli but not in unstimulated cultures, indicating that activation signals are required. Furthermore, the production of IFN- , by T cells is crucial as production was not increased in LPS-stimulated, purified adherent monocytes/macrophages without the addition of exogenous IFN- ,. These results provide evidence that p38 MAPK has an unexpected suppressive effect on IL-12p40 gene transcription, and suggests interplay between p38 MAPK- and IFN- , -mediated signals in the regulation of IL-12 production by monocytes/macrophages. Furthermore, the importance of IL-12 as a key immunoregulatory cytokine suggests that the clinical application of pyrinidyl imidazole inhibitors, such as SB203580, may need to be reassessed. [source]

TRAF6 knockdown promotes survival and inhibits inflammatory response to lipopolysaccharides in rat primary renal proximal tubule cells

ACTA PHYSIOLOGICA, Issue 3 2010
S. Liu
Abstract Aim:, TRAF6 is a unique adaptor protein of the tumour necrosis factor receptor-associated factor family that mediates both tumour necrosis factor receptor (TNFR) and interleukin-1 receptor/Toll-like receptor (IL-1R/TLR) signalling. Activation of IL-1R/TLR and TNFR pathways in renal tubular cells contributes to renal injury. This study aimed to investigate if blockade of lipopolysaccharide (LPS)-triggered TLR4 signalling by small interfering RNA (siRNA) targeting TRAF6 protects survival and inhibits inflammatory response in isolated rat renal proximal tubular cells (PTCs). Methods:, PTCs isolated from F344 rat kidneys were transfected with chemically synthesized siRNA targeting TRAF6 mRNA. Real-time quantitative PCR was applied to measure mRNA level of TRAF6, TNF-,, IL-6 and monocyte chemoattractant protein-1 (MCP-1). Protein levels of extracellular signal-regulated kinase (ERK), c-jun N-terminal kinase (JNK), p38 mitogen-activated protein kinase, caspase 3 and cleaved caspase 3 were evaluated by Western blotting. Cell viability was analysed with XTT reagents. Results:, We found that the TRAF6 gene was effectively silenced in PTCs using siRNA. TRAF6 knockdown resulted in reduced TNF-, and IL-6 mRNA expression upon LPS challenge. LPS-induced phosphorylation of JNK and p38 was attenuated in TRAF6 siRNA-transfected cells while the change in the phosphorylation of ERK was not remarkable. TRAF6 knockdown was associated with increased cell viability and reduced protein level of cleaved caspase-3, both, in the absence and presence of LPS. Conclusion:, Our studies suggest that TRAF6 knockdown may inhibit inflammatory response and promote cell survival upon LPS challenge in primary rat proximal renal tubular cells. [source]

The birth and postnatal development of purinergic signalling

ACTA PHYSIOLOGICA, Issue 2 2010
G. Burnstock
Abstract The purinergic signalling system is one of the most ancient and arguably the most widespread intercellular signalling system in living tissues. In this review we present a detailed account of the early developments and current status of purinergic signalling. We summarize the current knowledge on purinoceptors, their distribution and role in signal transduction in various tissues in physiological and pathophysiological conditions. [source]

Modulation and metamodulation of synapses by adenosine

ACTA PHYSIOLOGICA, Issue 2 2010
J. A. Ribeiro
Abstract The presence of adenosine in all nervous system cells (neurones and glia) together with its intensive release following insults makes adenosine as a sort of ,regulator' of synaptic communication, leading to the homeostatic coordination of brain function. Besides the direct actions of adenosine on the neurosecretory mechanisms, to tune neurotransmitter release, adenosine receptors interact with other receptors as well as with transporters as part of its attempt to fine-tune synaptic transmission. This review will focus on examples of the different ways adenosine can use to modulate or metamodulate synapses, in other words, to trigger or brake the action of some neurotransmitters and neuromodulators, to cross-talk with other G protein-coupled receptors, with ionotropic receptors and with receptor kinases as well as with transporters. Most of these interactions occur through A2A receptors, which in spite of their low density in some brain areas, such as the hippocampus, may function as amplifiers of the signalling of other mediators at synapses. [source]

Resistance exercise increases leg muscle protein synthesis and mTOR signalling independent of sex

ACTA PHYSIOLOGICA, Issue 1 2010
H. C. Dreyer
Abstract Aim:, Sex differences are evident in human skeletal muscle as the cross-sectional area of individual muscle fibres is greater in men than in women. We have recently shown that resistance exercise stimulates mammalian target of rapamycin (mTOR) signalling and muscle protein synthesis in humans during early post-exercise recovery. Therefore, the aim of this study was to determine if sex influences the muscle protein synthesis response during recovery from resistance exercise. Methods:, Seventeen subjects, nine male and eight female, were studied in the fasted state before, during and for 2 h following a bout of high-intensity leg resistance exercise. Mixed muscle protein fractional synthetic rate was measured using stable isotope techniques and mTOR signalling was assessed by immunoblotting from repeated vastus lateralis muscle biopsy samples. Results:, Post-exercise muscle protein synthesis increased by 52% in the men and by 47% in the women (P < 0.05) and was not different between groups (P > 0.05). Akt phosphorylation increased in both groups at 1 h post-exercise (P < 0.05) and returned to baseline during 2 h post-exercise with no differences between groups (P > 0.05). Phosphorylation of mTOR and its downstream effector S6K1 increased significantly and similarly between groups during post-exercise recovery (P < 0.05). eEF2 phosphorylation decreased at 1- and 2 h post-exercise (P < 0.05) to a similar extent in both groups. Conclusion:, The contraction-induced increase in early post-exercise mTOR signalling and muscle protein synthesis is independent of sex and appears to not play a role in the sexual dimorphism of leg skeletal muscle in young men and women. [source]

Role of ataxia telangiectasia mutated in insulin signalling of muscle-derived cell lines and mouse soleus

ACTA PHYSIOLOGICA, Issue 4 2010
I. Jeong
Abstract Aim:, Ataxia telangiectasia mutated (ATM) reportedly plays a role in insulin-stimulated activation of Akt in some cell types but not in others. The role of ATM in insulin signalling has not been firmly resolved for skeletal muscle cells, for which Akt phosphorylation is a pivotal step in stimulation of glucose transport. Accordingly, our aim was to determine the role of ATM in insulin effects for cell lines derived from skeletal muscle and for skeletal muscle. Methods:, We examined insulin effects in L6 myotubes, mouse soleus, C2C12 myotubes and differentiated rhabdomyosarcoma (RD) cells in the presence and absence of a low concentration (1 ,m) of the ATM inhibitor KU55933. We also compared insulin signalling in C2C12 cells expressing shRNA against ATM and control cell lines (empty vector; cells expressing non-targeting shRNA). Results:, In L6 myotubes and mouse soleus muscle, KU55933 inhibited insulin-stimulated phosphorylation of the 160 kDa substrate of Akt (AS160) despite no effect on Akt. In contrast, KU55933 prevented insulin-stimulated Akt phosphorylation in C2C12 myotubes. Furthermore, C2C12 myotubes expressing shRNA against ATM displayed reduced insulin-stimulated Akt phosphorylation compared to controls. KU55933 also decreased insulin-stimulated Akt phosphorylation in differentiated RD cells. Conclusion:, These model-dependent differences in the role of ATM in insulin action demonstrate a role of ATM in insulin-stimulated phosphorylation of Akt (in C2C12 and RD cells) but also allow the elucidation of a novel, Akt-independent role of ATM (in L6 myotubes and mouse soleus, at the level of AS160) in insulin signalling. [source]

Contraction-induced changes in skeletal muscle Na+,K+ pump mRNA expression , importance of exercise intensity and Ca2+ -mediated signalling

ACTA PHYSIOLOGICA, Issue 4 2010
N. B. Nordsborg
Abstract Aim:, To investigate if exercise intensity and Ca2+ signalling regulate Na+,K+ pump mRNA expression in skeletal muscle. Methods:, The importance of exercise intensity was evaluated by having trained and untrained humans perform intense intermittent and prolonged exercise. The importance of Ca2+ signalling was investigated by electrical stimulation of rat soleus and extensor digitorum longus (EDL) muscles in combination with studies of cell cultures. Results:, Intermittent cycling exercise at ,85% of VO2peak increased (P < 0.05) ,1 and ,1 mRNA expression ,2-fold in untrained and trained subjects. In trained subjects, intermittent exercise at ,70% of VO2peak resulted in a less (P < 0.05) pronounced increase (,1.4-fold; P < 0.05) for ,1 and no change in ,1 mRNA. Prolonged low intensity exercise increased (P < 0.05) mRNA expression of ,1 ,3.0-fold and ,2 ,1.8-fold in untrained but not in trained subjects. Electrical stimulation of rat soleus, but not EDL, muscle increased (P < 0.05) ,1 mRNA expression, but not when combined with KN62 and cyclosporin A incubation. Ionomycin incubation of cultured primary rat skeletal muscle cells increased (P < 0.05) ,1 and reduced (P < 0.001) ,2 mRNA expression and these responses were abolished (P < 0.05) by co-incubation with cyclosporin A or KN62. Conclusion:, (1) Exercise-induced increases in Na+,K+ pump ,1 and ,1 mRNA expression in trained subjects are more pronounced after high- than after moderate- and low-intensity exercise. (2) Both prolonged low and short-duration high-intensity exercise increase ,1 mRNA expression in untrained subjects. (3) Ca2+i regulates ,1 mRNA expression in oxidative muscles via Ca2+/calmodulin-dependent protein kinase (CaMK) and calcineurin signalling pathways. [source]

Hypocretin/orexin and energy expenditure

ACTA PHYSIOLOGICA, Issue 3 2010
J. A. Teske
Abstract The hypocretins or orexins are endogenous neuropeptides synthesized in discrete lateral, perifornical and dorsal hypothalamic neurones. These multi-functional neuropeptides modulate energy homeostasis, arousal, stress, reward, reproduction and cardiovascular function. This review summarizes the role of hypocretins in modulating non-sleep-related energy expenditure with specific focus on the augmentation of whole body energy expenditure as well as hypocretin-induced physical activity and sympathetic outflow. We compare the efficacy of hypocretin-1 and 2 on energy expenditure and evaluate whether the literature implicates hypocretin signalling though the hypocretin-1 and -2 receptor as having shared and or functionally specific physiological effects. Thus far data suggest that hypocretin-1 has a more robust stimulatory effect relative to hypocretin-2. Furthermore, hypocretin-1 receptor predominantly mediates behaviours known to influence energy expenditure. Further studies on the hypocretin-2 receptor are needed. [source]

Role of orexin in the regulation of glucose homeostasis

ACTA PHYSIOLOGICA, Issue 3 2010
H. Tsuneki
Abstract Orexin-A (hypocretin-1) and orexin-B (hypocretin-2) are hypothalamic neuropeptides that play key roles in the regulation of wakefulness, feeding, reward, autonomic functions and energy homeostasis. To control these functions indispensable for survival, orexin-expressing neurones integrate peripheral metabolic signals, interact with many types of neurones in the brain and modulate their activities via the activation of orexin-1 receptor or orexin-2 receptor. In addition, a new functional role of orexin is emerging in the regulation of insulin and leptin sensitivities responsible for whole-body glucose metabolism. Recent evidence indicates that orexin efficiently protects against the development of peripheral insulin resistance induced by ageing or high-fat feeding in mice. In particular, the orexin receptor-2 signalling appears to confer resistance to diet-induced obesity and insulin insensitivity by improving leptin sensitivity. In fact, the expression of orexin gene is known to be down-regulated by hyperglycaemia in the rodent model of diabetes, such as ob/ob and db/db mice. Moreover, the levels of orexin receptor-2 mRNA have been shown to decline in the brain of mice along with ageing. These suggest that hyperglycaemia due to insulin insensitivity during ageing or by habitual consumption of a high-fat diet leads to the reduction in orexin expression in the hypothalamus, thereby further exacerbating peripheral insulin resistance. Therefore, orexin receptor controlling hypothalamic insulin/leptin actions may be a new target for possible future treatment of hyperglycaemia in patients with type 2 diabetes. [source]

Effects of short-term food deprivation on orexin-A-induced intestinal bicarbonate secretion in comparison with related secretagogues

ACTA PHYSIOLOGICA, Issue 3 2010
G. Flemström
Abstract Studies of gastrointestinal physiology in humans and intact animals are usually conducted after overnight fast. We compared the effects of orexin-A, vasoactive intestinal polypeptide (VIP), melatonin, serotonin, uroguanylin, ghrelin and prostaglandin E2 (PGE2) on duodenal bicarbonate secretion in fed and overnight fasted animals. This review is a summary of our findings. Secretagogues were administered by intra-arterial infusion or luminally (PGE2). Enterocyte intracellular calcium ([Ca2+]i) signalling was studied by fluorescence imaging. Total RNA was extracted, reverse transcripted to cDNA and expression of orexin receptors measured by quantitative real-time PCR. Orexin-A stimulates the duodenal secretion in continuously fed animals but not in food-deprived animals. Similarly, short-term fasting causes a 100-fold decrease in the amount of the muscarinic agonist bethanechol required for stimulation of secretion. In contrast, fasting does not affect secretory responses to intra-arterial VIP, melatonin, serotonin, uroguanylin and ghrelin, or that to luminal PGE2. Orexin-A induces [Ca2+]i signalling in enterocytes from fed rats but no significant [Ca2+]i responses occur in enterocytes from fasted animals. In addition, overnight fasting decreases the expression of mucosal orexin receptors. Short-term food deprivation thus decreases duodenal expression of orexin receptors and abolishes the secretory response to orexin-A as well as orexin-A-induced [Ca2+]i signalling. Fasting, furthermore, decreases mucosal sensitivity to bethanechol. The absence of declines in secretory responses to other secretagogues tested strongly suggests that short-term fasting does not affect the secretory capacity of the duodenal mucosa in general. Studies of intestinal secretion require particular evaluation with respect to feeding status. [source]

Rapid and easy semi-quantitative evaluation method for diacylglycerol and inositol-1,4,5-trisphosphate generation in orexin receptor signalling

ACTA PHYSIOLOGICA, Issue 3 2010
M. E. Ekholm
Abstract Aim:, Fluorescent protein-based indicators have enabled measurement of intracellular signals previously nearly inaccessible for studies. However, indicators showing intracellular translocation upon response suffer from serious limitations, especially the very time-consuming data collection. We therefore set out in this study to evaluate whether fixing and counting cells showing translocation could mend this issue. Methods:, Altogether three different genetically encoded indicators for diacylglycerol and inositol-1,4,5-trisphosphate were transiently expressed in Chinese hamster ovary cells stably expressing human OX1 orexin receptors. Upon stimulation with orexin-A, the cells were fixed with six different protocols. Results:, Different protocols showed clear differences in their ability to preserve the indicator's localization (i.e. translocation after stimulus) and its fluorescence, and the best results for each indicator were obtained with a different protocol. The concentration,response data obtained with cell counting are mostly comparable to the real-time translocation and biochemical data. Conclusion:, The counting method, as used here, works at single time point and looses the single-cell-quantitative aspect. However, it also has some useful properties. First, it easily allows processing of a 100- to 1000-fold higher cell numbers than real-time imaging producing statistically consistent population-quantitative data much faster. Secondly, it does not require expensive real-time imaging equipment. Fluorescence in fixed cells can also be quantitated, though this analysis would be more time-consuming than cell counting. Thirdly, in addition to the quantitative data collection, the method could be applied for identifying responsive cells. This might be very useful in identification of e.g. orexin-responding neurones in a large population of non-responsive cells in primary cultures. [source]

Cardioprotection of bradykinin at reperfusion involves transactivation of the epidermal growth factor receptor via matrix metalloproteinase-8

ACTA PHYSIOLOGICA, Issue 4 2009
C. Methner
Abstract Aim:, The endogenous autacoid bradykinin (BK) reportedly reduces myocardial infarct size when given exogenously at reperfusion. Muscarinic and opioid G-protein-coupled receptors are equally protective and have been shown to couple through a matrix metalloproteinase (MMP)-dependent transactivation of the epidermal growth factor receptor (EGFR). Here we test whether BK protects the rat heart through the EGFR by an MMP-dependent pathway. Methods:, Infarct size was measured in isolated perfused rat hearts undergoing 30 min regional ischaemia followed by 120 min reperfusion. In additional studies HL-1 cardiomyocytes were loaded with tetramethylrhodamine ethyl to measure their mitochondrial membrane potential (,m). Adding the calcium ionophore calcimycin, causes ,m-collapse presumably due to calcium-induced mitochondrial permeability transition. Results:, As expected, BK (100 nmol L,1) started 5 min prior to reperfusion reduced infarct size from 38.9 ± 2.0% of the ischaemic zone in control hearts to 22.2 ± 3.3% (P < 0.001). Co-infusing the EGFR inhibitor AG1478, the broad-spectrum MMP-inhibitor GM6001, or a highly selective MMP-8 inhibitor abolished BK's protection, thus suggesting an MMP-8-dependent EGFR transactivation in the signalling. Eighty minutes of exposure to calcimycin reduced the mean cell fluorescence to 37.4 ± 1.8% of untreated cells while BK could partly preserve the fluorescence and, hence, protect the cells (50.5 ± 2.3%, P < 0.001). The BK-induced mitochondrial protection could again be blocked by AG1478, GM6001 and MMP-8 inhibitor. Finally, Western blotting revealed that BK's protection was correlated with increased phosphorylation of EGFR and its downstream target Akt. Conclusion:, These results indicate that BK at reperfusion triggers its protective signalling pathway through MMP-8-dependent transactivation of the EGFR. [source]