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Sample records for distinct pathways mediate

  1. Distinct Hippocampal Pathways Mediate Dissociable Roles of Context in Memory Retrieval.

    Science.gov (United States)

    Xu, Chun; Krabbe, Sabine; Gründemann, Jan; Botta, Paolo; Fadok, Jonathan P; Osakada, Fumitaka; Saur, Dieter; Grewe, Benjamin F; Schnitzer, Mark J; Callaway, Edward M; Lüthi, Andreas

    2016-11-03

    Memories about sensory experiences are tightly linked to the context in which they were formed. Memory contextualization is fundamental for the selection of appropriate behavioral reactions needed for survival, yet the underlying neuronal circuits are poorly understood. By combining trans-synaptic viral tracing and optogenetic manipulation, we found that the ventral hippocampus (vHC) and the amygdala, two key brain structures encoding context and emotional experiences, interact via multiple parallel pathways. A projection from the vHC to the basal amygdala mediates fear behavior elicited by a conditioned context, whereas a parallel projection from a distinct subset of vHC neurons onto midbrain-projecting neurons in the central amygdala is necessary for context-dependent retrieval of cued fear memories. Our findings demonstrate that two fundamentally distinct roles of context in fear memory retrieval are processed by distinct vHC output pathways, thereby allowing for the formation of robust contextual fear memories while preserving context-dependent behavioral flexibility. Copyright © 2016 Elsevier Inc. All rights reserved.

  2. Syndecans promote integrin-mediated adhesion of mesenchymal cells in two distinct pathways

    DEFF Research Database (Denmark)

    Whiteford, James; Behrends, Volker; Kirby, Hishani

    2007-01-01

    and signaling through the cytoplasmic domain of syndecan-4. Here an alternate pathway mediated by the extracellular domains of syndecans-2 and -4 is characterized that is independent of both heparan sulphate and syndecan signaling. This pathway is restricted to mesenchymal cells and was not seen in any...... epithelial cell line tested, apart from vascular endothelia. The syndecan ectodomains coated as substrates promoted integrin-dependent attachment, spreading and focal adhesion formation. Syndecan-4 null cells were competent, as were fibroblasts compromised in heparan sulphate synthesis that were unable...

  3. Distinct pathways mediate the sorting of tail-anchored proteins to the plastid outer envelope.

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    Preetinder K Dhanoa

    Full Text Available BACKGROUND: Tail-anchored (TA proteins are a distinct class of membrane proteins that are sorted post-translationally to various organelles and function in a number of important cellular processes, including redox reactions, vesicular trafficking and protein translocation. While the molecular targeting signals and pathways responsible for sorting TA proteins to their correct intracellular destinations in yeasts and mammals have begun to be characterized, relatively little is known about TA protein biogenesis in plant cells, especially for those sorted to the plastid outer envelope. METHODOLOGY/PRINCIPAL FINDINGS: Here we investigated the biogenesis of three plastid TA proteins, including the 33-kDa and 34-kDa GTPases of the translocon at the outer envelope of chloroplasts (Toc33 and Toc34 and a novel 9-kDa protein of unknown function that we define here as an outer envelope TA protein (OEP9. Using a combination of in vivo and in vitro assays we show that OEP9 utilizes a different sorting pathway than that used by Toc33 and Toc34. For instance, while all three TA proteins interact with the cytosolic OEP chaperone/receptor, AKR2A, the plastid targeting information within OEP9 is distinct from that within Toc33 and Toc34. Toc33 and Toc34 also appear to differ from OEP9 in that their insertion is dependent on themselves and the unique lipid composition of the plastid outer envelope. By contrast, the insertion of OEP9 into the plastid outer envelope occurs in a proteinaceous-dependent, but Toc33/34-independent manner and membrane lipids appear to serve primarily to facilitate normal thermodynamic integration of this TA protein. CONCLUSIONS/SIGNIFICANCE: Collectively, the results provide evidence in support of at least two sorting pathways for plastid TA outer envelope proteins and shed light on not only the complex diversity of pathways involved in the targeting and insertion of proteins into plastids, but also the molecular mechanisms that underlie

  4. Transcriptome signature identifies distinct cervical pathways induced in lipopolysaccharide-mediated preterm birth.

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    Willcockson, Alexandra R; Nandu, Tulip; Liu, Cheuk-Lun; Nallasamy, Shanmugasundaram; Kraus, W Lee; Mahendroo, Mala

    2018-03-01

    With half a million babies born preterm each year in the USA and about 15 million worldwide, preterm birth (PTB) remains a global health issue. Preterm birth is a primary cause of infant morbidity and mortality and can impact lives long past infancy. The fact that there are numerous, and many currently unidentified, etiologies of PTB has hindered development of tools for risk evaluation and preventative therapies. Infection is estimated to be involved in nearly 40% of PTBs of known etiology; therefore, understanding how infection-mediated inflammation alters the cervical milieu and leads to preterm tissue biomechanical changes are questions of interest. Using RNA-seq, we identified enrichment of components involved in inflammasome activation and unique proteases in the mouse cervix during lipopolysaccharide (LPS)-mediated PTB and not physiologically at term before labor. Despite transcriptional induction of inflammasome components, there was no evidence of functional activation based on assessment of mature IL1B and IL18 proteins. The increased transcription of proteases that target both elastic fibers and collagen and concentration of myeloid-derived cells capable of protease synthesis in the cervical stroma support the structural disruption of elastic fibers as a functional output of protease activity. The recent demonstration that elastic fibers contribute to the biomechanical function of the pregnant cervix suggests their protease-induced disruption in the infection model of LPS-mediated PTB and may contribute to premature loss of mechanical competency and preterm delivery. Collectively, the transcriptomics and ultrastructural data provide new insights into the distinct mechanisms of premature cervical remodeling in response to infection.

  5. Two conserved modules of Schizosaccharomyces pombe Mediator regulate distinct cellular pathways

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    Linder, Tomas; Rasmussen, Nina; Samuelsen, Camilla O

    2008-01-01

    Mediator is an evolutionary conserved coregulator complex required for transcription of almost all RNA polymerase II-dependent genes. The Schizosaccharomyces pombe Mediator consists of two dissociable components-a core complex organized into a head and middle domain as well as the Cdk8 regulatory...... subcomplex. In this work we describe a functional characterization of the S. pombe Mediator. We report the identification of the S. pombe Med20 head subunit and the isolation of ts alleles of the core head subunit encoding med17+. Biochemical analysis of med8(ts), med17(ts), Deltamed18, Deltamed20...... and Deltamed27 alleles revealed a stepwise head domain molecular architecture. Phenotypical analysis of Cdk8 and head module alleles including expression profiling classified the Mediator mutant alleles into one of two groups. Cdk8 module mutants flocculate due to overexpression of adhesive cell...

  6. Distinct phosphotyrosines on a growth factor receptor bind to specific molecules that mediate different signaling pathways.

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    Fantl, W J; Escobedo, J A; Martin, G A; Turck, C W; del Rosario, M; McCormick, F; Williams, L T

    1992-05-01

    The receptor for platelet-derived growth factor (PDGF) binds two proteins containing SH2 domains, GTPase activating protein (GAP) and phosphatidylinositol 3-kinase (PI3-kinase). The sites on the receptor that mediate this interaction were identified by using phosphotyrosine-containing peptides representing receptor sequences to block specifically binding of either PI3-kinase or GAP. These results suggested that PI3-kinase binds two phosphotyrosine residues, each located in a 5 aa motif with an essential methionine at the fourth position C-terminal to the tyrosine. Point mutations at these sites caused a selective elimination of PI3-kinase binding and loss of PDGF-stimulated DNA synthesis. Mutation of the binding site for GAP prevented the receptor from associating with or phosphorylating GAP, but had no effect on PI3-kinase binding and little effect on DNA synthesis. Therefore, GAP and PI3-kinase interact with the receptor by binding to different phosphotyrosine-containing sequence motifs.

  7. p53 and ATF4 mediate distinct and additive pathways to skeletal muscle atrophy during limb immobilization

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    Fox, Daniel K.; Ebert, Scott M.; Bongers, Kale S.; Dyle, Michael C.; Bullard, Steven A.; Dierdorff, Jason M.; Kunkel, Steven D.

    2014-01-01

    Immobilization causes skeletal muscle atrophy via complex signaling pathways that are not well understood. To better understand these pathways, we investigated the roles of p53 and ATF4, two transcription factors that mediate adaptations to a variety of cellular stresses. Using mouse models, we demonstrate that 3 days of muscle immobilization induces muscle atrophy and increases expression of p53 and ATF4. Furthermore, muscle fibers lacking p53 or ATF4 are partially resistant to immobilization-induced muscle atrophy, and forced expression of p53 or ATF4 induces muscle fiber atrophy in the absence of immobilization. Importantly, however, p53 and ATF4 do not require each other to promote atrophy, and coexpression of p53 and ATF4 induces more atrophy than either transcription factor alone. Moreover, muscle fibers lacking both p53 and ATF4 are more resistant to immobilization-induced atrophy than fibers lacking only p53 or ATF4. Interestingly, the independent and additive nature of the p53 and ATF4 pathways allows for combinatorial control of at least one downstream effector, p21. Using genome-wide mRNA expression arrays, we identified p21 mRNA as a skeletal muscle transcript that is highly induced in immobilized muscle via the combined actions of p53 and ATF4. Additionally, in mouse muscle, p21 induces atrophy in a manner that does not require immobilization, p53 or ATF4, and p21 is required for atrophy induced by immobilization, p53, and ATF4. Collectively, these results identify p53 and ATF4 as essential and complementary mediators of immobilization-induced muscle atrophy and discover p21 as a critical downstream effector of the p53 and ATF4 pathways. PMID:24895282

  8. Activity-Dependent Ubiquitination of GluA1 Mediates a Distinct AMPAR Endocytosis and Sorting Pathway

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    Schwarz, Lindsay A.; Hall, Benjamin J.; Patrick, Gentry N.

    2010-01-01

    The accurate trafficking of AMPA receptors (AMPARs) to and from the synapse is a critical component of learning and memory in the brain, while dysfunction of AMPAR trafficking is hypothesized to be an underlying mechanism of Alzheimer’s disease. Previous work has shown that ubiquitination of integral membrane proteins is a common post-translational modification used to mediate endocytosis and endocytic sorting of surface proteins in eukaryotic cells. Here we report that mammalian AMPARs become ubiquitinated in response to their activation. Using a mutant of GluA1 that is unable to be ubiquitinated at lysines on its carboxy-terminus, we demonstrate that ubiquitination is required for internalization of surface AMPARs and their trafficking to the lysosome in response to the AMPAR agonist AMPA, but not for internalization of AMPARs in response to the NMDA receptor (NMDAR) agonist NMDA. Through over-expression or RNAi-mediated knockdown, we identify that a specific E3 ligase, Nedd4-1, is necessary for this process. Finally, we show that ubiquitination of GluA1 by Nedd4-1 becomes more prevalent as neurons mature. Together, these data show that ubiquitination of GluA1-containing AMPARs by Nedd4-1 mediates their endocytosis and trafficking to the lysosome. Furthermore, these results provide insight into how hippocampal neurons regulate AMPAR trafficking and degradation with high specificity in response to differing neuronal signaling cues, and suggest that changes to this pathway may occur as neurons mature. PMID:21148011

  9. Activity-dependent ubiquitination of GluA1 mediates a distinct AMPA receptor endocytosis and sorting pathway.

    Science.gov (United States)

    Schwarz, Lindsay A; Hall, Benjamin J; Patrick, Gentry N

    2010-12-08

    The accurate trafficking of AMPA receptors (AMPARs) to and from the synapse is a critical component of learning and memory in the brain, whereas dysfunction of AMPAR trafficking is hypothesized to be an underlying mechanism of Alzheimer's disease. Previous work has shown that ubiquitination of integral membrane proteins is a common posttranslational modification used to mediate endocytosis and endocytic sorting of surface proteins in eukaryotic cells. Here we report that mammalian AMPARs become ubiquitinated in response to their activation. Using a mutant of GluA1 that is unable to be ubiquitinated at lysines on its C-terminus, we demonstrate that ubiquitination is required for internalization of surface AMPARs and their trafficking to the lysosome in response to the AMPAR agonist AMPA but not for internalization of AMPARs in response to the NMDA receptor agonist NMDA. Through overexpression or RNA interference-mediated knockdown, we identify that a specific E3 ligase, Nedd4-1 (neural-precursor cell-expressed developmentally downregulated gene 4-1), is necessary for this process. Finally, we show that ubiquitination of GluA1 by Nedd4-1 becomes more prevalent as neurons mature. Together, these data show that ubiquitination of GluA1-containing AMPARs by Nedd4-1 mediates their endocytosis and trafficking to the lysosome. Furthermore, these results provide insight into how hippocampal neurons regulate AMPAR trafficking and degradation with high specificity in response to differing neuronal signaling cues and suggest that changes to this pathway may occur as neurons mature.

  10. Two Distinct Mediated Pathways to Disordered Eating in Response to Weight Stigmatization and Their Application to Prevention Programs

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    Simone, Melissa; Lockhart, Ginger

    2016-01-01

    Objective: Disordered eating behaviors among undergraduate women are common and, thus, are an important public health concern. Weight stigmatization, stress, and social withdrawal are often associated with disordered eating behaviors; however, it is unclear whether stress and social withdrawal act as mediators between weight stigmatization and…

  11. Raft-mediated trafficking of apical resident proteins occurs in both direct and transcytotic pathways in polarized hepatic cells : Role of distinct lipid microdomains

    NARCIS (Netherlands)

    Slimane, TA; Trugnan, G; van Ijzendoorn, SCD; Hoekstra, D

    In polarized hepatic cells, pathways and molecular principles mediating the flow of resident apical bile canalicular proteins have not yet been resolved. Herein, we have investigated apical trafficking of a glycosylphosphatidylinositol-linked and two single transmembrane domain proteins on the one

  12. Breast cancer oestrogen independence mediated by BCAR1 or BCAR3 genes is transmitted through mechanisms distinct from the oestrogen receptor signalling pathway or the epidermal growth factor receptor signalling pathway

    International Nuclear Information System (INIS)

    Dorssers, Lambert CJ; Agthoven, Ton van; Brinkman, Arend; Veldscholte, Jos; Smid, Marcel; Dechering, Koen J

    2005-01-01

    Tamoxifen is effective for endocrine treatment of oestrogen receptor-positive breast cancers but ultimately fails due to the development of resistance. A functional screen in human breast cancer cells identified two BCAR genes causing oestrogen-independent proliferation. The BCAR1 and BCAR3 genes both encode components of intracellular signal transduction, but their direct effect on breast cancer cell proliferation is not known. The aim of this study was to investigate the growth control mediated by these BCAR genes by gene expression profiling. We have measured the expression changes induced by overexpression of the BCAR1 or BCAR3 gene in ZR-75-1 cells and have made direct comparisons with the expression changes after cell stimulation with oestrogen or epidermal growth factor (EGF). A comparison with published gene expression data of cell models and breast tumours is made. Relatively few changes in gene expression were detected in the BCAR-transfected cells, in comparison with the extensive and distinct differences in gene expression induced by oestrogen or EGF. Both BCAR1 and BCAR3 regulate discrete sets of genes in these ZR-75-1-derived cells, indicating that the proliferation signalling proceeds along distinct pathways. Oestrogen-regulated genes in our cell model showed general concordance with reported data of cell models and gene expression association with oestrogen receptor status of breast tumours. The direct comparison of the expression profiles of BCAR transfectants and oestrogen or EGF-stimulated cells strongly suggests that anti-oestrogen-resistant cell proliferation is not caused by alternative activation of the oestrogen receptor or by the epidermal growth factor receptor signalling pathway

  13. RNA sequencing supports distinct reactive oxygen species-mediated pathways of apoptosis by high and low size mass fractions of Bay leaf (Lauris nobilis) in HT-29 cells.

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    Rodd, Annabelle L; Ververis, Katherine; Sayakkarage, Dheeshana; Khan, Abdul W; Rafehi, Haloom; Ziemann, Mark; Loveridge, Shanon J; Lazarus, Ross; Kerr, Caroline; Lockett, Trevor; El-Osta, Assam; Karagiannis, Tom C; Bennett, Louise E

    2015-08-01

    Anti-proliferative and pro-apoptotic effects of Bay leaf (Laurus nobilis) in mammalian cancer and HT-29 adenocarcinoma cells have been previously attributed to effects of polyphenolic and essential oil chemical species. Recently, we demonstrated differentiated growth-regulating effects of high (HFBL) versus low molecular mass (LFBL) aqueous fractions of bay leaf and now confirm by comparative effects on gene expression, that HFBL and LFBL suppress HT-29 growth by distinct mechanisms. Induction of intra-cellular lesions including DNA strand breakage by extra-cellular HFBL, invoked the hypothesis that iron-mediated reactive oxygen species with capacity to penetrate cell membrane, were responsible for HFBL-mediated effects, supported by equivalent effects of HFBL in combination with γ radiation. Activities of HFBL and LFBL were interpreted to reflect differentiated responses to iron-mediated reactive oxygen species (ROS), occurring either outside or inside cells. In the presence of LFBL, apoptotic death was relatively delayed compared with HFBL. ROS production by LFBL mediated p53-dependent apoptosis and recovery was suppressed by promoting G1/S phase arrest and failure of cellular tight junctions. In comparison, intra-cellular anti-oxidant protection exerted by LFBL was absent for extra-cellular HFBL (likely polysaccharide-rich), which potentiated more rapid apoptosis by producing DNA double strand breaks. Differentiated effects on expression of genes regulating ROS defense and chromatic condensation by LFBL versus HFBL, were observed. The results support ferrous iron in cell culture systems and potentially in vivo, can invoke different extra-cellular versus intra-cellular ROS-mediated chemistries, that may be regulated by exogenous, including dietary species.

  14. Distinct domains within the NITROGEN LIMITATION ADAPTATION protein mediate its subcellular localization and function in the nitrate-dependent phosphate homeostasis pathway

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    The NITROGEN LIMITATION ADAPTATION (NLA) protein is a RING-type E3 ubiquitin ligase that plays an essential role in the regulation of nitrogen and phosphate homeostasis. NLA is localized to two distinct subcellular sites, the plasma membrane and nucleus, and contains four distinct domains: i) a RING...

  15. Cdc42 and phosphoinositide 3-kinase drive Rac-mediated actin polymerization downstream of c-Met in distinct and common pathways

    DEFF Research Database (Denmark)

    Bosse, Tanja; Ehinger, Julia; Czuchra, Aleksandra

    2007-01-01

    -WASP. Instead, actin polymerization was driven by Arp2/3 complex activation through the WAVE complex downstream of Rac. Together, our data establish an intricate signaling network comprising as key molecules Cdc42 and PI3-kinase, which converge on Rac-mediated actin reorganization essential for Listeria...

  16. Microarray and pathway analysis reveal distinct mechanisms underlying cannabinoid-mediated modulation of LPS-induced activation of BV-2 microglial cells.

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    Ana Juknat

    Full Text Available Cannabinoids are known to exert immunosuppressive activities. However, the mechanisms which contribute to these effects are unknown. Using lipopolysaccharide (LPS to activate BV-2 microglial cells, we examined how Δ(9-tetrahydrocannabinol (THC, the major psychoactive component of marijuana, and cannabidiol (CBD the non-psychoactive component, modulate the inflammatory response. Microarray analysis of genome-wide mRNA levels was performed using Illumina platform and the resulting expression patterns analyzed using the Ingenuity Pathway Analysis to identify functional subsets of genes, and the Ingenuity System Database to denote the gene networks regulated by CBD and THC. From the 5338 transcripts that were differentially expressed across treatments, 400 transcripts were found to be upregulated by LPS, 502 by CBD+LPS and 424 by THC+LPS, while 145 were downregulated by LPS, 297 by CBD+LPS and 149 by THC+LPS, by 2-fold or more (p≤0.005. Results clearly link the effects of CBD and THC to inflammatory signaling pathways and identify new cannabinoid targets in the MAPK pathway (Dusp1, Dusp8, Dusp2, cell cycle related (Cdkn2b, Gadd45a as well as JAK/STAT regulatory molecules (Socs3, Cish, Stat1. The impact of CBD on LPS-stimulated gene expression was greater than that of THC. We attribute this difference to the fact that CBD highly upregulated several genes encoding negative regulators of both NFκB and AP-1 transcriptional activities, such as Trib3 and Dusp1 known to be modulated through Nrf2 activation. The CBD-specific expression profile reflected changes associated with oxidative stress and glutathione depletion via Trib3 and expression of ATF4 target genes. Furthermore, the CBD affected genes were shown to be controlled by nuclear factors usually involved in regulation of stress response and inflammation, mainly via Nrf2/Hmox1 axis and the Nrf2/ATF4-Trib3 pathway. These observations indicate that CBD, and less so THC, induce a cellular stress

  17. Diverse exocytic pathways for mast cell mediators.

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    Xu, Hao; Bin, Na-Ryum; Sugita, Shuzo

    2018-04-17

    Mast cells play pivotal roles in innate and adaptive immunities but are also culprits in allergy, autoimmunity, and cardiovascular diseases. Mast cells respond to environmental changes by initiating regulated exocytosis/secretion of various biologically active compounds called mediators (e.g. proteases, amines, and cytokines). Many of these mediators are stored in granules/lysosomes and rely on intricate degranulation processes for release. Mast cell stabilizers (e.g. sodium cromoglicate), which prevent such degranulation processes, have therefore been clinically employed to treat asthma and allergic rhinitis. However, it has become increasingly clear that different mast cell diseases often involve multiple mediators that rely on overlapping but distinct mechanisms for release. This review illustrates existing evidence that highlights the diverse exocytic pathways in mast cells. We also discuss strategies to delineate these pathways so as to identify unique molecular components which could serve as new drug targets for more effective and specific treatments against mast cell-related diseases. © 2018 The Author(s). Published by Portland Press Limited on behalf of the Biochemical Society.

  18. PHOTOBIOMODULATION-MEDIATED PATHWAY DIAGNOSTICS

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    TIMON CHENG-YI LIU

    2013-01-01

    Full Text Available Cellular pathways are ordinarily diagnosed with pathway inhibitors, related gene regulation, or fluorescent protein markers. They are also suggested to be diagnosed with pathway activation modulation of photobiomodulation (PBM in this paper. A PBM on a biosystem function depends on whether the biosystem is in its function-specific homeostasis (FSH. An FSH, a negative feedback response for the function to be performed perfectly, is maintained by its FSH-essential subfunctions and its FSH-non-essential subfunctions (FNSs. A function in its FSH or far from its FSH is called a normal or dysfunctional function. A direct PBM may self-adaptatively modulate a dysfunctional function until it is normal so that it can be used to discover the optimum pathways for an FSH to be established. An indirect PBM may self-adaptatively modulate a dysfunctional FNS of a normal function until the FNS is normal, and the normal function is then upgraded so that it can be used to discover the redundant pathways for a normal function to be upgraded.

  19. Dispositional optimism and sleep quality: a test of mediating pathways.

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    Uchino, Bert N; Cribbet, Matthew; de Grey, Robert G Kent; Cronan, Sierra; Trettevik, Ryan; Smith, Timothy W

    2017-04-01

    Dispositional optimism has been related to beneficial influences on physical health outcomes. However, its links to global sleep quality and the psychological mediators responsible for such associations are less studied. This study thus examined if trait optimism predicted global sleep quality, and if measures of subjective well-being were statistical mediators of such links. A community sample of 175 participants (93 men, 82 women) completed measures of trait optimism, depression, and life satisfaction. Global sleep quality was assessed using the Pittsburgh Sleep Quality Index. Results indicated that trait optimism was a strong predictor of better PSQI global sleep quality. Moreover, this association was mediated by depression and life satisfaction in both single and multiple mediator models. These results highlight the importance of optimism for the restorative process of sleep, as well as the utility of multiple mediator models in testing distinct psychological pathways.

  20. Non-secreted clusterin isoforms are translated in rare amounts from distinct human mRNA variants and do not affect Bax-mediated apoptosis or the NF-κB signaling pathway.

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    Hans Prochnow

    Full Text Available Clusterin, also known as apolipoprotein J, is expressed from a variety of tissues and implicated in pathological disorders such as neurodegenerative diseases, ischemia and cancer. In contrast to secretory clusterin (sCLU, which acts as an extracellular chaperone, the synthesis, subcellular localization and function(s of intracellular CLU isoforms is currently a matter of intense discussion. By investigating human CLU mRNAs we here unravel mechanisms leading to the synthesis of distinct CLU protein isoforms and analyze their subcellular localization and their impact on apoptosis and on NF-κB-activity. Quantitative PCR-analyses revealed the expression of four different stress-inducible CLU mRNA variants in non-cancer and cancer cell lines. In all cell lines variant 1 represents the most abundant mRNA, whereas all other variants collectively account for no more than 0.34% of total CLU mRNA, even under stressed conditions. Overexpression of CLU cDNAs combined with in vitro mutagenesis revealed distinct translational start sites including a so far uncharacterized non-canonical CUG start codon. We show that all exon 2-containing mRNAs encode sCLU and at least three non-glycosylated intracellular isoforms, CLU1‑449, CLU21‑449 and CLU34‑449, which all reside in the cytosol of unstressed and stressed HEK‑293 cells. The latter is the only form expressed from an alternatively spliced mRNA variant lacking exon 2. Functional analysis revealed that none of these cytosolic CLU forms modulate caspase-mediated intrinsic apoptosis or significantly affects TNF-α-induced NF-κB-activity. Therefore our data challenge some of the current ideas regarding the physiological functions of CLU isoforms in pathologies.

  1. NK cell activation: distinct stimulatory pathways counterbalancing inhibitory signals.

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    Bakker, A B; Wu, J; Phillips, J H; Lanier, L L

    2000-01-01

    A delicate balance between positive and negative signals regulates NK cell effector function. Activation of NK cells may be initiated by the triggering of multiple adhesion or costimulatory molecules, and can be counterbalanced by inhibitory signals induced by receptors for MHC class I. A common pathway of inhibitory signaling is provided by immunoreceptor tyrosine-based inhibitory motifs (ITIMs) in the cytoplasmic domains of these receptors which mediate the recruitment of SH2 domain-bearing tyrosine phosphate-1 (SHP-1). In contrast to the extensive progress that has been made regarding the negative regulation of NK cell function, our knowledge of the signals that activate NK cells is still poor. Recent studies of the activating receptor complexes have shed new light on the induction of NK cell effector function. Several NK receptors using novel adaptors with immunoreceptor tyrosine-based activation motifs (ITAMs) and with PI 3-kinase recruiting motifs have been implicated in NK cell stimulation.

  2. Tp53 gene mediates distinct dopaminergic neuronal damage in different dopaminergic neurotoxicant models

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    Tao Lu

    2017-01-01

    Full Text Available Tp53, a stress response gene, is involved in diverse cell death pathways and its activation is implicated in the pathogenesis of Parkinson's disease. However, whether the neuronal Tp53 protein plays a direct role in regulating dopaminergic (DA neuronal cell death or neuronal terminal damage in different neurotoxicant models is unknown. In our recent studies, in contrast to the global inhibition of Tp53 function by pharmacological inhibitors and in traditional Tp53 knock-out mice, we examined the effects of DA-specific Tp53 gene deletion after 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine and methamphetamine exposure. Our data suggests that the Tp53 gene might be involved in both neuronal apoptosis and neuronal terminal damage caused by different neurotoxicants. Additional results from other studies also suggest that as a master regulator of many pathways that regulate apoptosis and synaptic terminal damage, it is possible that Tp53 may function as a signaling hub to integrate different signaling pathways to mediate distinctive target pathways. Tp53 protein as a signaling hub might be able to evaluate the microenvironment of neurons, assess the forms and severities of injury incurred, and determine whether apoptotic cell death or neuronal terminal degeneration occurs. Identification of the precise mechanisms activated in distinct neuronal damage caused by different forms and severities of injuries might allow for development of specific Tp53 inhibitors or ways to modulate distinct downstream target pathways involved.

  3. Distinct metal-exchange pathways of doped Ag25 nanoclusters

    KAUST Repository

    Bootharaju, Megalamane Siddaramappa

    2016-09-09

    Atomically precise metal nanoclusters (NCs) containing more than one type of metal atom (i.e., doped or alloyed), due to synergistic effects, open new avenues for engineering the catalytic and optical properties of NCs in a manner that homometal NCs cannot. Unfortunately, it is still a major challenge to controllably introduce multimetallic dopants in NCs, understanding the dopants\\' positions, mechanism, and synergistic effects. To overcome these challenges, we designed a metal-exchange approach involving NCs as molecular templates and metal ions as the source of the incoming dopant. In particular, two structurally similar monodoped silver-rich NCs, [MAg24(SR)(18)](2-) (M = Pd/Pt and SR: thiolate), were synthesized as templates to study their mechanistic transformation in response to the introduction of gold atoms. The controllable incorporation of Au atoms into the MAg24 framework facilitated the elucidation of distinct doping pathways through high-resolution mass spectrometry, optical spectroscopy and elemental analysis. Interestingly, gold replaced the central Pd atom of [PdAg24(SR)(18)](2-) clusters to produce predominantly bimetallic [AuAg24(SR)(18)](-) clusters along with a minor product of an [Au2Ag23(SR)(18)](-) cluster. In contrast, the central Pt atom remained intact in [PtAg24(SR)(18)](2-) clusters, and gold replaced the noncentral Ag atoms to form trimetallic [AuxPtAg24-x(SR)(18)](2-) NCs, where x = 1-2, with a portion of the starting [PtAg24(SR)(18)](2-) NCs remaining. This study reveals some of the unusual metal-exchange pathways of doped NCs and the important role played by the initial metal dopant in directing the position of a second dopant in the final product.

  4. Uniform and Complementary Social Interaction: Distinct Pathways to Solidarity.

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    Koudenburg, Namkje; Postmes, Tom; Gordijn, Ernestine H; van Mourik Broekman, Aafke

    2015-01-01

    We examine how different forms of co-action give rise to feelings of solidarity. We propose that (a) coordinated action elicits a sense of solidarity, and (b) the process through which such solidarity emerges differs for different forms of co-action. We suggest that whether solidarity within groups emerges from uniform action (e.g. synchronizing, as when people speak in unison) or from more complementary forms of action (e.g. alternating, when speaking in turns) has important consequences for the emergent position of individuals within the group. Uniform action relies on commonality, leaving little scope for individuality. In complementary action each individual makes a distinctive contribution to the group, thereby increasing a sense of personal value to the group, which should contribute to the emergence of solidarity. The predictions receive support from five studies, in which we study groups in laboratory and field settings. Results show that both complementary and uniform co-action increase a sense of solidarity compared to control conditions. However, in the complementary action condition, but not in the uniform action (or synchrony) condition, the effect on feelings of solidarity is mediated by a sense of personal value to the group.

  5. Endocytic pathways mediating oligomeric Aβ42 neurotoxicity

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    Laxton Kevin

    2010-05-01

    Full Text Available Abstract Background One pathological hallmark of Alzheimer's disease (AD is amyloid plaques, composed primarily of amyloid-β peptide (Aβ. Over-production or diminished clearance of the 42 amino acid form of Aβ (Aβ42 in the brain leads to accumulation of soluble Aβ and plaque formation. Soluble oligomeric Aβ (oAβ has recently emerged to be as a likely proximal cause of AD. Results Here we demonstrate that endocytosis is critical in mediating oAβ42-induced neurotoxicity and intraneuronal accumulation of Aβ. Inhibition of clathrin function either with a pharmacological inhibitor, knock-down of clathrin heavy chain expression, or expression of the dominant-negative mutant of clathrin-assembly protein AP180 did not block oAβ42-induced neurotoxicity or intraneuronal accumulation of Aβ. However, inhibition of dynamin and RhoA by expression of dominant negative mutants reduced neurotoxicity and intraneuronal Aβ accumulation. Pharmacologic inhibition of the dynamin-mediated endocytic pathway by genistein also reduced neurotoxicity. Conclusions These data suggest that dynamin-mediated and RhoA-regulated endocytosis are integral steps for oligomeric Aβ42-induced neurotoxicity and intraneuronal Aβ accumulation.

  6. Distinct brain systems mediate the effects of nociceptive input and self-regulation on pain.

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    Choong-Wan Woo

    2015-01-01

    Full Text Available Cognitive self-regulation can strongly modulate pain and emotion. However, it is unclear whether self-regulation primarily influences primary nociceptive and affective processes or evaluative ones. In this study, participants engaged in self-regulation to increase or decrease pain while experiencing multiple levels of painful heat during functional magnetic resonance imaging (fMRI imaging. Both heat intensity and self-regulation strongly influenced reported pain, but they did so via two distinct brain pathways. The effects of stimulus intensity were mediated by the neurologic pain signature (NPS, an a priori distributed brain network shown to predict physical pain with over 90% sensitivity and specificity across four studies. Self-regulation did not influence NPS responses; instead, its effects were mediated through functional connections between the nucleus accumbens and ventromedial prefrontal cortex. This pathway was unresponsive to noxious input, and has been broadly implicated in valuation, emotional appraisal, and functional outcomes in pain and other types of affective processes. These findings provide evidence that pain reports are associated with two dissociable functional systems: nociceptive/affective aspects mediated by the NPS, and evaluative/functional aspects mediated by a fronto-striatal system.

  7. Two distinct and competitive pathways confer the cellcidal actions of artemisinins

    Directory of Open Access Journals (Sweden)

    Chen Sun

    2015-01-01

    Full Text Available The biological actions of artemisinin (ART, an antimalarial drug derived from Artemisia annua, remain poorly understood and controversial. Besides potent antimalarial activity, some of artemisinin derivatives (together with artemisinin, hereafter referred to as ARTs, in particular dihydroartemisinin (DHA, are also associated with anticancer and other antiparasitic activities. In this study, we used baker’s yeast Saccharomyces cerevisiae as cellular and genetic model to investigate the molecular and cellular properties of ARTs. Two clearly separable pathways exist. While all ARTs exhibit potent anti-mitochondrial actions as shown before, DHA exerts an additional strong heme-dependent, likely mitochondria-independent inhibitory action. More importantly, heme antagonizes the mitochondria-dependent cellcidal action. Indeed, when heme synthesis was inhibited, the mitochondria-dependent cellcidal action of ARTs could be dramatically strengthened, and significant yeast growth inhibition at as low as 100 nM ART, an increase of about 25 folds in sensitivity, was observed. We conclude that ARTs are endowed with two major and distinct types of properties: a potent and specific mitochondria-dependent reaction and a more general and less specific heme-mediated reaction. The competitive nature of these two actions could be explained by their shared source of the consumable ARTs, so that inhibition of the heme-mediated degradation pathway would enable more ARTs to be available for the mitochondrial action. These properties of ARTs can be used to interpret the divergent antimalarial and anticancer actions of ARTs.

  8. Distinct patterns of IFITM-mediated restriction of filoviruses, SARS coronavirus, and influenza A virus.

    Directory of Open Access Journals (Sweden)

    I-Chueh Huang

    2011-01-01

    Full Text Available Interferon-inducible transmembrane proteins 1, 2, and 3 (IFITM1, 2, and 3 are recently identified viral restriction factors that inhibit infection mediated by the influenza A virus (IAV hemagglutinin (HA protein. Here we show that IFITM proteins restricted infection mediated by the entry glycoproteins (GP(1,2 of Marburg and Ebola filoviruses (MARV, EBOV. Consistent with these observations, interferon-β specifically restricted filovirus and IAV entry processes. IFITM proteins also inhibited replication of infectious MARV and EBOV. We observed distinct patterns of IFITM-mediated restriction: compared with IAV, the entry processes of MARV and EBOV were less restricted by IFITM3, but more restricted by IFITM1. Moreover, murine Ifitm5 and 6 did not restrict IAV, but efficiently inhibited filovirus entry. We further demonstrate that replication of infectious SARS coronavirus (SARS-CoV and entry mediated by the SARS-CoV spike (S protein are restricted by IFITM proteins. The profile of IFITM-mediated restriction of SARS-CoV was more similar to that of filoviruses than to IAV. Trypsin treatment of receptor-associated SARS-CoV pseudovirions, which bypasses their dependence on lysosomal cathepsin L, also bypassed IFITM-mediated restriction. However, IFITM proteins did not reduce cellular cathepsin activity or limit access of virions to acidic intracellular compartments. Our data indicate that IFITM-mediated restriction is localized to a late stage in the endocytic pathway. They further show that IFITM proteins differentially restrict the entry of a broad range of enveloped viruses, and modulate cellular tropism independently of viral receptor expression.

  9. β1-adrenergic receptors activate two distinct signaling pathways in striatal neurons

    Science.gov (United States)

    Meitzen, John; Luoma, Jessie I.; Stern, Christopher M.; Mermelstein, Paul G.

    2010-01-01

    Monoamine action in the dorsal striatum and nucleus accumbens plays essential roles in striatal physiology. Although research often focuses on dopamine and its receptors, norepinephrine and adrenergic receptors are also crucial in regulating striatal function. While noradrenergic neurotransmission has been identified in the striatum, little is known regarding the signaling pathways activated by β-adrenergic receptors in this brain region. Using cultured striatal neurons, we characterized a novel signaling pathway by which activation of β1-adrenergic receptors leads to the rapid phosphorylation of cAMP Response Element Binding Protein (CREB), a transcription-factor implicated as a molecular switch underlying long-term changes in brain function. Norepinephrine-mediated CREB phosphorylation requires β1-adrenergic receptor stimulation of a receptor tyrosine kinase, ultimately leading to the activation of a Ras/Raf/MEK/MAPK/MSK signaling pathway. Activation of β1-adrenergic receptors also induces CRE-dependent transcription and increased c-fos expression. In addition, stimulation of β1-adrenergic receptors produces cAMP production, but surprisingly, β1-adrenergic receptor activation of adenylyl cyclase was not functionally linked to rapid CREB phosphorylation. These findings demonstrate that activation of β1-adrenergic receptors on striatal neurons can stimulate two distinct signaling pathways. These adrenergic actions can produce long-term changes in gene expression, as well as rapidly modulate cellular physiology. By elucidating the mechanisms by which norepinephrine and β1-adrenergic receptor activation affects striatal physiology, we provide the means to more fully understand the role of monoamines in modulating striatal function, specifically how norepinephrine and β1-adrenergic receptors may affect striatal physiology. PMID:21143600

  10. Distinct pathways of neural coupling for different basic emotions.

    Science.gov (United States)

    Tettamanti, Marco; Rognoni, Elena; Cafiero, Riccardo; Costa, Tommaso; Galati, Dario; Perani, Daniela

    2012-01-16

    Emotions are complex events recruiting distributed cortical and subcortical cerebral structures, where the functional integration dynamics within the involved neural circuits in relation to the nature of the different emotions are still unknown. Using fMRI, we measured the neural responses elicited by films representing basic emotions (fear, disgust, sadness, happiness). The amygdala and the associative cortex were conjointly activated by all basic emotions. Furthermore, distinct arrays of cortical and subcortical brain regions were additionally activated by each emotion, with the exception of sadness. Such findings informed the definition of three effective connectivity models, testing for the functional integration of visual cortex and amygdala, as regions processing all emotions, with domain-specific regions, namely: i) for fear, the frontoparietal system involved in preparing adaptive motor responses; ii) for disgust, the somatosensory system, reflecting protective responses against contaminating stimuli; iii) for happiness: medial prefrontal and temporoparietal cortices involved in understanding joyful interactions. Consistently with these domain-specific models, the results of the effective connectivity analysis indicate that the amygdala is involved in distinct functional integration effects with cortical networks processing sensorimotor, somatosensory, or cognitive aspects of basic emotions. The resulting effective connectivity networks may serve to regulate motor and cognitive behavior based on the quality of the induced emotional experience. Copyright © 2011. Published by Elsevier Inc.

  11. Distinct Signaling Mechanisms in Multiple Developmental Pathways by the SCRAMBLED Receptor of Arabidopsis1[OPEN

    Science.gov (United States)

    Kwak, Su-Hwan; Woo, Sooah; Lee, Myeong Min; Schiefelbein, John

    2014-01-01

    SCRAMBLED (SCM), a leucine-rich repeat receptor-like kinase in Arabidopsis (Arabidopsis thaliana), is required for positional signaling in the root epidermis and for tissue/organ development in the shoot. To further understand SCM action, we generated a series of kinase domain variants and analyzed their ability to complement scm mutant defects. We found that the SCM kinase domain, but not kinase activity, is required for its role in root epidermal patterning, supporting the view that SCM is an atypical receptor kinase. We also describe a previously uncharacterized role for SCM in fruit dehiscence, because mature siliques from scm mutants fail to open properly. Interestingly, the kinase domain of SCM appears to be dispensable for this developmental process. Furthermore, we found that most of the SCM kinase domain mutations dramatically inhibit inflorescence development. Because this process is not affected in scm null mutants, it is likely that SCM acts redundantly to regulate inflorescence size. The importance of distinct kinase residues for these three developmental processes provides an explanation for the maintenance of the conserved kinase domain in the SCM protein, and it may generally explain its conservation in other atypical kinases. Furthermore, these results indicate that individual leucine-rich repeat receptor-like kinases may participate in multiple pathways using distinct signaling mechanisms to mediate diverse cellular communication events. PMID:25136062

  12. Distinct signaling mechanisms in multiple developmental pathways by the SCRAMBLED receptor of Arabidopsis.

    Science.gov (United States)

    Kwak, Su-Hwan; Woo, Sooah; Lee, Myeong Min; Schiefelbein, John

    2014-10-01

    SCRAMBLED (SCM), a leucine-rich repeat receptor-like kinase in Arabidopsis (Arabidopsis thaliana), is required for positional signaling in the root epidermis and for tissue/organ development in the shoot. To further understand SCM action, we generated a series of kinase domain variants and analyzed their ability to complement scm mutant defects. We found that the SCM kinase domain, but not kinase activity, is required for its role in root epidermal patterning, supporting the view that SCM is an atypical receptor kinase. We also describe a previously uncharacterized role for SCM in fruit dehiscence, because mature siliques from scm mutants fail to open properly. Interestingly, the kinase domain of SCM appears to be dispensable for this developmental process. Furthermore, we found that most of the SCM kinase domain mutations dramatically inhibit inflorescence development. Because this process is not affected in scm null mutants, it is likely that SCM acts redundantly to regulate inflorescence size. The importance of distinct kinase residues for these three developmental processes provides an explanation for the maintenance of the conserved kinase domain in the SCM protein, and it may generally explain its conservation in other atypical kinases. Furthermore, these results indicate that individual leucine-rich repeat receptor-like kinases may participate in multiple pathways using distinct signaling mechanisms to mediate diverse cellular communication events. © 2014 American Society of Plant Biologists. All Rights Reserved.

  13. DMPD: Signalling pathways mediating type I interferon gene expression. [Dynamic Macrophage Pathway CSML Database

    Lifescience Database Archive (English)

    Full Text Available 17904888 Signalling pathways mediating type I interferon gene expression. Edwards M...hways mediating type I interferon gene expression. PubmedID 17904888 Title Signalling pathways...R, Slater L, Johnston SL. Microbes Infect. 2007 Sep;9(11):1245-51. Epub 2007 Jul 1. (.png) (.svg) (.html) (.csml) Show Signalling pat

  14. Distinct Neural Mechanisms Mediate Olfactory Memory Formation at Different Timescales

    Science.gov (United States)

    McNamara, Ann Marie; Magidson, Phillip D.; Linster, Christiane; Wilson, Donald A.; Cleland, Thomas A.

    2008-01-01

    Habituation is one of the oldest forms of learning, broadly expressed across sensory systems and taxa. Here, we demonstrate that olfactory habituation induced at different timescales (comprising different odor exposure and intertrial interval durations) is mediated by different neural mechanisms. First, the persistence of habituation memory is…

  15. Distinct RNAi Pathways in the Regulation of Physiology and Development in the Fungus Mucor circinelloides.

    Science.gov (United States)

    Ruiz-Vázquez, Rosa M; Nicolás, Francisco E; Torres-Martínez, Santiago; Garre, Victoriano

    2015-01-01

    The basal fungus Mucor circinelloides has become, in recent years, a valuable model to study RNA-mediated gene silencing or RNA interference (RNAi). Serendipitously discovered in the late 1900s, the gene silencing in M. circinelloides is a landscape of consensus and dissents. Although similar to other classical fungal models in the basic design of the essential machinery that is responsible for silencing of gene expression, the existence of small RNA molecules of different sizes generated during this process and the presence of a mechanism that amplifies the silencing signal, give it a unique identity. In addition, M. circinelloides combines the components of RNAi machinery to carry out functions that not only limit themselves to the defense against foreign genetic material, but it uses some of these elements to regulate the expression of its own genes. Thus, different combinations of RNAi elements produce distinct classes of endogenous small RNAs (esRNAs) that regulate different physiological and developmental processes in response to environmental signals. The recent discovery of a new RNAi pathway involved in the specific degradation of endogenous mRNAs, using a novel RNase protein, adds one more element to the exciting puzzle of the gene silencing in M. circinelloides, in addition to providing hints about the evolutionary origin of the RNAi mechanism. Copyright © 2015 Elsevier Inc. All rights reserved.

  16. Task-dependent activation of distinct fast and slow(er) motor pathways during motor imagery.

    Science.gov (United States)

    Keller, Martin; Taube, Wolfgang; Lauber, Benedikt

    2018-02-22

    Motor imagery and actual movements share overlapping activation of brain areas but little is known about task-specific activation of distinct motor pathways during mental simulation of movements. For real contractions, it was demonstrated that the slow(er) motor pathways are activated differently in ballistic compared to tonic contractions but it is unknown if this also holds true for imagined contractions. The aim of the present study was to assess the activity of fast and slow(er) motor pathways during mentally simulated movements of ballistic and tonic contractions. H-reflexes were conditioned with transcranial magnetic stimulation at different interstimulus intervals to assess the excitability of fast and slow(er) motor pathways during a) the execution of tonic and ballistic contractions, b) motor imagery of these contraction types, and c) at rest. In contrast to the fast motor pathways, the slow(er) pathways displayed a task-specific activation: for imagined ballistic as well as real ballistic contractions, the activation was reduced compared to rest whereas enhanced activation was found for imagined tonic and real tonic contractions. This study provides evidence that the excitability of fast and slow(er) motor pathways during motor imagery resembles the activation pattern observed during real contractions. The findings indicate that motor imagery results in task- and pathway-specific subliminal activation of distinct subsets of neurons in the primary motor cortex. Copyright © 2018 The Author(s). Published by Elsevier Inc. All rights reserved.

  17. Distinct mechanisms act in concert to mediate cell cycle arrest.

    Science.gov (United States)

    Toettcher, Jared E; Loewer, Alexander; Ostheimer, Gerard J; Yaffe, Michael B; Tidor, Bruce; Lahav, Galit

    2009-01-20

    In response to DNA damage, cells arrest at specific stages in the cell cycle. This arrest must fulfill at least 3 requirements: it must be activated promptly; it must be sustained as long as damage is present to prevent loss of genomic information; and after the arrest, cells must re-enter into the appropriate cell cycle phase to ensure proper ploidy. Multiple molecular mechanisms capable of arresting the cell cycle have been identified in mammalian cells; however, it is unknown whether each mechanism meets all 3 requirements or whether they act together to confer specific functions to the arrest. To address this question, we integrated mathematical models describing the cell cycle and the DNA damage signaling networks and tested the contributions of each mechanism to cell cycle arrest and re-entry. Predictions from this model were then tested with quantitative experiments to identify the combined action of arrest mechanisms in irradiated cells. We find that different arrest mechanisms serve indispensable roles in the proper cellular response to DNA damage over time: p53-independent cyclin inactivation confers immediate arrest, whereas p53-dependent cyclin downregulation allows this arrest to be sustained. Additionally, p21-mediated inhibition of cyclin-dependent kinase activity is indispensable for preventing improper cell cycle re-entry and endoreduplication. This work shows that in a complex signaling network, seemingly redundant mechanisms, acting in a concerted fashion, can achieve a specific cellular outcome.

  18. Chemically different non-thermal plasmas target distinct cell death pathways

    Czech Academy of Sciences Publication Activity Database

    Lunov, O.; Zablotskyy, V.; Chrupina, O.; Lunova, M.; Jirsa, M.; Dejneka, A.; Kubinová, Šárka

    2017-01-01

    Roč. 7, apr (2017), s. 600 ISSN 2045-2322 R&D Projects: GA MŠk(CZ) LO1309 Institutional support: RVO:68378041 Keywords : chemically different * non-thermal plasmas * target distinct cell death pathways Subject RIV: FP - Other Medical Disciplines OBOR OECD: Biophysics Impact factor: 4.259, year: 2016

  19. The moderator-mediator variable distinction in social psychological research: conceptual, strategic, and statistical considerations.

    Science.gov (United States)

    Baron, R M; Kenny, D A

    1986-12-01

    In this article, we attempt to distinguish between the properties of moderator and mediator variables at a number of levels. First, we seek to make theorists and researchers aware of the importance of not using the terms moderator and mediator interchangeably by carefully elaborating, both conceptually and strategically, the many ways in which moderators and mediators differ. We then go beyond this largely pedagogical function and delineate the conceptual and strategic implications of making use of such distinctions with regard to a wide range of phenomena, including control and stress, attitudes, and personality traits. We also provide a specific compendium of analytic procedures appropriate for making the most effective use of the moderator and mediator distinction, both separately and in terms of a broader causal system that includes both moderators and mediators.

  20. Mycobacteria exploit three genetically distinct DNA double-strand break repair pathways.

    Science.gov (United States)

    Gupta, Richa; Barkan, Daniel; Redelman-Sidi, Gil; Shuman, Stewart; Glickman, Michael S

    2011-01-01

    Bacterial pathogens rely on their DNA repair pathways to resist genomic damage inflicted by the host. DNA double-strand breaks (DSBs) are especially threatening to bacterial viability. DSB repair by homologous recombination (HR) requires nucleases that resect DSB ends and a strand exchange protein that facilitates homology search. RecBCD and RecA perform these functions in Escherichia coli and constitute the major pathway of error-free DSB repair. Mycobacteria, including the human pathogen M. tuberculosis, elaborate an additional error-prone pathway of DSB repair via non-homologous end-joining (NHEJ) catalysed by Ku and DNA ligase D (LigD). Little is known about the relative contributions of HR and NHEJ to mycobacterial chromosome repair, the factors that dictate pathway choice, or the existence of additional DSB repair pathways. Here we demonstrate that Mycobacterium smegmatis has three DSB repair pathway options: HR, NHEJ and a novel mechanism of single-strand annealing (SSA). Inactivation of NHEJ or SSA is compensated by elevated HR. We find that mycobacterial RecBCD does not participate in HR or confer resistance to ionizing radiation (IR), but is required for the RecA-independent SSA pathway. In contrast, the mycobacterial helicase-nuclease AdnAB participates in the RecA-dependent HR pathway, and is a major determinant of resistance to IR and oxidative DNA damage. These findings reveal distinctive features of mycobacterial DSB repair, most notably the dedication of the RecBCD and AdnAB helicase-nuclease machines to distinct repair pathways. © 2010 Blackwell Publishing Ltd.

  1. Different AMPA receptor subtypes mediate the distinct kinetic components of a biphasic EPSC in hippocampal interneurons

    Directory of Open Access Journals (Sweden)

    Todd eStincic

    2015-05-01

    Full Text Available CA1 hippocampal interneurons at the border between stratum radiatum and stratum lacunosum-moleculare have AMPA receptor (AMPAR-mediated excitatory postsynaptic currents (EPSCs that consist of two distinct phases: a typical fast component (FC, and a highly unusual slow component (SC that persists for hundreds of milliseconds. To determine whether these kinetically distinct components of the EPSC are mediated by distinct AMPAR subpopulations, we examined the relative contributions of GluA2-containing and –lacking AMPARs to the SC. GluA2-containing AMPARs mediated the majority of the FC whereas GluA2-lacking AMPARs preferentially generated the SC. When glutamate uptake through the glial glutamate transporter EAAT1 was inhibited, spill over-mediated AMPAR activation recruited an even slower third kinetic component that persisted for several seconds; however, this spillover-mediated current was mediated predominantly by GluA2-containing AMPARs and therefore was clearly distinct from the SC when uptake is intact. Thus, different AMPAR subpopulations that vary in GluA2 content mediate the distinct components of the AMPAR EPSC. The SC is developmentally downregulated in mice, declining after the second postnatal week. This downregulation affects both GluA2-containing and GluA2-lacking AMPARs mediating the SC, and is not accompanied by developmental changes in the GluA2 content of AMPARs underlying the FC. Thus, the downregulation of the SC appears to be independent of synaptic GluA2 expression, suggesting the involvement of another AMPAR subunit or an auxiliary protein. Our results therefore identify GluA2-dependent and GluA2-independent determinants of the SC: GluA2-lacking AMPARs preferentially contribute to the SC, while the developmental downregulation of the SC is independent of GluA2 content.

  2. Parallel prefrontal pathways reach distinct excitatory and inhibitory systems in memory-related rhinal cortices.

    Science.gov (United States)

    Bunce, Jamie G; Zikopoulos, Basilis; Feinberg, Marcia; Barbas, Helen

    2013-12-15

    To investigate how prefrontal cortices impinge on medial temporal cortices we labeled pathways from the anterior cingulate cortex (ACC) and posterior orbitofrontal cortex (pOFC) in rhesus monkeys to compare their relationship with excitatory and inhibitory systems in rhinal cortices. The ACC pathway terminated mostly in areas 28 and 35 with a high proportion of large terminals, whereas the pOFC pathway terminated mostly through small terminals in area 36 and sparsely in areas 28 and 35. Both pathways terminated in all layers. Simultaneous labeling of pathways and distinct neurochemical classes of inhibitory neurons, followed by analyses of appositions of presynaptic and postsynaptic fluorescent signal, or synapses, showed overall predominant association with spines of putative excitatory neurons, but also significant interactions with presumed inhibitory neurons labeled for calretinin, calbindin, or parvalbumin. In the upper layers of areas 28 and 35 the ACC pathway was associated with dendrites of neurons labeled with calretinin, which are thought to disinhibit neighboring excitatory neurons, suggesting facilitated hippocampal access. In contrast, in area 36 pOFC axons were associated with dendrites of calbindin neurons, which are poised to reduce noise and enhance signal. In the deep layers, both pathways innervated mostly dendrites of parvalbumin neurons, which strongly inhibit neighboring excitatory neurons, suggesting gating of hippocampal output to other cortices. These findings suggest that the ACC, associated with attention and context, and the pOFC, associated with emotional valuation, have distinct contributions to memory in rhinal cortices, in processes that are disrupted in psychiatric diseases. Copyright © 2013 Wiley Periodicals, Inc.

  3. Chemically different non-thermal plasmas target distinct cell death pathways

    Czech Academy of Sciences Publication Activity Database

    Lunov, Oleg; Zablotskyy, Vitaliy A.; Churpita, Olexandr; Lunova, M.; Jirsa, M.; Dejneka, Alexandr; Kubinová, Šárka

    2017-01-01

    Roč. 7, č. 1 (2017), s. 1-17, č. článku 600. ISSN 2045-2322 Grant - others:AV ČR(CZ) Fellowship J. E. Purkyně Institutional support: RVO:68378271 Keywords : chemically different * non-thermal plasmas * target distinct cell death pathways Subject RIV: BO - Biophysics OBOR OECD: Biophysics Impact factor: 4.259, year: 2016

  4. Formation of distinct inclusion bodies by inhibition of ubiquitin-proteasome and autophagy-lysosome pathways

    International Nuclear Information System (INIS)

    Lee, Junho; Yang, Kyu-Hwan; Joe, Cheol O.; Kang, Seok-Seong

    2011-01-01

    Research highlights: → Distinct inclusion bodies are developed by inhibition of UPP and ALP. → The inclusion bodies differ in morphology, localization and formation process. → The inclusion bodies are distinguishable by the localization of TSC2. → Inhibition of both UPP and ALP simultaneously induces those inclusion bodies. -- Abstract: Accumulation of misfolded proteins is caused by the impairment of protein quality control systems, such as ubiquitin-proteasome pathway (UPP) and autophagy-lysosome pathway (ALP). In this study, the formation of inclusion bodies was examined after the blockade of UPP and/or ALP in A549 cells. UPP inhibition induced a single and large inclusion body localized in microtubule-organizing center. Interestingly, however, ALP inhibition generated dispersed small inclusion bodies in the cytoplasm. Tuberous sclerosis complex 2 was selectively accumulated in the inclusion bodies of UPP-inhibited cells, but not those of ALP-inhibited cells. Blockade of transcription and translation entirely inhibited the formation of inclusion body induced by UPP inhibition, but partially by ALP inhibition. Moreover, the simultaneous inhibition of two protein catabolic pathways independently developed two distinct inclusion bodies within a single cell. These findings clearly demonstrated that dysfunction of each catabolic pathway induced formation and accumulation of unique inclusion bodies on the basis of morphology, localization and formation process in A549 cells.

  5. Distinct Calcium Signaling Pathways Regulate Calmodulin Gene Expression in Tobacco1

    Science.gov (United States)

    van der Luit, Arnold H.; Olivari, Claudio; Haley, Ann; Knight, Marc R.; Trewavas, Anthony J.

    1999-01-01

    Cold shock and wind stimuli initiate Ca2+ transients in transgenic tobacco (Nicotiana plumbaginifolia) seedlings (named MAQ 2.4) containing cytoplasmic aequorin. To investigate whether these stimuli initiate Ca2+ pathways that are spatially distinct, stress-induced nuclear and cytoplasmic Ca2+ transients and the expression of a stress-induced calmodulin gene were compared. Tobacco seedlings were transformed with a construct that encodes a fusion protein between nucleoplasmin (a major oocyte nuclear protein) and aequorin. Immunocytochemical evidence indicated targeting of the fusion protein to the nucleus in these plants, which were named MAQ 7.11. Comparison between MAQ 7.11 and MAQ 2.4 seedlings confirmed that wind stimuli and cold shock invoke separate Ca2+ signaling pathways. Partial cDNAs encoding two tobacco calmodulin genes, NpCaM-1 and NpCaM-2, were identified and shown to have distinct nucleotide sequences that encode identical polypeptides. Expression of NpCaM-1, but not NpCaM-2, responded to wind and cold shock stimulation. Comparison of the Ca2+ dynamics with NpCaM-1 expression after stimulation suggested that wind-induced NpCaM-1 expression is regulated by a Ca2+ signaling pathway operational predominantly in the nucleus. In contrast, expression of NpCaM-1 in response to cold shock is regulated by a pathway operational predominantly in the cytoplasm. PMID:10557218

  6. Astrocytes mediate synapse elimination through MEGF10 and MERTK pathways

    Science.gov (United States)

    Chung, Won-Suk; Clarke, Laura E.; Wang, Gordon X.; Stafford, Benjamin K.; Sher, Alexander; Chakraborty, Chandrani; Joung, Julia; Foo, Lynette C.; Thompson, Andrew; Chen, Chinfei; Smith, Stephen J.; Barres, Ben A.

    2013-12-01

    To achieve its precise neural connectivity, the developing mammalian nervous system undergoes extensive activity-dependent synapse remodelling. Recently, microglial cells have been shown to be responsible for a portion of synaptic pruning, but the remaining mechanisms remain unknown. Here we report a new role for astrocytes in actively engulfing central nervous system synapses. This process helps to mediate synapse elimination, requires the MEGF10 and MERTK phagocytic pathways, and is strongly dependent on neuronal activity. Developing mice deficient in both astrocyte pathways fail to refine their retinogeniculate connections normally and retain excess functional synapses. Finally, we show that in the adult mouse brain, astrocytes continuously engulf both excitatory and inhibitory synapses. These studies reveal a novel role for astrocytes in mediating synapse elimination in the developing and adult brain, identify MEGF10 and MERTK as critical proteins in the synapse remodelling underlying neural circuit refinement, and have important implications for understanding learning and memory as well as neurological disease processes.

  7. Monoclonal Antibodies Follow Distinct Aggregation Pathways During Production-Relevant Acidic Incubation and Neutralization

    DEFF Research Database (Denmark)

    Pedersen, Thomas Skamris; Tian, Xinsheng; Thorolfsson, Matthias

    2016-01-01

    and orthogonal analytical methods, including small-angle X-ray scattering and dynamic light scattering and supplemented the experimental data with crystal structure-based spatial aggregation propensity (SAP) calculations. RESULTS: We revealed distinct solution behaviors between the three mAb models: At acidic p......PURPOSE: Aggregation aspects of therapeutic monoclonal antibodies (mAbs) are of common concern to the pharmaceutical industry. Low pH treatment is applied during affinity purification and to inactivate endogenous retroviruses, directing interest to the mechanisms of acid-induced antibody...... distinguish between reversible and irreversible mAb aggregation pathways at early stages of acidic treatment....

  8. [Cognition-Emotion Interactions and Psychopathic Personality: Distinct Pathways to Antisocial and Violent Behavior].

    Science.gov (United States)

    Verona, Edelyn

    Researchers have long acknowledged heterogeneity among persons who exhibit antisocial and violent behaviours. The study of psychopathic personality or psychopathy can help elucidate this heterogeneity through examination of the different facets that constitute this disorder. In particular, the distinct correlates of the interpersonal-affective traits (Factor 1) and the impulsive-antisocial traits (Factor 2) of psychopathy suggest at least two possible pathways to antisocial behaviours. Building on basic studies in cognitive and affective neuroscience, we provide a focused, non-comprehensive review of work identifying the biopsychological mechanisms involved in these two pathways, with special attention to studies using event-related potential (ERP) methods. In specific, a series of studies are discussed which examined affective and cognitive processes that may distinguish offenders high on psychopathic traits from other offenders, with emphasis on alterations in emotion-cognition interactions related to each factor of psychopathy. The set of findings reviewed highlight a central conclusion: Factor 1 represents a pathway involving reduced emotional responding, exacerbated by attentional abnormalities, that make for a more deliberate and emotionally insensitive offender profile. In contrast, Factor 2 characterizes a pathway marked by emotional and behavioural dysregulation and cognitive control dysfunctions, particularly in emotional contexts. Implications for identifying etiological processes and the further understanding of antisocial and violent behaviours are discussed.

  9. Formation of distinct inclusion bodies by inhibition of ubiquitin-proteasome and autophagy-lysosome pathways.

    Science.gov (United States)

    Lee, Junho; Yang, Kyu-Hwan; Joe, Cheol O; Kang, Seok-Seong

    2011-01-14

    Accumulation of misfolded proteins is caused by the impairment of protein quality control systems, such as ubiquitin-proteasome pathway (UPP) and autophagy-lysosome pathway (ALP). In this study, the formation of inclusion bodies was examined after the blockade of UPP and/or ALP in A549 cells. UPP inhibition induced a single and large inclusion body localized in microtubule-organizing center. Interestingly, however, ALP inhibition generated dispersed small inclusion bodies in the cytoplasm. Tuberous sclerosis complex 2 was selectively accumulated in the inclusion bodies of UPP-inhibited cells, but not those of ALP-inhibited cells. Blockade of transcription and translation entirely inhibited the formation of inclusion body induced by UPP inhibition, but partially by ALP inhibition. Moreover, the simultaneous inhibition of two protein catabolic pathways independently developed two distinct inclusion bodies within a single cell. These findings clearly demonstrated that dysfunction of each catabolic pathway induced formation and accumulation of unique inclusion bodies on the basis of morphology, localization and formation process in A549 cells. Copyright © 2010 Elsevier Inc. All rights reserved.

  10. Azospirillum brasilense Chemotaxis Depends on Two Signaling Pathways Regulating Distinct Motility Parameters.

    Science.gov (United States)

    Mukherjee, Tanmoy; Kumar, Dhivya; Burriss, Nathan; Xie, Zhihong; Alexandre, Gladys

    2016-06-15

    The genomes of most motile bacteria encode two or more chemotaxis (Che) systems, but their functions have been characterized in only a few model systems. Azospirillum brasilense is a motile soil alphaproteobacterium able to colonize the rhizosphere of cereals. In response to an attractant, motile A. brasilense cells transiently increase swimming speed and suppress reversals. The Che1 chemotaxis pathway was previously shown to regulate changes in the swimming speed, but it has a minor role in chemotaxis and root surface colonization. Here, we show that a second chemotaxis system, named Che4, regulates the probability of swimming reversals and is the major signaling pathway for chemotaxis and wheat root surface colonization. Experimental evidence indicates that Che1 and Che4 are functionally linked to coordinate changes in the swimming motility pattern in response to attractants. The effect of Che1 on swimming speed is shown to enhance the aerotactic response of A. brasilense in gradients, likely providing the cells with a competitive advantage in the rhizosphere. Together, the results illustrate a novel mechanism by which motile bacteria utilize two chemotaxis pathways regulating distinct motility parameters to alter movement in gradients and enhance the chemotactic advantage. Chemotaxis provides motile bacteria with a competitive advantage in the colonization of diverse niches and is a function enriched in rhizosphere bacterial communities, with most species possessing at least two chemotaxis systems. Here, we identify the mechanism by which cells may derive a significant chemotactic advantage using two chemotaxis pathways that ultimately regulate distinct motility parameters. Copyright © 2016, American Society for Microbiology. All Rights Reserved.

  11. Histone Deacetylase Inhibitors Antagonize Distinct Pathways to Suppress Tumorigenesis of Embryonal Rhabdomyosarcoma.

    Directory of Open Access Journals (Sweden)

    Terra Vleeshouwer-Neumann

    Full Text Available Embryonal rhabdomyosarcoma (ERMS is the most common soft tissue cancer in children. The prognosis of patients with relapsed or metastatic disease remains poor. ERMS genomes show few recurrent mutations, suggesting that other molecular mechanisms such as epigenetic regulation might play a major role in driving ERMS tumor biology. In this study, we have demonstrated the diverse roles of histone deacetylases (HDACs in the pathogenesis of ERMS by characterizing effects of HDAC inhibitors, trichostatin A (TSA and suberoylanilide hydroxamic acid (SAHA; also known as vorinostat in vitro and in vivo. TSA and SAHA suppress ERMS tumor growth and progression by inducing myogenic differentiation as well as reducing the self-renewal and migratory capacity of ERMS cells. Differential expression profiling and pathway analysis revealed downregulation of key oncogenic pathways upon HDAC inhibitor treatment. By gain-of-function, loss-of-function, and chromatin immunoprecipitation (ChIP studies, we show that Notch1- and EphrinB1-mediated pathways are regulated by HDACs to inhibit differentiation and enhance migratory capacity of ERMS cells, respectively. Our study demonstrates that aberrant HDAC activity plays a major role in ERMS pathogenesis. Druggable targets in the molecular pathways affected by HDAC inhibitors represent novel therapeutic options for ERMS patients.

  12. Can a Mediator Moderate? Considering the Role of Time and Change in the Mediator-Moderator Distinction.

    Science.gov (United States)

    Karazsia, Bryan T; Berlin, Kristoffer S

    2018-01-01

    The concepts of mediation and moderation are important for specifying ways in which psychological treatments work and for whom they are most beneficial. Historically, the terms were confused and used interchangeably, so a rich body of scholarly literature makes clear their distinction. Researchers are also becoming increasingly aware that mediation and moderation can be integrated and that such integration can advance theory development and testing. One question that has not received sufficient attention is whether a mediator can simultaneously moderate. We tackle this question in this paper, and in doing so we expand on the MacArthur conceptualizations of mediation and moderation. The result is a presentation of a meta-theoretical model that illustrates how a construct that is initially a mediator can, not simultaneously but over time, evolve into a construct that moderates. When this occurs, a construct that changed for the better as a result of an intervention can later promote more positive change during a later intervention. Various implications of this novel paradigm for future research are discussed, including the importance of this model in the emerging context of managed health care. Copyright © 2017. Published by Elsevier Ltd.

  13. Understanding nanoparticle-mediated nucleation pathways of anisotropic nanoparticles

    Science.gov (United States)

    Laramy, Christine R.; Fong, Lam-Kiu; Jones, Matthew R.; O'Brien, Matthew N.; Schatz, George C.; Mirkin, Chad A.

    2017-09-01

    Several seed-mediated syntheses of low symmetry anisotropic nanoparticles yield broad product distributions with multiple defect structures. This observation challenges the role of the nanoparticle precursor as a seed for certain syntheses and suggests the possibility of alternate nucleation pathways. Herein, we report a method to probe the role of the nanoparticle precursor in anisotropic nanoparticle nucleation with compositional and structural 'labels' to track their fate. We use the synthesis of gold triangular nanoprisms (Au TPs) as a model system. We propose a mechanism in which, rather than acting as a template, the nanoparticle precursor catalyzes homogenous nucleation of Au TPs.

  14. Selective Modulation of Integrin-mediated Cell Migration by Distinct ADAM Family MembersV⃞

    Science.gov (United States)

    Huang, Jing; Bridges, Lance C.; White, Judith M.

    2005-01-01

    A disintegrin and a metalloprotease (ADAM) family members have been implicated in many biological processes. Although it is recognized that recombinant ADAM disintegrin domains can interact with integrins, little is known about ADAM-integrin interactions in cellular context. Here, we tested whether ADAMs can selectively regulate integrin-mediated cell migration. ADAMs were expressed in Chinese hamster ovary cells that express defined integrins (α4β1, α5β1, or both), and cell migration on full-length fibronectin or on its α4β1 or α5β1 binding fragments was studied. We found that ADAMs inhibit integrin-mediated cell migration in patterns dictated by the integrin binding profiles of their isolated disintegrin domains. ADAM12 inhibited cell migration mediated by the α4β1 but not the α5β1 integrin. ADAM17 had the reciprocal effect; it inhibited α5β1- but not α4β1-mediated cell migration. ADAM19 and ADAM33 inhibited migration mediated by both α4β1 and α5β1 integrins. A point mutation in the ADAM12 disintegrin loop partially reduced the inhibitory effect of ADAM12 on cell migration on the α4β1 binding fragment of fibronectin, whereas mutations that block metalloprotease activity had no effect. Our results indicate that distinct ADAMs can modulate cell migration mediated by specific integrins in a pattern dictated, at least in part, by their disintegrin domains. PMID:16079176

  15. Genetic approach identifies distinct asthma pathways in overweight vs normal weight children.

    Science.gov (United States)

    Butsch Kovacic, M; Martin, L J; Biagini Myers, J M; He, H; Lindsey, M; Mersha, T B; Khurana Hershey, G K

    2015-08-01

    The pathogenesis of asthma in the context of excess body weight may be distinct from asthma that develops in normal weight children. The study's objective was to explore the biology of asthma in the context of obesity and normal weight status using genetic methodologies. Associations between asthma and SNPs in 49 genes were assessed, as well as, interactions between SNPs and overweight status in child participants of the Greater Cincinnati Pediatric Clinic Repository. Asthma was significantly associated with weight (OR = 1.38; P = 0.037). The number of genes and the magnitude of their associations with asthma were notably greater when considering overweight children alone vs normal weight and overweight children together. When considering weight, distinct sets of asthma-associated genes were observed, many times with opposing effects. We demonstrated that the underlying heterogeneity of asthma is likely due in part to distinct pathogenetic pathways that depend on preceding/comorbid overweight and/or allergy. It is therefore important to consider both obesity and asthma when conducting studies of asthma. © 2015 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.

  16. Heat-shock stress activates a novel nuclear import pathway mediated by Hikeshi

    OpenAIRE

    Imamoto, Naoko; Kose, Shingo

    2012-01-01

    Cellular stresses significantly affect nuclear transport systems. Nuclear transport pathways mediated by importin β-family members, which are active under normal conditions, are downregulated. During thermal stress, a nuclear import pathway mediated by a novel carrier, which we named Hikeshi, becomes active. Hikeshi is not a member of the importin β family and mediates the nuclear import of Hsp70s. Unlike importin β family-mediated nuclear transport, the Hikeshi-mediated nuclear import of Hsp...

  17. Spermidine and resveratrol induce autophagy by distinct pathways converging on the acetylproteome

    DEFF Research Database (Denmark)

    Morselli, Eugenia; Mariño, Guillermo; Bennetzen, Martin V

    2011-01-01

    Autophagy protects organelles, cells, and organisms against several stress conditions. Induction of autophagy by resveratrol requires the nicotinamide adenine dinucleotide-dependent deacetylase sirtuin 1 (SIRT1). In this paper, we show that the acetylase inhibitor spermidine stimulates autophagy...... independent of SIRT1 in human and yeast cells as well as in nematodes. Although resveratrol and spermidine ignite autophagy through distinct mechanisms, these compounds stimulate convergent pathways that culminate in concordant modifications of the acetylproteome. Both agents favor convergent deacetylation...... and acetylation reactions in the cytosol and in the nucleus, respectively. Both resveratrol and spermidine were able to induce autophagy in cytoplasts (enucleated cells). Moreover, a cytoplasm-restricted mutant of SIRT1 could stimulate autophagy, suggesting that cytoplasmic deacetylation reactions dictate...

  18. Spermidine and resveratrol induce autophagy by distinct pathways converging on the acetylproteome.

    Science.gov (United States)

    Morselli, Eugenia; Mariño, Guillermo; Bennetzen, Martin V; Eisenberg, Tobias; Megalou, Evgenia; Schroeder, Sabrina; Cabrera, Sandra; Bénit, Paule; Rustin, Pierre; Criollo, Alfredo; Kepp, Oliver; Galluzzi, Lorenzo; Shen, Shensi; Malik, Shoaib Ahmad; Maiuri, Maria Chiara; Horio, Yoshiyuki; López-Otín, Carlos; Andersen, Jens S; Tavernarakis, Nektarios; Madeo, Frank; Kroemer, Guido

    2011-02-21

    Autophagy protects organelles, cells, and organisms against several stress conditions. Induction of autophagy by resveratrol requires the nicotinamide adenine dinucleotide-dependent deacetylase sirtuin 1 (SIRT1). In this paper, we show that the acetylase inhibitor spermidine stimulates autophagy independent of SIRT1 in human and yeast cells as well as in nematodes. Although resveratrol and spermidine ignite autophagy through distinct mechanisms, these compounds stimulate convergent pathways that culminate in concordant modifications of the acetylproteome. Both agents favor convergent deacetylation and acetylation reactions in the cytosol and in the nucleus, respectively. Both resveratrol and spermidine were able to induce autophagy in cytoplasts (enucleated cells). Moreover, a cytoplasm-restricted mutant of SIRT1 could stimulate autophagy, suggesting that cytoplasmic deacetylation reactions dictate the autophagic cascade. At doses at which neither resveratrol nor spermidine stimulated autophagy alone, these agents synergistically induced autophagy. Altogether, these data underscore the importance of an autophagy regulatory network of antagonistic deacetylases and acetylases that can be pharmacologically manipulated.

  19. Infectious Entry Pathway Mediated by the Human Endogenous Retrovirus K Envelope Protein.

    Science.gov (United States)

    Robinson, Lindsey R; Whelan, Sean P J

    2016-01-20

    Endogenous retroviruses (ERVs), the majority of which exist as degraded remnants of ancient viruses, comprise approximately 8% of the human genome. The youngest human ERVs (HERVs) belong to the HERV-K(HML-2) subgroup and were endogenized within the past 1 million years. The viral envelope protein (ENV) facilitates the earliest events of endogenization (cellular attachment and entry), and here, we characterize the requirements for HERV-K ENV to mediate infectious cell entry. Cell-cell fusion assays indicate that a minimum of two events are required for fusion, proteolytic processing by furin-like proteases and exposure to acidic pH. We generated an infectious autonomously replicating recombinant vesicular stomatitis virus (VSV) in which the glycoprotein was replaced by HERV-K ENV. HERV-K ENV imparts an endocytic entry pathway that requires dynamin-mediated membrane scission and endosomal acidification but is distinct from clathrin-dependent or macropinocytic uptake pathways. The lack of impediments to the replication of the VSV core in eukaryotic cells allowed us to broadly survey the HERV-K ENV-dictated tropism. Unlike extant betaretroviral envelopes, which impart a narrow species tropism, we found that HERV-K ENV mediates broad tropism encompassing cells from multiple mammalian and nonmammalian species. We conclude that HERV-K ENV dictates an evolutionarily conserved entry pathway and that the restriction of HERV-K to primate genomes reflects downstream stages of the viral replication cycle. Approximately 8% of the human genome is of retroviral origin. While many of those viral genomes have become inactivated, some copies of the most recently endogenized human retrovirus, HERV-K, can encode individual functional proteins. Here, we characterize the envelope protein (ENV) of the virus to define how it mediates infection of cells. We demonstrate that HERV-K ENV undergoes a proteolytic processing step and triggers membrane fusion in response to acidic pH--a strategy

  20. Molecular Signaling Pathways Mediating Osteoclastogenesis Induced by Prostate Cancer Cells

    International Nuclear Information System (INIS)

    Rafiei, Shahrzad; Komarova, Svetlana V

    2013-01-01

    Advanced prostate cancer commonly metastasizes to bone leading to osteoblastic and osteolytic lesions. Although an osteolytic component governed by activation of bone resorbing osteoclasts is prominent in prostate cancer metastasis, the molecular mechanisms of prostate cancer-induced osteoclastogenesis are not well-understood. We studied the effect of soluble mediators released from human prostate carcinoma cells on osteoclast formation from mouse bone marrow and RAW 264.7 monocytes. Soluble factors released from human prostate carcinoma cells significantly increased viability of naïve bone marrow monocytes, as well as osteoclastogenesis from precursors primed with receptor activator of nuclear factor κ-B ligand (RANKL). The prostate cancer-induced osteoclastogenesis was not mediated by RANKL as it was not inhibited by osteoprotegerin (OPG). However inhibition of TGFβ receptor I (TβRI), or macrophage-colony stimulating factor (MCSF) resulted in attenuation of prostate cancer-induced osteoclastogenesis. We characterized the signaling pathways induced in osteoclast precursors by soluble mediators released from human prostate carcinoma cells. Prostate cancer factors increased basal calcium levels and calcium fluctuations, induced nuclear localization of nuclear factor of activated t-cells (NFAT)c1, and activated prolonged phosphorylation of ERK1/2 in RANKL-primed osteoclast precursors. Inhibition of calcium signaling, NFATc1 activation, and ERK1/2 phosphorylation significantly reduced the ability of prostate cancer mediators to stimulate osteoclastogenesis. This study reveals the molecular mechanisms underlying the direct osteoclastogenic effect of prostate cancer derived factors, which may be beneficial in developing novel osteoclast-targeting therapeutic approaches

  1. Extracellular Monomeric and Aggregated Tau Efficiently Enter Human Neurons through Overlapping but Distinct Pathways

    Directory of Open Access Journals (Sweden)

    Lewis D. Evans

    2018-03-01

    Full Text Available Summary: In Alzheimer’s disease, neurofibrillary tangle pathology appears to spread along neuronal connections, proposed to be mediated by the release and uptake of abnormal, disease-specific forms of microtubule-binding protein tau MAPT. It is currently unclear whether transfer of tau between neurons is a toxic gain-of-function process in dementia or reflects a constitutive biological process. We report two entry mechanisms for monomeric tau to human neurons: a rapid dynamin-dependent phase typical of endocytosis and a second, slower actin-dependent phase of macropinocytosis. Aggregated tau entry is independent of actin polymerization and largely dynamin dependent, consistent with endocytosis and distinct from macropinocytosis, the major route for aggregated tau entry reported for non-neuronal cells. Anti-tau antibodies abrogate monomeric tau entry into neurons, but less efficiently in the case of aggregated tau, where internalized tau carries antibody with it into neurons. These data suggest that tau entry to human neurons is a physiological process and not a disease-specific phenomenon. : In contrast with predictions that transfer of the microtubule-associated protein tau between neurons is a toxic gain-of-function process in dementia, Evans et al. show that healthy human neurons efficiently take up both normal and aggregated tau, by distinct but overlapping uptake mechanisms. Keywords: Alzheimer’s disease, frontotemporal dementia, Tau, MAPT, iPSC, endocytosis, human neurons, intracellular transport

  2. Distinct signalling properties of insulin receptor substrate (IRS)-1 and IRS-2 in mediating insulin/IGF-1 action.

    Science.gov (United States)

    Rabiee, Atefeh; Krüger, Marcus; Ardenkjær-Larsen, Jacob; Kahn, C Ronald; Emanuelli, Brice

    2018-07-01

    Insulin/IGF-1 action is driven by a complex and highly integrated signalling network. Loss-of-function studies indicate that the major insulin/IGF-1 receptor substrate (IRS) proteins, IRS-1 and IRS-2, mediate different biological functions in vitro and in vivo, suggesting specific signalling properties despite their high degree of homology. To identify mechanisms contributing to the differential signalling properties of IRS-1 and IRS-2 in the mediation of insulin/IGF-1 action, we performed comprehensive mass spectrometry (MS)-based phosphoproteomic profiling of brown preadipocytes from wild type, IRS-1 -/- and IRS-2 -/- mice in the basal and IGF-1-stimulated states. We applied stable isotope labeling by amino acids in cell culture (SILAC) for the accurate quantitation of changes in protein phosphorylation. We found ~10% of the 6262 unique phosphorylation sites detected to be regulated by IGF-1. These regulated sites included previously reported substrates of the insulin/IGF-1 signalling pathway, as well as novel substrates including Nuclear Factor I X and Semaphorin-4B. In silico prediction suggests the protein kinase B (PKB), protein kinase C (PKC), and cyclin-dependent kinase (CDK) as the main mediators of these phosphorylation events. Importantly, we found preferential phosphorylation patterns depending on the presence of either IRS-1 or IRS-2, which was associated with specific sets of kinases involved in signal transduction downstream of these substrates such as PDHK1, MAPK3, and PKD1 for IRS-1, and PIN1 and PKC beta for IRS-2. Overall, by generating a comprehensive phosphoproteomic profile from brown preadipocyte cells in response to IGF-1 stimulation, we reveal both common and distinct insulin/IGF-1 signalling events mediated by specific IRS proteins. Copyright © 2018 Elsevier Inc. All rights reserved.

  3. Distinct configurations of protein complexes and biochemical pathways revealed by epistatic interaction network motifs

    LENUS (Irish Health Repository)

    Casey, Fergal

    2011-08-22

    Abstract Background Gene and protein interactions are commonly represented as networks, with the genes or proteins comprising the nodes and the relationship between them as edges. Motifs, or small local configurations of edges and nodes that arise repeatedly, can be used to simplify the interpretation of networks. Results We examined triplet motifs in a network of quantitative epistatic genetic relationships, and found a non-random distribution of particular motif classes. Individual motif classes were found to be associated with different functional properties, suggestive of an underlying biological significance. These associations were apparent not only for motif classes, but for individual positions within the motifs. As expected, NNN (all negative) motifs were strongly associated with previously reported genetic (i.e. synthetic lethal) interactions, while PPP (all positive) motifs were associated with protein complexes. The two other motif classes (NNP: a positive interaction spanned by two negative interactions, and NPP: a negative spanned by two positives) showed very distinct functional associations, with physical interactions dominating for the former but alternative enrichments, typical of biochemical pathways, dominating for the latter. Conclusion We present a model showing how NNP motifs can be used to recognize supportive relationships between protein complexes, while NPP motifs often identify opposing or regulatory behaviour between a gene and an associated pathway. The ability to use motifs to point toward underlying biological organizational themes is likely to be increasingly important as more extensive epistasis mapping projects in higher organisms begin.

  4. Autism spectrum disorders and drug addiction: Common pathways, common molecules, distinct disorders?

    Directory of Open Access Journals (Sweden)

    Patrick E. Rothwell

    2016-02-01

    Full Text Available Autism spectrum disorders (ASDs and drug addiction do not share substantial comorbidity or obvious similarities in etiology or symptomatology. It is thus surprising that a number of recent studies implicate overlapping neural circuits and molecular signaling pathways in both disorders. The purpose of this review is to highlight this emerging intersection and consider implications for understanding the pathophysiology of these seemingly distinct disorders. One area of overlap involves neural circuits and neuromodulatory systems in the striatum and basal ganglia, which play an established role in addiction and reward but are increasingly implicated in clinical and preclinical studies of ASDs. A second area of overlap relates to molecules like Fragile X mental retardation protein (FMRP and methyl CpG-binding protein-2 (MECP2, which are best known for their contribution to the pathogenesis of syndromic ASDs, but have recently been shown to regulate behavioral and neurobiological responses to addictive drug exposure. These shared pathways and molecules point to common dimensions of behavioral dysfunction, including the repetition of behavioral patterns and aberrant reward processing. The synthesis of knowledge gained through parallel investigations of ASDs and addiction may inspire the design of new therapeutic interventions to correct common elements of striatal dysfunction.

  5. Autism Spectrum Disorders and Drug Addiction: Common Pathways, Common Molecules, Distinct Disorders?

    Science.gov (United States)

    Rothwell, Patrick E

    2016-01-01

    Autism spectrum disorders (ASDs) and drug addiction do not share substantial comorbidity or obvious similarities in etiology or symptomatology. It is thus surprising that a number of recent studies implicate overlapping neural circuits and molecular signaling pathways in both disorders. The purpose of this review is to highlight this emerging intersection and consider implications for understanding the pathophysiology of these seemingly distinct disorders. One area of overlap involves neural circuits and neuromodulatory systems in the striatum and basal ganglia, which play an established role in addiction and reward but are increasingly implicated in clinical and preclinical studies of ASDs. A second area of overlap relates to molecules like Fragile X mental retardation protein (FMRP) and methyl CpG-binding protein-2 (MECP2), which are best known for their contribution to the pathogenesis of syndromic ASDs, but have recently been shown to regulate behavioral and neurobiological responses to addictive drug exposure. These shared pathways and molecules point to common dimensions of behavioral dysfunction, including the repetition of behavioral patterns and aberrant reward processing. The synthesis of knowledge gained through parallel investigations of ASDs and addiction may inspire the design of new therapeutic interventions to correct common elements of striatal dysfunction.

  6. Distinct protease pathways control cell shape and apoptosis in v-src-transformed quail neuroretina cells

    International Nuclear Information System (INIS)

    Neel, Benjamin D.; Aouacheria, Abdel; Nouvion, Anne-Laure; Ronot, Xavier; Gillet, Germain

    2005-01-01

    Intracellular proteases play key roles in cell differentiation, proliferation and apoptosis. In nerve cells, little is known about their relative contribution to the pathways which control cell physiology, including cell death. Neoplastic transformation of avian neuroretina cells by p60 v-src tyrosine kinase results in dramatic morphological changes and deregulation of apoptosis. To identify the proteases involved in the cellular response to p60 v-src , we evaluated the effect of specific inhibitors of caspases, calpains and the proteasome on cell shape changes and apoptosis induced by p60 v-src inactivation in quail neuroretina cells transformed by tsNY68, a thermosensitive strain of Rous sarcoma virus. We found that the ubiquitin-proteasome pathway is recruited early after p60 v-src inactivation and is critical for morphological changes, whereas caspases are essential for cell death. This study provides evidence that distinct intracellular proteases are involved in the control of the morphology and fate of v-src-transformed cells

  7. Steroidal androgens and nonsteroidal, tissue-selective androgen receptor modulator, S-22, regulate androgen receptor function through distinct genomic and nongenomic signaling pathways.

    Science.gov (United States)

    Narayanan, Ramesh; Coss, Christopher C; Yepuru, Muralimohan; Kearbey, Jeffrey D; Miller, Duane D; Dalton, James T

    2008-11-01

    Androgen receptor (AR) ligands are important for the development and function of several tissues and organs. However, the poor oral bioavailability, pharmacokinetic properties, and receptor cross-reactivity of testosterone, coupled with side effects, place limits on its clinical use. Selective AR modulators (SARMs) elicit anabolic effects in muscle and bone, sparing reproductive organs like the prostate. However, molecular mechanisms underlying the tissue selectivity remain ambiguous. We performed a variety of in vitro studies to compare and define the molecular mechanisms of an aryl propionamide SARM, S-22, as compared with dihydrotestosterone (DHT). Studies indicated that S-22 increased levator ani muscle weight but decreased the size of prostate in rats. Analysis of the upstream intracellular signaling events indicated that S-22 and DHT mediated their actions through distinct pathways. Modulation of these pathways altered the recruitment of AR and its cofactors to the PSA enhancer in a ligand-dependent fashion. In addition, S-22 induced Xenopus laevis oocyte maturation and rapid phosphorylation of several kinases, through pathways distinct from steroids. These studies reveal novel differences in the molecular mechanisms by which S-22, a nonsteroidal SARM, and DHT mediate their pharmacological effects.

  8. Phyllodes tumors with and without fibroadenoma-like areas display distinct genomic features and may evolve through distinct pathways.

    Science.gov (United States)

    Pareja, Fresia; Geyer, Felipe C; Kumar, Rahul; Selenica, Pier; Piscuoglio, Salvatore; Ng, Charlotte K Y; Burke, Kathleen A; Edelweiss, Marcia; Murray, Melissa P; Brogi, Edi; Weigelt, Britta; Reis-Filho, Jorge S

    2017-01-01

    Breast fibroepithelial lesions (fibroadenomas and phyllodes tumors) are underpinned by recurrent MED12 exon 2 mutations, which are more common in fibroadenomas and benign phyllodes tumors. TERT promoter hotspot mutations have been documented in phyllodes tumors, and found to be more frequent in borderline and malignant lesions. Several lines of evidence suggest that a subset of phyllodes tumors might arise from fibroadenomas. Here we sought to investigate the genetic differences between phyllodes tumors with fibroadenoma-like areas vs. those without. We retrieved data for 16 borderline/ malignant phyllodes tumors, including seven phyllodes tumors with fibroadenoma-like areas and nine phyllodes tumors without fibroadenoma-like areas, which had been previously subjected to targeted capture massively parallel sequencing. Whilst MED12 exon 2 mutations were significantly more frequent in tumors with fibroadenoma-like areas (71 vs. 11%), an enrichment in genetic alterations targeting bona fide cancer genes was found in those without fibroadenoma-like areas, in particular in EGFR mutations and amplifications (78 vs. 14%). No significant difference in the frequency of TERT genetic alterations was observed (71% in cases with fibroadenoma-like areas vs 56% in those without fibroadenoma-like areas). Our data suggest that the development of phyllodes tumors might follow two different evolutionary pathways: a MED12 -mutant pathway that involves the progression from a fibroadenoma to a malignant phyllodes tumor; and a MED12 -wild-type pathway, where malignant phyllodes tumors arise de novo through the acquisition of genetic alterations targeting cancer genes. Additional studies are warranted to confirm our observations and define whether the outcome differs between both pathways.

  9. RIG-I-like receptor-induced IRF3 mediated pathway of apoptosis (RIPA: a new antiviral pathway

    Directory of Open Access Journals (Sweden)

    Saurabh Chattopadhyay

    2016-11-01

    Full Text Available Abstract The innate immune response is the first line of host defense to eliminate viral infection. Pattern recognition receptors in the cytosol, such as RIG-I-like receptors (RLR and Nod-like receptors (NLR, and membrane bound Toll like receptors (TLR detect viral infection and initiate transcription of a cohort of antiviral genes, including interferon (IFN and interferon stimulated genes (ISGs, which ultimately block viral replication. Another mechanism to reduce viral spread is through RIPA, the RLR-induced IRF3-mediated pathway of apoptosis, which causes infected cells to undergo premature death. The transcription factor IRF3 can mediate cellular antiviral responses by both inducing antiviral genes and triggering apoptosis through the activation of RIPA. The mechanism of IRF3 activation in RIPA is distinct from that of transcriptional activation; it requires linear polyubiquitination of specific lysine residues of IRF3. Using RIPA-active, but transcriptionally inactive, IRF3 mutants, it was shown that RIPA can prevent viral replication and pathogenesis in mice.

  10. Ultrasound Microbubble Treatment Enhances Clathrin-Mediated Endocytosis and Fluid-Phase Uptake through Distinct Mechanisms.

    Directory of Open Access Journals (Sweden)

    Farnaz Fekri

    Full Text Available Drug delivery to tumors is limited by several factors, including drug permeability of the target cell plasma membrane. Ultrasound in combination with microbubbles (USMB is a promising strategy to overcome these limitations. USMB treatment elicits enhanced cellular uptake of materials such as drugs, in part as a result of sheer stress and formation of transient membrane pores. Pores formed upon USMB treatment are rapidly resealed, suggesting that other processes such as enhanced endocytosis may contribute to the enhanced material uptake by cells upon USMB treatment. How USMB regulates endocytic processes remains incompletely understood. Cells constitutively utilize several distinct mechanisms of endocytosis, including clathrin-mediated endocytosis (CME for the internalization of receptor-bound macromolecules such as Transferrin Receptor (TfR, and distinct mechanism(s that mediate the majority of fluid-phase endocytosis. Tracking the abundance of TfR on the cell surface and the internalization of its ligand transferrin revealed that USMB acutely enhances the rate of CME. Total internal reflection fluorescence microscopy experiments revealed that USMB treatment altered the assembly of clathrin-coated pits, the basic structural units of CME. In addition, the rate of fluid-phase endocytosis was enhanced, but with delayed onset upon USMB treatment relative to the enhancement of CME, suggesting that the two processes are distinctly regulated by USMB. Indeed, vacuolin-1 or desipramine treatment prevented the enhancement of CME but not of fluid phase endocytosis upon USMB, suggesting that lysosome exocytosis and acid sphingomyelinase, respectively, are required for the regulation of CME but not fluid phase endocytosis upon USMB treatment. These results indicate that USMB enhances both CME and fluid phase endocytosis through distinct signaling mechanisms, and suggest that strategies for potentiating the enhancement of endocytosis upon USMB treatment may

  11. Pediatric-type nodal follicular lymphoma: a biologically distinct lymphoma with frequent MAPK pathway mutations.

    Science.gov (United States)

    Louissaint, Abner; Schafernak, Kristian T; Geyer, Julia T; Kovach, Alexandra E; Ghandi, Mahmoud; Gratzinger, Dita; Roth, Christine G; Paxton, Christian N; Kim, Sunhee; Namgyal, Chungdak; Morin, Ryan; Morgan, Elizabeth A; Neuberg, Donna S; South, Sarah T; Harris, Marian H; Hasserjian, Robert P; Hochberg, Ephraim P; Garraway, Levi A; Harris, Nancy Lee; Weinstock, David M

    2016-08-25

    Pediatric-type nodal follicular lymphoma (PTNFL) is a variant of follicular lymphoma (FL) characterized by limited-stage presentation and invariably benign behavior despite often high-grade histological appearance. It is important to distinguish PTNFL from typical FL in order to avoid unnecessary treatment; however, this distinction relies solely on clinical and pathological criteria, which may be variably applied. To define the genetic landscape of PTNFL, we performed copy number analysis and exome and/or targeted sequencing of 26 PTNFLs (16 pediatric and 10 adult). The most commonly mutated gene in PTNFL was MAP2K1, encoding MEK1, with a mutation frequency of 43%. All MAP2K1 mutations were activating missense mutations localized to exons 2 and 3, which encode negative regulatory and catalytic domains, respectively. Missense mutations in MAPK1 (2/22) and RRAS (1/22) were identified in cases that lacked MAP2K1 mutations. The second most commonly mutated gene in PTNFL was TNFRSF14, with a mutation frequency of 29%, similar to that seen in limited-stage typical FL (P = .35). PTNFL was otherwise genomically bland and specifically lacked recurrent mutations in epigenetic modifiers (eg, CREBBP, KMT2D). Copy number aberrations affected a mean of only 0.5% of PTNFL genomes, compared with 10% of limited-stage typical FL genomes (P < .02). Importantly, the mutational profiles of PTNFLs in children and adults were highly similar. Together, these findings define PTNFL as a biologically and clinically distinct indolent lymphoma of children and adults characterized by a high prevalence of MAPK pathway mutations and a near absence of mutations in epigenetic modifiers. © 2016 by The American Society of Hematology.

  12. ANGUSTIFOLIA mediates one of the multiple SCRAMBLED signaling pathways regulating cell growth pattern in Arabidopsis thaliana.

    Science.gov (United States)

    Kwak, Su-Hwan; Song, Sang-Kee; Lee, Myeong Min; Schiefelbein, John

    2015-09-25

    In Arabidopsis thaliana, an atypical leucine-rich repeat receptor-like kinase, SCRAMBLED (SCM), is required for multiple developmental processes including root epidermal cell fate determination, silique dehiscence, inflorescence growth, ovule morphogenesis, and tissue morphology. Previous work suggested that SCM regulates these multiple pathways using distinct mechanisms via interactions with specific downstream factors. ANGUSTIFOLIA (AN) is known to regulate cell and tissue morphogenesis by influencing cortical microtubule arrangement, and recently, the AN protein was reported to interact with the SCM protein. Therefore, we examined whether AN might be responsible for mediating some of the SCM-dependent phenotypes. We discovered that both scm and an mutant lines cause an abnormal spiral or twisting growth of roots, but only the scm mutant affected root epidermal patterning. The siliques of the an and scm mutants also exhibited spiral growth, as previously reported, but only the scm mutant altered silique dehiscence. Interestingly, we discovered that the spiral growth of roots and siliques of the scm mutant is rescued by a truncated SCM protein that lacks its kinase domain, and that a juxtamembrane domain of SCM was sufficient for AN binding in the yeast two-hybrid analysis. These results suggest that the AN protein is one of the critical downstream factors of SCM pathways specifically responsible for mediating its effects on cell/tissue morphogenesis through cortical microtubule arrangement. Copyright © 2015 Elsevier Inc. All rights reserved.

  13. Dendritic cells and hepatocytes use distinct pathways to process protective antigen from plasmodium in vivo.

    Directory of Open Access Journals (Sweden)

    Ian A Cockburn

    2011-03-01

    Full Text Available Malaria-protective CD8+ T cells specific for the circumsporozoite (CS protein are primed by dendritic cells (DCs after sporozoite injection by infected mosquitoes. The primed cells then eliminate parasite liver stages after recognizing the CS epitopes presented by hepatocytes. To define the in vivo processing of CS by DCs and hepatocytes, we generated parasites carrying a mutant CS protein containing the H-2K(b epitope SIINFEKL, and evaluated the T cell response using transgenic and mutant mice. We determined that in both DCs and hepatocytes CS epitopes must reach the cytosol and use the TAP transporters to access the ER. Furthermore, we used endosomal mutant (3d and cytochrome c treated mice to address the role of cross-presentation in the priming and effector phases of the T cell response. We determined that in DCs, CS is cross-presented via endosomes while, conversely, in hepatocytes protein must be secreted directly into the cytosol. This suggests that the main targets of protective CD8+ T cells are parasite proteins exported to the hepatocyte cytosol. Surprisingly, however, secretion of the CS protein into hepatocytes was not dependent upon parasite-export (Pexel/VTS motifs in this protein. Together, these results indicate that the presentation of epitopes to CD8+ T cells follows distinct pathways in DCs when the immune response is induced and in hepatocytes during the effector phase.

  14. Separate transcriptionally regulated pathways specify distinct classes of sister dendrites in a nociceptive neuron.

    Science.gov (United States)

    O'Brien, Barbara M J; Palumbos, Sierra D; Novakovic, Michaela; Shang, Xueying; Sundararajan, Lakshmi; Miller, David M

    2017-12-15

    The dendritic processes of nociceptive neurons transduce external signals into neurochemical cues that alert the organism to potentially damaging stimuli. The receptive field for each sensory neuron is defined by its dendritic arbor, but the mechanisms that shape dendritic architecture are incompletely understood. Using the model nociceptor, the PVD neuron in C. elegans, we determined that two types of PVD lateral branches project along the dorsal/ventral axis to generate the PVD dendritic arbor: (1) Pioneer dendrites that adhere to the epidermis, and (2) Commissural dendrites that fasciculate with circumferential motor neuron processes. Previous reports have shown that the LIM homeodomain transcription factor MEC-3 is required for all higher order PVD branching and that one of its targets, the claudin-like membrane protein HPO-30, preferentially promotes outgrowth of pioneer branches. Here, we show that another MEC-3 target, the conserved TFIIA-like zinc finger transcription factor EGL-46, adopts the alternative role of specifying commissural dendrites. The known EGL-46 binding partner, the TEAD transcription factor EGL-44, is also required for PVD commissural branch outgrowth. Double mutants of hpo-30 and egl-44 show strong enhancement of the lateral branching defect with decreased numbers of both pioneer and commissural dendrites. Thus, HPO-30/Claudin and EGL-46/EGL-44 function downstream of MEC-3 and in parallel acting pathways to direct outgrowth of two distinct classes of PVD dendritic branches. Copyright © 2017 Elsevier Inc. All rights reserved.

  15. Possible distinct molecular carcinogenic pathways for bladder cancer in Ukraine, before and after the Chernobyl disaster.

    Science.gov (United States)

    Morimura, Keiichirou; Romanenko, Alina; Min, Wei; Salim, Elsayed I; Kinoshita, Anna; Wanibuchi, Hideki; Vozianov, Alexander; Fukushima, Shoji

    2004-04-01

    After the Chernobyl accident in 1986, the incidence of urinary bladder cancers in the Ukraine increased gradually from 26.2 to 43.3 per 100,000 people between 1986 and 2001. In the areas of low level but persistent cesium-137 (137Cs) radio-contamination, a unique atypical radiation-related urinary bladder cystitis named 'Chernobyl cystitis', a possible pre-neoplastic condition in humans, has been detected. We have previously documented high incidences of bladder lesions, including severe dysplasias and/or carcinoma in situ, in association with this cystitis and correlating with oxidative DNA damage. To further investigate the molecular mechanisms underlying bladder carcinogenesis with this specific etiology, mutation analysis of p53 gene (exon 5-8) was performed for 11 and 18 paraffin-embedded bladder cancers in Ukrainians, respectively collected before and after the Chernobyl disaster. DNAs were extracted and subjected to nested PCR-single-strand conformational polymorphism analysis followed by direct DNA sequencing, as well as p53 immunohistochemistry (IHC). The incidences of p53 gene mutation were 54.5 and 16.7% for before and after the Chernobyl disaster, respectively, the difference being statistically significant. Also a tendency for higher p53 IHC score was apparent in the earlier group of lesions. No significant difference was noted for the proportions of historical types. These results point to possible distinct molecular carcinogenic pathways of bladder cancer formation, before and after the Chernobyl disaster, on the basis of variation in p53 gene alteration.

  16. Substrate-Trapped Interactors of PHD3 and FIH Cluster in Distinct Signaling Pathways

    Directory of Open Access Journals (Sweden)

    Javier Rodriguez

    2016-03-01

    Full Text Available Amino acid hydroxylation is a post-translational modification that regulates intra- and inter-molecular protein-protein interactions. The modifications are regulated by a family of 2-oxoglutarate- (2OG dependent enzymes and, although the biochemistry is well understood, until now only a few substrates have been described for these enzymes. Using quantitative interaction proteomics, we screened for substrates of the proline hydroxylase PHD3 and the asparagine hydroxylase FIH, which regulate the HIF-mediated hypoxic response. We were able to identify hundreds of potential substrates. Enrichment analysis revealed that the potential substrates of both hydroxylases cluster in the same pathways but frequently modify different nodes of signaling networks. We confirm that two proteins identified in our screen, MAPK6 (Erk3 and RIPK4, are indeed hydroxylated in a FIH- or PHD3-dependent mechanism. We further determined that FIH-dependent hydroxylation regulates RIPK4-dependent Wnt signaling, and that PHD3-dependent hydroxylation of MAPK6 protects the protein from proteasomal degradation.

  17. Experimentally-derived fibroblast gene signatures identify molecular pathways associated with distinct subsets of systemic sclerosis patients in three independent cohorts.

    Directory of Open Access Journals (Sweden)

    Michael E Johnson

    Full Text Available Genome-wide expression profiling in systemic sclerosis (SSc has identified four 'intrinsic' subsets of disease (fibroproliferative, inflammatory, limited, and normal-like, each of which shows deregulation of distinct signaling pathways; however, the full set of pathways contributing to this differential gene expression has not been fully elucidated. Here we examine experimentally derived gene expression signatures in dermal fibroblasts for thirteen different signaling pathways implicated in SSc pathogenesis. These data show distinct and overlapping sets of genes induced by each pathway, allowing for a better understanding of the molecular relationship between profibrotic and immune signaling networks. Pathway-specific gene signatures were analyzed across a compendium of microarray datasets consisting of skin biopsies from three independent cohorts representing 80 SSc patients, 4 morphea, and 26 controls. IFNα signaling showed a strong association with early disease, while TGFβ signaling spanned the fibroproliferative and inflammatory subsets, was associated with worse MRSS, and was higher in lesional than non-lesional skin. The fibroproliferative subset was most strongly associated with PDGF signaling, while the inflammatory subset demonstrated strong activation of innate immune pathways including TLR signaling upstream of NF-κB. The limited and normal-like subsets did not show associations with fibrotic and inflammatory mediators such as TGFβ and TNFα. The normal-like subset showed high expression of genes associated with lipid signaling, which was absent in the inflammatory and limited subsets. Together, these data suggest a model by which IFNα is involved in early disease pathology, and disease severity is associated with active TGFβ signaling.

  18. Triglyceride-mediated pathways and coronary disease: collaborative analysis of 101 studies

    DEFF Research Database (Denmark)

    Sarwar, Nadeem; Sandhu, Manjinder S; Ricketts, Sally L

    2010-01-01

    Whether triglyceride-mediated pathways are causally relevant to coronary heart disease is uncertain. We studied a genetic variant that regulates triglyceride concentration to help judge likelihood of causality.......Whether triglyceride-mediated pathways are causally relevant to coronary heart disease is uncertain. We studied a genetic variant that regulates triglyceride concentration to help judge likelihood of causality....

  19. Yarrowia lipolytica vesicle-mediated protein transport pathways

    Directory of Open Access Journals (Sweden)

    Beckerich Jean-Marie

    2007-11-01

    Full Text Available Abstract Background Protein secretion is a universal cellular process involving vesicles which bud and fuse between organelles to bring proteins to their final destination. Vesicle budding is mediated by protein coats; vesicle targeting and fusion depend on Rab GTPase, tethering factors and SNARE complexes. The Génolevures II sequencing project made available entire genome sequences of four hemiascomycetous yeasts, Yarrowia lipolytica, Debaryomyces hansenii, Kluyveromyces lactis and Candida glabrata. Y. lipolytica is a dimorphic yeast and has good capacities to secrete proteins. The translocation of nascent protein through the endoplasmic reticulum membrane was well studied in Y. lipolytica and is largely co-translational as in the mammalian protein secretion pathway. Results We identified S. cerevisiae proteins involved in vesicular secretion and these protein sequences were used for the BLAST searches against Génolevures protein database (Y. lipolytica, C. glabrata, K. lactis and D. hansenii. These proteins are well conserved between these yeasts and Saccharomyces cerevisiae. We note several specificities of Y. lipolytica which may be related to its good protein secretion capacities and to its dimorphic aspect. An expansion of the Y. lipolytica Rab protein family was observed with autoBLAST and the Rab2- and Rab4-related members were identified with BLAST against NCBI protein database. An expansion of this family is also found in filamentous fungi and may reflect the greater complexity of the Y. lipolytica secretion pathway. The Rab4p-related protein may play a role in membrane recycling as rab4 deleted strain shows a modification of colony morphology, dimorphic transition and permeability. Similarly, we find three copies of the gene (SSO encoding the plasma membrane SNARE protein. Quantification of the percentages of proteins with the greatest homology between S. cerevisiae, Y. lipolytica and animal homologues involved in vesicular

  20. Distinct aspects of frontal lobe structure mediate age-related differences in fluid intelligence and multitasking

    Science.gov (United States)

    Kievit, Rogier A.; Davis, Simon W.; Mitchell, Daniel J.; Taylor, Jason R.; Duncan, John; Tyler, Lorraine K.; Brayne, Carol; Bullmore, Ed; Calder, Andrew; Cusack, Rhodri; Dalgleish, Tim; Matthews, Fiona; Marslen-Wilson, William; Rowe, James; Shafto, Meredith; Campbell, Karen; Cheung, Teresa; Geerligs, Linda; McCarrey, Anna; Tsvetanov, Kamen; Williams, Nitin; Bates, Lauren; Emery, Tina; Erzinçlioglu, Sharon; Gadie, Andrew; Gerbase, Sofia; Georgieva, Stanimira; Hanley, Claire; Parkin, Beth; Troy, David; Allen, Jodie; Amery, Gillian; Amunts, Liana; Barcroft, Anne; Castle, Amanda; Dias, Cheryl; Dowrick, Jonathan; Fair, Melissa; Fisher, Hayley; Goulding, Anna; Grewal, Adarsh; Hale, Geoff; Hilton, Andrew; Johnson, Frances; Johnston, Patricia; Kavanagh-Williamson, Thea; Kwasniewska, Magdalena; McMinn, Alison; Norman, Kim; Penrose, Jessica; Roby, Fiona; Rowland, Diane; Sargeant, John; Squire, Maggie; Stevens, Beth; Stoddart, Aldabra; Stone, Cheryl; Thompson, Tracy; Yazlik, Ozlem; Barnes, Dan; Dixon, Marie; Hillman, Jaya; Mitchell, Joanne; Villis, Laura; Henson, Richard N.A.

    2014-01-01

    Ageing is characterized by declines on a variety of cognitive measures. These declines are often attributed to a general, unitary underlying cause, such as a reduction in executive function owing to atrophy of the prefrontal cortex. However, age-related changes are likely multifactorial, and the relationship between neural changes and cognitive measures is not well-understood. Here we address this in a large (N=567), population-based sample drawn from the Cambridge Centre for Ageing and Neuroscience (Cam-CAN) data. We relate fluid intelligence and multitasking to multiple brain measures, including grey matter in various prefrontal regions and white matter integrity connecting those regions. We show that multitasking and fluid intelligence are separable cognitive abilities, with differential sensitivities to age, which are mediated by distinct neural subsystems that show different prediction in older versus younger individuals. These results suggest that prefrontal ageing is a manifold process demanding multifaceted models of neurocognitive ageing. PMID:25519467

  1. Distinct mechanisms of loss of IFN-gamma mediated HLA class I inducibility in two melanoma cell lines

    International Nuclear Information System (INIS)

    Rodríguez, Teresa; Méndez, Rosa; Del Campo, Ana; Jiménez, Pilar; Aptsiauri, Natalia; Garrido, Federico; Ruiz-Cabello, Francisco

    2007-01-01

    The inability of cancer cells to present antigen on the cell surface via MHC class I molecules is one of the mechanisms by which tumor cells evade anti-tumor immunity. Alterations of Jak-STAT components of interferon (IFN)-mediated signaling can contribute to the mechanism of cell resistance to IFN, leading to lack of MHC class I inducibility. Hence, the identification of IFN-γ-resistant tumors may have prognostic and/or therapeutic relevance. In the present study, we investigated a mechanism of MHC class I inducibility in response to IFN-γ treatment in human melanoma cell lines. Basal and IFN-induced expression of HLA class I antigens was analyzed by means of indirect immunofluorescence flow cytometry, Western Blot, RT-PCR, and quantitative real-time RT-PCR (TaqMan ® Gene Expression Assays). In demethylation studies cells were cultured with 5-aza-2'-deoxycytidine. Electrophoretic Mobility Shift Assay (EMSA) was used to assay whether IRF-1 promoter binding activity is induced in IFN-γ-treated cells. Altered IFN-γ mediated HLA-class I induction was observed in two melanoma cells lines (ESTDAB-004 and ESTDAB-159) out of 57 studied, while treatment of these two cell lines with IFN-α led to normal induction of HLA class I antigen expression. Examination of STAT-1 in ESTDAB-004 after IFN-γ treatment demonstrated that the STAT-1 protein was expressed but not phosphorylated. Interestingly, IFN-α treatment induced normal STAT-1 phosphorylation and HLA class I expression. In contrast, the absence of response to IFN-γ in ESTDAB-159 was found to be associated with alterations in downstream components of the IFN-γ signaling pathway. We observed two distinct mechanisms of loss of IFN-γ inducibility of HLA class I antigens in two melanoma cell lines. Our findings suggest that loss of HLA class I induction in ESTDAB-004 cells results from a defect in the earliest steps of the IFN-γ signaling pathway due to absence of STAT-1 tyrosine-phosphorylation, while absence

  2. The effect of chronic seaweed subsidies on herbivory: plant-mediated fertilization pathway overshadows lizard-mediated predator pathways.

    Science.gov (United States)

    Piovia-Scott, Jonah; Spiller, David A; Takimoto, Gaku; Yang, Louie H; Wright, Amber N; Schoener, Thomas W

    2013-08-01

    Flows of energy and materials link ecosystems worldwide and have important consequences for the structure of ecological communities. While these resource subsidies typically enter recipient food webs through multiple channels, most previous studies focussed on a single pathway of resource input. We used path analysis to evaluate multiple pathways connecting chronic marine resource inputs (in the form of seaweed deposits) and herbivory in a shoreline terrestrial ecosystem. We found statistical support for a fertilization effect (seaweed increased foliar nitrogen content, leading to greater herbivory) and a lizard numerical response effect (seaweed increased lizard densities, leading to reduced herbivory), but not for a lizard diet-shift effect (seaweed increased the proportion of marine-derived prey in lizard diets, but lizard diet was not strongly associated with herbivory). Greater seaweed abundance was associated with greater herbivory, and the fertilization effect was larger than the combined lizard effects. Thus, the bottom-up, plant-mediated effect of fertilization on herbivory overshadowed the top-down effects of lizard predators. These results, from unmanipulated shoreline plots with persistent differences in chronic seaweed deposition, differ from those of a previous experimental study of the short-term effects of a pulse of seaweed deposition: while the increase in herbivory in response to chronic seaweed deposition was due to the fertilization effect, the short-term increase in herbivory in response to a pulse of seaweed deposition was due to the lizard diet-shift effect. This contrast highlights the importance of the temporal pattern of resource inputs in determining the mechanism of community response to resource subsidies.

  3. Subtypes of the Type II Pit Pattern Reflect Distinct Molecular Subclasses in the Serrated Neoplastic Pathway.

    Science.gov (United States)

    Aoki, Hironori; Yamamoto, Eiichiro; Yamano, Hiro-O; Sugai, Tamotsu; Kimura, Tomoaki; Tanaka, Yoshihito; Matsushita, Hiro-O; Yoshikawa, Kenjiro; Takagi, Ryo; Harada, Eiji; Nakaoka, Michiko; Yoshida, Yuko; Harada, Taku; Sudo, Gota; Eizuka, Makoto; Yorozu, Akira; Kitajima, Hiroshi; Niinuma, Takeshi; Kai, Masahiro; Nojima, Masanori; Suzuki, Hiromu; Nakase, Hiroshi

    2018-03-15

    Colorectal serrated lesions (SLs) are important premalignant lesions whose clinical and biological features are not fully understood. We aimed to establish accurate colonoscopic diagnosis and treatment of SLs through evaluation of associations among the morphological, pathological, and molecular characteristics of SLs. A total of 388 premalignant and 18 malignant colorectal lesions were studied. Using magnifying colonoscopy, microsurface structures were assessed based on Kudo's pit pattern classification system, and the Type II pit pattern was subcategorized into classical Type II, Type II-Open (Type II-O) and Type II-Long (Type II-L). BRAF/KRAS mutations and DNA methylation of CpG island methylator phenotype (CIMP) markers (MINT1, - 2, - 12, - 31, p16, and MLH1) were analyzed through pyrosequencing. Type II-O was tightly associated with sessile serrated adenoma/polyps (SSA/Ps) with BRAF mutation and CIMP-high. Most lesions with simple Type II or Type II-L were hyperplastic polyps, while mixtures of Type II or Type II-L plus more advanced pit patterns (III/IV) were characteristic of traditional serrated adenomas (TSAs). Type II-positive TSAs frequently exhibited BRAF mutation and CIMP-low, while Type II-L-positive TSAs were tightly associated with KRAS mutation and CIMP-low. Analysis of lesions containing both premalignant and cancerous components suggested Type II-L-positive TSAs may develop into KRAS-mutated/CIMP-low/microsatellite stable cancers, while Type II-O-positive SSA/Ps develop into BRAF-mutated/CIMP-high/microsatellite unstable cancers. These results suggest that Type II subtypes reflect distinct molecular subclasses in the serrated neoplasia pathway and that they could be useful hallmarks for identifying SLs at high risk of developing into CRC.

  4. Distinct Internalization Pathways of Human Amylin Monomers and Its Cytotoxic Oligomers in Pancreatic Cells

    Science.gov (United States)

    Trikha, Saurabh; Jeremic, Aleksandar M.

    2013-01-01

    Toxic human amylin oligomers and aggregates are implicated in the pathogenesis of type 2 diabetes mellitus (TTDM). Although recent studies have shown that pancreatic cells can recycle amylin monomers and toxic oligomers, the exact uptake mechanism and trafficking routes of these molecular forms and their significance for amylin toxicity are yet to be determined. Using pancreatic rat insulinoma (RIN-m5F) beta (β)-cells and human islets as model systems we show that monomers and oligomers cross the plasma membrane (PM) through both endocytotic and non-endocytotic (translocation) mechanisms, the predominance of which is dependent on amylin concentrations and incubation times. At low (≤100 nM) concentrations, internalization of amylin monomers in pancreatic cells is completely blocked by the selective amylin-receptor (AM-R) antagonist, AC-187, indicating an AM-R dependent mechanism. In contrast at cytotoxic (µM) concentrations monomers initially (1 hour) enter pancreatic cells by two distinct mechanisms: translocation and macropinocytosis. However, during the late stage (24 hours) monomers internalize by a clathrin-dependent but AM-R and macropinocytotic independent pathway. Like monomers a small fraction of the oligomers initially enter cells by a non-endocytotic mechanism. In contrast a majority of the oligomers at both early (1 hour) and late times (24 hours) traffic with a fluid-phase marker, dextran, to the same endocytotic compartments, the uptake of which is blocked by potent macropinocytotic inhibitors. This led to a significant increase in extra-cellular PM accumulation, in turn potentiating amylin toxicity in pancreatic cells. Our studies suggest that macropinocytosis is a major but not the only clearance mechanism for both amylin’s molecular forms, thereby serving a cyto-protective role in these cells. PMID:24019897

  5. Orexins control intestinal glucose transport by distinct neuronal, endocrine, and direct epithelial pathways.

    Science.gov (United States)

    Ducroc, Robert; Voisin, Thierry; El Firar, Aadil; Laburthe, Marc

    2007-10-01

    Orexins are neuropeptides involved in energy homeostasis. We investigated the effect of orexin A (OxA) and orexin B (OxB) on intestinal glucose transport in the rat. Injection of orexins led to a decrease in the blood glucose level in oral glucose tolerance tests (OGTTs). Effects of orexins on glucose entry were analyzed in Ussing chambers using the Na(+)-dependent increase in short-circuit current (Isc) to quantify jejunal glucose transport. The rapid and marked increase in Isc induced by luminal glucose was inhibited by 10 nmol/l OxA or OxB (53 and 59%, respectively). Response curves to OxA and OxB were not significantly different with half-maximal inhibitory concentrations at 0.9 and 0.4 nmol/l, respectively. On the one hand, OxA-induced inhibition of Isc was reduced by the neuronal blocker tetrodotoxin (TTX) and by a cholecystokinin (CCK) 2R antagonist, indicating involvement of neuronal and endocrine CCK-releasing cells. The OX(1)R antagonist SB334867 had no effect on OxA-induced inhibition, which is likely to occur via a neuronal and/or endocrine OX(2)R. On the other hand, SB334867 induced a significant right shift of the concentration-effect curve for OxB. This OxB-preferring OX(1)R pathway was not sensitive to TTX or to CCKR antagonists, suggesting that OxB may act directly on enterocytic OX(1)R. These distinct effects of OxA and OxB are consistent with the expression of OX(1)R and OX(2)R mRNA in the epithelial and nonepithelial tissues, respectively. Our data delineate a new function for orexins as inhibitors of intestinal glucose absorption and provide a new basis for orexin-induced short-term control of energy homeostasis.

  6. Distinct parietal sites mediate the influences of mood, arousal, and their interaction on human recognition memory.

    Science.gov (United States)

    Greene, Ciara M; Flannery, Oliver; Soto, David

    2014-12-01

    The two dimensions of emotion, mood valence and arousal, have independent effects on recognition memory. At present, however, it is not clear how those effects are reflected in the human brain. Previous research in this area has generally dealt with memory for emotionally valenced or arousing stimuli, but the manner in which interacting mood and arousal states modulate responses in memory substrates remains poorly understood. We investigated memory for emotionally neutral items while independently manipulating mood valence and arousal state by means of music exposure. Four emotional conditions were created: positive mood/high arousal, positive mood/low arousal, negative mood/high arousal, and negative mood/low arousal. We observed distinct effects of mood valence and arousal in parietal substrates of recognition memory. Positive mood increased activity in ventral posterior parietal cortex (PPC) and orbitofrontal cortex, whereas arousal condition modulated activity in dorsal PPC and the posterior cingulate. An interaction between valence and arousal was observed in left ventral PPC, notably in a parietal area distinct from the those identified for the main effects, with a stronger effect of mood on recognition memory responses here under conditions of relative high versus low arousal. We interpreted the PPC activations in terms of the attention-to-memory hypothesis: Increased arousal may lead to increased top-down control of memory, and hence dorsal PPC activation, whereas positive mood valence may result in increased activity in ventral PPC regions associated with bottom-up attention to memory. These findings indicate that distinct parietal sites mediate the influences of mood, arousal, and their interplay during recognition memory.

  7. The Striatal Balancing Act in Drug Addiction: Distinct Roles of Direct and Indirect Pathway Medium Spiny Neurons

    Directory of Open Access Journals (Sweden)

    Mary Kay eLobo

    2011-07-01

    Full Text Available The striatum plays a key role in mediating the acute and chronic effects of addictive drugs, with drugs of abuse causing long-lasting molecular and cellular alterations in both dorsal striatum and nucleus accumbens (ventral striatum. Despite the wealth of research on the biological actions of abused drugs in striatum, until recently, the distinct roles of the striatum’s two major subtypes of medium spiny neuron (MSN in drug addiction remained elusive. Recent advances in cell-type specific technologies, including fluorescent reporter mice, transgenic or knockout mice, and viral-mediated gene transfer, have advanced the field toward a more comprehensive understanding of the two MSN subtypes in the long-term actions of drugs of abuse. Here we review progress in defining the distinct molecular and functional contributions of the two MSN subtypes in mediating addiction.

  8. HDAC4 preserves skeletal muscle structure following long-term denervation by mediating distinct cellular responses.

    Science.gov (United States)

    Pigna, Eva; Renzini, Alessandra; Greco, Emanuela; Simonazzi, Elena; Fulle, Stefania; Mancinelli, Rosa; Moresi, Viviana; Adamo, Sergio

    2018-02-24

    Denervation triggers numerous molecular responses in skeletal muscle, including the activation of catabolic pathways and oxidative stress, leading to progressive muscle atrophy. Histone deacetylase 4 (HDAC4) mediates skeletal muscle response to denervation, suggesting the use of HDAC inhibitors as a therapeutic approach to neurogenic muscle atrophy. However, the effects of HDAC4 inhibition in skeletal muscle in response to long-term denervation have not been described yet. To further study HDAC4 functions in response to denervation, we analyzed mutant mice in which HDAC4 is specifically deleted in skeletal muscle. After an initial phase of resistance to neurogenic muscle atrophy, skeletal muscle with a deletion of HDAC4 lost structural integrity after 4 weeks of denervation. Deletion of HDAC4 impaired the activation of the ubiquitin-proteasome system, delayed the autophagic response, and dampened the OS response in skeletal muscle. Inhibition of the ubiquitin-proteasome system or the autophagic response, if on the one hand, conferred resistance to neurogenic muscle atrophy; on the other hand, induced loss of muscle integrity and inflammation in mice lacking HDAC4 in skeletal muscle. Moreover, treatment with the antioxidant drug Trolox prevented loss of muscle integrity and inflammation in in mice lacking HDAC4 in skeletal muscle, despite the resistance to neurogenic muscle atrophy. These results reveal new functions of HDAC4 in mediating skeletal muscle response to denervation and lead us to propose the combined use of HDAC inhibitors and antioxidant drugs to treat neurogenic muscle atrophy.

  9. Structural white-matter connections mediating distinct behavioral components of spatial neglect in right brain-damaged patients.

    Science.gov (United States)

    Vaessen, Maarten J; Saj, Arnaud; Lovblad, Karl-Olof; Gschwind, Markus; Vuilleumier, Patrik

    2016-04-01

    Spatial neglect is a neuropsychological syndrome in which patients fail to perceive and orient to stimuli located in the space contralateral to the lesioned hemisphere. It is characterized by a wide heterogeneity in clinical symptoms which can be grouped into distinct behavioral components correlating with different lesion sites. Moreover, damage to white-matter (WM) fiber tracts has been suggested to disconnect brain networks that mediate different functions associated with spatial cognition and attention. However, it remains unclear what WM pathways are associated with functionally dissociable neglect components. In this study we examined nine patients with a focal right hemisphere stroke using a series of neuropsychological tests and diffusion tensor imaging (DTI) in order to disentangle the role of specific WM pathways in neglect symptoms. First, following previous work, the behavioral test scores of patients were factorized into three independent components reflecting perceptual, exploratory, and object-centered deficits in spatial awareness. We then examined the structural neural substrates of these components by correlating indices of WM integrity (fractional anisotropy) with the severity of deficits along each profile. Several locations in the right parietal and frontal WM correlated with neuropsychological scores. Fiber tracts projecting from these locations indicated that posterior parts of the superior longitudinal fasciculus (SLF), as well as nearby callosal fibers connecting ipsilateral and contralateral parietal areas, were associated with perceptual spatial deficits, whereas more anterior parts of SLF and inferior fronto-occipital fasciculus (IFOF) were predominantly associated with object-centered deficits. In addition, connections between frontal areas and superior colliculus were found to be associated with the exploratory deficits. Our results provide novel support to the view that neglect may result from disconnection lesions in distributed

  10. Mutant Allele-Specific Uncoupling of PENETRATION3 Functions Reveals Engagement of the ATP-Binding Cassette Transporter in Distinct Tryptophan Metabolic Pathways1[OPEN

    Science.gov (United States)

    Lu, Xunli; Dittgen, Jan; Piślewska-Bednarek, Mariola; Molina, Antonio; Schneider, Bernd; Doubský, Jan; Schneeberger, Korbinian; Schulze-Lefert, Paul

    2015-01-01

    Arabidopsis (Arabidopsis thaliana) PENETRATION (PEN) genes quantitatively contribute to the execution of different forms of plant immunity upon challenge with diverse leaf pathogens. PEN3 encodes a plasma membrane-resident pleiotropic drug resistance-type ATP-binding cassette transporter and is thought to act in a pathogen-inducible and PEN2 myrosinase-dependent metabolic pathway in extracellular defense. This metabolic pathway directs the intracellular biosynthesis and activation of tryptophan-derived indole glucosinolates for subsequent PEN3-mediated efflux across the plasma membrane at pathogen contact sites. However, PEN3 also functions in abiotic stress responses to cadmium and indole-3-butyric acid (IBA)-mediated auxin homeostasis in roots, raising the possibility that PEN3 exports multiple functionally unrelated substrates. Here, we describe the isolation of a pen3 allele, designated pen3-5, that encodes a dysfunctional protein that accumulates in planta like wild-type PEN3. The specific mutation in pen3-5 uncouples PEN3 functions in IBA-stimulated root growth modulation, callose deposition induced with a conserved peptide epitope of bacterial flagellin (flg22), and pathogen-inducible salicylic acid accumulation from PEN3 activity in extracellular defense, indicating the engagement of multiple PEN3 substrates in different PEN3-dependent biological processes. We identified 4-O-β-d-glucosyl-indol-3-yl formamide (4OGlcI3F) as a pathogen-inducible, tryptophan-derived compound that overaccumulates in pen3 leaf tissue and has biosynthesis that is dependent on an intact PEN2 metabolic pathway. We propose that a precursor of 4OGlcI3F is the PEN3 substrate in extracellular pathogen defense. These precursors, the shared indole core present in IBA and 4OGlcI3F, and allele-specific uncoupling of a subset of PEN3 functions suggest that PEN3 transports distinct indole-type metabolites in distinct biological processes. PMID:26023163

  11. BAD-mediated apoptotic pathway is associated with human cancer development.

    Science.gov (United States)

    Stickles, Xiaomang B; Marchion, Douglas C; Bicaku, Elona; Al Sawah, Entidhar; Abbasi, Forough; Xiong, Yin; Bou Zgheib, Nadim; Boac, Bernadette M; Orr, Brian C; Judson, Patricia L; Berry, Amy; Hakam, Ardeshir; Wenham, Robert M; Apte, Sachin M; Berglund, Anders E; Lancaster, Johnathan M

    2015-04-01

    The malignant transformation of normal cells is caused in part by aberrant gene expression disrupting the regulation of cell proliferation, apoptosis, senescence and DNA repair. Evidence suggests that the Bcl-2 antagonist of cell death (BAD)-mediated apoptotic pathway influences cancer chemoresistance. In the present study, we explored the role of the BAD-mediated apoptotic pathway in the development and progression of cancer. Using principal component analysis to derive a numeric score representing pathway expression, we evaluated clinico-genomic datasets (n=427) from corresponding normal, pre-invasive and invasive cancers of different types, such as ovarian, endometrial, breast and colon cancers in order to determine the associations between the BAD-mediated apoptotic pathway and cancer development. Immunofluorescence was used to compare the expression levels of phosphorylated BAD [pBAD (serine-112, -136 and -155)] in immortalized normal and invasive ovarian, colon and breast cancer cells. The expression of the BAD-mediated apoptotic pathway phosphatase, PP2C, was evaluated by RT-qPCR in the normal and ovarian cancer tissue samples. The growth-promoting effects of pBAD protein levels in the immortalized normal and cancer cells were assessed using siRNA depletion experiments with MTS assays. The expression of the BAD-mediated apoptotic pathway was associated with the development and/or progression of ovarian (n=106, pBAD-mediated apoptotic pathway is thus associated with the development of human cancers likely influenced by the protein levels of pBAD.

  12. Cartography of Pathway Signal Perturbations Identifies Distinct Molecular Pathomechanisms in Malignant and Chronic Lung Diseases

    Science.gov (United States)

    Arakelyan, Arsen; Nersisyan, Lilit; Petrek, Martin; Löffler-Wirth, Henry; Binder, Hans

    2016-01-01

    Lung diseases are described by a wide variety of developmental mechanisms and clinical manifestations. Accurate classification and diagnosis of lung diseases are the bases for development of effective treatments. While extensive studies are conducted toward characterization of various lung diseases at molecular level, no systematic approach has been developed so far. Here we have applied a methodology for pathway-centered mining of high throughput gene expression data to describe a wide range of lung diseases in the light of shared and specific pathway activity profiles. We have applied an algorithm combining a Pathway Signal Flow (PSF) algorithm for estimation of pathway activity deregulation states in lung diseases and malignancies, and a Self Organizing Maps algorithm for classification and clustering of the pathway activity profiles. The analysis results allowed clearly distinguish between cancer and non-cancer lung diseases. Lung cancers were characterized by pathways implicated in cell proliferation, metabolism, while non-malignant lung diseases were characterized by deregulations in pathways involved in immune/inflammatory response and fibrotic tissue remodeling. In contrast to lung malignancies, chronic lung diseases had relatively heterogeneous pathway deregulation profiles. We identified three groups of interstitial lung diseases and showed that the development of characteristic pathological processes, such as fibrosis, can be initiated by deregulations in different signaling pathways. In conclusion, this paper describes the pathobiology of lung diseases from systems viewpoint using pathway centered high-dimensional data mining approach. Our results contribute largely to current understanding of pathological events in lung cancers and non-malignant lung diseases. Moreover, this paper provides new insight into molecular mechanisms of a number of interstitial lung diseases that have been studied to a lesser extent. PMID:27200087

  13. Three distinct developmental pathways for adaptive and two IFN-γ-producing γδ T subsets in adult thymus

    DEFF Research Database (Denmark)

    Buus, Terkild Brink; Ødum, Niels; Geisler, Carsten

    2017-01-01

    -producing γδ T cells (γδNKT). Developmental progression towards both IFN-γ-producing subsets can be induced by TCR signalling, and each pathway results in thymic emigration at a different stage. Finally, we show that γδT1 cells are the predominating IFN-γ-producing subset developing in the adult thymus. Thus......, this study maps out three distinct development pathways that result in the programming of γδTn, γδT1 and γδNKT cells.......Murine γδ T cells include subsets that are programmed for distinct effector functions during their development in the thymus. Under pathological conditions, different γδ T cell subsets can be protective or can exacerbate a disease. Here we show that CD117, CD200 and CD371, together with other...

  14. β-Adrenergic induced SR Ca2+ leak is mediated by an Epac-NOS pathway.

    Science.gov (United States)

    Pereira, Laëtitia; Bare, Dan J; Galice, Samuel; Shannon, Thomas R; Bers, Donald M

    2017-07-01

    Cardiac β-adrenergic receptors (β-AR) and Ca 2+ -Calmodulin dependent protein kinase (CaMKII) regulate both physiological and pathophysiological Ca 2+ signaling. Elevated diastolic Ca 2+ leak from the sarcoplasmic reticulum (SR) contributes to contractile dysfunction in heart failure and to arrhythmogenesis. β-AR activation is known to increase SR Ca 2+ leak via CaMKII-dependent phosphorylation of the ryanodine receptor. Two independent and reportedly parallel pathways have been implicated in this β-AR-CaMKII cascade, one involving exchange protein directly activated by cAMP (Epac2) and another involving nitric oxide synthase 1 (NOS1). Here we tested whether Epac and NOS function in a single series pathway to increase β-AR induced and CaMKII-dependent SR Ca 2+ leak. Leak was measured as both Ca 2+ spark frequency and tetracaine-induced shifts in SR Ca 2+ , in mouse and rabbit ventricular myocytes. Direct Epac activation by 8-CPT (8-(4-chlorophenylthio)-2'-O-methyl-cAMP) mimicked β-AR-induced SR Ca 2+ leak, and both were blocked by NOS inhibition. The same was true for myocyte CaMKII activation (assessed via a FRET-based reporter) and ryanodine receptor phosphorylation. Inhibitor and phosphorylation studies also implicated phosphoinositide 3-kinase (PI3K) and protein kinase B (Akt) downstream of Epac and above NOS activation in this pathway. We conclude that these two independently characterized parallel pathways function mainly via a single series arrangement (β-AR-cAMP-Epac-PI3K-Akt-NOS1-CaMKII) to mediate increased SR Ca 2+ leak. Thus, for β-AR activation the cAMP-PKA branch effects inotropy and lusitropy (by effects on Ca 2+ current and SR Ca 2+ -ATPase), this cAMP-Epac-NOS pathway increases pathological diastolic SR Ca 2+ leak. This pathway distinction may allow novel SR Ca 2+ leak therapeutic targeting in treatment of arrhythmias in heart failure that spare the inotropic and lusitropic effects of the PKA branch. Copyright © 2017 Elsevier Ltd. All

  15. DMPD: Signal transduction pathways mediated by the interaction of CpG DNA withToll-like receptor 9. [Dynamic Macrophage Pathway CSML Database

    Lifescience Database Archive (English)

    Full Text Available 14751759 Signal transduction pathways mediated by the interaction of CpG DNA withTo...;16(1):17-22. (.png) (.svg) (.html) (.csml) Show Signal transduction pathways mediated by the interaction of... CpG DNA withToll-like receptor 9. PubmedID 14751759 Title Signal transduction pathways media

  16. Vasopressin up-regulates the expression of growth-related immediate-early genes via two distinct EGF receptor transactivation pathways

    Science.gov (United States)

    Fuentes, Lida Q.; Reyes, Carlos E.; Sarmiento, José M.; Villanueva, Carolina I.; Figueroa, Carlos D.; Navarro, Javier; González, Carlos B.

    2008-01-01

    Activation of V1a receptor triggers the expression of growth-related immediate-early genes (IEGs), including c-Fos and Egr-1. Here we found that pre-treatment of rat vascular smooth muscle A-10 cell line with the EGF receptor inhibitor AG1478 or the over-expression of an EGFR dominant negative mutant (HEBCD533) blocked the vasopressin-induced expression of IEGs, suggesting that activation of these early genes mediated by V1a receptor is via transactivation of the EGF receptor. Importantly, the inhibition of the metalloproteinases, which catalyzed the shedding of the EGF receptor agonist HB-EGF, selectively blocked the vasopressin-induced expression c-Fos. On the other hand, the inhibition of c-Src selectively blocked the vasopressin-induced expression of Egr-1. Interestingly, in contrast to the expression of c-Fos, the expression of Egr-1 was mediated via the Ras/MEK/MAPK-dependent signalling pathway. Vasopressin-triggered expression of both genes required the release of intracellular calcium, activation of PKC and β-arrestin 2. These findings demonstrated that vasopressin up-regulated the expression of c-Fos and Erg-1 via transactivation of two distinct EGF receptor-dependent signalling pathways. PMID:18571897

  17. tBuLi-Mediated One-Pot Direct Highly Selective Cross-Coupling of Two Distinct Aryl Bromides

    NARCIS (Netherlands)

    Vila, Carlos; Cembellin, Sara; Hornillos, Valentin; Giannerini, Massimo; Fananas-Mastral, Martin; Feringa, Ben L.

    2015-01-01

    A Pd-catalyzed direct cross-coupling of two distinct aryl bromides mediated by tBuLi is described. The use of [Pd-PEPPSI-IPr] or [Pd-PEPPSI-IPent] as catalyst allows for the efficient one-pot synthesis of unsymmetrical biaryls at room temperature. The key for this selective cross-coupling is the use

  18. CHARACTERISTICS OF SIGNALING PATHWAYS MEDIATING A CYTOTOXIC EFFECT OF DENDRITIC CELLS UPON ACTIVATED Т LYMPHOCYTES AND NK CELLS

    Directory of Open Access Journals (Sweden)

    T. V. Tyrinova

    2012-01-01

    Full Text Available Abstract. Cytotoxic/pro-apoptogenic effects of IFNα-induced dendritic cells (IFN-DCs directed against Т-lymphocytes and NK cells were investigated in healthy donors. Using an allogenic MLC system, it was revealed that IFN-DCs induce apoptosis of both activated CD4+ and CD8+ T-lymphocytes, and NK cells. Apoptosis of CD4+ and CD8+ T-lymphocytes induced by their interaction with IFN-DCs was mediated by various signaling pathways. In particular, activated CD4+Т-lymphocytes were most sensitive to TRAIL- и Fas/ FasL-transduction pathways, whereas activated CD8+ T-lymphocytes were induced to apoptosis via TNFα-mediated pathway. PD-1/B7-H1-signaling pathway also played a distinct role in cytotoxic activity of IFNDCs towards both types of T lymphocytes and activated NK cells. The pro-apoptogenic/cytotoxic activity of IFN-DC against activated lymphocytes may be regarded as a mechanism of a feedback regulation aimed at restriction of immune response and maintenance of immune homeostasis. Moreover, upregulation of proapoptogenic molecules on DCs under pathological conditions may lead to suppression of antigen-specific response, thus contributing to the disease progression.

  19. Two Distinct Pathways to Development of Squamous Cell Carcinoma of the Vulva

    International Nuclear Information System (INIS)

    Ueda, Y.; Enomoto, T.; Kimura, T.; Yoshino, K.; Fujita, M.; Kimura, T.

    2011-01-01

    Squamous cell carcinoma (SCC) accounts for approximately 95% of the malignant tumors of the vaginal vulva and is mostly found in elderly women. The future numbers of patients with vulva r SCC is expected to rise, mainly because of the proportional increase in the average age of the general population. Two different pathways for vulva r SCC have been put forth. The first pathway is triggered by infection with a high-risk-type Human Papillomavirus (HPV). Integration of the HPV DNA into the host genome leads to the development of a typical vulva r intraepithelial neoplasia (VIN), accompanied with overexpression ofP14"ARF and P16"INK4A . This lesion subsequently forms a warty- or basaloid-type SCC. The HPV vaccine is a promising new tool for prevention of this HPV related SCC of the vulva. The second pathway is HPV-independent. Keratinizing SCC develops within a background of lichen sclerosus (LS) through a differentiated VIN. It has a different set of genetic alterations than those in the first pathway, including p53 mutations, allelic imbalances (AI), and microsatellite instability (MSI). Further clinical and basic research is still required to understand and prevent vulvar SCC. Capsule. Two pathway for pathogenesis of squamous cell carcinoma of the value are reviewed.

  20. Two Distinct Pathways to Development of Squamous Cell Carcinoma of the Vulva

    Directory of Open Access Journals (Sweden)

    Yutaka Ueda

    2011-01-01

    Full Text Available Squamous cell carcinoma (SCC accounts for approximately 95% of the malignant tumors of the vaginal vulva and is mostly found in elderly women. The future numbers of patients with vulvar SCC is expected to rise, mainly because of the proportional increase in the average age of the general population. Two different pathways for vulvar SCC have been put forth. The first pathway is triggered by infection with a high-risk-type Human Papillomavirus (HPV. Integration of the HPV DNA into the host genome leads to the development of a typical vulvar intraepithelial neoplasia (VIN, accompanied with overexpression of p14ARF and p16INK4A. This lesion subsequently forms a warty- or basaloid-type SCC. The HPV vaccine is a promising new tool for prevention of this HPV related SCC of the vulva. The second pathway is HPV-independent. Keratinizing SCC develops within a background of lichen sclerosus (LS through a differentiated VIN. It has a different set of genetic alterations than those in the first pathway, including p53 mutations, allelic imbalances (AI, and microsatellite instability (MSI. Further clinical and basic research is still required to understand and prevent vulvar SCC. Capsule. Two pathway for pathogenesis of squamous cell carcinoma of the value are reviewed.

  1. Tyrosine kinase chromosomal translocations mediate distinct and overlapping gene regulation events

    International Nuclear Information System (INIS)

    Kim, Hani; Gillis, Lisa C; Jarvis, Jordan D; Yang, Stuart; Huang, Kai; Der, Sandy; Barber, Dwayne L

    2011-01-01

    Leukemia is a heterogeneous disease commonly associated with recurrent chromosomal translocations that involve tyrosine kinases including BCR-ABL, TEL-PDGFRB and TEL-JAK2. Most studies on the activated tyrosine kinases have focused on proximal signaling events, but little is known about gene transcription regulated by these fusions. Oligonucleotide microarray was performed to compare mRNA changes attributable to BCR-ABL, TEL-PDGFRB and TEL-JAK2 after 1 week of activation of each fusion in Ba/F3 cell lines. Imatinib was used to control the activation of BCR-ABL and TEL-PDGFRB, and TEL-JAK2-mediated gene expression was examined 1 week after Ba/F3-TEL-JAK2 cells were switched to factor-independent conditions. Microarray analysis revealed between 800 to 2000 genes induced or suppressed by two-fold or greater by each tyrosine kinase, with a subset of these genes commonly induced or suppressed among the three fusions. Validation by Quantitative PCR confirmed that eight genes (Dok2, Mrvi1, Isg20, Id1, gp49b, Cxcl10, Scinderin, and collagen Vα1(Col5a1)) displayed an overlapping regulation among the three tested fusion proteins. Stat1 and Gbp1 were induced uniquely by TEL-PDGFRB. Our results suggest that BCR-ABL, TEL-PDGFRB and TEL-JAK2 regulate distinct and overlapping gene transcription profiles. Many of the genes identified are known to be involved in processes associated with leukemogenesis, including cell migration, proliferation and differentiation. This study offers the basis for further work that could lead to an understanding of the specificity of diseases caused by these three chromosomal translocations

  2. Neurobiological Correlates and Predictors of Two Distinct Personality Trait Pathways to Escalated Alcohol Use

    Directory of Open Access Journals (Sweden)

    Malak Abu Shakra

    2018-01-01

    Interpretation: This double dissociation provides evidence of distinct neurobiological profiles in a priori identified personality trait-based risk groups for AUDs, and links these signatures to clinically relevant substance use outcomes at follow-up. AUD subtypes might benefit from motivationally and personality-specific ameliorative and preventative interventions.

  3. Distinct functions for the Drosophila piRNA pathway in genome maintenance and telomere protection.

    Directory of Open Access Journals (Sweden)

    Jaspreet S Khurana

    2010-12-01

    Full Text Available Transposons and other selfish DNA elements can be found in all phyla, and mobilization of these elements can compromise genome integrity. The piRNA (PIWI-interacting RNA pathway silences transposons in the germline, but it is unclear if this pathway has additional functions during development. Here we show that mutations in the Drosophila piRNA pathway genes, armi, aub, ago3, and rhi, lead to extensive fragmentation of the zygotic genome during the cleavage stage of embryonic divisions. Additionally, aub and armi show defects in telomere resolution during meiosis and the cleavage divisions; and mutations in lig-IV, which disrupt non-homologous end joining, suppress these fusions. By contrast, lig-IV mutations enhance chromosome fragmentation. Chromatin immunoprecipitation studies show that aub and armi mutations disrupt telomere binding of HOAP, which is a component of the telomere protection complex, and reduce expression of a subpopulation of 19- to 22-nt telomere-specific piRNAs. Mutations in rhi and ago3, by contrast, do not block HOAP binding or production of these piRNAs. These findings uncover genetically separable functions for the Drosophila piRNA pathway. The aub, armi, rhi, and ago3 genes silence transposons and maintain chromosome integrity during cleavage-stage embryonic divisions. However, the aub and armi genes have an additional function in assembly of the telomere protection complex.

  4. Distinctive Modulation of Dopamine Release in the Nucleus Accumbens Shell Mediated by Dopamine and Acetylcholine Receptors.

    Science.gov (United States)

    Shin, Jung Hoon; Adrover, Martin F; Alvarez, Veronica A

    2017-11-15

    Nucleus accumbens (NAc) shell shows unique dopamine (DA) signals in vivo and plays a unique role in DA-dependent behaviors such as reward-motivated learning and the response to drugs of abuse. A disynaptic mechanism for DA release was reported and shown to require synchronized firing of cholinergic interneurons (CINs) and activation of nicotinic acetylcholine (ACh) receptors (nAChRs) in DA neuron (DAN) axons. The properties of this disynaptic mechanism of DA transmission are not well understood in the NAc shell. In this study, in vitro fast-scan cyclic voltammetry was used to examine the modulation of DA transmission evoked by CINs firing in the shell of mice and compared with other striatal regions. We found that DA signals in the shell displayed significant degree of summation in response to train stimulation of CINs, contrary to core and dorsal striatum. The summation was amplified by a D2-like receptor antagonist and experiments with mice with targeted deletion of D2 receptors to DANs or CINs revealed that D2 receptors in CINs mediate a fast inhibition observed within 100 ms of the first pulse, whereas D2 autoreceptors in DAN terminals are engaged in a slower inhibition that peaks at ∼500 ms. ACh also contributes to the use-dependent inhibition of DA release through muscarinic receptors only in the shell, where higher activity of acetylcholinesterase minimizes nAChR desensitization and promotes summation. These findings show that DA signals are modulated differentially by endogenous DA and ACh in the shell, which may underlie the unique features of shell DA signals in vivo SIGNIFICANCE STATEMENT The present study reports that dopamine (DA) release evoked by activation of cholinergic interneurons displays a high degree of summation in the shell and shows unique modulation by endogenous DA and acetylcholine. Desensitization of nicotinic receptors, which is a prevailing mechanism for use-dependent inhibition in the nucleus accumbens core and dorsal striatum, is

  5. Role of Host Type IA Phosphoinositide 3-Kinase Pathway Components in Invasin-Mediated Internalization of Yersinia enterocolitica.

    Science.gov (United States)

    Dowd, Georgina C; Bhalla, Manmeet; Kean, Bernard; Thomas, Rowan; Ireton, Keith

    2016-06-01

    Many bacterial pathogens subvert mammalian type IA phosphoinositide 3-kinase (PI3K) in order to induce their internalization into host cells. How PI3K promotes internalization is not well understood. Also unclear is whether type IA PI3K affects different pathogens through similar or distinct mechanisms. Here, we performed an RNA interference (RNAi)-based screen to identify components of the type IA PI3K pathway involved in invasin-mediated entry of Yersinia enterocolitica, an enteropathogen that causes enteritis and lymphadenitis. The 69 genes targeted encode known upstream regulators or downstream effectors of PI3K. A similar RNAi screen was previously performed with the food-borne bacterium Listeria monocytogenes The results of the screen with Y. enterocolitica indicate that at least nine members of the PI3K pathway are needed for invasin-mediated entry. Several of these proteins, including centaurin-α1, Dock180, focal adhesion kinase (FAK), Grp1, LL5α, LL5β, and PLD2 (phospholipase D2), were recruited to sites of entry. In addition, centaurin-α1, FAK, PLD2, and mTOR were required for remodeling of the actin cytoskeleton during entry. Six of the human proteins affecting invasin-dependent internalization also promote InlB-mediated entry of L. monocytogenes Our results identify several host proteins that mediate invasin-induced effects on the actin cytoskeleton and indicate that a subset of PI3K pathway components promote internalization of both Y. enterocolitica and L. monocytogenes. Copyright © 2016, American Society for Microbiology. All Rights Reserved.

  6. Distinct DNA repair pathways involving RecA and nonhomologous end joining in Mycobacterium smegmatis.

    Science.gov (United States)

    Korycka-Machala, Malgorzata; Brzostek, Anna; Rozalska, Sylwia; Rumijowska-Galewicz, Anna; Dziedzic, Renata; Bowater, Richard; Dziadek, Jaroslaw

    2006-05-01

    Mycobacterium smegmatis was used to study the relationship between DNA repair processes involving RecA and nonhomologous end joining (NHEJ). The effect of gene deletions in recA and/or in two genes involved in NHEJ (ku and ligD) was tested on the ability of bacteria to join breaks in plasmids transformed into them and in their response to chemicals that damage DNA. The results provide in vivo evidence that only NHEJ is required for the repair of noncompatible DNA ends. By contrast, the response of mycobacteria to mitomycin C preferentially involved a RecA-dependent pathway.

  7. IAPs Regulate Distinct Innate Immune Pathways to Co-ordinate the Response to Bacterial Peptidoglycans.

    Science.gov (United States)

    Stafford, Che A; Lawlor, Kate E; Heim, Valentin J; Bankovacki, Aleksandra; Bernardini, Jonathan P; Silke, John; Nachbur, Ueli

    2018-02-06

    Inhibitors of apoptosis (IAPs) proteins are critical regulators of innate immune signaling pathways and therefore have potential as drug targets. X-linked IAP (XIAP) and cellular IAP1 and IAP2 (cIAP1 and cIAP2) are E3 ligases that have been shown to be required for signaling downstream of NOD2, an intracellular receptor for bacterial peptidoglycan. We used genetic and biochemical approaches to compare the responses of IAP-deficient mice and cells to NOD2 stimulation. In all cell types tested, XIAP is the only IAP required for signaling immediately downstream of NOD2, while cIAP1 and cIAP2 are dispensable for NOD2-induced nuclear factor κB (NF-κB) and mitogen-activated protein kinase (MAPK) activation. However, mice lacking cIAP1 or TNFR1 have a blunted cytokine response to NOD2 stimulation. We conclude that cIAPs regulate NOD2-dependent autocrine TNF signaling in vivo and highlight the importance of physiological context in the interplay of innate immune signaling pathways. Copyright © 2018 The Author(s). Published by Elsevier Inc. All rights reserved.

  8. Molecular design and nanoparticle-mediated intracellular delivery of functional proteins to target cellular pathways

    Science.gov (United States)

    Shah, Dhiral Ashwin

    functional proteins can be delivered intracellularly in vitro using nanoparticles and used to target key signaling proteins and regulate cell signaling pathways. The same concept of naturally occurring protein-protein interactions can also be implemented to selectively bring intracellular protein targets in close proximity to proteasomal degradation machinery in cells and effect their depletion from the cellular compartments. This approach will be able to not only target entire pool of proteins to ubiquitination-mediated degradation, but also to specific sub-pools of posttranslationally modified proteins in the cell, provided peptides having distinct binding affinities are identified for posttranslational modifications. This system can then be tested for intracellular protein delivery using nanoparticle carriers to identify roles of different posttranslational modifications on the protein's activity. In future work, we propose to develop a cellular detection system, based on GFP complementation, which can be used to evaluate the efficiency of different protein delivery carriers to internalize proteins into the cell cytosol. We envision the application of nanoscale materials as intracellular protein delivery vehicles to target diverse cell signaling pathways at the posttranslational level, and subsequent metabolic manipulation, which may have interesting therapeutic properties and can potentially target stem cell fate.

  9. Distinct neural pathways mediate alpha7 nicotinic acetylcholine receptor-dependent activation of the forebrain

    DEFF Research Database (Denmark)

    Thomsen, Morten S; Hay-Schmidt, Anders; Hansen, Henrik H

    2010-01-01

    alpha(7) nicotinic acetylcholine receptor (nAChR) agonists are candidates for the treatment of cognitive deficits in schizophrenia. Selective alpha(7) nAChR agonists, such as SSR180711, activate neurons in the medial prefrontal cortex (mPFC) and nucleus accumbens shell (ACCshell) in rats, regions...

  10. Transcriptomic Analysis of Lung Tissue from Cigarette Smoke-Induced Emphysema Murine Models and Human Chronic Obstructive Pulmonary Disease Show Shared and Distinct Pathways.

    Science.gov (United States)

    Yun, Jeong H; Morrow, Jarrett; Owen, Caroline A; Qiu, Weiliang; Glass, Kimberly; Lao, Taotao; Jiang, Zhiqiang; Perrella, Mark A; Silverman, Edwin K; Zhou, Xiaobo; Hersh, Craig P

    2017-07-01

    Although cigarette smoke (CS) is the primary risk factor for chronic obstructive pulmonary disease (COPD), the underlying molecular mechanisms for the significant variability in developing COPD in response to CS are incompletely understood. We performed lung gene expression profiling of two different wild-type murine strains (C57BL/6 and NZW/LacJ) and two genetic models with mutations in COPD genome-wide association study genes (HHIP and FAM13A) after 6 months of chronic CS exposure and compared the results to human COPD lung tissues. We identified gene expression patterns that correlate with severity of emphysema in murine and human lungs. Xenobiotic metabolism and nuclear erythroid 2-related factor 2-mediated oxidative stress response were commonly regulated molecular response patterns in C57BL/6, Hhip +/- , and Fam13a -/- murine strains exposed chronically to CS. The CS-resistant Fam13a -/- mouse and NZW/LacJ strain revealed gene expression response pattern differences. The Fam13a -/- strain diverged in gene expression compared with C57BL/6 control only after CS exposure. However, the NZW/LacJ strain had a unique baseline expression pattern, enriched for nuclear erythroid 2-related factor 2-mediated oxidative stress response and xenobiotic metabolism, and converged to a gene expression pattern similar to the more susceptible wild-type C57BL/6 after CS exposure. These results suggest that distinct molecular pathways may account for resistance to emphysema. Surprisingly, there were few genes commonly modulated in mice and humans. Our study suggests that gene expression responses to CS may be largely species and model dependent, yet shared pathways could provide biologically significant insights underlying individual susceptibility to CS.

  11. Curcumin mediates anticancer effects by modulating multiple cell signaling pathways.

    Science.gov (United States)

    Kunnumakkara, Ajaikumar B; Bordoloi, Devivasha; Harsha, Choudhary; Banik, Kishore; Gupta, Subash C; Aggarwal, Bharat B

    2017-08-01

    Curcumin, a component of a spice native to India, was first isolated in 1815 by Vogel and Pelletier from the rhizomes of Curcuma longa (turmeric) and, subsequently, the chemical structure of curcumin as diferuloylmethane was reported by Milobedzka et al. [(1910) 43., 2163-2170]. Since then, this polyphenol has been shown to exhibit antioxidant, anti-inflammatory, anticancer, antiviral, antibacterial, and antifungal activities. The current review primarily focuses on the anticancer potential of curcumin through the modulation of multiple cell signaling pathways. Curcumin modulates diverse transcription factors, inflammatory cytokines, enzymes, kinases, growth factors, receptors, and various other proteins with an affinity ranging from the pM to the mM range. Furthermore, curcumin effectively regulates tumor cell growth via modulation of numerous cell signaling pathways and potentiates the effect of chemotherapeutic agents and radiation against cancer. Curcumin can interact with most of the targets that are modulated by FDA-approved drugs for cancer therapy. The focus of this review is to discuss the molecular basis for the anticancer activities of curcumin based on preclinical and clinical findings. © 2017 The Author(s). Published by Portland Press Limited on behalf of the Biochemical Society.

  12. Two distinct phases of apoptosis in mammary gland involution: proteinase-independent and -dependent pathways

    Energy Technology Data Exchange (ETDEWEB)

    Lund, Leif R; Romer, John; Thomasset, Nicole; Solberg, Helene; Pyke, Charles; Bissell, Mina J; Dano, Keld; Werb, Zena

    1996-01-01

    Postlactational involution of the mammary gland is characterized by two distinct physiological events: apoptosis of the secretory, epithelial cells undergoing programmed cell death, and proteolytic degradation of the mammary gland basement membrane. We examined the spatial and temporal patterns of apoptotic cells in relation to those of proteinases during involution of the BALB/c mouse mammary gland. Apoptosis was almost absent during lactation but became evident at day 2 of involution, when {beta}-casein gene expression was still high. Apoptotic cells were then seen at least up to day 8 of involution, when {beta}-casein gene expression was being extinguished. Expression of sulfated glycoprotein-2 (SGP-2), interleukin-1{beta} converting enzyme (ICE) and tissue inhibitor of metalloproteinases-1 was upregulated at day 2, when apoptotic cells were seen initially. Expression of the matrix metalloproteinases gelatinase A and stromelysin-1 and the serine proteinase urokinase-type plasminogen activator, which was low during lactation, was strongly upregulated in parallel starting at day 4 after weaning, coinciding with start of the collapse of the lobulo-alveolar structures and the intensive tissue remodeling in involution. The major sites of mRNA synthesis for these proteinases were fibroblast-like cells in the periductal stroma and stromal cells surrounding the collapsed alveoli, suggesting that the degradative phase of involution is due to a specialized mesenchymal-epithelial interaction. To elucidate the functional role of these proteinases during involution, at the onset of weaning we treated mice systemically with the glucocorticoid hydrocortisone, which is known to inhibit mammary gland involution. Although the initial wave of apoptotic cells appeared in the lumina of the gland, the dramatic regression and tissue remodeling usually evident by day 5 was substantially inhibited by systemic treatment with hydrocortisone. mRNA and protein for gelatinase A, stromelysin

  13. Mediation pathways and effects of green structures on respiratory mortality via reducing air pollution

    OpenAIRE

    Shen, Yu-Sheng; Lung, Shih-Chun Candice

    2017-01-01

    Previous studies have shown both health and environmental benefits of green spaces, especially in moderating temperature and reducing air pollution. However, the characteristics of green structures have been overlooked in previous investigations. In addition, the mediation effects of green structures on respiratory mortality have not been assessed. This study explores the potential mediation pathways and effects of green structure characteristics on respiratory mortality through temperature, ...

  14. A chimeric prokaryotic pentameric ligand–gated channel reveals distinct pathways of activation

    Science.gov (United States)

    Schmandt, Nicolaus; Velisetty, Phanindra; Chalamalasetti, Sreevatsa V.; Stein, Richard A.; Bonner, Ross; Talley, Lauren; Parker, Mark D.; Mchaourab, Hassane S.; Yee, Vivien C.; Lodowski, David T.

    2015-01-01

    Recent high resolution structures of several pentameric ligand–gated ion channels have provided unprecedented details of their molecular architecture. However, the conformational dynamics and structural rearrangements that underlie gating and allosteric modulation remain poorly understood. We used a combination of electrophysiology, double electron–electron resonance (DEER) spectroscopy, and x-ray crystallography to investigate activation mechanisms in a novel functional chimera with the extracellular domain (ECD) of amine-gated Erwinia chrysanthemi ligand–gated ion channel, which is activated by primary amines, and the transmembrane domain of Gloeobacter violaceus ligand–gated ion channel, which is activated by protons. We found that the chimera was independently gated by primary amines and by protons. The crystal structure of the chimera in its resting state, at pH 7.0 and in the absence of primary amines, revealed a closed-pore conformation and an ECD that is twisted with respect to the transmembrane region. Amine- and pH-induced conformational changes measured by DEER spectroscopy showed that the chimera exhibits a dual mode of gating that preserves the distinct conformational changes of the parent channels. Collectively, our findings shed light on both conserved and divergent features of gating mechanisms in this class of channels, and will facilitate the design of better allosteric modulators. PMID:26415570

  15. Distinct Cell Guidance Pathways Controlled by the Rac and Rho GEF Domains of UNC-73/TRIO in Caenorhabditis elegans

    Science.gov (United States)

    Marcus-Gueret, Nancy; Schmidt, Kristopher L.; Stringham, Eve G.

    2012-01-01

    The cytoskeleton regulator UNC-53/NAV2 is required for both the anterior and posterior outgrowth of several neurons as well as that of the excretory cell while the kinesin-like motor VAB-8 is essential for most posteriorly directed migrations in Caenorhabditis elegans. Null mutations in either unc-53 or vab-8 result in reduced posterior excretory canal outgrowth, while double null mutants display an enhanced canal extension defect, suggesting the genes act in separate pathways to control this posteriorly directed outgrowth. Genetic analysis of putative interactors of UNC-53 or VAB-8, and cell-specific rescue experiments suggest that VAB-8, SAX-3/ROBO, SLT-1/Slit, and EVA-1 are functioning together in the outgrowth of the excretory canals, while UNC-53 appears to function in a parallel pathway with UNC-71/ADAM. The known VAB-8 interactor, the Rac/Rho GEF UNC-73/TRIO operates in both pathways, as isoform specific alleles exhibit enhancement of the phenotype in double-mutant combination with either unc-53 or vab-8. On the basis of these results, we propose a bipartite model for UNC-73/TRIO activity in excretory canal extension: a cell autonomous function that is mediated by the Rho-specific GEF domain of the UNC-73E isoform in conjunction with UNC-53 and UNC-71 and a cell nonautonomous function that is mediated by the Rac-specific GEF domain of the UNC-73B isoform, through partnering with VAB-8 and the receptors SAX-3 and EVA-1. PMID:21996675

  16. Regulators of the proteasome pathway, Uch37 and Rpn13, play distinct roles in mouse development.

    Directory of Open Access Journals (Sweden)

    Amin Al-Shami

    Full Text Available Rpn13 is a novel mammalian proteasomal receptor that has recently been identified as an amplification target in ovarian cancer. It can interact with ubiquitin and activate the deubiquitinating enzyme Uch37 at the 26S proteasome. Since neither Rpn13 nor Uch37 is an integral proteasomal subunit, we explored whether either protein is essential for mammalian development and survival. Deletion of Uch37 resulted in prenatal lethality in mice associated with severe defect in embryonic brain development. In contrast, the majority of Rpn13-deficient mice survived to adulthood, although they were smaller at birth and fewer in number than wild-type littermates. Absence of Rpn13 produced tissue-specific effects on proteasomal function: increased proteasome activity in adrenal gland and lymphoid organs, and decreased activity in testes and brain. Adult Rpn13(-/- mice reached normal body weight but had increased body fat content and were infertile due to defective gametogenesis. Additionally, Rpn13(-/- mice showed increased T-cell numbers, resembling growth hormone-mediated effects. Indeed, serum growth hormone and follicular stimulating hormone levels were significantly increased in Rpn13(-/- mice, while growth hormone receptor expression was reduced in the testes. In conclusion, this is the first report characterizing the physiological roles of Uch37 and Rpn13 in murine development and implicating a non-ATPase proteasomal protein, Rpn13, in the process of gametogenesis.

  17. Two distinct sensing pathways allow recognition of Klebsiella pneumoniae by Dictyostelium amoebae.

    Science.gov (United States)

    Lima, Wanessa C; Balestrino, Damien; Forestier, Christiane; Cosson, Pierre

    2014-03-01

    Recognition of bacteria by metazoans is mediated by receptors that recognize different types of microorganisms and elicit specific cellular responses. The soil amoebae Dictyostelium discoideum feeds upon a variable mixture of environmental bacteria, and it is expected to recognize and adapt to various food sources. To date, however, no bacteria-sensing mechanisms have been described. In this study, we isolated a Dictyostelium mutant (fspA KO) unable to grow in the presence of non-capsulated Klebsiella pneumoniae bacteria, but growing as efficiently as wild-type cells in the presence of other bacteria, such as Bacillus subtilis. fspA KO cells were also unable to respond to K. pneumoniae and more specifically to bacterially secreted folate in a chemokinetic assay, while they responded readily to B. subtilis. Remarkably, both WT and fspA KO cells were able to grow in the presence of capsulated LM21 K. pneumoniae, and responded to purified capsule, indicating that capsule recognition may represent an alternative, FspA-independent mechanism for K. pneumoniae sensing. When LM21 capsule synthesis genes were deleted, growth and chemokinetic response were lost for fspA KO cells, but not for WT cells. Altogether, these results indicate that Dictyostelium amoebae use specific recognition mechanisms to respond to different K. pneumoniae elements. © 2013 John Wiley & Sons Ltd.

  18. Cooperation of distinct Rac-dependent pathways to stabilise E-cadherin adhesion.

    Science.gov (United States)

    Erasmus, Jennifer C; Welsh, Natalie J; Braga, Vania M M

    2015-09-01

    The precise mechanisms via which Rac1 is activated by cadherin junctions are not fully known. In keratinocytes Rac1 activation by cadherin junctions requires EGFR signalling, but how EGFR does so is unclear. To address which activator could mediate E-cadherin signalling to Rac1, we investigated EGFR and two Rac1 GEFs, SOS1 and DOCK180. EGFR RNAi prevented junction-induced Rac1 activation and led to fragmented localization of E-cadherin at cadherin contacts. In contrast, depletion of another EGFR family member, ErbB3, did not interfere with either process. DOCK180 RNAi, but not SOS1, prevented E-cadherin-induced Rac1 activation. However, in a strong divergence from EGFR RNAi phenotype, DOCK180 depletion did not perturb actin recruitment or cadherin localisation at junctions. Rather, reduced DOCK180 levels impaired the resistance to mechanical stress of pre-formed cell aggregates. Thus, within the same cell type, EGFR and DOCK180 regulate Rac1 activation by newly-formed contacts, but control separate cellular events that cooperate to stabilise junctions. Copyright © 2015. Published by Elsevier Inc.

  19. Importance of Mediator complex in the regulation and integration of diverse signaling pathways in plants.

    Science.gov (United States)

    Samanta, Subhasis; Thakur, Jitendra K

    2015-01-01

    Basic transcriptional machinery in eukaryotes is assisted by a number of cofactors, which either increase or decrease the rate of transcription. Mediator complex is one such cofactor, and recently has drawn a lot of interest because of its integrative power to converge different signaling pathways before channeling the transcription instructions to the RNA polymerase II machinery. Like yeast and metazoans, plants do possess the Mediator complex across the kingdom, and its isolation and subunit analyses have been reported from the model plant, Arabidopsis. Genetic, and molecular analyses have unraveled important regulatory roles of Mediator subunits at every stage of plant life cycle starting from flowering to embryo and organ development, to even size determination. It also contributes immensely to the survival of plants against different environmental vagaries by the timely activation of its resistance mechanisms. Here, we have provided an overview of plant Mediator complex starting from its discovery to regulation of stoichiometry of its subunits. We have also reviewed involvement of different Mediator subunits in different processes and pathways including defense response pathways evoked by diverse biotic cues. Wherever possible, attempts have been made to provide mechanistic insight of Mediator's involvement in these processes.

  20. Neuropeptidomics Mass Spectrometry Reveals Signaling Networks Generated by Distinct Protease Pathways in Human Systems

    Science.gov (United States)

    Hook, Vivian; Bandeira, Nuno

    2015-12-01

    Neuropeptides regulate intercellular signaling as neurotransmitters of the central and peripheral nervous systems, and as peptide hormones in the endocrine system. Diverse neuropeptides of distinct primary sequences of various lengths, often with post-translational modifications, coordinate and integrate regulation of physiological functions. Mass spectrometry-based analysis of the diverse neuropeptide structures in neuropeptidomics research is necessary to define the full complement of neuropeptide signaling molecules. Human neuropeptidomics has notable importance in defining normal and dysfunctional neuropeptide signaling in human health and disease. Neuropeptidomics has great potential for expansion in translational research opportunities for defining neuropeptide mechanisms of human diseases, providing novel neuropeptide drug targets for drug discovery, and monitoring neuropeptides as biomarkers of drug responses. In consideration of the high impact of human neuropeptidomics for health, an observed gap in this discipline is the few published articles in human neuropeptidomics compared with, for example, human proteomics and related mass spectrometry disciplines. Focus on human neuropeptidomics will advance new knowledge of the complex neuropeptide signaling networks participating in the fine control of neuroendocrine systems. This commentary review article discusses several human neuropeptidomics accomplishments that illustrate the rapidly expanding diversity of neuropeptides generated by protease processing of pro-neuropeptide precursors occurring within the secretory vesicle proteome. Of particular interest is the finding that human-specific cathepsin V participates in producing enkephalin and likely other neuropeptides, indicating unique proteolytic mechanisms for generating human neuropeptides. The field of human neuropeptidomics has great promise to solve new mechanisms in disease conditions, leading to new drug targets and therapeutic agents for human

  1. Distinct roles of two anaplerotic pathways in glutamate production induced by biotin limitation in Corynebacterium glutamicum.

    Science.gov (United States)

    Sato, Hiroki; Orishimo, Keita; Shirai, Tomokazu; Hirasawa, Takashi; Nagahisa, Keisuke; Shimizu, Hiroshi; Wachi, Masaaki

    2008-07-01

    Corynebacterium glutamicum is a biotin auxotrophic bacterium in which glutamate production is induced under biotin-limited conditions. During glutamate production, anaplerotic reactions catalyzed by phosphoenolpyruvate carboxylase (PEPC) and a biotin-containing enzyme pyruvate carboxylase (PC) are believed to play an important role in supplying oxaloacetate in the tricarboxylic acid cycle. To understand the distinct roles of PEPC and PC on glutamate production by C. glutamicum, we observed glutamate production induced under biotin-limited conditions in the disruptants of the genes encoding PEPC (ppc) and PC (pyc), respectively. The pyc disruptant retained the ability to produce high amounts of glutamate, and lactate was simultaneously produced probably due to the increased intracellular pyruvate levels. On the other hand, the ppc knockout mutant could not produce glutamate. Additionally, glutamate production in the pyc disruptant was enhanced by overexpression of ppc rather than disruption of the lactate dehydrogenase gene (ldh), which is involved in lactate production. Metabolic flux analysis based on the 13C-labeling experiment and measurement of 13C-enrichment in glutamate using nuclear magnetic resonance spectroscopy revealed that the flux for anaplerotic reactions in the pyc disruptant was lower than that in the wild type, concomitantly increasing the flux for lactate formation. Moreover, overexpression of ppc increased this flux in both the pyc disruptant and the wild type. Our results suggest that the PEPC-catalyzed anaplerotic reaction is necessary for glutamate production induced under biotin-limited conditions, because PC is not active during glutamate production, and overexpression of ppc effectively enhances glutamate production under biotin-limited conditions.

  2. Specificity in mediated pathways by anxiety symptoms linking adolescent stress profiles to depressive symptoms: Results of a moderated mediation approach.

    Science.gov (United States)

    Anyan, Frederick; Bizumic, Boris; Hjemdal, Odin

    2018-03-01

    We investigated the specificity in mediated pathways that separately link specific stress dimensions through anxiety to depressive symptoms and the protective utility of resilience. Thus, this study goes beyond lumping together potential mediating and moderating processes that can explain the relations between stress and (symptoms of) psychopathology and the buffering effect of resilience. Ghanaian adolescents between 13 and 17 years (female = 285; male = 244) completed the Adolescent Stress Questionnaire (ASQ), Spielberger State Anxiety Inventory (STAI), Short Mood Feeling Questionnaire (SMFQ) and the Resilience Scale for Adolescents (READ). Independent samples t-test, multivariate analysis of covariance with follow-up tests and moderated mediation analyses were performed. Evidences were found for specificity in the associations between dimensions of adolescent stressors and depressive symptoms independent of transient anxiety. Transient anxiety partly accounted for the indirect effects of eight stress dimensions on depressive symptoms. Except stress of school attendance and school/leisure conflict, resilience moderated the indirect effects of specific stress dimensions on depressive symptoms. Results suggested differences in how Ghanaian adolescents view the various stress dimensions, and mediated pathways associated with anxiety and depressive symptoms. Use of cross-sectional data does not show causal process and temporal changes over time. Findings support and clarify the specificity in the interrelations and mediated pathways among dimensions of adolescent stress, transient anxiety, and depressive symptoms. Conditional process analyses shows that resilience does not only buffer direct, but also indirect psychological adversities. Interventions for good mental health may focus on low resilience subgroups in specific stress dimensions while minimizing transient anxiety. Copyright © 2017 Elsevier B.V. All rights reserved.

  3. Two Distinct Aerobic Methionine Salvage Pathways Generate Volatile Methanethiol in Rhodopseudomonas palustris

    Science.gov (United States)

    Miller, Anthony R.; North, Justin A.; Wildenthal, John A.

    2018-01-01

    ABSTRACT 5′-Methyl-thioadenosine (MTA) is a dead-end, sulfur-containing metabolite and cellular inhibitor that arises from S-adenosyl-l-methionine-dependent reactions. Recent studies have indicated that there are diverse bacterial methionine salvage pathways (MSPs) for MTA detoxification and sulfur salvage. Here, via a combination of gene deletions and directed metabolite detection studies, we report that under aerobic conditions the facultatively anaerobic bacterium Rhodopseudomonas palustris employs both an MTA-isoprenoid shunt identical to that previously described in Rhodospirillum rubrum and a second novel MSP, both of which generate a methanethiol intermediate. The additional R. palustris aerobic MSP, a dihydroxyacetone phosphate (DHAP)-methanethiol shunt, initially converts MTA to 2-(methylthio)ethanol and DHAP. This is identical to the initial steps of the recently reported anaerobic ethylene-forming MSP, the DHAP-ethylene shunt. The aerobic DHAP-methanethiol shunt then further metabolizes 2-(methylthio)ethanol to methanethiol, which can be directly utilized by O-acetyl-l-homoserine sulfhydrylase to regenerate methionine. This is in contrast to the anaerobic DHAP-ethylene shunt, which metabolizes 2-(methylthio)ethanol to ethylene and an unknown organo-sulfur intermediate, revealing functional diversity in MSPs utilizing a 2-(methylthio)ethanol intermediate. When MTA was fed to aerobically growing cells, the rate of volatile methanethiol release was constant irrespective of the presence of sulfate, suggesting a general housekeeping function for these MSPs up through the methanethiol production step. Methanethiol and dimethyl sulfide (DMS), two of the most important compounds of the global sulfur cycle, appear to arise not only from marine ecosystems but from terrestrial ones as well. These results reveal a possible route by which methanethiol might be biologically produced in soil and freshwater environments. PMID:29636438

  4. Dynamic microtubule organization and mitochondrial transport are regulated by distinct Kinesin-1 pathways

    Directory of Open Access Journals (Sweden)

    Anna Melkov

    2015-12-01

    Full Text Available The microtubule (MT plus-end motor kinesin heavy chain (Khc is well known for its role in long distance cargo transport. Recent evidence showed that Khc is also required for the organization of the cellular MT network by mediating MT sliding. We found that mutations in Khc and the gene of its adaptor protein, kinesin light chain (Klc resulted in identical bristle morphology defects, with the upper part of the bristle being thinner and flatter than normal and failing to taper towards the bristle tip. We demonstrate that bristle mitochondria transport requires Khc but not Klc as a competing force to dynein heavy chain (Dhc. Surprisingly, we demonstrate for the first time that Dhc is the primary motor for both anterograde and retrograde fast mitochondria transport. We found that the upper part of Khc and Klc mutant bristles lacked stable MTs. When following dynamic MT polymerization via the use of GFP-tagged end-binding protein 1 (EB1, it was noted that at Khc and Klc mutant bristle tips, dynamic MTs significantly deviated from the bristle parallel growth axis, relative to wild-type bristles. We also observed that GFP-EB1 failed to concentrate as a focus at the tip of Khc and Klc mutant bristles. We propose that the failure of bristle tapering is due to defects in directing dynamic MTs at the growing tip. Thus, we reveal a new function for Khc and Klc in directing dynamic MTs during polarized cell growth. Moreover, we also demonstrate a novel mode of coordination in mitochondrial transport between Khc and Dhc.

  5. Two Distinct Pathways for Metabolism of Theophylline and Caffeine Are Coexpressed in Pseudomonas putida CBB5▿ †

    Science.gov (United States)

    Yu, Chi Li; Louie, Tai Man; Summers, Ryan; Kale, Yogesh; Gopishetty, Sridhar; Subramanian, Mani

    2009-01-01

    Pseudomonas putida CBB5 was isolated from soil by enrichment on caffeine. This strain used not only caffeine, theobromine, paraxanthine, and 7-methylxanthine as sole carbon and nitrogen sources but also theophylline and 3-methylxanthine. Analyses of metabolites in spent media and resting cell suspensions confirmed that CBB5 initially N demethylated theophylline via a hitherto unreported pathway to 1- and 3-methylxanthines. NAD(P)H-dependent conversion of theophylline to 1- and 3-methylxanthines was also detected in the crude cell extracts of theophylline-grown CBB5. 1-Methylxanthine and 3-methylxanthine were subsequently N demethylated to xanthine. CBB5 also oxidized theophylline and 1- and 3-methylxanthines to 1,3-dimethyluric acid and 1- and 3-methyluric acids, respectively. However, these methyluric acids were not metabolized further. A broad-substrate-range xanthine-oxidizing enzyme was responsible for the formation of these methyluric acids. In contrast, CBB5 metabolized caffeine to theobromine (major metabolite) and paraxanthine (minor metabolite). These dimethylxanthines were further N demethylated to xanthine via 7-methylxanthine. Theobromine-, paraxanthine-, and 7-methylxanthine-grown cells also metabolized all of the methylxanthines mentioned above via the same pathway. Thus, the theophylline and caffeine N-demethylation pathways converged at xanthine via different methylxanthine intermediates. Xanthine was eventually oxidized to uric acid. Enzymes involved in theophylline and caffeine degradation were coexpressed when CBB5 was grown on theophylline or on caffeine or its metabolites. However, 3-methylxanthine-grown CBB5 cells did not metabolize caffeine, whereas theophylline was metabolized at much reduced levels to only methyluric acids. To our knowledge, this is the first report of theophylline N demethylation and coexpression of distinct pathways for caffeine and theophylline degradation in bacteria. PMID:19447909

  6. Chronic stress and peripheral pain: Evidence for distinct, region-specific changes in visceral and somatosensory pain regulatory pathways.

    Science.gov (United States)

    Zheng, Gen; Hong, Shuangsong; Hayes, John M; Wiley, John W

    2015-11-01

    Chronic stress alters the hypothalamic-pituitary-adrenal (HPA) axis and enhances visceral and somatosensory pain perception. It is unresolved whether chronic stress has distinct effects on visceral and somatosensory pain regulatory pathways. Previous studies reported that stress-induced visceral hyperalgesia is associated with reciprocal alterations of endovanilloid and endocannabinoid pain pathways in DRG neurons innervating the pelvic viscera. In this study, we compared somatosensory and visceral hyperalgesia with respect to differential responses of peripheral pain regulatory pathways in a rat model of chronic, intermittent stress. We found that chronic stress induced reciprocal changes in the endocannabinoid 2-AG (increased) and endocannabinoid degradation enzymes COX-2 and FAAH (decreased), associated with down-regulation of CB1 and up-regulation of TRPV1 receptors in L6-S2 DRG but not L4-L5 DRG neurons. In contrast, sodium channels Nav1.7 and Nav1.8 were up-regulated in L4-L5 but not L6-S2 DRGs in stressed rats, which was reproduced in control DRGs treated with corticosterone in vitro. The reciprocal changes of CB1, TRPV1 and sodium channels were cell-specific and observed in the sub-population of nociceptive neurons. Behavioral assessment showed that visceral hyperalgesia persisted, whereas somatosensory hyperalgesia and enhanced expression of Nav1.7 and Nav1.8 sodium channels in L4-L5 DRGs normalized 3 days after completion of the stress phase. These data indicate that chronic stress induces visceral and somatosensory hyperalgesia that involves differential changes in endovanilloid and endocannabinoid pathways, and sodium channels in DRGs innervating the pelvic viscera and lower extremities. These results suggest that chronic stress-induced visceral and lower extremity somatosensory hyperalgesia can be treated selectively at different levels of the spinal cord. Copyright © 2015 Elsevier Inc. All rights reserved.

  7. Identification of a novel Gnao-mediated alternate olfactory signaling pathway in murine OSNs

    Directory of Open Access Journals (Sweden)

    Paul eScholz

    2016-03-01

    Full Text Available It is generally agreed that in olfactory sensory neurons (OSNs, the binding of odorant molecules to their specific olfactory receptor (OR triggers a cAMP-dependent signaling cascade, activating cyclic-nucleotide gated (CNG channels. However, considerable controversy dating back more than 20 years has surrounded the question of whether alternate signaling plays a role in mammalian olfactory transduction. In this study, we demonstrate a specific alternate signaling pathway in Olfr73-expressing OSNs. Methylisoeugenol (MIEG and at least one other known weak Olfr73 agonist (Raspberry Ketone trigger a signaling cascade independent from the canonical pathway, leading to the depolarization of the cell. Interestingly, this pathway is mediated by Gnao activation, leading to Cl- efflux; however, the activation of adenylyl cyclase III (ACIII, the recruitment of Ca2+ from extra-or intracellular stores, and phosphatidylinositol 3-kinase-dependent signaling (PI signaling are not involved. Furthermore, we demonstrated that our newly identified pathway coexists with the canonical olfactory cAMP pathway in the same OSN and can be triggered by the same OR in a ligand-selective manner. We suggest that this pathway might reflect a mechanism for odor recognition predominantly used in early developmental stages before olfactory cAMP signaling is fully developed. Taken together, our findings support the existence of at least one odor-induced alternate signal transduction pathway in native OSNs mediated by Olfr73 in a ligand-selective manner.

  8. Floral pathway integrator gene expression mediates gradual transmission of environmental and endogenous cues to flowering time.

    Science.gov (United States)

    van Dijk, Aalt D J; Molenaar, Jaap

    2017-01-01

    The appropriate timing of flowering is crucial for the reproductive success of plants. Hence, intricate genetic networks integrate various environmental and endogenous cues such as temperature or hormonal statues. These signals integrate into a network of floral pathway integrator genes. At a quantitative level, it is currently unclear how the impact of genetic variation in signaling pathways on flowering time is mediated by floral pathway integrator genes. Here, using datasets available from literature, we connect Arabidopsis thaliana flowering time in genetic backgrounds varying in upstream signalling components with the expression levels of floral pathway integrator genes in these genetic backgrounds. Our modelling results indicate that flowering time depends in a quite linear way on expression levels of floral pathway integrator genes. This gradual, proportional response of flowering time to upstream changes enables a gradual adaptation to changing environmental factors such as temperature and light.

  9. Floral pathway integrator gene expression mediates gradual transmission of environmental and endogenous cues to flowering time

    Directory of Open Access Journals (Sweden)

    Aalt D.J. van Dijk

    2017-04-01

    Full Text Available The appropriate timing of flowering is crucial for the reproductive success of plants. Hence, intricate genetic networks integrate various environmental and endogenous cues such as temperature or hormonal statues. These signals integrate into a network of floral pathway integrator genes. At a quantitative level, it is currently unclear how the impact of genetic variation in signaling pathways on flowering time is mediated by floral pathway integrator genes. Here, using datasets available from literature, we connect Arabidopsis thaliana flowering time in genetic backgrounds varying in upstream signalling components with the expression levels of floral pathway integrator genes in these genetic backgrounds. Our modelling results indicate that flowering time depends in a quite linear way on expression levels of floral pathway integrator genes. This gradual, proportional response of flowering time to upstream changes enables a gradual adaptation to changing environmental factors such as temperature and light.

  10. EVIDENCE FOR EGFR PATHWAY MEDIATION OF CLEFT PALATE INDUCTION BY TCDD

    Science.gov (United States)

    EVIDENCE FOR EGFR PATHWAY MEDIATION OF CLEFT PALATE INDUCTION BY TCDD. B D Abbott, A R Buckalew, and K E Leffler. RTD, NHEERL, ORD,US EPA, RTP, NC, USA.2,3,7,8-Tetrachlorodibenzo-p-dioxin (TCDD) is teratogenic in C57BL/6J mice, producing cleft palate (CP) after exposure...

  11. USP21 regulates Hippo pathway activity by mediating MARK protein turnover

    DEFF Research Database (Denmark)

    Nguyen, Thanh Hung; Kugler, Jan-Michael; Loya, Anand Chainsukh

    2017-01-01

    observed in cancer and often correlates with worse survival. The activity and stability of Hippo pathway components, including YAP/TAZ, AMOT and LATS1/2, are regulated by ubiquitin-mediated protein degradation. Aberrant expression of ubiquitin ligase complexes that regulate the turnover of Hippo components...

  12. The ARG1-LIKE2 gene of Arabidopsis functions in a gravity signal transduction pathway that is genetically distinct from the PGM pathway

    Science.gov (United States)

    Guan, Changhui; Rosen, Elizabeth S.; Boonsirichai, Kanokporn; Poff, Kenneth L.; Masson, Patrick H.

    2003-01-01

    The arl2 mutants of Arabidopsis display altered root and hypocotyl gravitropism, whereas their inflorescence stems are fully gravitropic. Interestingly, mutant roots respond like the wild type to phytohormones and an inhibitor of polar auxin transport. Also, their cap columella cells accumulate starch similarly to wild-type cells, and mutant hypocotyls display strong phototropic responses to lateral light stimulation. The ARL2 gene encodes a DnaJ-like protein similar to ARG1, another protein previously implicated in gravity signal transduction in Arabidopsis seedlings. ARL2 is expressed at low levels in all organs of seedlings and plants. arl2-1 arg1-2 double mutant roots display kinetics of gravitropism similar to those of single mutants. However, double mutants carrying both arl2-1 and pgm-1 (a mutation in the starch-biosynthetic gene PHOSPHOGLUCOMUTASE) at the homozygous state display a more pronounced root gravitropic defect than the single mutants. On the other hand, seedlings with a null mutation in ARL1, a paralog of ARG1 and ARL2, behave similarly to the wild type in gravitropism and other related assays. Taken together, the results suggest that ARG1 and ARL2 function in the same gravity signal transduction pathway in the hypocotyl and root of Arabidopsis seedlings, distinct from the pathway involving PGM.

  13. Puma and Trail/Dr5 Pathways Control Radiation-Induced Apoptosis in Distinct Populations of Testicular Progenitors

    International Nuclear Information System (INIS)

    Coureuil, M.; Tavernier, M.; Barroca, V.; Fouchet, P.; Allemand, I.; Ugolin, N.; Chevillard, S.

    2010-01-01

    Spermatogonia- stem cells and progenitors of adult spermatogenesis- are killed through a p53-regulated apoptotic process after γ-irradiation but the death effectors are still poorly characterized. Our data demonstrate that both intrinsic and extrinsic apoptotic pathways are involved, and especially that spermatogonia can be split into two main populations, according to apoptotic effectors. Following irradiation both Dr5 and Puma genes are up-regulated in the α 6 -integrin-positive Side Population (SP) fraction, which is highly enriched in spermatogonia. Flow cytometric analysis confirms an increased number of Dr5-expressing SP cells, and Puma-β isoform accumulates in α 6 -integrin positive cellular extracts, enriched in spermatogonia. Trail -/- or Puma -/- spermatogonia display a reduced sensitivity to radiation-induced apoptosis. The TUNEL kinetics strongly suggest that the extrinsic and intrinsic pathways, via Trail/Dr5 and Puma respectively, could be engaged in distinct subpopulations of spermatogonia. Indeed flow cytometric studies show that Dr5 receptor is constitutively present on more than half of the undifferentiated progenitors (Kit - α 6 + SP) and half of the differentiated ones (Kit + α 6 + SP). In addition after irradiation, Puma is not detected in the Dr5-positive cellular fraction isolated by immuno-magnetic purification, while Puma is present in the Dr5-negative cell extracts. In conclusion, adult testicular progenitors are divided into distinct sub-populations by apoptotic effectors, independently of progenitor types (immature Kit-negative versus mature Kit-positive), underscoring differential radiosensitivities characterizing the stem cell/progenitors compartment. (authors)

  14. Importance of Mediator complex in the regulation and integration of diverse signaling pathways in plants

    Directory of Open Access Journals (Sweden)

    Subhasis eSamanta

    2015-09-01

    Full Text Available Basic transcriptional machinery in eukaryotes is assisted by a number of cofactors, which either increase or decrease the rate of transcription. Mediator complex is one such cofactor, and recently has drawn a lot of interest because of its integrative power to converge different signaling pathways before channelling the transcription instructions to the RNA polymerase II machinery. Like yeast and metazoans, plants do possess the Mediator complex across the kingdom, and its isolation and subunit analyses have been reported from the model plant, Arabidopsis. Genetic and molecular analyses have unravelled important regulatory roles of Mediator subunits at every stage of plant life cycle starting from flowering to embryo and organ development, to even size determination. It also contributes immensely to the survival of plants against different environmental vagaries by the timely activation of its resistance mechanisms. Here, we have provided an overview of plant Mediator complex starting from its discovery to regulation of stoichiometry of its subunits. We have also reviewed involvement of different Mediator subunits in different processes and pathways including defense response pathways evoked by diverse biotic cues. Wherever possible, attempts have been made to provide mechanistic insight of Mediator’s involvement in these processes.

  15. Acetylcholine produces contraction mediated by cyclooxigenase pathway in arterial vessels in the marine fish (Isacia conceptionis

    Directory of Open Access Journals (Sweden)

    FA. Moraga

    Full Text Available Preliminary studies showed that dorsal artery contraction mediated by acetylcholine (ACh is blocked with indomethacin in intertidal fish (G. laevifrons. Our objective was to characterize the cholinergic pathway in several artery vessels of the I. conceptionis. Afferent and efferent branchial, dorsal and mesenteric arteries were dissected of 6 juvenile specimens, isometric tension studies were done using doses response curves (DRC for Ach (10–13 to 10–3 M, and cholinergic pathways were obtained by blocking with atropine or indomethacin. CRC to ACh showed a pattern of high sensitivity only in efferente branchial artery and low sensibility in all vessels. Furthermore, these contractions were blocked in the presence of atropine and indomethacin in all vessels. Our results corroborate previous results observed in intertidal species that contraction induced by acetylcholine is mediated by receptors that activate a cyclooxygenase contraction pathway.

  16. RNF4-mediated polyubiquitination regulates the Fanconi anemia/BRCA pathway.

    Science.gov (United States)

    Xie, Jenny; Kim, Hyungjin; Moreau, Lisa A; Puhalla, Shannon; Garber, Judy; Al Abo, Muthana; Takeda, Shunichi; D'Andrea, Alan D

    2015-04-01

    The Fanconi anemia/BRCA (FA/BRCA) pathway is a DNA repair pathway that is required for excision of DNA interstrand cross-links. The 17 known FA proteins, along with several FA-associated proteins (FAAPs), cooperate in this pathway to detect, unhook, and excise DNA cross-links and to subsequently repair the double-strand breaks generated in the process. In the current study, we identified a patient with FA with a point mutation in FANCA, which encodes a mutant FANCA protein (FANCAI939S). FANCAI939S failed to bind to the FAAP20 subunit of the FA core complex, leading to decreased stability. Loss of FAAP20 binding exposed a SUMOylation site on FANCA at amino acid residue K921, resulting in E2 SUMO-conjugating enzyme UBC9-mediated SUMOylation, RING finger protein 4-mediated (RNF4-mediated) polyubiquitination, and proteasome-mediated degradation of FANCA. Mutation of the SUMOylation site of FANCA rescued the expression of the mutant protein. Wild-type FANCA was also subject to SUMOylation, RNF4-mediated polyubiquitination, and degradation, suggesting that regulated release of FAAP20 from FANCA is a critical step in the normal FA pathway. Consistent with this model, cells lacking RNF4 exhibited interstrand cross-linker hypersensitivity, and the gene encoding RNF4 was epistatic with the other genes encoding members of the FA/BRCA pathway. Together, the results from our study underscore the importance of analyzing unique patient-derived mutations for dissecting complex DNA repair processes.

  17. Adaptive immunity alters distinct host feeding pathways during nematode induced inflammation, a novel mechanism in parasite expulsion.

    Directory of Open Access Journals (Sweden)

    John J Worthington

    2013-01-01

    Full Text Available Gastrointestinal infection is often associated with hypophagia and weight loss; however, the precise mechanisms governing these responses remain poorly defined. Furthermore, the possibility that alterations in feeding during infection may be beneficial to the host requires further study. We used the nematode Trichinella spiralis, which transiently inhabits the small intestine before migrating to skeletal muscle, as a biphasic model of infection to determine the cellular and molecular pathways controlling feeding during enteric and peripheral inflammation. Through the infection of genetically modified mice lacking cholecystokinin, Tumor necrosis factor α receptors and T and B-cells, we observed a biphasic hypophagic response to infection resulting from two separate immune-driven mechanisms. The enteroendocrine I-cell derived hormone cholecystokinin is an essential mediator of initial hypophagia and is induced by CD4+ T-cells during enteritis. In contrast, the second hypophagic response is extra-intestinal and due to the anorectic effects of TNFα during peripheral infection of the muscle. Moreover, via maintaining naive levels of the adipose secreted hormone leptin throughout infection we demonstrate a novel feedback loop in the immunoendocrine axis. Immune driven I-cell hyperplasia and resultant weight loss leads to a reduction in the inflammatory adipokine leptin, which in turn heightens protective immunity during infection. These results characterize specific immune mediated mechanisms which reduce feeding during intestinal or peripheral inflammation. Importantly, the molecular mediators of each phase are entirely separate. The data also introduce the first evidence that I-cell hyperplasia is an adaptively driven immune response that directly impinges on the outcome to infection.

  18. Early experiences mediate distinct adult gene expression and reproductive programs in Caenorhabditis elegans

    Science.gov (United States)

    Ow, Maria C.; Nichitean, Alexandra M.; Dorus, Steve; Hall, Sarah E.

    2018-01-01

    Environmental stress during early development in animals can have profound effects on adult phenotypes via programmed changes in gene expression. Using the nematode C. elegans, we demonstrated previously that adults retain a cellular memory of their developmental experience that is manifested by differences in gene expression and life history traits; however, the sophistication of this system in response to different environmental stresses, and how it dictates phenotypic plasticity in adults that contribute to increased fitness in response to distinct environmental challenges, was unknown. Using transcriptional profiling, we show here that C. elegans adults indeed retain distinct cellular memories of different environmental conditions. We identified approximately 500 genes in adults that entered dauer due to starvation that exhibit significant opposite (“seesaw”) transcriptional phenotypes compared to adults that entered dauer due to crowding, and are distinct from animals that bypassed dauer. Moreover, we show that two-thirds of the genes in the genome experience a 2-fold or greater seesaw trend in gene expression, and based upon the direction of change, are enriched in large, tightly linked regions on different chromosomes. Importantly, these transcriptional programs correspond to significant changes in brood size depending on the experienced stress. In addition, we demonstrate that while the observed seesaw gene expression changes occur in both somatic and germline tissue, only starvation-induced changes require a functional GLP-4 protein necessary for germline development, and both programs require the Argonaute CSR-1. Thus, our results suggest that signaling between the soma and the germ line can generate phenotypic plasticity as a result of early environmental experience, and likely contribute to increased fitness in adverse conditions and the evolution of the C. elegans genome. PMID:29447162

  19. Early experiences mediate distinct adult gene expression and reproductive programs in Caenorhabditis elegans.

    Directory of Open Access Journals (Sweden)

    Maria C Ow

    2018-02-01

    Full Text Available Environmental stress during early development in animals can have profound effects on adult phenotypes via programmed changes in gene expression. Using the nematode C. elegans, we demonstrated previously that adults retain a cellular memory of their developmental experience that is manifested by differences in gene expression and life history traits; however, the sophistication of this system in response to different environmental stresses, and how it dictates phenotypic plasticity in adults that contribute to increased fitness in response to distinct environmental challenges, was unknown. Using transcriptional profiling, we show here that C. elegans adults indeed retain distinct cellular memories of different environmental conditions. We identified approximately 500 genes in adults that entered dauer due to starvation that exhibit significant opposite ("seesaw" transcriptional phenotypes compared to adults that entered dauer due to crowding, and are distinct from animals that bypassed dauer. Moreover, we show that two-thirds of the genes in the genome experience a 2-fold or greater seesaw trend in gene expression, and based upon the direction of change, are enriched in large, tightly linked regions on different chromosomes. Importantly, these transcriptional programs correspond to significant changes in brood size depending on the experienced stress. In addition, we demonstrate that while the observed seesaw gene expression changes occur in both somatic and germline tissue, only starvation-induced changes require a functional GLP-4 protein necessary for germline development, and both programs require the Argonaute CSR-1. Thus, our results suggest that signaling between the soma and the germ line can generate phenotypic plasticity as a result of early environmental experience, and likely contribute to increased fitness in adverse conditions and the evolution of the C. elegans genome.

  20. A CCR2 macrophage endocytic pathway mediates extravascular fibrin clearance in vivo

    DEFF Research Database (Denmark)

    Motley, Michael P; Madsen, Daniel H; Jürgensen, Henrik J

    2016-01-01

    cellular endocytosis and lysosomal targeting, revealing a novel intracellular pathway for extravascular fibrin degradation. A C-C chemokine receptor type 2 (CCR2)-positive macrophage subpopulation constituted the majority of fibrin-uptaking cells. Consequently, cellular fibrin uptake was diminished...... by elimination of CCR2-expressing cells. The CCR2-positive macrophage subtype was different from collagen-internalizing M2-like macrophages. Cellular fibrin uptake was strictly dependent on plasminogen and plasminogen activator. Surprisingly, however, fibrin endocytosis was unimpeded by the absence of the fibrin...... subsets of macrophages employing distinct molecular pathways....

  1. Distinct muscarinic acetylcholine receptor subtypes mediate pre- and postsynaptic effects in rat neocortex

    Directory of Open Access Journals (Sweden)

    Gigout Sylvain

    2012-04-01

    Full Text Available Abstract Background Cholinergic transmission has been implicated in learning, memory and cognition. However, the cellular effects induced by muscarinic acetylcholine receptors (mAChRs activation are poorly understood in the neocortex. We investigated the effects of the cholinergic agonist carbachol (CCh and various agonists and antagonists on neuronal activity in rat neocortical slices using intracellular (sharp microelectrode and field potential recordings. Results CCh increased neuronal firing but reduced synaptic transmission. The increase of neuronal firing was antagonized by pirenzepine (M1/M4 mAChRs antagonist but not by AF-DX 116 (M2/M4 mAChRs antagonist. Pirenzepine reversed the depressant effect of CCh on excitatory postsynaptic potential (EPSP but had marginal effects when applied before CCh. AF-DX 116 antagonized the depression of EPSP when applied before or during CCh. CCh also decreased the paired-pulse inhibition of field potentials and the inhibitory conductances mediated by GABAA and GABAB receptors. The depression of paired-pulse inhibition was antagonized or prevented by AF-DX 116 or atropine but only marginally by pirenzepine. The inhibitory conductances were unaltered by xanomeline (M1/M4 mAChRs agonist, yet the CCh-induced depression was antagonized by AF-DX 116. Linopirdine, a selective M-current blocker, mimicked the effect of CCh on neuronal firing. However, linopirdine had no effect on the amplitude of EPSP or on the paired-pulse inhibition, indicating that M-current is involved in the increase of neuronal excitability but neither in the depression of EPSP nor paired-pulse inhibition. Conclusions These data indicate that the three effects are mediated by different mAChRs, the increase in firing being mediated by M1 mAChR, decrease of inhibition by M2 mAChR and depression of excitatory transmission by M4 mAChR. The depression of EPSP and increase of neuronal firing might enhance the signal-to-noise ratio, whereas the

  2. Transcription Profiles Reveal Sugar and Hormone Signaling Pathways Mediating Flower Induction in Apple (Malus domestica Borkh.).

    Science.gov (United States)

    Xing, Li-Bo; Zhang, Dong; Li, You-Mei; Shen, Ya-Wen; Zhao, Cai-Ping; Ma, Juan-Juan; An, Na; Han, Ming-Yu

    2015-10-01

    Flower induction in apple (Malus domestica Borkh.) is regulated by complex gene networks that involve multiple signal pathways to ensure flower bud formation in the next year, but the molecular determinants of apple flower induction are still unknown. In this research, transcriptomic profiles from differentiating buds allowed us to identify genes potentially involved in signaling pathways that mediate the regulatory mechanisms of flower induction. A hypothetical model for this regulatory mechanism was obtained by analysis of the available transcriptomic data, suggesting that sugar-, hormone- and flowering-related genes, as well as those involved in cell-cycle induction, participated in the apple flower induction process. Sugar levels and metabolism-related gene expression profiles revealed that sucrose is the initiation signal in flower induction. Complex hormone regulatory networks involved in cytokinin (CK), abscisic acid (ABA) and gibberellic acid pathways also induce apple flower formation. CK plays a key role in the regulation of cell formation and differentiation, and in affecting flowering-related gene expression levels during these processes. Meanwhile, ABA levels and ABA-related gene expression levels gradually increased, as did those of sugar metabolism-related genes, in developing buds, indicating that ABA signals regulate apple flower induction by participating in the sugar-mediated flowering pathway. Furthermore, changes in sugar and starch deposition levels in buds can be affected by ABA content and the expression of the genes involved in the ABA signaling pathway. Thus, multiple pathways, which are mainly mediated by crosstalk between sugar and hormone signals, regulate the molecular network involved in bud growth and flower induction in apple trees. © The Author 2015. Published by Oxford University Press on behalf of Japanese Society of Plant Physiologists.

  3. How does MBCT for depression work? studying cognitive and affective mediation pathways.

    Directory of Open Access Journals (Sweden)

    Tim Batink

    Full Text Available Mindfulness based cognitive therapy (MBCT is a non-pharmacological intervention to reduce current symptoms and to prevent recurrence of major depressive disorder. At present, it is not well understood which underlying mechanisms during MBCT are associated with its efficacy. The current study (n = 130 was designed to examine the roles of mindfulness skills, rumination, worry and affect, and the interplay between those factors, in the mechanisms of change in MBCT for residual depressive symptoms. An exploratory but systematic approach was chosen using Sobel-Goodman mediation analyses to identify mediators on the pathway from MBCT to reduction in depressive symptoms. We replicated earlier findings that therapeutic effects of MBCT are mediated by changes in mindfulness skills and worry. Second, results showed that changes in momentary positive and negative affect significantly mediated the efficacy of MBCT, and also mediated the effect of worry on depressive symptoms. Third, within the group of patients with a prior history of ≤ 2 episodes of MDD, predominantly changes in cognitive and to a lesser extent affective processes mediated the effect of MBCT. However, within the group of patients with a prior history of ≥ 3 episodes of MDD, only changes in affect were significant mediators for the effect of MBCT.[corrected] Nederlands Trial Register NTR1084.

  4. Distinct apoptotic blocks mediate resistance to panHER inhibitors in HER2+ breast cancer cells.

    Science.gov (United States)

    Karakas, Bahriye; Ozmay, Yeliz; Basaga, Huveyda; Gul, Ozgur; Kutuk, Ozgur

    2018-05-04

    Despite the development of novel targeted therapies, de novo or acquired chemoresistance remains a significant factor for treatment failure in breast cancer therapeutics. Neratinib and dacomitinib are irreversible panHER inhibitors, which block their autophosphorylation and downstream signaling. Moreover, neratinib and dacomitinib have been shown to activate cell death in HER2-overexpressing cell lines. Here we showed that increased MCL1 and decreased BIM and PUMA mediated resistance to neratinib in ZR-75-30 and SKBR3 cells while increased BCL-XL and BCL-2 and decreased BIM and PUMA promoted neratinib resistance in BT474 cells. Cells were also cross-resistant to dacomitinib. BH3 profiles of HER2+ breast cancer cells efficiently predicted antiapoptotic protein dependence and development of resistance to panHER inhibitors. Reactivation of ERK1/2 was primarily responsible for acquired resistance in SKBR3 and ZR-75-30 cells. Adding specific ERK1/2 inhibitor SCH772984 to neratinib or dacomitinib led to increased apoptotic response in neratinib-resistant SKBR3 and ZR-75-30 cells, but we did not detect a similar response in neratinib-resistant BT474 cells. Accordingly, suppression of BCL-2/BCL-XL by ABT-737 was required in addition to ERK1/2 inhibition for neratinib- or dacomitinib-induced apoptosis in neratinib-resistant BT474 cells. Our results showed that different mitochondrial apoptotic blocks mediated acquired panHER inhibitor resistance in HER2+ breast cancer cell lines as well as highlighted the potential of BH3 profiling assay in prediction of panHER inhibitor resistance in breast cancer cells. Copyright © 2018 Elsevier B.V. All rights reserved.

  5. Therapeutic implications of chemokine-mediated pathways in atherosclerosis: realistic perspectives and utopias.

    Science.gov (United States)

    Apostolakis, Stavros; Amanatidou, Virginia; Spandidos, Demetrios A

    2010-09-01

    Current perspectives on the pathogenesis of atherosclerosis strongly support the involvement of inflammatory mediators in the establishment and progression of atherosclerostic lesions. Chemokine-mediated mechanisms are potent regulators of such processes by orchestrating the interactions of inflammatory cellular components of the peripheral blood with cellular components of the arterial wall. The increasing evidence supporting the role of chemokine pathways in atherosclerosis renders chemokine ligands and their receptors potential therapeutic targets. In the following review, we aim to highlight the special structural and functional features of chemokines and their receptors in respect to their roles in atherosclerosis, and examine to what extent available data can be applied in disease management practices.

  6. Distinct Signaling Pathways After Higher or Lower Doses of Radiation in Three Closely Related Human Lymphoblast Cell Lines

    International Nuclear Information System (INIS)

    Lu, T.-P.; Lai, L.-C.; Lin, B.-I.; Chen, L.-H.; Hsiao, T.-H.; Liber, Howard L.; Cook, John A.; Mitchell, James B.; Tsai, M.-H.; Chuang, Eric Y.

    2010-01-01

    Purpose: The tumor suppressor p53 plays an essential role in cellular responses to DNA damage caused by ionizing radiation; therefore, this study aims to further explore the role that p53 plays at different doses of radiation. Materials and Methods: The global cellular responses to higher-dose (10 Gy) and lower dose (iso-survival dose, i.e., the respective D0 levels) radiation were analyzed using microarrays in three human lymphoblast cell lines with different p53 status: TK6 (wild-type p53), NH32 (p53-null), and WTK1 (mutant p53). Total RNAs were extracted from cells harvested at 0, 1, 3, 6, 9, and 24 h after higher and lower dose radiation exposures. Template-based clustering, hierarchical clustering, and principle component analysis were applied to examine the transcriptional profiles. Results: Differential expression profiles between 10 Gy and iso-survival radiation in cells with different p53 status were observed. Moreover, distinct gene expression patterns were exhibited among these three cells after 10 Gy radiation treatment, but similar transcriptional responses were observed in TK6 and NH32 cells treated with iso-survival radiation. Conclusions: After 10 Gy radiation exposure, the p53 signaling pathway played an important role in TK6, whereas the NFkB signaling pathway appeared to replace the role of p53 in WTK1. In contrast, after iso-survival radiation treatment, E2F4 seemed to play a dominant role independent of p53 status. This study dissected the impacts of p53, NFkB and E2F4 in response to higher or lower doses of γ-irradiation.

  7. Endothelial and Neuronal Nitric Oxide Activate Distinct Pathways on Sympathetic Neurotransmission in Rat Tail and Mesenteric Arteries.

    Directory of Open Access Journals (Sweden)

    Joana Beatriz Sousa

    Full Text Available Nitric oxide (NO seems to contribute to vascular homeostasis regulating neurotransmission. This work aimed at assessing the influence of NO from different sources and respective intracellular pathways on sympathetic neurotransmission, in two vascular beds. Electrically-evoked [3H]-noradrenaline release was assessed in rat mesenteric and tail arteries in the presence of NO donors or endothelial/neuronal nitric oxide synthase (NOS inhibitors. The influence of NO on adenosine-mediated effects was also studied using selective antagonists for adenosine receptors subtypes. Location of neuronal NOS (nNOS was investigated by immunohistochemistry (with specific antibodies for nNOS and for Schwann cells and Confocal Microscopy. Results indicated that: 1 in mesenteric arteries, noradrenaline release was reduced by NO donors and it was increased by nNOS inhibitors; the effect of NO donors was only abolished by the adenosine A1 receptors antagonist; 2 in tail arteries, noradrenaline release was increased by NO donors and it was reduced by eNOS inhibitors; adenosine receptors antagonists were devoid of effect; 3 confocal microscopy showed nNOS staining in adventitial cells, some co-localized with Schwann cells. nNOS staining and its co-localization with Schwann cells were significantly lower in tail compared to mesenteric arteries. In conclusion, in mesenteric arteries, nNOS, mainly located in Schwann cells, seems to be the main source of NO influencing perivascular sympathetic neurotransmission with an inhibitory effect, mediated by adenosine A1 receptors activation. Instead, in tail arteries endothelial NO seems to play a more relevant role and has a facilitatory effect, independent of adenosine receptors activation.

  8. Early induction of NRF2 antioxidant pathway by RHBDF2 mediates rapid cutaneous wound healing.

    Science.gov (United States)

    Hosur, Vishnu; Burzenski, Lisa M; Stearns, Timothy M; Farley, Michelle L; Sundberg, John P; Wiles, Michael V; Shultz, Leonard D

    2017-04-01

    Rhomboid family protein RHBDF2, an upstream regulator of the epidermal growth factor (EGF) receptor signaling, has been implicated in cutaneous wound healing. However, the underlying molecular mechanisms are still emerging. In humans, a gain-of-function mutation in the RHBDF2 gene accelerates cutaneous wound healing in an EGFR-dependent manner. Likewise, a gain-of-function mutation in the mouse Rhbdf2 gene (Rhbdf2 cub/cub ) shows a regenerative phenotype (rapid ear-hole closure) resulting from constitutive activation of the EGFR pathway. Because the RHBDF2-regulated EGFR pathway is relevant to cutaneous wound healing in humans, we used Rhbdf2 cub/cub mice to investigate the biological networks and pathways leading to accelerated ear-hole closure, with the goal of identifying therapeutic targets potentially effective in promoting wound healing in humans. Comparative transcriptome analysis of ear pinna tissue from Rhbdf2 cub/cub and Rhbdf2 +/+ mice at 0h, 15min, 2h, and 24h post-wounding revealed an early induction of the nuclear factor E2-related factor 2 (NRF2)-mediated anti-oxidative pathway (0h and 15min), followed by the integrin-receptor aggregation pathway (2h) as early-stage events immediately and shortly after wounding in Rhbdf2 cub/cub mice. Additionally, we observed genes enriched for the Fc fragment of the IgG receptor IIIa (FCGR3A)-mediated phagocytosis pathway 24h post-wounding. Although cutaneous wound repair in healthy individuals is generally non-problematic, it can be severely impaired due to aging, diabetes, and chronic inflammation. This study suggests that activation of the NRF2-antioxidant pathway by rhomboid protein RHBDF2 might be beneficial in treating chronic non-healing wounds. Copyright © 2017 Elsevier Inc. All rights reserved.

  9. Antibody-mediated neutralization of Ebola virus can occur by two distinct mechanisms

    International Nuclear Information System (INIS)

    Shedlock, Devon J.; Bailey, Michael A.; Popernack, Paul M.; Cunningham, James M.; Burton, Dennis R.; Sullivan, Nancy J.

    2010-01-01

    Human Ebola virus causes severe hemorrhagic fever disease with high mortality and there is no vaccine or treatment. Antibodies in survivors occur early, are sustained, and can delay infection when transferred into nonhuman primates. Monoclonal antibodies (mAbs) from survivors exhibit potent neutralizing activity in vitro and are protective in rodents. To better understand targets and mechanisms of neutralization, we investigated a panel of mAbs shown previously to react with the envelope glycoprotein (GP). While one non-neutralizing mAb recognized a GP epitope in the nonessential mucin-like domain, the rest were specific for GP1, were neutralizing, and could be further distinguished by reactivity with secreted GP. We show that survivor antibodies, human KZ52 and monkey JP3K11, were specific for conformation-dependent epitopes comprising residues in GP1 and GP2 and that neutralization occurred by two distinct mechanisms; KZ52 inhibited cathepsin cleavage of GP whereas JP3K11 recognized the cleaved, fusion-active form of GP.

  10. Evidence of Multiple Mediating Pathways in Associations Between Constructs of Stigma and Self-Reported Suicide Attempts in a Cross-Sectional Study of Gay and Bisexual Men.

    Science.gov (United States)

    Salway, Travis; Gesink, Dionne; Ibrahim, Selahadin; Ferlatte, Olivier; Rhodes, Anne E; Brennan, David J; Marchand, Rick; Trussler, Terry

    2018-05-01

    Gay and bisexual men (GBM) are more likely to attempt suicide than heterosexual men. This disparity is commonly interpreted using minority stress theory; however, specific pathways from antigay stigma to suicidal behavior are poorly understood. We aimed to estimate associations between multiple constructs of stigma and suicide attempts among adult GBM, and to measure the proportion of these associations mediated by distinct suicide risk factors, thus identifying proximal points of intervention. Data were drawn from a Canadian community-based survey of adult GBM. Structural equation modeling was used to compare associations between three latent constructs-enacted stigma (e.g., discrimination, harassment), anticipated prejudice (worry about encountering antigay/bisexual prejudice), and sexuality concealment-and self-reported suicide attempts (last 12 months). Coefficients were estimated for direct, indirect, and total pathways and evaluated based on magnitude and statistical significance. The proportion of associations mediated by depression, drug/alcohol use, and social isolation was calculated using indirect paths. Among 7872 respondents, 3.4% reported a suicide attempt in the past 12 months. The largest total association was observed for enacted stigma, and this association was partially mediated by depression and drug/alcohol use. The total association of anticipated prejudice was relatively smaller and mediated by depression and social isolation. Concealment had an inverse association with suicide attempts as mediated by depression but was also positively associated with suicide attempts when mediated through social isolation. Multiple constructs of antigay stigma were associated with suicide attempts; however, mediating pathways differed by construct, suggesting that a combination of strategies is required to prevent suicide in adult GBM.

  11. Diet-induced obesity mediated by the JNK/DIO2 signal transduction pathway

    Science.gov (United States)

    Vernia, Santiago; Cavanagh-Kyros, Julie; Barrett, Tamera; Jung, Dae Young; Kim, Jason K.; Davis, Roger J.

    2013-01-01

    The cJun N-terminal kinase (JNK) signaling pathway is a key mediator of metabolic stress responses caused by consuming a high-fat diet, including the development of obesity. To test the role of JNK, we examined diet-induced obesity in mice with targeted ablation of Jnk genes in the anterior pituitary gland. These mice exhibited an increase in the pituitary expression of thyroid-stimulating hormone (TSH), an increase in the blood concentration of thyroid hormone (T4), increased energy expenditure, and markedly reduced obesity compared with control mice. The increased amount of pituitary TSH was caused by reduced expression of type 2 iodothyronine deiodinase (Dio2), a gene that is required for T4-mediated negative feedback regulation of TSH expression. These data establish a molecular mechanism that accounts for the regulation of energy expenditure and the development of obesity by the JNK signaling pathway. PMID:24186979

  12. Heterotrimeric G protein-dependent WNT-5A signaling to ERK1/2 mediates distinct aspects of microglia proinflammatory transformation

    Directory of Open Access Journals (Sweden)

    Halleskog Carina

    2012-05-01

    Full Text Available Abstract Background WNT-5A signaling in the central nervous system is important for morphogenesis, neurogenesis and establishment of functional connectivity; the source of WNT-5A and its importance for cellular communication in the adult brain, however, are mainly unknown. We have previously investigated the inflammatory effects of WNT/β-catenin signaling in microglia in Alzheimer's disease. WNT-5A, however, generally recruits β-catenin-independent signaling. Thus, we aim here to characterize the role of WNT-5A and downstream signaling pathways for the inflammatory transformation of the brain's macrophages, the microglia. Methods Mouse brain sections were used for immunohistochemistry. Primary isolated microglia and astrocytes were employed to characterize the WNT-induced inflammatory transformation and underlying intracellular signaling pathways by immunoblotting, quantitative mRNA analysis, proliferation and invasion assays. Further, measurements of G protein activation by [γ-35 S]GTP binding, examination of calcium fluxes and cyclic AMP production were used to define intracellular signaling pathways. Results Astrocytes in the adult mouse brain express high levels of WNT-5A, which could serve as a novel astroglia-microglia communication pathway. The WNT-5A-induced proinflammatory microglia response is characterized by increased expression of inducible nitric oxide synthase, cyclooxygenase-2, cytokines, chemokines, enhanced invasive capacity and proliferation. Mapping of intracellular transduction pathways reveals that WNT-5A activates heterotrimeric Gi/o proteins to reduce cyclic AMP levels and to activate a Gi/o protein/phospholipase C/calcium-dependent protein kinase/extracellular signal-regulated kinase 1/2 (ERK1/2 axis. We show further that WNT-5A-induced ERK1/2 signaling is responsible for distinct aspects of the proinflammatory transformation, such as matrix metalloprotease 9/13 expression, invasion and proliferation. Conclusions

  13. Multiple repair pathways mediate tolerance to chemotherapeutic cross-linking agents in vertebrate cells.

    Science.gov (United States)

    Nojima, Kuniharu; Hochegger, Helfrid; Saberi, Alihossein; Fukushima, Toru; Kikuchi, Koji; Yoshimura, Michio; Orelli, Brian J; Bishop, Douglas K; Hirano, Seiki; Ohzeki, Mioko; Ishiai, Masamichi; Yamamoto, Kazuhiko; Takata, Minoru; Arakawa, Hiroshi; Buerstedde, Jean-Marie; Yamazoe, Mitsuyoshi; Kawamoto, Takuo; Araki, Kasumi; Takahashi, Jun A; Hashimoto, Nobuo; Takeda, Shunichi; Sonoda, Eiichiro

    2005-12-15

    Cross-linking agents that induce DNA interstrand cross-links (ICL) are widely used in anticancer chemotherapy. Yeast genetic studies show that nucleotide excision repair (NER), Rad6/Rad18-dependent postreplication repair, homologous recombination, and cell cycle checkpoint pathway are involved in ICL repair. To study the contribution of DNA damage response pathways in tolerance to cross-linking agents in vertebrates, we made a panel of gene-disrupted clones from chicken DT40 cells, each defective in a particular DNA repair or checkpoint pathway, and measured the sensitivities to cross-linking agents, including cis-diamminedichloroplatinum (II) (cisplatin), mitomycin C, and melphalan. We found that cells harboring defects in translesion DNA synthesis (TLS), Fanconi anemia complementation groups (FANC), or homologous recombination displayed marked hypersensitivity to all the cross-linking agents, whereas NER seemed to play only a minor role. This effect of replication-dependent repair pathways is distinctively different from the situation in yeast, where NER seems to play a major role in dealing with ICL. Cells deficient in Rev3, the catalytic subunit of TLS polymerase Polzeta, showed the highest sensitivity to cisplatin followed by fanc-c. Furthermore, epistasis analysis revealed that these two mutants work in the same pathway. Our genetic comprehensive study reveals a critical role for DNA repair pathways that release DNA replication block at ICLs in cellular tolerance to cross-linking agents and could be directly exploited in designing an effective chemotherapy.

  14. Natural products induce a G protein-mediated calcium pathway activating p53 in cancer cells

    Energy Technology Data Exchange (ETDEWEB)

    Ginkel, Paul R. van; Yan, Michael B. [UW Carbone Cancer Center, University of Wisconsin, Madison, WI 53792 (United States); Department of Ophthalmology and Visual Sciences, University of Wisconsin, Madison, WI 53792 (United States); Bhattacharya, Saswati [UW Carbone Cancer Center, University of Wisconsin, Madison, WI 53792 (United States); Department of Ophthalmology and Visual Sciences, University of Wisconsin, Madison, WI 53792 (United States); Department of Pediatrics, University of Wisconsin, Madison, WI 53792 (United States); Polans, Arthur S., E-mail: aspolans@wisc.edu [UW Carbone Cancer Center, University of Wisconsin, Madison, WI 53792 (United States); Department of Ophthalmology and Visual Sciences, University of Wisconsin, Madison, WI 53792 (United States); Kenealey, Jason D. [UW Carbone Cancer Center, University of Wisconsin, Madison, WI 53792 (United States); Department of Ophthalmology and Visual Sciences, University of Wisconsin, Madison, WI 53792 (United States); Department of Nutrition, Dietetics and Food Science, Brigham Young University, Provo, UT 84602 (United States)

    2015-11-01

    Paclitaxel, etoposide, vincristine and doxorubicin are examples of natural products being used as chemotherapeutics but with adverse side effects that limit their therapeutic window. Natural products derived from plants and having low toxicity, such as quercetin, resveratrol, epigallocatechin gallate and piceatannol, have been shown to inhibit tumor cell growth both in vitro and in pre-clinical models of cancer, but their mechanisms of action have not been fully elucidated, thus restricting their use as prototypes for developing synthetic analogs with improved anti-cancer properties. We and others have demonstrated that one of the earliest and consistent events upon exposure of tumor cells to these less toxic natural products is a rise in cytoplasmic calcium, activating several pro-apoptotic pathways. We describe here a G protein/inositol 1,4,5-trisphosphate pathway (InsP3) in MDA-MB-231 human breast cancer cells that mediates between these less toxic natural products and the release of calcium from the endoplasmic reticulum. Further, we demonstrate that this elevation of intracellular calcium modulates p53 activity and the subsequent transcription of several pro-apoptotic genes encoding PIG8, CD95, PIDD, TP53INP, RRM2B, Noxa, p21 and PUMA. We conclude from our findings that less toxic natural products likely bind to a G protein coupled receptor that activates a G protein-mediated and calcium-dependent pathway resulting selectively in tumor cell death. - Highlights: • Natural products having low toxicity increase cytoplasmic calcium in cancer cells. • A G-protein/IP{sub 3} pathway mediates the release of calcium from the ER. • The elevation of intracellular calcium modulates p53 activity. • p53 and other Ca{sup 2+}-dependent pro-apoptotic pathways inhibit cancer cell growth.

  15. Simultaneous modulation of the intrinsic and extrinsic pathways by simvastatin in mediating prostate cancer cell apoptosis

    International Nuclear Information System (INIS)

    Goc, Anna; Kochuparambil, Samith T; Al-Husein, Belal; Al-Azayzih, Ahmad; Mohammad, Shuaib; Somanath, Payaningal R

    2012-01-01

    Recent studies suggest the potential benefits of statins as anti-cancer agents. Mechanisms by which statins induce apoptosis in cancer cells are not clear. We previously showed that simvastatin inhibit prostate cancer cell functions and tumor growth. Molecular mechanisms by which simvastatin induce apoptosis in prostate cancer cells is not completely understood. Effect of simvastatin on PC3 cell apoptosis was compared with docetaxel using apoptosis, TUNEL and trypan blue viability assays. Protein expression of major candidates of the intrinsic pathway downstream of simvastatin-mediated Akt inactivation was analyzed. Gene arrays and western analysis of PC3 cells and tumor lysates were performed to identify the candidate genes mediating extrinsic apoptosis pathway by simvastatin. Data indicated that simvastatin inhibited intrinsic cell survival pathway in PC3 cells by enhancing phosphorylation of Bad, reducing the protein expression of Bcl-2, Bcl-xL and cleaved caspases 9/3. Over-expression of PC3 cells with Bcl-2 or DN-caspase 9 did not rescue the simvastatin-induced apoptosis. Simvastatin treatment resulted in increased mRNA and protein expression of molecules such as TNF, Fas-L, Traf1 and cleaved caspase 8, major mediators of intrinsic apoptosis pathway and reduced protein levels of pro-survival genes Lhx4 and Nme5. Our study provides the first report that simvastatin simultaneously modulates intrinsic and extrinsic pathways in the regulation of prostate cancer cell apoptosis in vitro and in vivo, and render reasonable optimism that statins could become an attractive anti-cancer agent

  16. Natural products induce a G protein-mediated calcium pathway activating p53 in cancer cells

    International Nuclear Information System (INIS)

    Ginkel, Paul R. van; Yan, Michael B.; Bhattacharya, Saswati; Polans, Arthur S.; Kenealey, Jason D.

    2015-01-01

    Paclitaxel, etoposide, vincristine and doxorubicin are examples of natural products being used as chemotherapeutics but with adverse side effects that limit their therapeutic window. Natural products derived from plants and having low toxicity, such as quercetin, resveratrol, epigallocatechin gallate and piceatannol, have been shown to inhibit tumor cell growth both in vitro and in pre-clinical models of cancer, but their mechanisms of action have not been fully elucidated, thus restricting their use as prototypes for developing synthetic analogs with improved anti-cancer properties. We and others have demonstrated that one of the earliest and consistent events upon exposure of tumor cells to these less toxic natural products is a rise in cytoplasmic calcium, activating several pro-apoptotic pathways. We describe here a G protein/inositol 1,4,5-trisphosphate pathway (InsP3) in MDA-MB-231 human breast cancer cells that mediates between these less toxic natural products and the release of calcium from the endoplasmic reticulum. Further, we demonstrate that this elevation of intracellular calcium modulates p53 activity and the subsequent transcription of several pro-apoptotic genes encoding PIG8, CD95, PIDD, TP53INP, RRM2B, Noxa, p21 and PUMA. We conclude from our findings that less toxic natural products likely bind to a G protein coupled receptor that activates a G protein-mediated and calcium-dependent pathway resulting selectively in tumor cell death. - Highlights: • Natural products having low toxicity increase cytoplasmic calcium in cancer cells. • A G-protein/IP 3 pathway mediates the release of calcium from the ER. • The elevation of intracellular calcium modulates p53 activity. • p53 and other Ca 2+ -dependent pro-apoptotic pathways inhibit cancer cell growth.

  17. Structural plasticity mediates distinct GAP-dependent GTP hydrolysis mechanisms in Rab33 and Rab5.

    Science.gov (United States)

    Majumdar, Soneya; Acharya, Abhishek; Prakash, Balaji

    2017-12-01

    The classical GTP hydrolysis mechanism, as seen in Ras, employs a catalytic glutamine provided in cis by the GTPase and an arginine supplied in trans by a GTPase activating protein (GAP). The key idea emergent from a large body of research on small GTPases is that GTPases employ a variety of different hydrolysis mechanisms; evidently, these variations permit diverse rates of GTPase inactivation, crucial for temporal regulation of different biological processes. Recently, we unified these variations and argued that a steric clash between active site residues (corresponding to positions 12 and 61 of Ras) governs whether a GTPase utilizes the cis-Gln or the trans-Gln (from the GAP) for catalysis. As the cis-Gln encounters a steric clash, the Rab GTPases employ the so-called dual finger mechanism where the interacting GAP supplies a trans-Gln for catalysis. Using experimental and computational methods, we demonstrate how the cis-Gln of Rab33 overcomes the steric clash when it is stabilized by a residue in the vicinity. In effect, this demonstrates how both cis-Gln- and trans-Gln-mediated mechanisms could operate in the same GTPase in different contexts, i.e. depending on the GAP that regulates its action. Interestingly, in the case of Rab5, which possesses a higher intrinsic GTP hydrolysis rate, a similar stabilization of the cis-Gln appears to overcome the steric clash. Taken together with the mechanisms seen for Rab1, it is evident that the observed variations in Rab and their GAP partners allow structural plasticity, or in other words, the choice of different catalytic mechanisms. © 2017 Federation of European Biochemical Societies.

  18. Potential mediating pathways through which sports participation relates to reduced risk of suicidal ideation.

    Science.gov (United States)

    Taliaferro, Lindsay A; Rienzo, Barbara A; Miller, M David; Pigg, R Morgan; Dodd, Virginia J

    2010-09-01

    Suicide ranks as the third leading cause of death for American youth. Researchers examining sport participation and suicidal behavior have regularly found inverse relationships. This study represents the first effort to test a model depicting potential mechanisms through which sport participation relates to reduced risk of suicidal ideation. The participants were 450 undergraduate students. Measures assessed participants' involvement in university-run sports and other activities; frequency of physical activity; and perceived social support, self-esteem, depression, hopelessness, loneliness, and suicidal ideation. Regression analyses confirmed a path model and tested for mediation effects. Vigorous activity mediated relationships between sport participation and self-esteem and depression; and self-esteem and depression mediated the relationship between vigorous activity and suicidal ideation. Social support mediated relationships between sport participation and depression, hopelessness, and loneliness; and each of these risk factors partially mediated the relationship between social support and suicidal ideation. However no variable fully mediated the relationship between sport participation and suicidal ideation. This study provides a foundation for research designed to examine pathways through which sport participation relates to reduced risk of suicidal behavior.

  19. Naja nigricollis CMS-9 enhances the mitochondria-mediated death pathway in adaphostin-treated human leukaemia U937 cells.

    Science.gov (United States)

    Chen, Ying-Jung; Wang, Jeh-Jeng; Chang, Long-Sen

    2011-11-01

    1. The aim of the present study was to explore the effect of the Naja nigricollis phospholipase A(2) CMS-9 on adaphostin-induced death of human leukaemia U937 cells. 2. Leukaemia U937 cells (Bcr/Abl-negative cells) were treated with adaphostin (0-10 μmol/L) and CMS-9 (0-1 μmol/L). The effects of CMS-9, adaphostin and their combination on cell viability, the generation reactive oxygen species (ROS), [Ca(2+) ](i) , p38 mitogen-activated protein kinase (MAPK) activation, Akt and extracellular signal-regulated kinase (ERK) inactivation, mitochondrial membrane potential (ΔΨ(m) ) and Bcl-2 family proteins were analysed. 3. Both adaphostin and CMS-9 induced U937 cell apoptosis, characterized by dissipation of ΔΨ(m) and ROS generation. Combined treatment further increased ΔΨ(m) loss and reduced the viability of adaphostin-treated cells. Unlike in CMS-9-treated cells, in adaphostin-treated cells ROS-induced increases in [Ca(2+) ](i) were observed. CMS-9-induced ROS generation resulted in p38 MAPK activation, whereas adaphostin treatment elicited ROS/Ca(2+) -mediated inactivation of Akt and ERK. Moreover, Akt was found to be involved in ERK phosphorylation. Suppression of p38 MAPK activation blocked CMS-9-induced ΔΨ(m) loss and Bcl-xL downregulation. Overexpression of constitutively active Akt and mitogen-activated protein kinase kinase (MEK) 1 rescued adaphostin-induced ΔΨ(m) loss and Bcl-2 downregulation. Similarly, CMS-9 augmented adaphostin toxicity in human leukaemia K562 cells via increased mitochondrial alterations. 4. The results suggest that two distinct pathways mediate adaphostin- and CMS-9-induced mitochondrial damage (i.e. the ROS-Ca(2+) -Akt-ERK and ROS-p38 MAPK pathways, respectively). These distinct pathway explain the augmentation by CMS-9 of ΔΨ(m) loss and apoptosis in adaphostin-treated U937 cells. © 2011 The Authors. Clinical and Experimental Pharmacology and Physiology © 2011 Blackwell Publishing Asia Pty Ltd.

  20. The role of the TOR pathway in mediating the link between nutrition and longevity.

    Science.gov (United States)

    Lushchak, Oleh; Strilbytska, Olha; Piskovatska, Veronika; Storey, Kenneth B; Koliada, Alexander; Vaiserman, Alexander

    2017-06-01

    The target of rapamycin (TOR) pathway integrates signals from extracellular and intracellular agents, such as growth factors, nutrients, mediators of energy balance, oxygen availability and other environmental cues. It allows the regulation of multiple cellular processes including protein and lipid synthesis, ribosome biogenesis, autophagy and metabolic processes. Being conserved across different phyla, TOR regulates longevity of various organisms in response to dietary conditions. In this review we described the main components of the TOR pathway and its upstream effectors and downstream processes in relation to aging. The potential contribution of the TOR pathway in lifespan-extending effects of varied dietary interventions, and the anti-aging drugs rapamycin and metformin direct or indirect regulation of TOR activity in yeasts, worms, flies and mammals are also discussed. Copyright © 2017 Elsevier B.V. All rights reserved.

  1. Nucleolus-derived mediators in oncogenic stress response and activation of p53-dependent pathways.

    Science.gov (United States)

    Stępiński, Dariusz

    2016-08-01

    Rapid growth and division of cells, including tumor ones, is correlated with intensive protein biosynthesis. The output of nucleoli, organelles where translational machineries are formed, depends on a rate of particular stages of ribosome production and on accessibility of elements crucial for their effective functioning, including substrates, enzymes as well as energy resources. Different factors that induce cellular stress also often lead to nucleolar dysfunction which results in ribosome biogenesis impairment. Such nucleolar disorders, called nucleolar or ribosomal stress, usually affect cellular functioning which in fact is a result of p53-dependent pathway activation, elicited as a response to stress. These pathways direct cells to new destinations such as cell cycle arrest, damage repair, differentiation, autophagy, programmed cell death or aging. In the case of impaired nucleolar functioning, nucleolar and ribosomal proteins mediate activation of the p53 pathways. They are also triggered as a response to oncogenic factor overexpression to protect tissues and organs against extensive proliferation of abnormal cells. Intentional impairment of any step of ribosome biosynthesis which would direct the cells to these destinations could be a strategy used in anticancer therapy. This review presents current knowledge on a nucleolus, mainly in relation to cancer biology, which is an important and extremely sensitive element of the mechanism participating in cellular stress reaction mediating activation of the p53 pathways in order to counteract stress effects, especially cancer development.

  2. Pathway Linking Internet Health Information Seeking to Better Health: A Moderated Mediation Study.

    Science.gov (United States)

    Jiang, Shaohai; Street, Richard L

    2017-08-01

    The Internet increasingly has been recognized as an important medium with respect to population health. However, little is known about the mechanisms that underlie the potential impact of health-related Internet use on health outcomes. Based on the three-stage model of health promotion using interactive media, this study empirically tested a moderated mediation pathway model. Results showed that the effect of Internet health information seeking on three health outcomes (general, emotional, and physical) was completely mediated by respondents' access to social support resources. In addition, users' online health information seeking experience positively moderated this mediation path. The findings have significant theoretical and practical implications for the design of Internet-based health promotion resources to improve health outcomes.

  3. Ficolin-3-mediated lectin complement pathway activation in patients with subarachnoid hemorrhage

    DEFF Research Database (Denmark)

    Zanier, Elisa R; Zangari, Rosalia; Munthe-Fog, Lea

    2014-01-01

    OBJECTIVES: To assess the involvement of ficolin-3, the main initiator of the lectin complement pathway (LCP), in subarachnoid hemorrhage (SAH) pathology and outcome. METHODS: In this preliminary exploratory study, plasma concentration of ficolin-3 and of ficolin-3-mediated functional LCP activity...... the World Federation of Neurosurgical Societies grading scale; vasospasm, defined as neuro-worsening with angiographic confirmation of vessel narrowing; cerebral ischemia, defined as hypodense lesion on CT scan performed before discharge; and 6-month outcome, assessed using the Glasgow Outcome Scale....... RESULTS: In patients, no changes were detected for ficolin-3 compared with controls. Notably, however, ficolin-3-mediated functional LCP activity was reduced. Low levels of plasma ficolin-3 and ficolin-3-mediated functional LCP activity were related to SAH severity, vasospasm, and cerebral ischemia...

  4. Rme1 is necessary for Mi-1-mediated resistance and acts early in the resistance pathway.

    Science.gov (United States)

    Martinez de Ilarduya, Oscar; Nombela, Gloria; Hwang, Chin-Feng; Williamson, Valerie M; Muñiz, Mariano; Kaloshian, Isgouhi

    2004-01-01

    The tomato gene Mi-1 confers resistance to root-knot nematodes (Meloidogyne spp.), potato aphid, and whitefly. Using genetic screens, we have isolated a mutant, rme1 (resistance to Meloidogyne spp.), compromised in resistance to M. javanica and potato aphid. Here, we show that the rme1 mutant is also compromised in resistance to M. incognita, M. arenaria, and whitefly. In addition, using an Agrobacterium-mediated transient assay in leaves to express constitutive gain-of-function mutant Pto(L205D), we demonstrated that the rme1 mutation is not compromised in Pto-mediated hypersensitive response. Moreover, the mutation in rme1 does not result in increased virulence of pathogenic Pseudomonas syringae or Mi-1-virulent M. incognita. Using a chimeric Mi-1 construct, Mi-DS4, which confers constitutive cell death phenotype and A. rhizogenes root transformation, we showed that the Mi-1-mediated cell death pathway is intact in this mutant. Our results indicate that Rme1 is required for Mi-1-mediated resistance and acts either at the same step in the signal transduction pathway as Mi-1 or upstream of Mi-1.

  5. Dihydroartemisinin induces apoptosis preferentially via a Bim-mediated intrinsic pathway in hepatocarcinoma cells.

    Science.gov (United States)

    Qin, Guiqi; Zhao, ChuBiao; Zhang, Lili; Liu, Hongyu; Quan, Yingyao; Chai, Liuying; Wu, Shengnan; Wang, Xiaoping; Chen, Tongsheng

    2015-08-01

    This report is designed to dissect the detail molecular mechanism by which dihydroartemisinin (DHA), a derivative of artemisinin, induces apoptosis in human hepatocellular carcinoma (HCC) cells. DHA induced a loss of the mitochondrial transmemberane potential (ΔΨm), release of cytochrome c, activation of caspases, and externalization of phosphatidylserine indicative of apoptosis induction. Compared with the modest inhibitory effects of silencing Bax, silencing Bak largely prevented DHA-induced ΔΨm collapse and apoptosis though DHA induced a commensurable activation of Bax and Bak, demonstrating a key role of the Bak-mediated intrinsic apoptosis pathway. DHA did not induce Bid cleavage and translocation from cytoplasm to mitochondria and had little effects on the expressions of Puma and Noxa, but did increase Bim and Bak expressions and decrease Mcl-1 expression. Furthermore, the cytotoxicity of DHA was remarkably reduced by silencing Bim, and modestly but significantly reduced by silencing Puma or Noxa. Silencing Bim or Noxa preferentially reduced DHA-induced Bak activation, while silencing Puma preferentially reduced DHA-induced Bax activation, demonstrating that Bim and to a lesser extent Noxa act as upstream mediators to trigger the Bak-mediated intrinsic apoptosis pathway. In addition, silencing Mcl-1 enhanced DHA-induced Bak activation and apoptosis. Taken together, our data demonstrate a crucial role of Bim in preferentially regulating the Bak/Mcl-1 rheostat to mediate DHA-induced apoptosis in HCC cells.

  6. Smad, but not MAPK, pathway mediates the expression of type I collagen in radiation induced fibrosis

    International Nuclear Information System (INIS)

    Yano, Hiroyuki; Hamanaka, Ryoji; Nakamura, Miki; Sumiyoshi, Hideaki; Matsuo, Noritaka; Yoshioka, Hidekatsu

    2012-01-01

    Highlights: ► We examine how radiation affects the expression level and signal pathway of collagen. ► TGF-β1 mRNA is elevated earlier than those of collagen genes after irradiation. ► Smad pathway mediates the expression of collagen in radiation induced fibrosis. ► MAPK pathways are not affected in the expression of collagen after irradiation. -- Abstract: Radiation induced fibrosis occurs following a therapeutic or accidental radiation exposure in normal tissues. Tissue fibrosis is the excessive accumulation of collagen and other extracellular matrix components. This study investigated how ionizing radiation affects the expression level and signal pathway of type I collagen. Real time RT-RCR showed that both α1and α2 chain of type I collagen mRNA were elevated from 48 h after irradiation with 10 Gy in NIH3T3 cells. The relative luciferase activities of both genes and type I collagen marker were elevated at 72 h. TGF-β1 mRNA was elevated earlier than those of type I collagen genes. A Western blot analysis showed the elevation of Smad phosphorylation at 72 h. Conversely, treatment with TGF-β receptor inhibitor inhibited the mRNA and relative luciferase activity of type I collagen. The phosphorylation of Smad was repressed with the inhibitor, and the luciferase activity was cancelled using a mutant construct of Smad binding site of α2(I) collagen gene. However, the MAPK pathways, p38, ERK1/2 and JNK, were not affected with specific inhibitors or siRNA. The data showed that the Smad pathway mediated the expression of type I collagen in radiation induced fibrosis.

  7. Smad, but not MAPK, pathway mediates the expression of type I collagen in radiation induced fibrosis

    Energy Technology Data Exchange (ETDEWEB)

    Yano, Hiroyuki [Department of Matrix Medicine, Oita University, 1-1 Idaigaoka Hasama-machi, Yufu, Oita 879-5593 (Japan); Division of Radioisotope Research, Department of Research Support, Research Promotion Project, Oita University, 1-1 Idaigaoka Hasama-machi, Yufu, Oita 879-5593 (Japan); Hamanaka, Ryoji; Nakamura, Miki [Cell Biology, Faculty of Medicine, Oita University, 1-1 Idaigaoka Hasama-machi, Yufu, Oita 879-5593 (Japan); Sumiyoshi, Hideaki; Matsuo, Noritaka [Department of Matrix Medicine, Oita University, 1-1 Idaigaoka Hasama-machi, Yufu, Oita 879-5593 (Japan); Yoshioka, Hidekatsu, E-mail: hidey@oita-u.ac.jp [Department of Matrix Medicine, Oita University, 1-1 Idaigaoka Hasama-machi, Yufu, Oita 879-5593 (Japan)

    2012-02-17

    Highlights: Black-Right-Pointing-Pointer We examine how radiation affects the expression level and signal pathway of collagen. Black-Right-Pointing-Pointer TGF-{beta}1 mRNA is elevated earlier than those of collagen genes after irradiation. Black-Right-Pointing-Pointer Smad pathway mediates the expression of collagen in radiation induced fibrosis. Black-Right-Pointing-Pointer MAPK pathways are not affected in the expression of collagen after irradiation. -- Abstract: Radiation induced fibrosis occurs following a therapeutic or accidental radiation exposure in normal tissues. Tissue fibrosis is the excessive accumulation of collagen and other extracellular matrix components. This study investigated how ionizing radiation affects the expression level and signal pathway of type I collagen. Real time RT-RCR showed that both {alpha}1and {alpha}2 chain of type I collagen mRNA were elevated from 48 h after irradiation with 10 Gy in NIH3T3 cells. The relative luciferase activities of both genes and type I collagen marker were elevated at 72 h. TGF-{beta}1 mRNA was elevated earlier than those of type I collagen genes. A Western blot analysis showed the elevation of Smad phosphorylation at 72 h. Conversely, treatment with TGF-{beta} receptor inhibitor inhibited the mRNA and relative luciferase activity of type I collagen. The phosphorylation of Smad was repressed with the inhibitor, and the luciferase activity was cancelled using a mutant construct of Smad binding site of {alpha}2(I) collagen gene. However, the MAPK pathways, p38, ERK1/2 and JNK, were not affected with specific inhibitors or siRNA. The data showed that the Smad pathway mediated the expression of type I collagen in radiation induced fibrosis.

  8. Distinct and shared cognitive functions mediate event- and time-based prospective memory impairment in normal ageing

    Science.gov (United States)

    Gonneaud, Julie; Kalpouzos, Grégoria; Bon, Laetitia; Viader, Fausto; Eustache, Francis; Desgranges, Béatrice

    2011-01-01

    Prospective memory (PM) is the ability to remember to perform an action at a specific point in the future. Regarded as multidimensional, PM involves several cognitive functions that are known to be impaired in normal aging. In the present study, we set out to investigate the cognitive correlates of PM impairment in normal aging. Manipulating cognitive load, we assessed event- and time-based PM, as well as several cognitive functions, including executive functions, working memory and retrospective episodic memory, in healthy subjects covering the entire adulthood. We found that normal aging was characterized by PM decline in all conditions and that event-based PM was more sensitive to the effects of aging than time-based PM. Whatever the conditions, PM was linked to inhibition and processing speed. However, while event-based PM was mainly mediated by binding and retrospective memory processes, time-based PM was mainly related to inhibition. The only distinction between high- and low-load PM cognitive correlates lays in an additional, but marginal, correlation between updating and the high-load PM condition. The association of distinct cognitive functions, as well as shared mechanisms with event- and time-based PM confirms that each type of PM relies on a different set of processes. PMID:21678154

  9. Acetylation within the N- and C-Terminal Domains of Src Regulates Distinct Roles of STAT3-Mediated Tumorigenesis.

    Science.gov (United States)

    Huang, Chao; Zhang, Zhe; Chen, Lihan; Lee, Hank W; Ayrapetov, Marina K; Zhao, Ting C; Hao, Yimei; Gao, Jinsong; Yang, Chunzhang; Mehta, Gautam U; Zhuang, Zhengping; Zhang, Xiaoren; Hu, Guohong; Chin, Y Eugene

    2018-06-01

    Posttranslational modifications of mammalian c-Src N-terminal and C-terminal domains regulate distinct functions. Myristoylation of G 2 controls its cell membrane association and phosphorylation of Y419/Y527 controls its activation or inactivation, respectively. We provide evidence that Src-cell membrane association-dissociation and catalytic activation-inactivation are both regulated by acetylation. In EGF-treated cells, CREB binding protein (CBP) acetylates an N-terminal lysine cluster (K5, K7, and K9) of c-Src to promote dissociation from the cell membrane. CBP also acetylates the C-terminal K401, K423, and K427 of c-Src to activate intrinsic kinase activity for STAT3 recruitment and activation. N-terminal domain phosphorylation (Y14, Y45, and Y68) of STAT3 by c-Src activates transcriptionally active dimers of STAT3. Moreover, acetyl-Src translocates into nuclei, where it forms the Src-STAT3 enhanceosome for gene regulation and cancer cell proliferation. Thus, c-Src acetylation in the N-terminal and C-terminal domains play distinct roles in Src activity and regulation. Significance: CBP-mediated acetylation of lysine clusters in both the N-terminal and C-terminal regions of c-Src provides additional levels of control over STAT3 transcriptional activity. Cancer Res; 78(11); 2825-38. ©2018 AACR . ©2018 American Association for Cancer Research.

  10. miR2Pathway: A Novel Analytical Method to Discover MicroRNA-mediated Dysregulated Pathways Involved in Hepatocellular Carcinoma.

    Science.gov (United States)

    Li, Chaoxing; Dinu, Valentin

    2018-03-22

    MicroRNAs (miRNAs) are small, non-coding RNAs involved in the regulation of gene expression at a post-transcriptional level. Recent studies have shown miRNAs as key regulators of a variety of biological processes, such as proliferation, differentiation, apoptosis, metabolism, etc. Aberrantly expressed miRNAs influence individual gene expression level, but rewired miRNA-mRNA connections can influence the activity of biological pathways. Here, we define rewired miRNA-mRNA connections as the differential (rewiring) effects on the activity of biological pathways between hepatocellular carcinoma (HCC) and normal phenotypes. Our work presented here uses a PageRank-based approach to measure the degree of miRNA-mediated dysregulation of biological pathways between HCC and normal samples based on rewired miRNA-mRNA connections. In our study, we regard the degree of miRNA-mediated dysregulation of biological pathways as disease risk of biological pathways. Therefore, we propose a new method, miR2Pathway, to measure and rank the degree of miRNA-mediated dysregulation of biological pathways by measuring the total differential influence of miRNAs on the activity of pathways between HCC and normal states. miR2Pathway proposed here systematically shows the first evidence for a mechanism of biological pathways being dysregulated by rewired miRNA-mRNA connections, and provides new insight into exploring mechanisms behind HCC. Thus, miR2Pathway is a novel method to identify and rank miRNA-dysregulated pathways in HCC. Copyright © 2018. Published by Elsevier Inc.

  11. ER-associated SNAREs and Sey1p mediate nuclear fusion at two distinct steps during yeast mating.

    Science.gov (United States)

    Rogers, Jason V; Arlow, Tim; Inkellis, Elizabeth R; Koo, Timothy S; Rose, Mark D

    2013-12-01

    During yeast mating, two haploid nuclei fuse membranes to form a single diploid nucleus. However, the known proteins required for nuclear fusion are unlikely to function as direct fusogens (i.e., they are unlikely to directly catalyze lipid bilayer fusion) based on their predicted structure and localization. Therefore we screened known fusogens from vesicle trafficking (soluble N-ethylmaleimide-sensitive factor attachment protein receptors [SNAREs]) and homotypic endoplasmic reticulum (ER) fusion (Sey1p) for additional roles in nuclear fusion. Here we demonstrate that the ER-localized SNAREs Sec20p, Ufe1p, Use1p, and Bos1p are required for efficient nuclear fusion. In contrast, Sey1p is required indirectly for nuclear fusion; sey1Δ zygotes accumulate ER at the zone of cell fusion, causing a block in nuclear congression. However, double mutants of Sey1p and Sec20p, Ufe1p, or Use1p, but not Bos1p, display extreme ER morphology defects, worse than either single mutant, suggesting that retrograde SNAREs fuse ER in the absence of Sey1p. Together these data demonstrate that SNAREs mediate nuclear fusion, ER fusion after cell fusion is necessary to complete nuclear congression, and there exists a SNARE-mediated, Sey1p-independent ER fusion pathway.

  12. Two distinct mTORC2-dependent pathways converge on Rac1 to drive breast cancer metastasis.

    Science.gov (United States)

    Morrison Joly, Meghan; Williams, Michelle M; Hicks, Donna J; Jones, Bayley; Sanchez, Violeta; Young, Christian D; Sarbassov, Dos D; Muller, William J; Brantley-Sieders, Dana; Cook, Rebecca S

    2017-06-30

    increased Rac1 activity through the Rac-GEF Tiam1, thus partially rescuing cell invasion/motility. The mTORC2 effector protein kinase C (PKC)α did rescue Rictor-mediated RhoGDI2 downregulation, partially rescuing Rac-guanosine triphosphate (GTP) and migration/motility. These findings suggest that mTORC2 uses two coordinated pathways to activate cell invasion/motility, both of which converge on Rac1. Akt signaling activates Rac1 through the Rac-GEF Tiam1, while PKC signaling dampens expression of the endogenous Rac1 inhibitor, RhoGDI2.

  13. Genomic and phenotypic characterization of myxoma virus from Great Britain reveals multiple evolutionary pathways distinct from those in Australia

    Science.gov (United States)

    Kerr, Peter J.; Cattadori, Isabella M.; Fitch, Adam; Geber, Adam; Liu, June; Sim, Derek G.; Boag, Brian; Ghedin, Elodie

    2017-01-01

    The co-evolution of myxoma virus (MYXV) and the European rabbit occurred independently in Australia and Europe from different progenitor viruses. Although this is the canonical study of the evolution of virulence, whether the genomic and phenotypic outcomes of MYXV evolution in Europe mirror those observed in Australia is unknown. We addressed this question using viruses isolated in the United Kingdom early in the MYXV epizootic (1954–1955) and between 2008–2013. The later UK viruses fell into three distinct lineages indicative of a long period of separation and independent evolution. Although rates of evolutionary change were almost identical to those previously described for MYXV in Australia and strongly clock-like, genome evolution in the UK and Australia showed little convergence. The phenotypes of eight UK viruses from three lineages were characterized in laboratory rabbits and compared to the progenitor (release) Lausanne strain. Inferred virulence ranged from highly virulent (grade 1) to highly attenuated (grade 5). Two broad disease types were seen: cutaneous nodular myxomatosis characterized by multiple raised secondary cutaneous lesions, or an amyxomatous phenotype with few or no secondary lesions. A novel clinical outcome was acute death with pulmonary oedema and haemorrhage, often associated with bacteria in many tissues but an absence of inflammatory cells. Notably, reading frame disruptions in genes defined as essential for virulence in the progenitor Lausanne strain were compatible with the acquisition of high virulence. Combined, these data support a model of ongoing host-pathogen co-evolution in which multiple genetic pathways can produce successful outcomes in the field that involve both different virulence grades and disease phenotypes, with alterations in tissue tropism and disease mechanisms. PMID:28253375

  14. Genomic and phenotypic characterization of myxoma virus from Great Britain reveals multiple evolutionary pathways distinct from those in Australia.

    Directory of Open Access Journals (Sweden)

    Peter J Kerr

    2017-03-01

    Full Text Available The co-evolution of myxoma virus (MYXV and the European rabbit occurred independently in Australia and Europe from different progenitor viruses. Although this is the canonical study of the evolution of virulence, whether the genomic and phenotypic outcomes of MYXV evolution in Europe mirror those observed in Australia is unknown. We addressed this question using viruses isolated in the United Kingdom early in the MYXV epizootic (1954-1955 and between 2008-2013. The later UK viruses fell into three distinct lineages indicative of a long period of separation and independent evolution. Although rates of evolutionary change were almost identical to those previously described for MYXV in Australia and strongly clock-like, genome evolution in the UK and Australia showed little convergence. The phenotypes of eight UK viruses from three lineages were characterized in laboratory rabbits and compared to the progenitor (release Lausanne strain. Inferred virulence ranged from highly virulent (grade 1 to highly attenuated (grade 5. Two broad disease types were seen: cutaneous nodular myxomatosis characterized by multiple raised secondary cutaneous lesions, or an amyxomatous phenotype with few or no secondary lesions. A novel clinical outcome was acute death with pulmonary oedema and haemorrhage, often associated with bacteria in many tissues but an absence of inflammatory cells. Notably, reading frame disruptions in genes defined as essential for virulence in the progenitor Lausanne strain were compatible with the acquisition of high virulence. Combined, these data support a model of ongoing host-pathogen co-evolution in which multiple genetic pathways can produce successful outcomes in the field that involve both different virulence grades and disease phenotypes, with alterations in tissue tropism and disease mechanisms.

  15. Genomic and phenotypic characterization of myxoma virus from Great Britain reveals multiple evolutionary pathways distinct from those in Australia.

    Science.gov (United States)

    Kerr, Peter J; Cattadori, Isabella M; Rogers, Matthew B; Fitch, Adam; Geber, Adam; Liu, June; Sim, Derek G; Boag, Brian; Eden, John-Sebastian; Ghedin, Elodie; Read, Andrew F; Holmes, Edward C

    2017-03-01

    The co-evolution of myxoma virus (MYXV) and the European rabbit occurred independently in Australia and Europe from different progenitor viruses. Although this is the canonical study of the evolution of virulence, whether the genomic and phenotypic outcomes of MYXV evolution in Europe mirror those observed in Australia is unknown. We addressed this question using viruses isolated in the United Kingdom early in the MYXV epizootic (1954-1955) and between 2008-2013. The later UK viruses fell into three distinct lineages indicative of a long period of separation and independent evolution. Although rates of evolutionary change were almost identical to those previously described for MYXV in Australia and strongly clock-like, genome evolution in the UK and Australia showed little convergence. The phenotypes of eight UK viruses from three lineages were characterized in laboratory rabbits and compared to the progenitor (release) Lausanne strain. Inferred virulence ranged from highly virulent (grade 1) to highly attenuated (grade 5). Two broad disease types were seen: cutaneous nodular myxomatosis characterized by multiple raised secondary cutaneous lesions, or an amyxomatous phenotype with few or no secondary lesions. A novel clinical outcome was acute death with pulmonary oedema and haemorrhage, often associated with bacteria in many tissues but an absence of inflammatory cells. Notably, reading frame disruptions in genes defined as essential for virulence in the progenitor Lausanne strain were compatible with the acquisition of high virulence. Combined, these data support a model of ongoing host-pathogen co-evolution in which multiple genetic pathways can produce successful outcomes in the field that involve both different virulence grades and disease phenotypes, with alterations in tissue tropism and disease mechanisms.

  16. Combined MEK and ERK inhibition overcomes therapy-mediated pathway reactivation in RAS mutant tumors.

    Directory of Open Access Journals (Sweden)

    Mark Merchant

    Full Text Available Mitogen-activated protein kinase (MAPK pathway dysregulation is implicated in >30% of all cancers, rationalizing the development of RAF, MEK and ERK inhibitors. While BRAF and MEK inhibitors improve BRAF mutant melanoma patient outcomes, these inhibitors had limited success in other MAPK dysregulated tumors, with insufficient pathway suppression and likely pathway reactivation. In this study we show that inhibition of either MEK or ERK alone only transiently inhibits the MAPK pathway due to feedback reactivation. Simultaneous targeting of both MEK and ERK nodes results in deeper and more durable suppression of MAPK signaling that is not achievable with any dose of single agent, in tumors where feedback reactivation occurs. Strikingly, combined MEK and ERK inhibition is synergistic in RAS mutant models but only additive in BRAF mutant models where the RAF complex is dissociated from RAS and thus feedback productivity is disabled. We discovered that pathway reactivation in RAS mutant models occurs at the level of CRAF with combination treatment resulting in a markedly more active pool of CRAF. However, distinct from single node targeting, combining MEK and ERK inhibitor treatment effectively blocks the downstream signaling as assessed by transcriptional signatures and phospho-p90RSK. Importantly, these data reveal that MAPK pathway inhibitors whose activity is attenuated due to feedback reactivation can be rescued with sufficient inhibition by using a combination of MEK and ERK inhibitors. The MEK and ERK combination significantly suppresses MAPK pathway output and tumor growth in vivo to a greater extent than the maximum tolerated doses of single agents, and results in improved anti-tumor activity in multiple xenografts as well as in two Kras mutant genetically engineered mouse (GEM models. Collectively, these data demonstrate that combined MEK and ERK inhibition is functionally unique, yielding greater than additive anti-tumor effects and

  17. Agrobacterium Mediated Transient Gene Silencing (AMTS) in Stevia rebaudiana: Insights into Steviol Glycoside Biosynthesis Pathway

    Science.gov (United States)

    Guleria, Praveen; Yadav, Sudesh Kumar

    2013-01-01

    Background Steviol glycoside biosynthesis pathway has emerged as bifurcation from ent-kaurenoic acid, substrate of methyl erythritol phosphate pathway that also leads to gibberellin biosynthesis. However, the genetic regulation of steviol glycoside biosynthesis has not been studied. So, in present study RNA interference (RNAi) based Agrobacterium mediated transient gene silencing (AMTS) approach was followed. SrKA13H and three SrUGTs (SrUGT85C2, SrUGT74G1 and SrUGT76G1) genes encoding ent-kaurenoic acid-13 hydroxylase and three UDP glycosyltransferases of steviol glycoside biosynthesis pathway were silenced in Stevia rebaudiana to understand its molecular mechanism and association with gibberellins. Methodology/Principal Findings RNAi mediated AMTS of SrKA13H and three SrUGTs has significantly reduced the expression of targeted endogenous genes as well as total steviol glycoside accumulation. While gibberellins (GA3) content was significantly enhanced on AMTS of SrUGT85C2 and SrKA13H. Silencing of SrKA13H and SrUGT85C2 was found to block the metabolite flux of steviol glycoside pathway and shifted it towards GA3 biosynthesis. Further, molecular docking of three SrUGT proteins has documented highest affinity of SrUGT76G1 for the substrates of alternate pathways synthesizing steviol glycosides. This could be a plausible reason for maximum reduction in steviol glycoside content on silencing of SrUGT76G1 than other genes. Conclusions SrKA13H and SrUGT85C2 were identified as regulatory genes influencing carbon flux between steviol glycoside and gibberellin biosynthesis. This study has also documented the existence of alternate steviol glycoside biosynthesis route. PMID:24023961

  18. Agrobacterium mediated transient gene silencing (AMTS in Stevia rebaudiana: insights into steviol glycoside biosynthesis pathway.

    Directory of Open Access Journals (Sweden)

    Praveen Guleria

    Full Text Available Steviol glycoside biosynthesis pathway has emerged as bifurcation from ent-kaurenoic acid, substrate of methyl erythritol phosphate pathway that also leads to gibberellin biosynthesis. However, the genetic regulation of steviol glycoside biosynthesis has not been studied. So, in present study RNA interference (RNAi based Agrobacterium mediated transient gene silencing (AMTS approach was followed. SrKA13H and three SrUGTs (SrUGT85C2, SrUGT74G1 and SrUGT76G1 genes encoding ent-kaurenoic acid-13 hydroxylase and three UDP glycosyltransferases of steviol glycoside biosynthesis pathway were silenced in Stevia rebaudiana to understand its molecular mechanism and association with gibberellins.RNAi mediated AMTS of SrKA13H and three SrUGTs has significantly reduced the expression of targeted endogenous genes as well as total steviol glycoside accumulation. While gibberellins (GA3 content was significantly enhanced on AMTS of SrUGT85C2 and SrKA13H. Silencing of SrKA13H and SrUGT85C2 was found to block the metabolite flux of steviol glycoside pathway and shifted it towards GA3 biosynthesis. Further, molecular docking of three SrUGT proteins has documented highest affinity of SrUGT76G1 for the substrates of alternate pathways synthesizing steviol glycosides. This could be a plausible reason for maximum reduction in steviol glycoside content on silencing of SrUGT76G1 than other genes.SrKA13H and SrUGT85C2 were identified as regulatory genes influencing carbon flux between steviol glycoside and gibberellin biosynthesis. This study has also documented the existence of alternate steviol glycoside biosynthesis route.

  19. Silver Nanoparticles Induce HePG-2 Cells Apoptosis Through ROS-Mediated Signaling Pathways

    Science.gov (United States)

    Zhu, Bing; Li, Yinghua; Lin, Zhengfang; Zhao, Mingqi; Xu, Tiantian; Wang, Changbing; Deng, Ning

    2016-04-01

    Recently, silver nanoparticles (AgNPs) have been shown to provide a novel approach to overcome tumors, especially those of hepatocarcinoma. However, the anticancer mechanism of silver nanoparticles is unclear. Thus, the purpose of this study was to estimate the effect of AgNPs on proliferation and activation of ROS-mediated signaling pathway on human hepatocellular carcinoma HePG-2 cells. A simple chemical method for preparing AgNPs with superior anticancer activity has been showed in this study. AgNPs were detected by transmission electronic microscopy (TEM) and energy dispersive X-ray (EDX). The size distribution and zeta potential of silver nanoparticles were detected by Zetasizer Nano. The average size of AgNPs (2 nm) observably increased the cellular uptake by endocytosis. AgNPs markedly inhibited the proliferation of HePG-2 cells through induction of apoptosis with caspase-3 activation and PARP cleavage. AgNPs with dose-dependent manner significantly increased the apoptotic cell population (sub-G1). Furthermore, AgNP-induced apoptosis was found dependent on the overproduction of reactive oxygen species (ROS) and affecting of MAPKs and AKT signaling and DNA damage-mediated p53 phosphorylation to advance HePG-2 cells apoptosis. Therefore, our results show that the mechanism of ROS-mediated signaling pathways may provide useful information in AgNP-induced HePG-2 cell apoptosis.

  20. Proteomic analysis of the signaling pathway mediated by the heterotrimeric Gα protein Pga1 of Penicillium chrysogenum.

    Science.gov (United States)

    Carrasco-Navarro, Ulises; Vera-Estrella, Rosario; Barkla, Bronwyn J; Zúñiga-León, Eduardo; Reyes-Vivas, Horacio; Fernández, Francisco J; Fierro, Francisco

    2016-10-06

    The heterotrimeric Gα protein Pga1-mediated signaling pathway regulates the entire developmental program in Penicillium chrysogenum, from spore germination to the formation of conidia. In addition it participates in the regulation of penicillin biosynthesis. We aimed to advance the understanding of this key signaling pathway using a proteomics approach, a powerful tool to identify effectors participating in signal transduction pathways. Penicillium chrysogenum mutants with different levels of activity of the Pga1-mediated signaling pathway were used to perform comparative proteomic analyses by 2D-DIGE and LC-MS/MS. Thirty proteins were identified which showed differences in abundance dependent on Pga1 activity level. By modifying the intracellular levels of cAMP we could establish cAMP-dependent and cAMP-independent pathways in Pga1-mediated signaling. Pga1 was shown to regulate abundance of enzymes in primary metabolic pathways involved in ATP, NADPH and cysteine biosynthesis, compounds that are needed for high levels of penicillin production. An in vivo phosphorylated protein containing a pleckstrin homology domain was identified; this protein is a candidate for signal transduction activity. Proteins with possible roles in purine metabolism, protein folding, stress response and morphogenesis were also identified whose abundance was regulated by Pga1 signaling. Thirty proteins whose abundance was regulated by the Pga1-mediated signaling pathway were identified. These proteins are involved in primary metabolism, stress response, development and signal transduction. A model describing the pathways through which Pga1 signaling regulates different cellular processes is proposed.

  1. Positive and negative affect as predictors of urge to smoke: temporal factors and mediational pathways.

    Science.gov (United States)

    Leventhal, Adam M; Greenberg, Jodie B; Trujillo, Michael A; Ameringer, Katherine J; Lisha, Nadra E; Pang, Raina D; Monterosso, John

    2013-03-01

    Elucidating interrelations between prior affective experience, current affective state, and acute urge to smoke could inform affective models of addiction motivation and smoking cessation treatment development. This study tested the hypothesis that prior levels of positive (PA) and negative (NA) affect predict current smoking urge via a mediational pathway involving current state affect. We also explored if tobacco deprivation moderated affect-urge relations and compared the effects of PA and NA on smoking urge to one another. At a baseline session, smokers reported affect experienced over the preceding few weeks. At a subsequent experimental session, participants were randomly assigned to 12-hr tobacco deprived (n = 51) or nondeprived (n = 69) conditions and reported state affect and current urge. Results revealed a mediational pathway whereby prior NA reported at baseline predicted state NA at the experimental session, which in turn predicted current urge. This mediational pathway was found primarily for an urge subtype indicative of urgent need to smoke and desire to smoke for NA relief, was stronger in the deprived (vs. nondeprived) condition, and remained significant after controlling for PA. Prior PA and current state PA were inversely associated with current urge; however, these associations were eliminated after controlling for NA. These results cohere with negative reinforcement models of addiction and with prior research and suggest that: (a) NA plays a stronger role in smoking motivation than PA; (b) state affect is an important mechanism linking prior affective experience to current urge; and (c) affect management interventions may attenuate smoking urge in individuals with a history of affective disturbance. (PsycINFO Database Record (c) 2013 APA, all rights reserved).

  2. The Ebola virus glycoprotein mediates entry via a non-classical dynamin-dependent macropinocytic pathway

    International Nuclear Information System (INIS)

    Mulherkar, Nirupama; Raaben, Matthijs; Torre, Juan Carlos de la; Whelan, Sean P.; Chandran, Kartik

    2011-01-01

    Ebola virus (EBOV) has been reported to enter cultured cell lines via a dynamin-2-independent macropinocytic pathway or clathrin-mediated endocytosis. The route(s) of productive EBOV internalization into physiologically relevant cell types remain unexplored, and viral-host requirements for this process are incompletely understood. Here, we use electron microscopy and complementary chemical and genetic approaches to demonstrate that the viral glycoprotein, GP, induces macropinocytic uptake of viral particles into cells. GP's highly-glycosylated mucin domain is dispensable for virus-induced macropinocytosis, arguing that interactions between other sequences in GP and the host cell surface are responsible. Unexpectedly, we also found a requirement for the large GTPase dynamin-2, which is proposed to be dispensable for several types of macropinocytosis. Our results provide evidence that EBOV uses an atypical dynamin-dependent macropinocytosis-like entry pathway to enter Vero cells, adherent human peripheral blood-derived monocytes, and a mouse dendritic cell line.

  3. LPA, HGF, and EGF utilize distinct combinations of signaling pathways to promote migration and invasion of MDA-MB-231 breast carcinoma cells

    International Nuclear Information System (INIS)

    Harrison, Susan MW; Knifley, Teresa; Chen, Min; O’Connor, Kathleen L

    2013-01-01

    Various pathways impinge on the actin-myosin pathway to facilitate cell migration and invasion including members of the Rho family of small GTPases and MAPK. However, the signaling components that are considered important for these processes vary substantially within the literature with certain pathways being favored. These distinctions in signaling pathways utilized are often attributed to differences in cell type or physiological conditions; however, these attributes have not been systematically assessed. To address this question, we analyzed the migration and invasion of MDA-MB-231 breast carcinoma cell line in response to various stimuli including lysophosphatidic acid (LPA), hepatocyte growth factor (HGF) and epidermal growth factor (EGF) and determined the involvement of select signaling pathways that impact myosin light chain phosphorylation. LPA, a potent stimulator of the Rho-ROCK pathway, surprisingly did not require the Rho-ROCK pathway to stimulate migration but instead utilized Rac and MAPK. In contrast, LPA-stimulated invasion required Rho, Rac, and MAPK. Of these three major pathways, EGF-stimulated MDA-MB-231 migration and invasion required Rho; however, Rac was essential only for invasion and MAPK was dispensable for migration. HGF signaling, interestingly, utilized the same pathways for migration and invasion, requiring Rho but not Rac signaling. Notably, the dependency of HGF-stimulated migration and invasion as well as EGF-stimulated invasion on MAPK was subject to the inhibitors used. As expected, myosin light chain kinase (MLCK), a convergence point for MAPK and Rho family GTPase signaling, was required for all six conditions. These observations suggest that, while multiple signaling pathways contribute to cancer cell motility, not all pathways operate under all conditions. Thus, our study highlights the plasticity of cancer cells to adapt to multiple migratory cues

  4. Selenium-Mediated Dehalogenation of Halogenated Nucleosides and its Relevance to the DNA Repair Pathway.

    Science.gov (United States)

    Mondal, Santanu; Manna, Debasish; Mugesh, Govindasamy

    2015-08-03

    Halogenated nucleosides can be incorporated into the newly synthesized DNA of replicating cells and therefore are commonly used in the detection of proliferating cells in living tissues. Dehalogenation of these modified nucleosides is one of the key pathways involved in DNA repair mediated by the uracil-DNA glycosylase. Herein, we report the first example of a selenium-mediated dehalogenation of halogenated nucleosides. We also show that the mechanism for the debromination is remarkably different from that of deiodination and that the presence of a ribose or deoxyribose moiety in the nucleosides facilitates the deiodination. The results described herein should help in understanding the metabolism of halogenated nucleosides in DNA and RNA. © 2015 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  5. TLR2-dependent inhibition of macrophage responses to IFN-gamma is mediated by distinct, gene-specific mechanisms.

    Directory of Open Access Journals (Sweden)

    Sarah A Benson

    2009-07-01

    Full Text Available Mycobacterium tuberculosis uses multiple mechanisms to avoid elimination by the immune system. We have previously shown that M. tuberculosis can inhibit selected macrophage responses to IFN-gamma through TLR2-dependent and -independent mechanisms. To specifically address the role of TLR2 signaling in mediating this inhibition, we stimulated macrophages with the specific TLR2/1 ligand Pam(3CSK(4 and assayed responses to IFN-gamma. Pam(3CSK(4 stimulation prior to IFN-gamma inhibited transcription of the unrelated IFN-gamma-inducible genes, CIITA and CXCL11. Surface expression of MHC class II and secretion of CXCL11 were greatly reduced as well, indicating that the reduction in transcripts had downstream effects. Inhibition of both genes required new protein synthesis. Using chromatin immunoprecipitation, we found that TLR2 stimulation inhibited IFN-gamma-induced RNA polymerase II binding to the CIITA and CXCL11 promoters. Furthermore, TATA binding protein was unable to bind the TATA box of the CXCL11 promoter, suggesting that assembly of transcriptional machinery was disrupted. However, TLR2 stimulation affected chromatin modifications differently at each of the inhibited promoters. Histone H3 and H4 acetylation was reduced at the CIITA promoter but unaffected at the CXCL11 promoter. In addition, NF-kappaB signaling was required for inhibition of CXCL11 transcription, but not for inhibition of CIITA. Taken together, these results indicate that TLR2-dependent inhibition of IFN-gamma-induced gene expression is mediated by distinct, gene-specific mechanisms that disrupt binding of the transcriptional machinery to the promoters.

  6. Mediation pathways and effects of green structures on respiratory mortality via reducing air pollution.

    Science.gov (United States)

    Shen, Yu-Sheng; Lung, Shih-Chun Candice

    2017-02-23

    Previous studies have shown both health and environmental benefits of green spaces, especially in moderating temperature and reducing air pollution. However, the characteristics of green structures have been overlooked in previous investigations. In addition, the mediation effects of green structures on respiratory mortality have not been assessed. This study explores the potential mediation pathways and effects of green structure characteristics on respiratory mortality through temperature, primary and secondary air pollutants separately using partial least squares model with data from Taiwan. The measurable characteristics of green structure include the largest patch percentage, landscape proportion, aggregation, patch distance, and fragmentation. The results showed that mortality of pneumonia and chronic lower respiratory diseases could be reduced by minimizing fragmentation and increasing the largest patch percentage of green structure, and the mediation effects are mostly through reducing air pollutants rather than temperature. Moreover, a high proportion of but fragmented green spaces would increase secondary air pollutants and enhance health risks; demonstrating the deficiency of traditional greening policy with primary focus on coverage ratio. This is the first research focusing on mediation effects of green structure characteristics on respiratory mortality, revealing that appropriate green structure planning can be a useful complementary strategy in environmental health management.

  7. Proteomic analysis of the signaling pathway mediated by the heterotrimeric G? protein Pga1 of Penicillium chrysogenum

    OpenAIRE

    Carrasco-Navarro, Ulises; Vera-Estrella, Rosario; Barkla, Bronwyn J.; Z??iga-Le?n, Eduardo; Reyes-Vivas, Horacio; Fern?ndez, Francisco J.; Fierro, Francisco

    2016-01-01

    Background The heterotrimeric G? protein Pga1-mediated signaling pathway regulates the entire developmental program in Penicillium chrysogenum, from spore germination to the formation of conidia. In addition it participates in the regulation of penicillin biosynthesis. We aimed to advance the understanding of this key signaling pathway using a proteomics approach, a powerful tool to identify effectors participating in signal transduction pathways. Results Penicillium chrysogenum mutants with ...

  8. MAPKs are essential upstream signaling pathways in proteolytic cartilage degradation--divergence in pathways leading to aggrecanase and MMP-mediated articular cartilage degradation

    DEFF Research Database (Denmark)

    Sondergaard, B-C; Schultz, N; Madsen, S H

    2010-01-01

    Matrix metalloproteinases (MMPs) and aggrecanases are essential players in cartilage degradation. However, the signaling pathways that results in MMP and/or aggrecanase synthesis and activation are not well understood. We investigated the molecular events leading to MMP- and aggrecanase-mediated ......Matrix metalloproteinases (MMPs) and aggrecanases are essential players in cartilage degradation. However, the signaling pathways that results in MMP and/or aggrecanase synthesis and activation are not well understood. We investigated the molecular events leading to MMP- and aggrecanase......-mediated cartilage degradation....

  9. Mediating pathways and gender differences between shift work and subjective cognitive function.

    Science.gov (United States)

    Wong, Imelda S; Smith, Peter M; Ibrahim, Selahadin; Mustard, Cameron A; Gignac, Monique A M

    2016-11-01

    Increased injury risk among shift workers is often attributed to cognitive function deficits that come about as a result of sleep disruptions. However, little is known about the intermediate influences of other factors (eg, work stress, health) which may affect this relationship. In addition, gender differences in these the complex relationships have not been fully explored. The purpose of this study is to (1) identify the extent to which work and non-work factors mediate the relationship between shift work, sleep and subsequent subjective cognitive function; and (2) determine if the mediating pathways differ for men and women. Data from the 2010 National Population Health Survey was used to create a cross-sectional sample of 4255 employed Canadians. Using path modelling, we examined the direct and indirect relationships between shift work, sleep duration, sleep quality and subjective cognitive function. Multigroup analyses tested for significantly different pathways between men and women. Potential confounding effects of age and self-reported health and potential mediating effects of work stress were simultaneously examined. Work stress and sleep quality significantly mediated the effects of shift work on cognition. Age and health confounded the relationship between sleep quality and subjective cognition. No differences were found between men and women. Occupational health and safety programmes are needed to address stress and health factors, in addition to sleep hygiene, to effectively address cognitive function among shift workers. Published by the BMJ Publishing Group Limited. For permission to use (where not already granted under a licence) please go to http://www.bmj.com/company/products-services/rights-and-licensing/.

  10. College Men and Alcohol Use: Positive Alcohol Expectancies as a Mediator Between Distinct Masculine Norms and Alcohol Use.

    Science.gov (United States)

    Iwamoto, Derek Kenji; Corbin, William; Lejuez, Carl; MacPherson, Laura

    2014-01-01

    College men are more likely to engage in health-compromising behaviors including risky drinking behavior, and experience more alcohol-related problems, including violence and arrest, as compared to women. The study of masculine norms or societal expectations, defined as beliefs and values about what it means to be a man, is one promising area of investigation that may help explain within-group differences and differential rates of alcohol use among men. Using the gender social learning model, we investigated the role of positive alcohol expectancies as an underlying mediator between masculine norms and alcohol use among college men. Data from 804 college adult men ( Mean age = 20.43) were collected through a web-based assessment. Participants completed a self-report measure of binge drinking, frequency of drinking, quantity of drinks, conformity to masculine norms, and positive alcohol expectancies measures. Structural equation modeling was used to examine relations between masculine norms, alcohol expectancies and alcohol use. The masculine norms of "Playboy" and Risk-Taking were positively related to heavy alcohol use, while Emotional Control and Heterosexual Presentation were both negatively associated with alcohol use, after controlling for fraternity Greek status and positive expectancies. Playboy and Winning norms were positively associated with positive expectancies while Power Over Women was inversely related to positive expectancies which, in turn, were associated with heavier alcohol use. This study was a novel exploration into the multiple pathways and mediators through which positive alcohol expectancies may help explain and provide specificity to the masculinity and alcohol use relationship among college men.

  11. The HIV-1 envelope transmembrane domain binds TLR2 through a distinct dimerization motif and inhibits TLR2-mediated responses.

    Directory of Open Access Journals (Sweden)

    Eliran Moshe Reuven

    2014-08-01

    Full Text Available HIV-1 uses a number of means to manipulate the immune system, to avoid recognition and to highjack signaling pathways. HIV-1 infected cells show limited Toll-Like Receptor (TLR responsiveness via as yet unknown mechanisms. Using biochemical and biophysical approaches, we demonstrate that the trans-membrane domain (TMD of the HIV-1 envelope (ENV directly interacts with TLR2 TMD within the membrane milieu. This interaction attenuates TNFα, IL-6 and MCP-1 secretion in macrophages, induced by natural ligands of TLR2 both in in vitro and in vivo models. This was associated with decreased levels of ERK phosphorylation. Furthermore, mutagenesis demonstrated the importance of a conserved GxxxG motif in driving this interaction within the membrane milieu. The administration of the ENV TMD in vivo to lipotechoic acid (LTA/Galactosamine-mediated septic mice resulted in a significant decrease in mortality and in tissue damage, due to the weakening of systemic macrophage activation. Our findings suggest that the TMD of ENV is involved in modulation of the innate immune response during HIV infection. Furthermore, due to the high functional homology of viral ENV proteins this function may be a general character of viral-induced immune modulation.

  12. Distinct CPT-induced deaths in lung cancer cells caused by clathrin-mediated internalization of CP micelles

    Science.gov (United States)

    Liu, Yu-Sheng; Cheng, Ru-You; Lo, Yu-Lun; Hsu, Chin; Chen, Su-Hwei; Chiu, Chien-Chih; Wang, Li-Fang

    2016-02-01

    We previously synthesized a chondroitin sulfate-graft-poly(ε-caprolactone) copolymer (H-CP) with a high content of poly(ε-caprolactone) (18.7 mol%), which self-assembled in water into a rod-like micelle to encapsulate hydrophobic camptothecin (CPT) in the core (micelle/CPT) for tumor-targeted drug delivery. As a result of the recognition of the micelle by CD44, the micelle/CPT entered CRL-5802 cells efficiently and released CPT efficaciously, resulting in higher tumor suppression than commercial CPT-11. In this study, H1299 cells were found to have a higher CD44 expression than CRL-5802 cells. However, the lower CD44-expressing CRL-5802 cells had a higher percentage of cell death and higher cellular uptake of the micelle/CPT than the higher CD44-expressing H1299 cells. Examination of the internalization pathway of the micelle/CPT in the presence of different endocytic chemical inhibitors showed that the CRL-5802 cells involved clathrin-mediated endocytosis, which was not found in the H1299 cells. Analysis of the cell cycle of the two cell lines exposed to the micelle/CPT revealed that the CRL-5802 cells arrested mainly in the S phase and the H1299 cells arrested mainly in the G2-M phase. A consistent result was also found in the evaluation of γ-H2AX expression, which was about three-fold higher in the CRL-5802 cells than in the H1299 cells. A near-infrared dye, IR780, was encapsulated into the micelle to observe the in vivo biodistribution of the micelle/IR780 in tumor-bearing mice. The CRL-5802 tumor showed a higher fluorescence intensity than the H1299 tumor at any tracing time after 1 h. Thus we tentatively concluded that CRL-5802 cells utilized the clathrin-mediated internalization pathway and arrested in the S phase on exposure to the micelle/CPT; all are possible reasons for the better therapeutic outcome in CRL-5802 cells than in H1299 cells.We previously synthesized a chondroitin sulfate-graft-poly(ε-caprolactone) copolymer (H-CP) with a high content of

  13. Endoplasmic reticulum (ER) stress and cAMP/PKA pathway mediated Zn-induced hepatic lipolysis.

    Science.gov (United States)

    Song, Yu-Feng; Hogstrand, Christer; Wei, Chuan-Chuan; Wu, Kun; Pan, Ya-Xiong; Luo, Zhi

    2017-09-01

    The present study was performed to determine the effect of Zn exposure influencing endoplasmic reticulum (ER) stress, explore the underlying molecular mechanism of Zn-induced hepatic lipolysis in a fish species of significance for aquaculture, yellow catfish Pelteobagrus fulvidraco. We found that waterborne Zn exposure evoked ER stress and unfolded protein response (UPR), and activated cAMP/PKA pathway, and up-regulated hepatic lipolysis. The increase in ER stress and lipolysis were associated with activation of cAMP/PKA signaling pathway. Zn also induced an increase in intracellular Ca 2+ level, which could be partially prevented by dantrolene (RyR receptor inhibitor) and 2-APB (IP3 receptor inhibitor), demonstrating that the disturbed Ca 2+ homeostasis in ER contributed to ER stress and dysregulation of lipolysis. Inhibition of ER stress by PBA attenuated UPR, inhibited the activation of cAMP/PKA pathway and resulted in down-regulation of lipolysis. Inhibition of protein kinase RNA-activated-like ER kinase (PERK) by GSK2656157 and inositol-requiring enzyme (IRE) by STF-083010 differentially influenced Zn-induced changes of lipid metabolism, indicating that PERK and IRE pathways played different regulatory roles in Zn-induced lipolysis. Inhibition of PKA by H89 blocked the Zn-induced activation of cAMP/PKA pathway with a concomitant inhibition of ER stress-mediated lipolysis. Taken together, our findings highlight the importance of the ER stress-cAMP/PKA axis in Zn-induced lipolysis, which provides new insights into Zn toxicology in fish and probably in other vertebrates. Copyright © 2017 Elsevier Ltd. All rights reserved.

  14. Wnt/β-catenin signalling pathway mediated aberrant hippocampal neurogenesis in kainic acid-induced epilepsy.

    Science.gov (United States)

    Qu, Zhengyi; Su, Fang; Qi, Xueting; Sun, Jianbo; Wang, Hongcai; Qiao, Zhenkui; Zhao, Hong; Zhu, Yulan

    2017-10-01

    Temporal lobe epilepsy is a chronic disorder of nerve system, mainly characterized by hippocampal sclerosis with massive neuronal loss and severe gliosis. Aberrant neurogenesis has been shown in the epileptogenesis process of temporal lobe epilepsy. However, the molecular mechanisms underlying aberrant neurogenesis remain unclear. The roles of Wnt signalling cascade have been well established in neurogenesis during multiple aspects. Here, we used kainic acid-induced rat epilepsy model to investigate whether Wnt/β-catenin signalling pathway is involved in the aberrant neurogenesis in temporal lobe epilepsy. Immunostaining and western blotting results showed that the expression levels of β-catenin, Wnt3a, and cyclin D1, the key regulators in Wnt signalling pathway, were up-regulated during acute epilepsy induced by the injection of kainic acids, indicating that Wnt signalling pathway was activated in kainic acid-induced temporal lobe epilepsy. Moreover, BrdU labelling results showed that blockade of the Wnt signalling by knocking down β-catenin attenuated aberrant neurogenesis induced by kainic acids injection. Altogether, Wnt/β-catenin signalling pathway mediated hippocampal neurogenesis during epilepsy, which might provide new strategies for clinical treatment of temporal lobe epilepsy. Temporal lobe epilepsy is a chronic disorder of nerve system, mainly characterized by hippocampal sclerosis. Aberrant neurogenesis has been shown to involve in the epileptogenesis process of temporal lobe epilepsy. In the present study, we discovered that Wnt3a/β-catenin signalling pathway serves as a link between aberrant neurogenesis and underlying remodelling in the hippocampus, leading to temporal lobe epilepsy, which might provide new strategies for clinical treatment of temporal lobe epilepsy. Copyright © 2017 John Wiley & Sons, Ltd.

  15. Catabolite-mediated mutations in alternate toluene degradative pathways in Pseudomonas putida.

    Science.gov (United States)

    Leddy, M B; Phipps, D W; Ridgway, H F

    1995-01-01

    Pseudomonas putida 54g grew on mineral salts with toluene and exhibited catechol-2,3-dioxygenase (C23O) activity, indicating a meta pathway. After 10 to 15 days on toluene, nondegrading (Tol-) variants approached nearly 10% of total CFU. Auxotrophs were not detected among variants, suggesting selective loss of catabolic function(s). Variant formation was substrate dependent, since Tol- cells were observed on neither ethylbenzene, glucose, nor peptone-based media nor when toluene catabolism was suppressed by glucose. Unlike wild-type cells, variants did not grow on gasoline, toluene, benzene, ethylbenzene, benzoate, or catechol, suggesting loss of meta pathway function. Catabolic and C23O activities were restored to variants via transfer of a 78-mDa TOL-like plasmid from a wild-type Tol+ donor. Tests for reversion of variants to Tol+ were uniformly negative, suggesting possible delection or excision of catabolic genes. Deletions were confirmed in some variants by failure to hybridize with a DNA probe specific for the xylE gene encoding C23O. Cells grown on benzoate remained Tol+ but were C23O- and contained a plasmid of reduced size or were plasmid free, suggesting an alternate chromosomal catabolic pathway, also defective in variants. Cells exposed to benzyl alcohol, the initial oxidation product of toluene, accumulated > 13% variants in 5 days, even when cell division was repressed by nitrogen deprivation to abrogate selection processes. No variants formed in identical ethylbenzene-exposed controls. The results suggest that benzyl alcohol mediates irreversible defects in both a plasmid-associated meta pathway and an alternate chromosomal pathway. PMID:7642499

  16. Emergency room nurses' pathway to turnover intention: a moderated serial mediation analysis.

    Science.gov (United States)

    Bruyneel, Luk; Thoelen, Tom; Adriaenssens, Jef; Sermeus, Walter

    2017-04-01

    The aim of this study was to explore the association between the quality of the work environment, job characteristics, demographic characteristics and a pathway of job satisfaction, emotional exhaustion and turnover intention among nurses in emergency departments and perform subgroup analyses. Turnover intention among nurses is high. Multiple causes have been described, mostly in large studies of nurses working on general wards, often without considering complementarity of conceptual models and showing scant interest in the consistency of associations across subgroups of nurses. Cross-sectional multicentre survey. Convenience sample of 294 nurses in 11 Belgian emergency departments during 2014-2015. Indirect effects in the form of mediation and serial mediation were estimated to assess the association between work environment (Magnet model), job characteristics (Job Demand Control Support model) and turnover intention via job satisfaction and emotional exhaustion. Consistency of these indirect effects across subgroups of nurses was examined using moderated mediation analysis (conditional indirect effects). Several Magnet and Job Demand Control Support dimensions were related to turnover intention, either via job dissatisfaction (mediation) or via job satisfaction and emotional exhaustion (serial mediation). In the case of social support from supervisor, these indirect effects were only significant for female nurses, among whom turnover intention was higher. Last, nurses with more years of experience were less likely to indicate turnover intention. To maximize prevention of turnover intention at emergency departments, interventions could target early career nurses, work environment and job characteristics. Female nurses in particular may also benefit from improved social support from their supervisor. © 2016 John Wiley & Sons Ltd.

  17. Changes of TSPO-mediated mitophagy signaling pathway in learned helplessness mice.

    Science.gov (United States)

    Li, Dongmei; Zheng, Ji; Wang, Mingyang; Feng, Lu; Ren, Zhili; Liu, Yanyong; Yang, Nan; Zuo, Pingping

    2016-11-30

    Low response rate was witnessed with the present monoaminergic based antidepressants, urging a need for new therapeutic target identification. Accumulated evidences strongly suggest that mitochondrial deficit is implicated in major depression and 18kDa translocator protein (TSPO) plays an important role in regulating mitochondrial function. However the changes of TSPO and TSPO mediated mitophagy pathway in the depressive brain is unclear. In present study, a well validated animal model of depression, learned helplessness (LH), was employed to investigate the relevant changes. Significant behavioral changes were observed in the LH mice. Results showed that TSPO and other mitophagy related proteins, such as VDAC1, Pink1 and Beclin1 were significantly decreased by LH challenge. Moreover, KIFC2, relevant to the mitochondrial transport and Snap25, relevant to neurotransmitter vesicle release, were also obviously down-regulated in the LH mice, which further rendered supportive evidence for the existing mitochondrial dysfunction in LH mice. Present results demonstrated that LH induced depressive symptoms and affected TSPO-mediated mitophagy pathway, indicating a potential target candidate for depression treatment. Copyright © 2016 Elsevier Ireland Ltd. All rights reserved.

  18. Poly(I:C) reduces expression of JAM-A and induces secretion of IL-8 and TNF-α via distinct NF-κB pathways in human nasal epithelial cells

    International Nuclear Information System (INIS)

    Ohkuni, Tsuyoshi; Kojima, Takashi; Ogasawara, Noriko; Masaki, Tomoyuki; Fuchimoto, Jun; Kamekura, Ryuta; Koizumi, Jun-ichi; Ichimiya, Shingo; Murata, Masaki; Tanaka, Satoshi; Himi, Tetsuo; Sawada, Norimasa

    2011-01-01

    Human nasal epithelium is an important physical barrier and innate immune defense protecting against inhaled substances and pathogens. Toll-like receptor (TLR) signaling, which plays a key role in the innate immune response, has not been well characterized in human nasal epithelial cells (HNECs), including the epithelial tight junctional barrier. In the present study, mRNAs of TLR1-10 were detected in hTERT-transfected HNECs, which can be used as an indispensable and stable model of normal HNECs, similar to primary cultured HNECs. To investigate the changes of tight junction proteins and the signal transduction pathways via TLRs in HNECs in vitro, hTERT-transfected HNECs were treated with TLR2 ligand P 3 CSK 4 , TLR3 ligand poly(I:C), TLR4 ligand LPS, TLR7/8 ligand CL097, TLR8 ligand ssRNA40/LyoVec, and TLR9 ligand ODN2006. In hTERT-transfected HNECs, treatment with poly(I:C) significantly reduced expression of the tight junction protein JAM-A and induced secretion of proinflammatory cytokines IL-8 and TNF-α. Both the reduction of JAM-A expression and the induction of secretion of IL-8 and TNF-α after treatment with poly(I:C) were modulated by distinct signal transduction pathways via EGFR, PI3K, and p38 MAPK and finally regulated by a TLR3-mediated NF-κB pathway. The control of TLR3-mediated signaling pathways in HNECs may be important not only in infection by viral dsRNA but also in autoimmune diseases caused by endogenous dsRNA released from necrotic cells.

  19. Delineating the Maladaptive Pathways of Child Maltreatment: A Mediated Moderation Analysis of the Roles of Self Perception and Social Support

    Science.gov (United States)

    Appleyard, Karen; Yang, Chongming; Runyan, Desmond K.

    2014-01-01

    The current study investigated concurrent and longitudinal mediated and mediated moderation pathways among maltreatment, self perception (i.e., loneliness and self esteem), social support, and internalizing and externalizing behavior problems. For both genders, early childhood maltreatment (i.e., ages 0–6) was related directly to internalizing and externalizing behavior problems at age 6, and later maltreatment (i.e., ages 6–8) was directly related to internalizing and externalizing behavior problems at age 8. Results of concurrent mediation and mediated moderation indicated that early maltreatment was significantly related to internalizing and externalizing behavior problems at age 6 indirectly both through age 6 loneliness and self esteem for boys and through age 6 loneliness for girls. Significant moderation of the pathway from early maltreatment to self esteem, and, for boys, significant mediated moderation to emotional and behavioral problems were found, such that the mediated effect through self esteem varied across levels of social support, though in an unexpected direction. No significant longitudinal mediation or mediated moderation was found, however, between the age 6 mediators and moderator and internalizing or externalizing problems at age 8. The roles of the hypothesized mediating and moderating mechanisms are discussed, with implications for designing intervention and prevention programs. PMID:20423545

  20. Downregulation of toll-like receptor-mediated signalling pathways in oral lichen planus.

    Science.gov (United States)

    Sinon, Suraya H; Rich, Alison M; Parachuru, Venkata P B; Firth, Fiona A; Milne, Trudy; Seymour, Gregory J

    2016-01-01

    The objective of this study was to investigate the expression of Toll-like receptors (TLR) and TLR-associated signalling pathway genes in oral lichen planus (OLP). Initially, immunohistochemistry was used to determine TLR expression in 12 formalin-fixed archival OLP tissues with 12 non-specifically inflamed oral tissues as controls. RNA was isolated from further fresh samples of OLP and non-specifically inflamed oral tissue controls (n = 6 for both groups) and used in qRT(2)-PCR focused arrays to determine the expression of TLRs and associated signalling pathway genes. Genes with a statistical significance of ±two-fold regulation (FR) and a P-value < 0.05 were considered as significantly regulated. Significantly more TLR4(+) cells were present in the inflammatory infiltrate in OLP compared with the control tissues (P < 0.05). There was no statistically significant difference in the numbers of TLR2(+) and TLR8(+) cells between the groups. TLR3 was significantly downregulated in OLP (P < 0.01). TLR8 was upregulated in OLP, but the difference between the groups was not statistically significant. The TLR-mediated signalling-associated protein genes MyD88 and TIRAP were significantly downregulated (P < 0.01 and P < 0.05), as were IRAK1 (P < 0.05), MAPK8 (P < 0.01), MAP3K1 (P < 0.05), MAP4K4 (P < 0.05), REL (P < 0.01) and RELA (P < 0.01). Stress proteins HMGB1 and the heat shock protein D1 were significantly downregulated in OLP (P < 0.01). These findings suggest a downregulation of TLR-mediated signalling pathways in OLP lesions. © 2015 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.

  1. Genistein Stimulates Jejunum Chloride Secretion via an Akt-Mediated Pathway in Intact Female Mice

    Directory of Open Access Journals (Sweden)

    Lana Leung

    2015-02-01

    Full Text Available Background/Aims: We have previously shown that daily subcutaneous injections with the naturally occurring phytoestrogen genistein (600 mg genistein/kg body weight/day, 600G results in a significantly increased basal intestinal chloride, Cl-, secretion (Isc, a measure of transepithelial secretion in intact C57BL/6J female mice after 1-week of treatment, compared to controls (DMSO vehicle injected. Removal of endogenous estrogen via ovariectomy (OVX had no effect on the 600G-mediated increase in basal Isc. Methods: Given the estrogen-like characteristics of genistein, we compared the effects of daily estradiol (E2 injections (10 mg E2/kg body weight/day, 10E2 on basal Isc in intact and OVX mice. In intact mice, 10E2 was without effect on basal Isc, however, in OVX mice, 10E2 significantly increased basal Isc (mimicked 600G. The goal of the current study was to characterize the intracellular signaling pathways responsible for mediating 600G- or 10E2-stimulated increases in basal Isc in intact female or OVX mice. Results: We measured total protein expression in isolated segments of jejunum using western blot from the following six groups of mice; intact or OVX with; 600G, 10E2 or control. The proteins of interest were: Akt, p-Akt, p-PDK1, p-PTEN, p-c-Raf, p-GSK-3β, rap-1 and ERK1/2. All blots were normalized to GAPDH levels (n = 6-18/group. Conclusion: These data suggest that the presence of the endogenous sex steroid, estrogen, modifies the intracellular signaling pathway required to mediate Cl- secretion when the intestine is exposed to exogenous 600G or E2. These studies may have relevance for designing pharmacological tools for women with intestinal chloride secretory dysfunctions.

  2. Identification of potential pathway mediation targets in Toll-like receptor signaling.

    Directory of Open Access Journals (Sweden)

    Fan Li

    2009-02-01

    Full Text Available Recent advances in reconstruction and analytical methods for signaling networks have spurred the development of large-scale models that incorporate fully functional and biologically relevant features. An extended reconstruction of the human Toll-like receptor signaling network is presented herein. This reconstruction contains an extensive complement of kinases, phosphatases, and other associated proteins that mediate the signaling cascade along with a delineation of their associated chemical reactions. A computational framework based on the methods of large-scale convex analysis was developed and applied to this network to characterize input-output relationships. The input-output relationships enabled significant modularization of the network into ten pathways. The analysis identified potential candidates for inhibitory mediation of TLR signaling with respect to their specificity and potency. Subsequently, we were able to identify eight novel inhibition targets through constraint-based modeling methods. The results of this study are expected to yield meaningful avenues for further research in the task of mediating the Toll-like receptor signaling network and its effects.

  3. H19 mediates methotrexate resistance in colorectal cancer through activating Wnt/β-catenin pathway

    Energy Technology Data Exchange (ETDEWEB)

    Wu, Ke-feng [Guangdong Key Laboratory for Research and Development of Natural Drugs, Guangdong Medical University, Zhanjiang, Guangdong (China); Liang, Wei-Cheng [School of Biomedical Sciences, Faculty of Medicine, The Chinese University of Hong Kong, Hong Kong (China); Feng, Lu [Department of Orthopaedics & Traumatology, The Chinese University of Hong Kong, Prince of Wales Hospital, Shatin, Hong Kong (China); Pang, Jian-xin [School of Pharmaceutical Sciences, Southern Medical University, Guangzhou 510515 (China); Waye, Mary Miu-Yee [School of Biomedical Sciences, Faculty of Medicine, The Chinese University of Hong Kong, Hong Kong (China); Zhang, Jin-Fang [Department of Orthopaedics & Traumatology, The Chinese University of Hong Kong, Prince of Wales Hospital, Shatin, Hong Kong (China); Fu, Wei-Ming, E-mail: fuweiming76@smu.edu.cn [School of Pharmaceutical Sciences, Southern Medical University, Guangzhou 510515 (China)

    2017-01-15

    Colorectal cancer (CRC) is a common malignancy, most of which remain unresponsive to chemotherapy. As one of the earliest cytotoxic drugs, methotrexate (MTX) serves as an anti-metabolite and anti-folate chemotherapy for various cancers. Unfortunately, MTX resistance prevents its clinical application in cancer therapy. Thereby, overcoming the drug resistance is an alternative strategy to maximize the therapeutic efficacy of MTX in clinics. Long noncoding RNAs (lncRNAs) have gained widespread attention in recent years. More and more emerging evidences have demonstrated that they play important regulatory roles in various biological activities and disease progression including drug resistance. In the present study, a MTX-resistant colorectal cell line HT-29 (HT-29-R) was developed, which displayed the active proliferation and shortened cell cycle. LncRNA H19 was found to be significantly upregulated in this resistant cell line. Further investigation showed that H19 knockdown sensitized the MTX resistance in HT-29-R cells while its overexpression improved the MTX resistance in the parental cells, suggesting that H19 mediate MTX resistance. The Wnt/β-catenin signaling was activated in HT-29-R cells, and H19 knockdown suppressed this signaling in the parental cells. In conclusion, H19 mediated MTX resistance via activating Wnt/β-catenin signaling, which help to develop H19 as a promising therapeutic target for MTX resistant CRC. - Highlights: • A methotrexate (MTX) -resistant colorectal cancer cell line HT-29 (HT-29-R) has been developed. • H19 was upregulated in HT-29-R cells. • H19 mediated MTX resistance in colorectal cancer (CRC). • Wnt/β-catenin pathway was involved in the H19-mediated MTX resistance in CRC cells.

  4. H19 mediates methotrexate resistance in colorectal cancer through activating Wnt/β-catenin pathway

    International Nuclear Information System (INIS)

    Wu, Ke-feng; Liang, Wei-Cheng; Feng, Lu; Pang, Jian-xin; Waye, Mary Miu-Yee; Zhang, Jin-Fang; Fu, Wei-Ming

    2017-01-01

    Colorectal cancer (CRC) is a common malignancy, most of which remain unresponsive to chemotherapy. As one of the earliest cytotoxic drugs, methotrexate (MTX) serves as an anti-metabolite and anti-folate chemotherapy for various cancers. Unfortunately, MTX resistance prevents its clinical application in cancer therapy. Thereby, overcoming the drug resistance is an alternative strategy to maximize the therapeutic efficacy of MTX in clinics. Long noncoding RNAs (lncRNAs) have gained widespread attention in recent years. More and more emerging evidences have demonstrated that they play important regulatory roles in various biological activities and disease progression including drug resistance. In the present study, a MTX-resistant colorectal cell line HT-29 (HT-29-R) was developed, which displayed the active proliferation and shortened cell cycle. LncRNA H19 was found to be significantly upregulated in this resistant cell line. Further investigation showed that H19 knockdown sensitized the MTX resistance in HT-29-R cells while its overexpression improved the MTX resistance in the parental cells, suggesting that H19 mediate MTX resistance. The Wnt/β-catenin signaling was activated in HT-29-R cells, and H19 knockdown suppressed this signaling in the parental cells. In conclusion, H19 mediated MTX resistance via activating Wnt/β-catenin signaling, which help to develop H19 as a promising therapeutic target for MTX resistant CRC. - Highlights: • A methotrexate (MTX) -resistant colorectal cancer cell line HT-29 (HT-29-R) has been developed. • H19 was upregulated in HT-29-R cells. • H19 mediated MTX resistance in colorectal cancer (CRC). • Wnt/β-catenin pathway was involved in the H19-mediated MTX resistance in CRC cells.

  5. Single-cell analysis reveals a link between CD3- and CD59-mediated signaling pathways in Jurkat T cells

    International Nuclear Information System (INIS)

    Lipp, A. M.

    2012-01-01

    Elevation of intracellular free calcium concentration ([Ca2+]i) is a key signal during T cell activation and is commonly used as a read-out parameter for stimulation of T cell signaling. Upon T cell stimulation a variety of calcium signals is produced by individual cells of the T cell population and the type of calcium signal strongly influences cell fate decisions. The heterogeneous nature of T cells is masked in ensemble measurements, which highlights the need for single-cell measurements. In this study we used single-cell calcium measurements in Jurkat cells to investigate signaling pathways, which are triggered by different proteins, namely CD3 and CD59. By application of an automated cluster algorithm the presented assay provides unbiased analysis of a large data set of individual calcium time traces generated by the whole cell population. By using this method we could demonstrate that the Jurkat population generates heterogeneous calcium signals in a stimulus-dependent manner. Furthermore, our data revealed the existence of a link between CD3- and CD59-mediated signaling pathways. Single-cell calcium measurements in Jurkat cells expressing different levels of the T cell receptor (TCR) complex indicated that CD59-mediated calcium signaling is critically dependent on TCR surface expression levels. In addition, triggering CD59-mediated calcium signaling resulted in down-regulation of TCR surface expression levels, which is known to happen upon direct TCR triggering too. Moreover, by using siRNA-mediated protein knock-downs and protein knock-out Jurkat mutants we could show that CD3- and CD59-mediated calcium signaling require identical key proteins. We therefore explored by which mechanism CD59-mediated signaling couples into TCR-mediated signaling. Fluorescence recovery after photobleaching (FRAP) experiments and live-cell protein-protein interaction assays provided no evidence of a direct physical interaction between CD3- and CD59-mediated signaling pathways

  6. A STUDY OF INTERMEDIATES INVOLVED IN THE FOLDING PATHWAY FOR RECOMBINANT HUMAN MACROPHAGE COLONY-STIMULATING FACTOR (M-CSF) - EVIDENCE FOR 2 DISTINCT FOLDING PATHWAYS

    NARCIS (Netherlands)

    WILKINS, JA; CONE, J; RANDHAWA, ZI; WOOD, D; WARREN, MK; WITKOWSKA, HE

    The folding pathway for a 150-amino acid recombinant form of the dimeric cytokine human macrophage colony-stimulating factor (M-CSF) has been studied. All 14 cysteine residues in the biologically active homodimer are involved in disulfide linkages. The structural characteristics of folding

  7. Lhx6 delineates a pathway mediating innate reproductive behaviors from the amygdala to the hypothalamus.

    Science.gov (United States)

    Choi, Gloria B; Dong, Hong-Wei; Murphy, Andrew J; Valenzuela, David M; Yancopoulos, George D; Swanson, Larry W; Anderson, David J

    2005-05-19

    In mammals, innate reproductive and defensive behaviors are mediated by anatomically segregated connections between the amygdala and hypothalamus. This anatomic segregation poses the problem of how the brain integrates activity in these circuits when faced with conflicting stimuli eliciting such mutually exclusive behaviors. Using genetically encoded and conventional axonal tracers, we have found that the transcription factor Lhx6 delineates the reproductive branch of this pathway. Other Lhx proteins mark neurons in amygdalar nuclei implicated in defense. We have traced parallel projections from the posterior medial amygdala, activated by reproductive or defensive olfactory stimuli, respectively, to a point of convergence in the ventromedial hypothalamus. The opposite neurotransmitter phenotypes of these convergent projections suggest a "gate control" mechanism for the inhibition of reproductive behaviors by threatening stimuli. Our data therefore identify a potential neural substrate for integrating the influences of conflicting behavioral cues and a transcription factor family that may contribute to the development of this substrate.

  8. Multiple signaling pathways mediated by dopamine and calcium ionophore A23187 in human platelets

    International Nuclear Information System (INIS)

    Saeed, S.A.; Waqar, M.A.

    2009-01-01

    This study was undertaken to investigate the mechanism(s) of platelet aggregation induced by the synergistic action of dopamine (DA) and a Ca/sup +2/-ionophore, A23187. DA showed non significant effect on platelet aggregation over a wide range of concentrations (up to 500 micro M), but did potentiate the aggregation response of A23187. Aggregation induced by A23187 was inhibited by calcium channel blockers (diltiazem and verpamil), receptor blockers (chlorpromazine and haloperidol) and a cyclo-oxygenase inhibitor (indomethacin). However, the inhibitory effect of these blockers was more pronounced (with a selectivity ratio of 1.5-28) in the aggregation induced by synergistic effect of A23187 and DA. A phosphatidylinositol 3-kinase (P1 3-Kinase) inhibitor, wortmanin (1C/sub 50/. 25-30 nM), inhibited aggregation induced by either A23187 or DA and act synergistically. This synergistic effect on platelet aggregation is mediated through multiple signaling pathways. (author)

  9. A bacterial cyclic dinucleotide activates the cytosolic surveillance pathway and mediates innate resistance to tuberculosis.

    Science.gov (United States)

    Dey, Bappaditya; Dey, Ruchi Jain; Cheung, Laurene S; Pokkali, Supriya; Guo, Haidan; Lee, Jong-Hee; Bishai, William R

    2015-04-01

    Detection of cyclic-di-adenosine monophosphate (c-di-AMP), a bacterial second messenger, by the host cytoplasmic surveillance pathway (CSP) is known to elicit type I interferon (IFN) responses, which are crucial to antimicrobial defense. However, the mechanisms and role of c-di-AMP signaling in Mycobacterium tuberculosis virulence remain unclear. Here we show that resistance to tuberculosis requires CSP-mediated detection of c-di-AMP produced by M. tuberculosis and that levels of c-di-AMP modulate the fate of infection. We found that a di-adenylate cyclase (disA or dacA)-overexpressing M. tuberculosis strain that secretes excess c-di-AMP activates the interferon regulatory factor (IRF) pathway with enhanced levels of IFN-β, elicits increased macrophage autophagy, and exhibits substantial virulence attenuation in mice. We show that c-di-AMP-mediated IFN-β induction during M. tuberculosis infection requires stimulator of interferon genes (STING)-signaling. We observed that c-di-AMP induction of IFN-β is independent of the cytosolic nucleic acid receptor cyclic GMP-AMP (cGAMP) synthase (cGAS), but cGAS nevertheless contributes substantially to the overall IFN-β response to M. tuberculosis infection. In sum, our results reveal c-di-AMP to be a key mycobacterial pathogen-associated molecular pattern (PAMP) driving host type I IFN responses and autophagy. These findings suggest that modulating the levels of this small molecule may lead to novel immunotherapeutic strategies against tuberculosis.

  10. Surfactant protein D delays Fas- and TRAIL-mediated extrinsic pathway of apoptosis in T cells.

    Science.gov (United States)

    Djiadeu, Pascal; Kotra, Lakshmi P; Sweezey, Neil; Palaniyar, Nades

    2017-05-01

    Only a few extracellular soluble proteins are known to modulate apoptosis. We considered that surfactant-associated protein D (SP-D), an innate immune collectin present on many mucosal surfaces, could regulate apoptosis. Although SP-D is known to be important for immune cell homeostasis, whether SP-D affects apoptosis is unknown. In this study we aimed to determine the effects of SP-D on Jurkat T cells and human T cells dying by apoptosis. Here we show that SP-D binds to Jurkat T cells and delays the progression of Fas (CD95)-Fas ligand and TRAIL-TRAIL receptor induced, but not TNF-TNF receptor-mediated apoptosis. SP-D exerts its effects by reducing the activation of initiator caspase-8 and executioner caspase-3. SP-D also delays the surface exposure of phosphatidylserine. The effect of SP-D was ablated by the presence of caspase-8 inhibitor, but not by intrinsic pathway inhibitors. The binding ability of SP-D to dying cells decreases during the early stages of apoptosis, suggesting the release of apoptotic cell surface targets during apoptosis. SP-D also delays FasL-induced death of primary human T cells. SP-D delaying the progression of the extrinsic pathway of apoptosis could have important implications in regulating immune cell homeostasis at mucosal surfaces.

  11. Hypothalamic mTOR pathway mediates thyroid hormone-induced hyperphagia in hyperthyroidism.

    Science.gov (United States)

    Varela, Luis; Martínez-Sánchez, Noelia; Gallego, Rosalía; Vázquez, María J; Roa, Juan; Gándara, Marina; Schoenmakers, Erik; Nogueiras, Rubén; Chatterjee, Krishna; Tena-Sempere, Manuel; Diéguez, Carlos; López, Miguel

    2012-06-01

    Hyperthyroidism is characterized in rats by increased energy expenditure and marked hyperphagia. Alterations of thermogenesis linked to hyperthyroidism are associated with dysregulation of hypothalamic AMPK and fatty acid metabolism; however, the central mechanisms mediating hyperthyroidism-induced hyperphagia remain largely unclear. Here, we demonstrate that hyperthyroid rats exhibit marked up-regulation of the hypothalamic mammalian target of rapamycin (mTOR) signalling pathway associated with increased mRNA levels of agouti-related protein (AgRP) and neuropeptide Y (NPY), and decreased mRNA levels of pro-opiomelanocortin (POMC) in the arcuate nucleus of the hypothalamus (ARC), an area where mTOR co-localizes with thyroid hormone receptor-α (TRα). Central administration of thyroid hormone (T3) or genetic activation of thyroid hormone signalling in the ARC recapitulated hyperthyroidism effects on feeding and the mTOR pathway. In turn, central inhibition of mTOR signalling with rapamycin in hyperthyroid rats reversed hyperphagia and normalized the expression of ARC-derived neuropeptides, resulting in substantial body weight loss. The data indicate that in the hyperthyroid state, increased feeding is associated with thyroid hormone-induced up-regulation of mTOR signalling. Furthermore, our findings that different neuronal modulations influence food intake and energy expenditure in hyperthyroidism pave the way for a more rational design of specific and selective therapeutic compounds aimed at reversing the metabolic consequences of this disease. Copyright © 2012 Pathological Society of Great Britain and Ireland. Published by John Wiley & Sons, Ltd.

  12. 25-Hydroxycholesterol promotes fibroblast-mediated tissue remodeling through NF-κB dependent pathway

    International Nuclear Information System (INIS)

    Ichikawa, Tomohiro; Sugiura, Hisatoshi; Koarai, Akira; Kikuchi, Takashi; Hiramatsu, Masataka; Kawabata, Hiroki; Akamatsu, Keiichiro; Hirano, Tsunahiko; Nakanishi, Masanori; Matsunaga, Kazuto; Minakata, Yoshiaki; Ichinose, Masakazu

    2013-01-01

    Abnormal structural alterations termed remodeling, including fibrosis and alveolar wall destruction, are important features of the pathophysiology of chronic airway diseases such as chronic obstructive pulmonary disease (COPD) and asthma. 25-hydroxycholesterol (25-HC) is enzymatically produced by cholesterol 25-hydorxylase (CH25H) in macrophages and is reported to be involved in the formation of arteriosclerosis. We previously demonstrated that the expression of CH25H and production of 25HC were increased in the lungs of COPD. However, the role of 25-HC in lung tissue remodeling is unknown. In this study, we investigated the effect of 25-HC on fibroblast-mediated tissue remodeling using human fetal lung fibroblasts (HFL-1) in vitro. 25-HC significantly augmented α-smooth muscle actin (SMA) (P 1 production (P 1 release. These results suggest that 25-HC could contribute to fibroblast-mediated lung tissue remodeling by promoting myofibroblast differentiation and the excessive release of extracellular matrix protein and MMPs via an NF-κB-TGF-β dependent pathway

  13. Intercellular and intracellular signaling pathways mediating ionizing radiation-induced bystander effects

    International Nuclear Information System (INIS)

    Hamada, Nobuyuki; Hara, Takamitsu; Kobayashi, Yasuhiko; Matsumoto, Hideki

    2007-01-01

    A rapidly growing body of experimental evidence indicates that ionizing radiation induces biological effects in non-irradiated bystander cells that have received signals from adjacent or distant irradiated cells. This phenomenon, which has been termed the ionizing radiation-induced bystander effect, challenges the long-standing paradigm that radiation traversal through the nucleus of a cell is a prerequisite to elicit genetic damage or a biological response. Bystander effects have been observed in a number of experimental systems, and cells whose nucleus or cytoplasm is irradiated exert bystander responses. Bystander cells manifest a multitude of biological consequences, such as genetic and epigenetic changes, alterations in gene expression, activation of signal transduction pathways, and delayed effects in their progeny. Several mediating mechanisms have been proposed. These involve gap junction-mediated intercellular communication, secreted soluble factors, oxidative metabolism, plasma membrane-bound lipid rafts, and calcium fluxes. This paper reviews briefly the current knowledge of the bystander effect with a focus on proposed mechanisms. The potential benefit of bystander effects to cancer radiotherapy will also be discussed. (author)

  14. Arakeri’s Reflex: an Alternative Pathway for Dento-Cardiac Reflex Mediated Syncope

    Directory of Open Access Journals (Sweden)

    Veena Arali

    2010-03-01

    Full Text Available Introduction: Dentocardiac reflex, a variant of trigeminocardiac reflex elicited specifically during tooth extraction procedures in den-tal/maxillofacial surgery and is believed to cause syncope with an afferent link mediated by posterior superior alveolar nerve. Another variant of trigeminocardiac reflex which is also of interest to the oral and maxillofacial surgeon is oculocardiac reflex which can be triggered by direct or indirect manipulation of eye globe or muscles around it.The hypothesis: Excessive or injudicious pressure or manipulations around the maxillary first molars during extraction procedure are as-sociated with maximum incidence of bradycardia and hypotension than around incisor/ canine/ third molars. This is because; the pressure on eye globe and ophthalmic rectus muscle is maximum during extraction of first molar than incisor/canine and third molars. This observation led us to postulate an alternative pathway for dentocardiac reflex mediated syncope which may possibly justify the maxillary first molar region as a prone factor for the trigger. Evaluation of the hypothesis: Present hypothesis may not confer the specific factor responsible for switch in autonomic response in syncope origin during the tooth extraction procedure, but may provide a clue to where we should be looking.

  15. Visualization of oxytocin release that mediates paired pulse facilitation in hypothalamic pathways to brainstem autonomic neurons.

    Directory of Open Access Journals (Sweden)

    Ramón A Piñol

    Full Text Available Recent work has shown that oxytocin is involved in more than lactation and uterine contraction. The paraventricular nucleus of the hypothalamus (PVN contains neuroendocrine neurons that control the release of hormones, including vasopressin and oxytocin. Other populations of PVN neurons do not release hormones, but rather project to and release neurotransmitters onto other neurons in the CNS involved in fluid retention, thermoregulation, sexual behavior and responses to stress. Activation of oxytocin receptors can be cardioprotective and reduces the adverse cardiovascular consequences of anxiety and stress, yet how oxytocin can affect heart rate and cardiac function is unknown. While anatomical work has shown the presence of peptides, including oxytocin, in the projections from the PVN to parasympathetic nuclei, electrophysiological studies to date have only demonstrated release of glutamate and activation of fast ligand gated receptors in these pathways. In this study, using rats, we directly show, using sniffer CHO cells that express oxytocin receptors and the Ca2+ indicator R-GECO, that optogenetic activation of channelrhodopsin-2 (ChR2 expressing PVN fibers in the brainstem activates oxytocin receptors in the dorsomotor nucleus of the vagus (DMNV. We also demonstrate that while a single photoactivation of PVN terminals only activates glutamatergic receptors in brainstem cardiac vagal neurons (CVNs, neurons that dominate the neural control of heart rate, both the paired pulse facilitation, and sustained enhancement of glutamate release in this pathway is mediated by activation of oxytocin receptors. Our results provide direct evidence that a pathway from the PVN likely releases oxytocin and enhances short-term plasticity of this critical autonomic connection.

  16. Functions of the nonsense-mediated mRNA decay pathway in Drosophila development.

    Directory of Open Access Journals (Sweden)

    Mark M Metzstein

    2006-12-01

    Full Text Available Nonsense-mediated mRNA decay (NMD is a cellular surveillance mechanism that degrades transcripts containing premature translation termination codons, and it also influences expression of certain wild-type transcripts. Although the biochemical mechanisms of NMD have been studied intensively, its developmental functions and importance are less clear. Here, we describe the isolation and characterization of Drosophila "photoshop" mutations, which increase expression of green fluorescent protein and other transgenes. Mapping and molecular analyses show that photoshop mutations are loss-of-function mutations in the Drosophila homologs of NMD genes Upf1, Upf2, and Smg1. We find that Upf1 and Upf2 are broadly active during development, and they are required for NMD as well as for proper expression of dozens of wild-type genes during development and for larval viability. Genetic mosaic analysis shows that Upf1 and Upf2 are required for growth and/or survival of imaginal cell clones, but this defect can be overcome if surrounding wild-type cells are eliminated. By contrast, we find that the PI3K-related kinase Smg1 potentiates but is not required for NMD or for viability, implying that the Upf1 phosphorylation cycle that is required for mammalian and Caenorhabditis elegans NMD has a more limited role during Drosophila development. Finally, we show that the SV40 3' UTR, present in many Drosophila transgenes, targets the transgenes for regulation by the NMD pathway. The results establish that the Drosophila NMD pathway is broadly active and essential for development, and one critical function of the pathway is to endow proliferating imaginal cells with a competitive growth advantage that prevents them from being overtaken by other proliferating cells.

  17. Glioma cell fate decisions mediated by Dll1-Jag1-Fringe in Notch1 signaling pathway.

    Science.gov (United States)

    Shi, Xiaofei; Wang, Ruiqi

    2017-09-21

    The Notch family of proteins plays a vital role in determining cell fates, such as proliferation, differentiation, and apoptosis. It has been shown that Notch1 and its ligands, Dll1 and Jag1, are overexpressed in many glioma cell lines and primary human gliomas. The roles of Notch1 in some cancers have been firmly established, and recent data implicate that it plays important roles in glioma cell fate decisions. This paper focuses on devising a specific theoretical framework that incorporates Dll1, Jag1, and Fringe in Notch1 signaling pathway to explore their functional roles of these proteins in glioma cells in the tumorigenesis and progression of human gliomas, and to study how glioma cell fate decisions are modulated by both trans-activation and cis-inhibition. This paper presents a computational model for Notch1 signaling pathway in glioma cells. Based on the bifurcation analysis of the model, we show that how the glioma cell fate decisions are modulated by both trans-activation and cis-inhibition mediated by the Fringe protein, providing insight into the design and control principles of the Notch signaling system and the gliomas. This paper presents a computational model for Notch1 signaling pathway in glioma cells based on intertwined dynamics with cis-inhibition and trans-activation involving the proteins Notch1, Dll1, Jag1, and Fringe. The results show that how the glioma cell fate transitions are performed by the Notch1 signaling. Transition from grade III ∼ IV with significantly high Notch1 to grade I ∼ II with high Notch1, and then to normal cells by repressing the Fringe levels or decreasing the strength of enhancement induced by Fringe.

  18. Visualization of Oxytocin Release that Mediates Paired Pulse Facilitation in Hypothalamic Pathways to Brainstem Autonomic Neurons

    Science.gov (United States)

    Piñol, Ramón A.; Jameson, Heather; Popratiloff, Anastas; Lee, Norman H.; Mendelowitz, David

    2014-01-01

    Recent work has shown that oxytocin is involved in more than lactation and uterine contraction. The paraventricular nucleus of the hypothalamus (PVN) contains neuroendocrine neurons that control the release of hormones, including vasopressin and oxytocin. Other populations of PVN neurons do not release hormones, but rather project to and release neurotransmitters onto other neurons in the CNS involved in fluid retention, thermoregulation, sexual behavior and responses to stress. Activation of oxytocin receptors can be cardioprotective and reduces the adverse cardiovascular consequences of anxiety and stress, yet how oxytocin can affect heart rate and cardiac function is unknown. While anatomical work has shown the presence of peptides, including oxytocin, in the projections from the PVN to parasympathetic nuclei, electrophysiological studies to date have only demonstrated release of glutamate and activation of fast ligand gated receptors in these pathways. In this study, using rats, we directly show, using sniffer CHO cells that express oxytocin receptors and the Ca2+ indicator R-GECO, that optogenetic activation of channelrhodopsin-2 (ChR2) expressing PVN fibers in the brainstem activates oxytocin receptors in the dorsomotor nucleus of the vagus (DMNV). We also demonstrate that while a single photoactivation of PVN terminals only activates glutamatergic receptors in brainstem cardiac vagal neurons (CVNs), neurons that dominate the neural control of heart rate, both the paired pulse facilitation, and sustained enhancement of glutamate release in this pathway is mediated by activation of oxytocin receptors. Our results provide direct evidence that a pathway from the PVN likely releases oxytocin and enhances short-term plasticity of this critical autonomic connection. PMID:25379676

  19. Temporal expression profiling identifies pathways mediating effect of causal variant on phenotype.

    Directory of Open Access Journals (Sweden)

    Saumya Gupta

    2015-06-01

    of analyzing allele-specific transcriptional dynamics of mediating genes. Applications in higher eukaryotes can be valuable for inferring causal molecular pathways underlying complex dynamic processes, such as development, physiology and disease progression.

  20. Chlorpromazine-induced hepatotoxicity during inflammation is mediated by TIRAP-dependent signaling pathway in mice

    International Nuclear Information System (INIS)

    Gandhi, Adarsh; Guo, Tao; Shah, Pranav; Moorthy, Bhagavatula; Ghose, Romi

    2013-01-01

    Inflammation is a major component of idiosyncratic adverse drug reactions (IADRs). To understand the molecular mechanism of inflammation-mediated IADRs, we determined the role of the Toll-like receptor (TLR) signaling pathway in idiosyncratic hepatotoxicity of the anti-psychotic drug, chlorpromazine (CPZ). Activation of TLRs recruits the first adaptor protein, Toll-interleukin 1 receptor domain containing adaptor protein (TIRAP) to the TIR domain of TLRs leading to the activation of the downstream kinase, c-Jun-N-terminal kinase (JNK). Prolonged activation of JNK leads to cell-death. We hypothesized that activation of TLR2 by lipoteichoic acid (LTA) or TLR4 by lipopolysaccharide (LPS) will augment the hepatotoxicity of CPZ by TIRAP-dependent mechanism involving prolonged activation of JNK. Adult male C57BL/6, TIRAP +/+ and TIRAP −/− mice were pretreated with saline, LPS (2 mg/kg) or LTA (6 mg/kg) for 30 min or 16 h followed by CPZ (5 mg/kg) or saline (vehicle) up to 24 h. We found that treatment of mice with CPZ in presence of LPS or LTA leads to ∼ 3–4 fold increase in serum ALT levels, a marked reduction in hepatic glycogen content, significant induction of serum tumor necrosis factor (TNF) α and prolonged JNK activation, compared to LPS or LTA alone. Similar results were observed in TIRAP +/+ mice, whereas the effects of LPS or LTA on CPZ-induced hepatotoxicity were attenuated in TIRAP −/− mice. For the first time, we show that inflammation-mediated hepatotoxicity of CPZ is dependent on TIRAP, and involves prolonged JNK activation in vivo. Thus, TIRAP-dependent pathways may be targeted to predict and prevent inflammation-mediated IADRs. -- Highlights: ► Inflammation augments the toxicity of an idiosyncratic hepatotoxin chlorpromazine. ► Activation of Toll-like receptors by LPS or LTA induces chlorpromazine toxicity. ► Sustained stress kinase (JNK) activation is associated with chlorpromazine toxicity. ► These studies provide novel mechanistic

  1. Chlorpromazine-induced hepatotoxicity during inflammation is mediated by TIRAP-dependent signaling pathway in mice

    Energy Technology Data Exchange (ETDEWEB)

    Gandhi, Adarsh, E-mail: adarsh.gandhi@nih.gov [University of Houston, Department of Pharmacological and Pharmaceutical Sciences, College of Pharmacy, 1441 Moursund Street, Room 517, Houston, TX 77030 (United States); Guo, Tao, E-mail: tguo4@jhu.edu [University of Houston, Department of Pharmacological and Pharmaceutical Sciences, College of Pharmacy, 1441 Moursund Street, Room 517, Houston, TX 77030 (United States); Shah, Pranav [University of Houston, Department of Pharmacological and Pharmaceutical Sciences, College of Pharmacy, 1441 Moursund Street, Room 517, Houston, TX 77030 (United States); Moorthy, Bhagavatula [Baylor College of Medicine, Department of Pediatrics, 1102 Bates Avenue, Suite 530, Houston, TX 77030 (United States); Ghose, Romi, E-mail: rghose@uh.edu [University of Houston, Department of Pharmacological and Pharmaceutical Sciences, College of Pharmacy, 1441 Moursund Street, Room 517, Houston, TX 77030 (United States)

    2013-02-01

    Inflammation is a major component of idiosyncratic adverse drug reactions (IADRs). To understand the molecular mechanism of inflammation-mediated IADRs, we determined the role of the Toll-like receptor (TLR) signaling pathway in idiosyncratic hepatotoxicity of the anti-psychotic drug, chlorpromazine (CPZ). Activation of TLRs recruits the first adaptor protein, Toll-interleukin 1 receptor domain containing adaptor protein (TIRAP) to the TIR domain of TLRs leading to the activation of the downstream kinase, c-Jun-N-terminal kinase (JNK). Prolonged activation of JNK leads to cell-death. We hypothesized that activation of TLR2 by lipoteichoic acid (LTA) or TLR4 by lipopolysaccharide (LPS) will augment the hepatotoxicity of CPZ by TIRAP-dependent mechanism involving prolonged activation of JNK. Adult male C57BL/6, TIRAP{sup +/+} and TIRAP{sup −/−} mice were pretreated with saline, LPS (2 mg/kg) or LTA (6 mg/kg) for 30 min or 16 h followed by CPZ (5 mg/kg) or saline (vehicle) up to 24 h. We found that treatment of mice with CPZ in presence of LPS or LTA leads to ∼ 3–4 fold increase in serum ALT levels, a marked reduction in hepatic glycogen content, significant induction of serum tumor necrosis factor (TNF) α and prolonged JNK activation, compared to LPS or LTA alone. Similar results were observed in TIRAP{sup +/+} mice, whereas the effects of LPS or LTA on CPZ-induced hepatotoxicity were attenuated in TIRAP{sup −/−} mice. For the first time, we show that inflammation-mediated hepatotoxicity of CPZ is dependent on TIRAP, and involves prolonged JNK activation in vivo. Thus, TIRAP-dependent pathways may be targeted to predict and prevent inflammation-mediated IADRs. -- Highlights: ► Inflammation augments the toxicity of an idiosyncratic hepatotoxin chlorpromazine. ► Activation of Toll-like receptors by LPS or LTA induces chlorpromazine toxicity. ► Sustained stress kinase (JNK) activation is associated with chlorpromazine toxicity. ► These studies

  2. Distinct energetics and closing pathways for DNA polymerase β with 8-oxoG template and different incoming nucleotides

    Directory of Open Access Journals (Sweden)

    Wang Yanli

    2007-02-01

    Full Text Available Abstract Background 8-Oxoguanine (8-oxoG is a common oxidative lesion frequently encountered by DNA polymerases such as the repair enzyme DNA polymerase β (pol β. To interpret in atomic and energetic detail how pol β processes 8-oxoG, we apply transition path sampling to delineate closing pathways of pol β 8-oxoG complexes with dCTP and dATP incoming nucleotides and compare the results to those of the nonlesioned G:dCTP and G:dATPanalogues. Results Our analyses show that the closing pathways of the 8-oxoG complexes are different from one another and from the nonlesioned analogues in terms of the individual transition states along each pathway, associated energies, and the stability of each pathway's closed state relative to the corresponding open state. In particular, the closed-to-open state stability difference in each system establishes a hierarchy of stability (from high to low as G:C > 8-oxoG:C > 8-oxoG:A > G:A, corresponding to -3, -2, 2, 9 kBT, respectively. This hierarchy of closed state stability parallels the experimentally observed processing efficiencies for the four pairs. Network models based on the calculated rate constants in each pathway indicate that the closed species are more populated than the open species for 8-oxoG:dCTP, whereas the opposite is true for 8-oxoG:dATP. Conclusion These results suggest that the lower insertion efficiency (larger Km for dATP compared to dCTP opposite 8-oxoG is caused by a less stable closed-form of pol β, destabilized by unfavorable interactions between Tyr271 and the mispair. This stability of the closed vs. open form can also explain the higher insertion efficiency for 8-oxoG:dATP compared to the nonlesioned G:dATP pair, which also has a higher overall conformational barrier. Our study offers atomic details of the complexes at different states, in addition to helping interpret the different insertion efficiencies of dATP and dCTP opposite 8-oxoG and G.

  3. Distinct evolution of TLR-mediated dendritic cell cytokine secretion in patients with limited and diffuse cutaneous systemic sclerosis.

    NARCIS (Netherlands)

    Bon, L. van; Popa, C.; Huibens, R.J.F.; Vonk, M.C.; York, M.; Simms, R.; Hesselstrand, R.; Wuttge, D.M.; Lafyatis, R.; Radstake, T.R.D.J.

    2010-01-01

    BACKGROUND: Systemic sclerosis (SSc) is an autoimmune disease and accumulating evidence suggests a role for Toll-like receptor (TLR)-mediated activation of dendritic cells (DCs). OBJECTIVE: To map TLR-mediated cytokine responses of DCs from patients with SSc. METHODS: 45 patients with SSc were

  4. RhoA/ROCK Signaling Pathway Mediates Shuanghuanglian Injection-Induced Pseudo-allergic Reactions.

    Science.gov (United States)

    Han, Jiayin; Zhao, Yong; Zhang, Yushi; Li, Chunying; Yi, Yan; Pan, Chen; Tian, Jingzhuo; Yang, Yifei; Cui, Hongyu; Wang, Lianmei; Liu, Suyan; Liu, Jing; Deng, Nuo; Liang, Aihua

    2018-01-01

    SHLI-induced hypersensitivity reactions in both endothelial cells and mice indicating its protective effect. SHLI-induced pseudo-allergic reactions were mediated by the activation of the RhoA/ROCK signaling pathway. Conclusion : This study presents a novel mechanism of SHLI-induced immediate hypersensitivity reactions and suggests a potential therapeutic strategy to prevent the associated adverse reactions.

  5. Distinct UV-B and UV-A/blue light signal transduction pathways induce chalcone synthase gene expression in Arabidopsis cells

    International Nuclear Information System (INIS)

    Christie, J.M.; Jenkins, G.I.

    1996-01-01

    UV and blue light control the expression of flavonoid biosynthesis genes in a range of higher plants. To investigate the signal transduction processes involved in the induction of chalcone synthase (CHS) gene expression by UV-B and UV-A/blue light, we examined the, effects of specific agonists and inhibitors of known signaling components in mammalian systems in a photomixotrophic Arabidopsis cell suspension culture. CHS expression is induced specifically by these wavelengths in the cell culture, in a manner similar to that in mature Arabidopsis leaf tissue. Both the UV-B and UV-A/blue phototransduction processes involve calcium, although the elevation of cytosolic calcium is insufficient on its own to stimulate CHS expression. The UV-A/blue light induction of CHS expression does not appear to involve calmodulin, whereas the UV-B response does; this difference indicates that the signal transduction pathways are, at least in part, distinct. We provide evidence that both pathways involve reversible protein phosphorylation and require protein synthesis. The UV-B and UV-A/blue light signaling pathways are therefore different from the phytochrome signal transduction pathway regulating CHS expression in other species

  6. Feedback regulation on PTEN/AKT pathway by the ER stress kinase PERK mediated by interaction with the Vault complex

    DEFF Research Database (Denmark)

    Zhang, Wei; Neo, Suat Peng; Gunaratne, Jayantha

    2015-01-01

    The high proliferation rate of cancer cells, together with environmental factors such as hypoxia and nutrient deprivation can cause Endoplasmic Reticulum (ER) stress. The protein kinase PERK is an essential mediator in one of the three ER stress response pathways. Genetic and pharmacological inhi...

  7. Host cell virus entry mediated by Australian bat lyssavirus G envelope glycoprotein occurs through a clathrin-mediated endocytic pathway that requires actin and Rab5.

    Science.gov (United States)

    Weir, Dawn L; Laing, Eric D; Smith, Ina L; Wang, Lin-Fa; Broder, Christopher C

    2014-02-27

    Australian bat lyssavirus (ABLV), a rhabdovirus of the genus Lyssavirus which circulates in both pteropid fruit bats and insectivorous bats in mainland Australia, has caused three fatal human infections, the most recent in February 2013, manifested as acute neurological disease indistinguishable from clinical rabies. Rhabdoviruses infect host cells through receptor-mediated endocytosis and subsequent pH-dependent fusion mediated by their single envelope glycoprotein (G), but the specific host factors and pathways involved in ABLV entry have not been determined. ABLV internalization into HEK293T cells was examined using maxGFP-encoding recombinant vesicular stomatitis viruses (rVSV) that express ABLV G glycoproteins. A combination of chemical and molecular approaches was used to investigate the contribution of different endocytic pathways to ABLV entry. Dominant negative Rab GTPases were used to identify the endosomal compartment utilized by ABLV to gain entry into the host cell cytosol. Here we show that ABLV G-mediated entry into HEK293T cells was significantly inhibited by the dynamin-specific inhibitor dynasore, chlorpromazine, a drug that blocks clathrin-mediated endocytosis, and the actin depolymerizing drug latrunculin B. Over expression of dominant negative mutants of Eps15 and Rab5 also significantly reduced ABLV G-mediated entry into HEK293T cells. Chemical inhibitors of caveolae-dependent endocytosis and macropinocytosis and dominant negative mutants of Rab7 and Rab11 had no effect on ABLV entry. The predominant pathway utilized by ABLV for internalization into HEK293T cells is clathrin-and actin-dependent. The requirement of Rab5 for productive infection indicates that ABLV G-mediated fusion occurs within the early endosome compartment.

  8. Transgenic Mouse Lines Subdivide External Segment of the Globus Pallidus (GPe) Neurons and Reveal Distinct GPe Output Pathways

    Science.gov (United States)

    Mastro, Kevin J.; Bouchard, Rachel S.; Holt, Hiromi A. K.

    2014-01-01

    Cell-type diversity in the brain enables the assembly of complex neural circuits, whose organization and patterns of activity give rise to brain function. However, the identification of distinct neuronal populations within a given brain region is often complicated by a lack of objective criteria to distinguish one neuronal population from another. In the external segment of the globus pallidus (GPe), neuronal populations have been defined using molecular, anatomical, and electrophysiological criteria, but these classification schemes are often not generalizable across preparations and lack consistency even within the same preparation. Here, we present a novel use of existing transgenic mouse lines, Lim homeobox 6 (Lhx6)–Cre and parvalbumin (PV)–Cre, to define genetically distinct cell populations in the GPe that differ molecularly, anatomically, and electrophysiologically. Lhx6–GPe neurons, which do not express PV, are concentrated in the medial portion of the GPe. They have lower spontaneous firing rates, narrower dynamic ranges, and make stronger projections to the striatum and substantia nigra pars compacta compared with PV–GPe neurons. In contrast, PV–GPe neurons are more concentrated in the lateral portions of the GPe. They have narrower action potentials, deeper afterhyperpolarizations, and make stronger projections to the subthalamic nucleus and parafascicular nucleus of the thalamus. These electrophysiological and anatomical differences suggest that Lhx6–GPe and PV–GPe neurons participate in different circuits with the potential to contribute to different aspects of motor function and dysfunction in disease. PMID:24501350

  9. DMPD: IRAK1: a critical signaling mediator of innate immunity. [Dynamic Macrophage Pathway CSML Database

    Lifescience Database Archive (English)

    Full Text Available 17890055 IRAK1: a critical signaling mediator of innate immunity. Gottipati S, Rao ...IRAK1: a critical signaling mediator of innate immunity. PubmedID 17890055 Title IRAK1: a critical signaling mediator

  10. Analysis of Human TAAR8 and Murine Taar8b Mediated Signaling Pathways and Expression Profile

    Directory of Open Access Journals (Sweden)

    Jessica Mühlhaus

    2014-11-01

    Full Text Available The thyroid hormone derivative 3-iodothyronamine (3-T1AM exerts metabolic effects in vivo that contradict known effects of thyroid hormones. 3-T1AM acts as a trace amine-associated receptor 1 (TAAR1 agonist and activates Gs signaling in vitro. Interestingly, 3-T1AM-meditated in vivo effects persist in Taar1 knockout-mice indicating that further targets of 3-T1AM might exist. Here, we investigated another member of the TAAR family, the only scarcely studied mouse and human trace-amine-associated receptor 8 (Taar8b, TAAR8. By RT-qPCR and locked-nucleic-acid (LNA in situ hybridization, Taar8b expression in different mouse tissues was analyzed. Functionally, we characterized TAAR8 and Taar8b with regard to cell surface expression and signaling via different G-protein-mediated pathways. Cell surface expression was verified by ELISA, and cAMP accumulation was quantified by AlphaScreen for detection of Gs and/or Gi/o signaling. Activation of G-proteins Gq/11 and G12/13 was analyzed by reporter gene assays. Expression analyses revealed at most marginal Taar8b expression and no gender differences for almost all analyzed tissues. In heart, LNA-in situ hybridization demonstrated the absence of Taar8b expression. We could not identify 3-T1AM as a ligand for TAAR8 and Taar8b, but both receptors were characterized by a basal Gi/o signaling activity, a so far unknown signaling pathway for TAARs.

  11. HMGB1 mediates depressive behavior induced by chronic stress through activating the kynurenine pathway.

    Science.gov (United States)

    Wang, Bo; Lian, Yong-Jie; Su, Wen-Jun; Peng, Wei; Dong, Xin; Liu, Lin-Lin; Gong, Hong; Zhang, Ting; Jiang, Chun-Lei; Wang, Yun-Xia

    2017-11-28

    Our previous study has reported that the proactive secretion and role of central high mobility group box 1 (HMGB1) in lipopolysaccharide-induced depressive behavior. Here, the potential mechanism of HMGB1 mediating chronic-stress-induced depression through the kynurenine pathway (KP) was further explored both in vivo and in vitro. Depression model was established with the 4-week chronic unpredictable mild stress (CUMS). Sucrose preference and Barnes maze test were performed to reflect depressive behaviors. The ratio of kynurenine (KYN)/tryptophan (Trp) represented the enzyme activity of indoleamine-2,3-dioxygenase (IDO). Gene transcription and protein expression were assayed by real-time RT-PCR and western-blot or ELISA kit respectively. Along with depressive behaviors, HMGB1 concentrations in the hippocampus and serum substantially increased post 4-week CUMS exposure. Concurrent with the upregulated HMGB1 protein, the regulator of translocation of HMGB1, sirtuin 1 (SIRT1) concentration in the hippocampus remarkably increased. In addition to HMGB1 and SIRT1, IDO, the rate limiting enzyme of KP, was upregulated at the level of mRNA expression and enzyme activity in stressed hippocampi and LPS/HMGB1-treated hippocampal slices. The gene transcription of kynurenine monooxygenase (KMO) and kynureninase (KYNU) in the downstream of KP also increased both in vivo and in vitro. Mice treated with ethyl pyruvate (EP), the inhibitor of HMGB1 releasing, were observed with lower tendency of developing depressive behaviors and reduced activation of enzymes in KP. All of these experiments demonstrate that the role of HMGB1 on the induction of depressive behavior is mediated by KP activation. Copyright © 2017 Elsevier Inc. All rights reserved.

  12. A convergent and essential interneuron pathway for Mauthner-cell-mediated escapes.

    Science.gov (United States)

    Lacoste, Alix M B; Schoppik, David; Robson, Drew N; Haesemeyer, Martin; Portugues, Ruben; Li, Jennifer M; Randlett, Owen; Wee, Caroline L; Engert, Florian; Schier, Alexander F

    2015-06-01

    The Mauthner cell (M-cell) is a command-like neuron in teleost fish whose firing in response to aversive stimuli is correlated with short-latency escapes [1-3]. M-cells have been proposed as evolutionary ancestors of startle response neurons of the mammalian reticular formation [4], and studies of this circuit have uncovered important principles in neurobiology that generalize to more complex vertebrate models [3]. The main excitatory input was thought to originate from multisensory afferents synapsing directly onto the M-cell dendrites [3]. Here, we describe an additional, convergent pathway that is essential for the M-cell-mediated startle behavior in larval zebrafish. It is composed of excitatory interneurons called spiral fiber neurons, which project to the M-cell axon hillock. By in vivo calcium imaging, we found that spiral fiber neurons are active in response to aversive stimuli capable of eliciting escapes. Like M-cell ablations, bilateral ablations of spiral fiber neurons largely eliminate short-latency escapes. Unilateral spiral fiber neuron ablations shift the directionality of escapes and indicate that spiral fiber neurons excite the M-cell in a lateralized manner. Their optogenetic activation increases the probability of short-latency escapes, supporting the notion that spiral fiber neurons help activate M-cell-mediated startle behavior. These results reveal that spiral fiber neurons are essential for the function of the M-cell in response to sensory cues and suggest that convergent excitatory inputs that differ in their input location and timing ensure reliable activation of the M-cell, a feedforward excitatory motif that may extend to other neural circuits. Copyright © 2015 Elsevier Ltd. All rights reserved.

  13. 25-Hydroxycholesterol promotes fibroblast-mediated tissue remodeling through NF-κB dependent pathway

    Energy Technology Data Exchange (ETDEWEB)

    Ichikawa, Tomohiro [Third Department of Internal Medicine, Wakayama Medical University, School of Medicine, 811-1 Kimiidera, Wakayama 641-8509 (Japan); Sugiura, Hisatoshi, E-mail: sugiura@rm.med.tohoku.ac.jp [Department of Respiratory Medicine, Tohoku University Graduate School of Medicine, 1-1 Seiryo-machi, Aoba-ku, Sendai 980-8574 (Japan); Koarai, Akira; Kikuchi, Takashi; Hiramatsu, Masataka; Kawabata, Hiroki; Akamatsu, Keiichiro; Hirano, Tsunahiko; Nakanishi, Masanori; Matsunaga, Kazuto; Minakata, Yoshiaki [Third Department of Internal Medicine, Wakayama Medical University, School of Medicine, 811-1 Kimiidera, Wakayama 641-8509 (Japan); Ichinose, Masakazu [Department of Respiratory Medicine, Tohoku University Graduate School of Medicine, 1-1 Seiryo-machi, Aoba-ku, Sendai 980-8574 (Japan)

    2013-05-01

    Abnormal structural alterations termed remodeling, including fibrosis and alveolar wall destruction, are important features of the pathophysiology of chronic airway diseases such as chronic obstructive pulmonary disease (COPD) and asthma. 25-hydroxycholesterol (25-HC) is enzymatically produced by cholesterol 25-hydorxylase (CH25H) in macrophages and is reported to be involved in the formation of arteriosclerosis. We previously demonstrated that the expression of CH25H and production of 25HC were increased in the lungs of COPD. However, the role of 25-HC in lung tissue remodeling is unknown. In this study, we investigated the effect of 25-HC on fibroblast-mediated tissue remodeling using human fetal lung fibroblasts (HFL-1) in vitro. 25-HC significantly augmented α-smooth muscle actin (SMA) (P<0.001) and collagen I (P<0.001) expression in HFL-1. 25-HC also significantly enhanced the release and activation of matrix metallaoproteinase (MMP)-2 (P<0.001) and MMP-9 (P<0.001) without any significant effect on the production of tissue inhibitor of metalloproteinase (TIMP)-1 and TIMP-2. 25-HC stimulated transforming growth factor (TGF)-β{sub 1} production (P<0.01) and a neutralizing anti-TGF-β antibody restored these 25-HC-augmented pro-fibrotic responses. 25-HC significantly promoted the translocation of nuclear factor (NF)-κB p65 into the nuclei (P<0.01), but not phospholylated-c-jun, a complex of activator protein-1. Pharmacological inhibition of NF-κB restored the 25-HC-augmented pro-fibrotic responses and TGF-β{sub 1} release. These results suggest that 25-HC could contribute to fibroblast-mediated lung tissue remodeling by promoting myofibroblast differentiation and the excessive release of extracellular matrix protein and MMPs via an NF-κB-TGF-β dependent pathway.

  14. Light-Mediated Kinetic Control Reveals the Temporal Effect of the Raf/MEK/ERK Pathway in PC12 Cell Neurite Outgrowth

    Science.gov (United States)

    Zhang, Kai; Duan, Liting; Ong, Qunxiang; Lin, Ziliang; Varman, Pooja Mahendra; Sung, Kijung; Cui, Bianxiao

    2014-01-01

    It has been proposed that differential activation kinetics allows cells to use a common set of signaling pathways to specify distinct cellular outcomes. For example, nerve growth factor (NGF) and epidermal growth factor (EGF) induce different activation kinetics of the Raf/MEK/ERK signaling pathway and result in differentiation and proliferation, respectively. However, a direct and quantitative linkage between the temporal profile of Raf/MEK/ERK activation and the cellular outputs has not been established due to a lack of means to precisely perturb its signaling kinetics. Here, we construct a light-gated protein-protein interaction system to regulate the activation pattern of the Raf/MEK/ERK signaling pathway. Light-induced activation of the Raf/MEK/ERK cascade leads to significant neurite outgrowth in rat PC12 pheochromocytoma cell lines in the absence of growth factors. Compared with NGF stimulation, light stimulation induces longer but fewer neurites. Intermittent on/off illumination reveals that cells achieve maximum neurite outgrowth if the off-time duration per cycle is shorter than 45 min. Overall, light-mediated kinetic control enables precise dissection of the temporal dimension within the intracellular signal transduction network. PMID:24667437

  15. BRAF and RAS oncogenes regulate Rho GTPase pathways to mediate migration and invasion properties in human colon cancer cells: a comparative study

    Directory of Open Access Journals (Sweden)

    Shirasawa Senji

    2011-09-01

    Full Text Available Abstract Background Colorectal cancer is a common disease that involves genetic alterations, such as inactivation of tumour suppressor genes and activation of oncogenes. Among them are RAS and BRAF mutations, which rarely coexist in the same tumour. Individual members of the Rho (Ras homology GTPases contribute with distinct roles in tumour cell morphology, invasion and metastasis. The aim of this study is to dissect cell migration and invasion pathways that are utilised by BRAFV600E as compared to KRASG12V and HRASG12V oncoproteins. In particular, the role of RhoA (Ras homolog gene family, member A, Rac1 (Ras-related C3 botulinum toxin substrate 1 and Cdc42 (cell division cycle 42 in cancer progression induced by each of the three oncogenes is described. Methods Colon adenocarcinoma cells with endogenous as well as ectopically expressed or silenced oncogenic mutations of BRAFV600E, KRASG12V and HRASG12V were employed. Signalling pathways and Rho GTPases were inhibited with specific kinase inhibitors and siRNAs. Cell motility and invasion properties were correlated with cytoskeletal properties and Rho GTPase activities. Results Evidence presented here indicate that BRAFV600E significantly induces cell migration and invasion properties in vitro in colon cancer cells, at least in part through activation of RhoA GTPase. The relationship established between BRAFV600E and RhoA activation is mediated by the MEK-ERK pathway. In parallel, KRASG12V enhances the ability of colon adenocarcinoma cells Caco-2 to migrate and invade through filopodia formation and PI3K-dependent Cdc42 activation. Ultimately increased cell migration and invasion, mediated by Rac1, along with the mesenchymal morphology obtained through the Epithelial-Mesenchymal Transition (EMT were the main characteristics rendered by HRASG12V in Caco-2 cells. Moreover, BRAF and KRAS oncogenes are shown to cooperate with the TGFβ-1 pathway to provide cells with additional transforming

  16. Role of the Nrf2-heme oxygenase-1 pathway in silver nanoparticle-mediated cytotoxicity

    International Nuclear Information System (INIS)

    Kang, Su Jin; Ryoo, In-geun; Lee, Young Joon; Kwak, Mi-Kyoung

    2012-01-01

    Silver nanoparticles (nano-Ag) have been widely used in various commercial products including textiles, electronic appliances and biomedical products. However, there remains insufficient information on the potential risk of nano-Ag to human health and environment. In the current study, we have investigated the role of NF-E2-related factor 2 (Nrf2) transcription factor in nano-Ag-induced cytotoxicity. When Nrf2 expression was blocked using interring RNA expression in ovarian carcinoma cell line, nano-Ag treatment showed a substantial decrease in cell viability with concomitant increases in apoptosis and DNA damage compared to the control cells. Target gene analysis revealed that the expression of heme oxygenase-1 (HO-1) was highly elevated by nano-Ag in nonspecific shRNA expressing cells, while Nrf2 knockdown cells (NRF2i) did not increase HO-1 expression. The role of HO-1 in cytoprotection against nano-Ag was reinforced by results using pharmacological inducer of HO-1: cobalt protoporphyrin-mediated HO-1 activation in the NRF2i cells prevented nano-Ag-mediated cell death. Similarly, pharmacological or genetic inhibition of HO-1 in nonspecific control cells exacerbated nano-Ag toxicity. As the upstream signaling mechanism, nano-Ag required the phosphoinositide 3-kinase (PI3K) and p38MAPK signaling cascades for HO-1 induction. The treatment with either PI3K inhibitor or p38MAPK inhibitor suppressed HO-1 induction and intensified nano-Ag-induced cell death. Taken together, these results suggest that Nrf2-dependent HO-1 up-regulation plays a protective role in nano-Ag-induced DNA damage and consequent cell death. In addition, nano-Ag-mediated HO-1 induction is associated with the PI3K and p38MAPK signaling pathways. -- Highlights: ► Role of Nrf2 signaling in silver nanoparticle toxicity. ► Silver nanoparticle toxicity is increased by Nrf2 blockade. ► Nrf2-dependent HO-1 induction protects cells from silver nanoparticle toxicity. ► PI3K and p38MAPK cascades are

  17. Kidins220/ARMS as a functional mediator of multiple receptor signalling pathways.

    Science.gov (United States)

    Neubrand, Veronika E; Cesca, Fabrizia; Benfenati, Fabio; Schiavo, Giampietro

    2012-04-15

    An increasing body of evidence suggests that several membrane receptors--in addition to activating distinct signalling cascades--also engage in substantial crosstalk with each other, thereby adjusting their signalling outcome as a function of specific input information. However, little is known about the molecular mechanisms that control their coordination and integration of downstream signalling. A protein that is likely to have a role in this process is kinase-D-interacting substrate of 220 kDa [Kidins220, also known as ankyrin repeat-rich membrane spanning (ARMS), hereafter referred to as Kidins220/ARMS]. Kidins220/ARMS is a conserved membrane protein that is preferentially expressed in the nervous system and interacts with the microtubule and actin cytoskeleton. It interacts with neurotrophin, ephrin, vascular endothelial growth factor (VEGF) and glutamate receptors, and is a common downstream target of several trophic stimuli. Kidins220/ARMS is required for neuronal differentiation and survival, and its expression levels modulate synaptic plasticity. Kidins220/ARMS knockout mice show developmental defects mainly in the nervous and cardiovascular systems, suggesting a crucial role for this protein in modulating the cross talk between different signalling pathways. In this Commentary, we summarise existing knowledge regarding the physiological functions of Kidins220/ARMS, and highlight some interesting directions for future studies on the role of this protein in health and disease.

  18. Homocysteine elicits an M1 phenotype in murine macrophages through an EMMPRIN-mediated pathway.

    Science.gov (United States)

    Winchester, Lee J; Veeranki, Sudhakar; Givvimani, Srikanth; Tyagi, Suresh C

    2015-07-01

    Hyperhomocysteinemia (HHcy) is associated with inflammatory diseases and is known to increase the production of reactive oxygen species (ROS), matrix metalloproteinase (MMP)-9, and inducible nitric oxide synthase, and to decrease endothelial nitric oxide production. However, the impact of HHcy on macrophage phenotype differentiation is not well-established. It has been documented that macrophages have 2 distinct phenotypes: the "classically activated/destructive" (M1), and the "alternatively activated/constructive" (M2) subtypes. We hypothesize that HHcy increases M1 macrophage differentiation through extracellular matrix metalloproteinase inducer (EMMPRIN), a known inducer of matrix metalloproteinases. murine J774A.1 and Raw 264.7 macrophages were treated with 100 and 500 μmol/L Hcy, respectively, for 24 h. Samples were analyzed using Western blotting and immunocytochemistry. Homocysteine treatment increased cluster of differentiation 40 (CD40; M1 marker) in J774A.1 and Raw 264.7 macrophages. MMP-9 was induced in both cell lines. EMMPRIN protein expression was also increased in both cell lines. Blocking EMMPRIN function by pre-treating cells with anti-EMMPRIN antibody, with or without Hcy, resulted in significantly lower expression of CD40 in both cell lines by comparison with the controls. A DCFDA assay demonstrated increased ROS production in both cell lines with Hcy treatment when compared with the controls. Our results suggest that HHcy results in an increase of the M1 macrophage phenotype. This effect seems to be at least partially mediated by EMMPRIN induction.

  19. Suppression of the auxin response pathway enhances susceptibility to Phytophthora cinnamomi while phosphite-mediated resistance stimulates the auxin signalling pathway

    Science.gov (United States)

    2014-01-01

    Background Phytophthora cinnamomi is a devastating pathogen worldwide and phosphite (Phi), an analogue of phosphate (Pi) is highly effective in the control of this pathogen. Phi also interferes with Pi starvation responses (PSR), of which auxin signalling is an integral component. In the current study, the involvement of Pi and the auxin signalling pathways in host and Phi-mediated resistance to P. cinnamomi was investigated by screening the Arabidopsis thaliana ecotype Col-0 and several mutants defective in PSR and the auxin response pathway for their susceptibility to this pathogen. The response to Phi treatment was also studied by monitoring its effect on Pi- and the auxin response pathways. Results Here we demonstrate that phr1-1 (phosphate starvation response 1), a mutant defective in response to Pi starvation was highly susceptible to P. cinnamomi compared to the parental background Col-0. Furthermore, the analysis of the Arabidopsis tir1-1 (transport inhibitor response 1) mutant, deficient in the auxin-stimulated SCF (Skp1 − Cullin − F-Box) ubiquitination pathway was also highly susceptible to P. cinnamomi and the susceptibility of the mutants rpn10 and pbe1 further supported a role for the 26S proteasome in resistance to P. cinnamomi. The role of auxin was also supported by a significant (P < 0.001) increase in susceptibility of blue lupin (Lupinus angustifolius) to P. cinnamomi following treatment with the inhibitor of auxin transport, TIBA (2,3,5-triiodobenzoic acid). Given the apparent involvement of auxin and PSR signalling in the resistance to P. cinnamomi, the possible involvement of these pathways in Phi mediated resistance was also investigated. Phi (especially at high concentrations) attenuates the response of some Pi starvation inducible genes such as AT4, AtACP5 and AtPT2 in Pi starved plants. However, Phi enhanced the transcript levels of PHR1 and the auxin responsive genes (AUX1, AXR1and AXR2), suppressed the primary root

  20. Novel Hematopoietic Target Genes in the NRF2-Mediated Transcriptional Pathway

    Directory of Open Access Journals (Sweden)

    Michelle R. Campbell

    2013-01-01

    Full Text Available Nuclear factor- (erythroid-derived 2 like 2 (NFE2L2, NRF2 is a key transcriptional activator of the antioxidant response pathway and is closely related to erythroid transcription factor NFE2. Under oxidative stress, NRF2 heterodimerizes with small Maf proteins and binds cis-acting enhancer sequences found near oxidative stress response genes. Using the dietary isothiocyanate sulforaphane (SFN to activate NRF2, chromatin immunoprecipitation sequencing (ChIP-seq identified several hundred novel NRF2-mediated targets beyond its role in oxidative stress. Activated NRF2 bound the antioxidant response element (ARE in promoters of several known and novel target genes involved in iron homeostasis and heme metabolism, including known targets FTL and FTH1, as well as novel binding in the globin locus control region. Five novel NRF2 target genes were chosen for followup: AMBP, ABCB6, FECH, HRG-1 (SLC48A1, and TBXAS1. SFN-induced gene expression in erythroid K562 and lymphoid cells were compared for each target gene. NRF2 silencing showed reduced expression in lymphoid, lung, and hepatic cells. Furthermore, stable knockdown of NRF2 negative regulator KEAP1 in K562 cells resulted in increased NQO1, AMBP, and TBXAS1 expression. NFE2 binding sites in K562 cells revealed similar binding profiles as lymphoid NRF2 sites in all potential NRF2 candidates supporting a role for NRF2 in heme metabolism and erythropoiesis.

  1. O-GlcNAc regulates NEDD4-1 stability via caspase-mediated pathway

    International Nuclear Information System (INIS)

    Jiang, Kuan; Bai, Bingyang; Ta, Yajie; Zhang, Tingling; Xiao, Zikang; Wang, Peng George; Zhang, Lianwen

    2016-01-01

    O-GlcNAc modification of cytosolic and nuclear proteins regulates essential cellular processes such as stress responses, transcription, translation, and protein degradation. Emerging evidence indicates O-GlcNAcylation has a dynamic interplay with ubiquitination in cellular regulation. Here, we report that O-GlcNAc indirectly targets a vital E3 ubiquitin ligase enzyme of NEDD4-1. The protein level of NEDD4-1 is accordingly decreased following an increase of overall O-GlcNAc level upon PUGNAc or glucosamine stimulation. O-GlcNAc transferase (OGT) knockdown, overexpression and mutation results confirm that the stability of NEDD4-1 is negatively regulated by cellular O-GlcNAc. Moreover, the NEDD4-1 degradation induced by PUGNAc or GlcN is significantly inhibited by the caspase inhibitor. Our study reveals a regulation mechanism of NEDD4-1 stability by O-GlcNAcylation. - Highlights: • Reduced NEDD4-1 correlates with increased overall O-GlcNAc level. • OGT negatively regulates NEDD4-1 stability. • O-GlcNAc regulates NEDD4-1 through caspase-mediated pathway.

  2. Gut microbiome remodeling induces depressive-like behaviors through a pathway mediated by the host's metabolism.

    Science.gov (United States)

    Zheng, P; Zeng, B; Zhou, C; Liu, M; Fang, Z; Xu, X; Zeng, L; Chen, J; Fan, S; Du, X; Zhang, X; Yang, D; Yang, Y; Meng, H; Li, W; Melgiri, N D; Licinio, J; Wei, H; Xie, P

    2016-06-01

    Major depressive disorder (MDD) is the result of complex gene-environment interactions. According to the World Health Organization, MDD is the leading cause of disability worldwide, and it is a major contributor to the overall global burden of disease. However, the definitive environmental mechanisms underlying the pathophysiology of MDD remain elusive. The gut microbiome is an increasingly recognized environmental factor that can shape the brain through the microbiota-gut-brain axis. We show here that the absence of gut microbiota in germ-free (GF) mice resulted in decreased immobility time in the forced swimming test relative to conventionally raised healthy control mice. Moreover, from clinical sampling, the gut microbiotic compositions of MDD patients and healthy controls were significantly different with MDD patients characterized by significant changes in the relative abundance of Firmicutes, Actinobacteria and Bacteroidetes. Fecal microbiota transplantation of GF mice with 'depression microbiota' derived from MDD patients resulted in depression-like behaviors compared with colonization with 'healthy microbiota' derived from healthy control individuals. Mice harboring 'depression microbiota' primarily exhibited disturbances of microbial genes and host metabolites involved in carbohydrate and amino acid metabolism. This study demonstrates that dysbiosis of the gut microbiome may have a causal role in the development of depressive-like behaviors, in a pathway that is mediated through the host's metabolism.

  3. Nano Copper Induces Apoptosis in PK-15 Cells via a Mitochondria-Mediated Pathway.

    Science.gov (United States)

    Zhang, Hui; Chang, Zhenyu; Mehmood, Khalid; Abbas, Rao Zahid; Nabi, Fazul; Rehman, Mujeeb Ur; Wu, Xiaoxing; Tian, Xinxin; Yuan, Xiaodan; Li, Zhaoyang; Zhou, Donghai

    2018-01-01

    Nano-sized copper particles are widely used in various chemical, physical, and biological fields. However, earlier studies have shown that nano copper particles (40-100 μg/mL) can induce cell toxicity and apoptosis. Therefore, this study was conducted to investigate the role of nano copper in mitochondrion-mediated apoptosis in PK-15 cells. The cells were treated with different doses of nano copper (20, 40, 60, and 80 μg/mL) to determine the effects of apoptosis using acridine orange/ethidium bromide (AO/EB) fluorescence staining and a flow cytometry assay. The levels of malondialdehyde (MDA) and superoxide dismutase (SOD) in the PK-15 cells were examined using commercially available kits. Moreover, the mRNA levels of the Bax, Bid, Caspase-3, and CYCS genes were assessed by real-time PCR. The results revealed that nano copper exposure induced apoptosis and changed the mitochondrial membrane potential. In addition, nano copper significantly altered the levels of the Bax, Bid, Caspase-3, and CYCS genes at a concentration of 40 μg/mL. To summarize, nano copper significantly (P nano copper can play an important role in inducing the apoptotic pathway in PK-15 cells, which may be the mechanism by which nano copper induces nephrotoxicity.

  4. The OXI1 kinase pathway mediates Piriformospora indica-induced growth promotion in Arabidopsis.

    Directory of Open Access Journals (Sweden)

    Iris Camehl

    2011-05-01

    Full Text Available Piriformospora indica is an endophytic fungus that colonizes roots of many plant species and promotes growth and resistance to certain plant pathogens. Despite its potential use in agriculture, little is known on the molecular basis of this beneficial plant-fungal interaction. In a genetic screen for plants, which do not show a P. indica- induced growth response, we isolated an Arabidopsis mutant in the OXI1 (Oxidative Signal Inducible1 gene. OXI1 has been characterized as a protein kinase which plays a role in pathogen response and is regulated by H₂O₂ and PDK1 (3-PHOSPHOINOSITIDE-DEPENDENT PROTEIN KINASE1. A genetic analysis showed that double mutants of the two closely related PDK1.1 and PDK1.2 genes are defective in the growth response to P. indica. While OXI1 and PDK1 gene expression is upregulated in P. indica-colonized roots, defense genes are downregulated, indicating that the fungus suppresses plant defense reactions. PDK1 is activated by phosphatidic acid (PA and P. indica triggers PA synthesis in Arabidopsis plants. Under beneficial co-cultivation conditions, H₂O₂ formation is even reduced by the fungus. Importantly, phospholipase D (PLDα1 or PLDδ mutants, which are impaired in PA synthesis do not show growth promotion in response to fungal infection. These data establish that the P. indica-stimulated growth response is mediated by a pathway consisting of the PLD-PDK1-OXI1 cascade.

  5. UV-B Radiation Induces Root Bending Through the Flavonoid-Mediated Auxin Pathway in Arabidopsis.

    Science.gov (United States)

    Wan, Jinpeng; Zhang, Ping; Wang, Ruling; Sun, Liangliang; Wang, Wenying; Zhou, Huakun; Xu, Jin

    2018-01-01

    Ultraviolet (UV)-B radiation-induced root bending has been reported; however, the underlying mechanisms largely remain unclear. Here, we investigate whether and how auxin and flavonoids are involved in UV-B radiation-induced root bending in Arabidopsis using physiological, pharmacological, and genetic approaches. UV-B radiation modulated the direction of root growth by decreasing IAA biosynthesis and affecting auxin distribution in the root tips, where reduced auxin accumulation and asymmetric auxin distribution were observed. UV-B radiation increased the distribution of auxin on the nonradiated side of the root tips, promoting growth and causing root bending. Further analysis indicated that UV-B induced an asymmetric accumulation of flavonoids; this pathway is involved in modulating the accumulation and asymmetric distribution of auxin in root tips and the subsequent redirection of root growth by altering the distribution of auxin carriers in response to UV-B radiation. Taken together, our results indicate that UV-B radiation-induced root bending occurred through a flavonoid-mediated phototropic response to UV-B radiation.

  6. M2-like macrophages are responsible for collagen degradation through a mannose receptor–mediated pathway

    Science.gov (United States)

    Madsen, Daniel H.; Leonard, Daniel; Masedunskas, Andrius; Moyer, Amanda; Jürgensen, Henrik Jessen; Peters, Diane E.; Amornphimoltham, Panomwat; Selvaraj, Arul; Yamada, Susan S.; Brenner, David A.; Burgdorf, Sven; Engelholm, Lars H.; Behrendt, Niels; Holmbeck, Kenn; Weigert, Roberto

    2013-01-01

    Tissue remodeling processes critically depend on the timely removal and remodeling of preexisting collagen scaffolds. Nevertheless, many aspects related to the turnover of this abundant extracellular matrix component in vivo are still incompletely understood. We therefore took advantage of recent advances in optical imaging to develop an assay to visualize collagen turnover in situ and identify cell types and molecules involved in this process. Collagen introduced into the dermis of mice underwent cellular endocytosis in a partially matrix metalloproteinase–dependent manner and was subsequently routed to lysosomes for complete degradation. Collagen uptake was predominantly executed by a quantitatively minor population of M2-like macrophages, whereas more abundant Col1a1-expressing fibroblasts and Cx3cr1-expressing macrophages internalized collagen at lower levels. Genetic ablation of the collagen receptors mannose receptor (Mrc1) and urokinase plasminogen activator receptor–associated protein (Endo180 and Mrc2) impaired this intracellular collagen degradation pathway. This study demonstrates the importance of receptor-mediated cellular uptake to collagen turnover in vivo and identifies a key role of M2-like macrophages in this process. PMID:24019537

  7. Nuclear import of Nkx2-2 is mediated by multiple pathways

    International Nuclear Information System (INIS)

    Lin, Wenbo; Xu, PengPeng; Guo, YingYing; Jia, Qingjie; Tao, Tao

    2017-01-01

    Nkx2-2 homeoprotein is essential for the development of the central nervous system and pancreas. Although the nuclear localization signals of Nkx2-2 have been identified, the responsible transport receptor is still unknown. Here, we demonstrate that imp α1 not only interacts with Nkx2-2 but also transports it into the nucleus in vitro by acting together with imp β1. However, the nuclear import of Nkx2-2 in cells was not inhibited in response to knockdown expression of endogenous imp β1 or over-expression of Bimax2. Furthermore, imp β1 and imp 13, but not imp 4, directly interact with Nkx2-2 and are capable of transporting Nkx2-2 in an in vitro import assay. By GST pull-down assay, we demonstrate that mutation of NLS1 or NLS2 has no effect on interaction with imp α1 or imp 13, but significantly reduced binding to imp β1. Thus, the nuclear import of Nkx2-2 is mediated not only by the classical import pathway but also directly by imp β1 or imp 13.

  8. Diffused and sustained inhibitory effects of intestinal electrical stimulation on intestinal motility mediated via sympathetic pathway.

    Science.gov (United States)

    Zhao, Xiaotuan; Yin, Jieyun; Wang, Lijie; Chen, Jiande D Z

    2014-06-01

    The aims were to investigate the energy-dose response effect of intestinal electrical stimulation (IES) on small bowel motility, to compare the effect of forward and backward IES, and to explore the possibility of using intermittent IES and mechanism of IES on intestinal motility. Five dogs implanted with a duodenal cannula and one pair of intestinal serosal electrodes were studied in five sessions: 1) energy-dose response study; 2) forward IES; 3) backward IES; 4) intermittent IES vs. continuous IES; 5) administration of guanethidine. The contractile activity and tonic pressure of the small intestine were recorded. The duration of sustained effect after turning off IES was manually calculated. 1) IES with long pulse energy dose dependently inhibited contractile activity and tonic pressure of the small intestine (p intestine depended on the energy of IES delivered (p intestine. 5) Guanethidine blocked the inhibitory effect of IES on intestinal motility. IES with long pulses inhibits small intestinal motility; the effect is energy-dose dependent, diffused, and sustained. Intermittent IES has the same efficacy as the continuous IES in inhibiting small intestinal motility. Forward and backward IES have similar inhibitory effects on small bowel motility. This IES-induced inhibitory effect is mediated via the sympathetic pathway. © 2013 International Neuromodulation Society.

  9. Distinct Polymer Architecture Mediates Switching of Complement Activation Pathways at the Nanosphere-Serum Interface: Implications for Stealth Nanoparticle Engineering

    DEFF Research Database (Denmark)

    Hamad, I.; Al-Hanbali, O.; Hunter, A.C.

    2010-01-01

    Nanoparticles with surface projected polyethyleneoxide (PEO) chains in 'mushroom-brush' and "brush" configurations display stealth properties in systemic circulation and have numerous applications in site specific targeting for controlled drug delivery and release as well as diagnostic Imaging. W...... engineering and design of immunologically safer stealth and targetable nanosystems with polymers for use in clinical medicine....

  10. Linear and branched glyco-lipopeptide vaccines follow distinct cross-presentation pathways and generate different magnitudes of antitumor immunity.

    Directory of Open Access Journals (Sweden)

    Olivier Renaudet

    2010-06-01

    Full Text Available Glyco-lipopeptides, a form of lipid-tailed glyco-peptide, are currently under intense investigation as B- and T-cell based vaccine immunotherapy for many cancers. However, the cellular and molecular mechanisms of glyco-lipopeptides (GLPs immunogenicity and the position of the lipid moiety on immunogenicity and protective efficacy of GLPs remain to be determined.We have constructed two structural analogues of HER-2 glyco-lipopeptide (HER-GLP by synthesizing a chimeric peptide made of one universal CD4(+ epitope (PADRE and one HER-2 CD8(+ T-cell epitope (HER(420-429. The C-terminal end of the resulting CD4-CD8 chimeric peptide was coupled to a tumor carbohydrate B-cell epitope, based on a regioselectively addressable functionalized templates (RAFT, made of four alpha-GalNAc molecules. The resulting HER glyco-peptide (HER-GP was then linked to a palmitic acid moiety, attached either at the N-terminal end (linear HER-GLP-1 or in the middle between the CD4+ and CD8+ T cell epitopes (branched HER-GLP-2. We have investigated the uptake, processing and cross-presentation pathways of the two HER-GLP vaccine constructs, and assessed whether the position of linkage of the lipid moiety would affect the B- and T-cell immunogenicity and protective efficacy. Immunization of mice revealed that the linear HER-GLP-1 induced a stronger and longer lasting HER(420-429-specific IFN-gamma producing CD8(+ T cell response, while the branched HER-GLP-2 induced a stronger tumor-specific IgG response. The linear HER-GLP-1 was taken up easily by dendritic cells (DCs, induced stronger DCs maturation and produced a potent TLR- 2-dependent T-cell activation. The linear and branched HER-GLP molecules appeared to follow two different cross-presentation pathways. While regression of established tumors was induced by both linear HER-GLP-1 and branched HER-GLP-2, the inhibition of tumor growth was significantly higher in HER-GLP-1 immunized mice (p<0.005.These findings have

  11. Distinct abscisic acid signaling pathways for modulation of guard cell versus mesophyll cell potassium channels revealed by expression studies in Xenopus laevis oocytes

    Science.gov (United States)

    Sutton, F.; Paul, S. S.; Wang, X. Q.; Assmann, S. M.; Evans, M. L. (Principal Investigator)

    2000-01-01

    Regulation of guard cell ion transport by abscisic acid (ABA) and in particular ABA inhibition of a guard cell inward K(+) current (I(Kin)) is well documented. However, little is known concerning ABA effects on ion transport in other plant cell types. Here we applied patch clamp techniques to mesophyll cell protoplasts of fava bean (Vicia faba cv Long Pod) plants and demonstrated ABA inhibition of an outward K(+) current (I(Kout)). When mesophyll cell protoplast mRNA (mesophyll mRNA) was expressed in Xenopus laevis oocytes, I(Kout) was generated that displayed similar properties to I(Kout) observed from direct analysis of mesophyll cell protoplasts. I(Kout) expressed by mesophyll mRNA-injected oocytes was inhibited by ABA, indicating that the ABA signal transduction pathway observed in mesophyll cells was preserved in the frog oocytes. Co-injection of oocytes with guard cell protoplast mRNA and cRNA for KAT1, an inward K(+) channel expressed in guard cells, resulted in I(Kin) that was similarly inhibited by ABA. However, oocytes co-injected with mesophyll mRNA and KAT1 cRNA produced I(Kin) that was not inhibited by ABA. These results demonstrate that the mesophyll-encoded signaling mechanism could not substitute for the guard cell pathway. These findings indicate that mesophyll cells and guard cells use distinct and different receptor types and/or signal transduction pathways in ABA regulation of K(+) channels.

  12. β-Adrenergic Receptors Regulate the Acquisition and Consolidation Phases of Aversive Memory Formation Through Distinct, Temporally Regulated Signaling Pathways.

    Science.gov (United States)

    Schiff, Hillary C; Johansen, Joshua P; Hou, Mian; Bush, David E A; Smith, Emily K; Klein, JoAnna E; LeDoux, Joseph E; Sears, Robert M

    2017-03-01

    Memory formation requires the temporal coordination of molecular events and cellular processes following a learned event. During Pavlovian threat (fear) conditioning (PTC), sensory and neuromodulatory inputs converge on post-synaptic neurons within the lateral nucleus of the amygdala (LA). By activating an intracellular cascade of signaling molecules, these G-protein-coupled neuromodulatory receptors are capable of recruiting a diverse profile of plasticity-related proteins. Here we report that norepinephrine, through its actions on β-adrenergic receptors (βARs), modulates aversive memory formation following PTC through two molecularly and temporally distinct signaling mechanisms. Specifically, using behavioral pharmacology and biochemistry in adult rats, we determined that βAR activity during, but not after PTC training initiates the activation of two plasticity-related targets: AMPA receptors (AMPARs) for memory acquisition and short-term memory and extracellular regulated kinase (ERK) for consolidating the learned association into a long-term memory. These findings reveal that βAR activity during, but not following PTC sets in motion cascading molecular events for the acquisition (AMPARs) and subsequent consolidation (ERK) of learned associations.

  13. Do Trust and Sexual Intimacy Mediate Attachment's Pathway Toward Sexual Violence Occurring in Same Sex Romantic Relationships?

    Science.gov (United States)

    Gabbay, Nicolas; Lafontaine, Marie-France

    2017-07-01

    This study tested a serial mediation model examining how dyadic trust and sexual intimacy mediate the relationship between insecure romantic attachment and perpetrated sexual violence occurring between same sex intimate partners (sexual SSIPV). A community sample of adults ( N = 310; 203 women, 107 men) involved in a committed (6 months or longer) same sex romantic relationship completed an encrypted online questionnaire package which included psychometric measures designed to assess the aforementioned variables. Controlling for gender effects, analyses conducted using bootstrapping procedures supported full mediation pathways for both attachment anxiety and attachment avoidance. That is, attachment anxiety and avoidance were both directly associated to the perpetration of sexual SSIPV, and these relationships were both fully mediated by dyadic trust and sexual intimacy, in that respective order.

  14. Neonatal plasma polarizes TLR4-mediated cytokine responses towards low IL-12p70 and high IL-10 production via distinct factors.

    Directory of Open Access Journals (Sweden)

    Mirjam E Belderbos

    Full Text Available Human neonates are highly susceptible to infection, which may be due in part to impaired innate immune function. Neonatal Toll-like receptor (TLR responses are biased against the generation of pro-inflammatory/Th1-polarizing cytokines, yet the underlying mechanisms are incompletely defined. Here, we demonstrate that neonatal plasma polarizes TLR4-mediated cytokine production. When exposed to cord blood plasma, mononuclear cells (MCs produced significantly lower TLR4-mediated IL-12p70 and higher IL-10 compared to MC exposed to adult plasma. Suppression by neonatal plasma of TLR4-mediated IL-12p70 production, but not induction of TLR4-mediated IL-10 production, was maintained up to the age of 1 month. Cord blood plasma conferred a similar pattern of MC cytokine responses to TLR3 and TLR8 agonists, demonstrating activity towards both MyD88-dependent and MyD88-independent agonists. The factor causing increased TLR4-mediated IL-10 production by cord blood plasma was heat-labile, lost after protein depletion and independent of lipoprotein binding protein (LBP or soluble CD14 (sCD14. The factor causing inhibition of TLR4-mediated IL-12p70 production by cord blood plasma was resistant to heat inactivation or protein depletion and was independent of IL-10, vitamin D and prostaglandin E2. In conclusion, human neonatal plasma contains at least two distinct factors that suppress TLR4-mediated IL-12p70 production or induce IL-10 or production. Further identification of these factors will provide insight into the ontogeny of innate immune development and might identify novel targets for the prevention and treatment of neonatal infection.

  15. Neonatal Plasma Polarizes TLR4-Mediated Cytokine Responses towards Low IL-12p70 and High IL-10 Production via Distinct Factors

    Science.gov (United States)

    Belderbos, Mirjam E.; Levy, Ofer; Stalpers, Femke; Kimpen, Jan L.; Meyaard, Linde; Bont, Louis

    2012-01-01

    Human neonates are highly susceptible to infection, which may be due in part to impaired innate immune function. Neonatal Toll-like receptor (TLR) responses are biased against the generation of pro-inflammatory/Th1-polarizing cytokines, yet the underlying mechanisms are incompletely defined. Here, we demonstrate that neonatal plasma polarizes TLR4-mediated cytokine production. When exposed to cord blood plasma, mononuclear cells (MCs) produced significantly lower TLR4-mediated IL-12p70 and higher IL-10 compared to MC exposed to adult plasma. Suppression by neonatal plasma of TLR4-mediated IL-12p70 production, but not induction of TLR4-mediated IL-10 production, was maintained up to the age of 1 month. Cord blood plasma conferred a similar pattern of MC cytokine responses to TLR3 and TLR8 agonists, demonstrating activity towards both MyD88-dependent and MyD88-independent agonists. The factor causing increased TLR4-mediated IL-10 production by cord blood plasma was heat-labile, lost after protein depletion and independent of lipoprotein binding protein (LBP) or soluble CD14 (sCD14). The factor causing inhibition of TLR4-mediated IL-12p70 production by cord blood plasma was resistant to heat inactivation or protein depletion and was independent of IL-10, vitamin D and prostaglandin E2. In conclusion, human neonatal plasma contains at least two distinct factors that suppress TLR4-mediated IL-12p70 production or induce IL-10 or production. Further identification of these factors will provide insight into the ontogeny of innate immune development and might identify novel targets for the prevention and treatment of neonatal infection. PMID:22442690

  16. Co-ordinate regulation of distinct host cell signalling pathways by multifunctional enteropathogenic Escherichia coli effector molecules.

    Science.gov (United States)

    Kenny, Brendan; Ellis, Sarah; Leard, Alan D; Warawa, Jonathan; Mellor, Harry; Jepson, Mark A

    2002-05-01

    Enteropathogenic Escherichia coli (EPEC) is a major cause of paediatric diarrhoea and a model for the family of attaching and effacing (A/E) pathogens. A/E pathogens encode a type III secretion system to transfer effector proteins into host cells. The EPEC Tir effector protein acts as a receptor for the bacterial surface protein intimin and is involved in the formation of Cdc42-independent, actin-rich pedestal structures beneath the adhered bacteria. In this paper, we demonstrate that EPEC binding to HeLa cells also induces Tir-independent, cytoskeletal rearrangement evidenced by the early, transient formation of filopodia-like structures at sites of infection. Filopodia formation is dependent on expression of the EPEC Map effector molecule - a protein that targets mitochondria and induces their dysfunction. We show that Map-induced filopodia formation is independent of mitochondrial targeting and is abolished by cellular expression of the Cdc42 inhibitory WASP-CRIB domain, demonstrating that Map has at least two distinct functions in host cells. The transient nature of the filopodia is related to an ability of EPEC to downregulate Map-induced cell signalling that, like pedestal formation, was dependent on both Tir and intimin proteins. The ability of Tir to downregulate filopodia was impaired by disrupting a putative GTPase-activating protein (GAP) motif, suggesting that Tir may possess such a function, with its interaction with intimin triggering this activity. Furthermore, we also found that Map-induced cell signalling inhibits pedestal formation, revealing that the cellular effects of Tir and Map must be co-ordinately regulated during infection. Possible implications of the multifunctional nature of EPEC effector molecules in pathogenesis are discussed.

  17. Expression of three topologically distinct membrane proteins elicits unique stress response pathways in the yeast Saccharomyces cerevisiae.

    Science.gov (United States)

    Buck, Teresa M; Jordan, Rick; Lyons-Weiler, James; Adelman, Joshua L; Needham, Patrick G; Kleyman, Thomas R; Brodsky, Jeffrey L

    2015-06-01

    Misfolded membrane proteins are retained in the endoplasmic reticulum (ER) and are subject to ER-associated degradation, which clears the secretory pathway of potentially toxic species. While the transcriptional response to environmental stressors has been extensively studied, limited data exist describing the cellular response to misfolded membrane proteins. To this end, we expressed and then compared the transcriptional profiles elicited by the synthesis of three ER retained, misfolded ion channels: The α-subunit of the epithelial sodium channel, ENaC, the cystic fibrosis transmembrane conductance regulator, CFTR, and an inwardly rectifying potassium channel, Kir2.1, which vary in their mass, membrane topologies, and quaternary structures. To examine transcriptional profiles in a null background, the proteins were expressed in yeast, which was previously used to examine the degradation requirements for each substrate. Surprisingly, the proteins failed to induce a canonical unfolded protein response or heat shock response, although messages encoding several cytosolic and ER lumenal protein folding factors rose when αENaC or CFTR was expressed. In contrast, the levels of these genes were unaltered by Kir2.1 expression; instead, the yeast iron regulon was activated. Nevertheless, a significant number of genes that respond to various environmental stressors were upregulated by all three substrates, and compared with previous microarray data we deduced the existence of a group of genes that reflect a novel misfolded membrane protein response. These data indicate that aberrant proteins in the ER elicit profound yet unique cellular responses. Copyright © 2015 the American Physiological Society.

  18. Distinct signalling pathways of murine histamine H1- and H4-receptors expressed at comparable levels in HEK293 cells.

    Directory of Open Access Journals (Sweden)

    Silke Beermann

    Full Text Available Histamine (HA is recognized by its target cells via four G-protein-coupled receptors, referred to as histamine H1-receptor (H1R, H2R, H3R, and H4R. Both H1R and H4R exert pro-inflammatory functions. However, their signal transduction pathways have never been analyzed in a directly comparable manner side by side. Moreover, the analysis of pharmacological properties of the murine orthologs, representing the main targets of pre-clinical research, is very important. Therefore, we engineered recombinant HEK293 cells expressing either mouse (mH1R or mH4R at similar levels and analyzed HA-induced signalling in these cells. HA induced intracellular calcium mobilization via both mH1R and mH4R, with the mH1R being much more effective. Whereas cAMP accumulation was potentiated via the mH1R, it was reduced via the mH4R. The regulation of both second messengers via the H4R, but not the H1R, was sensitive to pertussis toxin (PTX. The mitogen-activated protein kinases (MAPKs ERK 1/2 were massively activated downstream of both receptors and demonstrated a functional involvement in HA-induced EGR-1 gene expression. The p38 MAPK was moderately activated via both receptors as well, but was functionally involved in HA-induced EGR-1 gene expression only in H4R-expressing cells. Surprisingly, in this system p38 MAPK activity reduced the HA-induced gene expression. In summary, using this system which allows a direct comparison of mH1R- and mH4R-induced signalling, qualitative and quantitative differences on the levels of second messenger generation and also in terms of p38 MAPK function became evident.

  19. Deciphering the ubiquitin-mediated pathway in apicomplexan parasites: a potential strategy to interfere with parasite virulence.

    Science.gov (United States)

    Ponts, Nadia; Yang, Jianfeng; Chung, Duk-Won Doug; Prudhomme, Jacques; Girke, Thomas; Horrocks, Paul; Le Roch, Karine G

    2008-06-11

    Reversible modification of proteins through the attachment of ubiquitin or ubiquitin-like modifiers is an essential post-translational regulatory mechanism in eukaryotes. The conjugation of ubiquitin or ubiquitin-like proteins has been demonstrated to play roles in growth, adaptation and homeostasis in all eukaryotes, with perturbation of ubiquitin-mediated systems associated with the pathogenesis of many human diseases, including cancer and neurodegenerative disorders. Here we describe the use of an HMM search of functional Pfam domains found in the key components of the ubiquitin-mediated pathway necessary to activate and reversibly modify target proteins in eight apicomplexan parasitic protozoa for which complete or late-stage genome projects exist. In parallel, the same search was conducted on five model organisms, single-celled and metazoans, to generate data to validate both the search parameters employed and aid paralog classification in Apicomplexa. For each of the 13 species investigated, a set of proteins predicted to be involved in the ubiquitylation pathway has been identified and demonstrates increasing component members of the ubiquitylation pathway correlating with organism and genome complexity. Sequence homology and domain architecture analyses facilitated prediction of apicomplexan-specific protein function, particularly those involved in regulating cell division during these parasite's complex life cycles. This study provides a comprehensive analysis of proteins predicted to be involved in the apicomplexan ubiquitin-mediated pathway. Given the importance of such pathway in a wide variety of cellular processes, our data is a key step in elucidating the biological networks that, in part, direct the pathogenicity of these parasites resulting in a massive impact on global health. Moreover, apicomplexan-specific adaptations of the ubiquitylation pathway may represent new therapeutic targets for much needed drugs against apicomplexan parasites.

  20. Transforming growth factor β-induced expression of chondroitin sulfate proteoglycans is mediated through non-Smad signaling pathways.

    Science.gov (United States)

    Jahan, Naima; Hannila, Sari S

    2015-01-01

    The expression of chondroitin sulfate proteoglycans (CSPGs) by reactive astrocytes is a major factor contributing to glial scarring and regenerative failure after spinal cord injury, but the molecular mechanisms underlying CSPG expression remain largely undefined. One contributing factor is transforming growth factor β (TGFβ), which is upregulated after injury and has been shown to induce expression of CSPGs in vitro. TGFβ typically mediates its effects through the Smad2/3 signaling pathway, and it has been suggested that this pathway is responsible for CSPG expression. However, there is evidence that TGFβ can also activate non-Smad signaling pathways. In this study, we report that TGFβ-induced expression of three different CSPGs--neurocan, brevican, and aggrecan--is mediated through non-Smad signaling pathways. We observed significant increases in TGFβ-induced expression of neurocan, brevican, and aggrecan following siRNA knockdown of Smad2 or Smad4, which indicates that Smad signaling is not required for the expression of these CSPGs. In addition, we show that neurocan, aggrecan, and brevican levels are significantly reduced when TGFβ is administered in the presence of either the PI3K inhibitor LY294002 or the mTOR inhibitor rapamycin, but not the MEK1/2 inhibitor U0126. This suggests that TGFβ mediates this effect through non-Smad-dependent activation of the PI3K-Akt-mTOR signaling pathway, and targeting this pathway may therefore be an effective means of reducing CSPG expression in the injured CNS. Copyright © 2014 Elsevier Inc. All rights reserved.

  1. The Hippo pathway controls border cell migration through distinct mechanisms in outer border cells and polar cells of the Drosophila ovary.

    Science.gov (United States)

    Lin, Tzu-Huai; Yeh, Tsung-Han; Wang, Tsu-Wei; Yu, Jenn-Yah

    2014-11-01

    The Hippo pathway is a key signaling cascade in controlling organ size. The core components of this pathway are two kinases, Hippo (Hpo) and Warts (Wts), and a transcriptional coactivator, Yorkie (Yki). Yes-associated protein (YAP, a Yki homolog in mammals) promotes epithelial-mesenchymal transition and cell migration in vitro. Here, we use border cells in the Drosophila ovary as a model to study Hippo pathway functions in cell migration in vivo. During oogenesis, polar cells secrete Unpaired (Upd), which activates JAK/STAT signaling of neighboring cells and specifies them into outer border cells. The outer border cells form a cluster with polar cells and undergo migration. We find that hpo and wts are required for migration of the border cell cluster. In outer border cells, overexpression of hpo disrupts polarization of the actin cytoskeleton and attenuates migration. In polar cells, knockdown of hpo and wts or overexpression of yki impairs border cell induction and disrupts migration. These manipulations in polar cells reduce JAK/STAT activity in outer border cells. Expression of upd-lacZ is increased and decreased in yki and hpo mutant polar cells, respectively. Furthermore, forced expression of upd in polar cells rescues defects of border cell induction and migration caused by wts knockdown. These results suggest that Yki negatively regulates border cell induction by inhibiting JAK/STAT signaling. Together, our data elucidate two distinct mechanisms of the Hippo pathway in controlling border cell migration: (1) in outer border cells, it regulates polarized distribution of the actin cytoskeleton; (2) in polar cells, it regulates upd expression to control border cell induction and migration. Copyright © 2014 by the Genetics Society of America.

  2. Mechanistically Distinct Pathways of Divergent Regulatory DNA Creation Contribute to Evolution of Human-Specific Genomic Regulatory Networks Driving Phenotypic Divergence of Homo sapiens.

    Science.gov (United States)

    Glinsky, Gennadi V

    2016-09-19

    Thousands of candidate human-specific regulatory sequences (HSRS) have been identified, supporting the hypothesis that unique to human phenotypes result from human-specific alterations of genomic regulatory networks. Collectively, a compendium of multiple diverse families of HSRS that are functionally and structurally divergent from Great Apes could be defined as the backbone of human-specific genomic regulatory networks. Here, the conservation patterns analysis of 18,364 candidate HSRS was carried out requiring that 100% of bases must remap during the alignments of human, chimpanzee, and bonobo sequences. A total of 5,535 candidate HSRS were identified that are: (i) highly conserved in Great Apes; (ii) evolved by the exaptation of highly conserved ancestral DNA; (iii) defined by either the acceleration of mutation rates on the human lineage or the functional divergence from non-human primates. The exaptation of highly conserved ancestral DNA pathway seems mechanistically distinct from the evolution of regulatory DNA segments driven by the species-specific expansion of transposable elements. Genome-wide proximity placement analysis of HSRS revealed that a small fraction of topologically associating domains (TADs) contain more than half of HSRS from four distinct families. TADs that are enriched for HSRS and termed rapidly evolving in humans TADs (revTADs) comprise 0.8-10.3% of 3,127 TADs in the hESC genome. RevTADs manifest distinct correlation patterns between placements of human accelerated regions, human-specific transcription factor-binding sites, and recombination rates. There is a significant enrichment within revTAD boundaries of hESC-enhancers, primate-specific CTCF-binding sites, human-specific RNAPII-binding sites, hCONDELs, and H3K4me3 peaks with human-specific enrichment at TSS in prefrontal cortex neurons (P sapiens is driven by the evolution of human-specific genomic regulatory networks via at least two mechanistically distinct pathways of creation of

  3. Distinct dictation of Japanese encephalitis virus-induced neuroinflammation and lethality via triggering TLR3 and TLR4 signal pathways.

    Directory of Open Access Journals (Sweden)

    Young Woo Han

    2014-09-01

    Full Text Available Japanese encephalitis (JE is major emerging neurologic disease caused by JE virus. To date, the impact of TLR molecules on JE progression has not been addressed. Here, we determined whether each TLR modulates JE, using several TLR-deficient mouse strains (TLR2, TLR3, TLR4, TLR7, TLR9. Surprisingly, among the tested TLR-deficient mice there were contrasting results in TLR3(-/- and TLR4(-/- mice, i.e. TLR3(-/- mice were highly susceptible to JE, whereas TLR4(-/- mice showed enhanced resistance to JE. TLR3 ablation induced severe CNS inflammation characterized by early infiltration of inflammatory CD11b(+Ly-6Chigh monocytes along with profoundly increased viral burden, proinflammatory cytokine/chemokine expression as well as BBB permeability. In contrast, TLR4(-/- mice showed mild CNS inflammation manifested by reduced viral burden, leukocyte infiltration and proinflammatory cytokine expression. Interestingly, TLR4 ablation provided potent in vivo systemic type I IFN innate response, as well as ex vivo type I IFN production associated with strong induction of antiviral PRRs (RIG-I, MDA5, transcription factors (IRF-3, IRF-7, and IFN-dependent (PKR, Oas1, Mx and independent ISGs (ISG49, ISG54, ISG56 by alternative activation of IRF3 and NF-κB in myeloid-derived DCs and macrophages, as compared to TLR3(-/- myeloid-derived cells which were more permissive to viral replication through impaired type I IFN innate response. TLR4 ablation also appeared to mount an enhanced type I IFN innate and humoral, CD4(+ and CD8(+ T cell responses, which were mediated by altered immune cell populations (increased number of plasmacytoid DCs and NK cells, reduced CD11b(+Ly-6C(high monocytes and CD4(+Foxp3(+ Treg number in lymphoid tissue. Thus, potent type I IFN innate and adaptive immune responses in the absence of TLR4 were closely coupled with reduced JE lethality. Collectively, these results suggest that a balanced triggering of TLR signal array by viral components

  4. Bifidobacterium breve UCC2003 metabolises the human milk oligosaccharides lacto-N-tetraose and lacto-N-neo-tetraose through overlapping, yet distinct pathways

    Science.gov (United States)

    James, Kieran; Motherway, Mary O’Connell; Bottacini, Francesca; van Sinderen, Douwe

    2016-01-01

    In this study, we demonstrate that the prototype B. breve strain UCC2003 possesses specific metabolic pathways for the utilisation of lacto-N-tetraose (LNT) and lacto-N-neotetraose (LNnT), which represent the central moieties of Type I and Type II human milk oligosaccharides (HMOs), respectively. Using a combination of experimental approaches, the enzymatic machinery involved in the metabolism of LNT and LNnT was identified and characterised. Homologs of the key genetic loci involved in the utilisation of these HMO substrates were identified in B. breve, B. bifidum, B. longum subsp. infantis and B. longum subsp. longum using bioinformatic analyses, and were shown to be variably present among other members of the Bifidobacterium genus, with a distinct pattern of conservation among human-associated bifidobacterial species. PMID:27929046

  5. Mapping glucose-mediated gut-to-brain signalling pathways in humans.

    Science.gov (United States)

    Little, Tanya J; McKie, Shane; Jones, Richard B; D'Amato, Massimo; Smith, Craig; Kiss, Orsolya; Thompson, David G; McLaughlin, John T

    2014-08-01

    Previous fMRI studies have demonstrated that glucose decreases the hypothalamic BOLD response in humans. However, the mechanisms underlying the CNS response to glucose have not been defined. We recently demonstrated that the slowing of gastric emptying by glucose is dependent on activation of the gut peptide cholecystokinin (CCK1) receptor. Using physiological functional magnetic resonance imaging this study aimed to determine the whole brain response to glucose, and whether CCK plays a central role. Changes in blood oxygenation level-dependent (BOLD) signal were monitored using fMRI in 12 healthy subjects following intragastric infusion (250ml) of: 1M glucose+predosing with dexloxiglumide (CCK1 receptor antagonist), 1M glucose+placebo, or 0.9% saline (control)+placebo, in a single-blind, randomised fashion. Gallbladder volume, blood glucose, insulin, and GLP-1 and CCK concentrations were determined. Hunger, fullness and nausea scores were also recorded. Intragastric glucose elevated plasma glucose, insulin, and GLP-1, and reduced gall bladder volume (an in vivo assay for CCK secretion). Glucose decreased BOLD signal, relative to saline, in the brainstem and hypothalamus as well as the cerebellum, right occipital cortex, putamen and thalamus. The timing of the BOLD signal decrease was negatively correlated with the rise in blood glucose and insulin levels. The glucose+dex arm highlighted a CCK1-receptor dependent increase in BOLD signal only in the motor cortex. Glucose induces site-specific differences in BOLD response in the human brain; the brainstem and hypothalamus show a CCK1 receptor-independent reduction which is likely to be mediated by a circulatory effect of glucose and insulin, whereas the motor cortex shows an early dexloxiglumide-reversible increase in signal, suggesting a CCK1 receptor-dependent neural pathway. Copyright © 2014. Published by Elsevier Inc.

  6. Mapping glucose-mediated gut-to-brain signalling pathways in humans☆

    Science.gov (United States)

    Little, Tanya J.; McKie, Shane; Jones, Richard B.; D'Amato, Massimo; Smith, Craig; Kiss, Orsolya; Thompson, David G.; McLaughlin, John T.

    2014-01-01

    Objectives Previous fMRI studies have demonstrated that glucose decreases the hypothalamic BOLD response in humans. However, the mechanisms underlying the CNS response to glucose have not been defined. We recently demonstrated that the slowing of gastric emptying by glucose is dependent on activation of the gut peptide cholecystokinin (CCK1) receptor. Using physiological functional magnetic resonance imaging this study aimed to determine the whole brain response to glucose, and whether CCK plays a central role. Experimental design Changes in blood oxygenation level-dependent (BOLD) signal were monitored using fMRI in 12 healthy subjects following intragastric infusion (250 ml) of: 1 M glucose + predosing with dexloxiglumide (CCK1 receptor antagonist), 1 M glucose + placebo, or 0.9% saline (control) + placebo, in a single-blind, randomised fashion. Gallbladder volume, blood glucose, insulin, and GLP-1 and CCK concentrations were determined. Hunger, fullness and nausea scores were also recorded. Principal observations Intragastric glucose elevated plasma glucose, insulin, and GLP-1, and reduced gall bladder volume (an in vivo assay for CCK secretion). Glucose decreased BOLD signal, relative to saline, in the brainstem and hypothalamus as well as the cerebellum, right occipital cortex, putamen and thalamus. The timing of the BOLD signal decrease was negatively correlated with the rise in blood glucose and insulin levels. The glucose + dex arm highlighted a CCK1-receptor dependent increase in BOLD signal only in the motor cortex. Conclusions Glucose induces site-specific differences in BOLD response in the human brain; the brainstem and hypothalamus show a CCK1 receptor-independent reduction which is likely to be mediated by a circulatory effect of glucose and insulin, whereas the motor cortex shows an early dexloxiglumide-reversible increase in signal, suggesting a CCK1 receptor-dependent neural pathway. PMID:24685436

  7. The Hippo pathway mediates inhibition of vascular smooth muscle cell proliferation by cAMP.

    Science.gov (United States)

    Kimura, Tomomi E; Duggirala, Aparna; Smith, Madeleine C; White, Stephen; Sala-Newby, Graciela B; Newby, Andrew C; Bond, Mark

    2016-01-01

    Inhibition of vascular smooth muscle cell (VSMC) proliferation by intracellular cAMP prevents excessive neointima formation and hence angioplasty restenosis and vein-graft failure. These protective effects are mediated via actin-cytoskeleton remodelling and subsequent regulation of gene expression by mechanisms that are incompletely understood. Here we investigated the role of components of the growth-regulatory Hippo pathway, specifically the transcription factor TEAD and its co-factors YAP and TAZ in VSMC. Elevation of cAMP using forskolin, dibutyryl-cAMP or the physiological agonists, Cicaprost or adenosine, significantly increased phosphorylation and nuclear export YAP and TAZ and inhibited TEAD-luciferase report gene activity. Similar effects were obtained by inhibiting RhoA activity with C3-transferase, its downstream kinase, ROCK, with Y27632, or actin-polymerisation with Latrunculin-B. Conversely, expression of constitutively-active RhoA reversed the inhibitory effects of forskolin on TEAD-luciferase. Forskolin significantly inhibited the mRNA expression of the pro-mitogenic genes, CCN1, CTGF, c-MYC and TGFB2 and this was reversed by expression of constitutively-active YAP or TAZ phospho-mutants. Inhibition of YAP and TAZ function with RNAi or Verteporfin significantly reduced VSMC proliferation. Furthermore, the anti-mitogenic effects of forskolin were reversed by overexpression of constitutively-active YAP or TAZ. Taken together, these data demonstrate that cAMP-induced actin-cytoskeleton remodelling inhibits YAP/TAZ-TEAD dependent expression of pro-mitogenic genes in VSMC. This mechanism contributes novel insight into the anti-mitogenic effects of cAMP in VSMC and suggests a new target for intervention. Copyright © 2015 The Authors. Published by Elsevier Ltd.. All rights reserved.

  8. DHEA-induced ovarian hyperfibrosis is mediated by TGF-β signaling pathway.

    Science.gov (United States)

    Wang, Daojuan; Wang, Wenqing; Liang, Qiao; He, Xuan; Xia, Yanjie; Shen, Shanmei; Wang, Hongwei; Gao, Qian; Wang, Yong

    2018-01-10

    The polycystic ovary syndrome (PCOS) is a common metabolic and endocrine disorder with pathological mechanisms remain unclear. The following study investigates the ovarian hyperfibrosis forming via transforming growth factor-β (TGF-β) signaling pathway in Dehydroepiandrosterone (DHEA)- induced polycystic ovary syndrome (PCOS) rat model. We furthermore explored whether TGF-βRI inhibitor (SB431542) decreases ovarian fibrosis by counterbalancing the expression of fibrotic biomarkers. Thirty female Sprague-Dawley rats were randomly divided into Blank group (n = 6), Oil group (n = 6), and Oil + DHEA-induced model group (n = 6 + 12). The model groups were established by subcutaneous injection of DHEA for 35 consecutive days. The 12 successful model rats were additionally divided in vehicle group (n = 6) and SB431542-treated group (n = 6). Vehicle group and SB431542-treated group, served as administration group and were intraperitoneally injected with DMSO and SB431542 for additional 14 consecutive days. Ovarian morphology, fibrin and collagen localization and expression in ovaries were detected using H&E staining, immunohistochemistry and Sirius red staining. The ovarian protein and RNA were examined using Western blot and RT-PCR. In DHEA-induced ovary in rat, fibrin and collagen had significantly higher levels, while the main fibrosis markers (TGF-β, CTGF, fibronectin, a-SMA) were obviously upregulated. SB431542 significantly reduced the expression of pro-fibrotic molecules (TGF-β, Smad3, Smad2, a-SMA) and increased anti-fibrotic factor MMP2. TGF-βRI inhibitor (SB431542) inhibits the downstream signaling molecules of TGF-β and upregulates MMP2, which in turn prevent collagen deposition. Moreover, ovarian hyperfibrosis in DHEA-induced PCOS rat model could be improved by TGF-βRI inhibitor (SB431542) restraining the transcription of accelerating fibrosis genes and modulating EMT mediator.

  9. Distinct signalling properties of insulin receptor substrate (IRS)-1 and IRS-2 in mediating insulin/IGF-1 action

    DEFF Research Database (Denmark)

    Rabiee, Atefeh; Krüger, Marcus; Ardenkjær-Larsen, Jacob

    2018-01-01

    Insulin/IGF-1 action is driven by a complex and highly integrated signalling network. Loss-of-function studies indicate that the major insulin/IGF-1 receptor substrate (IRS) proteins, IRS-1 and IRS-2, mediate different biological functions in vitro and in vivo, suggesting specific signalling...... properties despite their high degree of homology. To identify mechanisms contributing to the differential signalling properties of IRS-1 and IRS-2 in the mediation of insulin/IGF-1 action, we performed comprehensive mass spectrometry (MS)-based phosphoproteomic profiling of brown preadipocytes from wild type......, IRS-1-/-and IRS-2-/-mice in the basal and IGF-1-stimulated states. We applied stable isotope labeling by amino acids in cell culture (SILAC) for the accurate quantitation of changes in protein phosphorylation. We found ~10% of the 6262 unique phosphorylation sites detected to be regulated by IGF-1...

  10. DMPD: Complement-mediated phagocytosis--the role of Syk. [Dynamic Macrophage Pathway CSML Database

    Lifescience Database Archive (English)

    Full Text Available 16754322 Complement-mediated phagocytosis--the role of Syk. Tohyama Y, Yamamura H. ...IUBMB Life. 2006 May-Jun;58(5-6):304-8. (.png) (.svg) (.html) (.csml) Show Complement-mediated phagocytosis-...-the role of Syk. PubmedID 16754322 Title Complement-mediated phagocytosis--the role of Syk. Authors Tohyama

  11. DMPD: Negative regulation of cytoplasmic RNA-mediated antiviral signaling. [Dynamic Macrophage Pathway CSML Database

    Lifescience Database Archive (English)

    Full Text Available 18703349 Negative regulation of cytoplasmic RNA-mediated antiviral signaling. Komur...Show Negative regulation of cytoplasmic RNA-mediated antiviral signaling. PubmedID 18703349 Title Negative r...egulation of cytoplasmic RNA-mediated antiviral signaling. Authors Komuro A, Bamm

  12. DMPD: Fragments of extracellular matrix as mediators of inflammation. [Dynamic Macrophage Pathway CSML Database

    Lifescience Database Archive (English)

    Full Text Available 18243041 Fragments of extracellular matrix as mediators of inflammation. Adair-Kirk...l) Show Fragments of extracellular matrix as mediators of inflammation. PubmedID 18243041 Title Fragments of... extracellular matrix as mediators of inflammation. Authors Adair-Kirk TL, Senior

  13. The BDNF/TrkB Signaling Pathway Is Involved in Heat Hyperalgesia Mediated by Cdk5 in Rats

    OpenAIRE

    Zhang, Hong-Hai; Zhang, Xiao-Qin; Xue, Qing-Sheng; Yan-Luo,; Huang, Jin-Lu; Zhang, Su; Shao, Hai-Jun; Lu, Han; Wang, Wen-Yuan; Yu, Bu-Wei

    2014-01-01

    Background Cyclin-dependent kinase 5 (Cdk5) has been shown to play an important role in mediating inflammation-induced heat hyperalgesia. However, the underlying mechanism remains unclear. The aim of this study was to determine whether roscovitine, an inhibitor of Cdk5, could reverse the heat hyperalgesia induced by peripheral injection of complete Freund's adjuvant (CFA) via the brain-derived neurotrophic factor (BDNF)-tyrosine kinase B (TrkB) signaling pathway in the dorsal horn of the spin...

  14. Dietary Blueberry and Bifidobacteria Attenuate Nonalcoholic Fatty Liver Disease in Rats by Affecting SIRT1-Mediated Signaling Pathway

    Science.gov (United States)

    Ren, Tingting; Huang, Chao; Cheng, Mingliang

    2014-01-01

    NAFLD model rats were established and divided into NAFLD model (MG group), SIRT1 RNAi (SI group), blueberry juice (BJ group), blueberry juice + bifidobacteria (BJB group), blueberry juice + SIRT1 RNAi (BJSI group), and blueberry juice + bifidobacteria + SIRT1 RNAi groups (BJBSI group). A group with normal rats was a control group (CG). BJB group ameliorated NAFLD, which was better than BJ group (P Blueberry juice and bifidobacteria improve NAFLD by activating SIRTI-mediating signaling pathway. PMID:25544867

  15. A pivotal role of the jasmonic acid signal pathway in mediating radiation-induced bystander effects in Arabidopsis thaliana

    Energy Technology Data Exchange (ETDEWEB)

    Wang, Ting; Xu, Wei; Deng, Chenguang; Xu, Shaoxin; Li, Fanghua; Wu, Yuejin; Wu, Lijun [Key Laboratory of Ion Beam Bioengineering, Hefei Institutes of Physical Science, Chinese Academy of Sciences, Hefei 230031 (China); Key Laboratory of Environmental Toxicology and Pollution Control Technology of Anhui Province, Hefei 230031 (China); Bian, Po, E-mail: bianpo@ipp.ac.cn [Key Laboratory of Ion Beam Bioengineering, Hefei Institutes of Physical Science, Chinese Academy of Sciences, Hefei 230031 (China); Key Laboratory of Environmental Toxicology and Pollution Control Technology of Anhui Province, Hefei 230031 (China)

    2016-09-15

    Highlights: • The JA signal pathway plays a pivotal role in mediating radiation-induced bystander effects in Arabidopsis thaliana. • The JA signal pathway is involved in both the generation of bystander signals in irradiated roots and radiation responses in bystander aerial plants. • Over-accumulation of endogenous JA enhances the radiosensitivity of plants in terms of RIBE. - Abstract: Although radiation-induced bystander effects (RIBE) in Arabidopsis thaliana have been well demonstrated in vivo, little is known about their underlying mechanisms, particularly with regard to the participating signaling molecules and signaling pathways. In higher plants, jasmonic acid (JA) and its bioactive derivatives are well accepted as systemic signal transducers that are produced in response to various environmental stresses. It is therefore speculated that the JA signal pathway might play a potential role in mediating radiation-induced bystander signaling of root-to-shoot. In the present study, pretreatment of seedlings with Salicylhydroxamic acid, an inhibitor of lipoxigenase (LOX) in JA biosynthesis, significantly suppressed RIBE-mediated expression of the AtRAD54 gene. After root irradiation, the aerial parts of A. thaliana mutants deficient in JA biosynthesis (aos) and signaling cascades (jar1-1) showed suppressed induction of the AtRAD54 and AtRAD51 genes and TSI and 180-bp repeats, which have been extensively used as endpoints of bystander genetic and epigenetic effects in plants. These results suggest an involvement of the JA signal pathway in the RIBE of plants. Using the root micro-grafting technique, the JA signal pathway was shown to participate in both the generation of bystander signals in irradiated root cells and radiation responses in the bystander aerial parts of plants. The over-accumulation of endogenous JA in mutant fatty acid oxygenation up-regulated 2 (fou2), in which mutation of the Two Pore Channel 1 (TPC1) gene up-regulates expression of the LOX

  16. Silencing of Pokemon enhances caspase-dependent apoptosis via fas- and mitochondria-mediated pathways in hepatocellular carcinoma cells.

    Science.gov (United States)

    Zhang, Yu-Qin; Xiao, Chuan-Xing; Lin, Bi-Yun; Shi, Ying; Liu, Yun-Peng; Liu, Jing-Jing; Guleng, Bayasi; Ren, Jian-Lin

    2013-01-01

    The role of Pokemon (POK erythroid myeloid ontogenic actor), a recently identified POK transcription factor with proto-oncogenic activity, in hepatocellular carcinogenesis has only been assessed by a few studies. Our previous study revealed that Pokemon is overexpressed in hepatocellular carcinomas (HCC) and promotes HCC cell proliferation and migration via an AKT- and ERK- dependent manner. In the present study, we used the TUNEL assay and FACS analysis to demonstrate that oxaliplatin induced apoptosis was significantly increased in cells with silenced Pokemon. Western blots showed that p53 expression and phosphorylation were significantly increased in Pokemon defective cells, thereby initiating the mitochondria-mediated and death receptor-mediated apoptotic pathways. In the mitochondria-mediated pathway, expression of pro-apoptotic Bcl-2 family members (including Bad, Bid, Bim and Puma) as well as AIF was increased and decreasing the mitochondrial membrane potential resulted in cytochrome C released from mitochondrial in HepG2 si-Pokemon cells. In addition, upon oxaliplatin treatment of Pokemon-silenced cells, the FAS receptor, FADD and their downstream targets caspase-10 and caspase-8 were activated, causing increased release of caspase-8 active fragments p18 and p10. Increased activated caspase-8-mediated cleavage and activation of downstream effector caspases such as caspase-9 and caspase-3 was observed in HepG2 si-Pokemon cells as compared to control. Therefore, Pokemon might serve as an important mediator of crosstalk between intrinsic and extrinsic apoptotic pathways in HCC cells. Moreover, our findings suggest that Pokemon could be an attractive therapeutic target gene for human cancer therapy.

  17. Silencing of Pokemon enhances caspase-dependent apoptosis via fas- and mitochondria-mediated pathways in hepatocellular carcinoma cells.

    Directory of Open Access Journals (Sweden)

    Yu-Qin Zhang

    Full Text Available The role of Pokemon (POK erythroid myeloid ontogenic actor, a recently identified POK transcription factor with proto-oncogenic activity, in hepatocellular carcinogenesis has only been assessed by a few studies. Our previous study revealed that Pokemon is overexpressed in hepatocellular carcinomas (HCC and promotes HCC cell proliferation and migration via an AKT- and ERK- dependent manner. In the present study, we used the TUNEL assay and FACS analysis to demonstrate that oxaliplatin induced apoptosis was significantly increased in cells with silenced Pokemon. Western blots showed that p53 expression and phosphorylation were significantly increased in Pokemon defective cells, thereby initiating the mitochondria-mediated and death receptor-mediated apoptotic pathways. In the mitochondria-mediated pathway, expression of pro-apoptotic Bcl-2 family members (including Bad, Bid, Bim and Puma as well as AIF was increased and decreasing the mitochondrial membrane potential resulted in cytochrome C released from mitochondrial in HepG2 si-Pokemon cells. In addition, upon oxaliplatin treatment of Pokemon-silenced cells, the FAS receptor, FADD and their downstream targets caspase-10 and caspase-8 were activated, causing increased release of caspase-8 active fragments p18 and p10. Increased activated caspase-8-mediated cleavage and activation of downstream effector caspases such as caspase-9 and caspase-3 was observed in HepG2 si-Pokemon cells as compared to control. Therefore, Pokemon might serve as an important mediator of crosstalk between intrinsic and extrinsic apoptotic pathways in HCC cells. Moreover, our findings suggest that Pokemon could be an attractive therapeutic target gene for human cancer therapy.

  18. Descending serotonergic facilitation mediated by spinal 5-HT3 receptors engages spinal rapamycin-sensitive pathways in the rat

    Science.gov (United States)

    Asante, Curtis O.; Dickenson, Anthony H.

    2010-01-01

    We have recently reported the importance of spinal rapamycin-sensitive pathways in maintaining persistent pain-like states. A descending facilitatory drive mediated through spinal 5-HT3 receptors (5-HT3Rs) originating from superficial dorsal horn NK1-expressing neurons and that relays through the parabrachial nucleus and the rostroventral medial medulla to act on deep dorsal horn neurons is known be important in maintaining these pain-like states. To determine if spinal rapamycin-sensitive pathways are activated by a descending serotonergic drive, we investigated the effects of spinally administered rapamycin on responses of deep dorsal horn neurons that had been pre-treated with the selective 5-HT3R antagonist ondansetron. We also investigated the effects of spinally administered cell cycle inhibitor (CCI)-779 (a rapamycin ester analogue) on deep dorsal horn neurons from rats with carrageenan-induced inflammation of the hind paw. Unlike some other models of persistent pain, this model does not involve an altered 5-HT3R-mediated descending serotonergic drive. We found that the inhibitory effects of rapamycin were significantly reduced for neuronal responses to mechanical and thermal stimuli when the spinal cord was pre-treated with ondansetron. Furthermore, CCI-779 was found to be ineffective in attenuating spinal neuronal responses to peripheral stimuli in carrageenan-treated rats. Therefore, we conclude that 5-HT3R-mediated descending facilitation is one requirement for activation of rapamycin-sensitive pathways that contribute to persistent pain-like states. PMID:20709148

  19. Different gene-expression profiles for the poorly differentiated carcinoma and the highly differentiated papillary adenocarcinoma in mammary glands support distinct metabolic pathways

    International Nuclear Information System (INIS)

    Eilon, Tali; Barash, Itamar

    2008-01-01

    Deregulation of Stat5 in the mammary gland of transgenic mice causes tumorigenesis. Poorly differentiated carcinoma and highly differentiated papillary adenocarcinoma tumors evolve. To distinguish the genes and elucidate the cellular processes and metabolic pathways utilized to preserve these phenotypes, gene-expression profiles were analyzed. Mammary tumors were excised from transgenic mice carrying a constitutively active variant of Stat5, or a Stat5 variant lacking s transactivation domain. These tumors displayed either the carcinoma or the papillary adenocarcinoma phenotypes. cRNAs, prepared from each tumor were hybridized to an Affymetrix GeneChip ® Mouse Genome 430A 2.0 array. Gene-ontology analysis, hierarchical clustering and biological-pathway analysis were performed to distinct the two types of tumors. Histopathology and immunofluorescence staining complemented the comparison between the tumor phenotypes. The nucleus-cytoskeleton-plasma membrane axis is a major target for differential gene expression between phenotypes. In the carcinoma, stronger expression of genes coding for specific integrins, cytoskeletal proteins and calcium-binding proteins highlight cell-adhesion and motility features of the tumor cells. This is supported by the higher expression of genes involved in O-glycan synthesis, TGF-β, activin, their receptors and Smad3, as well as the Notch ligands and members of the γ-secretase complex that enable Notch nuclear localization. The Wnt pathway was also a target for differential gene expression. Higher expression of genes encoding the degradation complex of the canonical pathway and limited TCF expression in the papillary adenocarcinoma result in membranal accumulation of β-catenin, in contrast to its nuclear translocation in the carcinoma. Genes involved in cell-cycle arrest at G1 and response to DNA damage were more highly expressed in the papillary adenocarcinomas, as opposed to favored G2/M regulation in the carcinoma tumors. At least

  20. Excessive Time on Social Networking Sites and Disordered Eating Behaviors Among Undergraduate Students: Appearance and Weight Esteem as Mediating Pathways.

    Science.gov (United States)

    Murray, Marisa; Maras, Danijela; Goldfield, Gary S

    2016-12-01

    Social networking sites (SNS) are a popular form of communication among undergraduate students. Body image concerns and disordered eating behaviors are also quite prevalent among this population. Maladaptive use of SNS has been associated with disordered eating behaviors; however, the mechanisms remain unclear. The present study examined if body image concerns (e.g., appearance and weight esteem) mediate the relationship between excessive time spent on SNS and disordered eating behaviors (restrained and emotional eating). The sample included 383 (70.2 percent female) undergraduate students (mean age = 23.08 years, standard deviation = 3.09) who completed self-report questionnaires related to SNS engagement, body image, disordered eating behaviors, and demographics. Parallel multiple mediation and moderated mediation analyses revealed that lower weight and appearance esteem mediated the relationship between excessive time on SNS and restrained eating for males and females, whereas appearance esteem mediated the relationship between excessive time on SNS and emotional eating for females only. The study adds to the literature by highlighting mediational pathways and gender differences. Intervention research is needed to determine if teaching undergraduate students more adaptive ways of using SNS or reducing exposure to SNS reduces body dissatisfaction and disordered eating in this high-risk population.

  1. Neutralized adenovirus-immune complexes can mediate effective gene transfer via an Fc receptor-dependent infection pathway.

    Science.gov (United States)

    Leopold, Philip L; Wendland, Rebecca L; Vincent, Theresa; Crystal, Ronald G

    2006-10-01

    Neutralization of adenovirus (Ad) by anti-Ad neutralizing antibodies in serum involves formation of Ad-immune complexes that prevent the virus from interacting with target cells. We hypothesized that Ad-immune complexes likely contain viable Ad vectors which, although no longer capable of gaining access to receptors on target cells, may be able to express transgenes in cells bearing Fc receptors for immunoglobulins, i.e., that antibody-based "neutralization" of Ad vectors may be circumvented by the Fc receptor pathway. To test this hypothesis, we expressed the Fcgamma receptor IIA (FcgammaR) in A549 lung epithelial cells or human dermal fibroblasts and evaluated gene transfer in the presence of human neutralizing anti-Ad serum. FcgammaR-expressing cells bound and internalized copious amounts of Ad, with a distinct population of internalized Ad trafficking to the nucleus. The dose-response curves for inhibition of gene transfer revealed that FcgammaR-expressing cells required a more-than-10-fold higher concentration of anti-Ad serum to achieve 50% inhibition of Ad-encoded beta-galactosidase expression compared with non-FcgammaR-expressing cells. The discrepancy between neutralization of Ad during infection of FcgammaR-expressing cells and neutralization of Ad during infection of non-FcgammaR-expressing cells occurred with either heat-inactivated or non-heat-inactivated sera, was blocked by addition of purified Fc domain protein, and did not require the cytoplasmic domain of FcgammaR, suggesting that immune complex internalization proceeded via endocytosis rather than phagocytosis. FcgammaR-mediated infection by Ad-immune complexes did not require expression of the coxsackie virus-Ad receptor (CAR) since similar data were obtained when CAR-deficient human dermal fibroblasts were engineered to express FcgammaR. However, interaction of the Ad penton base with cell surface integrins contributed to the difference in neutralization between FcgammaR-expressing and non

  2. TOR Pathway-Mediated Juvenile Hormone Synthesis Regulates Nutrient-Dependent Female Reproduction in Nilaparvata lugens (Stål).

    Science.gov (United States)

    Lu, Kai; Chen, Xia; Liu, Wen-Ting; Zhou, Qiang

    2016-03-28

    The "target of rapamycin" (TOR) nutritional signaling pathway and juvenile hormone (JH) regulation of vitellogenesis has been known for a long time. However, the interplay between these two pathways regulating vitellogenin (Vg) expression remains obscure. Here, we first demonstrated the key role of amino acids (AAs) in activation of Vg synthesis and egg development in Nilaparvata lugens using chemically defined artificial diets. AAs induced the expression of TOR and S6K (S6 kinase), whereas RNAi-mediated silencing of these two TOR pathway genes and rapamycin application strongly inhibited the AAs-induced Vg synthesis. Furthermore, knockdown of Rheb (Ras homologue enriched in brain), TOR, S6K and application of rapamycin resulted in a dramatic reduction in the mRNA levels of jmtN (juvenile hormone acid methyltransferase, JHAMT). Application of JH III on the RNAi (Rheb and TOR) and rapamycin-treated females partially rescued the Vg expression. Conversely, knockdown of either jmtN or met (methoprene-tolerant, JH receptor) and application of JH III had no effects on mRNA levels of Rheb, TOR and S6K and phosphorylation of S6K. In summary, our results demonstrate that the TOR pathway induces JH biosynthesis that in turn regulates AAs-mediated Vg synthesis in N. lugens.

  3. Longitudinal pathways from early maternal depression to children's dysregulated representations: a moderated mediation analysis of harsh parenting and gender.

    Science.gov (United States)

    Martoccio, Tiffany L; Brophy-Herb, Holly E; Maupin, Angela N; Robinson, Joann L

    2016-01-01

    There is some evidence linking maternal depression, harsh parenting, and children's internal representations of attachment, yet, longitudinal examinations of these relationships and differences in the developmental pathways between boys and girls are lacking. Moderated mediation growth curves were employed to examine harsh parenting as a mechanism underlying the link between maternal depression and children's dysregulated representations using a nationally-representative, economically-vulnerable sample of mothers and their children (n = 575; 49% boys, 51% girls). Dysregulation representations were measured using the MacArthur Story Stem Battery at five years of age (M = 5.14, SD = 0.29). Harsh parenting mediated the association between early maternal depression and dysregulated representations for girls. Though initial harsh parenting was a significant mediator for boys, a stronger direct effect of maternal depression to dysregulated representations emerged over time. Results are discussed in terms of their implications for intervention efforts aimed at promoting early supportive parenting.

  4. Distinct Escape Pathway by Hepatitis C Virus Genotype 1a from a Dominant CD8+ T Cell Response by Selection of Altered Epitope Processing.

    Science.gov (United States)

    Walker, Andreas; Skibbe, Kathrin; Steinmann, Eike; Pfaender, Stephanie; Kuntzen, Thomas; Megger, Dominik A; Groten, Svenja; Sitek, Barbara; Lauer, Georg M; Kim, Arthur Y; Pietschmann, Thomas; Allen, Todd M; Timm, Joerg

    2016-01-01

    Antiviral CD8(+) T cells are a key component of the adaptive immune response against HCV, but their impact on viral control is influenced by preexisting viral variants in important target epitopes and the development of viral escape mutations. Immunodominant epitopes highly conserved across genotypes therefore are attractive for T cell based prophylactic vaccines. Here, we characterized the CD8(+) T cell response against the highly conserved HLA-B*51-restricted epitope IPFYGKAI1373-1380 located in the helicase domain of NS3 in people who inject drugs (PWID) exposed predominantly to HCV genotypes 1a and 3a. Despite this epitope being conserved in both genotypes, the corresponding CD8(+) T cell response was detected only in PWID infected with genotype 3a and HCV-RNA negative PWID, but not in PWID infected with genotype 1a. In genotype 3a, the detection of strong CD8(+) T cell responses was associated with epitope variants in the autologous virus consistent with immune escape. Analysis of viral sequences from multiple cohorts confirmed HLA-B*51-associated escape mutations inside the epitope in genotype 3a, but not in genotype 1a. Here, a distinct substitution in the N-terminal flanking region located 5 residues upstream of the epitope (S1368P; P = 0.00002) was selected in HLA-B*51-positive individuals. Functional assays revealed that the S1368P substitution impaired recognition of target cells presenting the endogenously processed epitope. The results highlight that, despite an epitope being highly conserved between two genotypes, there are major differences in the selected viral escape pathways and the corresponding T cell responses. HCV is able to evolutionary adapt to CD8(+) T cell immune pressure in multiple ways. Beyond selection of mutations inside targeted epitopes, this study demonstrates that HCV inhibits epitope processing by modification of the epitope flanking region under T cell immune pressure. Selection of a substitution five amino acids upstream of the

  5. Agonist-induced internalisation of the glucagon-like peptide-1 receptor is mediated by the Gαq pathway.

    Science.gov (United States)

    Thompson, Aiysha; Kanamarlapudi, Venkateswarlu

    2015-01-01

    The glucagon-like peptide-1 receptor (GLP-1R) is a G-protein-coupled receptor (GPCR) and an important target in the treatment of type 2 diabetes mellitus (T2DM). Upon stimulation with agonist, the GLP-1R signals through both Gαs and Gαq coupled pathways to stimulate insulin secretion. The agonist-induced GLP-1R internalisation has recently been shown to be important for insulin secretion. However, the molecular mechanisms underlying GLP-1R internalisation remain unknown. The aim of this study was to determine the role of GLP-1R downstream signalling pathways in its internalisation. Agonist-induced human GLP-1R (hGLP-1R) internalisation and activity were examined using a number of techniques including immunoblotting, ELISA, immunofluorescence and luciferase assays to determine cAMP production, intracellular Ca(2+) accumulation and ERK phosphorylation. Agonist-induced hGLP-1R internalisation is dependent on caveolin-1 and dynamin. Inhibition of the Gαq pathway but not the Gαs pathway affected hGLP-1R internalisation. Consistent with this, hGLP-1R mutant T149M and small-molecule agonists (compound 2 and compound B), which activate only the Gαs pathway, failed to induce internalisation of the receptor. Chemical inhibitors of the Gαq pathway, PKC and ERK phosphorylation significantly reduced agonist-induced hGLP-1R internalisation. These inhibitors also suppressed agonist-induced ERK1/2 phosphorylation demonstrating that the phosphorylated ERK acts downstream of the Gαq pathway in the hGLP-1R internalisation. In summary, agonist-induced hGLP-1R internalisation is mediated by the Gαq pathway. The internalised hGLP-1R stimulates insulin secretion from pancreatic β-cells, indicating the importance of GLP-1 internalisation for insulin secretion. Copyright © 2014 Elsevier Inc. All rights reserved.

  6. SREBP-1c overactivates ROS-mediated hepatic NF-κB inflammatory pathway in dairy cows with fatty liver.

    Science.gov (United States)

    Li, Xinwei; Huang, Weikun; Gu, Jingmin; Du, Xiliang; Lei, Lin; Yuan, Xue; Sun, Guoquan; Wang, Zhe; Li, Xiaobing; Liu, Guowen

    2015-10-01

    Dairy cows with fatty liver are characterized by hepatic lipid accumulation and a severe inflammatory response. Sterol receptor element binding protein-1c (SREBP-1c) and nuclear factor κB (NF-κB) are components of the main pathways for controlling triglyceride (TG) accumulation and inflammatory levels, respectively. A previous study demonstrated that hepatic inflammatory levels are positively correlated with hepatic TG content. We therefore speculated that SREBP-1c might play an important role in the overactivation of the hepatic NF-κB inflammatory pathway in cows with fatty liver. Compared with healthy cows, cows with fatty liver exhibited severe hepatic injury and high blood concentrations of the inflammatory cytokines TNF-α, IL-6 and IL-1β. Hepatic SREBP-1c-mediated lipid synthesis and the NF-κB inflammatory pathway were both overinduced in cows with fatty liver. In vitro, treatment with non-esterified fatty acids (NEFA) further increased SREBP-1c expression and NF-κB pathway activation, which then promoted TG and inflammatory cytokine synthesis. SREBP-1c overexpression overactivated the NF-κB inflammatory pathway in hepatocytes by increasing ROS content and not through TLR4. Furthermore, SREBP-1c silencing decreased ROS content and further attenuated the activation of the NEFA-induced NF-κB pathway, thereby decreasing TNF-α, IL-6 and IL-1β synthesis. SREBP-1c-overexpressing mice exhibited hepatic steatosis and an overinduced hepatic NF-κB pathway. Taken together, these results indicate that SREBP-1c enhances the NEFA-induced overactivation of the NF-κB inflammatory pathway by increasing ROS in cow hepatocytes, thereby further increasing hepatic inflammatory injury in cows with fatty liver. Copyright © 2015. Published by Elsevier Inc.

  7. Degradation of sulfadimethoxine catalyzed by laccase with soybean meal extract as natural mediator: Mechanism and reaction pathway.

    Science.gov (United States)

    Liang, Shangtao; Luo, Qi; Huang, Qingguo

    2017-08-01

    Natural laccase-mediator systems have been well recognized as an eco-friendly and energy-saving approach in environmental remediation, whose further application is however limited by the high cost of natural mediators and relatively long treatment time span. This study evaluated the water extract of soybean meal, a low-cost compound system, in mediating the laccase catalyzed degradation of a model contaminant of emerging concern, sulfadimethoxine (SDM), and demonstrated it as a promising alternative mediator for soil and water remediation. Removal of 73.3% and 65.6% was achieved in 9 h using soybean meal extract (SBE) as the mediating system for laccase-catalyzed degradation of sulfadimethoxine at the concentration of 1 ppm and 10 ppm, respectively. Further degradation of sulfadimethoxine was observed with multiple SBE additions. Using SBE as mediator increased the 9-h removal of SDM at 1 ppm initial concentration by 52.9%, 49.4%, and 36.3% in comparison to the system mediated by 1-Hydroxybenzotriazole (HBT), p-Coumaric acid (COU) and 2,2'-azinobis(3-ethylbenzthiazoline-6-sulfonate) (ABTS), respectively. With the detection of stable coupling products formed with radical scavenger (5,5-Dimethyl-1-pyrroline N-oxide, DMPO), three phenolic compounds (vanillin, apocynin, and daidzein) in SBE were confirmed to serve as mediators for Trametes versicolor laccase. Reaction pathways were proposed based on the results of High Resolution Mass Spectrometry. SO 2 excursion happened during SDM transformation, leading to elimination of antimicrobial activity. Therefore, as a natural, phenol rich, and affordable compound system, the future application of SBE in wastewater and soil remediation is worth exploring. Copyright © 2017 Elsevier Ltd. All rights reserved.

  8. The Mediator subunit SFR6/MED16 controls defence gene expression mediated by salicylic acid and jasmonate responsive pathways.

    Science.gov (United States)

    Wathugala, Deepthi L; Hemsley, Piers A; Moffat, Caroline S; Cremelie, Pieter; Knight, Marc R; Knight, Heather

    2012-07-01

    • Arabidopsis SENSITIVE TO FREEZING6 (SFR6) controls cold- and drought-inducible gene expression and freezing- and osmotic-stress tolerance. Its identification as a component of the MEDIATOR transcriptional co-activator complex led us to address its involvement in other transcriptional responses. • Gene expression responses to Pseudomonas syringae, ultraviolet-C (UV-C) irradiation, salicylic acid (SA) and jasmonic acid (JA) were investigated in three sfr6 mutant alleles by quantitative real-time PCR and susceptibility to UV-C irradiation and Pseudomonas infection were assessed. • sfr6 mutants were more susceptible to both Pseudomonas syringae infection and UV-C irradiation. They exhibited correspondingly weaker PR (pathogenesis-related) gene expression than wild-type Arabidopsis following these treatments or after direct application of SA, involved in response to both UV-C and Pseudomonas infection. Other genes, however, were induced normally in the mutants by these treatments. sfr6 mutants were severely defective in expression of plant defensin genes in response to JA; ectopic expression of defensin genes was provoked in wild-type but not sfr6 by overexpression of ERF5. • SFR6/MED16 controls both SA- and JA-mediated defence gene expression and is necessary for tolerance of Pseudomonas syringae infection and UV-C irradiation. It is not, however, a universal regulator of stress gene transcription and is likely to mediate transcriptional activation of specific regulons only. © 2012 The Authors. New Phytologist © 2012 New Phytologist Trust.

  9. Anti-apoptotic effect of heat shock protein 90 on hypoxia-mediated cardiomyocyte damage is mediated via the phosphatidylinositol 3-kinase/AKT pathway.

    Science.gov (United States)

    Wang, Wei; Peng, Yizhi; Wang, Yuanyuan; Zhao, Xiaohui; Yuan, Zhiqiang

    2009-09-01

    1. Hypoxia-induced cardiomyocyte apoptosis contributes significantly to cardiac dysfunction following trauma, shock and burn injury. There is evidence that heat shock protein (HSP) 90 is anti-apoptotic in cardiomyocytes subjected to a variety of apoptotic stimuli. Because HSP90 acts as an upstream regulator of the serine/threonine protein kinase Akt survival pathway during cellular stress, we hypothesized that HSP90 exerts a cardioprotective effect via the phosphatidylinositol 3-kinase (PI3-K)/Akt pathway. 2. Neonatal rat cardiomyocytes were subjected to normoxia or hypoxia in the absence or presence of the HSP90 inhibitor geldanamycin (1 μg/mL). Cardiomyocyte apoptosis was assessed by release of lactate dehydrogenase (LDH), terminal deoxyribonucleotidyl transferase-mediated dUTP-digoxigenin nick end-labelling (TUNEL) staining and caspase 3 activity. Expression of HSP90, Akt, Bad and cytochrome c release was determined by western blot analysis. 3. Following exposure of cells to hypoxia, HSP90 was markedly elevated in a time-dependent manner, reaching a peak at 6 h (eightfold increase). Geldanamycin significantly increased hypoxia-induced release of LDH by 114%, the percentage of apoptotic cardiomyocytes by 102% and caspase 3 activity by 78%. Pretreatment of cells with geldanamycin also suppressed phosphorylation of both Akt and its downstream target Bad, but promoted the mitochondrial release of cytochrome c. 4. In conclusion, HSP90 activity is enhanced in cardiomyocytes following hypoxic insult. The anti-apoptotic effect of HSP90 on cardiomyocytes subjected to hypoxia is mediated, at least in part, by the PI3-K/Akt pathway. Key words: apoptosis, cardiomyocyte, heart failure, heat shock protein 90, hypoxia, phosphatidylinositol 3-kinase/Akt signalling pathway, serine/threonine protein kinase Akt.

  10. Sodium fluoride induces apoptosis in mouse embryonic stem cells through ROS-dependent and caspase- and JNK-mediated pathways

    International Nuclear Information System (INIS)

    Nguyen Ngoc, Tam Dan; Son, Young-Ok; Lim, Shin-Saeng; Shi, Xianglin; Kim, Jong-Ghee; Heo, Jung Sun; Choe, Youngji; Jeon, Young-Mi; Lee, Jeong-Chae

    2012-01-01

    Sodium fluoride (NaF) is used as a source of fluoride ions in diverse applications. Fluoride salt is an effective prophylactic for dental caries and is an essential element required for bone health. However, fluoride is known to cause cytotoxicity in a concentration-dependent manner. Further, no information is available on the effects of NaF on mouse embryonic stem cells (mESCs). We investigated the mode of cell death induced by NaF and the mechanisms involved. NaF treatment greater than 1 mM reduced viability and DNA synthesis in mESCs and induced cell cycle arrest in the G 2 /M phase. The addition of NaF induced cell death mainly by apoptosis rather than necrosis. Catalase (CAT) treatment significantly inhibited the NaF-mediated cell death and also suppressed the NaF-mediated increase in phospho-c-Jun N-terminal kinase (p-JNK) levels. Pre-treatment with SP600125 or z-VAD-fmk significantly attenuated the NaF-mediated reduction in cell viability. In contrast, intracellular free calcium chelator, but not of sodium or calcium ion channel blockers, facilitated NaF-induced toxicity in the cells. A JNK specific inhibitor (SP600125) prevented the NaF-induced increase in growth arrest and the DNA damage-inducible protein 45α. Further, NaF-mediated loss of mitochondrial membrane potential was apparently inhibited by pifithrin-α or CAT inhibitor. These findings suggest that NaF affects viability of mESCs in a concentration-dependent manner, where more than 1 mM NaF causes apoptosis through hydroxyl radical-dependent and caspase- and JNK-mediated pathways. -- Highlights: ► The mode of NaF-induced cell death and the mechanisms involved were examined. ► NaF induced mainly apoptotic death of mouse embryonic stem cells (mESCs). ► NaF induced mitochondrial-mediated and caspase-dependent apoptosis. ► JNK- and p53-mediated pathways are involved in NaF-mediated apoptosis in the cells. ► ROS are the up-stream effector in NaF-mediated activation of JNK and p53 in mESCs.

  11. Sodium fluoride induces apoptosis in mouse embryonic stem cells through ROS-dependent and caspase- and JNK-mediated pathways

    Energy Technology Data Exchange (ETDEWEB)

    Nguyen Ngoc, Tam Dan [Institute of Oral Biosciences and School of Dentistry (BK21 Program), Chonbuk National University, Jeonju 561-756 (Korea, Republic of); Son, Young-Ok [Graduate Center for Toxicology, School of Medicine, University of Kentucky, Lexington, KY 40536-0305 (United States); Lim, Shin-Saeng [Institute of Oral Biosciences and School of Dentistry (BK21 Program), Chonbuk National University, Jeonju 561-756 (Korea, Republic of); Department of Bioactive Material Sciences and Research Center of Bioactive Materials, Chonbuk National University, Jeonju 561-756 (Korea, Republic of); Shi, Xianglin [Graduate Center for Toxicology, School of Medicine, University of Kentucky, Lexington, KY 40536-0305 (United States); Kim, Jong-Ghee [Institute of Oral Biosciences and School of Dentistry (BK21 Program), Chonbuk National University, Jeonju 561-756 (Korea, Republic of); Heo, Jung Sun [Department of Maxillofacial Biomedical Engineering and Institute of Oral Biology, School of Dentistry, Kyung Hee University, Seoul 130-701 (Korea, Republic of); Choe, Youngji [Institute of Oral Biosciences and School of Dentistry (BK21 Program), Chonbuk National University, Jeonju 561-756 (Korea, Republic of); Jeon, Young-Mi, E-mail: young@jbnu.ac.kr [Institute of Oral Biosciences and School of Dentistry (BK21 Program), Chonbuk National University, Jeonju 561-756 (Korea, Republic of); Lee, Jeong-Chae, E-mail: leejc88@jbnu.ac.kr [Institute of Oral Biosciences and School of Dentistry (BK21 Program), Chonbuk National University, Jeonju 561-756 (Korea, Republic of); Graduate Center for Toxicology, School of Medicine, University of Kentucky, Lexington, KY 40536-0305 (United States); Department of Bioactive Material Sciences and Research Center of Bioactive Materials, Chonbuk National University, Jeonju 561-756 (Korea, Republic of)

    2012-03-15

    Sodium fluoride (NaF) is used as a source of fluoride ions in diverse applications. Fluoride salt is an effective prophylactic for dental caries and is an essential element required for bone health. However, fluoride is known to cause cytotoxicity in a concentration-dependent manner. Further, no information is available on the effects of NaF on mouse embryonic stem cells (mESCs). We investigated the mode of cell death induced by NaF and the mechanisms involved. NaF treatment greater than 1 mM reduced viability and DNA synthesis in mESCs and induced cell cycle arrest in the G{sub 2}/M phase. The addition of NaF induced cell death mainly by apoptosis rather than necrosis. Catalase (CAT) treatment significantly inhibited the NaF-mediated cell death and also suppressed the NaF-mediated increase in phospho-c-Jun N-terminal kinase (p-JNK) levels. Pre-treatment with SP600125 or z-VAD-fmk significantly attenuated the NaF-mediated reduction in cell viability. In contrast, intracellular free calcium chelator, but not of sodium or calcium ion channel blockers, facilitated NaF-induced toxicity in the cells. A JNK specific inhibitor (SP600125) prevented the NaF-induced increase in growth arrest and the DNA damage-inducible protein 45α. Further, NaF-mediated loss of mitochondrial membrane potential was apparently inhibited by pifithrin-α or CAT inhibitor. These findings suggest that NaF affects viability of mESCs in a concentration-dependent manner, where more than 1 mM NaF causes apoptosis through hydroxyl radical-dependent and caspase- and JNK-mediated pathways. -- Highlights: ► The mode of NaF-induced cell death and the mechanisms involved were examined. ► NaF induced mainly apoptotic death of mouse embryonic stem cells (mESCs). ► NaF induced mitochondrial-mediated and caspase-dependent apoptosis. ► JNK- and p53-mediated pathways are involved in NaF-mediated apoptosis in the cells. ► ROS are the up-stream effector in NaF-mediated activation of JNK and p53 in mESCs.

  12. GABA, not glycine, mediates inhibition of latent respiratory motor pathways after spinal cord injury

    OpenAIRE

    Zimmer, M. Beth; Goshgarian, Harry G.

    2006-01-01

    Previous work has shown that latent respiratory motor pathways known as crossed phrenic pathways are inhibited via a spinal inhibitory process; however, the underlying mechanisms remain unknown. The present study investigated whether spinal GABA-A and/or glycine receptors are involved in the inhibition of the crossed phrenic pathways after a C2 spinal cord hemisection injury. Under ketamine/xylazine anesthesia, adult, female, Sprague Dawley rats were hemisected at the C2 spinal cord level. Fo...

  13. An archaeal CRISPR type III-B system exhibiting distinctive RNA targeting features and mediating dual RNA and DNA interference

    DEFF Research Database (Denmark)

    Peng, Wenfang; Feng, Mingxia; Feng, Xu

    2015-01-01

    CRISPR-Cas systems provide a small RNA-based mechanism to defend against invasive genetic elements in archaea and bacteria. To investigate the in vivo mechanism of RNA interference by two type III-B systems (Cmr-α and Cmr-β) in Sulfolobus islandicus, a genetic assay was developed using plasmids...... carrying an artificial mini-CRISPR (AC) locus with a single spacer. After pAC plasmids were introduced into different strains, Northern analyses confirmed that mature crRNAs were produced from the plasmid-borne CRISPR loci, which then guided gene silencing to target gene expression. Spacer mutagenesis....... islandicus Cmr-α mediated transcription-dependent DNA interference, the Cmr-α constitutes the first CRISPR system exhibiting dual targeting of RNA and DNA....

  14. Stanniocalcin-1 Protects a Mouse Model from Renal Ischemia-Reperfusion Injury by Affecting ROS-Mediated Multiple Signaling Pathways.

    Science.gov (United States)

    Liu, Dajun; Shang, Huiping; Liu, Ying

    2016-07-12

    Stanniocalcin-1 (STC-1) protects against renal ischemia-reperfusion injury (RIRI). However, the molecular mechanisms remain widely unknown. STC-1 inhibits reactive oxygen species (ROS), whereas most ROS-mediated pathways are associated with ischemic injury. Therefore, to explore the mechanism, the effects of STC-1 on ROS-medicated pathways were studied. Non-traumatic vascular clamps were used to establish RIRI mouse models. The serum levels of STC-1, interleukin-6 (IL-6), interferon (IFN) γ, P53, and capase-3 were measured by ELISA kits. Superoxide dismutase (SOD) and malondialdehyde (MDA) were measured by fluorescence spectrofluorometer. All these molecules changed significantly in a RIRI model mouse when compared with those in a sham control. Kidney cells were isolated from sham and model mice. STC-1 was overexpressed or knockout in these kidney cells. The molecules in ROS-medicated pathways were measured by real-time quantitative PCR and Western blot. The results showed that STC-1 is an effective ROS scavenger. The serum levels of STC-1, MDA and SOD activity were increased while the serum levels of IL-6, iIFN-γ, P53, and capase-3 were decreased in a model group when compared with a sham control (p ROS-mediated molecules. Therefore, STC-1 maybe improve anti-inflammation, anti-oxidant and anti-apoptosis activities by affecting ROS-mediated pathways, especially the phospho-modifications of the respective proteins, resulting in the increase of SOD and reduce of capase-3, p53, IL-6 and IFN-γ.

  15. Protein tyrosine kinase and mitogen-activated protein kinase signalling pathways contribute to differences in heterophil-mediated innate immune responsiveness between two lines of broilers

    Science.gov (United States)

    Protein tyrosine phosphorylation mediates signal transduction of cellular processes, with protein tyrosine kinases (PTKs) regulating virtually all signaling events. The mitogen-activated protein kinase (MAPK) super-family consists of three conserved pathways that convert receptor activation into ce...

  16. Examining a pathway for hormone mediated maternal effects - Yolk testosterone affects androgen receptor expression and endogenous testosterone production in young chicks (Gallus gallus domesticus)

    NARCIS (Netherlands)

    Pfannkuche, K. A.; Gahr, M.; Weites, I. M.; Riedstra, B.; Wolf, C.; Groothuis, T. G. G.

    2011-01-01

    In vertebrates maternal androgens can substantially influence developing offspring, inducing both short and long term changes in physiology and behavior, including androgen sensitive traits. However, how the effects of maternal hormones are mediated remains unknown. Two possible pathways are that

  17. DMPD: Modulation of Toll-interleukin 1 receptor mediated signaling. [Dynamic Macrophage Pathway CSML Database

    Lifescience Database Archive (English)

    Full Text Available 15662540 Modulation of Toll-interleukin 1 receptor mediated signaling. Li X, Qin J.... J Mol Med. 2005 Apr;83(4):258-66. Epub 2005 Jan 21. (.png) (.svg) (.html) (.csml) Show Modulation of Toll-i...nterleukin 1 receptor mediated signaling. PubmedID 15662540 Title Modulation of Toll-interleukin 1 receptor

  18. Caffeine Inhibits the Activation of Hepatic Stellate Cells Induced by Acetaldehyde via Adenosine A2A Receptor Mediated by the cAMP/PKA/SRC/ERK1/2/P38 MAPK Signal Pathway

    Science.gov (United States)

    Yang, Wanzhi; Wang, Qi; Zhao, Han; Yang, Feng; Lv, Xiongwen; Li, Jun

    2014-01-01

    Hepatic stellate cell (HSC) activation is an essential event during alcoholic liver fibrosis. Evidence suggests that adenosine aggravates liver fibrosis via the adenosine A2A receptor (A2AR). Caffeine, which is being widely consumed during daily life, inhibits the action of adenosine. In this study, we attempted to validate the hypothesis that caffeine influences acetaldehyde-induced HSC activation by acting on A2AR. Acetaldehyde at 50, 100, 200, and 400 μM significantly increased HSC-T6 cells proliferation, and cell proliferation reached a maximum at 48 h after exposure to 200 μM acetaldehyde. Caffeine and the A2AR antagonist ZM241385 decreased the cell viability and inhibited the expression of procollagen type I and type III in acetaldehyde-induced HSC-T6 cells. In addition, the inhibitory effect of caffeine on the expression of procollagen type I was regulated by A2AR-mediated signal pathway involving cAMP, PKA, SRC, and ERK1/2. Interestingly, caffeine’s inhibitory effect on the expression of procollagen type III may depend upon the A2AR-mediated P38 MAPK-dependent pathway. Conclusions: Caffeine significantly inhibited acetaldehyde-induced HSC-T6 cells activation by distinct A2AR mediated signal pathway via inhibition of cAMP-PKA-SRC-ERK1/2 for procollagen type I and via P38 MAPK for procollagen type III. PMID:24682220

  19. The octadecanoid signalling pathway in plants mediates a response to ultraviolet radiation

    International Nuclear Information System (INIS)

    Conconi, A.; Smerdon, M.J.; Howe, G.A.; Ryan, C.A.

    1996-01-01

    Many plant genes that respond to environmental and developmental changes are regulated by jasmonic acid, which is derived from linolenic acid via the octadecanoid pathway. Linolenic acid is an important fatty-acid constituent of membranes in most plant species and its intracellular levels increase in response to certain signals. Here we report that irradiation of tomato leaves with ultraviolet light induces the expression of several plant defensive genes that are normally activated through the octadecanoid pathway after wounding. The response to ultraviolet light is blocked by an inhibitor of the octadecanoid pathway and it does not occur in a tomato mutant defective in this pathway. The ultraviolet irradiation maximally induces the defence genes at levels where cyclobutane pyrimidine dimer formation, an indicator of DNA damage, is less than 0.2 dimers per gene. Our evidence indicates that this plant defence response to certain wavelengths of ultraviolet radiation requires the activation of the octadecanoid defence signalling pathway. (author)

  20. Two Distinct Approaches for CRISPR-Cas9-Mediated Gene Editing in Cryptococcus neoformans and Related Species.

    Science.gov (United States)

    Wang, Ping

    2018-06-27

    Cryptococcus neoformans and related species are encapsulated basidiomycetous fungi that cause meningoencephalitis in individuals with immune deficiency. This pathogen has a tractable genetic system; however, gene disruption via electroporation remains difficult, while biolistic transformation is often limited by lack of multiple genetic markers and the high initial cost of equipment. The approach using clustered regularly interspaced short palindromic repeats (CRISPR) and CRISPR-associated protein 9 (Cas9) has become the technology of choice for gene editing in many organisms due to its simplicity, efficiency, and versatility. The technique has been successfully demonstrated in C. neoformans and Cryptococcus deneoformans in which two DNA plasmids expressing either the Streptococcus pyogenes CAS9 gene or the guide RNA (gRNA) were employed. However, potential adverse effects due to constitutive expression and the time-consuming process of constructing vectors to express each gRNA remain as a primary barrier for wide adaptation. This report describes the delivery of preassembled CRISPR-Cas9-gRNA ribonucleoproteins (RNPs) via electroporation that is able to generate edited mutant alleles. RNP-mediated CRISPR-Cas9 was used to replace the wild-type GIB2 gene encoding a Gβ-like/RACK1 Gib2 protein with a gib2 :: NAT allele via homologous recombination in both C. neoformans and C. deneoformans In addition, a DNA plasmid (pCnCas9:U6-gRNA) that expresses both Cas9 and gRNA, allowing for convenient yet low-cost DNA-mediated gene editing, is described. pCnCas9:U6-gRNA contains an endogenous U6 promoter for gRNA expression and restriction sites for one-step insertion of a gRNA. These approaches and resources provide new opportunities to accelerate genetic studies of Cryptococcus species. IMPORTANCE For genetic studies of the Cryptococcus genus, generation of mutant strains is often hampered by a limited number of selectable genetic markers, the tedious process of vector

  1. TSLP signaling pathway map: a platform for analysis of TSLP-mediated signaling.

    Science.gov (United States)

    Zhong, Jun; Sharma, Jyoti; Raju, Rajesh; Palapetta, Shyam Mohan; Prasad, T S Keshava; Huang, Tai-Chung; Yoda, Akinori; Tyner, Jeffrey W; van Bodegom, Diederik; Weinstock, David M; Ziegler, Steven F; Pandey, Akhilesh

    2014-01-01

    Thymic stromal lymphopoietin (TSLP) is a four-helix bundle cytokine that plays a critical role in the regulation of immune responses and in the differentiation of hematopoietic cells. TSLP signals through a heterodimeric receptor complex consisting of an interleukin-7 receptor α chain and a unique TSLP receptor (TSLPR) [also known as cytokine receptor-like factor 2 (CRLF2)]. Cellular targets of TSLP include dendritic cells, B cells, mast cells, regulatory T (Treg) cells and CD4+ and CD8+ T cells. The TSLP/TSLPR axis can activate multiple signaling transduction pathways including the JAK/STAT pathway and the PI-3 kinase pathway. Aberrant TSLP/TSLPR signaling has been associated with a variety of human diseases including asthma, atopic dermatitis, nasal polyposis, inflammatory bowel disease, eosinophilic eosophagitis and, most recently, acute lymphoblastic leukemia. A centralized resource of the TSLP signaling pathway cataloging signaling events is not yet available. In this study, we present a literature-annotated resource of reactions in the TSLP signaling pathway. This pathway map is publicly available through NetPath (http://www.netpath.org/), an open access signal transduction pathway resource developed previously by our group. This map includes 236 molecules and 252 reactions that are involved in TSLP/TSLPR signaling pathway. We expect that the TSLP signaling pathway map will provide a rich resource to study the biology of this important cytokine as well as to identify novel therapeutic targets for diseases associated with dysregulated TSLP/TSLPR signaling. Database URL: http://www.netpath.org/pathways?path_id=NetPath_24.

  2. Distinct unfolded protein responses mitigate or mediate effects of nonlethal deprivation of C. elegans sleep in different tissues.

    Science.gov (United States)

    Sanders, Jarred; Scholz, Monika; Merutka, Ilaria; Biron, David

    2017-08-28

    Disrupting sleep during development leads to lasting deficits in chordates and arthropods. To address lasting impacts of sleep deprivation in Caenorhabditis elegans, we established a nonlethal deprivation protocol. Deprivation triggered protective insulin-like signaling and two unfolded protein responses (UPRs): the mitochondrial (UPR mt ) and the endoplasmic reticulum (UPR ER ) responses. While the latter is known to be triggered by sleep deprivation in rodent and insect brains, the former was not strongly associated with sleep deprivation previously. We show that deprivation results in a feeding defect when the UPR mt is deficient and in UPR ER -dependent germ cell apoptosis. In addition, when the UPR ER is deficient, deprivation causes excess twitching in vulval muscles, mirroring a trend caused by loss of egg-laying command neurons. These data show that nonlethal deprivation of C. elegans sleep causes proteotoxic stress. Unless mitigated, distinct types of deprivation-induced proteotoxicity can lead to anatomically and genetically separable lasting defects. The relative importance of different UPRs post-deprivation likely reflects functional, developmental, and genetic differences between the respective tissues and circuits.

  3. Leukemia-Associated Nup214 Fusion Proteins Disturb the XPO1-Mediated Nuclear-Cytoplasmic Transport Pathway and Thereby the NF-κB Signaling Pathway.

    Science.gov (United States)

    Saito, Shoko; Cigdem, Sadik; Okuwaki, Mitsuru; Nagata, Kyosuke

    2016-07-01

    Nuclear-cytoplasmic transport through nuclear pore complexes is mediated by nuclear transport receptors. Previous reports have suggested that aberrant nuclear-cytoplasmic transport due to mutations or overexpression of nuclear pore complexes and nuclear transport receptors is closely linked to diseases. Nup214, a component of nuclear pore complexes, has been found as chimeric fusion proteins in leukemia. Among various Nup214 fusion proteins, SET-Nup214 and DEK-Nup214 have been shown to be engaged in tumorigenesis, but their oncogenic mechanisms remain unclear. In this study, we examined the functions of the Nup214 fusion proteins by focusing on their effects on nuclear-cytoplasmic transport. We found that SET-Nup214 and DEK-Nup214 interact with exportin-1 (XPO1)/CRM1 and nuclear RNA export factor 1 (NXF1)/TAP, which mediate leucine-rich nuclear export signal (NES)-dependent protein export and mRNA export, respectively. SET-Nup214 and DEK-Nup214 decreased the XPO1-mediated nuclear export of NES proteins such as cyclin B and proteins involved in the NF-κB signaling pathway by tethering XPO1 onto nuclear dots where Nup214 fusion proteins are localized. We also demonstrated that SET-Nup214 and DEK-Nup214 expression inhibited NF-κB-mediated transcription by abnormal tethering of the complex containing p65 and its inhibitor, IκB, in the nucleus. These results suggest that SET-Nup214 and DEK-Nup214 perturb the regulation of gene expression through alteration of the nuclear-cytoplasmic transport system. Copyright © 2016, American Society for Microbiology. All Rights Reserved.

  4. Comparative Transcriptomic and Proteomic Analyses Reveal a FluG-Mediated Signaling Pathway Relating to Asexual Sporulation of Antrodia camphorata.

    Science.gov (United States)

    Li, Hua-Xiang; Lu, Zhen-Ming; Zhu, Qing; Gong, Jin-Song; Geng, Yan; Shi, Jin-Song; Xu, Zheng-Hong; Ma, Yan-He

    2017-09-01

    Medicinal mushroom Antrodia camphorata sporulate large numbers of arthroconidia in submerged fermentation, which is rarely reported in basidiomycetous fungi. Nevertheless, the molecular mechanisms underlying this asexual sporulation (conidiation) remain unclear. Here, we used comparative transcriptomic and proteomic approaches to elucidate possible signaling pathway relating to the asexual sporulation of A. camphorata. First, 104 differentially expressed proteins and 2586 differential cDNA sequences during the culture process of A. camphorata were identified by 2DE and RNA-seq, respectively. By applying bioinformatics analysis, a total of 67 genes which might play roles in the sporulation were obtained, and 18 of these genes, including fluG, sfgA, SfaD, flbA, flbB, flbC, flbD, nsdD, brlA, abaA, wetA, ganB, fadA, PkaA, veA, velB, vosA, and stuA might be involved in a potential FluG-mediated signaling pathway. Furthermore, the mRNA expression levels of the 18 genes in the proposed FluG-mediated signaling pathway were analyzed by quantitative real-time PCR. In summary, our study helps elucidate the molecular mechanisms underlying the asexual sporulation of A. camphorata, and provides also useful transcripts and proteome for further bioinformatics study of this valuable medicinal mushroom. © 2017 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  5. Myeloid-derived suppressor activity is mediated by monocytic lineages maintained by continuous inhibition of extrinsic and intrinsic death pathways.

    Science.gov (United States)

    Haverkamp, Jessica M; Smith, Amber M; Weinlich, Ricardo; Dillon, Christopher P; Qualls, Joseph E; Neale, Geoffrey; Koss, Brian; Kim, Young; Bronte, Vincenzo; Herold, Marco J; Green, Douglas R; Opferman, Joseph T; Murray, Peter J

    2014-12-18

    Nonresolving inflammation expands a heterogeneous population of myeloid suppressor cells capable of inhibiting T cell function. This heterogeneity has confounded the functional dissection of individual myeloid subpopulations and presents an obstacle for antitumor immunity and immunotherapy. Using genetic manipulation of cell death pathways, we found the monocytic suppressor-cell subset, but not the granulocytic subset, requires continuous c-FLIP expression to prevent caspase-8-dependent, RIPK3-independent cell death. Development of the granulocyte subset requires MCL-1-mediated control of the intrinsic mitochondrial death pathway. Monocytic suppressors tolerate the absence of MCL-1 provided cytokines increase expression of the MCL-1-related protein A1. Monocytic suppressors mediate T cell suppression, whereas their granulocytic counterparts lack suppressive function. The loss of the granulocytic subset via conditional MCL-1 deletion did not alter tumor incidence implicating the monocytic compartment as the functionally immunosuppressive subset in vivo. Thus, death pathway modulation defines the development, survival, and function of myeloid suppressor cells. Copyright © 2014 Elsevier Inc. All rights reserved.

  6. The role of Ca(2+) mediated signaling pathways on the effect of taurine against Streptococcus uberis infection.

    Science.gov (United States)

    Dai, Bin; Zhang, Jinqiu; Liu, Ming; Lu, Jinye; Zhang, Yuanshu; Xu, Yuanyuan; Miao, Jinfeng; Yin, Yulong

    2016-08-30

    To provide insight into the mechanisms of taurine attenuation of pro-inflammatory response in mouse mammary epithelial cell line (EpH4-Ev, purchased by ATCC, USA) after Streptococcus uberis (S. uberis, 0140J) challenge, we infected MECs with S. uberis (2.5×10(7)cfumL(-1), MOI=10) for 3h and quantified changes in TLR-2 and calcium (Ca(2+)) mediated signaling pathways. The results indicate that S. uberis infection significantly increases the expression of TLR-2, intracellular Ca(2+) levels, PLC-γ1 and PKC-α, the activities of transcription factors NF-κB and NFAT, and related cytokines (TNF-α, IL-1β, IL-6, G-CSF, IL-2, KC, IL-15, FasL, MCP-1, and LIX) in culture supernatants. Taurine administration downregulated all these indices, the activities of NF-κB and NFAT. Cytokine secretions were similar using special PKC inhibitor Go 6983 and NFAT inhibitor VIVIT. Our data indicate that S. uberis infection induces pro-inflammatory response of MECs through a TLR-2 mediated signaling pathway. In addition, taurine can prevent MEC damage by affecting both PLC-γ1-Ca(2+)-PKC-α-NF-κB and PLC-γ1-Ca(2+)-NFATs signaling pathways. This is the first report to demonstrate the mechanisms of taurine attenuated pro-inflammatory response in MECs after S. uberis challenge. Copyright © 2016 Elsevier B.V. All rights reserved.

  7. Distinct sperm nucleus behaviors between genotypic and temperature-dependent sex determination males are associated with replication and expression-related pathways in a gynogenetic fish.

    Science.gov (United States)

    Zhu, Yao-Jun; Li, Xi-Yin; Zhang, Jun; Li, Zhi; Ding, Miao; Zhang, Xiao-Juan; Zhou, Li; Gui, Jian-Fang

    2018-06-05

    Coexistence and transition of diverse sex determination strategies have been revealed in some ectothermic species, but the variation between males caused by different sex determination strategies and the underlying mechanism remain unclear. Here, we used the gynogenetic gibel carp (Carassius gibelio) with both genotypic sex determination (GSD) and temperature-dependent sex determination (TSD) strategies to illustrate this issue. We found out that males of GSD and TSD in gibel carp had similar morphology, testicular histology, sperm structure and sperm vitality. However, when maternal individuals were mated with males of GSD, sperm nucleus swelling and fusing with the female pronucleus were observed in the fertilized eggs. On the contrary, when maternal individuals were mated with males of TSD, sperm nucleus remained in the condensed status throughout the whole process. Subsequently, semen proteomics analysis unveiled that DNA replication and gene expression-related pathways were inhibited in the sperm from males of TSD compared to males of GSD, and most differentially expressed proteins associated with DNA replication, transcription and translation were down-regulated. Moreover, via BrdU incorporation and immunofluorescence detection, male nucleus replication was revealed to be present in the fertilized eggs by the sperm from males of GSD, but absent in the fertilized eggs by the sperm from males of TSD. These findings indicate that DNA replication and gene expression-related pathways are associated with the distinct sperm nucleus development behaviors in fertilized eggs in response to the sperm from males of GSD and TSD. And this study is the first attempt to screen the differences between males determined via GSD and TSD in gynogenetic species, which might give a hint for understanding evolutionary adaption of diverse sex determination mechanisms in unisexual vertebrates.

  8. PSM/SH2-B distributes selected mitogenic receptor signals to distinct components in the PI3-kinase and MAP kinase signaling pathways.

    Science.gov (United States)

    Deng, Youping; Xu, Hu; Riedel, Heimo

    2007-02-15

    The Pro-rich, PH, and SH2 domain containing mitogenic signaling adapter PSM/SH2-B has been implicated as a cellular partner of various mitogenic receptor tyrosine kinases and related signaling mechanisms. Here, we report in a direct comparison of three peptide hormones, that PSM participates in the assembly of distinct mitogenic signaling complexes in response to insulin or IGF-I when compared to PDGF in cultured normal fibroblasts. The complex formed in response to insulin or IGF-I involves the respective peptide hormone receptor and presumably the established components leading to MAP kinase activation. However, our data suggest an alternative link from the PDGF receptor via PSM directly to MEK1/2 and consequently also to p44/42 activation, possibly through a scaffold protein. At least two PSM domains participate, the SH2 domain anticipated to link PSM to the respective receptor and the Pro-rich region in an association with an unidentified downstream component resulting in direct MEK1/2 and p44/42 regulation. The PDGF receptor signaling complex formed in response to PDGF involves PI 3-kinase in addition to the same components and interactions as described for insulin or IGF-I. PSM associates with PI 3-kinase via p85 and in addition the PSM PH domain participates in the regulation of PI 3-kinase activity, presumably through membrane interaction. In contrast, the PSM Pro-rich region appears to participate only in the MAP kinase signal. Both pathways contribute to the mitogenic response as shown by cell proliferation, survival, and focus formation. PSM regulates p38 MAP kinase activity in a pathway unrelated to the mitogenic response.

  9. STAT1 pathway mediates amplification of metastatic potential and resistance to therapy.

    Directory of Open Access Journals (Sweden)

    Nikolai N Khodarev

    Full Text Available BACKGROUND: Traditionally IFN/STAT1 signaling is connected with an anti-viral response and pro-apoptotic tumor-suppressor functions. Emerging functions of a constitutively activated IFN/STAT1 pathway suggest an association with an aggressive tumor phenotype. We hypothesized that tumor clones that constitutively overexpress this pathway are preferentially selected by the host microenvironment due to a resistance to STAT1-dependent cytotoxicity and demonstrate increased metastatic ability combined with increased resistance to genotoxic stress. METHODOLOGY/PRINCIPAL FINDINGS: Here we report that clones of B16F1 tumors grown in the lungs of syngeneic C57BL/6 mice demonstrate variable transcriptional levels of IFN/STAT1 pathway expression. Tumor cells that constitutively overexpress the IFN/STAT1 pathway (STAT1(H genotype are selected by the lung microenvironment. STAT1(H tumor cells also demonstrate resistance to IFN-gamma (IFNgamma, ionizing radiation (IR, and doxorubicin relative to parental B16F1 and low expressors of the IFN/STAT1 pathway (STAT1(L genotype. Stable knockdown of STAT1 reversed the aggressive phenotype and decreased both lung colonization and resistance to genotoxic stress. CONCLUSIONS: Our results identify a pathway activated by tumor-stromal interactions thereby selecting for pro-metastatic and therapy-resistant tumor clones. New therapies targeted against the IFN/STAT1 signaling pathway may provide an effective strategy to treat or sensitize aggressive tumor clones to conventional cancer therapies and potentially prevent distant organ colonization.

  10. Molecular pathway profiling of T lymphocyte signal transduction pathways; Th1 and Th2 genomic fingerprints are defined by TCR and CD28-mediated signaling

    Directory of Open Access Journals (Sweden)

    Smeets Ruben L

    2012-03-01

    Full Text Available Abstract Background T lymphocytes are orchestrators of adaptive immunity. Naïve T cells may differentiate into Th1, Th2, Th17 or iTreg phenotypes, depending on environmental co-stimulatory signals. To identify genes and pathways involved in differentiation of Jurkat T cells towards Th1 and Th2 subtypes we performed comprehensive transcriptome analyses of Jurkat T cells stimulated with various stimuli and pathway inhibitors. Results from these experiments were validated in a human experimental setting using whole blood and purified CD4+ Tcells. Results Calcium-dependent activation of T cells using CD3/CD28 and PMA/CD3 stimulation induced a Th1 expression profile reflected by increased expression of T-bet, RUNX3, IL-2, and IFNγ, whereas calcium-independent activation via PMA/CD28 induced a Th2 expression profile which included GATA3, RXRA, CCL1 and Itk. Knock down with siRNA and gene expression profiling in the presence of selective kinase inhibitors showed that proximal kinases Lck and PKCθ are crucial signaling hubs during T helper cell activation, revealing a clear role for Lck in Th1 development and for PKCθ in both Th1 and Th2 development. Medial signaling via MAPkinases appeared to be less important in these pathways, since specific inhibitors of these kinases displayed a minor effect on gene expression. Translation towards a primary, whole blood setting and purified human CD4+ T cells revealed that PMA/CD3 stimulation induced a more pronounced Th1 specific, Lck and PKCθ dependent IFNγ production, whereas PMA/CD28 induced Th2 specific IL-5 and IL-13 production, independent of Lck activation. PMA/CD3-mediated skewing towards a Th1 phenotype was also reflected in mRNA expression of the master transcription factor Tbet, whereas PMA/CD28-mediated stimulation enhanced GATA3 mRNA expression in primary human CD4+ Tcells. Conclusions This study identifies stimulatory pathways and gene expression profiles for in vitro skewing of T helper cell

  11. The mitochondrial fatty acid synthesis (mtFASII) pathway is capable of mediating nuclear-mitochondrial cross talk through the PPAR system of transcriptional activation

    International Nuclear Information System (INIS)

    Parl, Angelika; Mitchell, Sabrina L.; Clay, Hayley B.; Reiss, Sara; Li, Zhen; Murdock, Deborah G.

    2013-01-01

    Highlights: •The function of the mitochondria fatty acid synthesis pathway is partially unknown. •Overexpression of the pathway causes transcriptional activation through PPARs. •Knock down of the pathway attenuates that activation. •The last enzyme in the pathway regulates its own transcription. •Products of the mtFASII pathway are able to drive nuclear transcription. -- Abstract: Mammalian cells contain two fatty acid synthesis pathways, the cytosolic FASI pathway, and the mitochondrial FASII pathway. The selection behind the conservation of the mitochondrial pathway is not completely understood, given the presence of the cytosolic FAS pathway. In this study, we show through heterologous gene reporter systems and PCR-based arrays that overexpression of MECR, the last step in the mtFASII pathway, causes modulation of gene expression through the PPAR pathway. Electromobility shift assays (EMSAs) demonstrate that overexpression of MECR causes increased binding of PPARs to DNA, while cell fractionation and imaging studies show that MECR remains localized to the mitochondria. Interestingly, knock down of the mtFASII pathway lessens the effect of MECR on this transcriptional modulation. Our data are most consistent with MECR-mediated transcriptional activation through products of the mtFASII pathway, although we cannot rule out MECR acting as a coactivator. Further investigation into the physiological relevance of this communication will be necessary to better understand some of the phenotypic consequences of deficits in this pathway observed in animal models and human disease

  12. The mitochondrial fatty acid synthesis (mtFASII) pathway is capable of mediating nuclear-mitochondrial cross talk through the PPAR system of transcriptional activation

    Energy Technology Data Exchange (ETDEWEB)

    Parl, Angelika; Mitchell, Sabrina L.; Clay, Hayley B.; Reiss, Sara; Li, Zhen; Murdock, Deborah G., E-mail: deborah.murdock@vanderbilt.edu

    2013-11-15

    Highlights: •The function of the mitochondria fatty acid synthesis pathway is partially unknown. •Overexpression of the pathway causes transcriptional activation through PPARs. •Knock down of the pathway attenuates that activation. •The last enzyme in the pathway regulates its own transcription. •Products of the mtFASII pathway are able to drive nuclear transcription. -- Abstract: Mammalian cells contain two fatty acid synthesis pathways, the cytosolic FASI pathway, and the mitochondrial FASII pathway. The selection behind the conservation of the mitochondrial pathway is not completely understood, given the presence of the cytosolic FAS pathway. In this study, we show through heterologous gene reporter systems and PCR-based arrays that overexpression of MECR, the last step in the mtFASII pathway, causes modulation of gene expression through the PPAR pathway. Electromobility shift assays (EMSAs) demonstrate that overexpression of MECR causes increased binding of PPARs to DNA, while cell fractionation and imaging studies show that MECR remains localized to the mitochondria. Interestingly, knock down of the mtFASII pathway lessens the effect of MECR on this transcriptional modulation. Our data are most consistent with MECR-mediated transcriptional activation through products of the mtFASII pathway, although we cannot rule out MECR acting as a coactivator. Further investigation into the physiological relevance of this communication will be necessary to better understand some of the phenotypic consequences of deficits in this pathway observed in animal models and human disease.

  13. RNF4-mediated polyubiquitination regulates the Fanconi anemia/BRCA pathway

    OpenAIRE

    Xie, Jenny; Kim, Hyungjin; Moreau, Lisa A.; Puhalla, Shannon; Garber, Judy; Al Abo, Muthana; Takeda, Shunichi; D’Andrea, Alan D.

    2015-01-01

    The Fanconi anemia/BRCA (FA/BRCA) pathway is a DNA repair pathway that is required for excision of DNA interstrand cross-links. The 17 known FA proteins, along with several FA-associated proteins (FAAPs), cooperate in this pathway to detect, unhook, and excise DNA cross-links and to subsequently repair the double-strand breaks generated in the process. In the current study, we identified a patient with FA with a point mutation in FANCA, which encodes a mutant FANCA protein (FANCAI939S). FANCA...

  14. Ursolic acid-mediated changes in glycolytic pathway promote cytotoxic autophagy and apoptosis in phenotypically different breast cancer cells.

    Science.gov (United States)

    Lewinska, Anna; Adamczyk-Grochala, Jagoda; Kwasniewicz, Ewa; Deregowska, Anna; Wnuk, Maciej

    2017-06-01

    Plant-derived pentacyclic triterpenotids with multiple biological activities are considered as promising candidates for cancer therapy and prevention. However, their mechanisms of action are not fully understood. In the present study, we have analyzed the effects of low dose treatment (5-20 µM) of ursolic acid (UA) and betulinic acid (BA) on breast cancer cells of different receptor status, namely MCF-7 (ER + , PR +/- , HER2 - ), MDA-MB-231 (ER - , PR - , HER2 - ) and SK-BR-3 (ER - , PR - , HER2 + ). UA-mediated response was more potent than BA-mediated response. Triterpenotids (5-10 µM) caused G0/G1 cell cycle arrest, an increase in p21 levels and SA-beta-galactosidase staining that was accompanied by oxidative stress and DNA damage. UA (20 µM) also diminished AKT signaling that affected glycolysis as judged by decreased levels of HK2, PKM2, ATP and lactate. UA-induced energy stress activated AMPK that resulted in cytotoxic autophagy and apoptosis. UA-mediated elevation in nitric oxide levels and ATM activation may also account for AMPK activation-mediated cytotoxic response. Moreover, UA-promoted apoptosis was associated with decreased pERK1/2 signals and the depolarization of mitochondrial membrane potential. Taken together, we have shown for the first time that UA at low micromolar range may promote its anticancer action by targeting glycolysis in phenotypically distinct breast cancer cells.

  15. Two Sets of Piwi Proteins Are Involved in Distinct sRNA Pathways Leading to Elimination of Germline-Specific DNA

    Directory of Open Access Journals (Sweden)

    Dominique I. Furrer

    2017-07-01

    Full Text Available Piwi proteins and piRNAs protect eukaryotic germlines against the spread of transposons. During development in the ciliate Paramecium, two Piwi-dependent sRNA classes are involved in the elimination of transposons and transposon-derived DNA: scan RNAs (scnRNAs, associated with Ptiwi01 and Ptiwi09, and iesRNAs, whose binding partners we now identify as Ptiwi10 and Ptiwi11. scnRNAs derive from the maternal genome and initiate DNA elimination during development, whereas iesRNAs continue DNA targeting until the removal process is complete. Here, we show that scnRNAs and iesRNAs are processed by distinct Dicer-like proteins and bind Piwi proteins in a mutually exclusive manner, suggesting separate biogenesis pathways. We also demonstrate that the PTIWI10 gene is transcribed from the developing nucleus and that its transcription depends on prior DNA excision, suggesting a mechanism of gene expression control triggered by the removal of short DNA segments interrupting the gene.

  16. The BDNF/TrkB signaling pathway is involved in heat hyperalgesia mediated by Cdk5 in rats.

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    Hong-Hai Zhang

    Full Text Available Cyclin-dependent kinase 5 (Cdk5 has been shown to play an important role in mediating inflammation-induced heat hyperalgesia. However, the underlying mechanism remains unclear. The aim of this study was to determine whether roscovitine, an inhibitor of Cdk5, could reverse the heat hyperalgesia induced by peripheral injection of complete Freund's adjuvant (CFA via the brain-derived neurotrophic factor (BDNF-tyrosine kinase B (TrkB signaling pathway in the dorsal horn of the spinal cord in rats.Heat hyperalgesia induced by peripheral injection of CFA was significantly reversed by roscovitine, TrkB-IgG, and the TrkB inhibitor K252a, respectively. Furthermore, BDNF was significantly increased from 0.5 h to 24 h after CFA injection in the spinal cord dorsal horn. Intrathecal adminstration of the Cdk5 inhibitor roscovitine had no obvious effects on BDNF levels. Increased TrkB protein level was significantly reversed by roscovitine between 0.5 h and 6 h after CFA injection. Cdk5 and TrkB co-immunoprecipitation results suggested Cdk5 mediates the heat hyperalgesia induced by CFA injection by binding with TrkB, and the binding between Cdk5 and TrkB was markedly blocked by intrathecal adminstration of roscovitine.Our data suggested that the BDNF-TrkB signaling pathway was involved in CFA-induced heat hyperalgesia mediated by Cdk5. Roscovitine reversed the heat hyperalgesia induced by peripheral injection of CFA by blocking BDNF/TrkB signaling pathway, suggesting that severing the close crosstalk between Cdk5 and the BDNF/TrkB signaling cascade may present a potential target for anti-inflammatory pain.

  17. Exploration of molecular pathways mediating electric field-directed Schwann cell migration by RNA-Seq

    Science.gov (United States)

    Yao, Li; Li, Yongchao; Knapp, Jennifer; Smith, Peter

    2015-01-01

    In peripheral nervous systems, Schwann cells wrap around axons of motor and sensory neurons to form the myelin sheath. Following spinal cord injury, Schwann cells regenerate and migrate to the lesion and are involved in the spinal cord regeneration process. Transplantation of Schwann cells into injured neural tissue results in enhanced spinal axonal regeneration. Effective directional migration of Schwann cells is critical in the neural regeneration process. In this study, we report that Schwann cells migrate anodally in an applied electric field (EF). The directedness and displacement of anodal migration increased significantly when the strength of the EF increased from 50 mV/mm to 200 mV/mm. The EF did not significantly affect the cell migration speed. To explore the genes and signaling pathways that regulate cell migration in EFs, we performed a comparative analysis of differential gene expression between cells stimulated with an EF (100 mV/mm) and those without using next-generation RNA sequencing, verified by RT-qPCR. Based on the cut-off criteria (FC > 1.2, q cells versus EF-stimulated cells. A Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway analysis found that compared to the control group, 21 pathways are down-regulated, while 10 pathways are up-regulated. Differentially expressed genes participate in multiple cellular signaling pathways involved in the regulation of cell migration, including pathways of regulation of actin cytoskeleton, focal adhesion, and PI3K-Akt. PMID:25557037

  18. B7h-expressing dendritic cells and plasma B cells mediate distinct outcomes of ICOS costimulation in T cell-dependent antibody responses

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    Larimore Kevin

    2012-06-01

    Full Text Available Abstract Background The ICOS-B7h costimulatory receptor-ligand pair is required for germinal center formation, the production of isotype-switched antibodies, and antibody affinity maturation in response to T cell-dependent antigens. However, the potentially distinct roles of regulated B7h expression on B cells and dendritic cells in T cell-dependent antibody responses have not been defined. Results We generated transgenic mice with lineage-restricted B7h expression to assess the cell-type specific roles of B7h expression on B cells and dendritic cells in regulating T cell-dependent antibody responses. Our results show that endogenous B7h expression is reduced on B cells after activation in vitro and is also reduced in vivo on antibody-secreting plasma B cells in comparison to both naïve and germinal center B cells from which they are derived. Increasing the level of B7h expression on activated and plasma B cells in B-B7hTg mice led to an increase in the number of antibody-secreting plasma cells generated after immunization and a corresponding increase in the concentration of antigen-specific high affinity serum IgG antibodies of all isotypes, without affecting the number of responding germinal center B cells. In contrast, ICOS costimulation mediated by dendritic cells in DC-B7hTg mice contributed to germinal center formation and selectively increased IgG2a production without affecting the overall magnitude of antibody responses. Conclusions Using transgenic mice with lineage-restricted B7h expression, we have revealed distinct roles of ICOS costimulation mediated by dendritic cells and B cells in the regulation of T cell-dependent antibody responses.

  19. Astrocytes protect neurons against methylmercury via ATP/P2Y(1) receptor-mediated pathways in astrocytes.

    Science.gov (United States)

    Noguchi, Yusuke; Shinozaki, Youichi; Fujishita, Kayoko; Shibata, Keisuke; Imura, Yoshio; Morizawa, Yosuke; Gachet, Christian; Koizumi, Schuichi

    2013-01-01

    Methylmercury (MeHg) is a well known environmental pollutant that induces serious neuronal damage. Although MeHg readily crosses the blood-brain barrier, and should affect both neurons and glial cells, how it affects glia or neuron-to-glia interactions has received only limited attention. Here, we report that MeHg triggers ATP/P2Y1 receptor signals in astrocytes, thereby protecting neurons against MeHg via interleukin-6 (IL-6)-mediated pathways. MeHg increased several mRNAs in astrocytes, among which IL-6 was the highest. For this, ATP/P2Y1 receptor-mediated mechanisms were required because the IL-6 production was (i) inhibited by a P2Y1 receptor antagonist, MRS2179, (ii) abolished in astrocytes obtained from P2Y1 receptor-knockout mice, and (iii) mimicked by exogenously applied ATP. In addition, (iv) MeHg released ATP by exocytosis from astrocytes. As for the intracellular mechanisms responsible for IL-6 production, p38 MAP kinase was involved. MeHg-treated astrocyte-conditioned medium (ACM) showed neuro-protective effects against MeHg, which was blocked by anti-IL-6 antibody and was mimicked by the application of recombinant IL-6. As for the mechanism of neuro-protection by IL-6, an adenosine A1 receptor-mediated pathway in neurons seems to be involved. Taken together, when astrocytes sense MeHg, they release ATP that autostimulates P2Y1 receptors to upregulate IL-6, thereby leading to A1 receptor-mediated neuro-protection against MeHg.

  20. A pivotal role of the jasmonic acid signal pathway in mediating radiation-induced bystander effects in Arabidopsis thaliana.

    Science.gov (United States)

    Wang, Ting; Xu, Wei; Deng, Chenguang; Xu, Shaoxin; Li, Fanghua; Wu, Yuejin; Wu, Lijun; Bian, Po

    Although radiation-induced bystander effects (RIBE) in Arabidopsis thaliana have been well demonstrated in vivo, little is known about their underlying mechanisms, particularly with regard to the participating signaling molecules and signaling pathways. In higher plants, jasmonic acid (JA) and its bioactive derivatives are well accepted as systemic signal transducers that are produced in response to various environmental stresses. It is therefore speculated that the JA signal pathway might play a potential role in mediating radiation-induced bystander signaling of root-to-shoot. In the present study, pretreatment of seedlings with Salicylhydroxamic acid, an inhibitor of lipoxigenase (LOX) in JA biosynthesis, significantly suppressed RIBE-mediated expression of the AtRAD54 gene. After root irradiation, the aerial parts of A. thaliana mutants deficient in JA biosynthesis (aos) and signaling cascades (jar1-1) showed suppressed induction of the AtRAD54 and AtRAD51 genes and TSI and 180-bp repeats, which have been extensively used as endpoints of bystander genetic and epigenetic effects in plants. These results suggest an involvement of the JA signal pathway in the RIBE of plants. Using the root micro-grafting technique, the JA signal pathway was shown to participate in both the generation of bystander signals in irradiated root cells and radiation responses in the bystander aerial parts of plants. The over-accumulation of endogenous JA in mutant fatty acid oxygenation up-regulated 2 (fou2), in which mutation of the Two Pore Channel 1 (TPC1) gene up-regulates expression of the LOX and allene oxide synthase (AOS) genes, inhibited RIBE-mediated expression of the AtRAD54 gene, but up-regulated expression of the AtKU70 and AtLIG4 genes in the non-homologous end joining (NHEJ) pathway. Considering that NHEJ is employed by plants with increased DNA damage, the switch from HR to NHEJ suggests that over-accumulation of endogenous JA might enhance the radiosensitivity of plants

  1. Tissue factor pathway inhibitor for prediction of placenta-mediated adverse pregnancy outcomes in high-risk women: AngioPred study.

    Directory of Open Access Journals (Sweden)

    Aurélie Di Bartolomeo

    Full Text Available The study aimed to evaluate if the rate of tissue factor pathway inhibitor during pregnancy and following delivery could be a predictive factor for placenta-mediated adverse pregnancy outcomes in high-risk women.This was a prospective multicentre cohort study of 200 patients at a high risk of occurrence or recurrence of placenta-mediated adverse pregnancy outcomes conducted between June 2008 and October 2010. Measurements of tissue factor pathway inhibitor resistance (normalized ratio and tissue factor pathway inhibitor activity were performed for the last 72 patients at 20, 24, 28, 32, and 36 weeks of gestation and during the postpartum period.Overall, 15 patients presented a placenta-mediated adverse pregnancy outcome. There was no difference in normalized tissue factor pathway inhibitor ratios between patients with and without placenta-mediated adverse pregnancy outcomes during pregnancy and in the post-partum period. Patients with placenta-mediated adverse pregnancy outcomes had tissue factor pathway inhibitor activity rates that were significantly higher than those in patients without at as early as 24 weeks of gestation. The same results were observed following delivery.Among high-risk women, the tissue factor pathway inhibitor activity of patients with gestational vascular complications is higher than that in other patients. Hence, these markers could augment a screening strategy that includes an analysis of angiogenic factors as well as clinical and ultrasound imaging with Doppler measurement of the uterine arteries.

  2. Cre-Mediated Stress Affects Sirtuin Expression Levels, Peroxisome Biogenesis and Metabolism, Antioxidant and Proinflammatory Signaling Pathways

    Science.gov (United States)

    Xiao, Yu; Karnati, Srikanth; Qian, Guofeng; Nenicu, Anca; Fan, Wei; Tchatalbachev, Svetlin; Höland, Anita; Hossain, Hamid; Guillou, Florian; Lüers, Georg H.; Baumgart-Vogt, Eveline

    2012-01-01

    Cre-mediated excision of loxP sites is widely used in mice to manipulate gene function in a tissue-specific manner. To analyze phenotypic alterations related to Cre-expression, we have used AMH-Cre-transgenic mice as a model system. Different Cre expression levels were obtained by investigation of C57BL/6J wild type as well as heterozygous and homozygous AMH-Cre-mice. Our results indicate that Cre-expression itself in Sertoli cells already has led to oxidative stress and lipid peroxidation (4-HNE lysine adducts), inducing PPARα/γ, peroxisome proliferation and alterations of peroxisome biogenesis (PEX5, PEX13 and PEX14) as well as metabolic proteins (ABCD1, ABCD3, MFP1, thiolase B, catalase). In addition to the strong catalase increase, a NRF2- and FOXO3-mediated antioxidative response (HMOX1 of the endoplasmic reticulum and mitochondrial SOD2) and a NF-κB activation were noted. TGFβ1 and proinflammatory cytokines like IL1, IL6 and TNFα were upregulated and stress-related signaling pathways were induced. Sertoli cell mRNA-microarray analysis revealed an increase of TNFR2-signaling components. 53BP1 recruitment and expression levels for DNA repair genes as well as for p53 were elevated and the ones for related sirtuin deacetylases affected (SIRT 1, 3-7) in Sertoli cells. Under chronic Cre-mediated DNA damage conditions a strong downregulation of Sirt1 was observed, suggesting that the decrease of this important coordinator between DNA repair and metabolic signaling might induce the repression release of major transcription factors regulating metabolic and cytokine-mediated stress pathways. Indeed, caspase-3 was activated and increased germ cell apoptosis was observed, suggesting paracrine effects. In conclusion, the observed wide stress-induced effects and metabolic alterations suggest that it is essential to use the correct control animals (Cre/Wt) with matched Cre expression levels to differentiate between Cre-mediated and specific gene-knock out-mediated

  3. Cre-mediated stress affects sirtuin expression levels, peroxisome biogenesis and metabolism, antioxidant and proinflammatory signaling pathways.

    Directory of Open Access Journals (Sweden)

    Yu Xiao

    Full Text Available Cre-mediated excision of loxP sites is widely used in mice to manipulate gene function in a tissue-specific manner. To analyze phenotypic alterations related to Cre-expression, we have used AMH-Cre-transgenic mice as a model system. Different Cre expression levels were obtained by investigation of C57BL/6J wild type as well as heterozygous and homozygous AMH-Cre-mice. Our results indicate that Cre-expression itself in Sertoli cells already has led to oxidative stress and lipid peroxidation (4-HNE lysine adducts, inducing PPARα/γ, peroxisome proliferation and alterations of peroxisome biogenesis (PEX5, PEX13 and PEX14 as well as metabolic proteins (ABCD1, ABCD3, MFP1, thiolase B, catalase. In addition to the strong catalase increase, a NRF2- and FOXO3-mediated antioxidative response (HMOX1 of the endoplasmic reticulum and mitochondrial SOD2 and a NF-κB activation were noted. TGFβ1 and proinflammatory cytokines like IL1, IL6 and TNFα were upregulated and stress-related signaling pathways were induced. Sertoli cell mRNA-microarray analysis revealed an increase of TNFR2-signaling components. 53BP1 recruitment and expression levels for DNA repair genes as well as for p53 were elevated and the ones for related sirtuin deacetylases affected (SIRT 1, 3-7 in Sertoli cells. Under chronic Cre-mediated DNA damage conditions a strong downregulation of Sirt1 was observed, suggesting that the decrease of this important coordinator between DNA repair and metabolic signaling might induce the repression release of major transcription factors regulating metabolic and cytokine-mediated stress pathways. Indeed, caspase-3 was activated and increased germ cell apoptosis was observed, suggesting paracrine effects. In conclusion, the observed wide stress-induced effects and metabolic alterations suggest that it is essential to use the correct control animals (Cre/Wt with matched Cre expression levels to differentiate between Cre-mediated and specific gene-knock out-mediated

  4. Demodex canis regulates cholinergic system mediated immunosuppressive pathways in canine demodicosis.

    Science.gov (United States)

    Kumari, P; Nigam, R; Singh, A; Nakade, U P; Sharma, A; Garg, S K; Singh, S K

    2017-09-01

    Demodex canis infestation in dogs remains one of the main challenges in veterinary dermatology. The exact pathogenesis of canine demodicosis is unknown but an aberration in immune status is considered very significant. No studies have underpinned the nexus between induction of demodicosis and neural immunosuppressive pathways so far. We have evaluated the involvement of cholinergic pathways in association with cytokines regulation as an insight into the immuno-pathogenesis of canine demodicosis in the present study. Remarkable elevations in circulatory immunosuppressive cytokine interleukin-10 and cholinesterase activity were observed in dogs with demodicosis. Simultaneously, remarkable reduction in circulatory pro-inflammatory cytokine tumour necrosis factor-alpha level was observed in dogs with demodicosis. Findings of the present study evidently suggest that Demodex mites might be affecting the cholinergic pathways to induce immunosuppression in their host and then proliferate incessantly in skin microenvironment to cause demodicosis.

  5. The nitric oxide-sensitive p21Ras-ERK pathway mediates S-nitrosoglutathione-induced apoptosis

    International Nuclear Information System (INIS)

    Tsujita, Maristela; Batista, Wagner L.; Ogata, Fernando T.; Stern, Arnold; Monteiro, Hugo P.; Arai, Roberto J.

    2008-01-01

    p21Ras protein plays a critical role in cellular signaling that induces either cell cycle progression or apoptosis. Nitric oxide (NO) has been consistently reported to activate p21Ras through the redox sensitive cysteine residue (118). In this study, we demonstrated that the p21Ras-ERK pathway regulates THP-1 monocyte/macrophage apoptosis induced by S-nitrosoglutathione (SNOG). This was apparent from studies in THP-1 cells expressing NO-insensitive p21Ras (p21Ras C118S ) where the pro-apoptotic action of SNOG was almost abrogated. Three major MAP kinase pathways (ERK, JNK, and p38) that are downstream to p21Ras were investigated. It was observed that only the activation of ERK1/2 MAP kinases by SNOG in THP-1 cells was attributable to p21Ras. The inhibition of the ERK pathway by PD98059 markedly attenuated apoptosis in SNOG-treated THP-1 cells, but had a marginal effect on SNOG-treated THP-1 cells expressing NO-insensitive p21Ras. The inhibition of the JNK and p38 pathways by selective inhibitors had no marked effects on the percentage of apoptosis. The induction of p21Waf1 expression by SNOG was observed in THP-1 cells harboring mutant and wild-type p21Ras, however in cells expressing mutant Ras, the expression of p21Waf1 was significantly attenuated. The treatment of THP-1 cells expressing wild-type p21Ras with PD98059 resulted in significant attenuation of p21Waf1 expression. These results indicate that the redox sensitive p21Ras-ERK pathway plays a critical role in sensing and delivering the pro-apoptotic signaling mediated by SNOG

  6. The effect of tributyltin chloride on Caenorhabditis elegans germline is mediated by a conserved DNA damage checkpoint pathway.

    Science.gov (United States)

    Cheng, Zhe; Tian, Huimin; Chu, Hongran; Wu, Jianjian; Li, Yingying; Wang, Yanhai

    2014-03-21

    Tributyltin (TBT), one of the environmental pollutants, has been shown to impact the reproduction of animals. However, due to the lack of appropriate animal model, analysis of the affected molecular pathways in germ cells is lagging and has been particularly challenging. In the present study, we investigated the effects of tributyltin chloride (TBTCL) on the nematode Caenorhabditis elegans germline. We show that exposure of C. elegans to TBTCL causes significantly elevated level of sterility and embryonic lethality. TBTCL exposure results in an increased number of meiotic DNA double-strand breaks in germ cells, subsequently leading to activated DNA damage checkpoint. Exposing C. elegans to TBTCL causes dose- and time-dependent germline apoptosis. This apoptotic response was blocked in loss-of-function mutants of hus-1 (op241), mrt-2 (e2663) and p53/cep-1 (gk138), indicating that checkpoints and p53 are essential for mediating TBTCL-induced germ cell apoptosis. Moreover, TBTCL exposure can inhibit germ cell proliferation, which is also mediated by the conserved checkpoint pathway. We thereby propose that TBT exhibits its effects on the germline by inducing DNA damage and impaired maintenance of genomic integrity. Copyright © 2014 The Authors. Published by Elsevier Ireland Ltd.. All rights reserved.

  7. Can the TLR-4-Mediated Signaling Pathway Be “A Key Inflammatory Promoter for Sporadic TAA”?

    Directory of Open Access Journals (Sweden)

    Giovanni Ruvolo

    2014-01-01

    Full Text Available Thoracic aorta shows with advancing age various changes and a progressive deterioration in structure and function. As a result, vascular remodeling (VR and medial degeneration (MD occur as pathological entities responsible principally for the sporadic TAA onset. Little is known about their genetic, molecular, and cellular mechanisms. Recent evidence is proposing the strong role of a chronic immune/inflammatory process in their evocation and progression. Thus, we evaluated the potential role of Toll like receptor- (TLR- 4-mediated signaling pathway and its polymorphisms in sporadic TAA. Genetic, immunohistochemical, and biochemical analyses were assessed. Interestingly, the rs4986790 TLR4 polymorphism confers a higher susceptibility for sporadic TAA (OR=14.4, P=0.0008 and it represents, together with rs1799752 ACE, rs3918242 MMP-9, and rs2285053 MMP-2 SNPs, an independent sporadic TAA risk factor. In consistency with these data, a significant association was observed between their combined risk genotype and sporadic TAA. Cases bearing this risk genotype showed higher systemic inflammatory mediator levels, significant inflammatory/immune infiltrate, a typical MD phenotype, lower telomere length, and positive correlations with histopatological abnormalities, hypertension, smoking, and ageing. Thus, TLR4 pathway should seem to have a key role in sporadic TAA. It might represent a potential useful tool for preventing and monitoring sporadic TAA and developing personalized treatments.

  8. Regulation of the Nampt-mediated NAD salvage pathway and its therapeutic implications in pancreatic cancer.

    Science.gov (United States)

    Ju, Huai-Qiang; Zhuang, Zhuo-Nan; Li, Hao; Tian, Tian; Lu, Yun-Xin; Fan, Xiao-Qiang; Zhou, Hai-Jun; Mo, Hai-Yu; Sheng, Hui; Chiao, Paul J; Xu, Rui-Hua

    2016-08-28

    Nicotinamide adenine dinucleotide (NAD) is a crucial cofactor for the redox reactions in the metabolic pathways of cancer cells that have elevated aerobic glycolysis (Warburg effect). Cancer cells are reported to rely on NAD recycling and inhibition of the NAD salvage pathway causes metabolic collapse and cell death. However, the underlying regulatory mechanisms and clinical implications for the NAD salvage pathway in pancreatic ductal adenocarcinoma (PDAC) remain unclear. This study showed that the expression of Nampt, the rate-limiting enzyme of the NAD salvage pathway, was significantly increased in PDAC cells and PDAC tissues. Additionally, inhibition of Nampt impaired tumor growth in vitro and tumorigenesis in vivo, which was accompanied by a decreased cellular NAD level and glycolytic activity. Mechanistically, the Nampt expression was independent of Kras and p16 status, but it was directly regulated by miR-206, which was inversely correlated with the expression of Nampt in PDAC tissues. Importantly, pharmacological inhibition of Nampt by its inhibitor, FK866, significantly enhanced the antitumor activity of gemcitabine in PDAC cells and in orthotopic xenograft mouse models. In conclusion, the present study revealed a novel regulatory mechanism for Nampt in PDAC and suggested that Nampt inhibition may override gemcitabine resistance by decreasing the NAD level and suppressing glycolytic activity, warranting further clinical investigation for pancreatic cancer treatment. Copyright © 2016 Elsevier Ireland Ltd. All rights reserved.

  9. Delineating the maladaptive pathways of child maltreatment: a mediated moderation analysis of the roles of self-perception and social support.

    Science.gov (United States)

    Appleyard, Karen; Yang, Chongming; Runyan, Desmond K

    2010-05-01

    The current study investigated concurrent and longitudinal mediated and mediated moderation pathways among maltreatment, self-perception (i.e., loneliness and self-esteem), social support, and internalizing and externalizing behavior problems. For both genders, early childhood maltreatment (i.e., ages 0-6) was related directly to internalizing and externalizing behavior problems at age 6, and later maltreatment (i.e., ages 6-8) was directly related to internalizing and externalizing behavior problems at age 8. Results of concurrent mediation and mediated moderation indicated that early maltreatment was significantly related to internalizing and externalizing behavior problems at age 6 indirectly both through age 6 loneliness and self-esteem for boys and through age 6 loneliness for girls. Significant moderation of the pathway from early maltreatment to self-esteem, and for boys, significant mediated moderation to emotional and behavioral problems were found, such that the mediated effect through self-esteem varied across levels of social support, though in an unexpected direction. No significant longitudinal mediation or mediated moderation was found, however, between the age 6 mediators and moderator and internalizing or externalizing problems at age 8. The roles of the hypothesized mediating and moderating mechanisms are discussed, with implications for designing intervention and prevention programs.

  10. Gendered Pathways? Gender, Mediating Factors, and the Gap in Boys' and Girls' Substance Use

    Science.gov (United States)

    Whaley, Rachel Bridges; Hayes-Smith, Justin; Hayes-Smith, Rebecca

    2013-01-01

    A gender gap in alcohol and drug use exists but is somewhat smaller than the gender gap in other forms of delinquency. This article extends studies that examine the gender-delinquency relationship to substance use in particular and estimate the extent to which major risk and protective factors mediate the association between gender and alcohol and…

  11. Characterization of murine melanocortin receptors mediating adipocyte lipolysis and examination of signalling pathways involved

    DEFF Research Database (Denmark)

    Møller, Cathrine Laustrup; Raun, Kirsten; Jacobsen, Marianne Lambert

    2011-01-01

    hormone (a-MSH) generated from proopiomelanocortin (POMC), as well as synthetic MSH analogues to stimulate lipolysis in murine 3T3-L1 adipocytes it is shown that MC2R and MC5R are lipolytic mediators in differentiated 3T3-L1 adipocytes. Involvement of cAMP, phosphorylated extracellular signal...

  12. Antigen presentation by resting B cells. Radiosensitivity of the antigen-presentation function and two distinct pathways of T cell activation

    International Nuclear Information System (INIS)

    Ashwell, J.D.; DeFranco, A.L.; Paul, W.E.; Schwartz, R.H.

    1984-01-01

    In this report we have examined the ability of small resting B cells to act as antigen-presenting cells (APC) to antigen-specific MHC-restricted T cells as assessed by either T cell proliferation or T cell-dependent B cell stimulation. We found that 10 of 14 in vitro antigen-specific MHC-restricted T cell clones and lines and three of four T cell hybridomas could be induced to either proliferate or secrete IL-2 in the presence of lightly irradiated (1,000 rads) purified B cells and the appropriate foreign antigen. All T cell lines and hybridomas were stimulated to proliferate or make IL-2 by macrophage- and dendritic cell-enriched populations and all T cells tested except one hybridoma caused B cell activation when stimulated with B cells as APC. Furthermore, lightly irradiated, highly purified syngeneic B cells were as potent a source of APC for inducing B cell activation as were low density dendritic and macrophage-enriched cells. Lymph node T cells freshly taken from antigen-primed animals were also found to proliferate when cultured with purified B cells and the appropriate antigen. This APC function was easily measured when the cells were irradiated with 1,000 rads, but was greatly diminished or absent when they were irradiated with 3,300 rads. In addition, this radiosensitivity allowed us to easily distinguish B cell antigen presentation from presentation by the dendritic cell and macrophage, as the latter was resistant to 3,300 rads. Finally, one T cell clone that failed to proliferate when B cells were used as APC was able to recruit allogeneic B cells to proliferate in the presence of syngeneic B cells and the appropriate antigen. This result suggests that there are at least two distinct pathways of activation in T cells, one that leads to T cell proliferation and one that leads to the secretion of B cell recruitment factor(s)

  13. Interleukin-17A promotes MUC5AC expression and goblet cell hyperplasia in nasal polyps via the Act1-mediated pathway.

    Directory of Open Access Journals (Sweden)

    Wentong Xia

    Full Text Available BACKGROUND: Recent studies demonstrated that nasal polyps (NP patients in China and other Asian regions possessed distinct Th17-dominant inflammation and enhanced tissue remodeling. However, the mechanism underlying these observations is not fully understood. This study sought to evaluate the association of interleukin (IL-17A with MUC5AC expression and goblet cell hyperplasia in Chinese NP patients and to characterize the signaling pathway underlying IL-17A-induced MUC5AC expression in vitro. METHOD: We enrolled 25 NP patients and 22 normal controls and examined the expression of IL-17A, MUC5AC and act1 in polyp tissues by immunohistochemical (IHC staining, quantitative polymerase chain reaction (qPCR and western blot. Moreover, by using an in vitro culture system of polyp epithelial cells (PECs, IL-17A-induced gene expression was screened in cultured PECs by DNA microarray. The expression of IL-17RA, IL-17RC, act1 and MUC5AC and the activation of the MAPK pathway (ERK, p38 and JNK, were further examined in cultured PECs and NCI-H292 cells by qPCR and western blotting, respectively. RESULTS: We found that increased IL-17A production was significantly correlated with MUC5AC and act1 expression and goblet cell hyperplasia in polyp tissues (p<0.05. IL-17A significantly stimulated the expression of IL-17RA, IL-17RC, act1 and MUC5AC, and the activation of the MAPK pathway in cultured PECs and NCI-H292 cells (p<0.05. In addition, IL-17RA, IL-17RC and act1 siRNA significantly blocked IL-17A-induced MUC5AC production in vitro (p<0.05. CONCLUSION: Our results suggest that IL-17A plays a crucial role in stimulating the production of MUC5AC and goblet cell hyperplasia through the act1-mediated signaling pathway and may suggest a promising strategy for the management of Th17-dominant NP patients.

  14. Pathway from Delirium to Death: Potential In-Hospital Mediators of Excess Mortality.

    Science.gov (United States)

    Dharmarajan, Kumar; Swami, Sunil; Gou, Ray Y; Jones, Richard N; Inouye, Sharon K

    2017-05-01

    (1) To determine the relationship of incident delirium during hospitalization with 90-day mortality; (2) to identify potential in-hospital mediators through which delirium increases 90-day mortality. Analysis of data from Project Recovery, a controlled clinical trial of a delirium prevention intervention from 1995 to 1998 with follow-up through 2000. Large academic hospital. Patients ≥70 years old without delirium at hospital admission who were at intermediate-to-high risk of developing delirium and received usual care only. (1) Incident delirium; (2) potential mediators of delirium on death including use of restraining devices (physical restraints, urinary catheters), development of hospital acquired conditions (HACs) (falls, pressure ulcers), and exposure to other noxious insults (sleep deprivation, acute malnutrition, dehydration, aspiration pneumonia); (3) death within 90 days of admission. Among 469 patients, 70 (15%) developed incident delirium. These patients were more likely to experience restraining devices (37% vs 16%, P delirium was 4.2 (95% CI = 2.8-6.3) in bivariable analyses, increased in a graded manner with additional exposures to restraining devices, HACs, and other noxious insults, and declined by 10.9% after addition of these potential mediator categories, providing evidence of mediation. Restraining devices, HACs, and additional noxious insults were more frequent among patients with delirium, increased mortality in a graded manner, and were responsible for a significant percentage of the association of delirium with death. Additional efforts to prevent potential downstream mediators through which delirium increases mortality may help to improve outcomes among hospitalized older adults. © 2016, Copyright the Authors Journal compilation © 2016, The American Geriatrics Society.

  15. Hepatitis C virus infection induces apoptosis through a Bax-triggered, mitochondrion-mediated, caspase 3-dependent pathway.

    Science.gov (United States)

    Deng, Lin; Adachi, Tetsuya; Kitayama, Kikumi; Bungyoku, Yasuaki; Kitazawa, Sohei; Ishido, Satoshi; Shoji, Ikuo; Hotta, Hak

    2008-11-01

    We previously reported that cells harboring the hepatitis C virus (HCV) RNA replicon as well as those expressing HCV NS3/4A exhibited increased sensitivity to suboptimal doses of apoptotic stimuli to undergo mitochondrion-mediated apoptosis (Y. Nomura-Takigawa, et al., J. Gen. Virol. 87:1935-1945, 2006). Little is known, however, about whether or not HCV infection induces apoptosis of the virus-infected cells. In this study, by using the chimeric J6/JFH1 strain of HCV genotype 2a, we demonstrated that HCV infection induced cell death in Huh7.5 cells. The cell death was associated with activation of caspase 3, nuclear translocation of activated caspase 3, and cleavage of DNA repair enzyme poly(ADP-ribose) polymerase, which is known to be an important substrate for activated caspase 3. These results suggest that HCV-induced cell death is, in fact, apoptosis. Moreover, HCV infection activated Bax, a proapoptotic member of the Bcl-2 family, as revealed by its conformational change and its increased accumulation on mitochondrial membranes. Concomitantly, HCV infection induced disruption of mitochondrial transmembrane potential, followed by mitochondrial swelling and release of cytochrome c from mitochondria. HCV infection also caused oxidative stress via increased production of mitochondrial superoxide. On the other hand, HCV infection did not mediate increased expression of glucose-regulated protein 78 (GRP78) or GRP94, which are known as endoplasmic reticulum (ER) stress-induced proteins; this result suggests that ER stress is not primarily involved in HCV-induced apoptosis in our experimental system. Taken together, our present results suggest that HCV infection induces apoptosis of the host cell through a Bax-triggered, mitochondrion-mediated, caspase 3-dependent pathway(s).

  16. TRAF1 Coordinates Polyubiquitin Signaling to Enhance Epstein-Barr Virus LMP1-Mediated Growth and Survival Pathway Activation.

    Directory of Open Access Journals (Sweden)

    Hannah Greenfeld

    2015-05-01

    Full Text Available The Epstein-Barr virus (EBV encoded oncoprotein Latent Membrane Protein 1 (LMP1 signals through two C-terminal tail domains to drive cell growth, survival and transformation. The LMP1 membrane-proximal TES1/CTAR1 domain recruits TRAFs to activate MAP kinase, non-canonical and canonical NF-kB pathways, and is critical for EBV-mediated B-cell transformation. TRAF1 is amongst the most highly TES1-induced target genes and is abundantly expressed in EBV-associated lymphoproliferative disorders. We found that TRAF1 expression enhanced LMP1 TES1 domain-mediated activation of the p38, JNK, ERK and canonical NF-kB pathways, but not non-canonical NF-kB pathway activity. To gain insights into how TRAF1 amplifies LMP1 TES1 MAP kinase and canonical NF-kB pathways, we performed proteomic analysis of TRAF1 complexes immuno-purified from cells uninduced or induced for LMP1 TES1 signaling. Unexpectedly, we found that LMP1 TES1 domain signaling induced an association between TRAF1 and the linear ubiquitin chain assembly complex (LUBAC, and stimulated linear (M1-linked polyubiquitin chain attachment to TRAF1 complexes. LMP1 or TRAF1 complexes isolated from EBV-transformed lymphoblastoid B cell lines (LCLs were highly modified by M1-linked polyubiqutin chains. The M1-ubiquitin binding proteins IKK-gamma/NEMO, A20 and ABIN1 each associate with TRAF1 in cells that express LMP1. TRAF2, but not the cIAP1 or cIAP2 ubiquitin ligases, plays a key role in LUBAC recruitment and M1-chain attachment to TRAF1 complexes, implicating the TRAF1:TRAF2 heterotrimer in LMP1 TES1-dependent LUBAC activation. Depletion of either TRAF1, or the LUBAC ubiquitin E3 ligase subunit HOIP, markedly impaired LCL growth. Likewise, LMP1 or TRAF1 complexes purified from LCLs were decorated by lysine 63 (K63-linked polyubiqutin chains. LMP1 TES1 signaling induced K63-polyubiquitin chain attachment to TRAF1 complexes, and TRAF2 was identified as K63-Ub chain target. Co-localization of M1- and K63

  17. Role of SIRT1-mediated mitochondrial and Akt pathways in glioblastoma cell death induced by Cotinus coggygria flavonoid nanoliposomes

    Directory of Open Access Journals (Sweden)

    Wang G

    2015-08-01

    phosphorylated p53. Together, these results indicated SIRT1/p53-mediated cell death was induced by CCF-NLs, but not by extracellular signal-regulated kinase, in DBTRG-05MG cells. Overall, this study suggested caspase-dependent activation of both the intrinsic and extrinsic signaling pathways, probably through blockade of the SIRT1/p53-mediated mitochondrial and Akt pathways to exert the proapoptotic effect of CCF-NLs in DBTRG-05MG GBM cells. Keywords: Cotinus coggygria flavonoid nanoliposomes, cell death, SIRT1, mitochondrial, PI3K/Akt pathway

  18. Nanocurcumin-Mediated Down-Regulation of Telomerase Via Stimulating TGFβ1 Signaling Pathway in Hepatocellular Carcinoma Cells

    Science.gov (United States)

    Shariati, Molood; Hajigholami, Samira; Veisi Malekshahi, Ziba; Entezari, Maliheh; Bodaghabadi, Narges; Sadeghizadeh, Majid

    2017-10-10

    Curcumin, extracted from turmeric, represents enormous potential to serve as an anticancer agent. Telomerase is viewed as a prominent molecular target of curcumin, and Transforming growth factor-β1 (TGFβ1) has proven to be a major inhibitory signaling pathway for telomerase activity. In the current study, we aimed to explore suppressive effects of nanocurcumin on telomerase expression through TGFβ1 pathway in a hepatocellular carcinoma cell line (Huh7). MTT assay was used to determine the effect of nonocurcumin on viability of Huh7 cells. RT-PCR was used to analyze the gene expression patterns. MTT assay revealed that nanocurcumin acts in a dose- and time-dependent manner to diminish the cell viability. RT-PCR analysis indicated that nanocurcumin results in augmentation of TGFβ1 72 hours post treatment and leads to the reduction of telomerase expression 48 and 72 hours post exposure. Also, up-regulation of Smad3 and E2F1 and down-regulation of Smad7 confirmed the effect of nanocurcumin on intermediate components of TGFβ1 pathway. Furthermore, transfection of the proximal promoter of telomerase triggered a significant reduction in luciferase activity. The data from the present study lead us to develop a deeper understanding of the mechanisms underlying nanocurcumin-mediated regulation of telomerase expression, thereby presenting a new perspective to the landscape of using nanocurcumin as a cancer-oriented therapeutic agent.

  19. Death receptor pathways mediate targeted and non-targeted effects of ionizing radiations in breast cancer cells

    International Nuclear Information System (INIS)

    Luce, A.; Courtin, A.; Levalois, C.; Altmeyer-Morel, S.; Chevillard, S.; Lebeau, J.; Romeo, P.H.

    2009-01-01

    Delayed cell death by mitotic catastrophe is a frequent mode of solid tumor cell death after γ-irradiation, a widely used treatment of cancer. Whereas the mechanisms that underlie the early γ-irradiation-induced cell death are well documented, those that drive the delayed cell death are largely unknown. Here we show that the Fas, tumor necrosis factor-related apoptosis-inducing ligand (TRAIL) and tumor necrosis factor (TNF)-α death receptor pathways mediate the delayed cell death observed after γ-irradiation of breast cancer cells. Early after irradiation, we observe the increased expression of Fas, TRAIL-R and TNF-R that first sensitizes cells to apoptosis. Later, the increased expression of FasL, TRAIL and TNF-α permit the apoptosis engagement linked to mitotic catastrophe. Treatments with TNF-α, TRAIL or anti-Fas antibody, early after radiation exposure, induce apoptosis, whereas the neutralization of the three death receptors pathways impairs the delayed cell death. We also show for the first time that irradiated breast cancer cells excrete soluble forms of the three ligands that can induce the death of sensitive bystander cells. Overall, these results define the molecular basis of the delayed cell death of irradiated cancer cells and identify the death receptors pathways as crucial actors in apoptosis induced by targeted as well as non-targeted effects of ionizing radiation. (authors)

  20. The phosphatidylinositol-3 kinase pathway is not essential for insulin-like growth factor I receptor-mediated clonogenic radioresistance

    International Nuclear Information System (INIS)

    Yu, Dong; Watanabe, Hiroshi; Shibuya, Hitoshi; Miura, Masahiko

    2002-01-01

    The insulin-like growth factor I receptor (IGF-IR) is known to induce clonogenic radioresistance in cells following ionizing irradiation. To explore the downstream signaling pathways, we focused on the phosphatidylinositol-3 kinase (PI3-K) pathway, which is thought to be the primary cell survival signal originating from the receptor. For this purpose, R- cells deficient in the endogenous IGF-IR were used as a recipient of the human IGF-IR with or without mutations at potential PI3-K activation sites: NPXY 950 and Y 1316 XXM. Mutats with double mutation at Y950/Y1316 exhibited not abrogated, but reduced activation of insulin receptor substance-1 (IRS-1), PI3-K, and Akt upon IGF-I stimulation. However, the mutants had the same clonogenic radioresistance as cells with wild type (WT) receptors. Neither wortmannin nor LY294002, specific inhibitors of PI3-K, affected the radioresistance of cells with WT receptors at concentrations specific for PI3-K. Collectively, these results indicate that the PI3-K pathway is not essential for IGF-IR-mediated clonogenic radioresistance. (author)

  1. Metropolitan isolation segregation and Black-White disparities in very preterm birth: a test of mediating pathways and variance explained.

    Science.gov (United States)

    Kramer, Michael R; Cooper, Hannah L; Drews-Botsch, Carolyn D; Waller, Lance A; Hogue, Carol R

    2010-12-01

    Residential isolation segregation (a measure of residential inter-racial exposure) has been associated with rates of preterm birth (births (32-36 weeks) raise questions about whether this association is similar across gestational ages, and through what pathways it might be mediated. Hierarchical Bayesian models were fit to answer three questions: is the isolation-prematurity association similar for very and moderately preterm birth; is this association mediated by maternal chronic disease, socioeconomic status, or metropolitan area crime and poverty rates; and how much of the geographic variation in Black-White very preterm birth disparities is explained by isolation segregation? Singleton births to Black and White women in 231 U.S. metropolitan statistical areas in 2000-2002 were analyzed and isolation segregation was calculated for each. We found that among Black women, isolation is associated with very preterm birth and moderately preterm birth. The association may be partially mediated by individual level socioeconomic characteristics and metropolitan level violent crime rates. There is no association between segregation and prematurity among White women. Isolation segregation explains 28% of the geographic variation in Black-White very preterm birth disparities. Our findings highlight the importance of isolation segregation for the high-burden outcome of very preterm birth, but unexplained excess risk for prematurity among Black women is substantial. Copyright © 2010 Elsevier Ltd. All rights reserved.

  2. Ethnic identity and mental health in American Indian youth: examining mediation pathways through self-esteem, and future optimism.

    Science.gov (United States)

    Smokowski, Paul R; Evans, Caroline B R; Cotter, Katie L; Webber, Kristina C

    2014-03-01

    Mental health functioning in American Indian youth is an understudied topic. Given the increased rates of depression and anxiety in this population, further research is needed. Using multiple group structural equation modeling, the current study illuminates the effect of ethnic identity on anxiety symptoms, depressive symptoms, and externalizing behavior in a group of Lumbee adolescents and a group of Caucasian, African American, and Latino/Hispanic adolescents. This study examined two possible pathways (i.e., future optimism and self-esteem) through which ethnic identity is associated with adolescent mental health. The sample (N = 4,714) is 28.53% American Indian (Lumbee) and 51.38% female. The study findings indicate that self-esteem significantly mediated the relationships between ethnic identity and anxiety symptoms, depressive symptoms, and externalizing behavior for all racial/ethnic groups (i.e., the total sample). Future optimism significantly mediated the relationship between ethnic identity and externalizing behavior for all racial/ethnic groups and was a significant mediator between ethnic identity and depressive symptoms for American Indian youth only. Fostering ethnic identity in all youth serves to enhance mental health functioning, but is especially important for American Indian youth due to the collective nature of their culture.

  3. Angiotensin-II-induced Muscle Wasting is Mediated by 25-Hydroxycholesterol via GSK3β Signaling Pathway

    Directory of Open Access Journals (Sweden)

    Congcong Shen

    2017-02-01

    Full Text Available While angiotensin II (ang II has been implicated in the pathogenesis of cardiac cachexia (CC, the molecules that mediate ang II's wasting effect have not been identified. It is known TNF-α level is increased in patients with CC, and TNF-α release is triggered by ang II. We therefore hypothesized that ang II induced muscle wasting is mediated by TNF-α. Ang II infusion led to skeletal muscle wasting in wild type (WT but not in TNF alpha type 1 receptor knockout (TNFR1KO mice, suggesting that ang II induced muscle loss is mediated by TNF-α through its type 1 receptor. Microarray analysis identified cholesterol 25-hydroxylase (Ch25h as the down stream target of TNF-α. Intraperitoneal injection of 25-hydroxycholesterol (25-OHC, the product of Ch25h, resulted in muscle loss in C57BL/6 mice, accompanied by increased expression of atrogin-1, MuRF1 and suppression of IGF-1/Akt signaling pathway. The identification of 25-OHC as an inducer of muscle wasting has implications for the development of specific treatment strategies in preventing muscle loss.

  4. A Mitochondria-Dependent Pathway Mediates the Apoptosis of GSE-Induced Yeast

    OpenAIRE

    Cao, Sishuo; Xu, Wentao; Zhang, Nan; Wang, Yan; Luo, YunBo; He, Xiaoyun; Huang, Kunlun

    2012-01-01

    Grapefruit seed extract (GSE), which has powerful anti-fungal activity, can induce apoptosis in S. cerevisiae. The yeast cells underwent apoptosis as determined by testing for apoptotic markers of DNA cleavage and typical chromatin condensation by Terminal Deoxynucleotidyl Transferase-mediated dUTP Nick End Labeling (TUNEL) and 4,6'-diaminidino-2-phenylindole (DAPI) staining and electron microscopy. The changes of ΔΨmt (mitochondrial transmembrane potential) and ROS (reactive oxygen species) ...

  5. Engineered Aedes aegypti JAK/STAT Pathway-Mediated Immunity to Dengue Virus.

    Directory of Open Access Journals (Sweden)

    Natapong Jupatanakul

    2017-01-01

    Full Text Available We have developed genetically modified Ae. aegypti mosquitoes that activate the conserved antiviral JAK/STAT pathway in the fat body tissue, by overexpressing either the receptor Dome or the Janus kinase Hop by the blood feeding-induced vitellogenin (Vg promoter. Transgene expression inhibits infection with several dengue virus (DENV serotypes in the midgut as well as systemically and in the salivary glands. The impact of the transgenes Dome and Hop on mosquito longevity was minimal, but it resulted in a compromised fecundity when compared to wild-type mosquitoes. Overexpression of Dome and Hop resulted in profound transcriptome regulation in the fat body tissue as well as the midgut tissue, pinpointing several expression signatures that reflect mechanisms of DENV restriction. Our transcriptome studies and reverse genetic analyses suggested that enrichment of DENV restriction factor and depletion of DENV host factor transcripts likely accounts for the DENV inhibition, and they allowed us to identify novel factors that modulate infection. Interestingly, the fat body-specific activation of the JAK/STAT pathway did not result in any enhanced resistance to Zika virus (ZIKV or chikungunya virus (CHIKV infection, thereby indicating a possible specialization of the pathway's antiviral role.

  6. Embryotoxicity Caused by DON-Induced Oxidative Stress Mediated by Nrf2/HO-1 Pathway

    Directory of Open Access Journals (Sweden)

    Miao Yu

    2017-06-01

    Full Text Available Deoxynivalenol (DON belongs to the type B group of trichothecenes family, which is composed of sesquiterpenoid metabolites produced by Fusarium and other fungi in grain. DON may cause various toxicities, such as cytotoxicity, immunotoxicity, genotoxicity as well as teratogenicity and carcinogenicity. In the present study, we focus on a hypothesis that DON alters the expressions of Nrf2/HO-1 pathway by inducing embryotoxicity in C57BL/6 mouse (5.0, 2.5, 1.0, and 0 mg/kg/day and BeWo cell lines (0 and 50 nM; 3 h, 12 h and 24 h. Our results indicate that DON treatment in mice during pregnancy leads to ROS accumulation in the placenta, which results in embryotoxicity. At the same time Nrf2/HO-1 pathway is up-regulated by ROS to protect placenta cells from oxidative damage. In DON-treated BeWo cells, the level of ROS has time–effect and dose–effect relationships with HO-1 expression. Moderate increase in HO-1 protects the cell from oxidative damage, while excessive increase in HO-1 aggravates the oxidative damage, which is called in some studies the “threshold effect”. Therefore, oxidative stress may be the critical molecular mechanism for DON-induced embryotoxicity. Besides, Nrf2/HO-1 pathway accompanied by the “threshold effect” also plays an important role against DON-induced oxidative damage in this process.

  7. Distinct Paths for Basic Amino Acid Export in Escherichia coli: YbjE (LysO) Mediates Export of L-Lysine.

    Science.gov (United States)

    Pathania, Amit; Sardesai, Abhijit A

    2015-06-15

    In Escherichia coli, argO encodes an exporter for L-arginine (Arg) and its toxic analogue canavanine (CAN), and its transcriptional activation and repression, by Arg and L-lysine (Lys), respectively, are mediated by the regulator ArgP. Accordingly argO and argP mutants are CAN supersensitive (CAN(ss)). We report the identification of ybjE as a gene encoding a predicted inner membrane protein that mediates export of Lys, and our results confirm the previous identification with a different approach of YbjE as a Lys exporter, reported by Ueda and coworkers (T. Ueda, Y. Nakai, Y. Gunji, R. Takikawa, and Y. Joe, U.S. patents 7,629,142 B2 [December 2009] and 8,383,363 B1 [February 2013] and European patent 1,664,318 B1 [September 2009]). ybjE was isolated as a multicopy suppressor of the CAN(ss) phenotype of a strain lacking ArgO. The absence of YbjE did not confer a CAN(ss) phenotype but instead conferred hypersensitivity to the lysine antimetabolite thialysine and led to growth inhibition by the dipeptide lysylalanine, which is associated with elevated cellular Lys content. YbjE overproduction resulted in Lys excretion and syntrophic cross-feeding of a Lys auxotroph. Constitutive overexpression of argO promoted Lys cross-feeding that is indicative of a latent Lys export potential of ArgO. Arg modestly repressed ybjE transcription in an ArgR-dependent manner, and ArgR displayed Arg-sensitive binding to the ybjE promoter region in vitro. Our studies suggest that the reciprocal repression of argO and ybjE, respectively, by Lys and Arg confers the specificity for basic amino acid export by distinct paths and that such cross-repression contributes to maintenance of cytoplasmic Arg/Lys balance. We propose that YbjE be redesignated LysO. This work ascribes a lysine export function to the product of the ybjE gene of Escherichia coli, leading to a physiological scenario wherein two proteins, ArgO and YbjE, perform the task of separately exporting arginine and lysine

  8. Associations between socioeconomic status and allostatic load: effects of neighborhood poverty and tests of mediating pathways.

    Science.gov (United States)

    Schulz, Amy J; Mentz, Graciela; Lachance, Laurie; Johnson, Jonetta; Gaines, Causandra; Israel, Barbara A

    2012-09-01

    We examined relationships between neighborhood poverty and allostatic load in a low- to moderate-income multiracial urban community. We tested the hypothesis that neighborhood poverty is associated with allostatic load, controlling for household poverty. We also examined the hypotheses that this association was mediated by psychosocial stress and health-related behaviors. We conducted multilevel analyses using cross-sectional data from a probability sample survey in Detroit, Michigan (n = 919) and the 2000 US Census. The outcome measure was allostatic load. Independent variables included neighborhood and household poverty, psychosocial stress, and health-related behaviors. Covariates included neighborhood and individual demographic characteristics. Neighborhood poverty was positively associated with allostatic load (P poverty and controlling for potential confounders. Relationships between neighborhood poverty were mediated by self-reported neighborhood environment stress but not by health-related behaviors. Neighborhood poverty is associated with wear and tear on physiological systems, and this relationship is mediated through psychosocial stress. These relationships are evident after accounting for household poverty levels. Efforts to promote health equity should focus on neighborhood poverty, associated stressful environmental conditions, and household poverty.

  9. Akt Kinase-Mediated Checkpoint of cGAS DNA Sensing Pathway

    Directory of Open Access Journals (Sweden)

    Gil Ju Seo

    2015-10-01

    Full Text Available Upon DNA stimulation, cyclic GMP-AMP synthase (cGAS synthesizes the second messenger cyclic GMP-AMP (cGAMP that binds to the STING, triggering antiviral interferon-β (IFN-β production. However, it has remained undetermined how hosts regulate cGAS enzymatic activity after the resolution of DNA immunogen. Here, we show that Akt kinase plays a negative role in cGAS-mediated anti-viral immune response. Akt phosphorylated the S291 or S305 residue of the enzymatic domain of mouse or human cGAS, respectively, and this phosphorylation robustly suppressed its enzymatic activity. Consequently, expression of activated Akt led to the reduction of cGAMP and IFN-β production and the increase of herpes simplex virus 1 replication, whereas treatment with Akt inhibitor augmented cGAS-mediated IFN-β production. Furthermore, expression of the phosphorylation-resistant cGAS S291A mutant enhanced IFN-β production upon DNA stimulation, HSV-1 infection, and vaccinia virus infection. Our study identifies an Akt kinase-mediated checkpoint to fine-tune hosts’ immune responses to DNA stimulation.

  10. Akt Kinase-Mediated Checkpoint of cGAS DNA Sensing Pathway.

    Science.gov (United States)

    Seo, Gil Ju; Yang, Aerin; Tan, Brandon; Kim, Sungyoon; Liang, Qiming; Choi, Younho; Yuan, Weiming; Feng, Pinghui; Park, Hee-Sung; Jung, Jae U

    2015-10-13

    Upon DNA stimulation, cyclic GMP-AMP synthase (cGAS) synthesizes the second messenger cyclic GMP-AMP (cGAMP) that binds to the STING, triggering antiviral interferon-β (IFN-β) production. However, it has remained undetermined how hosts regulate cGAS enzymatic activity after the resolution of DNA immunogen. Here, we show that Akt kinase plays a negative role in cGAS-mediated anti-viral immune response. Akt phosphorylated the S291 or S305 residue of the enzymatic domain of mouse or human cGAS, respectively, and this phosphorylation robustly suppressed its enzymatic activity. Consequently, expression of activated Akt led to the reduction of cGAMP and IFN-β production and the increase of herpes simplex virus 1 replication, whereas treatment with Akt inhibitor augmented cGAS-mediated IFN-β production. Furthermore, expression of the phosphorylation-resistant cGAS S291A mutant enhanced IFN-β production upon DNA stimulation, HSV-1 infection, and vaccinia virus infection. Our study identifies an Akt kinase-mediated checkpoint to fine-tune hosts' immune responses to DNA stimulation. Copyright © 2015 The Authors. Published by Elsevier Inc. All rights reserved.

  11. Mediatization

    DEFF Research Database (Denmark)

    Hjarvard, Stig

    2017-01-01

    Mediatization research shares media effects studies' ambition of answering the difficult questions with regard to whether and how media matter and influence contemporary culture and society. The two approaches nevertheless differ fundamentally in that mediatization research seeks answers...... to these general questions by distinguishing between two concepts: mediation and mediatization. The media effects tradition generally considers the effects of the media to be a result of individuals being exposed to media content, i.e. effects are seen as an outcome of mediated communication. Mediatization...... research is concerned with long-term structural changes involving media, culture, and society, i.e. the influences of the media are understood in relation to how media are implicated in social and cultural changes and how these processes come to create new conditions for human communication and interaction...

  12. The Association of CXC Receptor 4 Mediated Signaling Pathway with Oxaliplatin-Resistant Human Colorectal Cancer Cells.

    Directory of Open Access Journals (Sweden)

    Wen-Shih Huang

    Full Text Available The stromal cell-derived factor-1 (SDF-1/CXC receptor 4 (CXCR4 axis plays an important role in tumor angiogenesis and invasiveness in colorectal cancer (CRC progression. In addition, metastatic CRC remains one of the most difficult human malignancies to treat because of its chemoresistant behavior. However, the mechanism by which correlation occurs between CXCR4 and the clinical response of CRC to chemotherapy remains unknown. We generated chemoresistant cells with increasing doses of oxaliplatin (OXA and 5-Fluorouracil (5FU to develop resistance at a clinical dose. We found that the putative markers did not change in the parental cells, but HCT-116/OxR and HCT-116/5-FUR were more aggressive and had higher tumor growth (demonstrated by wound healing, chemotaxis assay, and a nude mice xenograft model with the use of oxaliplatin. Apoptosis induced by oxaliplatin treatment was significantly decreased in HCT-116/OxR compared to the parental cells. Moreover, HCT-116/OxR cells displayed increased levels of p-gp, p-Akt p-ERK, p-IKBβ, CXCR4, and Bcl-2, but they also significantly inhibited the apoptotic pathways when compared to the parental strain. We evaluated the molecular mechanism governing the signaling pathway associated with anti-apoptosis activity and the aggressive status of chemoresistant cells. Experiments involving specific inhibitors demonstrated that the activation of the pathways associated with CXCR4, ERK1/2 mitogen-activated protein kinase (MAPK, and phosphatidylinositol 3-kinase (PI3K/Akt is critical to the functioning of the HCT-116/OxR and HCT-116/5-FUR characteristics of chemosensitivity. These findings elucidate the mechanism of CXCR4/PI3K/Akt downstream signaling and provide strategies to inhibit CXCR4 mediated signaling pathway in order to overcome CRC's resistance to chemotherapy.

  13. Arsenic trioxide mediates HAPI microglia inflammatory response and subsequent neuron apoptosis through p38/JNK MAPK/STAT3 pathway

    Energy Technology Data Exchange (ETDEWEB)

    Mao, Jiamin [Department of Environmental Health, School of Public Health, Nantong University, Nantong, Jiangsu 226001 (China); Yang, Jianbing [Department of Pediatrics, Affiliated Hospital of Nantong University, Nantong, Jiangsu 226001 (China); Zhang, Yan [Department of Nutrition and Food Hygiene, School of Public Health, Nantong University, Nantong, Jiangsu 226001 (China); Li, Ting [Department of Environmental Health, School of Public Health, Nantong University, Nantong, Jiangsu 226001 (China); Wang, Cheng [Department of Nutrition and Food Hygiene, School of Public Health, Nantong University, Nantong, Jiangsu 226001 (China); Xu, Lingfei; Hu, Qiaoyun; Wang, Xiaoke; Jiang, Shengyang [Department of Environmental Health, School of Public Health, Nantong University, Nantong, Jiangsu 226001 (China); Nie, Xiaoke [Department of Nutrition and Food Hygiene, School of Public Health, Nantong University, Nantong, Jiangsu 226001 (China); Chen, Gang, E-mail: chengang@ntu.edu.cn [Department of Environmental Health, School of Public Health, Nantong University, Nantong, Jiangsu 226001 (China)

    2016-07-15

    Arsenic is a widely distributed toxic metalloid all over the world. Inorganic arsenic species are supposed to affect astrocytic functions and to cause neuron apoptosis in CNS. Microglias are the key cell type involved in innate immune responses in CNS, and microglia activation has been linked to inflammation and neurotoxicity. In this study, using ELISA, we showed that Arsenic trioxide up-regulated the expression and secretion of IL-1β in a dose-dependent manner and a time-dependent manner in cultured HAPI microglia cells. The secretion of IL-1β caused the apoptosis of SH-SY5Y. These pro-inflammatory responses were inhibited by the STAT3 blocker, AG490 and P38/JNK MAPK blockers SB202190, SP600125. Further, Arsenic trioxide exposure could induce phosphorylation and activation of STAT3, and the translocation of STAT3 from the cytosol to the nucleus in this HAPI microglia cell line. Thus, the STAT3 signaling pathway can be activated after Arsenic trioxide treatment. However, P38/JNK MAPK blockers SB202190, SP600125 also obviously attenuated STAT3 activation and transnuclear transport induced by Arsenic trioxide. In concert with these results, we highlighted that the secretion of IL-1β and STAT3 activation induced by Arsenic trioxide can be mediated by elevation of P38/JNK MAPK in HAPI microglia cells and then induced the toxicity of neurons. - Highlights: • Arsenic trioxide exposure induced expression of IL-β in HAPI microglia. • Arsenic trioxide exposure induced activation of MAPK pathways in HAPI microglia. • Arsenic trioxide exposure induced activation of STAT3 pathways in HAPI microglia. • The expression of IL-β though P38/JNK MAPK/STAT3 pathways in HAPI microglia.

  14. Arsenic trioxide mediates HAPI microglia inflammatory response and subsequent neuron apoptosis through p38/JNK MAPK/STAT3 pathway

    International Nuclear Information System (INIS)

    Mao, Jiamin; Yang, Jianbing; Zhang, Yan; Li, Ting; Wang, Cheng; Xu, Lingfei; Hu, Qiaoyun; Wang, Xiaoke; Jiang, Shengyang; Nie, Xiaoke; Chen, Gang

    2016-01-01

    Arsenic is a widely distributed toxic metalloid all over the world. Inorganic arsenic species are supposed to affect astrocytic functions and to cause neuron apoptosis in CNS. Microglias are the key cell type involved in innate immune responses in CNS, and microglia activation has been linked to inflammation and neurotoxicity. In this study, using ELISA, we showed that Arsenic trioxide up-regulated the expression and secretion of IL-1β in a dose-dependent manner and a time-dependent manner in cultured HAPI microglia cells. The secretion of IL-1β caused the apoptosis of SH-SY5Y. These pro-inflammatory responses were inhibited by the STAT3 blocker, AG490 and P38/JNK MAPK blockers SB202190, SP600125. Further, Arsenic trioxide exposure could induce phosphorylation and activation of STAT3, and the translocation of STAT3 from the cytosol to the nucleus in this HAPI microglia cell line. Thus, the STAT3 signaling pathway can be activated after Arsenic trioxide treatment. However, P38/JNK MAPK blockers SB202190, SP600125 also obviously attenuated STAT3 activation and transnuclear transport induced by Arsenic trioxide. In concert with these results, we highlighted that the secretion of IL-1β and STAT3 activation induced by Arsenic trioxide can be mediated by elevation of P38/JNK MAPK in HAPI microglia cells and then induced the toxicity of neurons. - Highlights: • Arsenic trioxide exposure induced expression of IL-β in HAPI microglia. • Arsenic trioxide exposure induced activation of MAPK pathways in HAPI microglia. • Arsenic trioxide exposure induced activation of STAT3 pathways in HAPI microglia. • The expression of IL-β though P38/JNK MAPK/STAT3 pathways in HAPI microglia.

  15. BID links ferroptosis to mitochondrial cell death pathways

    NARCIS (Netherlands)

    Neitemeier, Sandra; Jelinek, Anja; Laino, Vincenzo; Hoffmann, Lena; Eisenbach, Ina; Eying, Roman; Ganjam, Goutham K; Dolga, Amalia M; Oppermann, Sina; Culmsee, Carsten

    2017-01-01

    Ferroptosis has been defined as an oxidative and iron-dependent pathway of regulated cell death that is distinct from caspase-dependent apoptosis and established pathways of death receptor-mediated regulated necrosis. While emerging evidence linked features of ferroptosis induced e.g. by

  16. UPP mediated Diabetic Retinopathy via ROS/PARP and NF-κB inflammatory factor pathways.

    Science.gov (United States)

    Luo, D-W; Zheng, Z; Wang, H; Fan, Y; Chen, F; Sun, Y; Wang, W-J; Sun, T; Xu, X

    2015-01-01

    Diabetic retinopathy (DR) is a leading cause of blindness in adults at working age. Human diabetic retinopathy is characterized by the basement membrane thick, pericytes loss, microaneurysms formation, retina neovascularization and vitreous hemorrhage. To investigate whether UPP activated ROS/PARP and NF-κB inflammatory factor pathways in Diabetic Retinopathy, human retinal endothelial cells (HRECs) and rats with streptozotocin-induced diabetes were used to determine the effect of UPP on ROS generation, cell apoptosis, mitochondrial membrane potential (ΔΨm) and inflammatory factor protein expression, through flow cytometry assay, immunohistochemistry, Real-time PCR, Western blot analysis and ELISA. The levels of ROS and apoptosis and the expressions of UPP (Ub and E3) and inflammatory factor protein were increased in high glucose-induced HRECs and retina of diabetic rats, while ΔΨm was decreased. The UPP inhibitor and UbshRNA could attenuate these effects through inhibiting the pathway of ROS/PARP and the expression of NF-κB inflammatory factors, and the increased UPP was a result of high glucose-induced increase of ROS generation and NF-κBp65 expression, accompanied with the decrease of ΔΨm. Clinical study showed the overexpression of UPP and detachment of epiretinal membranes in proliferative DR (PDR) patients. It has been indicated that the pathogenic effect of UPP on DR was involved in the increase of ROS generation and NF-κB expression, which associated with the ROS/PARP and NF-κB inflammatory factor pathways. Our study supports a new insight for further application of UPP inhibitor in DR treatment.

  17. Trophoblast cell fusion and differentiation are mediated by both the protein kinase C and a pathways.

    Directory of Open Access Journals (Sweden)

    Waka Omata

    Full Text Available The syncytiotrophoblast of the human placenta is an epithelial barrier that interacts with maternal blood and is a key for the transfer of nutrients and other solutes to the developing fetus. The syncytiotrophoblast is a true syncytium and fusion of progenitor cytotrophoblasts is the cardinal event leading to the formation of this layer. BeWo cells are often used as a surrogate for cytotrophoblasts, since they can be induced to fuse, and then express certain differentiation markers associated with trophoblast syncytialization. Dysferlin, a syncytiotrophoblast membrane repair protein, is up-regulated in BeWo cells induced to fuse by treatment with forskolin; this fusion is thought to occur through cAMP/protein kinase A-dependent mechanisms. We hypothesized that dysferlin may also be up-regulated in response to fusion through other pathways. Here, we show that BeWo cells can also be induced to fuse by treatment with an activator of protein kinase C, and that this fusion is accompanied by increased expression of dysferlin. Moreover, a dramatic synergistic increase in dysferlin expression is observed when both the protein kinase A and protein kinase C pathways are activated in BeWo cells. This synergy in fusion is also accompanied by dramatic increases in mRNA for the placental fusion proteins syncytin 1, syncytin 2, as well as dysferlin. Dysferlin, however, was shown to be dispensable for stimulus-induced BeWo cell syncytialization, since dysferlin knockdown lines fused to the same extent as control cells. The classical trophoblast differentiation marker human chorionic gonadotropin was also monitored and changes in the expression closely parallel that of dysferlin in all of the experimental conditions employed. Thus different biochemical markers of trophoblast fusion behave in concert supporting the hypothesis that activation of both protein kinase C and A pathways lead to trophoblastic differentiation.

  18. Bmh1p (14-3-3) mediates pathways associated with virulence in Candida albicans.

    Science.gov (United States)

    Kelly, Michelle N; Johnston, Douglas A; Peel, Bethany A; Morgan, Timothy W; Palmer, Glen E; Sturtevant, Joy E

    2009-05-01

    The ability of the pathogenic fungus Candida albicans to cause disease requires rapid adaptation to changes in the host environment and to an evolving host immune response. The identification of 'virulence factors' using in vitro characterization of mutant strains has traditionally relied on a common set of phenotypic and biochemical assays (most often performed at 30 degrees C) and the subsequent correlation with their corresponding virulence in mouse models of disease. Utilizing a panel of isogenic mutants for the multifunctional signal-modulating 14-3-3 protein (Bmh1p), we have found that specific mutations affect a variety of different pathways currently associated with virulence, including those involved with the formation of filaments, as well as interaction with host immune cells. Surprisingly, our studies revealed that deficiencies in many of these pathways do not always correlate with virulence in a mouse model of disseminated infection. Mutations within the binding pocket of Bmh1p that affect the ability of the protein to efficiently bind ligand had varying effects on the results of a number of in vitro and in vivo assays. The capability, in vitro, to filament in embedment conditions, and to filament and form chlamydospores under microaerophilic conditions on cornmeal agar, does not correlate with virulence. It is likely that only a subset of hyphal signalling pathways is actually required for the establishment of infection in the disseminated mouse model. Most importantly, our results suggest that the delayed onset of log-phase [corrected] growth in vitro at 37 degrees C, and not at 30 degrees C, results in an inability of these mutants to rapidly adjust to environmental changes in vivo and may be responsible for their increased clearance and reduced virulence. It is critical, therefore, that future in vitro studies of putative virulence factors in C. albicans include careful characterization at physiological temperatures.

  19. Pathways from Religion to Health: Mediation by Psychosocial and Lifestyle Mechanisms.

    Science.gov (United States)

    Morton, Kelly R; Lee, Jerry W; Martin, Leslie R

    2017-02-01

    Religiosity, often measured as attendance at religious services, is linked to better physical health and longevity though the mechanisms linking the two are debated. Potential explanations include: a healthier lifestyle, increased social support from congregational members, and/or more positive emotions. Thus far, these mechanisms have not been tested simultaneously in a single model though they likely operate synergistically. We test this model predicting all-cause mortality in Seventh-day Adventists, a denomination that explicitly promotes a healthy lifestyle. This allows the more explicit health behaviors linked to the religious doctrine (e.g., healthy diet) to be compared with other mechanisms not specific to religious doctrine (e.g., social support and positive emotions). Finally, this study examines both Church Activity (including worship attendance and church responsibilities) and Religious Engagement (coping, importance, and intrinsic beliefs). Religious Engagement is more is more inner-process focused (vs. activity-based) and less likely to be confounded with age and its associated functional status limitations, although it should be noted that age is controlled in the present study. The findings suggest that Religious Engagement and Church Activity operate through the mediators of health behavior, emotion, and social support to decrease mortality risk. All links between Religious Engagement and mortality are positive but indirect through positive Religious Support, Emotionality, and lifestyle mediators. However, Church Activity has a direct positive effect on mortality as well as indirect effects through, Religious Support, Emotionality, and lifestyle mediators (diet and exercise). The models were invariant by gender and for both Blacks and Whites.

  20. Subcutaneous white adipocytes express a light sensitive signaling pathway mediated via a melanopsin/TRPC channel axis.

    Science.gov (United States)

    Ondrusova, Katarina; Fatehi, Mohammad; Barr, Amy; Czarnecka, Zofia; Long, Wentong; Suzuki, Kunimasa; Campbell, Scott; Philippaert, Koenraad; Hubert, Matthew; Tredget, Edward; Kwan, Peter; Touret, Nicolas; Wabitsch, Martin; Lee, Kevin Y; Light, Peter E

    2017-11-27

    Subcutaneous white adipose tissue (scWAT) is the major fat depot in humans and is a central player in regulating whole body metabolism. Skin exposure to UV wavelengths from sunlight is required for Vitamin D synthesis and pigmentation, although it is plausible that longer visible wavelengths that penetrate the skin may regulate scWAT function. In this regard, we discovered a novel blue light-sensitive current in human scWAT that is mediated by melanopsin coupled to transient receptor potential canonical cation channels. This pathway is activated at physiological intensities of light that penetrate the skin on a sunny day. Daily exposure of differentiated adipocytes to blue light resulted in decreased lipid droplet size, increased basal lipolytic rate and alterations in adiponectin and leptin secretion. Our results suggest that scWAT function may be directly under the influence of ambient sunlight exposure and may have important implications for our current understanding of adipocyte biology. (150 words).

  1. The BRO proteins of Bombyx mori nucleopolyhedrovirus are nucleocytoplasmic shuttling proteins that utilize the CRM1-mediated nuclear export pathway

    International Nuclear Information System (INIS)

    Kang, Won Kyung; Kurihara, Masaaki; Matsumoto, Shogo

    2006-01-01

    The BRO proteins of Bombyx mori nucleopolyhedrovirus (BmNPV) display a biphasic pattern of intracellular localization during infection. At early times, they reside in the nucleus but then show both cytoplasmic and nuclear localization as the infection proceeds. Therefore, we examined the possibility of nuclear export. Using inhibitors, we reveal that BmNPV BRO proteins shuttle between the nucleus and cytoplasm. Mutations on the leucine-rich region of BRO proteins resulted in nuclear accumulation of transiently expressed proteins, suggesting that this region functions as a CRM1-dependent nuclear export signal (NES). On the contrary, mutant BRO-D with an altered NES did not show nuclear accumulation in infected cells, although protein production seemed to be reduced. RT-PCR analysis showed that the lower level of protein production was due to a reduction in RNA synthesis. Taken together, our results suggest that BRO proteins are nucleocytoplasmic shuttling proteins that utilize the CRM1-mediated nuclear export pathway

  2. [Ursodeoxycholic acid induced apoptosis of human hepatoma cells HepG2 and SMMC-7721 bymitochondrial-mediated pathway].

    Science.gov (United States)

    Wu, Duan; Zhou, Jianyin; Yin, Zhenyu; Liu, Pingguo; Zhao, Yilin; Liu, Jianming; Wang, Xiaomin

    2014-12-02

    To explore the effects and underlying mechanisms of ursodeoxycholic acid on human hepatoma cells. HepG2 and SMMC-7721 HCC cell lines were respectively treated with ursodeoxycholic acid. And cell proliferation, apoptosis and the expression of Bax/Bcl-2 gene were detected by methyl thiazolyl tetrazolium (MTT), inverted microscopy, fluorescent microscopy, flow cytometry and Western blot. Ursodeoxycholic acid significantly inhibited the proliferation of human hepatoma cells in a concentration- and time-dependent manner. The half maximal inhibitory concentrations (IC50) of HepG2 and SMMC-7721 were 397.3 and 387.7 µg/ml respectively after a 48-hour treatment of 400 µg /ml ursodeoxycholic acid. And it also induced the apoptosis of HepG2 and SMMC-7721 cells, up-regulated Bax gene and down-regulated Bcl-2 gene. Ursodeoxycholic acid inhibits the proliferation of hepatoma cells and induce apoptosis by mitochondrial-mediated pathway.

  3. Atractylenolide I-mediated Notch pathway inhibition attenuates gastric cancer stem cell traits

    Energy Technology Data Exchange (ETDEWEB)

    Ma, Li; Mao, Rurong; Shen, Ke; Zheng, Yuanhong; Li, Yueqi [State Key Laboratory of Bioreactor Engineering and Shanghai Key Laboratory of New Drug Design, School of Pharmacy, East China University of Science and Technology, #268, 130 Meilong Road, Shanghai 200237 (China); Liu, Jianwen, E-mail: liujian@ecust.edu.cn [State Key Laboratory of Bioreactor Engineering and Shanghai Key Laboratory of New Drug Design, School of Pharmacy, East China University of Science and Technology, #268, 130 Meilong Road, Shanghai 200237 (China); Ni, Lei, E-mail: nilei625@yahoo.com [Department of Respiration, Ruijin Hospital, Shanghai Jiaotong University School of Medicine, 197 Ruijin Road II, Shanghai 200025 (China)

    2014-07-18

    Highlights: • This paper supports the anti-tumor effects of AT-I on gastric cancer in vitro. • AT-I attenuates gastric cancer stem cell traits. • It is the systematic study regarding AT-I suppression of Notch pathway in GC and GCSLCs. - Abstract: Atractylenolide I (AT-I), one of the main naturally occurring compounds of Rhizoma Atractylodis Macrocephalae, has remarkable anti-cancer effects on various cancers. However, its effects on the treatment of gastric cancer remain unclear. Via multiple cellular and molecular approaches, we demonstrated that AT-I could potently inhibit cancer cell proliferation and induce apoptosis through inactivating Notch pathway. AT-I treatment led to the reduction of expressions of Notch1, Jagged1, and its downstream Hes1/ Hey1. Our results showed that AT-I inhibited the self-renewal capacity of gastric stem-like cells (GCSLCs) by suppression of their sphere formation capacity and cell viability. AT-I attenuated gastric cancer stem cell (GCSC) traits partly through inactivating Notch1, leading to reducing the expressions of its downstream target Hes1, Hey1 and CD44 in vitro. Collectively, our results suggest that AT-I might develop as a potential therapeutic drug for the treatment of gastric cancer.

  4. MVP interacts with YPEL4 and inhibits YPEL4-mediated activities of the ERK signal pathway.

    Science.gov (United States)

    Liang, Pei; Wan, Yongqi; Yan, Yan; Wang, Yuequn; Luo, Na; Deng, Yun; Fan, Xiongwei; Zhou, Junmei; Li, Yongqing; Wang, Zequn; Yuan, Wuzhou; Tang, Ming; Mo, Xiaoyang; Wu, Xiushan

    2010-06-01

    Human YPEL4 is a member of YPEL family. It contains a Yippee domain, which is a putative zinc-finger-like, metal-binding domain. The human YPEL4 gene maps to chromosome 11q12.1, is ubiquitously expressed in adult tissues, and encodes a nuclear protein of 127 amino acids, the function of which remains unknown. To gain insights into the cellular function of this protein, we searched for YPEL4-interacting proteins using a yeast two-hybrid screen. The major vault protein (MVP), a lung resistance associated protein, was identified as a binding partner of YPEL4. The interaction between YPEL4 and MVP in mammalian cells was further demonstrated by a series of biochemical assays including the mammalian two-hybrid assay, GST pull-down assay, co-immunoprecipitation assay, and immunocytochemistry. Using a reporter system, we found that MVP can inhibit YPEL4's ability to activate Elk-1 in the MAPK signaling pathway. This study provides new clues for understanding the molecular mechanism of YPEL4 in cell division and signal transduction pathways and should be helpful for understanding molecular functions of the YPEL family.

  5. Atractylenolide I-mediated Notch pathway inhibition attenuates gastric cancer stem cell traits

    International Nuclear Information System (INIS)

    Ma, Li; Mao, Rurong; Shen, Ke; Zheng, Yuanhong; Li, Yueqi; Liu, Jianwen; Ni, Lei

    2014-01-01

    Highlights: • This paper supports the anti-tumor effects of AT-I on gastric cancer in vitro. • AT-I attenuates gastric cancer stem cell traits. • It is the systematic study regarding AT-I suppression of Notch pathway in GC and GCSLCs. - Abstract: Atractylenolide I (AT-I), one of the main naturally occurring compounds of Rhizoma Atractylodis Macrocephalae, has remarkable anti-cancer effects on various cancers. However, its effects on the treatment of gastric cancer remain unclear. Via multiple cellular and molecular approaches, we demonstrated that AT-I could potently inhibit cancer cell proliferation and induce apoptosis through inactivating Notch pathway. AT-I treatment led to the reduction of expressions of Notch1, Jagged1, and its downstream Hes1/ Hey1. Our results showed that AT-I inhibited the self-renewal capacity of gastric stem-like cells (GCSLCs) by suppression of their sphere formation capacity and cell viability. AT-I attenuated gastric cancer stem cell (GCSC) traits partly through inactivating Notch1, leading to reducing the expressions of its downstream target Hes1, Hey1 and CD44 in vitro. Collectively, our results suggest that AT-I might develop as a potential therapeutic drug for the treatment of gastric cancer

  6. Electroacupuncture Exerts Neuroprotection through Caveolin-1 Mediated Molecular Pathway in Intracerebral Hemorrhage of Rats.

    Science.gov (United States)

    Li, Hui-Qin; Li, Yan; Chen, Zi-Xian; Zhang, Xiao-Guang; Zheng, Xia-Wei; Yang, Wen-Ting; Chen, Shuang; Zheng, Guo-Qing

    2016-01-01

    Spontaneous intracerebral hemorrhage (ICH) is one of the most devastating types of stroke. Here, we aim to demonstrate that electroacupuncture on Baihui (GV20) exerts neuroprotection for acute ICH possibly via the caveolin-1/matrix metalloproteinase/blood-brain barrier permeability pathway. The model of ICH was established by using collagenase VII. Rats were randomly divided into three groups: Sham-operation group, Sham electroacupuncture group, and electroacupuncture group. Each group was further divided into 4 subgroups according to the time points of 6 h, 1 d, 3 d, and 7 d after ICH. The methods were used including examination of neurological deficit scores according to Longa's scale, measurement of blood-brain barrier permeability through Evans Blue content, in situ immunofluorescent detection of caveolin-1 in brains, western blot analysis of caveolin-1 in brains, and in situ zymography for measuring matrix metalloproteinase-2/9 activity in brains. Compared with Sham electroacupuncture group, electroacupuncture group has resulted in a significant improvement in neurological deficit scores and in a reduction in Evans Blue content, expression of caveolin-1, and activity of matrix metalloproteinase-2/9 at 6 h, 1 d, 3 d, and 7 d after ICH ( P electroacupuncture on GV20 can improve neurological deficit scores and reduce blood-brain barrier permeability after ICH, and the mechanism possibly targets caveolin-1/matrix metalloproteinase/blood-brain barrier permeability pathway.

  7. HECTD3 Mediates an HSP90-Dependent Degradation Pathway for Protein Kinase Clients

    Directory of Open Access Journals (Sweden)

    Zhaobo Li

    2017-06-01

    Full Text Available Inhibition of the ATPase cycle of the HSP90 chaperone promotes ubiquitylation and proteasomal degradation of its client proteins, which include many oncogenic protein kinases. This provides the rationale for HSP90 inhibitors as cancer therapeutics. However, the mechanism by which HSP90 ATPase inhibition triggers ubiquitylation is not understood, and the E3 ubiquitin ligases involved are largely unknown. Using a siRNA screen, we have identified components of two independent degradation pathways for the HSP90 client kinase CRAF. The first requires CUL5, Elongin B, and Elongin C, while the second requires the E3 ligase HECTD3, which is also involved in the degradation of MASTL and LKB1. HECTD3 associates with HSP90 and CRAF in cells via its N-terminal DOC domain, which is mutationally disrupted in tumor cells with activated MAP kinase signaling. Our data implicate HECTD3 as a tumor suppressor modulating the activity of this important oncogenic signaling pathway.

  8. Development pathways from abusive parenting to delinquency: the mediating role of depression and aggression.

    Science.gov (United States)

    You, Sukkyung; Lim, Sun Ah

    2015-08-01

    This study investigated the long-term relationship between abusive parenting and adolescent mental health, and the path to delinquent behavior. Longitudinal data from 5th through 7th graders from the Korean Children and Youth Panel Survey (KCYPS) were analyzed to examine if abusive parenting was a predictor of early adolescent delinquency behavior, via aggression and depression as mediating factors. The results were as follows. First, parental abuse (both emotional and physical) was found to have significant effects on children's psychosocial factors (aggression and depression), while parental neglect (both emotional and physical) had significant effects on depression alone and not on aggression. Second, aggression exerted significant effects on both violent and non-violent delinquent behaviors, while depression had a significant effect on only non-violent delinquent behaviors. Third, children's psychosocial factors (aggression and depression) played significant mediating roles between earlier abusive parenting and delinquent behaviors. Fourth, for children living in a family with their grandparents, paths from abusive parenting, psychosocial adaptation, and later delinquent behaviors were not significant, implying that living with grandparents played a protective factor in these relationships. Copyright © 2015 Elsevier Ltd. All rights reserved.

  9. Bridge mediated two-electron transfer reactions: Analysis of stepwise and concerted pathways

    International Nuclear Information System (INIS)

    Petrov, E.G.; May, V.

    2004-01-01

    A theory of nonadiabatic donor (D)-acceptor (A) two-electron transfer (TET) mediated by a single regular bridge (B) is developed. The presence of different intermediate two-electron states connecting the reactant state D -- BA with the product state DBA -- results in complex multiexponential kinetics. The conditions are discussed at which a reduction to two-exponential as well as single-exponential kinetics becomes possible. For the latter case the rate K TET is calculated, which describes the bridge-mediated reaction as an effective two-electron D-A transfer. In the limit of small populations of the intermediate TET states D - B - A, DB -- A, D - BA - , and DB - A - , K TET is obtained as a sum of the rates K TET (step) and K TET (sup) . The first rate describes stepwise TET originated by transitions of a single electron. It starts at D -- BA and reaches DBA -- via the intermediate state D - BA - . These transitions cover contributions from sequential as well as superexchange reactions all including reduced bridge states. In contrast, a specific two-electron superexchange mechanism from D -- BA to DBA -- defines K TET (sup) . An analytic dependence of K TET (step) and K TET (sup) on the number of bridging units is presented and different regimes of D-A TET are studied

  10. The translocation of fullerenic nanoparticles into lysosome via the pathway of clathrin-mediated endocytosis

    International Nuclear Information System (INIS)

    Li Wei; Chen Chunying; Ye Chang; Zhao Yuliang; Chen Zhen; Meng Huan; Gao Yuxi; Yuan Hui; Xing Genmei; Zhao Feng; Chai Zhifang; Wei Taotao; Zhang Xujia; Yang Fuyu; Lao Fang; Han Dong; Tang Xianhua; Zhang Yingge

    2008-01-01

    Manufactured fullerene nanoparticles easily enter into cells and hence have been rapidly developed for biomedical uses. However, it is generally unknown which route the nanoparticles undergo when crossing cell membranes and where they localize to the intracellular compartments. Herein we have used both microscopic imaging and biological techniques to explore the processes of [C 60 (C(COOH) 2 ) 2 ] n nanoparticles across cellular membranes and their intracellular translocation in 3T3 L1 and RH-35 living cells. The fullerene nanoparticles are quickly internalized by the cells and then routed to the cytoplasm with punctate localization. Upon entering the cell, they are synchronized to lysosome-like vesicles. The [C 60 (C(COOH) 2 ) 2 ] n nanoparticles entering cells are mainly via endocytosis with time-, temperature- and energy-dependent manners. The cellular uptake of [C 60 (C(COOH) 2 ) 2 ] n nanoparticles was found to be clathrin-mediated but not caveolae-mediated endocytosis. The endocytosis mechanism and the subcellular target location provide key information for the better understanding and predicting of the biomedical function of fullerene nanoparticles inside cells

  11. Porcine parvovirus infection induces apoptosis in PK-15 cells through activation of p53 and mitochondria-mediated pathway

    International Nuclear Information System (INIS)

    Zhang, Hongling; Huang, Yong; Du, Qian; Luo, Xiaomao; Zhang, Liang; Zhao, Xiaomin; Tong, Dewen

    2015-01-01

    Highlights: • PPV reduces PK-15 cells viability by inducing apoptosis. • PPV infection induces apoptosis through mitochondria-mediated pathway. • PPV infection activates p53 to regulate the mitochondria apoptotic signaling. - Abstract: Porcine parvovirus (PPV) infection has been reported to induce the cytopathic effects (CPE) in some special host cells and contribute the occurrence of porcine parvovirus disease, but the molecular mechanisms underlying PPV-induced CPE are not clear. In this study, we investigated the morphological and molecular changes of porcine kidney cell line (PK-15 cells) infected with PPV. The results showed that PPV infection inhibited the viability of PK-15 cells in a time and concentration dependent manner. PPV infection induced typical apoptotic features including chromatin condensation, apoptotic body formation, nuclear fragmentation, and Annexin V-binding activity. Further studies showed that Bax was increased and translocated to mitochondria, whereas Bcl-2 was decreased in PPV-infected cells, which caused mitochondrial outer-membrane permeabilization, resulting in the release of mitochondrial cytochrome c, followed by caspase-9 and caspase-3 activation. However, the expression of Fas and Fas ligand (FasL) did not appear significant changes in the process of PPV-induced apoptosis. Moreover, PPV infection activated p53 signaling, which was involved in the activation of apoptotic signaling induced by PPV infection via regulation of Bax and Bcl-2. Taken together, our results demonstrated that PPV infection induced apoptosis in PK-15 cells through activation of p53 and mitochondria-mediated apoptosis pathway. This study may contribute to shed light on the molecular pathogenesis of PPV infection

  12. Porcine parvovirus infection induces apoptosis in PK-15 cells through activation of p53 and mitochondria-mediated pathway

    Energy Technology Data Exchange (ETDEWEB)

    Zhang, Hongling; Huang, Yong; Du, Qian; Luo, Xiaomao; Zhang, Liang; Zhao, Xiaomin; Tong, Dewen, E-mail: dwtong@nwsuaf.edu.cn

    2015-01-09

    Highlights: • PPV reduces PK-15 cells viability by inducing apoptosis. • PPV infection induces apoptosis through mitochondria-mediated pathway. • PPV infection activates p53 to regulate the mitochondria apoptotic signaling. - Abstract: Porcine parvovirus (PPV) infection has been reported to induce the cytopathic effects (CPE) in some special host cells and contribute the occurrence of porcine parvovirus disease, but the molecular mechanisms underlying PPV-induced CPE are not clear. In this study, we investigated the morphological and molecular changes of porcine kidney cell line (PK-15 cells) infected with PPV. The results showed that PPV infection inhibited the viability of PK-15 cells in a time and concentration dependent manner. PPV infection induced typical apoptotic features including chromatin condensation, apoptotic body formation, nuclear fragmentation, and Annexin V-binding activity. Further studies showed that Bax was increased and translocated to mitochondria, whereas Bcl-2 was decreased in PPV-infected cells, which caused mitochondrial outer-membrane permeabilization, resulting in the release of mitochondrial cytochrome c, followed by caspase-9 and caspase-3 activation. However, the expression of Fas and Fas ligand (FasL) did not appear significant changes in the process of PPV-induced apoptosis. Moreover, PPV infection activated p53 signaling, which was involved in the activation of apoptotic signaling induced by PPV infection via regulation of Bax and Bcl-2. Taken together, our results demonstrated that PPV infection induced apoptosis in PK-15 cells through activation of p53 and mitochondria-mediated apoptosis pathway. This study may contribute to shed light on the molecular pathogenesis of PPV infection.

  13. The acute neutrophil response mediated by S100 alarmins during vaginal Candida infections is independent of the Th17-pathway.

    Science.gov (United States)

    Yano, Junko; Kolls, Jay K; Happel, Kyle I; Wormley, Floyd; Wozniak, Karen L; Fidel, Paul L

    2012-01-01

    Vulvovaginal candidiasis (VVC) caused by Candida albicans affects a significant number of women during their reproductive ages. Clinical observations revealed that a robust vaginal polymorphonuclear neutrophil (PMN) migration occurs in susceptible women, promoting pathological inflammation without affecting fungal burden. Evidence to date in the mouse model suggests that a similar acute PMN migration into the vagina is mediated by chemotactic S100A8 and S100A9 alarmins produced by vaginal epithelial cells in response to Candida. Based on the putative role for the Th17 response in mucosal candidiasis as well as S100 alarmin induction, this study aimed to determine whether the Th17 pathway plays a role in the S100 alarmin-mediated acute inflammation during VVC using the experimental mouse model. For this, IL-23p19(-/-), IL-17RA(-/-) and IL-22(-/-) mice were intravaginally inoculated with Candida, and vaginal lavage fluids were evaluated for fungal burden, PMN infiltration, the presence of S100 alarmins and inflammatory cytokines and chemokines. Compared to wild-type mice, the cytokine-deficient mice showed comparative levels of vaginal fungal burden and PMN infiltration following inoculation. Likewise, inoculated mice of all strains with substantial PMN infiltration exhibited elevated levels of vaginal S100 alarmins in both vaginal epithelia and secretions in the vaginal lumen. Finally, cytokine analyses of vaginal lavage fluid from inoculated mice revealed equivalent expression profiles irrespective of the Th17 cytokine status or PMN response. These data suggest that the vaginal S100 alarmin response to Candida does not require the cells or cytokines of the Th17 lineage, and therefore, the immunopathogenic inflammatory response during VVC occurs independently of the Th17-pathway.

  14. A Recombinant Fragment of Human Surfactant Protein D induces Apoptosis in Pancreatic Cancer Cell Lines via Fas-Mediated Pathway.

    Science.gov (United States)

    Kaur, Anuvinder; Riaz, Muhammad Suleman; Murugaiah, Valarmathy; Varghese, Praveen Mathews; Singh, Shiv K; Kishore, Uday

    2018-01-01

    Human surfactant protein D (SP-D) is a potent innate immune molecule, which is emerging as a key molecule in the recognition and clearance of altered and non-self targets. Previous studies have shown that a recombinant fragment of human SP-D (rfhSP-D) induced apoptosis via p53-mediated apoptosis pathway in an eosinophilic leukemic cell line, AML14.3D10. Here, we report the ability of rfhSP-D to induce apoptosis via TNF-α/Fas-mediated pathway regardless of the p53 status in human pancreatic adenocarcinoma using Panc-1 (p53 mt ), MiaPaCa-2 (p53 mt ), and Capan-2 (p53 wt ) cell lines. Treatment of these cell lines with rfhSP-D for 24 h caused growth arrest in G1 cell cycle phase and triggered transcriptional upregulation of pro-apoptotic factors such as TNF-α and NF-κB. Translocation of NF-κB from the cytoplasm into the nucleus of pancreatic cancer cell lines was observed via immunofluorescence microscopy following treatment with rfhSP-D as compared to the untreated cells. The rfhSP-D treatment caused upregulation of pro-apoptotic marker Fas, as analyzed via qPCR and western blot, which then triggered caspase cascade, as evident from cleavage of caspase 8 and 3 analyzed via western blot at 48 h. The cell number following the rfhSP-D treatment was reduced in the order of Panc-1 (~67%) > MiaPaCa-2 (~60%) > Capan-2 (~35%). This study appears to suggest that rfhSP-D can potentially be used to therapeutically target pancreatic cancer cells irrespective of their p53 phenotype.

  15. Anti-inflammatory effects of benfotiamine are mediated through the regulation of the arachidonic acid pathway in macrophages.

    Science.gov (United States)

    Shoeb, Mohammad; Ramana, Kota V

    2012-01-01

    Benfotiamine, a lipid-soluble analogue of vitamin B1, is a potent antioxidant that is used as a food supplement for the treatment of diabetic complications. Our recent study (U.C. Yadav et al., Free Radic. Biol. Med. 48:1423-1434, 2010) indicates a novel role for benfotiamine in the prevention of bacterial endotoxin, lipopolysaccharide (LPS)-induced cytotoxicity and inflammatory response in murine macrophages. Nevertheless, it remains unclear how benfotiamine mediates anti-inflammatory effects. In this study, we investigated the anti-inflammatory role of benfotiamine in regulating arachidonic acid (AA) pathway-generated inflammatory lipid mediators in RAW264.7 macrophages. Benfotiamine prevented the LPS-induced activation of cPLA2 and release of AA metabolites such as leukotrienes, prostaglandin E2, thromboxane 2 (TXB2), and prostacyclin (PGI2) in macrophages. Further, LPS-induced expression of AA-metabolizing enzymes such as COX-2, LOX-5, TXB synthase, and PGI2 synthase was significantly blocked by benfotiamine. Furthermore, benfotiamine prevented the LPS-induced phosphorylation of ERK1/2 and expression of transcription factors NF-κB and Egr-1. Benfotiamine also prevented the LPS-induced oxidative stress and protein-HNE adduct formation. Most importantly, compared to specific COX-2 and LOX-5 inhibitors, benfotiamine significantly prevented LPS-induced macrophage death and monocyte adhesion to endothelial cells. Thus, our studies indicate that the dual regulation of the COX and LOX pathways in AA metabolism could be a novel mechanism by which benfotiamine exhibits its potential anti-inflammatory response. Copyright © 2011 Elsevier Inc. All rights reserved.

  16. Anti-Inflammatory Effects of Benfotiamine are Mediated Through the Regulation of Arachidonic Acid Pathway in Macrophages

    Science.gov (United States)

    Shoeb, Mohammad; Ramana, Kota V

    2011-01-01

    Benfotiamine, a lipid-soluble analogue of vitamin B1, is a potent anti-oxidant that is used as a food supplement for the treatment of diabetic complications. Our recent study indicates a novel role of benfotiamine in the prevention of bacterial endotoxin, lipopolysaccharide (LPS)-induced cytotoxicity and inflammatory response in murine macrophages. Nevertheless, it remains unclear how benfotiamine mediates anti-inflammatory effects. In this study, we investigated the anti-inflammatory role of benfotiamine in regulating the arachidonic acid (AA) pathway generated inflammatory lipid mediators in RAW 264.7 macrophages. Benfotiamine prevented the LPS-induced activation of cPLA2 and release of AA metabolites such as leukotrienes (LTB4), prostaglandin E2 (PGE2), thromboxanes 2 (TXB2) and prostacyclin (PGI2) in macrophages. Further, LPS-induced expressions of AA metabolizing enzymes such as COX-2, LOX-5, TXB synthase and PGI2 synthase were significantly blocked by benfotiamine. Furthermore, benfotiamine prevented the LPS-induced phosphorylation of ERK1/2 and expression of transcription factors NF-kB, and Egr-1. Benfotiamine also prevented the LPS-induced oxidative stress and protein-HNE adducts formation. Most importantly, as compared to specific COX-2 and LOX-5 inhibitors, benfotiamine significantly prevented the LPS-induced macrophage death and monocytes adhesion to endothelial cells. Thus, our studies indicate that the dual regulation of COX and LOX pathways in AA metabolism could be a novel mechanism by which benfotiamine exhibits its potential anti-inflammatory response. PMID:22067901

  17. Dysregulation of protein degradation pathways may mediate the liver injury and phospholipidosis associated with a cationic amphiphilic antibiotic drug

    International Nuclear Information System (INIS)

    Mosedale, Merrie; Wu, Hong; Kurtz, C. Lisa; Schmidt, Stephen P.; Adkins, Karissa; Harrill, Alison H.

    2014-01-01

    A large number of antibiotics are known to cause drug-induced liver injury in the clinic; however, interpreting clinical risk is not straightforward owing to a lack of predictivity of the toxicity by standard preclinical species and a poor understanding of the mechanisms of toxicity. An example is PF-04287881, a novel ketolide antibiotic that caused elevations in liver function tests in Phase I clinical studies. In this study, a mouse diversity panel (MDP), comprised of 34 genetically diverse, inbred mouse strains, was utilized to model the toxicity observed with PF-04287881 treatment and investigate potential mechanisms that may mediate the liver response. Significant elevations in serum alanine aminotransferase (ALT) levels in PF-04287881-treated animals relative to vehicle-treated controls were observed in the majority (88%) of strains tested following a seven day exposure. The average fold elevation in ALT varied by genetic background and correlated with microscopic findings of hepatocellular hypertrophy, hepatocellular single cell necrosis, and Kupffer cell vacuolation (confirmed as phospholipidosis) in the liver. Global liver mRNA expression was evaluated in a subset of four strains to identify transcript and pathway differences that distinguish susceptible mice from resistant mice in the context of PF-04287881 treatment. The protein ubiquitination pathway was highly enriched among genes associated with PF-04287881-induced hepatocellular necrosis. Expression changes associated with PF-04287881-induced phospholipidosis included genes involved in drug transport, phospholipid metabolism, and lysosomal function. The findings suggest that perturbations in genes involved in protein degradation leading to accumulation of oxidized proteins may mediate the liver injury induced by this drug. - Highlights: • Identified susceptible and resistant mouse strains to liver injury induced by a CAD • Liver injury characterized by single cell necrosis, and phospholipidosis

  18. Dysregulation of protein degradation pathways may mediate the liver injury and phospholipidosis associated with a cationic amphiphilic antibiotic drug

    Energy Technology Data Exchange (ETDEWEB)

    Mosedale, Merrie [Hamner-University of North Carolina Institute for Drug Safety Sciences, The Hamner Institutes for Health Sciences, Research Triangle Park, NC 27709 (United States); Wu, Hong [Drug Safety Research and Development, Pfizer Global Research and Development, Groton, CT06340 (United States); Kurtz, C. Lisa [Hamner-University of North Carolina Institute for Drug Safety Sciences, The Hamner Institutes for Health Sciences, Research Triangle Park, NC 27709 (United States); Schmidt, Stephen P. [Drug Safety Research and Development, Pfizer Global Research and Development, Groton, CT06340 (United States); Adkins, Karissa, E-mail: Karissa.Adkins@pfizer.com [Drug Safety Research and Development, Pfizer Global Research and Development, Groton, CT06340 (United States); Harrill, Alison H. [Hamner-University of North Carolina Institute for Drug Safety Sciences, The Hamner Institutes for Health Sciences, Research Triangle Park, NC 27709 (United States); University of Arkansas for Medical Sciences, Little Rock, AR72205 (United States)

    2014-10-01

    A large number of antibiotics are known to cause drug-induced liver injury in the clinic; however, interpreting clinical risk is not straightforward owing to a lack of predictivity of the toxicity by standard preclinical species and a poor understanding of the mechanisms of toxicity. An example is PF-04287881, a novel ketolide antibiotic that caused elevations in liver function tests in Phase I clinical studies. In this study, a mouse diversity panel (MDP), comprised of 34 genetically diverse, inbred mouse strains, was utilized to model the toxicity observed with PF-04287881 treatment and investigate potential mechanisms that may mediate the liver response. Significant elevations in serum alanine aminotransferase (ALT) levels in PF-04287881-treated animals relative to vehicle-treated controls were observed in the majority (88%) of strains tested following a seven day exposure. The average fold elevation in ALT varied by genetic background and correlated with microscopic findings of hepatocellular hypertrophy, hepatocellular single cell necrosis, and Kupffer cell vacuolation (confirmed as phospholipidosis) in the liver. Global liver mRNA expression was evaluated in a subset of four strains to identify transcript and pathway differences that distinguish susceptible mice from resistant mice in the context of PF-04287881 treatment. The protein ubiquitination pathway was highly enriched among genes associated with PF-04287881-induced hepatocellular necrosis. Expression changes associated with PF-04287881-induced phospholipidosis included genes involved in drug transport, phospholipid metabolism, and lysosomal function. The findings suggest that perturbations in genes involved in protein degradation leading to accumulation of oxidized proteins may mediate the liver injury induced by this drug. - Highlights: • Identified susceptible and resistant mouse strains to liver injury induced by a CAD • Liver injury characterized by single cell necrosis, and phospholipidosis

  19. Bone Morphogenetic Proteins and myostatin pathways: key mediator of human sarcopenia.

    Science.gov (United States)

    Scimeca, Manuel; Piccirilli, Eleonora; Mastrangeli, Francesca; Rao, Cecilia; Feola, Maurizio; Orlandi, Augusto; Gasbarra, Elena; Bonanno, Elena; Tarantino, Umberto

    2017-02-15

    Sarcopenia, osteoporosis and osteoarthritis are the most frequent musculoskeletal disorders affecting older people. The main aim of this study was to test the hypothesis that the balance between BMPs and myostatin pathways regulates the age-related muscle degeneration in OP and OA patients. To this end, we investigated the relationship among the expression of BMP-2/4-7, myostatin and phosphorylated Smads1-5-8 and the muscle quality, evaluated in term of fibers atrophy and satellite cells activity. In this retrospective study, we collected 123 biopsies of vastus lateralis: 48 biopsies from patients who underwent hip arthroplasty for subcapital fractures of the femur (OP), 55 biopsies from patients who underwent hip arthroplasty for osteoarthritis (OA) and 20 biopsies from patients who underwent hip arthroplasty for high-energy hip fractures (CTRL). Muscle biopsies were fixed in 4% paraformaldehyde and paraffin embedded. Serial sections were used for morphometrical and immunohistochemical analysis (BMP/2/4-7, myostatin, Smads1-5-8, Pax7 and myogenin). In addition, 1 mm 3 of muscle tissue of each patient was embedded in epon for ultrastructural study. Morphometric data indicated an increase of the number of atrophic fibers in OP patients compare to OA. In line with these data, we found an high regenerative potential in muscle tissues of OA patients due to the significant amount of both Pax7 and myogenin positive satellite cells detected in OA group. In addition, our data showed the decrease of BMP2/4 and -7 expression in OP patients compared to both OA group and CTRL. Conversely, OP patients were characterized by high levels of myostatin expression. A different expression profile was also found for phosphorylated Smad1-5-8 between OP and OA patients. In particular, OP patients showed a low number of positive phosphorylated Smad1-5-8 nuclei. The identification of molecular pathways involved in the pathogenesis of sarcopenia open new prospective for the development of

  20. Meta-analysis of global transcriptomics reveals conserved genetic pathways of Quercetin and Tannic acid mediated longevity in C. elegans

    Directory of Open Access Journals (Sweden)

    Kerstin ePietsch

    2012-04-01

    Full Text Available Recent research has highlighted that the polyphenols Quercetin and Tannic acid are capable of extending the lifespan of C. elegans. To gain a deep understanding of the underlying molecular genetics, we analyzed the global transcriptional patterns of nematodes exposed to Quercetin or Tannic acid concentrations that are non-effective (in lifespan extension, lifespan extending or toxic. By means of an intricate meta-analysis it was possible to compare the transcriptomes of polyphenol exposure to recently published data sets derived from i longevity mutants or ii infection. This detailed comparative in silico analysis facilitated the identification of compound specific and overlapping transcriptional profiles and allowed the formulation of mechanistic models of Quercetin and Tannic acid mediated longevity. Lifespan extension due to Quercetin was predominantly driven by the metabolome, TGF-beta signaling, Insulin-like signaling and the p38 MAPK pathway and Tannic acid’s impact involved, in part, the amino acid metabolism and was modulated by the TGF-beta and the p38 MAPK pathways. DAF-12, which integrates TGF-beta and Insulin-like downstream signaling, therefore seems to be a crucial regulator for both polyphenols.

  1. Rhein Elicits In Vitro Cytotoxicity in Primary Human Liver HL-7702 Cells by Inducing Apoptosis through Mitochondria-Mediated Pathway

    Directory of Open Access Journals (Sweden)

    Guy-Armel Bounda

    2015-01-01

    Full Text Available Objective. To study rhein-induced apoptosis signaling pathway and to investigate its molecular mechanisms in primary human hepatic cells. Results. Cell viability of HL-7702 cells treated with rhein showed significant decrease in dose-dependent manner. Following rhein treatment (25 μM, 50 μM, and 100 μM for 12 h, the detection of apoptotic cells was significantly analyzed by flow cytometry and nuclear morphological changes by Hoechst 33258, respectively. Fatty degeneration studies showed upregulation level of the relevant hepatic markers (P < 0.01. Caspase activities expressed significant upregulation of caspase-3, caspase-9, and caspase-8. Moreover, apoptotic cells by rhein were significantly inhibited by Z-LEHD-FMK and Z-DEVD-FMK, caspase-9 inhibitor, and caspase-3 inhibitor, respectively. Overproduction of reactive oxygen species, lipid peroxidation, and loss of mitochondrial membrane potential were detected by fluorometry. Additionally, NAC, a ROS scavenger, significantly attenuated rhein-induced oxidative damage in HL-7702 cells. Furthermore, real-time qPCR results showed significant upregulation of p53, PUMA, Apaf-1, and Casp-9 and Casp-3 mRNA, with no significant changes of Fas and Cytochrome-c. Immunoblotting revealed significant Cytochrome-c release from mitochondria into cytosol and no change in Fas expression. Conclusion. Taken together, these observations suggested that rhein could induce apoptosis in HL-7702 cells via mitochondria-mediated signal pathway with involvement of oxidative stress mechanism.

  2. Regulation of apoptosis by resveratrol through JAK/STAT and mitochondria mediated pathway in human epidermoid carcinoma A431 cells

    International Nuclear Information System (INIS)

    Madan, Esha; Prasad, Sahdeo; Roy, Preeti; George, Jasmine; Shukla, Yogeshwer

    2008-01-01

    Resveratrol (trans-3,4',5-trihydroxystilbene), a polyphenolic phytoalexin present mainly in grapes, red wine and berries, is known to possess strong chemopreventive and anticancer properties. Here, we demonstrated the anti-proliferative and apoptosis-inducing activities of resveratrol in human epidermoid carcinoma A431 cells. Resveratrol has cytotoxic effects through inhibiting cellular proliferation of A431 cells, which leads to the induction of apoptosis, as evident by an increase in the fraction of cells in the sub-G 1 phase of the cell cycle and Annexin-V binding of externalized phosphatidylserine. Results revealed that inhibition of proliferation is associated with regulation of the JAK/STAT pathway, where resveratrol prevents phosphorylation of JAK, thereby inhibiting STAT1 phosphorylation. Furthermore, resveratrol treatment actively stimulated reactive oxygen species (ROS) and mitochondrial membrane depolarization. Consequently, an imbalance in the Bax/Bcl-2 ratio triggered the caspase cascade and subsequent cleavage of PARP, thereby shifting the balance in favor of apoptosis. These observations indicate that resveratrol treatment inhibits JAK/STAT-mediated gene transcription and induce the mitochondrial cell death pathway.

  3. The function of DREAM gene mediated by NF-kB signal pathway in the pathogenesis of osteosarcoma.

    Science.gov (United States)

    Qian, Jize; Lv, Mingbo; Zhang, Wenbo

    2017-01-01

    To explore the function of DREAM gene mediated by NF-kB signal pathway in the pathogenesis of osteosarcoma. This study included 13 Sprague Dawley (SD) rats with osteosarcoma (treatment group) and 13 healthy rats (control group). NF-kB, DREAM and P105 mRNAs expression levels were determined using quantitative PCR (qPCR). The expression levels of NF-kB, DREAM and P105 proteins were evaluated using ELISA and western blot. Also, DREAM protein expression in rats was determined by immunofluorescence. NF-kB and DREAM levels in the treatment group were significantly higher than those in the control group (pkB and DREAM expression levels in the treatment group were significantly higher than in the control group (pkB and DREAM levels in the treatment group were 4.3±0.12 μg/l and 6.8±0.21 μg/l, respectively. These levels in the control group were 0.96±0.11 μg/l and 1.25±0.18 μg/l, respectively. P105 expression level in the treatment group was 0.37±0.11 μg/l which was significantly lower than that in the control group (1.63±0.21 μg/l) (pkB signal pathway promoted the expression of DREAM gene and also promoted the pathogenesis and worsening of osteosarcoma.

  4. Deprivation of L-Arginine Induces Oxidative Stress Mediated Apoptosis in Leishmania donovani Promastigotes: Contribution of the Polyamine Pathway

    Science.gov (United States)

    Mandal, Abhishek; Das, Sushmita; Roy, Saptarshi; Ghosh, Ayan Kumar; Sardar, Abul Hasan; Verma, Sudha; Saini, Savita; Singh, Ruby; Abhishek, Kumar; Kumar, Ajay; Mandal, Chitra; Das, Pradeep

    2016-01-01

    The growth and survival of intracellular parasites depends on the availability of extracellular nutrients. Deprivation of nutrients viz glucose or amino acid alters redox balance in mammalian cells as well as some lower organisms. To further understand the relationship, the mechanistic role of L-arginine in regulation of redox mediated survival of Leishmania donovani promastigotes was investigated. L-arginine deprivation from the culture medium was found to inhibit cell growth, reduce proliferation and increase L-arginine uptake. Relative expression of enzymes, involved in L-arginine metabolism, which leads to polyamine and trypanothione biosynthesis, were downregulated causing decreased production of polyamines in L-arginine deprived parasites and cell death. The resultant increase in reactive oxygen species (ROS), due to L-arginine deprivation, correlated with increased NADP+/NADPH ratio, decreased superoxide dismutase (SOD) level, increased lipid peroxidation and reduced thiol content. A deficiency of L-arginine triggered phosphatidyl serine externalization, a change in mitochondrial membrane potential, release of intracellular calcium and cytochrome-c. This finally led to DNA damage in Leishmania promastigotes. In summary, the growth and survival of Leishmania depends on the availability of extracellular L-arginine. In its absence the parasite undergoes ROS mediated, caspase-independent apoptosis-like cell death. Therefore, L-arginine metabolism pathway could be a probable target for controlling the growth of Leishmania parasites and disease pathogenesis. PMID:26808657

  5. LKB1 mediates the development of conventional and innate T cells via AMP-dependent kinase autonomous pathways.

    Directory of Open Access Journals (Sweden)

    Marouan Zarrouk

    Full Text Available The present study has examined the role of the serine/threonine kinase LKB1 in the survival and differentiation of CD4/8 double positive thymocytes. LKB1-null DPs can respond to signals from the mature α/β T-cell-antigen receptor and initiate positive selection. However, in the absence of LKB1, thymocytes fail to mature to conventional single positive cells causing severe lymphopenia in the peripheral lymphoid tissues. LKB1 thus appears to be dispensable for positive selection but important for the maturation of positively selected thymocytes. LKB1 also strikingly prevented the development of invariant Vα14 NKT cells and innate TCR αβ gut lymphocytes. Previous studies with gain of function mutants have suggested that the role of LKB1 in T cell development is mediated by its substrate the AMP-activated protein kinase (AMPK. The present study now analyses the impact of AMPK deletion in DP thymocytes and shows that the role of LKB1 during the development of both conventional and innate T cells is mediated by AMPK-independent pathways.

  6. Intense picosecond pulsed electric fields induce apoptosis through a mitochondrial-mediated pathway in HeLa cells

    Science.gov (United States)

    HUA, YUAN-YUAN; WANG, XIAO-SHU; ZHANG, YU; YAO, CHEN-GUO; ZHANG, XI-MING; XIONG, ZHENG-AI

    2012-01-01

    The application of pulsed electric fields (PEF) is emerging as a new technique for tumor therapy. Picosecond pulsed electric fields (psPEF) can be transferred to target deep tissue non-invasively and precisely, but the research of the biological effects of psPEF on cells is limited. Electric theory predicts that intense psPEF will target mitochondria and lead to changes in transmembrane potential, therefore, it is hypothesized that it can induce mitochondrial-mediated apoptosis. HeLa cells were exposed to psPEF in this study to investigate this hypothesis. MTT assay demonstrated that intense psPEF significantly inhibited the proliferation of HeLa cells in a dose-dependent manner. Typical characteristics of apoptosis in HeLa cells were observed, using transmission electron microscopy. Loss of mitochondrial transmembrane potential was explored using laser scanning confocal microscopy with Rhodamine-123 (Rh123) staining. Furthermore, the mitochondrial apoptotic events were also confirmed by western blot analysis for the release of cytochrome C and apoptosis-inducing factor from mitochondria into the cytosol. In addition, activation of caspase-3, caspase-9, upregulation of Bax, p53 and downregulation of Bcl-2 were observed in HeLa cells also indicating apoptosis. Taken together, these results demonstrate that intense psPEF induce cell apoptosis through a mitochondrial-mediated pathway. PMID:22307872

  7. Short-term psychosocial stress protects photoreceptors from damage via corticosterone-mediated activation of the AKT pathway.

    Science.gov (United States)

    Forkwa, Tembei K; Neumann, Inga D; Tamm, Ernst R; Ohlmann, Andreas; Reber, Stefan O

    2014-02-01

    Apoptotic death of photoreceptors in hereditary retinal degenerations can be prevented by neuroprotective molecules. Here, we report that adrenal glucocorticoids (GC) released during psychosocial stress protect photoreceptors from apoptosis after light damage. Psychosocial stress is known to be the main type of stressor humans are exposed to and was induced here in mice by 10h of chronic subordinate colony housing (CSC). Photoreceptor damage was generated by subsequent exposure to white light. Short-term psychosocial stress prior to illumination significantly reduced the number of apoptotic photoreceptors, an effect that was absent in adrenalectomized (ADX) mice. The neuroprotective effect was completely restored in ADX mice substituted with GC. Moreover, phosphorylation of retinal AKT increased following CSC or exogenous GC treatment, an effect that was again absent in ADX mice exposed to CSC. Finally, inhibition of AKT signaling with triciribine blocked the stress- and GC-mediated neuroprotective effects on photoreceptors. In summary, we provide evidence that 1) short-term psychosocial stress protects photoreceptors from light-induced damage and 2) the protective effect is most likely mediated by GC-induced activation of the AKT signaling pathway. Copyright © 2013 Elsevier Inc. All rights reserved.

  8. Non-thermal plasma induces mitochondria-mediated apoptotic signaling pathway via ROS generation in HeLa cells.

    Science.gov (United States)

    Li, Wei; Yu, K N; Ma, Jie; Shen, Jie; Cheng, Cheng; Zhou, Fangjian; Cai, Zhiming; Han, Wei

    2017-11-01

    Non-thermal plasma (NTP) has been proposed as a novel therapeutic method for anticancer treatment. Although increasing evidence suggests that NTP selectively induces apoptosis in some types of tumor cells, the molecular mechanisms underlying this phenomenon remain unclear. In this study, we further investigated possible molecular mechanisms for NTP-induced apoptosis of HeLa cells. The results showed that NTP exposure significantly inhibited the growth and viability of HeLa cells. Morphological observation and flow cytometry analysis demonstrated that NTP exposure induced HeLa cell apoptosis. NTP exposure also activated caspase-9 and caspase-3, which subsequently cleaved poly (ADP- ribose) polymerase. Furthermore, NTP exposure suppressed Bcl-2 expression, enhanced Bax expression and translocation to mitochondria, activated mitochondria-mediated apoptotic pathway, followed by the release of cytochrome c. Further studies showed that NTP treatment led to ROS generation, whereas blockade of ROS generation by N-acetyl-l-cysteine (NAC, ROS scavengers) significantly prevented NTP-induced mitochondrial alteration and subsequent apoptosis of HeLa cells via suppressing Bax translocation, cytochrome c and caspase-3 activation. Taken together, our results indicated that NTP exposure induced mitochondria-mediated intrinsic apoptosis of HeLa cells was activated by ROS generation. These findings provide insights to the therapeutic potential and clinical research of NTP as a novel tool in cervical cancer treatment. Copyright © 2017. Published by Elsevier Inc.

  9. Transmitters and pathways mediating inhibition of spinal itch-signaling neurons by scratching and other counterstimuli.

    Directory of Open Access Journals (Sweden)

    Tasuku Akiyama

    Full Text Available Scratching relieves itch, but the underlying neural mechanisms are poorly understood. We presently investigated a role for the inhibitory neurotransmitters GABA and glycine in scratch-evoked inhibition of spinal itch-signaling neurons in a mouse model of chronic dry skin itch. Superficial dorsal horn neurons ipsilateral to hindpaw dry skin treatment exhibited a high level of spontaneous firing that was significantly attenuated by cutaneous scratching, pinch and noxious heat. Scratch-evoked inhibition was nearly abolished by spinal delivery of the glycine antagonist, strychnine, and was markedly attenuated by respective GABA(A and GABA(B antagonists bicuculline and saclofen. Scratch-evoked inhibition was also significantly attenuated (but not abolished by interruption of the upper cervical spinal cord, indicating the involvement of both segmental and suprasegmental circuits that engage glycine- and GABA-mediated inhibition of spinal itch-signaling neurons by noxious counterstimuli.

  10. Iodinated contrast media induce neutrophil apoptosis through a mitochondrial and caspase mediated pathway.

    LENUS (Irish Health Repository)

    Fanning, N F

    2012-02-03

    Iodinated contrast media (ICM) can induce apoptosis (programmed cell death) in renal, myocardial and endothelial cells. Following intravascular injection, circulating immune cells are exposed to high concentrations of ICM. As neutrophils constitutively undergo apoptosis we hypothesized that ICM may adversely affect neutrophil survival. Our aim was to investigate the effect of ICM on neutrophil apoptosis. Neutrophils were isolated from healthy subjects and cultured in vitro with ionic (diatrizoate and ioxaglate) and non-ionic (iohexol and iotrolan) ICM. The effect of ICM on neutrophil apoptosis in both unstimulated and lipopolysaccharide-stimulated neutrophils was determined by annexin V flow cytometry. The influence of physicochemical properties of the different ICM on apoptosis of neutrophils was also studied. We further investigated the effects of ICM on key intracellular signal pathways, including p38 mitogen-activated protein kinase (MAPK) by Western blotting, and mitochondrial depolarization and caspase activity by flow cytometry. Isoiodine concentrations (20 mg ml(-1)) of ionic (diatrizoate 69.6+\\/-2.9%; ioxaglate 58.9+\\/-2.0%) and non-ionic (iohexol 57.3+\\/-2.9%; iotrolan 57.1+\\/-2.6%) ICM significantly induced neutrophil apoptosis over control levels (47.7+\\/-1.4%). The apoptotic effect of ICM was influenced by their chemical structure, with ionic ICM having a more significant (p<0.01) apoptotic effect than non-ionic ICM (p<0.05). Furthermore, ICM reversed the anti-apoptotic effect of lipopolysaccharide (1000 ng ml(-1)) treated neutrophils to control levels (23.0+\\/-3.5% to 61.2+\\/-5.3%; n=4; p<0.05). These agents induce apoptosis through a p38 MAPK independent pathway that results in mitochondrial depolarization, and is dependent on caspase activation. As neutrophils play a central role in host response to infection and injury, ICM, through induction of neutrophil apoptosis, could have a significant deleterious effect on host immune defence and

  11. Electroacupuncture Exerts Neuroprotection through Caveolin-1 Mediated Molecular Pathway in Intracerebral Hemorrhage of Rats

    Directory of Open Access Journals (Sweden)

    Hui-Qin Li

    2016-01-01

    Full Text Available Spontaneous intracerebral hemorrhage (ICH is one of the most devastating types of stroke. Here, we aim to demonstrate that electroacupuncture on Baihui (GV20 exerts neuroprotection for acute ICH possibly via the caveolin-1/matrix metalloproteinase/blood-brain barrier permeability pathway. The model of ICH was established by using collagenase VII. Rats were randomly divided into three groups: Sham-operation group, Sham electroacupuncture group, and electroacupuncture group. Each group was further divided into 4 subgroups according to the time points of 6 h, 1 d, 3 d, and 7 d after ICH. The methods were used including examination of neurological deficit scores according to Longa’s scale, measurement of blood-brain barrier permeability through Evans Blue content, in situ immunofluorescent detection of caveolin-1 in brains, western blot analysis of caveolin-1 in brains, and in situ zymography for measuring matrix metalloproteinase-2/9 activity in brains. Compared with Sham electroacupuncture group, electroacupuncture group has resulted in a significant improvement in neurological deficit scores and in a reduction in Evans Blue content, expression of caveolin-1, and activity of matrix metalloproteinase-2/9 at 6 h, 1 d, 3 d, and 7 d after ICH (P<0.05. In conclusion, the present results suggested that electroacupuncture on GV20 can improve neurological deficit scores and reduce blood-brain barrier permeability after ICH, and the mechanism possibly targets caveolin-1/matrix metalloproteinase/blood-brain barrier permeability pathway.

  12. Gambogic acid inhibits multiple myeloma mediated osteoclastogenesis through suppression of chemokine receptor CXCR4 signaling pathways.

    Science.gov (United States)

    Pandey, Manoj K; Kale, Vijay P; Song, Chunhua; Sung, Shen-shu; Sharma, Arun K; Talamo, Giampaolo; Dovat, Sinisa; Amin, Shantu G

    2014-10-01

    Bone disease, characterized by the presence of lytic lesions and osteoporosis is the hallmark of multiple myeloma (MM). Stromal cell-derived factor 1α (SDF-1α) and its receptor, CXC chemokine receptor 4 (CXCR4), has been implicated as a regulator of bone resorption, suggesting that agents that can suppress SDF1α/CXCR4 signaling might inhibit osteoclastogenesis, a process closely linked to bone resorption. We, therefore, investigated whether gambogic acid (GA), a xanthone, could inhibit CXCR4 signaling and suppress osteoclastogenesis induced by MM cells. Through docking studies we predicted that GA directly interacts with CXCR4. This xanthone down-regulates the expression of CXCR4 on MM cells in a dose- and time-dependent manner. The down-regulation of CXCR4 was not due to proteolytic degradation, but rather GA suppresses CXCR4 mRNA expression by inhibiting nuclear factor-kappa B (NF-κB) DNA binding. This was further confirmed by quantitative chromatin immunoprecipitation assay, as GA inhibits p65 binding at the CXCR4 promoter. GA suppressed SDF-1α-induced chemotaxis of MM cells and downstream signaling of CXCR4 by inhibiting phosphorylation of Akt, p38, and Erk1/2 in MM cells. GA abrogated the RANKL-induced differentiation of macrophages to osteoclasts in a dose- and time-dependent manner. In addition, we found that MM cells induced differentiation of macrophages to osteoclasts, and that GA suppressed this process. Importantly, suppression of osteoclastogenesis by GA was mediated through IL-6 inhibition. Overall, our results show that GA is a novel inhibitor of CXCR4 expression and has a strong potential to suppress osteoclastogenesis mediated by MM cells. Published by Elsevier Inc.

  13. Nano-MnO2-mediated transformation of triclosan with humic molecules present: kinetics, products, and pathways.

    Science.gov (United States)

    Sun, Kai; Li, Shunyao; Waigi, Michael Gatheru; Huang, Qingguo

    2018-05-01

    It has been shown that manganese dioxide (MnO 2 ) can mediate transformation of phenolic contaminants to form phenoxyl radical intermediates, and subsequently, these intermediates intercouple to form oligomers via covalent binding. However, the reaction kinetics and transformation mechanisms of phenolic contaminants with humic molecules present in nano-MnO 2 -mediated systems were still unclear. In this study, it was proven that nano-MnO 2 were effective in transforming triclosan under acidic conditions (pH 3.5-5.0) during manganese reduction, and the apparent pseudo first-order kinetics rate constants (k = 0.0599-1.5314 h -1 ) increased as the pH decreased. In particular, the transformation of triclosan by nano-MnO 2 was enhanced in the presence of low-concentration humic acid (1-10 mg L -1 ). The variation in the absorption of humic molecules at 275 nm supported possible covalent binding between humic molecules and triclosan in the nano-MnO 2 -mediated systems. A total of four main intermediate products were identified by high-resolution mass spectrometry (HRMS), regardless of humic molecules present in the systems or not. These products correspond to a suite of radical intercoupling reactions (dimers and trimers), ether cleavage (2,4-dichlorophenol), and oxidation to quinone-like products, triggered by electron transfer from triclosan molecules to nano-MnO 2 . A possible reaction pathway in humic acid solutions, including homo-coupling, decomposition, oxidation, and cross-coupling, was proposed. Our findings provide valuable information regarding the environmental fate and transformation mechanism of triclosan by nano-MnO 2 in complex water matrices.

  14. NIK is required for NF-κB-mediated induction of BAG3 upon inhibition of constitutive protein degradation pathways.

    Science.gov (United States)

    Rapino, F; Abhari, B A; Jung, M; Fulda, S

    2015-03-12

    Recently, we reported that induction of the co-chaperone Bcl-2-associated athanogene 3 (BAG3) is critical for recovery of rhabdomyosarcoma (RMS) cells after proteotoxic stress upon inhibition of the two constitutive protein degradation pathways, that is, the ubiquitin-proteasome system by Bortezomib and the aggresome-autophagy system by histone deacetylase 6 (HDAC6) inhibitor ST80. In the present study, we investigated the molecular mechanisms mediating BAG3 induction under these conditions. Here, we identify nuclear factor-kappa B (NF-κB)-inducing kinase (NIK) as a key mediator of ST80/Bortezomib-stimulated NF-κB activation and transcriptional upregulation of BAG3. ST80/Bortezomib cotreatment upregulates mRNA and protein expression of NIK, which is accompanied by an initial increase in histone H3 acetylation. Importantly, NIK silencing by siRNA abolishes NF-κB activation and BAG3 induction by ST80/Bortezomib. Furthermore, ST80/Bortezomib cotreatment stimulates NF-κB transcriptional activity and upregulates NF-κB target genes. Genetic inhibition of NF-κB by overexpression of dominant-negative IκBα superrepressor (IκBα-SR) or by knockdown of p65 blocks the ST80/Bortezomib-stimulated upregulation of BAG3 mRNA and protein expression. Interestingly, inhibition of lysosomal activity by Bafilomycin A1 inhibits ST80/Bortezomib-stimulated IκBα degradation, NF-κB activation and BAG3 upregulation, indicating that IκBα is degraded via the lysosome in the presence of Bortezomib. Thus, by demonstrating a critical role of NIK in mediating NF-κB activation and BAG3 induction upon ST80/Bortezomib cotreatment, our study provides novel insights into mechanisms of resistance to proteotoxic stress in RMS.

  15. The anti-epidermal growth factor receptor (EGFR) monoclonal antibody, C225, enhances radiation-induced apoptosis in primary glioma cell lines through mediation of MAPK/JNK/p38 signaling pathways

    International Nuclear Information System (INIS)

    Chakravarti, A.; Noll, E.; Black, P.M.; Loeffler, J.S.

    2001-01-01

    after C225 + RT compared to lines treated by RT alone (D0=2.0 vs. 2.8, respectively). The other 5 of 12 glioma cell lines demonstrated resistance to C225-mediated radiosensitization. Treatment of these 5 cell lines by combined RT + C225 failed to result in any enhancement of apoptosis or reduction in clonogenic survival compared to RT alone. C225 demonstrated distinct effects on the signaling pathways of responsive vs. resistant glioma cell lines. All 12 glioma cell lines demonstrated reduced levels of phospho-EGFR, demonstrating antagonism of EGFR by C225. JNK and p38 activation states, as measured by quantitative Western analysis using phospho-specific antibodies, were enhanced by 2.5-3.0 fold in the 7 responsive cell lines after RT + C225, compared to no measurable induction in the 5 resistant lines. The absence of JNK/p38 activation in the 5 resistant lines was reflected by a concomitant absence of phospho-MKK4 and MKK 3/6 induction after RT + C225, implying the presence of signaling deficits further upstream of JNK/p38 not directly regulated by EGFR activity. MAPK activity (p44/p42) was reduced 1.5-2.0 fold in the responsive lines compared to a more modest 0.5 fold in the resistant lines. Conclusion: C225 appears to enhance radiation response of a distinct subset of primary glioma cell lines through activation of JNK/p38 pathways and downregulation of signaling through MAPK. However, EGFR-expressing glioma cell lines resistant to C225-mediated enhancement of radiation-induced apoptosis demonstrated MAPK/JNK/p38 signaling that was not as directly impacted by inhibition of EGFR. It is critical to better understand the upstream signals mediating activation of these pathways to develop more effective strategies for enhancing radiation response

  16. Cdk1, PKCδ and calcineurin-mediated Drp1 pathway contributes to mitochondrial fission-induced cardiomyocyte death

    International Nuclear Information System (INIS)

    Zaja, Ivan; Bai, Xiaowen; Liu, Yanan; Kikuchi, Chika; Dosenovic, Svjetlana; Yan, Yasheng; Canfield, Scott G.; Bosnjak, Zeljko J.

    2014-01-01

    Highlights: • Drp1-mediated increased mitochondrial fission but not fusion is involved the cardiomyocyte death during anoxia-reoxygenation injury. • Reactive oxygen species are upstream initiators of mitochondrial fission. • Increased mitochondrial fission is resulted from Cdk1-, PKCδ-, and calcineurin-mediated Drp1 pathways. - Abstract: Myocardial ischemia–reperfusion (I/R) injury is one of the leading causes of death and disability worldwide. Mitochondrial fission has been shown to be involved in cardiomyocyte death. However, molecular machinery involved in mitochondrial fission during I/R injury has not yet been completely understood. In this study we aimed to investigate molecular mechanisms of controlling activation of dynamin-related protein 1 (Drp1, a key protein in mitochondrial fission) during anoxia-reoxygenation (A/R) injury of HL1 cardiomyocytes. A/R injury induced cardiomyocyte death accompanied by the increases of mitochondrial fission, reactive oxygen species (ROS) production and activated Drp1 (pSer616 Drp1), and decrease of inactivated Drp1 (pSer637 Drp1) while mitochondrial fusion protein levels were not significantly changed. Blocking Drp1 activity with mitochondrial division inhibitor mdivi1 attenuated cell death, mitochondrial fission, and Drp1 activation after A/R. Trolox, a ROS scavenger, decreased pSer616 Drp1 level and mitochondrial fission after A/R. Immunoprecipitation assay further indicates that cyclin dependent kinase 1 (Cdk1) and protein kinase C isoform delta (PKCδ) bind Drp1, thus increasing mitochondrial fission. Inhibiting Cdk1 and PKCδ attenuated the increases in pSer616 Drp1, mitochondrial fission, and cardiomyocyte death. FK506, a calcineurin inhibitor, blocked the decrease in expression of inactivated pSer637 Drp1 and mitochondrial fission. Our findings reveal the following novel molecular mechanisms controlling mitochondrial fission during A/R injury of cardiomyocytes: (1) ROS are upstream initiators of

  17. Cdk1, PKCδ and calcineurin-mediated Drp1 pathway contributes to mitochondrial fission-induced cardiomyocyte death

    Energy Technology Data Exchange (ETDEWEB)

    Zaja, Ivan [Department of Anesthesiology, Medical College of Wisconsin, Milwaukee, WI 53226 (United States); Bai, Xiaowen, E-mail: xibai@mcw.edu [Department of Anesthesiology, Medical College of Wisconsin, Milwaukee, WI 53226 (United States); Liu, Yanan; Kikuchi, Chika; Dosenovic, Svjetlana; Yan, Yasheng; Canfield, Scott G. [Department of Anesthesiology, Medical College of Wisconsin, Milwaukee, WI 53226 (United States); Bosnjak, Zeljko J. [Department of Anesthesiology, Medical College of Wisconsin, Milwaukee, WI 53226 (United States); Department of Physiology, Medical College of Wisconsin, Milwaukee, WI 53226 (United States)

    2014-10-31

    Highlights: • Drp1-mediated increased mitochondrial fission but not fusion is involved the cardiomyocyte death during anoxia-reoxygenation injury. • Reactive oxygen species are upstream initiators of mitochondrial fission. • Increased mitochondrial fission is resulted from Cdk1-, PKCδ-, and calcineurin-mediated Drp1 pathways. - Abstract: Myocardial ischemia–reperfusion (I/R) injury is one of the leading causes of death and disability worldwide. Mitochondrial fission has been shown to be involved in cardiomyocyte death. However, molecular machinery involved in mitochondrial fission during I/R injury has not yet been completely understood. In this study we aimed to investigate molecular mechanisms of controlling activation of dynamin-related protein 1 (Drp1, a key protein in mitochondrial fission) during anoxia-reoxygenation (A/R) injury of HL1 cardiomyocytes. A/R injury induced cardiomyocyte death accompanied by the increases of mitochondrial fission, reactive oxygen species (ROS) production and activated Drp1 (pSer616 Drp1), and decrease of inactivated Drp1 (pSer637 Drp1) while mitochondrial fusion protein levels were not significantly changed. Blocking Drp1 activity with mitochondrial division inhibitor mdivi1 attenuated cell death, mitochondrial fission, and Drp1 activation after A/R. Trolox, a ROS scavenger, decreased pSer616 Drp1 level and mitochondrial fission after A/R. Immunoprecipitation assay further indicates that cyclin dependent kinase 1 (Cdk1) and protein kinase C isoform delta (PKCδ) bind Drp1, thus increasing mitochondrial fission. Inhibiting Cdk1 and PKCδ attenuated the increases in pSer616 Drp1, mitochondrial fission, and cardiomyocyte death. FK506, a calcineurin inhibitor, blocked the decrease in expression of inactivated pSer637 Drp1 and mitochondrial fission. Our findings reveal the following novel molecular mechanisms controlling mitochondrial fission during A/R injury of cardiomyocytes: (1) ROS are upstream initiators of

  18. Proximate Mediators of Microvascular Dysfunction at the Blood-Brain Barrier: Neuroinflammatory Pathways to Neurodegeneration

    Directory of Open Access Journals (Sweden)

    Barry W. Festoff

    2017-01-01

    Full Text Available Current projections are that by 2050 the numbers of people aged 65 and older with Alzheimer’s disease (AD in the US may increase threefold while dementia is projected to double every 20 years reaching ~115 million by 2050. AD is clinically characterized by progressive dementia and neuropathologically by neuronal and synapse loss, accumulation of amyloid plaques, and neurofibrillary tangles (NFTs in specific brain regions. The preclinical or presymptomatic stage of AD-related brain changes may begin over 20 years before symptoms occur, making development of noninvasive biomarkers essential. Distinct from neuroimaging and cerebrospinal fluid biomarkers, plasma or serum biomarkers can be analyzed to assess (i the presence/absence of AD, (ii the risk of developing AD, (iii the progression of AD, or (iv AD response to treatment. No unifying theory fully explains the neurodegenerative brain lesions but neuroinflammation (a lethal stressor for healthy neurons is universally present. Current consensus is that the earlier the diagnosis, the better the chance to develop treatments that influence disease progression. In this article we provide a detailed review and analysis of the role of the blood-brain barrier (BBB and damage-associated molecular patterns (DAMPs as well as coagulation molecules in the onset and progression of these neurodegenerative disorders.

  19. Multiple repair pathways mediate cellular tolerance to resveratrol-induced DNA damage.

    Science.gov (United States)

    Liu, Ying; Wu, Xiaohua; Hu, Xiaoqing; Chen, Ziyuan; Liu, Hao; Takeda, Shunichi; Qing, Yong

    2017-08-01

    Resveratrol (RSV) has been reported to exert health benefits for the prevention and treatment of many diseases, including cancer. The anticancer mechanisms of RSV seem to be complex and may be associated with genotoxic potential. To better understand the genotoxic mechanisms, we used wild-type (WT) and a panel of isogenic DNA-repair deficient DT40 cell lines to identify the DNA damage effects and molecular mechanisms of cellular tolerance to RSV. Our results showed that RSV induced significant formation of γ-H2AX foci and chromosome aberrations (CAs) in WT cells, suggesting direct DNA damage effects. Comparing the survival of WT with isogenic DNA-repair deficient DT40 cell lines demonstrated that single strand break repair (SSBR) deficient cell lines of Parp1 -/- , base excision repair (BER) deficient cell lines of Polβ -/- , homologous recombination (HR) mutants of Brca1 -/- and Brca2 -/- and translesion DNA synthesis (TLS) mutants of Rev3 -/- and Rad18 -/- were more sensitive to RSV. The sensitivities of cells were associated with enhanced DNA damage comparing the accumulation of γ-H2AX foci and number of CAs of isogenic DNA-repair deficient DT40 cell lines with WT cells. These results clearly demonstrated that RSV-induced DNA damage in DT40 cells, and multiple repair pathways including BER, SSBR, HR and TLS, play critical roles in response to RSV- induced genotoxicity. Copyright © 2017. Published by Elsevier Ltd.

  20. Autism as early neurodevelopmental disorder: evidence for an sAPPα-mediated anabolic pathway

    Directory of Open Access Journals (Sweden)

    Debomoy K Lahiri

    2013-06-01

    Full Text Available Autism is a neurodevelopmental disorder marked by social skills and communication deficits and interfering repetitive behavior. Intellectual disability often accompanies autism. In addition to behavioral deficits, autism is characterized by neuropathology and brain overgrowth. Increased intracranial volume often accompanies this brain growth. We have found that the Alzheimer’s disease (AD associated amyloid-β precursor protein (APP, especially its neuroprotective processing product, secreted APP α (sAPPα, is elevated in persons with autism. This has led to the anabolic hypothesis of autism etiology, in which neuronal overgrowth in the brain results in interneuronal misconnections that may underlie multiple autism symptoms. We review the contribution of research in brain volume and of APP to the anabolic hypothesis, and relate APP to other proteins and pathways that have already been directly associated with autism, such as fragile X mental retardation protein (FMRP, Ras small GTPase/Extracellular Signal-Regulated Kinase (Ras/ERK, and phosphoinositide 3 kinase/protein kinase B/mammalian target of rapamycin (PI3K/Akt/mTOR. We also present additional evidence of MRI intracranial measurements in favor of the anabolic hypothesis. Finally, since it appears that APP’s involvement in autism is part of a multi-partner network, we extend this concept into the inherently interactive realm of epigenetics. We speculate that the underlying molecular abnormalities that influence APP’s contribution to autism are epigenetic markers overlaid onto potentially vulnerable gene sequences due to environmental influence.

  1. Autism as early neurodevelopmental disorder: evidence for an sAPPα-mediated anabolic pathway.

    Science.gov (United States)

    Lahiri, Debomoy K; Sokol, Deborah K; Erickson, Craig; Ray, Balmiki; Ho, Chang Y; Maloney, Bryan

    2013-01-01

    Autism is a neurodevelopmental disorder marked by social skills and communication deficits and interfering repetitive behavior. Intellectual disability often accompanies autism. In addition to behavioral deficits, autism is characterized by neuropathology and brain overgrowth. Increased intracranial volume often accompanies this brain growth. We have found that the Alzheimer's disease (AD) associated amyloid-β precursor protein (APP), especially its neuroprotective processing product, secreted APP α, is elevated in persons with autism. This has led to the "anabolic hypothesis" of autism etiology, in which neuronal overgrowth in the brain results in interneuronal misconnections that may underlie multiple autism symptoms. We review the contribution of research in brain volume and of APP to the anabolic hypothesis, and relate APP to other proteins and pathways that have already been directly associated with autism, such as fragile X mental retardation protein, Ras small GTPase/extracellular signal-regulated kinase, and phosphoinositide 3 kinase/protein kinase B/mammalian target of rapamycin. We also present additional evidence of magnetic resonance imaging intracranial measurements in favor of the anabolic hypothesis. Finally, since it appears that APP's involvement in autism is part of a multi-partner network, we extend this concept into the inherently interactive realm of epigenetics. We speculate that the underlying molecular abnormalities that influence APP's contribution to autism are epigenetic markers overlaid onto potentially vulnerable gene sequences due to environmental influence.

  2. The Protective Effect of Gangliosides on Lead (Pb)-Induced Neurotoxicity Is Mediated by Autophagic Pathways.

    Science.gov (United States)

    Meng, Hongtao; Wang, Lan; He, Junhong; Wang, Zhufeng

    2016-03-25

    Lead (Pb) is a ubiquitous environmental and industrial pollutant and can affect intelligence development and the learning ability and memory of children. Therefore, necessary measures should be taken to protect the central nervous system (CNS) from Pb toxicity. Gangliosides are sialic acid-containing glycosphingolipids that are constituents of mammalian cell membranes and are more abundantly expressed in the CNS. Studies have shown that gangliosides constitute a useful tool in the attempt to promote functional recovery of CNS and can reverse Pb-induced impairments of synaptic plasticity in rats. However, the detailed mechanisms have yet to be fully understood. In our present study, we tried to investigate the role of gangliosides in Pb-induced injury in hippocampus neurons and to further confirm the detailed mechanism. Our results show that Pb-induced injuries in the spatial reference memory were associated with a reduction of cell viability and cell apoptosis, and treatment with gangliosides markedly ameliorated the Pb-induced injury by inhibition of apoptosis action. Gangliosides further attenuated Pb-induced the abnormal autophagic process by regulation of mTOR pathways. In summary, our study establishes the efficacy of gangliosides as neuroprotective agents and provides a strong rationale for further studies on the underlying mechanisms of their neuroprotective functions.

  3. The Protective Effect of Gangliosides on Lead (Pb-Induced Neurotoxicity Is Mediated by Autophagic Pathways

    Directory of Open Access Journals (Sweden)

    Hongtao Meng

    2016-03-01

    Full Text Available Lead (Pb is a ubiquitous environmental and industrial pollutant and can affect intelligence development and the learning ability and memory of children. Therefore, necessary measures should be taken to protect the central nervous system (CNS from Pb toxicity. Gangliosides are sialic acid-containing glycosphingolipids that are constituents of mammalian cell membranes and are more abundantly expressed in the CNS. Studies have shown that gangliosides constitute a useful tool in the attempt to promote functional recovery of CNS and can reverse Pb-induced impairments of synaptic plasticity in rats. However, the detailed mechanisms have yet to be fully understood. In our present study, we tried to investigate the role of gangliosides in Pb-induced injury in hippocampus neurons and to further confirm the detailed mechanism. Our results show that Pb-induced injuries in the spatial reference memory were associated with a reduction of cell viability and cell apoptosis, and treatment with gangliosides markedly ameliorated the Pb-induced injury by inhibition of apoptosis action. Gangliosides further attenuated Pb-induced the abnormal autophagic process by regulation of mTOR pathways. In summary, our study establishes the efficacy of gangliosides as neuroprotective agents and provides a strong rationale for further studies on the underlying mechanisms of their neuroprotective functions.

  4. Mechanism of Anti-glioblastoma Effect of Temzolomide Involved in ROS-Mediated SIRT 1 Pathway

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    Yuan Jiang

    2014-03-01

    Full Text Available Objective: To explore the new molecular mechanism of anti-tumor effect of temzolomide (TMZon glioblastoma cell strain. Methods: MTT methods and Hoechst 33342 staining method were applied to determine the effect of TMZ on the proliferation and apoptosis of glioblastoma cell strains U251 and SHG44, while flow cytometry was used to detect the impact of TMZ on cellular cycles. Additionally, DCFH-DA probe was adopted to test intracellular reactive oxygen species (ROS level while Real-time PCR and Western blot tests were applied to determine the influence of TMZ on SIRT1 expression. Results: TMZ in different concentrations added into glioblastoma cell strain for 72 h could concentration-dependently inhibit the proliferation of glioblastoma cells, 100 μmol/L of which could also block cells in phase G2/M and improve cellular apoptosis. In addition, TMZ could evidently increase intracellular ROS level so as to activate SIRT1. Conclusion: The mechanism of anti-tumor effect of TMZ on glioblastoma may be associated with ROS-induced SIRT1 pathway, providing theoretical basis for the clinical efficacy of TMZ.

  5. A "Bony" Proposition: Pathways Mediating Responses to Simulated Weightlessness and Radiation

    Science.gov (United States)

    Tahimic, Candice; Globus, Ruth

    2016-01-01

    There is evidence that weightlessness and radiation, two elements of the spaceflight environment, can lead to detrimental changes in human musculoskeletal tissue, including bone loss and muscle atrophy. This bone loss is thought to be brought about by the increased activity of bone-resorbing osteoclasts and functional changes in bone-forming osteoblasts, cells that give rise to mature osteocytes. My current area of research focuses on understanding the mechanistic basis for the responses of bone to the spaceflight environment using earth-based animal and cellular models. The overarching goal is to identify molecular targets to prevent bone loss in space exploration and earth-based scenarios of radiotherapy, accidental radiation exposure and reduced mobility. In this talk, I will highlight two signaling pathways that potentially play a role in the response of bone to spaceflight-like conditions. Firstly, I will discuss the role of insulin-like growth factor 1 (IGF1) signaling as it pertains to the recovery of bone from simulated weightlessness (rodent hindlimb unloading model). Secondly, I will share recent findings from our study that aims to understand the emerging role of autophagy in maintaining the balance between bone formation and resorption (bone homeostasis) as well as normal skeletal structure.

  6. Rac1 Guides Porf-2 to Wnt Pathway to Mediate Neural Stem Cell Proliferation

    Directory of Open Access Journals (Sweden)

    Xi-Tao Yang

    2017-06-01

    Full Text Available The molecular and cellular mechanisms underlying the anti-proliferative effects of preoptic regulator factor 2 (Porf-2 on neural stem cells (NSCs remain largely unknown. Here, we found that Porf-2 inhibits the activity of ras-related C3 botulinum toxin substrate 1 (Rac1 protein in hippocampus-derived rat NSCs. Reduced Rac1 activity impaired the nuclear translocation of β-catenin, ultimately causing a repression of NSCs proliferation. Porf-2 knockdown enhanced NSCs proliferation but not in the presence of small molecule inhibitors of Rac1 or Wnt. At the same time, the repression of NSCs proliferation caused by Porf-2 overexpression was counteracted by small molecule activators of Rac1 or Wnt. By using a rat optic nerve crush model, we observed that Porf-2 knockdown enhanced the recovery of visual function. In particular, optic nerve injury in rats led to increased Wnt family member 3a (Wnt3a protein expression, which we found responsible for enhancing Porf-2 knockdown-induced NSCs proliferation. These findings suggest that Porf-2 exerts its inhibitory effect on NSCs proliferation via Rac1-Wnt/β-catenin pathway. Porf-2 may therefore represent and interesting target for optic nerve injury recovery and therapy.

  7. Polysaccharide of Dendrobium huoshanense activates macrophages via toll-like receptor 4-mediated signaling pathways.

    Science.gov (United States)

    Xie, Song-Zi; Hao, Ran; Zha, Xue-Qiang; Pan, Li-Hua; Liu, Jian; Luo, Jian-Ping

    2016-08-01

    The present work aimed at investigating the pattern recognition receptor (PRR) and immunostimulatory mechanism of a purified Dendrobium huoshanense polysaccharide (DHP). We found that DHP could bind to the surface of macrophages and stimulate macrophages to secrete NO, TNF-α and IL-1β. To unravel the mechanism for the binding of DHP to macrophages, flow cytometry, confocal laser-scanning microscopy, affinity electrophoresis, SDS-PAGE and western blotting were employed to verify the type of PRR responsible for the recognition of DHP by RAW264.7 macrophages and peritoneal macrophages of C3H/HeN and C3H/HeJ macrophages. Results showed that toll-like receptor 4 (TLR4) was an essential receptor for macrophages to directly bind DHP. Further, the phosphorylation of ERK, JNK, Akt and p38 were observed to be time-dependently promoted by DHP, as well as the nuclear translocation of NF-κB p65. These results suggest that DHP activates macrophages via its direct binding to TLR4 to trigger TLR4 signaling pathways. Copyright © 2016 Elsevier Ltd. All rights reserved.

  8. Exosome proteomics reveals transcriptional regulator proteins with potential to mediate downstream pathways.

    Science.gov (United States)

    Ung, Timothy H; Madsen, Helen J; Hellwinkel, Justin E; Lencioni, Alex M; Graner, Michael W

    2014-11-01

    Exosomes are virus-sized, membrane-enclosed vesicles with origins in the cellular endosomal system, but are released extracellularly. As a population, these tiny vesicles carry relatively enormous amounts of information in their protein, lipid and nucleic acid content, and the vesicles can have profound impacts on recipient cells. This review employs publically-available data combined with gene ontology applications to propose a novel concept, that exosomes transport transcriptional and translational machinery that may have direct impacts on gene expression in recipient cells. Here, we examine the previously published proteomic contents of medulloblastoma-derived exosomes, focusing on transcriptional regulators; we found that there are numerous proteins that may have potential roles in transcriptional and translational regulation with putative influence on downstream, cancer-related pathways. We expanded this search to all of the proteins in the Vesiclepedia database; using gene ontology approaches, we see that these regulatory factors are implicated in many of the processes involved in cancer initiation and progression. This information suggests that some of the effects of exosomes on recipient cells may be due to the delivery of protein factors that can directly and fundamentally change the transcriptional landscape of the cells. Within a tumor environment, this has potential to tilt the advantage towards the cancer. © 2014 The Authors. Cancer Science published by Wiley Publishing Asia Pty Ltd on behalf of Japanese Cancer Association.

  9. PKC α regulates netrin-1/UNC5B-mediated survival pathway in bladder cancer

    International Nuclear Information System (INIS)

    Liu, Jiao; Kong, Chui-ze; Gong, Da-xin; Zhang, Zhe; Zhu, Yu-yan

    2014-01-01

    Netrin-1 and its receptor UNC5B play important roles in angiogenesis, embryonic development, cancer and inflammation. However, their expression patttern and biological roles in bladder cancer have not been well characterized. The present study aims to investigating the clinical significance of PKC α, netrin-1 and UNC5B in bladder cancer as well as their association with malignant biological behavior of cancer cells. Netrin-1 and UNC5B expression was examined in 120 bladder cancer specimens using immunohistochemistry and in 40 fresh cancer tissues by western blot. Immunofluorescence was performed in cancer cell lines. PKC α agonist PMA and PKC siRNA was employed in bladder cancer cells. CCK-8, wound healing assays and flow cytometry analysis were used to examine cell proliferation, migration and cell cycle, respectively. Netrin-1 expression was positively correlated with histological grade, T stage, metastasis and poor prognosis in bladder cancer tissues. Immunofluorescence showed elevated netrin-1 and decreased UNC5B expression in bladder cancer cells compared with normal bladder cell line. Furthermore, cell proliferation, migration and cell cycle progression were promoted with PMA treatment while inhibited by calphostin C. In addition, PMA treatment could induce while calphostin C reduce netrin-1 expression in bladder cancer cells. The present study identified netrin-1/UNC5B, which could be regulated by PKC signaling, was important mediators of bladder cancer progression

  10. A mitochondria-dependent pathway mediates the apoptosis of GSE-induced yeast.

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    Sishuo Cao

    Full Text Available Grapefruit seed extract (GSE, which has powerful anti-fungal activity, can induce apoptosis in S. cerevisiae. The yeast cells underwent apoptosis as determined by testing for apoptotic markers of DNA cleavage and typical chromatin condensation by Terminal Deoxynucleotidyl Transferase-mediated dUTP Nick End Labeling (TUNEL and 4,6'-diaminidino-2-phenylindole (DAPI staining and electron microscopy. The changes of ΔΨmt (mitochondrial transmembrane potential and ROS (reactive oxygen species indicated that the mitochondria took part in the apoptotic process. Changes in this process detected by metabonomics and proteomics revealed that the yeast cells tenaciously resisted adversity. Proteins related to redox, cellular structure, membrane, energy and DNA repair were significantly increased. In this study, the relative changes in the levels of proteins and metabolites showed the tenacious resistance of yeast cells. However, GSE induced apoptosis in the yeast cells by destruction of the mitochondrial 60 S ribosomal protein, L14-A, and prevented the conversion of pantothenic acid to coenzyme A (CoA. The relationship between the proteins and metabolites was analyzed by orthogonal projections to latent structures (OPLS. We found that the changes of the metabolites and the protein changes had relevant consistency.

  11. Neuromodulatory Effect of Thymoquinone in Attenuating Glutamate-Mediated Neurotoxicity Targeting the Amyloidogenic and Apoptotic Pathways

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    Ibram Amin Fouad

    2018-04-01

    Full Text Available Overexposure of the glutamatergic N-methyl-d-aspartate (NMDA receptor to the excitatory neurotransmitter l-glutamic acid leads to neuronal cell death by excitotoxicity as a result of increased intracellular Ca2+, mitochondrial dysfunction, and apoptosis. Moreover, it was previously reported that prolonged activation of the NMDA receptor increased beta-amyloid (Aβ levels in the brain. Thymoquinone (TQ, the active constituent of Nigella sativa seeds, has been shown to have potent antioxidant and antiapoptotic effects. The aim of the present study was to explore the neuromodulatory effects of different doses of TQ (2.5 and 10 mg/kg against apoptotic cell death and Aβ formation resulting from glutamate administration in rats using vitamin E as a positive control. Behavioral changes were assessed using Y-maze and Morris water maze tests for evaluating spatial memory and cognitive functions. Caspase-3, Lactate dehydrogenase, Aβ-42, and cytochrome c gene expression were determined. TQ-treated groups showed significant decreases in the levels of all tested biochemical and behavioral parameters compared with the glutamate-treated group. These findings demonstrated that TQ has a promising neuroprotective activity against glutamate-induced neurotoxicity and this effect is mediated through its anti-amyloidogenic, antioxidant, and antiapoptotic activities.

  12. Immobilized laccase mediated dye decolorization and transformation pathway of azo dye acid red 27.

    Science.gov (United States)

    Chhabra, Meenu; Mishra, Saroj; Sreekrishnan, Trichur Ramaswamy

    2015-01-01

    Laccases have good potential as bioremediating agents and can be used continuously in the immobilized form like many other enzymes. In the present study, laccase from Cyathus bulleri was immobilized by entrapment in Poly Vinyl Alcohol (PVA) beads cross-linked with either nitrate or boric acid. Immobilized laccase was used for dye decolorization in both batch and continuous mode employing a packed bed column. The products of degradation of dye Acid Red 27 were identified by LC MS/MS analysis. The method led to very effective (90%) laccase immobilization and also imparted significant stability to the enzyme (more than 70% after 5 months of storage at 4°C). In batch decolorization, 90-95% decolorization was achieved of the simulated dye effluent for up to 10-20 cycles. Continuous decolorization in a packed bed bioreactor led to nearly 90% decolorization for up to 5 days. The immobilized laccase was also effective in decolorization and degradation of Acid Red 27 in the presence of a mediator. Four products of degradation were identified by LC-MS/MS analysis. The immobilized laccase in PVA-nitrate was concluded to be an effective agent in treatment of textile dye effluents.

  13. Propeptide-mediated inhibition of myostatin increases muscle mass through inhibiting proteolytic pathways in aged mice.

    Science.gov (United States)

    Collins-Hooper, Henry; Sartori, Roberta; Macharia, Raymond; Visanuvimol, Korntip; Foster, Keith; Matsakas, Antonios; Flasskamp, Hannah; Ray, Steve; Dash, Philip R; Sandri, Marco; Patel, Ketan

    2014-09-01

    Mammalian aging is accompanied by a progressive loss of skeletal muscle, a process called sarcopenia. Myostatin, a secreted member of the transforming growth factor-β family of signaling molecules, has been shown to be a potent inhibitor of muscle growth. Here, we examined whether muscle growth could be promoted in aged animals by antagonizing the activity of myostatin through the neutralizing activity of the myostatin propeptide. We show that a single injection of an AAV8 virus expressing the myostatin propeptide induced an increase in whole body weights and all muscles examined within 7 weeks of treatment. Our cellular studies demonstrate that muscle enlargement was due to selective fiber type hypertrophy, which was accompanied by a shift toward a glycolytic phenotype. Our molecular investigations elucidate the mechanism underpinning muscle hypertrophy by showing a decrease in the expression of key genes that control ubiquitin-mediated protein breakdown. Most importantly, we show that the hypertrophic muscle that develops as a consequence of myostatin propeptide in aged mice has normal contractile properties. We suggest that attenuating myostatin signaling could be a very attractive strategy to halt and possibly reverse age-related muscle loss. © The Author 2014. Published by Oxford University Press on behalf of The Gerontological Society of America. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.

  14. A mitochondria-dependent pathway mediates the apoptosis of GSE-induced yeast.

    Science.gov (United States)

    Cao, Sishuo; Xu, Wentao; Zhang, Nan; Wang, Yan; Luo, YunBo; He, Xiaoyun; Huang, Kunlun

    2012-01-01

    Grapefruit seed extract (GSE), which has powerful anti-fungal activity, can induce apoptosis in S. cerevisiae. The yeast cells underwent apoptosis as determined by testing for apoptotic markers of DNA cleavage and typical chromatin condensation by Terminal Deoxynucleotidyl Transferase-mediated dUTP Nick End Labeling (TUNEL) and 4,6'-diaminidino-2-phenylindole (DAPI) staining and electron microscopy. The changes of ΔΨmt (mitochondrial transmembrane potential) and ROS (reactive oxygen species) indicated that the mitochondria took part in the apoptotic process. Changes in this process detected by metabonomics and proteomics revealed that the yeast cells tenaciously resisted adversity. Proteins related to redox, cellular structure, membrane, energy and DNA repair were significantly increased. In this study, the relative changes in the levels of proteins and metabolites showed the tenacious resistance of yeast cells. However, GSE induced apoptosis in the yeast cells by destruction of the mitochondrial 60 S ribosomal protein, L14-A, and prevented the conversion of pantothenic acid to coenzyme A (CoA). The relationship between the proteins and metabolites was analyzed by orthogonal projections to latent structures (OPLS). We found that the changes of the metabolites and the protein changes had relevant consistency.

  15. Redundant Vasodilator Pathways Underlying Radial Artery Flow-Mediated Dilation Are Preserved in Healthy Aging

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    Kevin D. Ballard

    2014-01-01

    Full Text Available Background. Blocking nitric oxide (NO and vasodilator prostanoids (PN does not consistently reduce flow-mediated dilation (FMD in young adults. The impact of aging on the contribution of NO and PG to FMD is unknown. Methods. FMD was measured in older adults (n=10, 65±3 y after arterial infusion of saline, N(G-monomethyl-L-arginine (L-NMMA, and ketorolac + L-NMMA. Data were compared to published data in young adults. Results. L-NMMA reduced FMD in older adults (8.9±3.6 to 5.9±3.7% although this was not statistically significant (P=0.08 and did not differ (P=0.74 from the reduction observed in young adults (10.0±3.8 to 7.6±4.7%; P=0.03. Blocking PN did not affect FMD in young or older adults. In older adults, L-NMMA reduced (n=6; range = 36–123% decrease, augmented (n=3; 10–122% increase, or did not change FMD (n=1; 0.4% increase. After PN blockade, FMD responses were reduced (n=2, augmented (n=6, or unaffected (n=1. Conclusions. NO or PN blockade did not consistently reduce FMD in healthy older adults, suggesting the existence of redundant vasodilator phenotypes as observed previously in young adults.

  16. Nuclear translocation and regulation of intranuclear distribution of cytoplasmic poly(A-binding protein are distinct processes mediated by two Epstein Barr virus proteins.

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    Richard Park

    Full Text Available Many viruses target cytoplasmic polyA binding protein (PABPC to effect widespread inhibition of host gene expression, a process termed viral host-shutoff (vhs. During lytic replication of Epstein Barr Virus (EBV we observed that PABPC was efficiently translocated from the cytoplasm to the nucleus. Translocated PABPC was diffusely distributed but was excluded from viral replication compartments. Vhs during EBV infection is regulated by the viral alkaline nuclease, BGLF5. Transfection of BGLF5 alone into BGLF5-KO cells or uninfected 293 cells promoted translocation of PAPBC that was distributed in clumps in the nucleus. ZEBRA, a viral bZIP protein, performs essential functions in the lytic program of EBV, including activation or repression of downstream viral genes. ZEBRA is also an essential replication protein that binds to viral oriLyt and interacts with other viral replication proteins. We report that ZEBRA also functions as a regulator of vhs. ZEBRA translocated PABPC to the nucleus, controlled the intranuclear distribution of PABPC, and caused global shutoff of host gene expression. Transfection of ZEBRA alone into 293 cells caused nuclear translocation of PABPC in the majority of cells in which ZEBRA was expressed. Co-transfection of ZEBRA with BGLF5 into BGLF5-KO cells or uninfected 293 cells rescued the diffuse intranuclear pattern of PABPC seen during lytic replication. ZEBRA mutants defective for DNA-binding were capable of regulating the intranuclear distribution of PABPC, and caused PABPC to co-localize with ZEBRA. One ZEBRA mutant, Z(S186E, was deficient in translocation yet was capable of altering the intranuclear distribution of PABPC. Therefore ZEBRA-mediated nuclear translocation of PABPC and regulation of intranuclear PABPC distribution are distinct events. Using a click chemistry-based assay for new protein synthesis, we show that ZEBRA and BGLF5 each function as viral host shutoff factors.

  17. Induction of rat hepatic zinc thionein by phorbol ester-mediated protein kinase C pathway

    Energy Technology Data Exchange (ETDEWEB)

    Garrett, S.H.; Funk, A.E.; Brady, F.O.

    1986-05-01

    Metallothionein (MT) exists in rat liver mainly as a zinc protein. The levels of this protein fluctuate in response to a variety of internal and external stimuli. Among these inducers of MT are metals, glucocorticoids, catecholamines, and polypeptide hormones. Metals and glucocorticoids are primary inducers of MT, while the others operate either via adenylate cyclase/cAMP/cAMP-dependent protein kinase, or via phospholipase C/inositol 1,4,5-triphosphate, diacylglycerol/Ca/sup 2 +/-dependent protein kinase, protein kinase C. The authors have examined the role of the protein kinase C pathway in the induction of MT by using a phorbol ester, 12-O-tetradecanoyl-phorbol 13-acetate (TPA), to activate it. In vivo TPA is a good inducer of Zn/sub 7/-MT with an ED/sub 0.5/ of 26.5 nmoles/kg b.w. Maximal levels reached were about 7..mu..g Zn in MT/g liver, an induction increase of 8 to 10-fold. An inactive compound, 4..beta..-phorbol, and the vehicle (DMSO) did not stimulate the synthesis of Zn/sub 7/-MT. This induction by TPA requires de novo protein synthesis, as demonstrated by a cycloheximide/(/sup 35/S)-cysteine experiment. TPA stimulated Zn incorporation by 8.6-fold and (/sup 35/S)-cysteine incorporation by 4.8-fold during an 11h induction. These increases were blocked 100% by treatment with cycloheximide at -1 and +5h. These experiments have been repeated in cultured hepatocytes, using (/sup 35/S)-cysteine incorporation, slab SDS-PAGE, and autoradiography to quantitate MT levels.

  18. Gastrointestinal stimulatory and uterotonic activities of dietary radish leaves extract are mediated through multiple pathways.

    Science.gov (United States)

    Ghayur, Muhammad Nabeel; Gilani, Anwarul Hassan

    2005-09-01

    Raphanus sativus, commonly known as radish, is a food plant known worldwide for its culinary and medicinal properties especially as a laxative and abortifacient. This study reports the gastrointestinal and uterine tone modulatory activities of the crude extract (Rl.Cr) of radish leaves. Rl.Cr, showing the presence of saponins and alkaloids, exhibited a spasmogenic effect (0.03-10 mg/mL) in isolated rabbit jejunum, rat stomach fundus and uterus which was partially blocked by atropine. In contrast, Rl.Cr was found to be devoid of any stimulatory effect in rat ileum, instead showed an inhibitory effect (0.1 mg/mL) on the ACh dose-response curves. A mild relaxant effect was also observed in rabbit jejunum at the lower doses (0.1-0.3 mg/mL) but not against K(+)-induced contractions, ruling out a calcium channel blocking effect. In guinea-pig ileum, Rl.Cr exhibited a stimulant effect resistant to atropine while sensitive to pyrilamine pretreatment. The aqueous fraction, showing a strong presence of saponins, was found to be more efficacious than the non-polar fractions in its spasmogenic effect. This study shows the presence of species-dependent gastrointestinal effects of radish mediated partially through cholinergic receptors in rabbit and rat tissues, but through histaminergic activation in the guinea-pig, providing a scientific basis for its use in gut and uterine affections while also giving a wider picture of the activity profile of radish by using different species of animals. Copyright 2005 John Wiley & Sons, Ltd.

  19. Platelet proteome reveals novel pathways of platelet activation and platelet-mediated immunoregulation in dengue.

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    Monique Ramos de Oliveira Trugilho

    2017-05-01

    proteome in dengue, and sheds light on new mechanisms of platelet activation and platelet-mediated immune and inflammatory responses.

  20. Serotonin-induced vasodilatation in the human forearm is mediated by the "nitric oxide-pathway": no evidence for involvement of the 5-HT3-receptor

    NARCIS (Netherlands)

    Bruning, T. A.; Chang, P. C.; Blauw, G. J.; Vermeij, P.; van Zwieten, P. A.

    1993-01-01

    The "nitric oxide (NO)-pathway" is presumed to be involved in acetylcholine (ACh)- and serotonin (5-hydroxytryptamine, 5-HT)-mediated vasodilatation. In addition, both the 5-HT-induced transient and persistent vasodilator responses in the forearm vascular bed are abolished by the

  1. Two novel WD40 domain–containing proteins, Ere1 and Ere2, function in the retromer-mediated endosomal recycling pathway

    Science.gov (United States)

    Shi, Yufeng; Stefan, Christopher J.; Rue, Sarah M.; Teis, David; Emr, Scott D.

    2011-01-01

    Regulated secretion, nutrient uptake, and responses to extracellular signals depend on cell-surface proteins that are internalized and recycled back to the plasma membrane. However, the underlying mechanisms that govern membrane protein recycling to the cell surface are not fully known. Using a chemical-genetic screen in yeast, we show that the arginine transporter Can1 is recycled back to the cell surface via two independent pathways mediated by the sorting nexins Snx4/41/42 and the retromer complex, respectively. In addition, we identify two novel WD40-domain endosomal recycling proteins, Ere1 and Ere2, that function in the retromer pathway. Ere1 is required for Can1 recycling via the retromer-mediated pathway, but it is not required for the transport of other retromer cargoes, such as Vps10 and Ftr1. Biochemical studies reveal that Ere1 physically interacts with internalized Can1. Ere2 is present in a complex containing Ere1 on endosomes and functions as a regulator of Ere1. Taken together, our results suggest that Snx4/41/42 and the retromer comprise two independent pathways for the recycling of internalized cell-surface proteins. Moreover, a complex containing the two novel proteins Ere1 and Ere2 mediates cargo-specific recognition by the retromer pathway. PMID:21880895

  2. Rim Pathway-Mediated Alterations in the Fungal Cell Wall Influence Immune Recognition and Inflammation.

    Science.gov (United States)

    Ost, Kyla S; Esher, Shannon K; Leopold Wager, Chrissy M; Walker, Louise; Wagener, Jeanette; Munro, Carol; Wormley, Floyd L; Alspaugh, J Andrew

    2017-01-31

    mask immunogenic epitopes. We have created a fungal strain with a targeted mutation in a pH response pathway that is unable to properly organize its cell wall, resulting in a dramatic immune reaction during infection. This mutant cell wall is defective in hiding important cell wall components, such as the chito-oligomers chitin and chitosan. By creating a series of cell wall mutants, we demonstrated that the degree of chito-oligomer exposure correlates with the intensity of innate immune cell activation. This activation requires a combination of host receptors to recognize and respond to these infecting microorganisms. Therefore, these experiments explored host-pathogen interactions that determine the degree of the subsequent inflammatory response and the likely outcome of infection. Copyright © 2017 Ost et al.

  3. The S100A10 Pathway Mediates an Occult Hyperfibrinolytic Subtype in Trauma Patients.

    Science.gov (United States)

    Gall, Lewis S; Vulliamy, Paul; Gillespie, Scarlett; Jones, Timothy F; Pierre, Rochelle S J; Breukers, Sabine E; Gaarder, Christine; Juffermans, Nicole P; Maegele, Marc; Stensballe, Jakob; Johansson, Pär I; Davenport, Ross A; Brohi, Karim

    2018-03-19

    To determine the characteristics of trauma patients with low levels of fibrinolysis as detected by viscoelastic hemostatic assay (VHA) and explore the underlying mechanisms of this subtype. Hyperfibrinolysis is a central component of acute traumatic coagulopathy but a group of patients present with low levels of VHA-detected fibrinolysis. There is concern that these patients may be at risk of thrombosis if empirically administered an antifibrinolytic agent. A prospective multicenter observational cohort study was conducted at 5 European major trauma centers. Blood was drawn on arrival, within 2 hours of injury, for VHA (rotation thromboelastometry [ROTEM]) and fibrinolysis plasma protein analysis including the fibrinolytic mediator S100A10. An outcomes-based threshold for ROTEM hypofibrinolysis was determined and patients grouped by this and by D-dimer (DD) levels. Nine hundred fourteen patients were included in the study. The VHA maximum lysis (ML) lower threshold was determined to be <5%. Heterogeneity existed among patients with low ML, with survivors sharing similar clinical and injury characteristics to patients with normal ML values (5-15%). Those who died were critically injured with a preponderance of traumatic brain injury and had a 7-fold higher DD level (died vs. survived: 103,170 vs. 13,672 ng/mL, P < 0.001). Patients with low ML and high DD demonstrated a hyperfibrinolytic biomarker profile, low tissue plasminogen activator levels but high plasma levels of S100A10. S100A10 was negatively correlated with %ML (r = -0.26, P < 0.001) and caused a significant reduction in %ML when added to whole blood ex-vivo. Patients presenting with low ML and low DD levels have low injury severity and normal outcomes. Conversely, patients with low ML but high DD levels are severely injured, functionally coagulopathic and have poor clinical outcomes. These patients have low tissue plasminogen activator levels and are not detectable by ROTEM. S100A10 is a cell

  4. αν and β1 Integrins mediate Aβ-induced neurotoxicity in hippocampal neurons via the FAK signaling pathway.

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    Hai-Yan Han

    Full Text Available αν and β1 integrins mediate Aβ-induced neurotoxicity in primary hippocampal neurons. We treated hippocampal neurons with 2.5 µg/mL 17E6 and 5 µg/mL ab58524, which are specific αν and β1 integrin antagonists, respectively, for 42 h prior to 10 µM Aβ treatment. Next, we employed small interfering RNA (siRNA to silence focal adhesion kinase (FAK, a downstream target gene of integrins. The siRNAs were designed with a target sequence, an MOI of 10 and the addition of 5 µg/mL polybrene. Under these conditions, the neurons were transfected and the apoptosis of different cell types was detected. Moreover, we used real-time PCR and Western blotting analyses to detect the expression of FAK and ρFAK genes in different cell types and investigated the underlying mechanism and signal pathway by which αν and β1 integrins mediate Aβ-induced neurotoxicity in hippocampal neurons. An MTT assay showed that both 17E6 and ab58524 significantly increased cell viability compared with the Aβ-treated neurons (P<0.01 and P<0.05, respectively. However, this protective effect was markedly attenuated after transfection with silencing FAK (siFAK. Moreover, TUNEL immunostaining and flow cytometry indicated that both 17E6 and ab58524 significantly protected hippocampal neurons against apoptosis induced by Aβ (P<0.05 compared with the Aβ-treated cells. However, this protective effect was reversed with siFAK treatment. Both the gene and protein expression of FAK increased after Aβ treatment. Interestingly, as the gene and protein levels of FAK decreased, the ρFAK protein expression markedly increased. Furthermore, both the gene and protein expression of FAK and ρFAK were significantly diminished. Thus, we concluded that both αν and β1 integrins interfered with Aβ-induced neurotoxicity in hippocampal neurons and that this mechanism partially contributes to the activation of the Integrin-FAK signaling pathway.

  5. Enteric Glia Mediate Neuron Death in Colitis Through Purinergic Pathways That Require Connexin-43 and Nitric OxideSummary

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    Isola A.M. Brown

    2016-01-01

    Full Text Available Background & Aims: The concept of enteric glia as regulators of intestinal homeostasis is slowly gaining acceptance as a central concept in neurogastroenterology. Yet how glia contribute to intestinal disease is still poorly understood. Purines generated during inflammation drive enteric neuron death by activating neuronal P2X7 purine receptors (P2X7R; triggering adenosine triphosphate (ATP release via neuronal pannexin-1 channels that subsequently recruits intracellular calcium ([Ca2+]i in surrounding enteric glia. We tested the hypothesis that the activation of enteric glia contributes to neuron death during inflammation. Methods: We studied neuroinflammation in vivo using the 2,4-dinitrobenzene sulfonic acid model of colitis and in situ using whole-mount preparations of human and mouse intestine. Transgenic mice with a targeted deletion of glial connexin-43 (Cx43 [GFAP::CreERT2+/−/Cx43f/f] were used to specifically disrupt glial signaling pathways. Mice deficient in inducible nitric oxide (NO synthase (iNOS−/− were used to study NO production. Protein expression and oxidative stress were measured using immunohistochemistry and in situ Ca2+ and NO imaging were used to monitor glial [Ca2+]i and [NO]i. Results: Purinergic activation of enteric glia drove [Ca2+]i responses and enteric neuron death through a Cx43-dependent mechanism. Neurotoxic Cx43 activity, driven by NO production from glial iNOS, was required for neuron death. Glial Cx43 opening liberated ATP and Cx43-dependent ATP release was potentiated by NO. Conclusions: Our results show that the activation of glial cells in the context of neuroinflammation kills enteric neurons. Mediators of inflammation that include ATP and NO activate neurotoxic pathways that converge on glial Cx43 hemichannels. The glial response to inflammatory mediators might contribute to the development of motility disorders. Keywords: Enteric Nervous System, Hemichannels

  6. Selenite exacerbates hepatic insulin resistance in mouse model of type 2 diabetes through oxidative stress-mediated JNK pathway

    Energy Technology Data Exchange (ETDEWEB)

    Zhou, Jun, E-mail: hustzhj@hust.edu.cn; Xu, Gang; Bai, Zhaoshuai; Li, Kaicheng; Yan, Junyan; Li, Fen; Ma, Shuai; Xu, Huibi; Huang, Kaixun, E-mail: hxxzrf@hust.edu.cn

    2015-12-15

    Recent evidence suggests a potential pro-diabetic effect of selenite treatment in type 2 diabetics; however, the underlying mechanisms remain elusive. Here we investigated the effects and the underlying mechanisms of selenite treatment in a nongenetic mouse model of type 2 diabetes. High-fat diet (HFD)/streptozotocin (STZ)-induced diabetic mice were orally gavaged with selenite at 0.5 or 2.0 mg/kg body weight/day or vehicle for 4 weeks. High-dose selenite treatment significantly elevated fasting plasma insulin levels and insulin resistance index, in parallel with impaired glucose tolerance, insulin tolerance and pyruvate tolerance. High-dose selenite treatment also attenuated hepatic IRS1/Akt/FoxO1 signaling and pyruvate kinase gene expressions, but elevated the gene expressions of phosphoenolpyruvate carboxyl kinase (PEPCK), glucose 6-phosphatase (G6Pase), peroxisomal proliferator-activated receptor-γ coactivator 1α (PGC-1α) and selenoprotein P (SelP) in the liver. Furthermore, high-dose selenite treatment caused significant increases in MDA contents, protein carbonyl contents, and a decrease in GSH/GSSG ratio in the liver, concurrent with enhanced ASK1/MKK4/JNK signaling. Taken together, these findings suggest that high-dose selenite treatment exacerbates hepatic insulin resistance in mouse model of type 2 diabetes, at least in part through oxidative stress-mediated JNK pathway, providing new mechanistic insights into the pro-diabetic effect of selenite in type 2 diabetes. - Highlights: • Selenite exacerbates hepatic insulin resistance in HFD/STZ-induced diabetic mice. • Selenite elevates hepatic gluconeogenesis and reduces glycolysis in diabetic mice. • Selenite exacerbates hepatic oxidative stress and triggers JNK signaling pathway. • Selenite elevates hepatic selenoprotein P expression in diabetic mice.

  7. Selenite exacerbates hepatic insulin resistance in mouse model of type 2 diabetes through oxidative stress-mediated JNK pathway

    International Nuclear Information System (INIS)

    Zhou, Jun; Xu, Gang; Bai, Zhaoshuai; Li, Kaicheng; Yan, Junyan; Li, Fen; Ma, Shuai; Xu, Huibi; Huang, Kaixun

    2015-01-01

    Recent evidence suggests a potential pro-diabetic effect of selenite treatment in type 2 diabetics; however, the underlying mechanisms remain elusive. Here we investigated the effects and the underlying mechanisms of selenite treatment in a nongenetic mouse model of type 2 diabetes. High-fat diet (HFD)/streptozotocin (STZ)-induced diabetic mice were orally gavaged with selenite at 0.5 or 2.0 mg/kg body weight/day or vehicle for 4 weeks. High-dose selenite treatment significantly elevated fasting plasma insulin levels and insulin resistance index, in parallel with impaired glucose tolerance, insulin tolerance and pyruvate tolerance. High-dose selenite treatment also attenuated hepatic IRS1/Akt/FoxO1 signaling and pyruvate kinase gene expressions, but elevated the gene expressions of phosphoenolpyruvate carboxyl kinase (PEPCK), glucose 6-phosphatase (G6Pase), peroxisomal proliferator-activated receptor-γ coactivator 1α (PGC-1α) and selenoprotein P (SelP) in the liver. Furthermore, high-dose selenite treatment caused significant increases in MDA contents, protein carbonyl contents, and a decrease in GSH/GSSG ratio in the liver, concurrent with enhanced ASK1/MKK4/JNK signaling. Taken together, these findings suggest that high-dose selenite treatment exacerbates hepatic insulin resistance in mouse model of type 2 diabetes, at least in part through oxidative stress-mediated JNK pathway, providing new mechanistic insights into the pro-diabetic effect of selenite in type 2 diabetes. - Highlights: • Selenite exacerbates hepatic insulin resistance in HFD/STZ-induced diabetic mice. • Selenite elevates hepatic gluconeogenesis and reduces glycolysis in diabetic mice. • Selenite exacerbates hepatic oxidative stress and triggers JNK signaling pathway. • Selenite elevates hepatic selenoprotein P expression in diabetic mice.

  8. Hedgehog pathway mediates early acceleration of liver regeneration induced by a novel two-staged hepatectomy in mice.

    Science.gov (United States)

    Langiewicz, Magda; Schlegel, Andrea; Saponara, Enrica; Linecker, Michael; Borger, Pieter; Graf, Rolf; Humar, Bostjan; Clavien, Pierre A

    2017-03-01

    ALPPS, a novel two-staged approach for the surgical removal of large/multiple liver tumors, combines portal vein ligation (PVL) with parenchymal transection. This causes acceleration of compensatory liver growth, enabling faster and more extensive tumor removal. We sought to identify the plasma factors thought to mediate the regenerative acceleration following ALPPS. We compared a mouse model of ALPPS against PVL and additional control surgeries (n=6 per group). RNA deep sequencing was performed to identify candidate molecules unique to ALPPS liver (n=3 per group). Recombinant protein and a neutralizing antibody combined with appropriate surgeries were used to explore candidate functions in ALPPS (n=6 per group). Indian hedgehog (IHH/Ihh) levels were assessed in human ALPPS patient plasma (n=6). ALPPS in mouse confirmed significant acceleration of liver regeneration relative to PVL (pIhh mRNA, coding for a secreted ligand inducing hedgehog signaling, was uniquely upregulated in ALPPS liver (pIhh plasma levels rose 4h after surgery (pIhh alone was sufficient to induce ALPPS-like acceleration of liver growth. Conversely, blocking Ihh markedly inhibited the accelerating effects of ALPPS. In the small cohort of ALPPS patients, IHH tended to be elevated early after surgery. Ihh and hedgehog pathway activation provide the first mechanistic insight into the acceleration of liver regeneration triggered by ALPPS surgery. The accelerating potency of recombinant Ihh, and its potential effect in human ALPPS may lead to a clinical role for this protein. ALPPS, a novel two-staged hepatectomy, accelerates liver regeneration, thereby helping to treat patients with otherwise unresectable liver tumors. The molecular mechanisms behind this accelerated regeneration are unknown. Here, we elucidate that Indian hedgehog, a secreted ligand important for fetal development, is a crucial mediator of the regenerative acceleration triggered by ALPPS surgery. Copyright © 2016. Published by

  9. Propofol reduced myocardial contraction of vertebrates partly by mediating the cyclic AMP-dependent protein kinase phosphorylation pathway

    International Nuclear Information System (INIS)

    Sun, Xiaotong; Zhang, Xinyu; Bo, Qiyu; Meng, Tao; Lei, Zhen; Li, Jingxin; Hou, Yonghao; Yu, Xiaoqian; Yu, Jingui

    2016-01-01

    Propofol inhibits myocardial contraction in a dose dependent manner. The present study is designed to examine the effect of propofol on PKA mediated myocardial contraction in the absence of adrenoreceptor agonist. The contraction of isolated rat heart was measured in the presence or absence of PKA inhibitor H89 or propofol, using a pressure transducer. The levels of cAMP and PKA kinase activity were detected by ELISA. The mRNA and total protein or phosphorylation level of PKA and downstream proteins were tested in the presence or absence of PKA inhibitor H89 or propofol, using RT-PCR, QPCR and western blotting. The phosphorylation level of PKA was examined thoroughly using immunofluorescence and PKA activity non-radioactive detection kit. Propofol induced a dose-dependent negative contractile response on the rat heart. The inhibitory effect of high concentration propofol (50 μM) with 45% decease of control could be partly reversed by the PKA inhibitor H89 (10 μM) and the depressant effect of propofol decreased from 45% to 10%. PKA kinase activity was inhibited by propofol in a dose-dependent manner. Propofol also induced a decrease in phosphorylation of PKA, which was also inhibited by H89, but did not alter the production of cAMP and the mRNA levels of PKA. The downstream proteins of PKA, PLN and RyR2 were phosphorylated to a lesser extent with propofol or H89 than control. These results demonstrated that propofol induced a negative myocardial contractile response partly by mediating the PKA phosphorylation pathway.

  10. Cross-talk between an activator of nuclear receptors-mediated transcription and the D1 dopamine receptor signaling pathway.

    Science.gov (United States)

    Schmidt, Azriel; Vogel, Robert; Rutledge, Su Jane; Opas, Evan E; Rodan, Gideon A; Friedman, Eitan

    2005-03-01

    Nuclear receptors are transcription factors that usually interact, in a ligand-dependent manner, with specific DNA sequences located within promoters of target genes. The nuclear receptors can also be controlled in a ligand-independent manner via the action of membrane receptors and cellular signaling pathways. 5-Tetradecyloxy-2-furancarboxylic acid (TOFA) was shown to stimulate transcription from the MMTV promoter via chimeric receptors that consist of the DNA binding domain of GR and the ligand binding regions of the PPARbeta or LXRbeta nuclear receptors (GR/PPARbeta and GR/LXRbeta). TOFA and hydroxycholesterols also modulate transcription from NF-kappaB- and AP-1-controlled reporter genes and induce neurite differentiation in PC12 cells. In CV-1 cells that express D(1) dopamine receptors, D(1) dopamine receptor stimulation was found to inhibit TOFA-stimulated transcription from the MMTV promoter that is under the control of chimeric GR/PPARbeta and GR/LXRbeta receptors. Treatment with the D(1) dopamine receptor antagonist, SCH23390, prevented dopamine-mediated suppression of transcription, and by itself increased transcription controlled by GR/LXRbeta. Furthermore, combined treatment of CV-1 cells with TOFA and SCH23390 increased transcription controlled by the GR/LXRbeta chimeric receptor synergistically. The significance of this in vitro synergy was demonstrated in vivo, by the observation that SCH23390 (but not haloperidol)-mediated catalepsy in rats was potentiated by TOFA, thus showing that an agent that mimics the in vitro activities of compounds that activate members of the LXR and PPAR receptor families can influence D1 dopamine receptor elicited responses.

  11. Heme activates TLR4-mediated inflammatory injury via MyD88/TRIF signaling pathway in intracerebral hemorrhage

    Directory of Open Access Journals (Sweden)

    Lin Sen

    2012-03-01

    Full Text Available Abstract Background Inflammatory injury plays a critical role in intracerebral hemorrhage (ICH-induced neurological deficits; however, the signaling pathways are not apparent by which the upstream cellular events trigger innate immune and inflammatory responses that contribute to neurological impairments. Toll-like receptor 4 (TLR4 plays a role in inflammatory damage caused by brain disorders. Methods In this study, we investigate the role of TLR4 signaling in ICH-induced inflammation. In the ICH model, a significant upregulation of TLR4 expression in reactive microglia has been demonstrated using real-time RT-PCR. Activation of microglia was detected by immunohistochemistry, cytokines were measured by ELISA, MyD88, TRIF and NF-κB were measured by Western blot and EMSA, animal behavior was evaluated by animal behavioristics. Results Compared to WT mice, TLR4−/− mice had restrained ICH-induced brain damage showing in reduced cerebral edema and lower neurological deficit scores. Quantification of cytokines including IL-6, TNF-α and IL-1β and assessment of macrophage infiltration in perihematoma tissues from TLR4−/−, MyD88−/− and TRIF−/− mice showed attenuated inflammatory damage after ICH. TLR4−/− mice also exhibited reduced MyD88 and TRIF expression which was accompanied by decreased NF-κB activity. This suggests that after ICH both MyD88 and TRIF pathways might be involved in TLR4-mediated inflammatory injury possibly via NF-κB activation. Exogenous hemin administration significantly increased TLR4 expression and microglial activation in cultures and also exacerbated brain injury in WT mice but not in TLR4−/− mice. Anti-TLR4 antibody administration suppressed hemin-induced microglial activation in cultures and in the mice model of ICH. Conclusions Our findings suggest that heme potentiates microglial activation via TLR4, in turn inducing NF-κB activation via the MyD88/TRIF signaling pathway, and ultimately

  12. Does tryptophan degradation along the kynurenine pathway mediate the association between pro-inflammatory immune activity and depressive symptoms?

    Science.gov (United States)

    Quak, Jacqueline; Doornbos, Bennard; Roest, Annelieke M; Duivis, Hester E; Vogelzangs, Nicole; Nolen, Willem A; Penninx, Brenda W J H; Kema, Ido P; de Jonge, Peter

    2014-07-01

    Several studies have suggested that induced tryptophan (TRP) degradation through the kynurenine (KYN) pathway by the enzyme indoleamine 2,3-dioxygenase (IDO) is implicated in the relation between depression and inflammation. We investigated the role of tryptophan degradation in the relationship between inflammatory markers and depressive symptoms in the Netherlands Study of Depression and Anxiety (NESDA) and hypothesized that tryptophan degradation would mediate (part of) this association. 2812 Participants of NESDA were included in this study including 1042 persons with current major depressive disorder (MDD). Assessments of C-reactive protein (CRP), interleukin (IL)-6, tumor-necrosis factor (TNF)-α, KYN and TRP were obtained from fasting blood samples at the baseline assessment. Tryptophan degradation was estimated by calculating the ratio [KYN/TRP]. Depressive symptoms were measured with the Inventory of Depressive Symptomatology. Significant associations between inflammation and depressive symptoms were found for CRP and IL-6, for the total group and the subgroup of patients with current MDD. Adjustment for KYN/TRP did not attenuate these associations. There were no significant indirect effects for CRP on depressive symptoms mediated by KYN/TRP for the whole group (B=-0.032; 95% CI: -0.103 to 0.028) and for the subgroup of patients with current MDD (B=0.059; 95% CI: -0.037 to 0.165). Also IL-6 did not indirectly affect depressive symptoms through KYN/TRP in the total group (B=-0.023; 95% CI: -0.093 to 0.045) and in the MDD subgroup B=0.052; 95% CI: -0.019 to 0.144). Finally, no significant relation between depressive symptoms and KYN/TRP was found in the whole group (β=-0.019, p=0.311) nor in the subgroup with MDD (β=0.025, p=0.424). We did not find indications for tryptophan degradation, measured by KYN/TRP, to mediate the relationship between inflammation and depressive symptoms. Copyright © 2014 Elsevier Ltd. All rights reserved.

  13. Nickel (II)-induced cytotoxicity and apoptosis in human proximal tubule cells through a ROS- and mitochondria-mediated pathway

    Energy Technology Data Exchange (ETDEWEB)

    Wang, Yi-Fen; Shyu, Huey-Wen [Department of Medical Laboratory Sciences and Biotechnology, Fooyin University, Kaohsiung, Taiwan (China); Chang, Yi-Chuang [Department of Nursing, Fooyin University, Kaohsiung, Taiwan (China); Tseng, Wei-Chang [Department of Medical Laboratory Sciences and Biotechnology, Fooyin University, Kaohsiung, Taiwan (China); Huang, Yeou-Lih [Department of Medical Laboratory Science and Biotechnology, Kaohsiung Medical University, Kaohsiung, Taiwan (China); Lin, Kuan-Hua; Chou, Miao-Chen; Liu, Heng-Ling [Department of Medical Laboratory Sciences and Biotechnology, Fooyin University, Kaohsiung, Taiwan (China); Chen, Chang-Yu, E-mail: mt037@mail.fy.edu.tw [Department of Medical Laboratory Sciences and Biotechnology, Fooyin University, Kaohsiung, Taiwan (China)

    2012-03-01

    Nickel compounds are known to be toxic and carcinogenic in kidney and lung. In this present study, we investigated the roles of reactive oxygen species (ROS) and mitochondria in nickel (II) acetate-induced cytotoxicity and apoptosis in the HK-2 human renal cell line. The results showed that the cytotoxic effects of nickel (II) involved significant cell death and DNA damage. Nickel (II) increased the generation of ROS and induced a noticeable reduction of mitochondrial membrane potential (MMP). Analysis of the sub-G1 phase showed a significant increase in apoptosis in HK-2 cells after nickel (II) treatment. Pretreatment with N-acetylcysteine (NAC) not only inhibited nickel (II)-induced cell death and DNA damage, but also significantly prevented nickel (II)-induced loss of MMP and apoptosis. Cell apoptosis triggered by nickel (II) was characterized by the reduced protein expression of Bcl-2 and Bcl-xL and the induced the protein expression of Bad, Bcl-Xs, Bax, cytochrome c and caspases 9, 3 and 6. The regulation of the expression of Bcl-2-family proteins, the release of cytochrome c and the activation of caspases 9, 3 and 6 were inhibited in the presence of NAC. These results suggest that nickel (II) induces cytotoxicity and apoptosis in HK-2 cells via ROS generation and that the mitochondria-mediated apoptotic signaling pathway may be involved in the positive regulation of nickel (II)-induced renal cytotoxicity.

  14. Integrin-mediated signal transduction linked to Ras pathway by GRB2 binding to focal adhesion kinase.

    Science.gov (United States)

    Schlaepfer, D D; Hanks, S K; Hunter, T; van der Geer, P

    The cytoplasmic focal adhesion protein-tyrosine kinase (FAK) localizes with surface integrin receptors at sites where cells attach to the extracellular matrix. Increased FAK tyrosine phosphorylation occurs upon integrin engagement with fibronectin. Here we show that adhesion of murine NIH3T3 fibroblasts to fibronectin promotes SH2-domain-mediated association of the GRB2 adaptor protein and the c-Src protein-tyrosine kinase (PTK) with FAK in vivo, and also results in activation of mitogen-activated protein kinase (MAPK). In v-Src-transformed NIH3T3, the association of v-Src, GRB2 and Sos with FAK is independent of cell adhesion to fibronectin. The GRB2 SH2 domain binds directly to tyrosine-phosphorylated FAK. Mutation of tyrosine residue 925 of FAK (YENV motif) to phenylalanine blocks GRB2 SH2-domain binding to FAK in vitro. Our results show that fibronectin binding to integrins on NIH3T3 fibroblasts promotes c-Src and FAK association and formation of an integrin-activated signalling complex. Phosphorylation of FAK at Tyr 925 upon fibronectin stimulation creates an SH2-binding site for GRB2 which may link integrin engagement to the activation of the Ras/MAPK signal transduction pathway.

  15. Ultraviolet-Ray-Induced Sea Cucumber (Stichopus japonicus) Melting Is Mediated by the Caspase-Dependent Mitochondrial Apoptotic Pathway.

    Science.gov (United States)

    Su, Li; Yang, Jing-Feng; Fu, Xi; Dong, Liang; Zhou, Da-Yong; Sun, Li-Ming; Gong, Zhenwei

    2018-01-10

    Sea cucumber body-wall melting occurs under certain circumstances. We have shown that apoptosis but not autolysis plays a critical role in the initial stage. However, it is still unclear how apoptosis is triggered in this process. In this study, we examined the levels of reactive oxygen species (ROS), the levels of B-cell lymphoma 2 (Bcl-2) and Bcl-2-associated X (Bax) proteins, the depolarization of mitochondrial transmembrane potentials, and cytochrome c (Cyt c) release during sea cucumber melting induced by ultraviolet (UV) exposure. We also investigated the contribution of caspase in this process by injecting a pan-caspase inhibitor. Our data showed that UV exposure stimulates ROS production, dysfunction of mitochondria, and the release of Cyt c in sea cucumber coelomic fluid cells and body walls. We found a decrease of Bcl-2 and increase of Bax in the mitochondria after UV exposure. We also demonstrated that these changes are associated with elevated caspase-9 and -3 activity. Finally, our data showed that the inhibition of caspases-9 and -3 using an inhibitor suppresses UV-induced sea cucumber melting. These results suggest that apoptosis during sea cucumber melting is mediated by mitochondrial dysfunction and follows the activation of the caspase-signaling pathway. This study presents a novel insight into the mechanism of sea cucumber melting.

  16. TNF-α- Mediated-p38-Dependent Signaling Pathway Contributes to Myocyte Apoptosis in Rats Subjected to Surgical Trauma

    Directory of Open Access Journals (Sweden)

    Huaxing Wu

    2015-03-01

    Full Text Available Background: The accumulation of cytokines in the plasma after trauma can induce myocyte apoptosis. We aimed to identify which cytokine(s present in the plasma responsible for myocyte apoptosis, and delineated the signal transduction mechanism in rats subjected to surgical trauma. Methods: Rats were randomized into two groups: control and trauma groups, which was divided into five subgroups: posttraumatic 0, 3, 6, 12, and 24 h subgroups. Cardiomyocytes isolated from traumatized rats were incubated with one of the factors for 12 h (normal plasma; Cytomix; TNF-α; IL-1β; IFN-γ; trauma plasma; anti-TNF-α antibody; SB203580. Myocyte apoptosis, cytokine levels, and MAPKs activation, as the primary experimental outcomes, were measured by TUNEL, flow cytometry, ELISA and Western blot, respectively. Results: Myocyte apoptosis was induced by surgical trauma during the early stage after trauma. Accompanying this change, plasma TNF-α, IL-1β, and IFN-γ levels were elevated in traumatized rats. Incubation of traumatized cardiomyocytes with cytomix or TNF-α alone induced myocyte apoptosis, and increased the activation of p38 and ERK1/2. Myocyte apoptosis and p38 activation were elevated in traumatized cardiomyocytes with trauma plasma, and these increases were partly abolished by anti-TNF-α antibody or SB203580. Conclusion: Our study demonstrated that there exists the TNF-α-mediated-p38-dependent signaling pathway that contributed to posttraumatic myocyte apoptosis of rats undergoing surgical trauma.

  17. Fisetin Alleviates Lipopolysaccharide-Induced Acute Lung Injury via TLR4-Mediated NF-κB Signaling Pathway in Rats.

    Science.gov (United States)

    Feng, Guang; Jiang, Ze-Yu; Sun, Bo; Fu, Jie; Li, Tian-Zuo

    2016-02-01

    Acute lung injury (ALI), a common component of systemic inflammatory disease, is a life-threatening condition without many effective treatments. Fisetin, a natural flavonoid from fruits and vegetables, was reported to have wide pharmacological properties such as anti-inflammatory, antioxidant, and anticancer activities. The aim of this study was to detect the effects of fisetin on lipopolysaccharide (LPS)-induced acute lung injury and investigate the potential mechanism. Fisetin was injected (1, 2, and 4 mg/kg, i.v.) 30 min before LPS administration (5 mg/kg, i.v.). Our results showed that fisetin effectively reduced the inflammatory cytokine release and total protein in bronchoalveolar lavage fluids (BALF), decreased the lung wet/dry ratios, and obviously improved the pulmonary histology in LPS-induced ALI. Furthermore, fisetin inhibited LPS-induced increases of neutrophils and macrophage infiltration and attenuated MPO activity in lung tissues. Additionally, fisetin could significantly inhibit the Toll-like receptor 4 (TLR4) expression and the activation of NF-κB in lung tissues. Our data indicates that fisetin has a protective effect against LPS-induced ALI via suppression of TLR4-mediated NF-κB signaling pathways, and fisetin may be a promising candidate for LPS-induced ALI treatment.

  18. Nickel (II)-induced cytotoxicity and apoptosis in human proximal tubule cells through a ROS- and mitochondria-mediated pathway

    International Nuclear Information System (INIS)

    Wang, Yi-Fen; Shyu, Huey-Wen; Chang, Yi-Chuang; Tseng, Wei-Chang; Huang, Yeou-Lih; Lin, Kuan-Hua; Chou, Miao-Chen; Liu, Heng-Ling; Chen, Chang-Yu

    2012-01-01

    Nickel compounds are known to be toxic and carcinogenic in kidney and lung. In this present study, we investigated the roles of reactive oxygen species (ROS) and mitochondria in nickel (II) acetate-induced cytotoxicity and apoptosis in the HK-2 human renal cell line. The results showed that the cytotoxic effects of nickel (II) involved significant cell death and DNA damage. Nickel (II) increased the generation of ROS and induced a noticeable reduction of mitochondrial membrane potential (MMP). Analysis of the sub-G1 phase showed a significant increase in apoptosis in HK-2 cells after nickel (II) treatment. Pretreatment with N-acetylcysteine (NAC) not only inhibited nickel (II)-induced cell death and DNA damage, but also significantly prevented nickel (II)-induced loss of MMP and apoptosis. Cell apoptosis triggered by nickel (II) was characterized by the reduced protein expression of Bcl-2 and Bcl-xL and the induced the protein expression of Bad, Bcl-Xs, Bax, cytochrome c and caspases 9, 3 and 6. The regulation of the expression of Bcl-2-family proteins, the release of cytochrome c and the activation of caspases 9, 3 and 6 were inhibited in the presence of NAC. These results suggest that nickel (II) induces cytotoxicity and apoptosis in HK-2 cells via ROS generation and that the mitochondria-mediated apoptotic signaling pathway may be involved in the positive regulation of nickel (II)-induced renal cytotoxicity.