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Sample records for expressed camp phosphodiesterase

  1. cAMP phosphodiesterase and activator protein of mammalian cAMP phosphodiesterase from Trypanosoma cruzi.

    PubMed

    Gonçalves, M F; Zingales, B; Colli, W

    1980-04-01

    Epimastigote forms of Trypanosoma cruzi contain a soluble cAMP phosphodiesterase. Optimal activity was found at pH 8.0 and in the presence of 5 mM Mn2+. Other cations were less efficient and did not give rise to an additional stimulation when added in the presence of optimal concentrations of Mn2+. The enzyme is not Ca2+ dependent. The apparent Km of the enzyme for the substrate is 40 microM and no kinetic evidence for the existence of two enzymes has been found. Theophylline and caffein did not inhibit the T. cruzi cAMP phosphodiesterase. The enzyme activity does not change during cell growth suggesting that the fluctuation observed in the levels of cAMP are largely a response to variations in adenylyl cyclase activity. The intracellular concentrations of cAMP ranged between 0.04--0.15 microM. No evidence that the T. cruzi cAMP phosphodiesterase is regulated by an endogenous activator could be found. However, T. cruzi contains a heat-stable, low molecular weight, non-dialysable protein that activates mammalian cAMP phosphodiesterase in the presence of Ca2+. The properties so far studied of such an activator suggest that it might be equivalent to other Ca2+-dependent regulators described in vertebrate and invertebrate species. PMID:6255327

  2. Regulation of cAMP by Phosphodiesterases in Erythrocytes

    PubMed Central

    Adderley, Shaquria P.; Sprague, Randy S.; Stephenson, Alan H.; Hanson, Madelyn S.

    2010-01-01

    The erythrocyte, a cell responsible for carrying and delivering oxygen in the body, has often been regarded as simply a vehicle for the circulation of hemoglobin. However, it has become evident that this cell also participates in the regulation of vascular caliber in the microcirculation via release of the potent vasodilator, adenosine triphosphate (ATP). The regulated release of ATP from erythrocytes occurs via a defined signaling pathway and requires increases in cyclic 3’ 5’ adenosine monophosphate (cAMP). It is well recognized that cAMP is a critical second messenger in diverse signaling pathways. In all cells increases in cAMP are localized and regulated by the activity of phosphodiesterases (PDEs). In erythrocytes activation of either β adrenergic receptors (β 2AR) or the prostacyclin receptor (IPR) results in increases in cAMP and ATP release. Receptor-mediated increases in cAMP are tightly regulated by distinct PDEs associated with each signaling pathway as shown by the finding that selective inhibitors of the PDEs localized to each pathway potentiate both increases in cAMP and ATP release. Here we review the profile of PDEs identified in erythrocytes, their association with specific signaling pathways and their role in the regulation of ATP release from these cells. Understanding the contribution of PDEs to the control of ATP release from erythrocytes identifies this cell as a potential target for the development of drugs for the treatment of vascular disease. PMID:20631411

  3. Temporal and spatial regulation of cAMP signaling in disease: role of cyclic nucleotide phosphodiesterases.

    PubMed

    Otero, Carolina; Peñaloza, Juan P; Rodas, Paula I; Fernández-Ramires, Ricardo; Velasquez, Luis; Jung, Juan E

    2014-12-01

    Since its discovery, cAMP has been proposed as one of the most versatile second messengers. The remarkable feature of cAMP to tightly control highly diverse physiological processes, including metabolism, homeostasis, secretion, muscle contraction, cell proliferation and migration, immune response, and gene transcription, is reflected by millions of different articles worldwide. Compartmentalization of cAMP in space and time, maintained by mainly phosphodiesterases, contributes to the maintenance of equilibrium inside the cell where one signal can trigger many different events. Novel cAMP sensors seem to carry out certain unexpected signaling properties of cAMP and thereby to permit delicate adaptations of biologic responses. Measuring space and time events with biosensors will increase our current knowledge on the pathophysiology of diseases, such as chronic obstructive pulmonary disease, asthma, cognitive impairment, cancer, and renal and heart failure. Further insights into the cAMP dynamics will help to optimize the pharmacological treatment for these diseases. PMID:24750474

  4. Electrostatic steering enhances the rate of cAMP binding to phosphodiesterase: Brownian dynamics modeling.

    PubMed

    Huang, Yu-ming M; Huber, Gary; McCammon, J Andrew

    2015-11-01

    Signaling in cells often involves co-localization of the signaling molecules. Most experimental evidence has shown that intracellular compartmentalization restricts the range of action of the second messenger, 3'-5'-cyclic adenosine monophosphate (cAMP), which is degraded by phosphodiesterases (PDEs). The objective of this study is to understand the details of molecular encounter that may play a role in efficient operation of the cAMP signaling apparatus. The results from electrostatic potential calculations and Brownian dynamics simulations suggest that positive potential of the active site from PDE enhances capture of diffusing cAMP molecules. This electrostatic steering between cAMP and the active site of a PDE plays a major role in the enzyme-substrate encounter, an effect that may be of significance in sequestering cAMP released from a nearby binding site or in attracting more freely diffusing cAMP molecules. PMID:26346301

  5. In vitro and in vivo characterization of the Pseudomonas aeruginosa cyclic AMP (cAMP) phosphodiesterase CpdA, required for cAMP homeostasis and virulence factor regulation.

    PubMed

    Fuchs, Erin L; Brutinel, Evan D; Klem, Erich R; Fehr, Anthony R; Yahr, Timothy L; Wolfgang, Matthew C

    2010-06-01

    Cyclic AMP (cAMP) is an important second messenger signaling molecule that controls a wide variety of eukaryotic and prokaryotic responses to extracellular cues. For cAMP-dependent signaling pathways to be effective, the intracellular cAMP concentration is tightly controlled at the level of synthesis and degradation. In the opportunistic human pathogen Pseudomonas aeruginosa, cAMP is a key regulator of virulence gene expression. To better understand the role of cAMP homeostasis in this organism, we identified and characterized the enzyme CpdA, a putative cAMP phosphodiesterase. We demonstrate that CpdA possesses 3',5'-cAMP phosphodiesterase activity in vitro and that it utilizes an iron-dependent catalytic mechanism. Deletion of cpdA results in the accumulation of intracellular cAMP and altered regulation of P. aeruginosa virulence traits. Further, we demonstrate that the cAMP-dependent transcription factor Vfr directly regulates cpdA expression in response to intracellular cAMP accumulation, thus providing a feedback mechanism for controlling cAMP levels and fine-tuning virulence factor expression. PMID:20348254

  6. Phosphodiesterase inhibition by a gastroprotective agent irsogladine: preferential blockade of cAMP hydrolysis.

    PubMed

    Kyoi, Takashi; Oka, Michiko; Noda, Kumiko; Ukai, Yojiro

    2004-08-27

    The effect of irsogladine [2,4-diamino-6-(2,5-dichlorophenyl)-s-triazine maleate], an antiulcer drug, on contents of cyclic nucleotides including cAMP and cGMP was investigated in rat stomachs. Irsogladine concentration-dependently increased cAMP content in rat glandula stomach. However, irsogladine at higher concentration (10(-5) M) was unable to further increase cAMP level in the presence of non-selective phosphodiesterase (PDE) inhibitor 3-isobutyl-1-methylxanthine, although 3-isobutyl-1-methylxanthine by itself increased cAMP level. On the other hand, irsogladine had no effect on the glandula cGMP content. Subsequently, the effect of irsogladine on the cyclic nucleotide degradation by purified bovine brain and heart PDEs was investigated. The cAMP degradation by purified bovine brain PDE was partially suppressed by PDE1 inhibitor vinpocetin, PDE2 inhibitor erythro-9-(2-hydroxy-3-nonyl)adenine hydrochloride and PDE4 inhibitor rolipram but not by PDE3 inhibitor cilostamide, and completely inhibited by 3-isobutyl-1-methylxanthine, suggesting that is attributed almost exclusively to PDE1, PDE2 and PDE4. Meanwhile, cGMP degradation by purified bovine brain PDE was partially suppressed by erythro-9-(2-hydroxy-3-nonyl)adenine hydrochloride. Irsogladine preferentially inhibited the response to cAMP degradation compared with cGMP degradation by this brain PDE. The cAMP degradation by bovine heart PDE was almost completely inhibited by the combination with vinpocetine and cilostamide, indicating that is mediated almost exclusively by PDE1 and PDE3. Irsogladine suppressed this cAMP degradation measured in the presence of vinpocetine to almost the same extent as that determined in the presence of cilostamide. These results indicate that irsogladine produces the increase of intracellular cAMP content via non-selective inhibition of PDE isozymes, which may be a key mechanism involved in its gastroprotective actions. PMID:15302227

  7. A cAMP Biosensor-Based High-Throughput Screening Assay for Identification of Gs-Coupled GPCR Ligands and Phosphodiesterase Inhibitors.

    PubMed

    Vedel, Line; Bräuner-Osborne, Hans; Mathiesen, Jesper Mosolff

    2015-08-01

    Cyclic adenosine 3',5'-monophosphate (cAMP) is an important second messenger, and quantification of intracellular cAMP levels is essential in studies of G protein-coupled receptors (GPCRs). The intracellular cAMP levels are regulated by the adenylate cyclase (AC) upon activation of either Gs- or Gi-coupled GPCRs, which leads to increased or decreased cAMP levels, respectively. Here we describe a real-time Förster resonance energy transfer (FRET)-based cAMP high-throughput screening (HTS) assay for identification and characterization of Gs-coupled GPCR ligands and phosphodiesterase (PDE) inhibitors in living cells. We used the β2-adrenergic receptor (β(2)AR) as a representative Gs-coupled receptor and characterized two cell lines with different expression levels. Low receptor expression allowed detection of desensitization kinetics and delineation of partial agonism, whereas high receptor expression resulted in prolonged signaling and enabled detection of weak partial agonists and/or ligands with low potency, which is highly advantageous in large HTS settings and hit identification. In addition, the assay enabled detection of β(2)AR inverse agonists and PDE inhibitors. High signal-to-noise ratios were also observed for the other representative Gs-coupled GPCRs tested, GLP-1R and GlucagonR. The FRET-based cAMP biosensor assay is robust, reproducible, and inexpensive with good Z factors and is highly applicable for HTS. PMID:25851033

  8. Studying mechanisms of cAMP and cyclic nucleotide phosphodiesterase signaling in Leydig cell function with phosphoproteomics.

    PubMed

    Golkowski, Martin; Shimizu-Albergine, Masami; Suh, Hyong Won; Beavo, Joseph A; Ong, Shao-En

    2016-07-01

    Many cellular processes are modulated by cyclic AMP and nucleotide phosphodiesterases (PDEs) regulate this second messenger by catalyzing its breakdown. The major unique function of testicular Leydig cells is to produce testosterone in response to luteinizing hormone (LH). Treatment of Leydig cells with PDE inhibitors increases cAMP levels and the activity of its downstream effector, cAMP-dependent protein kinase (PKA), leading to a series of kinase-dependent signaling and transcription events that ultimately increase testosterone release. We have recently shown that PDE4B and PDE4C as well as PDE8A and PDE8B are expressed in rodent Leydig cells and that combined inhibition of PDE4 and PDE8 leads to dramatically increased steroid biosynthesis. Here we investigated the effect of PDE4 and PDE8 inhibition on the molecular mechanisms of cAMP actions in a mouse MA10 Leydig cell line model with SILAC mass spectrometry-based phosphoproteomics. We treated MA10 cells either with PDE4 family specific inhibitor (Rolipram) and PDE8 family specific inhibitor (PF-04957325) alone or in combination and quantified the resulting phosphorylation changes at five different time points between 0 and 180min. We identified 28,336 phosphosites from 4837 proteins and observed significant regulation of 749 sites in response to PDE4 and PDE8 inhibitor treatment. Of these, 132 phosphosites were consensus PKA sites. Our data strongly suggest that PDE4 and PDE8 inhibitors synergistically regulate phosphorylation of proteins required for many different cellular processes, including cell cycle progression, lipid and glucose metabolism, transcription, endocytosis and vesicle transport. Our data suggests that cAMP, PDE4 and PDE8 coordinate steroidogenesis by acting on not one rate-limiting step but rather multiple pathways. Moreover, the pools of cAMP controlled by these PDEs also coordinate many other metabolic processes that may be regulated to assure timely and sufficient testosterone secretion

  9. Evolution of self-organisation in Dictyostelia by adaptation of a non-selective phosphodiesterase and a matrix component for regulated cAMP degradation

    PubMed Central

    Kawabe, Yoshinori; Weening, Karin E.; Marquay-Markiewicz, Jacques; Schaap, Pauline

    2012-01-01

    Dictyostelium discoideum amoebas coordinate aggregation and morphogenesis by secreting cyclic adenosine monophosphate (cAMP) pulses that propagate as waves through fields of cells and multicellular structures. To retrace how this mechanism for self-organisation evolved, we studied the origin of the cAMP phosphodiesterase PdsA and its inhibitor PdiA, which are essential for cAMP wave propagation. D. discoideum and other species that use cAMP to aggregate reside in group 4 of the four major groups of Dictyostelia. We found that groups 1-3 express a non-specific, low affinity orthologue of PdsA, which gained cAMP selectivity and increased 200-fold in affinity in group 4. A low affinity group 3 PdsA only partially restored aggregation of a D. discoideum pdsA-null mutant, but was more effective at restoring fruiting body morphogenesis. Deletion of a group 2 PdsA gene resulted in disruption of fruiting body morphogenesis, but left aggregation unaffected. Together, these results show that groups 1-3 use a low affinity PdsA for morphogenesis that is neither suited nor required for aggregation. PdiA belongs to a family of matrix proteins that are present in all Dictyostelia and consist mainly of cysteine-rich repeats. However, in its current form with several extensively modified repeats, PdiA is only present in group 4. PdiA is essential for initiating spiral cAMP waves, which, by organising large territories, generate the large fruiting structures that characterise group 4. We conclude that efficient cAMP-mediated aggregation in group 4 evolved by recruitment and adaptation of a non-selective phosphodiesterase and a matrix component into a system for regulated cAMP degradation. PMID:22357931

  10. Differential regulation of human platelet responses by cGMP inhibited and stimulated cAMP phosphodiesterases.

    PubMed

    Manns, J M; Brenna, K J; Colman, R W; Sheth, S B

    2002-05-01

    Platelets contain two cAMP phosphodiesterases (PDEs) which regulate intracellular cAMP levels, cGMP-inhibited cAMP PDE (PDE3A) and cGMP-stimulated PDE (PDE2A). Using the PDE3 inhibitor, milrinone and the PDE2 inhibitor, erythro-9-(2-hydroxyl-3-nonyl)adenine (EHNA), we have explored the contribution of each PDE to the regulation of platelet function. Inhibition of PDE2 resulted in higher levels of intracellular cAMP than inhibition of PDE3A suggesting this PDE may be the more important regulator of cAMP in human platelets. However, a concentration-dependent inhibition of agonist-induced aggregation was observed with milrinone while little effect was seen with EHNA. In addition, we observed a concentration-dependent inhibition in the increase of intracellular Ca2+ with PDE3 inhibition and significantly less with PDE2 inhibition. PDE3 inhibition also resulted in a concentration-dependent increase in cAMP-mediated phosphorylation of the vasodilator-stimulated phospho-protein (VASP) whereas there was no significant increase with PDE2 inhibition. In each of these experiments, synergism was noted with the combination of milrinone and EHNA. These results suggest that cAMP pools may be localized and the various PDEs regulate specific pools. These data also suggest that inhibitors of PDE3A may be more effective antiplatelet agents. PMID:12038792

  11. PDE4 cAMP phosphodiesterases: modular enzymes that orchestrate signalling cross-talk, desensitization and compartmentalization.

    PubMed Central

    Houslay, Miles D; Adams, David R

    2003-01-01

    cAMP is a second messenger that controls many key cellular functions. The only way to inactivate cAMP is to degrade it through the action of cAMP phosphodiesterases (PDEs). PDEs are thus poised to play a key regulatory role. PDE4 cAMP-specific phosphodiesterases appear to have specific functions with selective inhibitors serving as potent anti-inflammatory agents. The recent elucidation of the structure of the PDE4 catalytic unit allows for molecular insight into the mode of catalysis as well as substrate and inhibitor selectivity. The four PDE4 genes encode over 16 isoforms, each of which is characterized by a unique N-terminal region. PDE4 isoforms play a pivotal role in controlling functionally and spatially distinct pools of cAMP by virtue of their unique intracellular targeting. Targeting occurs by association with proteins, such as arrestins, SRC family tyrosyl kinases, A-kinase anchoring proteins ('AKAPs') and receptor for activated C kinase 1 ('RACK1'), and, in the case of isoform PDE4A1, by a specific interaction (TAPAS-1) with phosphatidic acid. PDE4 isoforms are 'designed' to be regulated by extracellular-signal-related protein kinase (ERK), which binds to anchor sites on the PDE4 catalytic domain that it phosphorylates. The upstream conserved region 1 (UCR1) and 2 (UCR2) modules that abut the PDE4 catalytic unit confer regulatory functions by orchestrating the functional outcome of phosphorylation by cAMP-dependent protein kinase ('PKA') and ERK. PDE4 enzymes stand at a crossroads that allows them to integrate various signalling pathways with that of cAMP in spatially distinct compartments. PMID:12444918

  12. β-Adrenergic cAMP Signals Are Predominantly Regulated by Phosphodiesterase Type 4 in Cultured Adult Rat Aortic Smooth Muscle Cells

    PubMed Central

    Nicolas, Valérie; Ji, Guangju; Fischmeister, Rodolphe; Leblais, Véronique

    2012-01-01

    Background We investigated the role of cyclic nucleotide phosphodiesterases (PDEs) in the spatiotemporal control of intracellular cAMP concentrations in rat aortic smooth muscle cells (RASMCs). Methodology/Principal Findings The rank order of PDE families contributing to global cAMP-PDE activity was PDE4> PDE3  =  PDE1. PDE7 mRNA expression but not activity was confirmed. The Fluorescence Resonance Energy Transfer (FRET)-based cAMP sensor, Epac1-camps, was used to monitor the time course of cytosolic cAMP changes. A pulse application of the β-adrenoceptor (β-AR) agonist isoproterenol (Iso) induced a transient FRET signal. Both β1- and β2-AR antagonists decreased the signal amplitude without affecting its kinetics. The non-selective PDE inhibitor (IBMX) dramatically increased the amplitude and delayed the recovery phase of Iso response, in agreement with a role of PDEs in degrading cAMP produced by Iso. Whereas PDE1, PDE3 and PDE7 blockades [with MIMX, cilostamide (Cil) and BRL 50481 (BRL), respectively] had no or minor effect on Iso response, PDE4 inhibition [with Ro-20-1724 (Ro)] strongly increased its amplitude and delayed its recovery. When Ro was applied concomitantly with MIMX or Cil (but not with BRL), the Iso response was drastically further prolonged. PDE4 inhibition similarly prolonged both β1- and β2-AR-mediated responses. When a membrane-targeted FRET sensor was used, PDE3 and PDE4 acted in a synergistic manner to hydrolyze the submembrane cAMP produced either at baseline or after β-AR stimulation. Conclusion/Significance Our study underlines the importance of cAMP-PDEs in the dynamic control of intracellular cAMP signals in RASMCs, and demonstrates the prominent role of PDE4 in limiting β-AR responses. PDE4 inhibition unmasks an effect of PDE1 and PDE3 on cytosolic cAMP hydrolyzis, and acts synergistically with PDE3 inhibition at the submembrane compartment. This suggests that mixed PDE4/PDE1 or PDE4/PDE3 inhibitors would be attractive to

  13. The Dunce cAMP phosphodiesterase PDE-4 negatively regulates G alpha(s)-dependent and G alpha(s)-independent cAMP pools in the Caenorhabditis elegans synaptic signaling network.

    PubMed

    Charlie, Nicole K; Thomure, Angela M; Schade, Michael A; Miller, Kenneth G

    2006-05-01

    Forward genetic screens for mutations that rescue the paralysis of ric-8 (Synembryn) reduction-of-function mutations frequently reveal mutations that cause hyperactivation of one or more components of the G alpha(s) pathway. Here, we report that one of these mutations strongly reduces the function of the Dunce cAMP phosphodiesterase PDE-4 by disrupting a conserved active site residue. Loss of function and neural overexpression of PDE-4 have profound and opposite effects on locomotion rate, but drug-response assays suggest that loss of PDE-4 function does not affect steady-state acetylcholine release or reception. Our genetic analysis suggests that PDE-4 regulates both G alpha(s)-dependent and G alpha(s)-independent cAMP pools in the neurons controlling locomotion rate. By immunostaining, PDE-4 is strongly expressed throughout the nervous system, where it localizes to small regions at the outside boundaries of synaptic vesicle clusters as well as intersynaptic regions. The synaptic subregions containing PDE-4 are distinct from those containing active zones, as indicated by costaining with an antibody against the long form of UNC-13. This highly focal subsynaptic localization suggests that PDE-4 may exert its effects by spatially regulating intrasynaptic cAMP pools. PMID:16624912

  14. RACK1 and β-arrestin2 attenuate dimerization of PDE4 cAMP phosphodiesterase PDE4D5.

    PubMed

    Bolger, Graeme B

    2016-07-01

    PDE4 family cAMP-selective cyclic nucleotide phosphodiesterases are important in the regulation of cAMP abundance in numerous systems, and thereby play an important role in the regulation of PKA and EPAC activity and the phosphorylation of CREB. We have used the yeast 2-hybrid system to demonstrate recently that long PDE4 isoforms form homodimers, consistent with data obtained recently by structural studies. The long PDE4 isoform PDE4D5 interacts selectively with β-arrestin2, implicated in the regulation of G-protein-coupled receptors and other cell signaling components, and also with the β-propeller protein RACK1. In the present study, we use 2-hybrid approaches to demonstrate that RACK1 and β-arrestin2 inhibit the dimerization of PDE4D5. We also show that serine-to-alanine mutations at PKA and ERK1/2 phosphorylation sites on PDE4D5 detectably ablate dimerization. Conversely, phospho-mimic serine-to-aspartate mutations at the MK2 and oxidative stress kinase sites ablate dimerization. Analysis of PDE4D5 that is locked into the dimeric configuration by the formation of a trans disulfide bond between Ser261 and Ser602 shows that RACK1 interacts strongly with both the monomeric and dimeric forms, but that β-arrestin2 interacts exclusively with the monomeric form. This is consistent with the concept that β-arrestin2 can preferentially recruit the monomeric, or "open," form of PDE4D5 to β2-adrenergic receptors, where it can regulate cAMP signaling. PMID:26257302

  15. Transcriptional regulation of 2',3'-cyclic nucleotide 3'-phosphodiesterase gene expression by cyclic AMP in C6 cells.

    PubMed

    Gravel, M; Gao, E; Hervouet-Zeiber, C; Parsons, V; Braun, P E

    2000-11-01

    It was recently shown that the two transcripts encoding the isoforms of 2',3'-cyclic nucleotide 3'-phosphodiesterase (CNP1 and CNP2) are differentially regulated during the process of oligodendrocyte maturation. In oligodendrocyte precursors, only CNP2 mRNA is present, whereas in differentiating oligodendrocytes, both CNP1 and CNP2 mRNAs are expressed. This pattern of CNP expression is likely due to stage-specific transcriptional regulation of the two CNP promoters during the process of oligodendrocyte differentiation. Here, we report the influence of increased intracellular cyclic AMP (cAMP) levels on the transcription of both CNP1 and CNP2 mRNAs in rat C6 glioma cells. We found that the transcription of CNP1 mRNA was significantly increased in comparison with that of CNP2 mRNA in cells treated with cAMP analogues to elevate intracellular cAMP levels. This up-regulation of CNP1 expression (a) is due to an increase of transcription, (b) requires de novo protein synthesis, and (c) requires the activity of protein kinase A. These results are physiologically significant and support the idea that a cAMP-mediated pathway is part of the molecular mechanisms regulating the expression of CNP1 in oligodendrocytes. The regulation of CNP1 promoter activity by cAMP was then investigated in stably transfected C6 cell lines containing various deletions of the CNP promoter directing the bacterial chloramphenicol acetyltransferase gene. We showed that the sequence between nucleotides -126 and -102 was essential for the cAMP-dependent induction of CNP1 expression. Gel retardation analysis showed that two protein-DNA complexes are formed between this sequence and nuclear factors from C6 cells treated or not treated with cAMP. This suggests that the induction of CNP1 mRNA transcription is not mediated by changes in binding of nuclear factors that interact directly with the -126/-102 sequence. Sequence analysis of this region revealed the presence of a putative activator protein-2 (AP

  16. Expression and Genetic Activation of Cyclic Di-GMP-Specific Phosphodiesterases in Escherichia coli

    PubMed Central

    Reinders, Alberto; Hee, Chee-Seng; Ozaki, Shogo; Mazur, Adam; Boehm, Alex; Schirmer, Tilman

    2015-01-01

    ABSTRACT Intracellular levels of the bacterial second messenger cyclic di-GMP (c-di-GMP) are controlled by antagonistic activities of diguanylate cyclases and phosphodiesterases. The phosphodiesterase PdeH was identified as a key regulator of motility in Escherichia coli, while deletions of any of the other 12 genes encoding potential phosphodiesterases did not interfere with motility. To analyze the roles of E. coli phosphodiesterases, we demonstrated that most of these proteins are expressed under laboratory conditions. We next isolated suppressor mutations in six phosphodiesterase genes, which reinstate motility in the absence of PdeH by reducing cellular levels of c-di-GMP. Expression of all mutant alleles also led to a reduction of biofilm formation. Thus, all of these proteins are bona fide phosphodiesterases that are capable of interfering with different c-di-GMP-responsive output systems by affecting the global c-di-GMP pool. This argues that E. coli possesses several phosphodiesterases that are inactive under laboratory conditions because they lack appropriate input signals. Finally, one of these phosphodiesterases, PdeL, was studied in more detail. We demonstrated that this protein acts as a transcription factor to control its own expression. Motile suppressor alleles led to a strong increase of PdeL activity and elevated pdeL transcription, suggesting that enzymatic activity and transcriptional control are coupled. In agreement with this, we showed that overall cellular levels of c-di-GMP control pdeL transcription and that this control depends on PdeL itself. We thus propose that PdeL acts both as an enzyme and as a c-di-GMP sensor to couple transcriptional activity to the c-di-GMP status of the cell. IMPORTANCE Most bacteria possess multiple diguanylate cyclases and phosphodiesterases. Genetic studies have proposed that these enzymes show signaling specificity by contributing to distinct cellular processes without much cross talk. Thus, spatial

  17. Dimerization of cAMP phosphodiesterase-4 (PDE4) in living cells requires interfaces located in both the UCR1 and catalytic unit domains

    PubMed Central

    Bolger, Graeme B.; Dunlop, Allan J.; Meng, Dong; Day, Jon P.; Klussmann, Enno; Baillie, George S.; Adams, David R.; Houslay, Miles D.

    2015-01-01

    PDE4 family cAMP phosphodiesterases play a pivotal role in determining compartmentalised cAMP signalling through targeted cAMP breakdown. Expressing the widely found PDE4D5 isoform, as both bait and prey in a yeast 2-hybrid system, we demonstrated interaction consistent with the notion that long PDE4 isoforms form dimers. Four potential dimerization sites were uncovered using a scanning peptide array approach, where a recombinant purified PDE4D5 fusion protein was used to probe a 25-mer library of overlapping peptides covering the entire PDE4D5 sequence. Key residues involved in PDE4D5 dimerization were defined using a site-directed mutagenesis programme directed by an alanine scanning peptide array approach. Critical residues stabilising PDE4D5 dimerization were defined within the regulatory UCR1 region found in long, but not short, PDE4 isoforms, namely the Arg173, Asn174 and Asn175 (DD1) cluster. Disruption of the DD1 cluster was not sufficient, in itself, to destabilise PDE4D5 homodimers. Instead, disruption of an additional interface, located on the PDE4 catalytic unit, was also required to convert PDE4D5 into a monomeric form. This second dimerization site on the conserved PDE4 catalytic unit is dependent upon a critical ion pair interaction. This involves Asp463 and Arg499 in PDE4D5, which interact in a trans fashion involving the two PDE4D5 molecules participating in the homodimer. PDE4 long isoforms adopt a dimeric state in living cells that is underpinned by two key contributory interactions, one involving the UCR modules and one involving an interface on the core catalytic domain. We propose that short forms do not adopt a dimeric configuration because, in the absence of the UCR1 module, residual engagement of the remaining core catalytic domain interface provides insufficient free energy to drive dimerization. The functioning of PDE4 long and short forms is thus poised to be inherently distinct due to this difference in quaternary structure. PMID

  18. Inhibition of Phosphodiesterase 2 Augments cGMP and cAMP Signaling to Ameliorate Pulmonary Hypertension

    PubMed Central

    Bubb, Kristen J; Trinder, Sarah L; Baliga, Reshma S; Patel, Jigisha; Clapp, Lucie H; MacAllister, Raymond J; Hobbs, Adrian J

    2014-01-01

    Background Pulmonary hypertension (PH) is a life-threatening disorder characterized by increased pulmonary artery pressure, remodeling of the pulmonary vasculature, and right ventricular failure. Loss of endothelium-derived nitric oxide (NO) and prostacyclin (PGI2) contributes to PH pathogenesis and current therapies are targeted to restore these pathways. Phosphodiesterases (PDEs) are a family of enzymes that break down cGMP and cAMP which underpin the bioactivity of NO and PGI2. The PDE5 inhibitor (PDE5i) sildenafil is licensed for PH, but a role for PDE2 in lung physiology and disease has yet to be established. Herein, we investigated whether PDE2 inhibition modulates pulmonary cyclic nucleotide signaling and ameliorates experimental PH. Methods and Results The selective PDE2 inhibitor BAY 60-7550 augmented atrial natriuretic peptide (ANP) and treprostinil -evoked pulmonary vascular relaxation in isolated arteries from chronically hypoxic rats. BAY 60-7550 prevented the onset of both hypoxia- and bleomycin-induced PH, and produced a significantly greater reduction in disease severity when given in combination with a neutral endopeptidase inhibitor (enhances endogenous natriuretic peptides), the PGI2 analogue treprostinil, inorganic nitrate (NO donor), or a PDE5i. Proliferation of pulmonary artery smooth muscle cells from PAH patients was reduced by BAY 60-7550, an effect further enhanced in the presence of ANP, NO and treprostinil. Conclusions PDE2 inhibition elicits pulmonary dilation, prevents pulmonary vascular remodeling, and reduces the RVH characteristic of PH. This favorable pharmacodynamic profile is dependent on natriuretic peptide bioactivity, and is additive with PGI2 analogues, PDE5i, and NO. PDE2 inhibition represents a viable, orally-active therapy for PH. PMID:24899690

  19. Traumatic Brain Injury Upregulates Phosphodiesterase Expression in the Hippocampus

    PubMed Central

    Wilson, Nicole M.; Titus, David J.; Oliva, Anthony A.; Furones, Concepcion; Atkins, Coleen M.

    2016-01-01

    Traumatic brain injury (TBI) results in significant impairments in hippocampal synaptic plasticity. A molecule critically involved in hippocampal synaptic plasticity, 3′,5′-cyclic adenosine monophosphate, is downregulated in the hippocampus after TBI, but the mechanism that underlies this decrease is unknown. To address this question, we determined whether phosphodiesterase (PDE) expression in the hippocampus is altered by TBI. Young adult male Sprague Dawley rats received sham surgery or moderate parasagittal fluid-percussion brain injury. Animals were analyzed by western blotting for changes in PDE expression levels in the hippocampus. We found that PDE1A levels were significantly increased at 30 min, 1 h and 6 h after TBI. PDE4B2 and 4D2 were also significantly increased at 1, 6, and 24 h after TBI. Additionally, phosphorylation of PDE4A was significantly increased at 6 and 24 h after TBI. No significant changes were observed in levels of PDE1B, 1C, 3A, 8A, or 8B between 30 min to 7 days after TBI. To determine the spatial profile of these increases, we used immunohistochemistry and flow cytometry at 24 h after TBI. PDE1A and phospho-PDE4A localized to neuronal cell bodies. PDE4B2 was expressed in neuronal dendrites, microglia and infiltrating CD11b+ immune cells. PDE4D was predominantly found in microglia and infiltrating CD11b+ immune cells. To determine if inhibition of PDE4 would improve hippocampal synaptic plasticity deficits after TBI, we treated hippocampal slices with rolipram, a pan-PDE4 inhibitor. Rolipram partially rescued the depression in basal synaptic transmission and converted a decaying form of long-term potentiation (LTP) into long-lasting LTP. Overall, these results identify several possible PDE targets for reducing hippocampal synaptic plasticity deficits and improving cognitive function acutely after TBI. PMID:26903822

  20. Traumatic Brain Injury Upregulates Phosphodiesterase Expression in the Hippocampus.

    PubMed

    Wilson, Nicole M; Titus, David J; Oliva, Anthony A; Furones, Concepcion; Atkins, Coleen M

    2016-01-01

    Traumatic brain injury (TBI) results in significant impairments in hippocampal synaptic plasticity. A molecule critically involved in hippocampal synaptic plasticity, 3',5'-cyclic adenosine monophosphate, is downregulated in the hippocampus after TBI, but the mechanism that underlies this decrease is unknown. To address this question, we determined whether phosphodiesterase (PDE) expression in the hippocampus is altered by TBI. Young adult male Sprague Dawley rats received sham surgery or moderate parasagittal fluid-percussion brain injury. Animals were analyzed by western blotting for changes in PDE expression levels in the hippocampus. We found that PDE1A levels were significantly increased at 30 min, 1 h and 6 h after TBI. PDE4B2 and 4D2 were also significantly increased at 1, 6, and 24 h after TBI. Additionally, phosphorylation of PDE4A was significantly increased at 6 and 24 h after TBI. No significant changes were observed in levels of PDE1B, 1C, 3A, 8A, or 8B between 30 min to 7 days after TBI. To determine the spatial profile of these increases, we used immunohistochemistry and flow cytometry at 24 h after TBI. PDE1A and phospho-PDE4A localized to neuronal cell bodies. PDE4B2 was expressed in neuronal dendrites, microglia and infiltrating CD11b(+) immune cells. PDE4D was predominantly found in microglia and infiltrating CD11b(+) immune cells. To determine if inhibition of PDE4 would improve hippocampal synaptic plasticity deficits after TBI, we treated hippocampal slices with rolipram, a pan-PDE4 inhibitor. Rolipram partially rescued the depression in basal synaptic transmission and converted a decaying form of long-term potentiation (LTP) into long-lasting LTP. Overall, these results identify several possible PDE targets for reducing hippocampal synaptic plasticity deficits and improving cognitive function acutely after TBI. PMID:26903822

  1. Characterization of inhibitors of phosphodiesterase 1C on a human cellular system.

    PubMed

    Dunkern, Torsten R; Hatzelmann, Armin

    2007-09-01

    Different inhibitors of the Ca(2+)/calmodulin-stimulated phosphodiesterase 1 family have been described and used for the examination of phosphodiesterase 1 in cellular, organ or animal models. However, the inhibitors described differ in potency and selectivity for the different phosphodiesterase family enzymes, and in part exhibit additional pharmacodynamic actions. In this study, we demonstrate that phosphodiesterase 1C is expressed in the human glioblastoma cell line A172 with regard to mRNA, protein and activity level, and that lower activities of phosphodiesterase 2, phosphodiesterase 3, phosphodiesterase 4 and phosphodiesterase 5 are also present. The identity of the phosphodiesterase 1C activity detected was verified by downregulation of the mRNA and protein through human phosphodiesterase 1C specific small interfering RNA. In addition, the measured K(m) values (cAMP, 1.7 microm; cGMP, 1.3 microm) are characteristic of phosphodiesterase 1C. We demonstrate that treatment with the Ca(2+) ionophore ionomycin increases intracellular Ca(2+) in a concentration-dependent way without affecting cell viability. Under conditions of enhanced intracellular Ca(2+) concentration, a rapid increase in cAMP levels caused by the adenylyl cyclase activator forskolin was abolished, indicating the involvement of Ca(2+)-activated phosphodiesterase 1C. The reduction of forskolin-stimulated cAMP levels was reversed by phosphodiesterase 1 inhibitors in a concentration-dependent way. Using this cellular system, we compared the cellular potency of published phosphodiesterase 1 inhibitors, including 8-methoxymethyl-3-isobutyl-1-methylxanthine, vinpocetine, SCH51866, and two established phosphodiesterase 1 inhibitors developed by Schering-Plough (named compounds 31 and 30). We demonstrate that up to 10 microm 8-methoxymethyl-3-isobutyl-1-methylxanthine and vinpocetine had no effect on the reduction of forskolin-stimulated cAMP levels by ionomycin, whereas the more selective and up to 10

  2. Challenge of human Jurkat T-cells with the adenylate cyclase activator forskolin elicits major changes in cAMP phosphodiesterase (PDE) expression by up-regulating PDE3 and inducing PDE4D1 and PDE4D2 splice variants as well as down-regulating a novel PDE4A splice variant.

    PubMed Central

    Erdogan, S; Houslay, M D

    1997-01-01

    The cAMP phosphodiesterase (PDE) 3 and PDE4 isoforms provide the major cAMP-hydrolysing PDE activities in Jurkat T-cells, with additional contributions from the PDE1 and PDE2 isoforms. Challenge of cells with the adenylate cyclase activator forskolin led to a rapid, albeit transient, increase in PDE3 activity occurring over the first 45 min, followed by a sustained increase in PDE3 activity which began after approximately 3 h and continued for at least 24 h. Only this second phase of increase in PDE3 activity was blocked by the transcriptional inhibitor actinomycin D. After approximately 3 h of exposure to forskolin, PDE4 activity had increased, via a process that could be inhibited by actinomycin D, and it remained elevated for at least a 24 h period. Such actions of forskolin were mimicked by cholera toxin and 8-bromo-cAMP. Forskolin increased intracellular cAMP concentrations in a time-dependent fashion and its action was enhanced when PDE induction was blocked with actinomycin D. Reverse transcription (RT)-PCR analysis, using generic primers designed to detect transcripts representing enzymically active products of the four PDE4 genes, identified transcripts for PDE4A and PDE4D but not for PDE4B or PDE4C in untreated Jurkat T-cells. Forskolin treatment did not induce transcripts for either PDE4B or PDE4C; however, it reduced the RT-PCR signal for PDE4A transcripts and markedly enhanced that for PDE4D transcripts. Using RT-PCR primers for PDE4 splice variants, a weak signal for PDE4D1 was evident in control cells whereas, in forskolin-treated cells, clear signals for both PDE4D1 and PDE4D2 were detected. RT-PCR analysis of the PDE4A species indicated that it was not the PDE4A isoform PDE-46 (PDE4A4B). Immunoblotting of control cells for PDE4 forms identified a single PDE4A species of approximately 118 kDa, which migrated distinctly from the PDE4A4B isoform PDE-46, with immunoprecipitation analyses showing that it provided all of the PDE4 activity in control

  3. β-Adrenergic receptor stimulation increases surface NKCC2 expression in rat thick ascending limbs in a process inhibited by phosphodiesterase 4.

    PubMed

    Haque, Mohammed Z; Caceres, Paulo S; Ortiz, Pablo A

    2012-11-01

    The thick ascending limb of the loop of Henle (THAL) reabsorbs ∼30% of the filtered NaCl in a process mediated by the apical Na-K-2Cl cotransporter NKCC2. Stimulation of β-adrenergic receptors in the THAL enhances NaCl reabsorption and increases intracellular cAMP. We found that intracellular cAMP stimulates NKCC2 trafficking to the apical membrane via protein kinase A (PKA). Several cAMP-specific phosphodiesterases (PDE) have been identified in rat THALs, and PDE4 decreases cAMP generated by β-adrenergic stimulation in other cells. However, it is not known whether β-adrenergic receptors activation stimulates NKCC2 trafficking. Thus we hypothesized that β-adrenergic receptor stimulation enhances THAL apical membrane NKCC2 expression via the PKA pathway and PDE4 blunts this effect. THAL suspensions were obtained from Sprague-Dawley rats, and surface NKCC2 expression was measured by surface biotinylation and Western blot. Incubation of THALs with the β-adrenergic receptor agonist isoproterenol at 0.5 and 1.0 μM increased surface NKCC2 by 17 ± 1 and 29 ± 5% respectively (P < 0.05). Preventing cAMP degradation with 3-isobutyl-methylxanthine (IBMX; a nonselective phosphodiesterase inhibitor) enhanced isoproterenol-stimulated surface NKCC2 expression to 51 ± 7% (P < 0.05 vs. isoproterenol). The β-adrenergic receptor antagonist propranolol or the PKA inhibitor H-89 completely blocked isoproterenol + IBMX-induced increase on surface NKCC2, while propranolol or H-89 alone had no effect. Selective inhibition of PDE4 with rolipram (20 μM) potentiated the effect of isoproterenol on surface NKCC2 and increased cAMP levels. We concluded that β-adrenergic receptor stimulation enhances surface NKCC2 expression in the THALs via PKA and PDE4 blunts this effect. PMID:22933300

  4. Iloprost- and isoproterenol-induced increases in cAMP are regulated by different phosphodiesterases in erythrocytes of both rabbits and humans

    PubMed Central

    Adderley, Shaquria P.; Dufaux, Eileen A.; Sridharan, Meera; Bowles, Elizabeth A.; Hanson, Madelyn S.; Stephenson, Alan H.; Ellsworth, Mary L.; Sprague, Randy S.

    2009-01-01

    Activation of the G protein Gs results in increases in cAMP, a necessary step in the pathway for ATP release from rabbit and human erythrocytes. In all cells, the level of cAMP is the product of its synthesis by adenylyl cyclase and its hydrolysis by phosphodiesterases (PDEs). Both iloprost (Ilo), a PGI2 analog, and isoproterenol (Iso), a β-agonist, stimulate receptor-mediated increases in cAMP in rabbit and human erythrocytes. However, the specific PDEs associated with each of these signaling pathways in the erythrocyte have not been fully characterized. Previously, we reported that PDE3B is present in rabbit and human erythrocyte membranes and that PDE3 inhibitors potentiate Ilo-induced increases in cAMP. Here we report that inhibitors of either PDE2 or PDE4, erythro-9-(2-hydroxy-3-nonyl)adenine (EHNA) and rolipram, respectively, potentiate Iso-induced increases in cAMP in rabbit and human erythrocytes. Importantly, these inhibitors had no effect on cAMP increases associated with the incubation of erythrocytes with Ilo. In addition, we establish, for the first time, the presence of PDE2A protein in rabbit and human erythrocyte membranes. Finally, we determined that preincubation of human erythrocytes with EHNA and rolipram together potentiate Iso-induced ATP release, whereas preincubation with cilostazol enhances Ilo-induced release of ATP. These results are consistent with the hypothesis that, in rabbit and human erythrocytes, Ilo-induced increases in cAMP and ATP release are regulated by PDE3, whereas those associated with Iso are regulated by the activities of both PDE2 and PDE4. These studies demonstrate that PDE activity in these cells is localized to specific signaling pathways. PMID:19252089

  5. Acute hypoxia modifies cAMP levels induced by inhibitors of phosphodiesterase-4 in rat carotid bodies, carotid arteries and superior cervical ganglia

    PubMed Central

    Nunes, Ana R; Batuca, Joana R; Monteiro, Emília C

    2010-01-01

    Background and purpose: Phosphodiesterase (PDE) inhibitors are useful to treat hypoxia-related diseases and are used in experiments studying the effects of oxygen on 3′-5′-cyclic adenosine monophosphate (cAMP) production. We studied the effects of acute hypoxia on cAMP accumulation induced by PDE inhibitors in oxygen-specific chemosensors, the carotid bodies (CBs) and in non-chemosensitive CB-related structures: carotid arteries (CAs) and superior cervical ganglia (SCG). Experimental approach: Concentration–response curves for the effects of a non-specific PDE inhibitor [isobutylmethylxanthine (IBMX) ], PDE4 selective inhibitors (rolipram, Ro 20-1724) and a PDE2 selective inhibitor (erythro-9-(2-hydroxy-3-nonyl)adenine) on cAMP levels were obtained in normoxic (20% O2/5% CO2) or hypoxic (5% O2/5% CO2) conditions. Key results: Responses to the PDE inhibitors were compatible with the presence of PDE4 in rat CBs, CAs and SCG but in the absence of PDE2 in CAs and CBs. Acute hypoxia enhanced the effects of IBMX and PDE4 inhibitors on cAMP accumulation in CAs and CBs. In SCG, acute hypoxia reduced cAMP accumulation induced by all the four PDE inhibitors tested. Differences between the effects of Ro 20-1724 and rolipram on cAMP were found in CAs and CBs during hypoxia. Conclusions and implications: The effects of PDE4 inhibitors could be potentiated or inhibited by acute hypoxia depending on the PDE isoforms of the tissue. The similarities between the characterization of PDE4 inhibitors at the CBs and CAs, under normoxia and hypoxia, did not support a specific role for cAMP in the oxygen-sensing machinery at the CB and suggested that no direct CB-mediated, hyperventilatory, adverse effects would be expected with administration of PDE4 inhibitors. PMID:20082613

  6. Role of cyclic nucleotide phosphodiesterase isoforms in cAMP compartmentation following β2-adrenergic stimulation of ICa,L in frog ventricular myocytes

    PubMed Central

    Jurevičius, Jonas; Skeberdis, V Arvydas; Fischmeister, Rodolphe

    2003-01-01

    The role of cyclic nucleotide phosphodiesterase (PDE) isoforms in the β2-adrenergic stimulation of the L-type Ca2+ current (ICa,L) was investigated in frog ventricular myocytes using double patch-clamp and double-barrelled microperfusion techniques. Isoprenaline (ISO, 1 nM to 10 μM) was applied on one half of the cell, either alone or in the presence of PDE inhibitors, and the local and distant responses of ICa,L were used to determine the gradient of local vs. distant cAMP concentration (α). IBMX (100 μM), a non-selective PDE inhibitor, reduced α from 40 to 4.4 indicating a 9-fold reduction in intracellular cAMP compartmentation when all PDE activity was blocked. While PDE1 and PDE2 inhibition had no effect, PDE3 inhibition by milrinone (3 μM) or PDE4 inhibition by Ro 20-1724 (3 μM) reduced α by 6- and 4-fold, respectively. A simultaneous application of milrinone and Ro 20-1724 produced a similar effect to IBMX, showing that PDE3 and PDE4 were the major PDEs accounting for cAMP compartmentation. Okadaic acid (3 μM), a non-selective phosphatase inhibitor, or H89 (1 μM), an inhibitor of cAMP-dependent protein kinase (PKA), had no effect on the distant response of ICa,L to ISO indicating that PDE activation by PKA played a minor role in cAMP compartmentation. Our results demonstrate that PDE activity determines the degree of cAMP compartmentation in frog ventricular cells upon β2-adrenergic stimulation. PDE3 and PDE4 subtypes play a major role in this process, and contribute equally to ensure a functional coupling of β2-adrenergic receptors with nearby Ca2+ channels via local elevations of cAMP. PMID:12815180

  7. Role of phosphodiesterase 4 expression in the Epac1 signaling-dependent skeletal muscle hypertrophic action of clenbuterol.

    PubMed

    Ohnuki, Yoshiki; Umeki, Daisuke; Mototani, Yasumasa; Shiozawa, Kouichi; Nariyama, Megumi; Ito, Aiko; Kawamura, Naoya; Yagisawa, Yuka; Jin, Huiling; Cai, Wenqian; Suita, Kenji; Saeki, Yasutake; Fujita, Takayuki; Ishikawa, Yoshihiro; Okumura, Satoshi

    2016-05-01

    Clenbuterol (CB), a selective β2-adrenergic receptor (AR) agonist, induces muscle hypertrophy and counteracts muscle atrophy. However, it is paradoxically less effective in slow-twitch muscle than in fast-twitch muscle, though slow-twitch muscle has a greater density of β-AR We recently demonstrated that Epac1 (exchange protein activated by cyclic AMP [cAMP]1) plays a pivotal role in β2-AR-mediated masseter muscle hypertrophy through activation of the Akt and calmodulin kinase II (CaMKII)/histone deacetylase 4 (HDAC4) signaling pathways. Here, we investigated the role of Epac1 in the differential hypertrophic effect of CB using tibialis anterior muscle (TA; typical fast-twitch muscle) and soleus muscle (SOL; typical slow-twitch muscle) of wild-type (WT) and Epac1-null mice (Epac1KO). The TA mass to tibial length (TL) ratio was similar in WT and Epac1KO at baseline and was significantly increased after CB infusion in WT, but not in Epac1KO The SOL mass to TL ratio was also similar in WT and Epac1KO at baseline, but CB-induced hypertrophy was suppressed in both mice. In order to understand the mechanism involved, we measured the protein expression levels of β-AR signaling-related molecules, and found that phosphodiesterase 4 (PDE4) expression was 12-fold greater in SOL than in TA These results are consistent with the idea that increased PDE4-mediated cAMP hydrolysis occurs in SOL compared to TA, resulting in a reduced cAMP concentration that is insufficient to activate Epac1 and its downstream Akt and CaMKII/HDAC4 hypertrophic signaling pathways in SOL of WT This scenario can account for the differential effects of CB on fast- and slow-twitch muscles. PMID:27207782

  8. Loss of phosphodiesterase 10A expression is associated with progression and severity in Parkinson's disease.

    PubMed

    Niccolini, Flavia; Foltynie, Thomas; Reis Marques, Tiago; Muhlert, Nils; Tziortzi, Andri C; Searle, Graham E; Natesan, Sridhar; Kapur, Shitij; Rabiner, Eugenii A; Gunn, Roger N; Piccini, Paola; Politis, Marios

    2015-10-01

    The mechanisms underlying neurodegeneration and loss of dopaminergic signalling in Parkinson's disease are still only partially understood. Phosphodiesterase 10A (PDE10A) is a basal ganglia expressed dual substrate enzyme, which regulates cAMP and cGMP signalling cascades, thus having a key role in the regulation of dopaminergic signalling in striatal pathways, and in promoting neuronal survival. This study aimed to assess in vivo the availability of PDE10A in patients with Parkinson's disease using positron emission tomography molecular imaging with (11)C-IMA107, a highly selective PDE10A radioligand. We studied 24 patients with levodopa-treated, moderate to advanced Parkinson's disease. Their positron emission tomography imaging data were compared to those from a group of 12 healthy controls. Parametric images of (11)C-IMA107 binding potential relative to non-displaceable binding (BPND) were generated from the dynamic (11)C-IMA107 scans using the simplified reference tissue model with the cerebellum as the reference tissue. Corresponding region of interest analysis showed lower mean (11)C-IMA107 BPND in the caudate (P < 0.001), putamen (P < 0.001) and globus pallidus (P = 0.025) in patients with Parkinson's disease compared to healthy controls, which was confirmed with voxel-based analysis. Longer Parkinson's duration correlated with lower (11)C-IMA107 BPND in the caudate (r = -0.65; P = 0.005), putamen (r = -0.51; P = 0.025), and globus pallidus (r = -0.47; P = 0.030). Higher Unified Parkinson's Disease Rating Scale part-III motor scores correlated with lower (11)C-IMA107 BPND in the caudate (r = -0.54; P = 0.011), putamen (r = -0.48; P = 0.022), and globus pallidus (r = -0.70; P < 0.001). Higher Unified Dyskinesia Rating Scale scores in those Parkinson's disease with levodopa-induced dyskinesias (n = 12), correlated with lower (11)C-IMA107 BPND in the caudate (r = -0.73; P = 0.031) and putamen (r = -0.74; P = 0.031). Our findings demonstrate striatal and

  9. Novel alternative splice variants of rat phosphodiesterase 7B showing unique tissue-specific expression and phosphorylation.

    PubMed Central

    Sasaki, Takashi; Kotera, Jun; Omori, Kenji

    2002-01-01

    cDNA species coding for novel variants of cyclic-AMP-specific phosphodiesterases (PDEs), namely the PDE7B family, were isolated from rats and characterized. Rat PDE7B1 (RNPDE7B1) was composed of 446 amino acid residues. Rat PDE7B2 (RNPDE7B2) and PDE7B3 (RNPDE7B3), which possessed unique N-terminal sequences, consisted of 359 and 459 residues respectively. Northern hybridization analysis showed that rat PDE7B transcripts were particularly abundant in the striatum and testis. PCR analyses revealed that rat PDE7B2 transcripts were restricted to the testis and that low levels of PDE7B3 transcripts were expressed in the heart, lung and skeletal muscle. In situ hybridization analysis demonstrated that rat PDE7B transcripts were expressed in striatal neurons and spermatocytes. In spermatocytes, rat PDE7B transcripts were expressed in a stage-specific manner during spermatogenesis. The K(m) values of recombinant rat PDE7B1, PDE7B2 and PDE7B3 for cAMP were 0.05, 0.07 and 0.05 microM respectively. Each rat PDE7B variant was the most sensitive to 3-isobutyl-1-methylxanthine (IC(50) 1.5-2.1 microM). Two phosphorylation sites for cAMP-dependent protein kinase (PKA) were found in rat PDE7B1 and PDE7B3, whereas rat PDE7B2 possessed one site. PKA-dependent phosphorylation was observed in C-terminal phosphorylation sites of three rat PDE7B variants, in addition to unique N-terminal regions of rat PDE7B1 and PDE7B3. Unique tissue distribution and PKA-dependent phosphorylation of PDE7B variants suggested that each variant has a specific role for cellular functions via cAMP signalling in various tissues. PMID:11772393

  10. Pregnancy induces a modulation of the cAMP phosphodiesterase 4-conformers ratio in human myometrium: consequences for the utero-relaxant effect of PDE4-selective inhibitors.

    PubMed

    Méhats, C; Tanguy, G; Paris, B; Robert, B; Pernin, N; Ferré, F; Leroy, M J

    2000-02-01

    The inhibitory impacts of RP 73401, a phosphodiesterase type 4 (PDE4) selective inhibitor of the second generation, versus rolipram, the prototypal PDE4 inhibitor, were evaluated and compared on cAMP phosphodiesterase (PDE) activity and contractility of the myometrium in nonpregnant and pregnant women. In enzymatic studies, RP 73401 and rolipram inhibited the cAMP PDE activity with significantly greater maximal efficiency in the myometrium of pregnant compared with nonpregnant women (75 versus 55%; P <.05). Although myometrial PDE4 presented a single class of interaction with RP 73401 [pD(2) (-log [IC(50)]) = -8.2], it exhibited at least two classes of interaction with rolipram (pD(2) = -8.2 and -5.6). In the myometrium of pregnant versus nonpregnant women, rolipram is significantly more efficacious in the concentration range >0.01 to 100 microM (P <.01), whereas no difference was observed for the concentration range <0.01 microM. In contractility studies, RP 73401 was equally effective in relaxing myometrial strips from both nonpregnant and pregnant women (pD(2) = -8.8). Conversely, the ability of rolipram to inhibit contractions of the myometrium in pregnant women was significantly lower (pD(2) = -7.2) compared with that in nonpregnant women (pD(2) = -8.2; P <.01). Concomitantly, in the myometrium of pregnant women, a rise in immunoreactive PDE4B2 signal was detected, whereas the PDE4D3 signal was less intense. These results demonstrate that parallel to an accumulation of PDE4B2 isoform, a modification in the ratio of PDE4 conformers HPDE4 and LPDE4 (conformer that binds rolipram with high and low affinity, respectively) occurs in the myometrium of near-term pregnant women with an increase of LPDE4 functionally implicated in the contractile process. Such modifications provide a strong rationale to propose LPDE4 as potential pharmacologic targets for the design of new tocolytic treatments. PMID:10640323

  11. Identification and characterization of the type-IVA cyclic AMP-specific phosphodiesterase RD1 as a membrane-bound protein expressed in cerebellum.

    PubMed

    Shakur, Y; Wilson, M; Pooley, L; Lobban, M; Griffiths, S L; Campbell, A M; Beattie, J; Daly, C; Houslay, M D

    1995-03-15

    An antiserum was generated against a dodecapeptide whose sequence is found at the C-terminus of a cyclic AMP (cAMP)-specific, type-IVA phosphodiesterase encoded by the rat 'dunc-like' cyclic AMP phosphodiesterase (RD1) cDNA. This antiserum identified a single approximately 73 kDa protein species upon immunoblotting of cerebellum homogenates. This species co-migrated upon SDS/PAGE with a single immunoreactive species observed in COS cells transfected with the cDNA for RD1. Native RD1 in cerebellum was found to be predominantly (approximately 93%) membrane-associated and could be found in isolated synaptosome populations, in particular those enriched in post-synaptic densities. Fractionation of lysed synaptosomes on sucrose density gradients identified RD1 as co-migrating with the plasma membrane marker 5'-nucleotidase. Laser scanning confocal and digital deconvolution immunofluorescence studies done on intact COS cells transfected with RD1 cDNA showed RD1 to be predominantly localized to plasma membranes but also associated with the Golgi apparatus and intracellular vesicles. RD1-specific antisera immunoprecipitated phosphodiesterase activity from solubilized cerebellum membranes. This activity had the characteristics expected of the type-IV cAMP phosphodiesterase RD1 in that it was cAMP specific, exhibited a low Km cAMP of 2.3 microM, high sensitivity to inhibition by 4-[3-(cyclopentoxyl)-4-methoxyphenyl]-2-pyrrolidone (rolipram) (Ki approximately 0.7 microM) and was unaffected by Ca2+/calmodulin and low concentrations of cyclic GMP. The phosphodiesterase activities of RD1 solubilized from both cerebellum and transfected COS cell membranes showed identical first-order thermal denaturation kinetics at 50 degrees C. Native RD1 from cerebellum was shown to be an integral protein in that it was solubilized using the non-ionic detergent Triton X-100 but not by either re-homogenization or high NaCl concentrations. The observation that hydroxylamine was unable to cause

  12. Angiotensin-converting enzyme inhibition prevents myocardial infarction-induced increase in renal cortical cGMP and cAMP phosphodiesterase activities.

    PubMed

    Clauss, François; Charloux, Anne; Piquard, François; Doutreleau, Stéphane; Talha, Samy; Zoll, Joffrey; Lugnier, Claire; Geny, Bernard

    2015-08-01

    We investigated whether myocardial infarction (MI) enhances renal phosphodiesterases (PDE) activities, investigating particularly the relative contribution of PDE1-5 isozymes in total PDE activity involved in both cGMP and cAMP pathways, and whether angiotensin-converting enzyme inhibition (ACEi) decreases such renal PDE hyperactivities. We also investigated whether ACEi might thereby improve atrial natriuretic peptide (ANP) efficiency. We studied renal cortical PDE1-5 isozyme activities in sham (SH)-operated, MI rats and in MI rats treated with perindopril (ACEi) 1 month after coronary artery ligation. Circulating atrial natriuretic peptide (ANP), its second intracellular messenger cyclic guanosine monophosphate (cGMP) and cGMP/ANP ratio were also determined. Cortical cGMP-PDE2 (80.3 vs. 65.1 pmol/min/mg) and cGMP-PDE1 (50.7 vs. 30.1 pmol/min/mg), and cAMP-PDE2 (161 vs. 104.1 pmol/min/mg) and cAMP-PDE4 (307.5 vs. 197.2 pmol/min/mg) activities were higher in MI than in SH rats. Despite increased ANP plasma level, ANP efficiency tended to be decreased in MI compared to SH rats. Perindopril restored PDE activities and tended to improve ANP efficiency in MI rats. One month after coronary ligation, perindopril treatment of MI rats prevents the increase in renal cortical PDE activities. This may contribute to increase renal ANP efficiency in MI rats. PMID:25939307

  13. Evidence that insulin and isoprenaline activate the cGMP-inhibited low-Km cAMP phosphodiesterase in rat fat cells by phosphorylation

    SciTech Connect

    Degerman, E.; Smith, C.J.; Tornqvist, H.; Vasta, V.; Belfrage, P.; Manganiello, V.C. )

    1990-01-01

    Incubation of intact rat fat cells with maximally effective concentrations of insulin (1 nM, 12 min) or isoprenaline (300 nM, 3 min) increased particulate cGMP- and cilostamide-inhibited, low-Km cAMP phosphodiesterase (cAMP-PDE) activity by about 50% and 100%, respectively. In 32P-labeled cells, these agents induced serine 32P-phosphorylation of a 135-kDa particulate protein and, to a variable and lesser extent, a 44-kDa protein, which were selectively immunoprecipitated by anti-cAMP-PDE, as analyzed by SDS/PAGE and autoradiography. In the absence of hormonal stimulation, little phosphorylation was detected (less than 10% of that with the hormones). The two phosphoproteins were identified as cAMP-PDE or a closely related molecule (in the case of the 44-kDa species, perhaps a proteolytic fragment) since (i) amounts of 32P in the immunoprecipitated 135-kDa protein paralleled enzyme inactivation, (ii) prior incubation of the anti-cAMP-PDE with the pure rat or bovine enzyme selectively blocked the immunoprecipitation of the phosphoproteins, (iii) 135- and 44-kDa proteins reacted with the anti-cAMP-PDE on Western immunoblots, and (iv) the two phosphoproteins copurified with cAMP-PDE activity through DEAE-Sephacel chromatography and were isolated by highly selective affinity chromatography on cilostamide-agarose. Thus, in fat cells, catecholamine- and insulin-induced activation of the cAMP-PDE may be mediated via phosphorylation by cAMP-dependent protein kinase and an insulin-activated serine protein kinase, respectively.

  14. Dysregulation of hepatic cAMP levels via altered Pde4b expression plays a critical role in alcohol-induced steatosis.

    PubMed

    Avila, Diana V; Barker, David F; Zhang, JingWen; McClain, Craig J; Barve, Shirish; Gobejishvili, Leila

    2016-09-01

    Alcohol-induced hepatic steatosis is a significant risk factor for progressive liver disease. Cyclic adenosine monophosphate (cAMP) signalling has been shown to significantly regulate lipid metabolism; however, the role of altered cAMP homeostasis in alcohol-mediated hepatic steatosis has never been studied. Our previous work demonstrated that increased expression of hepatic phosphodiesterase 4 (Pde4), which specifically hydrolyses and decreases cAMP levels, plays a pathogenic role in the development of liver inflammation/injury. The aim of this study was to examine the role of PDE4 in alcohol-induced hepatic steatosis. C57BL/6 wild-type and Pde4b knockout (Pde4b(-/-) ) mice were pair-fed control or ethanol liquid diets. One group of wild-type mice received rolipram, a PDE4-specific inhibitor, during alcohol feeding. We demonstrate for the first time that an early increase in PDE4 enzyme expression and a resultant decrease in hepatic cAMP levels are associated with the significant reduction in carnitine palmitoyltransferase 1A (Cpt1a) expression. Notably, alcohol-fed (AF) Pde4b(-/-) mice and AF wild-type mice treated with rolipram had significantly lower hepatic free fatty acid content compared with AF wild-type mice. Importantly, PDE4 inhibition in alcohol-fed mice prevented the decrease in hepatic Cpt1a expression via the Pparα/Sirt1/Pgc1α pathway. These results demonstrate that the alcohol- induced increase in hepatic Pde4, specifically Pde4b expression, and compromised cAMP signalling predispose the liver to impaired fatty acid oxidation and the development of steatosis. Moreover, these data also suggest that hepatic PDE4 may be a clinically relevant therapeutic target for the treatment of alcohol-induced hepatic steatosis. Copyright © 2016 Pathological Society of Great Britain and Ireland. Published by John Wiley & Sons, Ltd. PMID:27287961

  15. Hydrolysis of N-methyl-D-aspartate receptor-stimulated cAMP and cGMP by PDE4 and PDE2 phosphodiesterases in primary neuronal cultures of rat cerebral cortex and hippocampus.

    PubMed

    Suvarna, Neesha U; O'Donnell, James M

    2002-07-01

    Stimulation of N-methyl-D-aspartate (NMDA) receptors on neurons activates both cAMP and cGMP signaling pathways. Experiments were carried out to determine which phosphodiesterase (PDE) families are involved in the hydrolysis of the cyclic nucleotides formed via this mechanism, using primary neuronal cultures prepared from rat cerebral cortex and hippocampus. The nonselective PDE inhibitor 3-isobutyl-1-methylxanthine (IBMX) potentiated the ability of NMDA to increase cAMP and cGMP. However, among the family-selective inhibitors, only the PDE4 inhibitor rolipram enhanced the ability of NMDA to increase cAMP in the neurons. In contrast, only the PDE2 inhibitor erythro-9-(2-hydroxy-3-nonyl) adenine (EHNA) enhanced the ability of NMDA to increase cGMP. Neither adenosine nor an adenosine deaminase inhibitor mimicked the effect of EHNA; this suggests that EHNA's inhibition of PDE2, not its effects on adenosine metabolism, mediates its effects on NMDA-stimulated cGMP concentrations. The PDE inhibitor-augmented effects of NMDA on cAMP and cGMP formation were antagonized by 5-methyl-10,11-dihydro-5H-dibenzo[a,d] cyclohepten-5,10-imine maleate (MK-801), verifying NMDA receptor mediation. In contrast, only NMDA-mediated cGMP formation was affected by altering either nitric oxide signaling or guanylyl cyclase; this suggests that NMDA-induced changes in cAMP are not secondary to altered cGMP concentrations. Overall, the present findings indicate that cAMP and cGMP formed in neurons as a result of NMDA receptor stimulation are hydrolyzed by PDE4 and PDE2, respectively. Selective inhibitors of the two PDE families will differentially affect the functional consequences of activation of these two signaling pathways by NMDA receptor stimulation. PMID:12065724

  16. Sphingomyelinase-Like Phosphodiesterase 3b Expression Levels Determine Podocyte Injury Phenotypes in Glomerular Disease

    PubMed Central

    Yoo, Tae-Hyun; Pedigo, Christopher E.; Guzman, Johanna; Correa-Medina, Mayrin; Wei, Changli; Villarreal, Rodrigo; Mitrofanova, Alla; Leclercq, Farah; Faul, Christian; Li, Jing; Kretzler, Matthias; Nelson, Robert G.; Lehto, Markku; Forsblom, Carol; Groop, Per-Henrik; Reiser, Jochen; Burke, George William

    2015-01-01

    Diabetic kidney disease (DKD) is the most common cause of ESRD in the United States. Podocyte injury is an important feature of DKD that is likely to be caused by circulating factors other than glucose. Soluble urokinase plasminogen activator receptor (suPAR) is a circulating factor found to be elevated in the serum of patients with FSGS and causes podocyte αVβ3 integrin-dependent migration in vitro. Furthermore, αVβ3 integrin activation occurs in association with decreased podocyte-specific expression of acid sphingomyelinase-like phosphodiesterase 3b (SMPDL3b) in kidney biopsy specimens from patients with FSGS. However, whether suPAR-dependent αVβ3 integrin activation occurs in diseases other than FSGS and whether there is a direct link between circulating suPAR levels and SMPDL3b expression in podocytes remain to be established. Our data indicate that serum suPAR levels are also elevated in patients with DKD. However, unlike in FSGS, SMPDL3b expression was increased in glomeruli from patients with DKD and DKD sera-treated human podocytes, where it prevented αVβ3 integrin activation by its interaction with suPAR and led to increased RhoA activity, rendering podocytes more susceptible to apoptosis. In vivo, inhibition of acid sphingomyelinase reduced proteinuria in experimental DKD but not FSGS, indicating that SMPDL3b expression levels determined the podocyte injury phenotype. These observations suggest that SMPDL3b may be an important modulator of podocyte function by shifting suPAR-mediated podocyte injury from a migratory phenotype to an apoptotic phenotype and that it represents a novel therapeutic glomerular disease target. PMID:24925721

  17. Sphingomyelinase-like phosphodiesterase 3b expression levels determine podocyte injury phenotypes in glomerular disease.

    PubMed

    Yoo, Tae-Hyun; Pedigo, Christopher E; Guzman, Johanna; Correa-Medina, Mayrin; Wei, Changli; Villarreal, Rodrigo; Mitrofanova, Alla; Leclercq, Farah; Faul, Christian; Li, Jing; Kretzler, Matthias; Nelson, Robert G; Lehto, Markku; Forsblom, Carol; Groop, Per-Henrik; Reiser, Jochen; Burke, George William; Fornoni, Alessia; Merscher, Sandra

    2015-01-01

    Diabetic kidney disease (DKD) is the most common cause of ESRD in the United States. Podocyte injury is an important feature of DKD that is likely to be caused by circulating factors other than glucose. Soluble urokinase plasminogen activator receptor (suPAR) is a circulating factor found to be elevated in the serum of patients with FSGS and causes podocyte αVβ3 integrin-dependent migration in vitro. Furthermore, αVβ3 integrin activation occurs in association with decreased podocyte-specific expression of acid sphingomyelinase-like phosphodiesterase 3b (SMPDL3b) in kidney biopsy specimens from patients with FSGS. However, whether suPAR-dependent αVβ3 integrin activation occurs in diseases other than FSGS and whether there is a direct link between circulating suPAR levels and SMPDL3b expression in podocytes remain to be established. Our data indicate that serum suPAR levels are also elevated in patients with DKD. However, unlike in FSGS, SMPDL3b expression was increased in glomeruli from patients with DKD and DKD sera-treated human podocytes, where it prevented αVβ3 integrin activation by its interaction with suPAR and led to increased RhoA activity, rendering podocytes more susceptible to apoptosis. In vivo, inhibition of acid sphingomyelinase reduced proteinuria in experimental DKD but not FSGS, indicating that SMPDL3b expression levels determined the podocyte injury phenotype. These observations suggest that SMPDL3b may be an important modulator of podocyte function by shifting suPAR-mediated podocyte injury from a migratory phenotype to an apoptotic phenotype and that it represents a novel therapeutic glomerular disease target. PMID:24925721

  18. Uncovering the function of Disrupted in Schizophrenia 1 through interactions with the cAMP phosphodiesterase PDE4: Contributions of the Houslay lab to molecular psychiatry.

    PubMed

    Brandon, Nicholas J

    2016-07-01

    Nearly 10years ago the laboratory of Miles Houslay was part of a collaboration which identified and characterized the interaction between Disrupted in Schizophrenia 1 and phosphodiesterase type 4. This work has had significant impact on our thinking of psychiatric illness causation and the potential for therapeutics. PMID:26432168

  19. Phosphodiesterase 1A Modulates Cystogenesis in Zebrafish

    PubMed Central

    Ward, Christopher J.; Leightner, Amanda C.; Smith, Jordan L.; Agarwal, Reema; Harris, Peter C.; Torres, Vicente E.

    2014-01-01

    Substantial evidence indicates the importance of elevated cAMP in polycystic kidney disease (PKD). Accumulation of cAMP in cystic tissues may be, in part, caused by enhanced adenylyl cyclase activity, but inhibition of cAMP degradation by phosphodiesterases (PDE) likely has an important role, because cAMP is inactivated much faster than it is synthesized. PDE1 is the only PDE family activated by Ca2+, which is reduced in PKD cells. To assess the contribution of the PDE1A subfamily to renal cyst formation, we examined the expression and function of PDE1A in zebrafish. We identified two splice isoforms with alternative starts corresponding to human PDE1A1 and PDE1A4. Expression of the two isoforms varied in embryos and adult tissues, and both isoforms hydrolyzed cAMP with Ca2+/calmodulin dependence. Depletion of PDE1A in zebrafish embryos using splice- and translation-blocking morpholinos (MOs) caused pronephric cysts, hydrocephalus, and body curvature. Human PDE1A RNA and the PKA inhibitors, H89 and Rp-cAMPS, partially rescued phenotypes of pde1a morphants. Additionally, MO depletion of PDE1A aggravated phenotypes in pkd2 morphants, causing more severe body curvature, and human PDE1A RNA partially rescued pkd2 morphant phenotypes, pronephric cysts, hydrocephalus, and body curvature. Together, these data indicate the integral role of PDE1A and cAMP signaling in renal development and cystogenesis, imply that PDE1A activity is altered downstream of polycystin-2, and suggest that PDE1A is a viable drug target for PKD. PMID:24700876

  20. Loss of extra-striatal phosphodiesterase 10A expression in early premanifest Huntington's disease gene carriers.

    PubMed

    Wilson, Heather; Niccolini, Flavia; Haider, Salman; Marques, Tiago Reis; Pagano, Gennaro; Coello, Christopher; Natesan, Sridhar; Kapur, Shitij; Rabiner, Eugenii A; Gunn, Roger N; Tabrizi, Sarah J; Politis, Marios

    2016-09-15

    Huntington's disease (HD) is a monogenic neurodegenerative disorder with an underlying pathology involving the toxic effect of mutant huntingtin protein primarily in striatal and cortical neurons. Phosphodiesterase 10A (PDE10A) regulates intracellular signalling cascades, thus having a key role in promoting neuronal survival. Using positron emission tomography (PET) with [(11)C]IMA107, we investigated the in vivo extra-striatal expression of PDE10A in 12 early premanifest HD gene carriers. Image processing and kinetic modelling was performed using MIAKAT™. Parametric images of [(11)C]IMA107 non-displaceable binding potential (BPND) were generated from the dynamic [(11)C]IMA107 scans using the simplified reference tissue model with the cerebellum as the reference tissue for nonspecific binding. We set a threshold criterion for meaningful quantification of [(11)C]IMA107 BPND at 0.30 in healthy control data; regions meeting this criterion were designated as regions of interest (ROIs). MRI-based volumetric analysis showed no atrophy in ROIs. We found significant differences in mean ROIs [(11)C]IMA107 BPND between HD gene carriers and healthy controls. HD gene carriers had significant loss of PDE10A within the insular cortex and occipital fusiform gyrus compared to healthy controls. Insula and occipital fusiform gyrus are important brain areas for the regulation of cognitive and limbic function that is impaired in HD. Our findings suggest that dysregulation of PDE10A-mediated intracellular signalling could be an early phenomenon in the course of HD with relevance also for extra-striatal brain areas. PMID:27538642

  1. Two Phosphodiesterases from Ustilago Maydis Share Structural and Biochemical Properties with Non-Fungal Phosphodiesterases

    PubMed Central

    Agarwal, Charu; Schultz, David J.; Perlin, Michael H.

    2010-01-01

    The dependence of Protein Kinase A (PKA) activity on cAMP levels is an important facet of the dimorphic switch between budding and filamentous growth as well as for pathogenicity in some fungi. To better understand these processes in the pathogenic fungus Ustilago maydis, we characterized the structure and biochemical functions of two phosphodiesterase (PDE) genes. Phosphodiesterases are enzymes involved in cAMP turnover and thus, contribute to the regulation of the cAMP-PKA signaling pathway. Two predicted homologs of PDEs were identified in the genome of U. maydis and hypothesized to be involved in cAMP turnover, thus regulating activity of the PKA catalytic subunit. Both umpde1 and umpde2 genes contain domains associated with phosphodiesterase activity predicted by InterPro analysis. Biochemical characterization of recombinantly produced UmPde1 (U. maydis Phosphodiesterase I) and UmPde2 demonstrated that both enzymes have phosphodiesterase activity in vitro, yet neither was inhibited by the phosphodiesterase inhibitor IBMX. Moreover, UmPde1 is specific for cAMP, while UmPde2 has broader substrate specificity, utilizing cAMP and cGMP as substrates. In addition, UmPde2 was also found to have nucleotide phosphatase activity that was higher with GMP compared to AMP. These results demonstrate that UmPde1 is a bona fide phosphodiesterase, while UmPde2 has more general activity as a cyclic nucleotide phosphodiesterase and/or GMP/AMP phosphatase. Thus, UmPde1 and UmPde2 likely have important roles in cell morphology and development and share some characteristics with a variety of non-fungal phosphodiesterases. PMID:21687762

  2. Reduced expression of cytochrome oxidases largely explains cAMP inhibition of aerobic growth in Shewanella oneidensis

    PubMed Central

    Yin, Jianhua; Meng, Qiu; Fu, Huihui; Gao, Haichun

    2016-01-01

    Inhibition of bacterial growth under aerobic conditions by elevated levels of cyclic adenosine 3′,5′-monophosphate (cAMP), first revealed more than 50 years ago, was attributed to accumulation of toxic methylglyoxal (MG). Here, we report a Crp-dependent mechanism rather than MG accumulation that accounts for the phenotype in Shewanella oneidensis, an emerging research model for the bacterial physiology. We show that a similar phenotype can be obtained by removing CpdA, a cAMP phosphodiesterase that appears more effective than its Escherichia coli counterpart. Although production of heme c and cytochromes c is correlated well with cAMP levels, neither is sufficient for the retarded growth. Quantities of overall cytochromes c increased substantially in the presence of elevated cAMP, a phenomenon resembling cells respiring on non-oxygen electron acceptors. In contrast, transcription of Crp-dependent genes encoding both cytochromes bd and cbb3 oxidases is substantially repressed under the same condition. Overall, our results suggest that cAMP of elevated levels drives cells into a low-energetic status, under which aerobic respiration is inhibited. PMID:27076065

  3. Defining the role of a FYVE domain in the localization and activity of a cAMP phosphodiesterase implicated in osmoregulation in Trypanosoma cruzi.

    PubMed

    Schoijet, Alejandra C; Miranda, Kildare; Medeiros, Lia Carolina Soares; de Souza, Wanderley; Flawiá, Mirtha M; Torres, Héctor N; Pignataro, Omar P; Docampo, Roberto; Alonso, Guillermo D

    2011-01-01

    Intracellular levels of cyclic nucleotide second messengers are regulated predominantly by a large superfamily of phosphodiesterases (PDEs). Trypanosoma cruzi, the causative agent of Chagas disease, encodes four different PDE families. One of these PDEs, T. cruzi PDE C2 (TcrPDEC2) has been characterized as a FYVE domain containing protein. Here, we report a novel role for TcrPDEC2 in osmoregulation in T. cruzi and reveal the relevance of its FYVE domain. Our data show that treatment of epimastigotes with TcrPDEC2 inhibitors improves their regulatory volume decrease, whereas cells overexpressing this enzyme are unaffected by the same inhibitors. Consistent with these results, TcrPDEC2 localizes to the contractile vacuole complex, showing strong labelling in the region corresponding to the spongiome. Furthermore, transgenic parasites overexpressing a truncated version of TcrPDEC2 without the FYVE domain show a failure in its targeting to the contractile vacuole complex and a marked decrease in PDE activity, supporting the importance of this domain to the localization and activity of TcrPDEC2. Taking together, the results here presented are consistent with the importance of the cyclic AMP signalling pathway in regulatory volume decrease and implicate TcrPDEC2 as a specifically localized PDE involved in osmoregulation in T. cruzi. PMID:21166893

  4. Communication of cAMP by connexin43 gap junctions regulates osteoblast signaling and gene expression.

    PubMed

    Gupta, Aditi; Anderson, Hidayah; Buo, Atum M; Moorer, Megan C; Ren, Margaret; Stains, Joseph P

    2016-08-01

    Connexin43 (Cx43) containing gap junctions play an important role in bone homeostasis, yet little is known about the second messengers communicated by Cx43 among bone cells. Here, we used MC3T3-E1 pre-osteoblasts and UMR106 rat osteosarcoma cells to test the hypothesis that cAMP is a second messenger communicated by bone cells through Cx43 containing gap junctions in a manner that is sufficient to impact osteoblast function. Overexpression of Cx43 markedly enhanced the activity of a cAMP-response element driven transcriptional luciferase reporter (CRE-luc) and increased phospho-CREB and phospho-ERK1/2 levels following expression of a constitutively active Gsα or by treatment with prostaglandin E2 (PGE2), 3-Isobutyl-1-methyl xanthine (IBMX) or forskolin. The Cx43-dependent potentiation of signaling in PGE2 treated cells was not accompanied by a further increase in cAMP levels, suggesting that the cAMP was shared between cells rather than Cx43 enhancing cAMP production. To support this, we developed a novel assay in which one set of cells expressing constitutively active Gsα (donor cells) were co-cultured with a second set of cells expressing a CRE-luc reporter (acceptor cells). Using this assay, activation of a CRE-luc reporter in the acceptor cells was both Cx43- and cell contact-dependent, indicating communication of cAMP among cells. Finally, we showed that Cx43 increased the cAMP-dependent mRNA expression of receptor activator of nuclear factor kappa B ligand (RANKL) and enhanced the repression of the sclerostin mRNA, implying a potential mechanism for the modulation of tissue remodeling. In total, these data demonstrate that Cx43 can communicate cAMP between cells and, more importantly, that the communicated cAMP is sufficient to impact signal transduction cascades and the expression of key bone effector molecules between interconnected cells. PMID:27156839

  5. Interplay of Ca2+ and cAMP signaling in the insulin-secreting MIN6 beta-cell line.

    PubMed

    Landa, Luis R; Harbeck, Mark; Kaihara, Kelly; Chepurny, Oleg; Kitiphongspattana, Kajorn; Graf, Oliver; Nikolaev, Viacheslav O; Lohse, Martin J; Holz, George G; Roe, Michael W

    2005-09-01

    Ca2+ and cAMP are important second messengers that regulate multiple cellular processes. Although previous studies have suggested direct interactions between Ca2+ and cAMP signaling pathways, the underlying mechanisms remain unresolved. In particular, direct evidence for Ca2+-regulated cAMP production in living cells is incomplete. Genetically encoded fluorescence resonance energy transfer-based biosensors have made possible real-time imaging of spatial and temporal gradients of intracellular cAMP concentration in single living cells. Here, we used confocal microscopy, fluorescence resonance energy transfer, and insulin-secreting MIN6 cells expressing Epac1-camps, a biosynthetic unimolecular cAMP indicator, to better understand the role of intracellular Ca2+ in cAMP production. We report that depolarization with high external K+, tolbutamide, or glucose caused a rapid increase in cAMP that was dependent on extracellular Ca2+ and inhibited by nitrendipine, a Ca2+ channel blocker, or 2',5'-dideoxyadenosine, a P-site antagonist of transmembrane adenylate cyclases. Stimulation of MIN6 cells with glucose in the presence of tetraethylammonium chloride generated concomitant Ca2+ and cAMP oscillations that were abolished in the absence of extracellular Ca2+ and blocked by 2',5'-dideoxyadenosine or 3-isobutyl-1-methylxanthine, an inhibitor of phosphodiesterase. Simultaneous measurements of Ca2+ and cAMP concentrations with Fura-2 and Epac1-camps, respectively, revealed a close temporal and causal interrelationship between the increases in cytoplasmic Ca2+ and cAMP levels following membrane depolarization. These findings indicate highly coordinated interplay between Ca2+ and cAMP signaling in electrically excitable endocrine cells and suggest that Ca2+-dependent cAMP oscillations are derived from an increase in adenylate cyclase activity and periodic activation and inactivation of cAMP-hydrolyzing phosphodiesterase. PMID:15987680

  6. Calcitonin Induces Expression of the Inducible cAMP Early Repressor in Osteoclasts

    PubMed Central

    Yang, Maobin; Kream, Barbara E.

    2010-01-01

    The cAMP response element modulator gene (Crem) encodes a variety of transcriptional regulators including the inducible cAMP early repressor, ICER. We previously showed that Crem knockout mice, which are deficient in CREM and ICER factors, display slightly increased long bone mass and decreased osteoclast number. These data are consistent with the notion that Crem regulates bone mass in part through an effect on osteoclast formation and/or function. Since ICER is strongly induced by cAMP, we asked whether the calcium-regulating hormone calcitonin, which stimulates cAMP production and inhibits osteoclastic bone resorption, could induce ICER in osteoclasts. The monocytic cell line RAW264.7 was treated with receptor activator of NF-κB ligand (RANKL) to induce osteoclast formation. Calcitonin caused a time- and dose-dependent induction of ICER mRNA and an increase in ICER protein abundance in RANKL-treated RAW264.7 cells. Calcitonin also induced ICER mRNA and protein in osteoclasts derived from primary mouse bone marrow cell cultures. Calcitonin-treated osteoclasts showed immunoreactivity with an anti-CREM antibody. Calcitonin decreased the activity of wild type and Crem knockout osteoclasts in vitro, and this inhibitory effect was greater in Crem knockout osteoclasts. Furthermore, calcitonin decreased calcitonin receptor mRNA expression in wild type osteoclasts but not in Crem knockout osteoclasts. These data suggest that calcitonin induction of ICER in osteoclasts might regulate osteoclast activity. PMID:19016003

  7. Expression of Phosphodiesterase 4B cAMP-Specific Gene in Subjects With Cryptorchidism and Down's Syndrome.

    PubMed

    Salemi, Michele; Condorelli, Rosita A; La Vignera, Sandro; Castiglione, Roberto; Salluzzo, Maria Grazia; Bonaccorso, Carmela M; Vinci, Mirella; Bosco, Paolo; Romano, Carmelo; Campagna, Cristina; Romano, Corrado; Calogero, Aldo E

    2016-05-01

    Cryptorchidism represents a risk factor for infertility and germ cell testicular neoplasia. An increased rate of cryptorchidism has been reported in subjects with Down's syndrome. Cyclic nucleotide phosphodiesterases (PDEs) are important messengers that regulate and mediate a number of cellular responses to extracellular signals, such as neurotransmitters and hormones. PDE4B, cAMP-specific (PDE4B) gene which maps to chromosome 1p31.3 appears to be involved in schizophrenia, chronic psychiatric illness, learning, memory, and mood disturbances. Expression of PDE4 enzymes have been studied in testes of cryptorchid rats. Expression of PDE4B protein examination showed marked degenerative changes in the epithelial lining of the seminiferous tubules. These findings led us to evaluate PDE4 mRNA expression in leukocytes of peripheral blood of five men with DS and cryptorchidism and eleven subjects with DS without cryptorchidism compared with healthy men (controls) by quantitative Real Time PCR (qRT-PCR). This study showed that the PDE4B gene was downexpressed in men with DS and cryptorchidism compared to normal controls and DS without cryptorchidism. A lower expression of the PDE4B gene may be involved in the neurological abnormalities in subjects with Down's syndrome. Moreover, PDE4B gene may be involved in the testicular abnormalities of men with DS and cryptorchidism. PMID:25546171

  8. Alterations of Phosphodiesterases in Adrenocortical Tumors.

    PubMed

    Hannah-Shmouni, Fady; Faucz, Fabio R; Stratakis, Constantine A

    2016-01-01

    Alterations in the cyclic (c)AMP-dependent signaling pathway have been implicated in the majority of benign adrenocortical tumors (ACTs) causing Cushing syndrome (CS). Phosphodiesterases (PDEs) are enzymes that regulate cyclic nucleotide levels, including cyclic adenosine monophosphate (cAMP). Inactivating mutations and other functional variants in PDE11A and PDE8B, two cAMP-binding PDEs, predispose to ACTs. The involvement of these two genes in ACTs was initially revealed by a genome-wide association study in patients with micronodular bilateral adrenocortical hyperplasia. Thereafter, PDE11A or PDE8B genetic variants have been found in other ACTs, including macronodular adrenocortical hyperplasias and cortisol-producing adenomas. In addition, downregulation of PDE11A expression and inactivating variants of the gene have been found in hereditary and sporadic testicular germ cell tumors, as well as in prostatic cancer. PDEs confer an increased risk of ACT formation probably through, primarily, their action on cAMP levels, but other actions might be possible. In this report, we review what is known to date about PDE11A and PDE8B and their involvement in the predisposition to ACTs. PMID:27625633

  9. Alterations of Phosphodiesterases in Adrenocortical Tumors

    PubMed Central

    Hannah-Shmouni, Fady; Faucz, Fabio R.; Stratakis, Constantine A.

    2016-01-01

    Alterations in the cyclic (c)AMP-dependent signaling pathway have been implicated in the majority of benign adrenocortical tumors (ACTs) causing Cushing syndrome (CS). Phosphodiesterases (PDEs) are enzymes that regulate cyclic nucleotide levels, including cyclic adenosine monophosphate (cAMP). Inactivating mutations and other functional variants in PDE11A and PDE8B, two cAMP-binding PDEs, predispose to ACTs. The involvement of these two genes in ACTs was initially revealed by a genome-wide association study in patients with micronodular bilateral adrenocortical hyperplasia. Thereafter, PDE11A or PDE8B genetic variants have been found in other ACTs, including macronodular adrenocortical hyperplasias and cortisol-producing adenomas. In addition, downregulation of PDE11A expression and inactivating variants of the gene have been found in hereditary and sporadic testicular germ cell tumors, as well as in prostatic cancer. PDEs confer an increased risk of ACT formation probably through, primarily, their action on cAMP levels, but other actions might be possible. In this report, we review what is known to date about PDE11A and PDE8B and their involvement in the predisposition to ACTs.

  10. Diatom acclimation to elevated CO2 via cAMP signalling and coordinated gene expression

    NASA Astrophysics Data System (ADS)

    Hennon, Gwenn M. M.; Ashworth, Justin; Groussman, Ryan D.; Berthiaume, Chris; Morales, Rhonda L.; Baliga, Nitin S.; Orellana, Mónica V.; Armbrust, E. V.

    2015-08-01

    Diatoms are responsible for ~40% of marine primary productivity, fuelling the oceanic carbon cycle and contributing to natural carbon sequestration in the deep ocean. Diatoms rely on energetically expensive carbon concentrating mechanisms (CCMs) to fix carbon efficiently at modern levels of CO2 (refs , , ). How diatoms may respond over the short and long term to rising atmospheric CO2 remains an open question. Here we use nitrate-limited chemostats to show that the model diatom Thalassiosira pseudonana rapidly responds to increasing CO2 by differentially expressing gene clusters that regulate transcription and chromosome folding, and subsequently reduces transcription of photosynthesis and respiration gene clusters under steady-state elevated CO2. These results suggest that exposure to elevated CO2 first causes a shift in regulation, and then a metabolic rearrangement. Genes in one CO2-responsive cluster included CCM and photorespiration genes that share a putative cAMP-responsive cis-regulatory sequence, implying these genes are co-regulated in response to CO2, with cAMP as an intermediate messenger. We verified cAMP-induced downregulation of CCM gene δ-CA3 in nutrient-replete diatom cultures by inhibiting the hydrolysis of cAMP. These results indicate an important role for cAMP in downregulating CCM and photorespiration genes under elevated CO2 and provide insights into mechanisms of diatom acclimation in response to climate change.

  11. Imaging cytoplasmic cAMP in mouse brainstem neurons

    PubMed Central

    Mironov, SL; Skorova, E; Taschenberger, G; Hartelt, N; Nikolaev, VO; Lohse, MJ; Kügler, S

    2009-01-01

    Background cAMP is an ubiquitous second messenger mediating various neuronal functions, often as a consequence of increased intracellular Ca2+ levels. While imaging of calcium is commonly used in neuroscience applications, probing for cAMP levels has not yet been performed in living vertebrate neuronal tissue before. Results Using a strictly neuron-restricted promoter we virally transduced neurons in the organotypic brainstem slices which contained pre-Bötzinger complex, constituting the rhythm-generating part of the respiratory network. Fluorescent cAMP sensor Epac1-camps was expressed both in neuronal cell bodies and neurites, allowing us to measure intracellular distribution of cAMP, its absolute levels and time-dependent changes in response to physiological stimuli. We recorded [cAMP]i changes in the micromolar range after modulation of adenylate cyclase, inhibition of phosphodiesterase and activation of G-protein-coupled metabotropic receptors. [cAMP]i levels increased after membrane depolarisation and release of Ca2+ from internal stores. The effects developed slowly and reached their maximum after transient [Ca2+]i elevations subsided. Ca2+-dependent [cAMP]i transients were suppressed after blockade of adenylate cyclase with 0.1 mM adenylate cyclase inhibitor 2'5'-dideoxyadenosine and potentiated after inhibiting phosphodiesterase with isobutylmethylxanthine and rolipram. During paired stimulations, the second depolarisation and Ca2+ release evoked bigger cAMP responses. These effects were abolished after inhibition of protein kinase A with H-89 pointing to the important role of phosphorylation of calcium channels in the potentiation of [cAMP]i transients. Conclusion We constructed and characterized a neuron-specific cAMP probe based on Epac1-camps. Using viral gene transfer we showed its efficient expression in organotypic brainstem preparations. Strong fluorescence, resistance to photobleaching and possibility of direct estimation of [cAMP] levels using

  12. Expression of Tas1 Taste Receptors in Mammalian Spermatozoa: Functional Role of Tas1r1 in Regulating Basal Ca2+ and cAMP Concentrations in Spermatozoa

    PubMed Central

    Meyer, Dorke; Voigt, Anja; Widmayer, Patricia; Borth, Heike; Huebner, Sandra; Breit, Andreas; Marschall, Susan; de Angelis, Martin Hrabé; Boehm, Ulrich; Meyerhof, Wolfgang; Gudermann, Thomas; Boekhoff, Ingrid

    2012-01-01

    Background During their transit through the female genital tract, sperm have to recognize and discriminate numerous chemical compounds. However, our current knowledge of the molecular identity of appropriate chemosensory receptor proteins in sperm is still rudimentary. Considering that members of the Tas1r family of taste receptors are able to discriminate between a broad diversity of hydrophilic chemosensory substances, the expression of taste receptors in mammalian spermatozoa was examined. Methodology/Principal Findings The present manuscript documents that Tas1r1 and Tas1r3, which form the functional receptor for monosodium glutamate (umami) in taste buds on the tongue, are expressed in murine and human spermatozoa, where their localization is restricted to distinct segments of the flagellum and the acrosomal cap of the sperm head. Employing a Tas1r1-deficient mCherry reporter mouse strain, we found that Tas1r1 gene deletion resulted in spermatogenic abnormalities. In addition, a significant increase in spontaneous acrosomal reaction was observed in Tas1r1 null mutant sperm whereas acrosomal secretion triggered by isolated zona pellucida or the Ca2+ ionophore A23187 was not different from wild-type spermatozoa. Remarkably, cytosolic Ca2+ levels in freshly isolated Tas1r1-deficient sperm were significantly higher compared to wild-type cells. Moreover, a significantly higher basal cAMP concentration was detected in freshly isolated Tas1r1-deficient epididymal spermatozoa, whereas upon inhibition of phosphodiesterase or sperm capacitation, the amount of cAMP was not different between both genotypes. Conclusions/Significance Since Ca2+ and cAMP control fundamental processes during the sequential process of fertilization, we propose that the identified taste receptors and coupled signaling cascades keep sperm in a chronically quiescent state until they arrive in the vicinity of the egg - either by constitutive receptor activity and/or by tonic receptor activation by

  13. 2', 3'-cyclic nucleotide 3'-phosphodiesterase is expressed in dissociated rat cerebellar cells and included in the postsynaptic density fraction.

    PubMed

    Cho, Sun-Jung; Jung, Jae Seob; Jin, IngNyol; Moon, Il Soo

    2003-08-31

    We have shown by protein sequencing that the phosphotyrosine-containing 48 kDa protein band of the rat cerebellar postsynaptic density fraction (CBL-PSD) is 2', 3'-cyclic nucleotide 3'-phosphodiesterase 2 (CNP2). Immunoblot analysis indicated that both CNP1 and CNP2 isoforms are present in the CBL-PSD fraction, whereas there is little CNP2 in the forebrain (FB)-PSD fraction. Both isoforms in the CBL-PSD fraction were tyrosine-phosphorylated to a basal extent. They were efficiently dissociated from the complexes in the PSD fraction by salt, but not by non-ionic detergents such as n-octyl glucoside (OG) and Triton X-100. Immunocytochemistry of dissociated cerebellar cultures revealed patchy CNP staining in oligodendrocytes (OLs), Purkinje cells (PCs), and unidentified PSD95-positive cells, but no staining in granule cells (GCs). Our results indicate that both CNP1 and CNP2 are expressed in cerian populations of cerebellar cells in addition to OL, and that they are associated with complexes that are co-isolated with the PSD. PMID:14503857

  14. Expression and Characterization of a Novel Glycerophosphodiester Phosphodiesterase from Pyrococcus furiosus DSM 3638 That Possesses Lysophospholipase D Activity.

    PubMed

    Wang, Fanghua; Lai, Linhui; Liu, Yanhua; Yang, Bo; Wang, Yonghua

    2016-01-01

    Glycerophosphodiester phosphodiesterases (GDPD) are enzymes which degrade various glycerophosphodiesters to produce glycerol-3-phosphate and the corresponding alcohol moiety. Apart from this, a very interesting finding is that this enzyme could be used in the degradation of toxic organophosphorus esters, which has resulted in much attention on the biochemical and application research of GDPDs. In the present study, a novel GDPD from Pyrococcus furiosus DSM 3638 (pfGDPD) was successfully expressed in Escherichia coli and biochemically characterized. This enzyme hydrolyzed bis(p-nitrophenyl) phosphate, one substrate analogue of organophosphorus diester, with an optimal reaction temperature 55 °C and pH 8.5. The activity of pfGDPD was strongly dependent on existing of bivalent cations. It was strongly stimulated by Mn(2+) ions, next was Co(2+) and Ni(2+) ions. Further investigations were conducted on its substrate selectivity towards different phospholipids. The results indicated that except of glycerophosphorylcholine (GPC), this enzyme also possessed lysophospholipase D activity toward both sn1-lysophosphatidylcholine (1-LPC) and sn2-lysophosphatidylcholine (2-LPC). Higher activity was found for 1-LPC than 2-LPC; however, no hydrolytic activity was found for phosphatidylcholine (PC). Molecular docking based on the 3D-modeled structure of pfGDPD was conducted in order to provide a structural foundation for the substrate selectivity. PMID:27248999

  15. Expression and Characterization of a Novel Glycerophosphodiester Phosphodiesterase from Pyrococcus furiosus DSM 3638 That Possesses Lysophospholipase D Activity

    PubMed Central

    Wang, Fanghua; Lai, Linhui; Liu, Yanhua; Yang, Bo; Wang, Yonghua

    2016-01-01

    Glycerophosphodiester phosphodiesterases (GDPD) are enzymes which degrade various glycerophosphodiesters to produce glycerol-3-phosphate and the corresponding alcohol moiety. Apart from this, a very interesting finding is that this enzyme could be used in the degradation of toxic organophosphorus esters, which has resulted in much attention on the biochemical and application research of GDPDs. In the present study, a novel GDPD from Pyrococcus furiosus DSM 3638 (pfGDPD) was successfully expressed in Escherichia coli and biochemically characterized. This enzyme hydrolyzed bis(p-nitrophenyl) phosphate, one substrate analogue of organophosphorus diester, with an optimal reaction temperature 55 °C and pH 8.5. The activity of pfGDPD was strongly dependent on existing of bivalent cations. It was strongly stimulated by Mn2+ ions, next was Co2+ and Ni2+ ions. Further investigations were conducted on its substrate selectivity towards different phospholipids. The results indicated that except of glycerophosphorylcholine (GPC), this enzyme also possessed lysophospholipase D activity toward both sn1-lysophosphatidylcholine (1-LPC) and sn2-lysophosphatidylcholine (2-LPC). Higher activity was found for 1-LPC than 2-LPC; however, no hydrolytic activity was found for phosphatidylcholine (PC). Molecular docking based on the 3D-modeled structure of pfGDPD was conducted in order to provide a structural foundation for the substrate selectivity. PMID:27248999

  16. Acidosis Is a key regulator of osteoblast ecto‐nucleotidase pyrophosphatase/phosphodiesterase 1 (NPP1) expression and activity

    PubMed Central

    Key, Michelle L.; Hajjawi, Mark O.R.; Millán, José L.; Arnett, Timothy R.

    2015-01-01

    Previous work has shown that acidosis prevents bone nodule formation by osteoblasts in vitro by inhibiting mineralisation of the collagenous matrix. The ratio of phosphate (Pi) to pyrophosphate (PPi) in the bone microenvironment is a fundamental regulator of bone mineralisation. Both Pi and PPi, a potent inhibitor of mineralisation, are generated from extracellular nucleotides by the actions of ecto‐nucleotidases. This study investigated the expression and activity of ecto‐nucleotidases by osteoblasts under normal and acid conditions. We found that osteoblasts express mRNA for a number of ecto‐nucleotidases including NTPdase 1–6 (ecto‐nucleoside triphosphate diphosphohydrolase) and NPP1‐3 (ecto‐nucleotide pyrophosphatase/phosphodiesterase). The rank order of mRNA expression in differentiating rat osteoblasts (day 7) was Enpp1 > NTPdase 4 > NTPdase 6 > NTPdase 5 > alkaline phosphatase > ecto‐5‐nucleotidase > Enpp3 > NTPdase 1 > NTPdase 3 > Enpp2 > NTPdase 2. Acidosis (pH 6.9) upregulated NPP1 mRNA (2.8‐fold) and protein expression at all stages of osteoblast differentiation compared to physiological pH (pH 7.4); expression of other ecto‐nucleotidases was unaffected. Furthermore, total NPP activity was increased up to 53% in osteoblasts cultured in acid conditions (P < 0.001). Release of ATP, one of the key substrates for NPP1, from osteoblasts, was unaffected by acidosis. Further studies showed that mineralised bone formation by osteoblasts cultured from NPP1 knockout mice was increased compared with wildtypes (2.5‐fold, P < 0.001) and was partially resistant to the inhibitory effect of acidosis. These results indicate that increased NPP1 expression and activity might contribute to the decreased mineralisation observed when osteoblasts are exposed to acid conditions. J. Cell. Physiol. 230: 3049–3056, 2015. © 2015 The Authors. Journal of Cellular Physiology Published by Wiley Periodicals, Inc

  17. Exchange factors directly activated by cAMP mediate melanocortin 4 receptor-induced gene expression

    PubMed Central

    Glas, Evi; Mückter, Harald; Gudermann, Thomas; Breit, Andreas

    2016-01-01

    Gs protein-coupled receptors regulate many vital body functions by activation of cAMP response elements (CRE) via cAMP-dependent kinase A (PKA)-mediated phosphorylation of the CRE binding protein (CREB). Melanocortin 4 receptors (MC4R) are prototypical Gs-coupled receptors that orchestrate the hypothalamic control of food-intake and metabolism. Remarkably, the significance of PKA for MC4R-induced CRE-dependent transcription in hypothalamic cells has not been rigorously interrogated yet. In two hypothalamic cell lines, we observed that blocking PKA activity had only weak or no effects on reporter gene expression. In contrast, inhibitors of exchange factors directly activated by cAMP-1/2 (EPAC-1/2) mitigated MC4R-induced CRE reporter activation and mRNA induction of the CREB-dependent genes c-fos and thyrotropin-releasing hormone. Furthermore, we provide first evidence that extracellular-regulated kinases-1/2 (ERK-1/2) activated by EPACs and not PKA are the elusive CREB kinases responsible for MC4R-induced CREB/CRE activation in hypothalamic cells. Overall, these data emphasize the pivotal role of EPACs rather than PKA in hypothalamic gene expression elicited by a prototypical Gs-coupled receptor. PMID:27612207

  18. Selective Phosphodiesterase 4B Inhibitors: A Review

    PubMed Central

    Azam, Mohammed Afzal; Tripuraneni, Naga Srinivas

    2014-01-01

    Abstract Phosphodiesterase 4B (PDE4B) is a member of the phosphodiesterase family of proteins that plays a critical role in regulating intracellular levels of cyclic adenosine monophosphate (cAMP) by controlling its rate of degradation. It has been demonstrated that this isoform is involved in the orchestra of events which includes inflammation, schizophrenia, cancers, chronic obstructive pulmonary disease, contractility of the myocardium, and psoriatic arthritis. Phosphodiesterase 4B has constituted an interesting target for drug development. In recent years, a number of PDE4B inhibitors have been developed for their use as therapeutic agents. In this review, an up-to-date status of the inhibitors investigated for the inhibition of PDE4B has been given so that this rich source of structural information of presently known PDE4B inhibitors could be helpful in generating a selective and potent inhibitor of PDE4B. PMID:25853062

  19. A Universal Stress Protein (USP) in Mycobacteria Binds cAMP

    PubMed Central

    Banerjee, Arka; Adolph, Ramona S.; Gopalakrishnapai, Jayashree; Kleinboelting, Silke; Emmerich, Christiane; Steegborn, Clemens; Visweswariah, Sandhya S.

    2015-01-01

    Mycobacteria are endowed with rich and diverse machinery for the synthesis, utilization, and degradation of cAMP. The actions of cyclic nucleotides are generally mediated by binding of cAMP to conserved and well characterized cyclic nucleotide binding domains or structurally distinct cGMP-specific and -regulated cyclic nucleotide phosphodiesterase, adenylyl cyclase, and E. coli transcription factor FhlA (GAF) domain-containing proteins. Proteins with cyclic nucleotide binding and GAF domains can be identified in the genome of mycobacterial species, and some of them have been characterized. Here, we show that a significant fraction of intracellular cAMP is bound to protein in mycobacterial species, and by using affinity chromatography techniques, we identify specific universal stress proteins (USP) as abundantly expressed cAMP-binding proteins in slow growing as well as fast growing mycobacteria. We have characterized the biochemical and thermodynamic parameters for binding of cAMP, and we show that these USPs bind cAMP with a higher affinity than ATP, an established ligand for other USPs. We determined the structure of the USP MSMEG_3811 bound to cAMP, and we confirmed through structure-guided mutagenesis, the residues important for cAMP binding. This family of USPs is conserved in all mycobacteria, and we suggest that they serve as “sinks” for cAMP, making this second messenger available for downstream effectors as and when ATP levels are altered in the cell. PMID:25802331

  20. Molecular cloning and transient expression in COS7 cells of a novel human PDE4B cAMP-specific phosphodiesterase, HSPDE4B3.

    PubMed Central

    Huston, E; Lumb, S; Russell, A; Catterall, C; Ross, A H; Steele, M R; Bolger, G B; Perry, M J; Owens, R J; Houslay, M D

    1997-01-01

    5'-Rapid amplification of cDNA ends, done on poly(A)+ RNA from human U87 cells, was used to identify 420 bp of novel 5' sequence of a PDE4B cAMP-specific phosphodiesterase (PDE). This identified an open reading frame encoding a putative 721-residue 'long-form' PDE4B splice variant, which we term HSPDE4B3. HSPDE4B3 differs from the two known PDE4B forms by virtue of its unique 79-residue N-terminal region, compared with the unique N-terminal regions of 94 and 39 residues found in HSPDE4B1 and HSPDE4B2 respectively. In transfected COS7 cells the two long forms, HSPDE4B1 and HSPDE4B3, had molecular masses of approx. 104 and approx. 103 kDa respectively. Expressed in COS-7 cells, the three HSPDE4B isoforms were found in the high-speed supernatant (cytosol) fraction as well as both the high-speed pellet (P2) and low-speed pellet (P1) fractions. All isoforms showed similar Km values for cAMP hydrolysis (1.5-2.6 microM). The maximal activities of the soluble cytosolic activity of the two long forms were very similar, whereas that of the short form, HSPDE4B2, was approx. 4-fold higher. Particulate-associated HSPDE4B1 and HSPDE4B2 were less active (approx. 40%) than their cytosol forms, whereas particulate HSPDE4B3 was similar in activity to its cytosolic form. Particulate and cytosolic forms of HSPDE4B1 and HSPDE4B3 were similarly inhibited by rolipram ¿4-[3-(cyclopentoxyl)-4-methoxyphenyl]-2-pyrrolidone¿, the selective inhibitor of PDE4 (IC50 0.05-0.1 microM), whereas particulate-associated HSPDE4B2 was profoundly (approx. 10-fold) more sensitive (IC50 0.02 microM) to rolipram inhibition than its cytosolic form (IC50 0.2 microM). The various particulate-associated HSPDE4B isoforms showed very different susceptibilities to solubilization with the detergent Triton X-100 and high NaCl concentration. A novel cDNA, called pRPDE74, was obtained by screening a rat olfactory lobe cDNA library. This contained an open reading frame encoding a 721-residue protein that showed

  1. Atrazine Acts as an Endocrine Disrupter by Inhibiting cAMP-specific Phosphodiesterase-4

    PubMed Central

    Kucka, Marek; Pogrmic-Majkic, Kristina; Fa, Svetlana; Stojilkovic, Stanko S.; Kovacevic, Radmila

    2014-01-01

    Atrazine, one of the most commonly used herbicides worldwide, acts as an endocrine disruptor, but the mechanism of its action has not been characterized. In this study, we show that atrazine rapidly increases cAMP levels in cultured rat pituitary and testicular Leydig cells in a concentration-dependent manner, but less effectively than 3-isobutyl-1-methylxanthine, a competitive non-specific inhibitor of phosphodiesterases (PDEs). In forskolin (an activator of adenylyl cyclase)- and probenecid (an inhibitor of cyclic nucleotide transporters)-treated cells, but not in 3-isobutyl-1-methylxanthine-treated cells, atrazine further increased cAMP levels, indicating that inhibition of PDEs accounts for accumulation of cAMP. In contrast to cAMP, atrazine did not alter cGMP levels, further indicating that it inhibits cAMP-specific PDEs. Atrazine-induced changes in cAMP levels were sufficient to stimulate prolactin release in pituitary cells and androgen production in Leydig cells, indicating that it acts as an endocrine disrupter both in cells that secrete by exocytosis of prestored hormones and in cells that secrete by de novo hormone synthesis. Rolipram abolished the stimulatory effect of atrazine on cAMP release in both cell types, suggesting that it acts as an inhibitor of PDE4s, isoforms whose mRNA transcripts dominate in pituitary and Leydig cells together with mRNA for PDE8A. In contrast, immortalized lacto-somatotrophs showed low expression of these mRNA transcripts and several fold higher cAMP levels compared to normal pituitary cells, and atrazine was unable to further increase cAMP levels. These results indicate that atrazine acts as a general endocrine disrupter by inhibiting cAMP-specific PDE4s. PMID:23022511

  2. Phosphodiesterase Isoform Regulation of Cell Proliferation and Fluid Secretion in Autosomal Dominant Polycystic Kidney Disease.

    PubMed

    Pinto, Cibele S; Raman, Archana; Reif, Gail A; Magenheimer, Brenda S; White, Corey; Calvet, James P; Wallace, Darren P

    2016-04-01

    cAMP stimulates cell proliferation and Cl(-)-dependent fluid secretion, promoting the progressive enlargement of renal cysts in autosomal dominant polycystic kidney disease (ADPKD). Intracellular cAMP levels are determined by the balance of cAMP synthesis by adenylyl cyclases and degradation by phosphodiesterases (PDEs). Therefore, PDE isoform expression and activity strongly influence global and compartmentalized cAMP levels. We report here that PDE3 and PDE4 expression levels are lower in human ADPKD tissue and cells compared with those of normal human kidneys (NHKs), whereas PDE1 levels are not significantly different. Inhibition of PDE4 caused a greater increase in basal and vasopressin (AVP)-stimulated cAMP levels and Cl(-) secretion by ADPKD cells than inhibition of PDE1, and inhibition of PDE4 induced cyst-like dilations in cultured mouse Pkd1(-/-) embryonic kidneys. In contrast, inhibition of PDE1 caused greater stimulation of extracellular signal-regulated kinase (ERK) and proliferation of ADPKD cells than inhibition of PDE4, and inhibition of PDE1 enhanced AVP-induced ERK activation. Notably, inhibition of PDE1, the only family of Ca(2+)-regulated PDEs, also induced a mitogenic response to AVP in NHK cells, similar to the effect of restricting intracellular Ca(2+). PDE1 coimmunoprecipitated with B-Raf and A-kinase anchoring protein 79, and AVP increased this interaction in ADPKD but not NHK cells. These data suggest that whereas PDE4 is the major PDE isoform involved in the regulation of global intracellular cAMP and Cl(-) secretion, PDE1 specifically affects the cAMP signal to the B-Raf/MEK/ERK pathway and regulates AVP-induced proliferation of ADPKD cells. PMID:26289612

  3. Superiority of combined phosphodiesterase PDE3/PDE4 inhibition over PDE4 inhibition alone on glucocorticoid- and long-acting β2-adrenoceptor agonist-induced gene expression in human airway epithelial cells.

    PubMed

    BinMahfouz, Hawazen; Borthakur, Bibhusana; Yan, Dong; George, Tresa; Giembycz, Mark A; Newton, Robert

    2015-01-01

    Glucocorticoids, also known as corticosteroids, induce effector gene transcription as a part of their anti-inflammatory mechanisms of action. Such genomic effects can be significantly enhanced by long-acting β2-adrenoceptor agonists (LABAs) and may contribute to the clinical superiority of inhaled corticosteroid (ICS)/LABA combinations in asthma and chronic obstructive pulmonary disease (COPD) over ICSs alone. Using models of cAMP- and glucocorticoid-induced transcription in human bronchial epithelial BEAS-2B cells, we show that combining inhibitors of phosphodiesterase (PDE) 3 and PDE4 provides greater benefits compared with inhibiting either PDE alone. In respect to cAMP-dependent transcription, inhibitors of PDE3 (siguazodan, cilostazol) and PDE4 (rolipram, GSK256066, roflumilast N-oxide) each sensitized to the LABA, formoterol. This effect was magnified by dual PDE3 and PDE4 inhibition. Siguazodan plus rolipram was also more effective at inducing cAMP-dependent transcription than either inhibitor alone. Conversely, the concentration-response curve describing the enhancement of dexamethasone-induced, glucocorticoid response element-dependent transcription by formoterol was displaced to the left by PDE4, but not PDE3, inhibition. Overall, similar effects were described for bona fide genes, including RGS2, CD200, and CRISPLD2. Importantly, the combination of siguazodan plus rolipram prolonged the duration of gene expression induced by formoterol, dexamethasone, or dexamethasone plus formoterol. This was most apparent for RGS2, a bronchoprotective gene that may also reduce the proinflammatory effects of constrictor mediators. Collectively, these data provide a rationale for the use of PDE3 and PDE4 inhibitors in the treatment of COPD and asthma where they may enhance, sensitize, and prolong the effects of LABA/ICS combination therapies. PMID:25324049

  4. Expression, intracellular distribution and basis for lack of catalytic activity of the PDE4A7 isoform encoded by the human PDE4A cAMP-specific phosphodiesterase gene.

    PubMed Central

    Johnston, Lee Ann; Erdogan, Suat; Cheung, York Fong; Sullivan, Michael; Barber, Rachael; Lynch, Martin J; Baillie, George S; Van Heeke, Gino; Adams, David R; Huston, Elaine; Houslay, Miles D

    2004-01-01

    PDE4A7 is an isoform encoded by the human PDE4A cAMP-specific phosphodiesterase gene that fails to hydrolyse cAMP and whose transcripts are widely expressed. Removal of either the N- or C-terminal unique portions of PDE4A7 did not reconstitute catalytic activity, showing that they did not exert a chronic inhibitory effect. A chimera (Hyb2), formed by swapping the unique N-terminal portion of PDE4A7 with that of the active PDE4A4C form, was not catalytically active. However, one formed (Hyb1) by swapping the unique C-terminal portion of PDE4A7 with that common to all active PDE4 isoforms was catalytically active. Compared with the active PDE4A4B isoform, Hyb1 exhibited a similar K(m) value for cAMP and IC50 value for rolipram inhibition, but was less sensitive to inhibition by Ro-20-1724 and denbufylline, and considerably more sensitive to thermal denaturation. The unique C-terminal region of PDE4A7 was unable to support an active catalytic unit, whereas its unique N-terminal region can. The N-terminal portion of the PDE4 catalytic unit is essential for catalytic activity and can be supplied by either highly conserved sequence found in active PDE4 isoforms from all four PDE4 subfamilies or the unique N-terminal portion of PDE4A7. A discrete portion of the conserved C-terminal region in active PDE4A isoforms underpins their aberrant migration on SDS/PAGE. Unlike active PDE4A isoforms, PDE4A7 is exclusively localized to the P1 particulate fraction in cells. A region located within the C-terminal portion of active PDE4 isoforms prevents such exclusive targeting. Three functional regions in PDE4A isoforms are identified, which influence catalytic activity, subcellular targeting and conformational status. PMID:15025561

  5. Receptors that couple to 2 classes of G proteins increase cAMP and activate CFTR expressed in Xenopus oocytes.

    PubMed

    Uezono, Y; Bradley, J; Min, C; McCarty, N A; Quick, M; Riordan, J R; Chavkin, C; Zinn, K; Lester, H A; Davidson, N

    1993-01-01

    The cystic fibrosis transmembrane conductance regulator (CFTR), a Cl- channel activated by phosphorylation, was expressed in Xenopus oocytes along with various combinations of several other components of the cAMP signalling pathway. Activation of the coexpressed beta 2 adrenergic receptor increased cAMP and led to CFTR activation. The activation of CFTR (1) requires only short (15 s) exposure to isoproterenol, (2) occurs for agonist concentrations 100-1000 fold lower than those that produce cAMP increases detectable by a radioimmunoassay, (3) requires injection of only 5 pg of receptor cRNA per oocyte, and (4) can be increased further by coexpression of cRNA for adenylyl cyclase type II or III or for Gs alpha. In addition, CFTR activation and cAMP increases by beta 2 activation were enhanced by activation of the coexpressed 5HT1A receptor, which is thought to couple to Gi. The additional activation by the 5HT1A receptor was enhanced by coexpression of adenylyl cyclase type II but not with type III and may proceed via the beta gamma subunits of a G protein. The sensitivity of the assay system is also demonstrated by responses to vasoactive intestinal peptide and to pituitary adenylate cyclase-activating polypeptide in oocytes injected with cerebral cortex mRNA. PMID:7522902

  6. Misoprostol modulates cytokine expression through a cAMP pathway: Potential therapeutic implication for liver disease.

    PubMed

    Gobejishvili, Leila; Ghare, Smita; Khan, Rehan; Cambon, Alexander; Barker, David F; Barve, Shirish; McClain, Craig; Hill, Daniell

    2015-12-01

    Dysregulated cytokine metabolism plays a critical role in the pathogenesis of many forms of liver disease, including alcoholic and non-alcoholic liver disease. In this study we examined the efficacy of Misoprostol in modulating LPS-inducible TNFα and IL-10 expression in healthy human subjects and evaluated molecular mechanisms for Misoprostol modulation of cytokines in vitro. Healthy subjects were given 14day courses of Misoprostol at doses of 100, 200, and 300μg four times a day, in random order. Baseline and LPS-inducible cytokine levels were examined ex vivo in whole blood at the beginning and the end of the study. Additionally, in vitro studies were performed using primary human PBMCs and the murine macrophage cell line, RAW 264.7, to investigate underlying mechanisms of misoprostol on cytokine production. Administration of Misoprostol reduced LPS inducible TNF production by 29%, while increasing IL-10 production by 79% in human subjects with no significant dose effect on ex vivo cytokine activity; In vitro, the effect of Misoprostol was largely mediated by increased cAMP levels and consequent changes in CRE and NFκB activity, which are critical for regulating IL-10 and TNF expression. Additionally, chromatin immunoprecipitation (ChIP) studies demonstrated that Misoprostol treatment led to changes in transcription factor and RNA Polymerase II binding, resulting in changes in mRNA levels. In summary, Misoprostol was effective at beneficially modulating TNF and IL-10 levels both in vivo and in vitro; these studies suggest a potential rationale for Misoprostol use in ALD, NASH and other liver diseases where inflammation plays an etiologic role. PMID:26408955

  7. Lesbian camp: An unearthing.

    PubMed

    Nielsen, Elly-Jean

    2016-01-01

    Camp-a sensibility, a style, and a form of artistic self-expression-is an elusive concept said to be in the eye of the beholder. To refute Susan Sontag's ( 1966 ) claims that camp is apolitical and not especially homosexual, a number of recent scholarly works have been geared toward revealing camp's fundamental gayness. With the odd footnote aside, lesbian camp has been collapsed into the category of gay male camp, if not eclipsed entirely. Despite the negligible efforts made to legitimize lesbian camp, there are numerous salient cultural examples one might draw on to illustrate, typify, and substantiate a lesbian camp sensibility. I lay the ground work for this scholarly exercise by outlining various definitions and critiques of camp, and by discussing its history and application to queer theory. Then, to unveil lesbian camp, three non-mutually exclusive categories are discussed: classic, erotic, and radical. By gathering various strands of inquiry, and various textual examples (e.g., photography, artistic performances, and literary tropes), this article attempts to reach a more inclusive and nuanced understanding of lesbian camp. PMID:26701773

  8. Differential expression of 2':3'-cyclic nucleotide 3'-phosphodiesterase in cultured central, peripheral, and extraneural cells.

    PubMed

    Sprinkle, T J; McMorris, F A; Yoshino, J; DeVries, G H

    1985-07-01

    The relative levels of the central nervous system myelin marker enzyme 2':3'-cyclic nucleotide 3'-phosphodiesterase (EC 3.1.4.37, CNPase) were determined in neuroblastoma, astrocyte, oligodendrocyte and Schwann cell cultures and in freshly isolated human lymphocytes and platelets. The highest specific activities were associated with the cells that elaborate myelin membrane in the central and peripheral nervous system, oligodendrocytes and Schwann cells, respectively. Antiserum to bovine CNPase recognized both CNP1 and CNP2 in CNS myelin and human oligodendroglioma. In addition, a 53,000 dalton protein was evident on autoradiographs of immunoblotted PNS myelin and human oligodendroglioma proteins. Cultured rat oligodendrocyte, C6 and mouse NA neuroblastoma CNPase appear to share common determinants with the corresponding normal rat CNS enzyme. PMID:2995854

  9. Targeting Phosphodiesterases in Anti-platelet Therapy

    PubMed Central

    Rondina, Matthew T.; Weyrich, Andrew S.

    2013-01-01

    There are two primary modes of platelet inhibition: blockade of membrane receptors or neutralization of intracellular pathways. Both means of inhibition have proven benefits in the prevention and resolution of atherothrombotic events. With regard to intracellular inhibition, phosphodiesterases (PDEs) are fundamental for platelet function. Platelets possess several PDEs (PDE2, PDE3 and PDE5) that catalyze the hydrolysis of cyclic adenosine 3′-5′-monophosphate (cAMP) and cyclic guanosine 3′-5′-monophosphate (cGMP), thereby limiting the levels of intracellular nucleotides. PDE inhibitors, such as cilostazol and dipyridamole, dampen platelet function by increasing cAMP and cGMP levels. This review focuses on the roles of PDE inhibitors in modulating platelet function, with particular attention paid to drugs that have anti-platelet clinical indications. PMID:22918733

  10. Modulation of Polycystic Kidney Disease Severity by Phosphodiesterase 1 and 3 Subfamilies.

    PubMed

    Ye, Hong; Wang, Xiaofang; Sussman, Caroline R; Hopp, Katharina; Irazabal, Maria V; Bakeberg, Jason L; LaRiviere, Wells B; Manganiello, Vincent C; Vorhees, Charles V; Zhao, Haiqing; Harris, Peter C; van Deursen, Jan; Ward, Christopher J; Torres, Vicente E

    2016-05-01

    Aberrant intracellular calcium levels and increased cAMP signaling contribute to the development of polycystic kidney disease (PKD). cAMP can be hydrolyzed by various phosphodiesterases (PDEs). To examine the role of cAMP hydrolysis and the most relevant PDEs in the pathogenesis of PKD, we examined cyst development in Pde1- or Pde3-knockout mice on the Pkd2(-/WS25) background (WS25 is an unstable Pkd2 allele). These PDEs were selected because of their importance in cross-talk between calcium and cyclic nucleotide signaling (PDE1), control of cell proliferation and cystic fibrosis transmembrane conductance regulator (CFTR) -driven fluid secretion (PDE3), and response to vasopressin V2 receptor activation (both). In Pkd2(-/WS25) mice, knockout of Pde1a, Pde1c, or Pde3a but not of Pde1b or Pde3b aggravated the development of PKD and was associated with higher levels of protein kinase A-phosphorylated (Ser133) cAMP-responsive binding protein (P-CREB), activating transcription factor-1, and CREB-induced CRE modulator proteins in kidney nuclear preparations. Immunostaining also revealed higher expression of P-CREB in Pkd2(-/) (WS25);Pde1a(-/-), Pkd2(-) (/WS25);Pde1c(-/-), and Pkd2(-/) (WS25);Pde3a(-/-) kidneys. The cystogenic effect of desmopressin administration was markedly enhanced in Pkd2(-/WS25);Pde3a(-/-) mice, despite PDE3 accounting for only a small fraction of renal cAMP PDE activity. These observations show that calcium- and calmodulin-dependent PDEs (PDE1A and PDE1C) and PDE3A modulate the development of PKD, possibly through the regulation of compartmentalized cAMP pools that control cell proliferation and CFTR-driven fluid secretion. Treatments capable of increasing the expression or activity of these PDEs may, therefore, retard the development of PKD. PMID:26374610

  11. Atrazine acts as an endocrine disrupter by inhibiting cAMP-specific phosphodiesterase-4

    SciTech Connect

    Kucka, Marek; Pogrmic-Majkic, Kristina; Fa, Svetlana; Stojilkovic, Stanko S.; Kovacevic, Radmila

    2012-11-15

    Atrazine, one of the most commonly used herbicides worldwide, acts as an endocrine disruptor, but the mechanism of its action has not been characterized. In this study, we show that atrazine rapidly increases cAMP levels in cultured rat pituitary and testicular Leydig cells in a concentration-dependent manner, but less effectively than 3-isobutyl-1-methylxanthine, a competitive non-specific inhibitor of phosphodiesterases (PDEs). In forskolin (an activator of adenylyl cyclase)- and probenecid (an inhibitor of cyclic nucleotide transporters)-treated cells, but not in 3-isobutyl-1-methylxanthine-treated cells, atrazine further increased cAMP levels, indicating that inhibition of PDEs accounts for accumulation of cAMP. In contrast to cAMP, atrazine did not alter cGMP levels, further indicating that it inhibits cAMP-specific PDEs. Atrazine-induced changes in cAMP levels were sufficient to stimulate prolactin release in pituitary cells and androgen production in Leydig cells, indicating that it acts as an endocrine disrupter both in cells that secrete by exocytosis of prestored hormones and in cells that secrete by de novo hormone synthesis. Rolipram abolished the stimulatory effect of atrazine on cAMP release in both cell types, suggesting that it acts as an inhibitor of PDE4s, isoforms whose mRNA transcripts dominate in pituitary and Leydig cells together with mRNA for PDE8A. In contrast, immortalized lacto-somatotrophs showed low expression of these mRNA transcripts and several fold higher cAMP levels compared to normal pituitary cells, and atrazine was unable to further increase cAMP levels. These results indicate that atrazine acts as a general endocrine disrupter by inhibiting cAMP-specific PDE4s. -- Highlights: ► Atrazine stimulates cAMP accumulation in pituitary and Leydig cells. ► Atrazine also stimulates PRL and androgens secretion. ► Stimulatory effects of atrazine were abolished in cells with IBMX-inhibited PDEs. ► Atrazine specificity toward c

  12. Targeting Cyclic Nucleotide Phosphodiesterase in the Heart: Therapeutic Implications

    PubMed Central

    Miller, Clint L.

    2010-01-01

    The second messengers, cAMP and cGMP, regulate a number of physiological processes in the myocardium, from acute contraction/relaxation to chronic gene expression and cardiac structural remodeling. Emerging evidence suggests that multiple spatiotemporally distinct pools of cyclic nucleotides can discriminate specific cellular functions from a given cyclic nucleotide-mediated signal. Cyclic nucleotide phosphodiesterases (PDEs), by hydrolyzing intracellular cyclic AMP and/or cyclic GMP, control the amplitude, duration, and compartmentation of cyclic nucleotide signaling. To date, more than 60 different isoforms have been described and grouped into 11 broad families (PDE1–PDE11) based on differences in their structure, kinetic and regulatory properties, as well as sensitivity to chemical inhibitors. In the heart, PDE isozymes from at least six families have been investigated. Studies using selective PDE inhibitors and/or genetically manipulated animals have demonstrated that individual PDE isozymes play distinct roles in the heart by regulating unique cyclic nucleotide signaling microdomains. Alterations of PDE activity and/or expression have also been observed in various cardiac disease models, which may contribute to disease progression. Several family-selective PDE inhibitors have been used clinically or pre-clinically for the treatment of cardiac or vascular-related diseases. In this review, we will highlight both recent advances and discrepancies relevant to cardiovascular PDE expression, pathophysiological function, and regulation. In particular, we will emphasize how these properties influence current and future development of PDE inhibitors for the treatment of pathological cardiac remodeling and dysfunction. PMID:20632220

  13. Mechanically induced c-fos expression is mediated by cAMP in MC3T3-E1 osteoblasts

    NASA Technical Reports Server (NTRS)

    Fitzgerald, J.; Hughes-Fulford, M.

    1999-01-01

    In serum-deprived MC3T3-E1 osteoblasts, mechanical stimulation caused by mild (287 x g) centrifugation induced a 10-fold increase in mRNA levels of the proto-oncogene, c-fos. Induction of c-fos was abolished by the cAMP-dependent protein kinase inhibitor H-89, suggesting that the transient c-fos mRNA increase is mediated by cAMP. Down-regulation of protein kinase C (PKC) activity by chronic TPA treatment failed to significantly reduce c-fos induction, suggesting that TPA-sensitive isoforms of PKC are not responsible for c-fos up-regulation. In addition, 287 x g centrifugation increased intracellular prostaglandin E2 (PGE2) levels 2.8-fold (P<0. 005). Since we have previously shown that prostaglandin E2 (PGE2) can induce c-fos expression via a cAMP-mediated mechanism, we asked whether the increase in c-fos mRNA was due to centrifugation-induced PGE2 release. Pretreatment with the cyclooxygenase inhibitors indomethacin and flurbiprofen did not hinder the early induction of c-fos by mechanical stimulation. We conclude that c-fos expression induced by mild mechanical loading is dependent primarily on cAMP, not PKC, and initial induction of c-fos is not necessarily dependent on the action of newly synthesized PGE2.

  14. Antenatal Maternally-Administered Phosphodiesterase Type 5 Inhibitors Normalize eNOS Expression in the Fetal Lamb Model of Congenital Diaphragmatic Hernia

    PubMed Central

    Shue, Eveline H; Schecter, Samuel C.; Gong, Wenhui; Etemadi, Mozziyar; Johengen, Michael; Iqbal, Corey; Derderian, S. Christopher; Oishi, Peter; Fineman, Jeffrey R.; Miniati, Doug

    2013-01-01

    Purpose Pulmonary hypertension (pHTN), a main determinant of survival in congenital diaphragmatic hernia (CDH), results from in utero vascular remodeling. Phosphodiesterase type 5 (PDE5) inhibitors have never been used antenatally to treat pHTN. The purpose of this study is to determine if antenatal PDE5 inhibitors can prevent pHTN in the fetal lamb model of CDH. Methods CDH were created in pregnant ewes. Postoperatively, pregnant ewes received oral placebo or tadalafil, a PDE5 inhibitor, until delivery. Near term gestation, lambs underwent resuscitations, and lung tissue was snap frozen for protein analysis. Results Mean cGMP levels were 0.53±0.11 in placebo-treated fetal lambs and 1.73±0.21 in tadalafil-treated fetal lambs (p=0.002). Normalized expression of eNOS was 82±12% in Normal-Placebo, 61±5% in CDH-Placebo, 116±6% in Normal-Tadalafil, and 86±8% in CDH-Tadalafil lambs. Normalized expression of β-sGC was 105±15% in Normal-Placebo, 82±3% in CDH-Placebo, 158±16% in Normal-Tadalafil, and 86±8% in CDH-Tadalafil lambs. Endothelial NOS and β-sGC were significantly decreased in CDH (p = 0.0007 and 0.01 for eNOS and β-sGC, respectively), and tadalafil significantly increased eNOS expression (p = 0.0002). Conclusions PDE5 inhibitors can cross the placental barrier. β-sGC and eNOS are downregulated in fetal lambs with CDH. Antenatal PDE5 inhibitors normalize eNOS and may prevent in utero vascular remodeling in CDH. PMID:24439578

  15. Role of Phosphodiesterase 2 in Growth and Invasion of Human Malignant Melanoma Cells

    PubMed Central

    Hiramoto, Kenichi; Murata, Taku; Shimizu, Kasumi; Morita, Hiroshi; Inui, Madoka; Manganiello, Vincent C.; Tagawa, Toshiro; Arai, Naoya

    2014-01-01

    Cyclic nucleotide phosphodiesterases (PDEs) regulate the intracellular concentrations and effects of adenosine 3’,5’-cyclic monophosphate (cAMP) and guanosine 3’,5’-cyclic monophosphate (cGMP). The role of PDEs in malignant tumor cells is still uncertain. The role of PDEs, especially PDE2, in human malignant melanoma PMP cell line was examined in this study. In PMP cells, 8-bromo-cAMP, a cAMP analog, inhibited cell growth and invasion. However, 8-bromo-cGMP, a cGMP analog, had little or no effect. PDE2 and PDE4, but not PDE3, were expressed in PMP cells. Growth and invasion of PMP cells were inhibited by erythro-9-(2-Hydroxy-3-nonyl) adenine (EHNA), a specific PDE2 inhibitor, but not by rolipram, a specific PDE4 inhibitor. Moreover, cell growth and invasion were inhibited by transfection of small interfering RNAs (siRNAs) specific for PDE2A and a catalytically-dead mutant of PDE2A. After treating cells with EHNA or rolipram, intracellular cAMP concentrations were increased. Growth and invasion were stimulated by PKA14-22, a PKA inhibitor, and inhibited by N6-benzoyl-c AMP, a PKA specific cAMP analogue, whereas 8-(4-chlorophenylthio)-2’-O-methyl-cAMP, an Epac specific cAMP analogue, did not. Invasion, but not growth, was stimulated by A-kinase anchor protein (AKAP) St-Ht31 inhibitory peptide. Based on these results, PDE2 appears to play an important role in growth and invasion of the human malignant melanoma PMP cell line. Selectively suppressing PDE2 might possibly inhibit growth and invasion of other malignant tumor cell lines. PMID:24705027

  16. Role of phosphodiesterase 2 in growth and invasion of human malignant melanoma cells.

    PubMed

    Hiramoto, Kenichi; Murata, Taku; Shimizu, Kasumi; Morita, Hiroshi; Inui, Madoka; Manganiello, Vincent C; Tagawa, Toshiro; Arai, Naoya

    2014-09-01

    Cyclic nucleotide phosphodiesterases (PDEs) regulate the intracellular concentrations and effects of adenosine 3',5'-cyclic monophosphate (cAMP) and guanosine 3',5'-cyclic monophosphate (cGMP). The role of PDEs in malignant tumor cells is still uncertain. The role of PDEs, especially PDE2, in human malignant melanoma PMP cell line was examined in this study. In PMP cells, 8-bromo-cAMP, a cAMP analog, inhibited cell growth and invasion. However, 8-bromo-cGMP, a cGMP analog, had little or no effect. PDE2 and PDE4, but not PDE3, were expressed in PMP cells. Growth and invasion of PMP cells were inhibited by erythro-9-(2-hydroxy-3-nonyl) adenine (EHNA), a specific PDE2 inhibitor, but not by rolipram, a specific PDE4 inhibitor. Moreover, cell growth and invasion were inhibited by transfection of small interfering RNAs (siRNAs) specific for PDE2A and a catalytically-dead mutant of PDE2A. After treating cells with EHNA or rolipram, intracellular cAMP concentrations were increased. Growth and invasion were stimulated by PKA14-22, a PKA inhibitor, and inhibited by N(6)-benzoyl-c AMP, a PKA specific cAMP analog, whereas 8-(4-chlorophenylthio)-2'-O-methyl-cAMP, an Epac specific cAMP analog, did not. Invasion, but not growth, was stimulated by A-kinase anchor protein (AKAP) St-Ht31 inhibitory peptide. Based on these results, PDE2 appears to play an important role in growth and invasion of the human malignant melanoma PMP cell line. Selectively suppressing PDE2 might possibly inhibit growth and invasion of other malignant tumor cell lines. PMID:24705027

  17. cGMP Signaling, Phosphodiesterases and Major Depressive Disorder

    PubMed Central

    Reierson, Gillian W; Guo, Shuyu; Mastronardi, Claudio; Licinio, Julio; Wong, Ma-Li

    2011-01-01

    Deficits in neuroplasticity are hypothesized to underlie the pathophysiology of major depressive disorder (MDD): the effectiveness of antidepressants is thought to be related to the normalization of disrupted synaptic transmission and neurogenesis. The cyclic adenosine monophosphate (cAMP) signaling cascade has received considerable attention for its role in neuroplasticity and MDD. However components of a closely related pathway, the cyclic guanosine monophosphate (cGMP) have been studied with much lower intensity, even though this signaling transduction cascade is also expressed in the brain and the activity of this pathway has been implicated in learning and memory processes. Cyclic GMP acts as a second messenger; it amplifies signals received at postsynaptic receptors and activates downstream effector molecules resulting in gene expression changes and neuronal responses. Phosphodiesterase (PDE) enzymes degrade cGMP into 5’GMP and therefore they are involved in the regulation of intracellular levels of cGMP. Here we review a growing body of evidence suggesting that the cGMP signaling cascade warrants further investigation for its involvement in MDD and antidepressant action. PMID:22654729

  18. UCR1C is a novel activator of phosphodiesterase 4 (PDE4) long isoforms and attenuates cardiomyocyte hypertrophy

    PubMed Central

    Wang, Li; Burmeister, Brian T.; Johnson, Keven R.; Baillie, George S.; Karginov, Andrei V.; Skidgel, Randal A.; O’Bryan, John P.; Carnegie, Graeme K.

    2015-01-01

    Hypertrophy increases the risk of heart failure and arrhythmia. Prevention or reversal of the maladaptive hypertrophic phenotype has thus been proposed to treat heart failure. Chronic β-adrenergic receptor (β-AR) stimulation induces cardiomyocyte hypertrophy by elevating 3′, 5′-cyclic adenosine monophosphate (cAMP) levels and activating downstream effectors such protein kinase A (PKA). Conversely, hydrolysis of cAMP by phosphodiesterases (PDEs) spatiotemporally restricts cAMP signaling. Here, we demonstrate that PDE4, but not PDE3, is critical in regulating cardiomyocyte hypertrophy, and may represent a potential target for preventing maladaptive hypertrophy. We identify a sequence within the upstream conserved region 1 of PDE4D, termed UCR1C, as a novel activator of PDE4 long isoforms. UCR1C activates PDE4 in complex with A-Kinase anchoring protein (AKAP)-Lbc resulting in decreased PKA signaling facilitated by AKAP-Lbc. Expression of UCR1C in cardiomyocytes inhibits hypertrophy in response to chronic β-AR stimulation. This effect is partially due to inhibition of nuclear PKA activity, which decreases phosphorylation of the transcription factor cAMP response element-binding protein (CREB). In conclusion, PDE4 activation by UCR1C attenuates cardiomyocyte hypertrophy by specifically inhibiting nuclear PKA activity. PMID:25683917

  19. Angiotensin II increases phosphodiesterase 5A expression in vascular smooth muscle cells: A mechanism by which angiotensin II antagonizes cGMP signaling

    PubMed Central

    Kim, Dongsoo; Aizawa, Toru; Wei, Heng; Pi, Xinchun; Rybalkin, Sergei D.; Berk, Bradford C.; Yan, Chen

    2014-01-01

    Angiotensin II (Ang II) and nitric oxide (NO)/natriuretic peptide (NP) signaling pathways mutually regulate each other. Imbalance of Ang II and NO/NP has been implicated in the pathophysiology of many vascular diseases. cGMP functions as a key mediator in the interaction between Ang II and NO/NP. Cyclic nucleotide phosphodiesterase 5A (PDE5A) is important in modulating cGMP signaling by hydrolyzing cGMP in vascular smooth muscle cells (VSMC). Therefore, we examined whether Ang II negatively modulates intracellular cGMP signaling in VSMC by regulating PDE5A. Ang II rapidly and transiently increased PDE5A mRNA levels in rat aortic VSMC. Upregulation of PDE5A mRNA was associated with a time-dependent increase of both PDE5 protein expression and activity. Increased PDE5A mRNA level was transcription-dependent and mediated by the Ang II type 1 receptor. Ang II-mediated activation of extracellular signal-regulated kinases 1/2 (ERK1/2) was essential for Ang II-induced PDE5A upregulation. Pretreatment of VSMC with Ang II inhibited C-type NP (CNP) stimulated cGMP signaling, such as cGMP dependent protein kinase (PKG)-mediated phosphorylation of vasodilator-stimulated-phosphoprotein (VASP). Ang II-mediated inhibition of PKG was blocked when PDE5 activity was decreased by selective PDE5 inhibitors, suggesting that upregulation of PDE5A expression is an important mechanism for Ang II to attenuate cGMP signaling. PDE5A may also play a critical role in the growth promoting effects of Ang II because inhibition of PDE5A activity significantly decreased Ang II-stimulated VSMC growth. These observations establish a new mechanism by which Ang II antagonizes cGMP signaling and stimulates VSMC growth. PMID:15623434

  20. Increased cAMP levels modulate transforming growth factor-beta/Smad-induced expression of extracellular matrix components and other key fibroblast effector functions.

    PubMed

    Schiller, Meinhard; Dennler, Sylviane; Anderegg, Ulf; Kokot, Agatha; Simon, Jan C; Luger, Thomas A; Mauviel, Alain; Böhm, Markus

    2010-01-01

    cAMP is a key messenger of many hormones and neuropeptides, some of which modulate the composition of extracellular matrix. Treatment of human dermal fibroblasts with dibutyryl cyclic AMP and forskolin antagonized the inductive effects of transforming growth factor-beta (TGF-beta) on the expression of collagen, connective tissue growth factor, tissue inhibitor of matrix metalloproteinase-1, and plasminogen activator inhibitor type I, four prototypical TGF-beta-responsive genes. Increased intracellular cAMP prevented TGF-beta-induced Smad-specific gene transactivation, although TGF-beta-mediated Smad phosphorylation and nuclear translocation remained unaffected. However, increased cAMP levels abolished TGF-beta-induced interaction of Smad3 with its transcriptional co-activator cAMP-response element-binding protein (CREB)-binding protein (CBP)/p300. Overexpression of the transcriptional co-activator CBP/p300 rescued Smad-specific gene transcription in the presence of cAMP suggesting that sequestration of limited amounts of CBP/p300 by the activated cAMP/CREB pathway is the molecular basis of this inhibitory effect. These findings were extended by two functional assays. Increased intracellular cAMP levels suppressed the inductive activity of TGF-beta to contract mechanically unloaded collagen lattices and resulted in an attenuation of fibroblast migration of mechanically induced cell layer wounds. Of note, cAMP and TGF-beta synergistically induced hyaluronan synthase 2 (HAS2) expression and hyaluronan secretion, presumably via putative CREB-binding sites adjacent to Smad-binding sites within the HAS2 promoter. Our findings identify the cAMP pathway as a potent but differential and promoter-specific regulator of TGF-beta-mediated effects involved in extracellular matrix homeostasis. PMID:19858184

  1. Inhibition of hormone-sensitive lipase gene expression by cAMP and phorbol esters in 3T3-F442A and BFC-1 adipocytes.

    PubMed Central

    Plée-Gautier, E; Grober, J; Duplus, E; Langin, D; Forest, C

    1996-01-01

    Hormone-sensitive lipase (HSL) catalyses the rate-limiting step in adipocyte lipolysis. Short-term hormonal regulation of HSL activity is well characterized, whereas little is known about the control of HSL gene expression. We have measured HSL mRNA content of 3T3-F442A and BFC-1 adipocytes in response to the cAMP analogue 8-(4-chlorophenylthio)-cAMP (8-CPT-cAMP) and to the phorbol ester phorbol 12-myristate 13-acetate (PMA) by Northern blot, using a specific mouse cDNA fragment. Treatment of the cells for 12 or 6 h with, respectively, 0.5 mM 8-CPT-cAMP or 1 microM PMA produced a maximal decrease of about 60% in HSL mRNA. These effects were unaffected by the protein-synthesis inhibitor anisomycin, suggesting that cAMP and PMA actions were direct. The reduction in HSL mRNA was accompanied by a reduction in HSL total activity. The intracellular routes that cAMP and PMA follow for inducing such an effect seemed clearly independent. (i) After desensitization of the protein kinase C regulation pathway by a 24 h treatment of the cells with 1 microM PMA, PMA action was abolished whereas cAMP was still fully active. (ii) Treatment with saturating concentrations of both agents produced an additive effect. (iii) The synthetic glucocorticoid dexamethasone had no proper effect on HSL gene expression but potentiated cAMP action without affecting PMA action. cAMP inhibitory action on HSL is unexpected. Indeed, the second messenger of catecholamines is the main activator of HSL by phosphorylation. We envision that a long-term cAMP treatment of adipocytes induces a counter-regulatory process that reduces HSL content and, ultimately, limits fatty acid depletion from stored triacylglycerols. PMID:8836156

  2. Camp Chinkapin.

    ERIC Educational Resources Information Center

    Crook County School District, Prineville, OR.

    The Camp Chinkapin program, begun in 1957-58 as a pilot program for the State of Oregon, provides all sixth grade students in Crook County (Oregon) with a 5-day session in a resident camp setting in the early summer. The book serves as an introduction to and workbook for students attending the Crook County Outdoor Classroom at Suttle Lake. The…

  3. Cyclic AMP phosphodiesterase in Salmonella typhimurium: characteristics and physiological function.

    PubMed

    Botsford, J L

    1984-11-01

    The physiological function of cyclic AMP (cAMP) phosphodiesterase in Salmonella typhimurium was investigated with strains which were isogenic except for the cpd locus. In crude broken-cell extracts the properties of the enzyme were found to be similar to those reported for Escherichia coli. The specific activity in the mutant was less than 1% that in the wild type. Rates of cAMP production in the mutant were as much as twice those observed in the wild type. The amount of cAMP accumulated when cells grew overnight with limiting glucose was 4.5-fold greater in the mutant than in the wild type. The intracellular concentration of cAMP in the two strains was measured directly, using four different techniques to wash the cells to remove extracellular cAMP. The cAMP level in the cpd strain was only 25% greater than in the wild type. The functional concentration of the cAMP receptor protein-cAMP complex was estimated indirectly from the specific activity of beta-galactosidase in the two strains after introducing F'lac. When cells were grown with carbon sources permitting synthesis of different levels of cAMP, the specific activity of the enzyme was at most 25% greater in the cpd strain. The cpd strain was more sensitive to the effects of exogenous cAMP. Exogenous cAMP relieved both permanent and transient catabolite repression of the lac operon at lower concentrations in the cpd strain than in the wild type. When cells grew with glucose, glycerol, or ribose, exogenous cAMP inhibited growth of the mutant strain more than the wild type. PMID:6094495

  4. Constellation of HCN channels and cAMP regulating proteins in dendritic spines of the primate prefrontal cortex: potential substrate for working memory deficits in schizophrenia.

    PubMed

    Paspalas, Constantinos D; Wang, Min; Arnsten, Amy F T

    2013-07-01

    Schizophrenia associates with impaired prefrontal cortical (PFC) function and alterations in cyclic AMP (cAMP) signaling pathways. These include genetic insults to disrupted-in-schizophrenia (DISC1) and phosphodiesterases (PDE4s) regulating cAMP hydrolysis, and increased dopamine D1 receptor (D1R) expression that elevates cAMP. We used immunoelectron microscopy to localize DISC1, PDE4A, PDE4B, and D1R in monkey PFC and to view spatial interactions with hyperpolarization-activated cyclic nucleotide-gated (HCN) channels that gate network inputs when opened by cAMP. Physiological interactions between PDE4s and HCN channels were tested in recordings of PFC neurons in monkeys performing a spatial working memory task. The study reveals a constellation of cAMP-related proteins (DISC1, PDE4A, and D1R) and HCN channels next to excitatory synapses and the spine neck in thin spines of superficial PFC, where working memory microcircuits interconnect and spine loss is most evident in schizophrenia. In contrast, channels in dendrites were distant from synapses and cAMP-related proteins, and were associated with endosomal trafficking. The data suggest that a cAMP signalplex is selectively positioned in the spines to gate PFC pyramidal cell microcircuits. Single-unit recordings confirmed physiological interactions between cAMP and HCN channels, consistent with gating actions. These data may explain why PFC networks are especially vulnerable to genetic insults that dysregulate cAMP signaling. PMID:22693343

  5. Selective inhibition of two soluble adenosine cyclic 3',5'-phosphate phosphodiesterases partially purified from calf liver.

    PubMed

    Yamamoto, T; Lieberman, F; Osborne, J C; Manganiello, V C; Vaughan, M; Hidaka, H

    1984-02-14

    "Low Km" cAMP phosphodiesterase and cGMP-stimulated cyclic nucleotide phosphodiesterase activities were partially purified from calf liver supernatant by chromatography on DEAE-cellulose and DEAE-Sepharose and ammonium sulfate precipitation. The low Km phosphodiesterase was not retained on N6-H2N(CH2)2-cAMP-agarose and could be separated from the cGMP-stimulated phosphodiesterase which was absorbed by this matrix. From the proteins that did not bind, two distinct low Km cAMP phosphodiesterases were separated on Ultrogel AcA 34. One form (fraction C) hydrolyzed cAMP with an apparent Km of approximately 0.5 microM and was very sensitive to inhibition by cGMP. Lineweaver-Burk plots of cAMP hydrolysis by a second form (fraction B) were nonlinear, with an apparent low Km component of approximately 2 microM. This form was rather insensitive to inhibition by cGMP. With both fractions, hydrolysis of cAMP relative to cGMP was much greater at low (approximately 1 microM) than at high (approximately 100 microM) substrate concentrations. Maximal velocities for cAMP and cGMP were similar. From sedimentation equilibrium, the apparent weight-average molecular weight of fraction B was estimated as 174000, and that of fraction C was 85000. Another fraction (A) of cAMP phosphodiesterase eluted at the void volume of the AcA 34 column. On the basis of the relative affinities for cAMP and cGMP and inhibition by cGMP, fraction A is most likely an aggregated form of fraction B. No apparent interconversion of fractions A, B, or C was observed on high-performance liquid chromatography.(ABSTRACT TRUNCATED AT 250 WORDS) PMID:6324851

  6. Phosphodiesterase 8B and cyclic AMP signaling in the adrenal cortex.

    PubMed

    Leal, Leticia Ferro; Szarek, Eva; Faucz, Fabio; Stratakis, Constantine A

    2015-09-01

    Bilateral adrenocortical hyperplasia (BAH) in humans and mice has been recently linked to phosphodiesterase (PDE) 8B (PDE8B) and 11 (PDE11A) defects. These findings have followed the discovery that defects of primary genes of the cyclic monophosphatase (cAMP) signaling pathway, such as guanine nucleotide binding alpha subunit and PRKAR1A, are involved in the pathogenesis of BAH in humans; complete absence of Prkar1a in the adrenal cortex of mice also led to pathology that mimicked the human disease. Here, we review the most recent findings in human and mouse studies on PDE8B, a cAMP-specific PDE that appears to be highly expressed in the adrenal cortex and whose deficiency may underlie predisposition to BAH and possibly other human diseases. PMID:25971952

  7. Cytokine-Induced S-Nitrosylation of Soluble Guanylyl Cyclase and Expression of Phosphodiesterase 1A Contribute to Dysfunction of Longitudinal Smooth Muscle Relaxation

    PubMed Central

    Rajagopal, Senthilkumar; Nalli, Ancy D.; Kumar, Divya P.; Bhattacharya, Sayak; Hu, Wenhui; Mahavadi, Sunila; Grider, John R.

    2015-01-01

    The effect of proinflammatory cytokines on the expression and activity of soluble guanylyl cyclase (sGC) and cGMP–phosphodiesterases (PDEs) was determined in intestinal longitudinal smooth muscle. In control muscle cells, cGMP levels are regulated via activation of sGC and PDE5; the activity of the latter is regulated via feedback phosphorylation by cGMP-dependent protein kinase. In muscle cells isolated from muscle strips cultured with interleukin-1β (IL-1β) or tumor necrosis factor α (TNF-α) or obtained from the colon of TNBS (2,4,6-trinitrobenzene sulfonic acid)-treated mice, expression of inducible nitric oxide synthase (iNOS) was induced and sGC was S-nitrosylated, resulting in attenuation of nitric oxide (NO)–induced sGC activity and cGMP formation. The effect of cytokines on sGC S-nitrosylation and activity was blocked by the iNOS inhibitor 1400W [N-([3-(aminomethyl)phenyl]methyl)ethanimidamide dihydrochloride]. The effect of cytokines on cGMP levels measured in the absence of IBMX (3-isobutyl-1-methylxanthine), however, was partly reversed by 1400W or PDE1 inhibitor vinpocetine and completely reversed by a combination of 1400W and vinpocetine. Expression of PDE1A was induced and was accompanied by an increase in PDE1A activity in muscle cells isolated from muscle strips cultured with IL-1β or TNF-α or obtained from the colon of TNBS-treated mice; the effect of cytokines on PDE1 expression and activity was blocked by MG132 (benzyl N-[(2S)-4-methyl-1-[[(2S)-4-methyl-1-[[(2S)-4-methyl-1-oxopentan-2-yl]amino]-1-oxopentan-2-yl]amino]-1-oxopentan-2-yl]carbamate), an inhibitor of nuclear factor κB activity. NO-induced muscle relaxation was inhibited in longitudinal muscle cells isolated from muscle strips cultured with IL-1β or TNF-α or obtained from the colon of TNBS-treated mice, and this inhibition was completely reversed by the combination of both 1400W and vinpocetine. Inhibition of smooth muscle relaxation during inflammation reflects the

  8. Relationship between Platelet PPARs, cAMP Levels, and P-Selectin Expression: Antiplatelet Activity of Natural Products

    PubMed Central

    Fuentes, Eduardo; Palomo, Iván

    2013-01-01

    Platelets are no longer considered simply as cells participating in thrombosis. In atherosclerosis, platelets are regulators of multiple processes, with the recruitment of inflammatory cells towards the lesion sites, inflammatory mediators release, and regulation of endothelial function. The antiplatelet therapy has been used for a long time in an effort to prevent and treat cardiovascular diseases. However, limited efficacy in some patients, drug resistance, and side effects are limitations of current antiplatelet therapy. In this context, a large number of natural products (polyphenols, terpenoids, alkaloids, and fatty acids) have been reported with antiplatelet activity. In this sense, the present paper describes mechanisms of antiplatelet action of natural products on platelet P-selectin expression through cAMP levels and its role as peroxisome proliferator-activated receptors agonists. PMID:24324520

  9. CCR-08-0827 Version 2 Targeted inhibition of cyclic AMP phosphodiesterase-4 promotes brain tumor regression

    PubMed Central

    Goldhoff, Patricia; Warrington, Nicole; Limbrick, David D.; Hope, Andrew; Woerner, B. Mark; Jackson, Erin; Perry, Arie; Piwnica-Worms, David; Rubin, Joshua B.

    2008-01-01

    Statement of Clinical Relevance Therapies that can overcome the resistance of malignant brain tumors would be a major clinical advance. Here, we investigate the role of cAMP Phosphodiesterase-4 in stimulating brain tumor growth and the therapeutic utility of cAMP Phosphodiesterase-4 inhibition in the treatment of malignant brain tumors. Cyclic AMP Phosphodiesterase-4 was widely expressed in human brain tumors of glial and neuronal lineage, and forced expression of PDE4A1 accelerated intracranial glioblastoma and medulloblastoma xenograft growth. Moreover, targeted inhibition of PDE4, in combination with standard radiation and chemotherapy, induced a unique regression of established intracranial glioblastoma xenografts. These findings identify PDE4 as a novel molecular target for brain tumor therapy and indicate that PDE4 inhibition should be evaluated in clinical trials for malignant brain tumors. Purpose As favorable outcomes from malignant brain tumors remain limited by poor survival and treatment-related toxicity, novel approaches to cure are essential. Previously, we identified the cyclic AMP phosphodiesterase-4 (PDE4) inhibitor Rolipram as a potent anti-tumor agent. Here, we investigate the role of PDE4 in brain tumors and examine the utility of PDE4 as a therapeutic target. Experimental Design Immunohistochemistry was used to evaluate the expression pattern of a subfamily of PDE4, PDE4A, in multiple brain tumor types. To evaluate the effect of PDE4A on growth, a brain-specific isoform, PDE4A1 was overexpressed in xenografts of Daoy medulloblastoma and U87 glioblastoma cells. To determine therapeutic potential of PDE4 inhibition, Rolipram, temozolomide, and radiation were tested alone and in combination on mice bearing intracranial U87 xenografts. Results We found that PDE4A is expressed in medulloblastoma, glioblastoma, oligodendroglioma, ependymoma and meningioma. Moreover, when PDE4A1 was overexpressed in Daoy medulloblastoma and U87 glioblastoma cells, in

  10. Profiling of functional phosphodiesterase in mesangial cells using a CRE-SEAP-based reporting system

    PubMed Central

    Zhu, Ying; Yao, Jian; Meng, Yiman; Kasai, Ayumi; Hiramatsu, Nobuhiko; Hayakawa, Kunihiro; Miida, Takashi; Takeda, Masayuki; Okada, Masahiko; Kitamura, Masanori

    2006-01-01

    Phosphodiesterases (PDEs) are critically implicated in the regulation of mesangial cell function, but profile of functional PDEs in mesangial cells is still unclear. In this study, we investigated roles of individual PDEs in the regulation of mesangial cell behavior by the cAMP pathway. Reporter mesangial cells that express secreted alkaline phosphatase (SEAP) under the control of the cAMP response element (CRE) were exposed to selective PDE inhibitors in the presence or absence of cAMP, and activity of CRE, expression of CRE-regulated protein, mitogenesis and cell survival were examined. Exposure of reporter cells to cAMP-elevating agents resulted in time- and concentration-dependent activation of CRE. Treatment of the cells with any PDE inhibitors alone did not induce CRE activation. Under stimulation with 8-bromo-cAMP or 8-bromo-cGMP, however, inhibitors of PDE2, PDE3, PDE4 and PDE5 enhanced activation of CRE. Inhibition of PDE1 or PDE6 did not affect the CRE activation. Among different combinations tested, only inhibitors of PDE3 and PDE4 cooperatively increased the level of intracellular cAMP, activity of protein kinase A, activation of CRE, and CRE-regulated protein, connexin43. Concomitant inhibition of PDE3 and PDE4 attenuated mitogen-induced activation of extracellular signal-regulated kinases and cell proliferation. Under serum deprivation, combinational inhibition of PDE3 and PDE4 exclusively caused activation of caspase-3 and apoptosis. The present data elucidated that PDE3 and PDE4 play critical roles in the regulation of mesangial cell function. PDE3 and PDE4 were identified as the novel, antiapoptotic machinery that supports survival of mesangial cells. PMID:16751794

  11. Mechanisms Restricting Diffusion of Intracellular cAMP

    PubMed Central

    Agarwal, Shailesh R.; Clancy, Colleen E.; Harvey, Robert D.

    2016-01-01

    Although numerous receptors stimulate cAMP production in a wide array of cells, many elicit distinct, highly localized responses, implying that the subcellular distribution of cAMP is not uniform. One often used explanation is that phosphodiesterases, which breakdown cAMP, act as functional barriers limiting diffusion. However, several studies refute the notion that this is sufficient, suggesting that phosphodiesterase-independent movement of cAMP must occur at rates slower than free diffusion. But, until now this has never been demonstrated. Using Raster Image Correlation Spectroscopy (RICS), we measured the diffusion coefficient of a fluorescently-labeled cAMP derivative (φ450-cAMP) as well as other fluorescent molecules in order to investigate the role that molecular size, cell morphology, and buffering by protein kinase A (PKA) play in restricting cAMP mobility in different cell types. Our results demonstrate that cytosolic movement of cAMP is indeed much slower than the rate of free diffusion and that interactions with PKA, especially type II PKA associated with mitochondria, play a significant role. These findings have important implications with respect to cAMP signaling in all cells. PMID:26795432

  12. Identification and Localization of the Cyclic Nucleotide Phosphodiesterase 10A in Bovine Testis and Mature Spermatozoa

    PubMed Central

    Goupil, Serge; Maréchal, Loïze; El Hajj, Hassan; Tremblay, Marie-Ève; Richard, François J.

    2016-01-01

    In mammals, adenosine 3’, 5’-cyclic monophosphate (cAMP) is known to play highly important roles in sperm motility and acrosomal exocytosis. It is known to act through protein phosphorylation via PRKA and through the activation of guanine nucleotide exchange factors like EPAC. Sperm intracellular cAMP levels depend on the activity of adenylyl cyclases, mostly SACY, though transmembrane-containing adenylyl cyclases are also present, and on the activity of cyclic nucleotide phosphodiesterases (PDE) whose role is to degrade cAMP into 5’-AMP. The PDE superfamily is subdivided into 11 families (PDE1 to 11), which act on either cAMP or cGMP, or on both cAMP and cGMP although with different enzymatic properties. PDE10, which is more effective on cAMP than cGMP, has been known for almost 15 years and is mostly studied in the brain where it is associated with neurological disorders. Although a high level of PDE10A gene expression is observed in the testis, information on the identity of the isoforms or on the cell type that express the PDE10 protein is lacking. The objective of this study was to identify the PDE10A isoforms expressed in the testis and germ cells, and to determine the presence and localization of PDE10A in mature spermatozoa. As a sub-objective, since PDE10A transcript variants were reported strictly through analyses of bovine genomic sequence, we also wanted to determine the nucleotide and amino acid sequences by experimental evidence. Using RT-PCR, 5’- and 3’-RACE approaches we clearly show that PDE10A transcript variants X3 and X5 are expressed in bovine testis as well as in primary spermatocytes and spermatids. We also reveal using a combination of immunological techniques and proteomics analytical tools that the PDE10A isoform X4 is present in the area of the developing acrosome of spermatids and of the acrosome of mature spermatozoa. PMID:27548062

  13. The ever unfolding story of cAMP signaling in trypanosomatids: vive la difference!

    PubMed Central

    Tagoe, Daniel N. A.; Kalejaiye, Titilola D.; de Koning, Harry P.

    2015-01-01

    Kinetoplastids are unicellular, eukaryotic, flagellated protozoans containing the eponymous kinetoplast. Within this order, the family of trypanosomatids are responsible for some of the most serious human diseases, including Chagas disease (Trypanosoma cruzi), sleeping sickness (Trypanosoma brucei spp.), and leishmaniasis (Leishmania spp). Although cAMP is produced during the life cycle stages of these parasites, its signaling pathways are very different from those of mammals. The absence of G-protein-coupled receptors, the presence of structurally different adenylyl cyclases, the paucity of known cAMP effector proteins and the stringent need for regulation of cAMP in the small kinetoplastid cells all suggest a significantly different biochemical pathway and likely cell biology. However, each of the main kinetoplastid parasites express four class 1-type cyclic nucleotide-specific phosphodiesterases (PDEA-D), which have highly similar catalytic domains to that of human PDEs. To date, only TbrPDEB, expressed as two slightly different isoforms TbrPDEB1 and B2, has been found to be essential when ablated. Although the genomes contain reasonably well conserved genes for catalytic and regulatory domains of protein kinase A, these have been shown to have varied structural and functional roles in the different species. Recent discovery of a role of cAMP/AMP metabolism in a quorum-sensing signaling pathway in T. brucei, and the identification of downstream cAMP Response Proteins (CARPs) whose expression levels correlate with sensitivity to PDE inhibitors, suggests a complex signaling cascade. The interplay between the roles of these novel CARPs and the quorum-sensing signaling pathway on cell division and differentiation makes for intriguing cell biology and a new paradigm in cAMP signal transduction, as well as potential targets for trypanosomatid-specific cAMP pathway-based therapeutics. PMID:26441645

  14. Anti-platelet therapy: phosphodiesterase inhibitors

    PubMed Central

    Gresele, Paolo; Momi, Stefania; Falcinelli, Emanuela

    2011-01-01

    Inhibition of platelet aggregation can be achieved either by the blockade of membrane receptors or by interaction with intracellular signalling pathways. Cyclic adenosine 3′,5′-monophosphate (cAMP) and cyclic guanosine 3′,5′-monophosphate (cGMP) are two critical intracellular second messengers provided with strong inhibitory activity on fundamental platelet functions. Phosphodiesterases (PDEs), by catalysing the hydrolysis of cAMP and cGMP, limit the intracellular levels of cyclic nucleotides, thus regulating platelet function. The inhibition of PDEs may therefore exert a strong platelet inhibitory effect. Platelets possess three PDE isoforms (PDE2, PDE3 and PDE5), with different selectivity for cAMP and cGMP. Several nonselective or isoenzyme-selective PDE inhibitors have been developed, and some of them have entered clinical use as antiplatelet agents. This review focuses on the effect of PDE2, PDE3 and PDE5 inhibitors on platelet function and on the evidence for an antithrombotic action of some of them, and in particular of dipyridamole and cilostazol. PMID:21649691

  15. Cyclic Nucleotide Phosphodiesterases: important signaling modulators and therapeutic targets

    PubMed Central

    Ahmad, Faiyaz; Murata, Taku; Simizu, Kasumi; Degerman, Eva; Maurice, Donald; Manganiello, Vincent

    2014-01-01

    By catalyzing hydrolysis of cAMP and cGMP, cyclic nucleotide phosphodiesterases are critical regulators of their intracellular concentrations and their biological effects. Since these intracellular second messengers control many cellular homeostatic processes, dysregulation of their signals and signaling pathways initiate or modulate pathophysiological pathways related to various disease states, including erectile dysfunction, pulmonary hypertension, acute refractory cardiac failure, intermittent claudication, chronic obstructive pulmonary disease, and psoriasis. Alterations in expression of PDEs and PDE-gene mutations (especially mutations in PDE6, PDE8B, PDE11A and PDE4) have been implicated in various diseases and cancer pathologies. PDEs also play important role in formation and function of multi-molecular signaling/regulatory complexes called signalosomes. At specific intracellular locations, individual PDEs, together with pathway-specific signaling molecules, regulators, and effectors, are incorporated into specific signalosomes, where they facilitate and regulate compartmentalization of cyclic nucleotide signaling pathways and specific cellular functions. Currently, only a limited number of PDE inhibitors (PDE3, PDE4, PDE5 inhibitors) are used in clinical practice. Future paths to novel drug discovery include the crystal structure-based design approach, which has resulted in generation of more effective family-selective inhibitors, as well as burgeoning development of strategies to alter compartmentalized cyclic nucleotide signaling pathways by selectively targeting individual PDEs and their signalosome partners. PMID:25056711

  16. Astro Camp

    NASA Technical Reports Server (NTRS)

    1995-01-01

    Children who attend NASA's summer Astro Camp at Stennis Space Center enjoy a week of fun-filled activities. Campers learn what it is like to be a couple of inches taller in space and go through an astronaut obstacle course. They also learn how to build their own model rockets, which are launched on the last day of each camp. Campers also attend field trips to places such as the Challenger Learning Center at the Louisiana Arts and Science Center in Baton Rouge. Four weeks of Astro Camp are held during the summer each year-two camps for 8- to 10-year-olds and two for 11- to 13-year olds.

  17. Astro Camp

    NASA Technical Reports Server (NTRS)

    1996-01-01

    Children who attend NASA's summer Astro Camp at Stennis Space Center enjoy a week of fun-filled activities. Campers learn what it feels like to be a couple of inches taller in space and treck through an astronaut obstacle course. They also have the opportunity to build their own model rockets, which are then launched on the last day of each camp. Campers also travel on field trips to places such as the Challenger Learning Center at the Louisiana Arts and Science Center in Baton Rouge. Four weeks of Astro Camp are held each year during the summer-two camps for 8- to 10-year-olds and two for 11- to 13-year olds.

  18. Novel reciprocal regulation of cAMP signaling and apoptosis by orphan G-protein-coupled receptor GPRC5A gene expression

    SciTech Connect

    Hirano, Minoru; Zang, Liqing; Oka, Takehiko; Ito, Yoshiyuki; Shimada, Yasuhito; Nishimura, Yuhei; Tanaka, Toshio . E-mail: tanaka@doc.medic.mie-u.ac.jp

    2006-12-08

    GPRC5A is a member of G-protein-coupled receptors, which was originally identified as an all-trans-retinoic acid-induced gene. Although recent studies reported that this gene was highly expressed in the cancer cell lines and that GPRC5A might positively regulate cell proliferation, its mechanism remains unknown. We investigated the upstream and downstream signaling of GPRC5A and its biological function, and found that cAMP signaling is the novel GPRC5A induction pathway. When GPRC5A gene was overexpressed, intracellular cAMP concentration was decreased, and Gs{alpha} gene expression was downregulated. On the other hand, RNA interference of GPRC5A increased mRNA levels of Gs{alpha} and intracellular cAMP, reduced cell number, and induced apoptosis. Conversely, cell number was increased by GPRC5A overexpression. We first report the novel negative feedback model of cAMP signaling through GPRC5A gene expression. This evidence explains one of the mechanisms of the GPRC5A-regulated cell growth in some cancer cell lines.

  19. Phosphodiesterase 7 Inhibition Preserves Dopaminergic Neurons in Cellular and Rodent Models of Parkinson Disease

    PubMed Central

    Morales-Garcia, Jose A.; Redondo, Miriam; Alonso-Gil, Sandra; Gil, Carmen; Perez, Concepción; Martinez, Ana; Santos, Angel; Perez-Castillo, Ana

    2011-01-01

    Background Phosphodiesterase 7 plays a major role in down-regulation of protein kinase A activity by hydrolyzing cAMP in many cell types. This cyclic nucleotide plays a key role in signal transduction in a wide variety of cellular responses. In the brain, cAMP has been implicated in learning, memory processes and other brain functions. Methodology/Principal Findings Here we show a novel function of phosphodiesterase 7 inhibition on nigrostriatal dopaminergic neuronal death. We found that S14, a heterocyclic small molecule inhibitor of phosphodiesterase 7, conferred significant neuronal protection against different insults both in the human dopaminergic cell line SH-SY5Y and in primary rat mesencephalic cultures. S14 treatment also reduced microglial activation, protected dopaminergic neurons and improved motor function in the lipopolysaccharide rat model of Parkinson disease. Finally, S14 neuroprotective effects were reversed by blocking the cAMP signaling pathways that operate through cAMP-dependent protein kinase A. Conclusions/Significance Our findings demonstrate that phosphodiesterase 7 inhibition can protect dopaminergic neurons against different insults, and they provide support for the therapeutic potential of phosphodiesterase 7 inhibitors in the treatment of neurodegenerative disorders, particularly Parkinson disease. PMID:21390306

  20. Cyclic Amp phosphodiesterase activity in normal and inflamed human dental pulp.

    PubMed

    Spoto, G; Menna, V; Serra, E; Santoleri, F; Perfetti, G; Ciavarelli, L; Trentini, P

    2004-01-01

    Cyclic AMP phosphodiesterase (cAMP PDE) seems to be important in pulp tissues. High levels of cAMP PDE have been demonstrated to be in dental pulp cells. In the present study cAMP PDE activity was analyzed in normal healthy human dental pulps, in reversible pulpitis and in irreversible pulpitis. Enzymatic cAMP PDE control values for normal healthy pulps were 12.14 +/- 3.74 nmols/mg of proteins. In reversible pulpitis the cAMP PDE activity increased almost 2.5 times. In irreversible pulpitis specimens the values increased 4.5 times compared with normal healthy pulps activity. The differences between the groups (control vs. reversible pulpitis and vs. irreversible pulpitis) were statistically significant. These results could point to a role of cAMP PDE in the initial pulp response after injury. PMID:16857100

  1. The Accordant Trend of Both Parameters (rgs Expression and cAMP Content) Follows the Pattern of Development of Fruiting Body in Volvariella volvacea.

    PubMed

    Lu, Yuanping; Lian, Lingdan; Guo, Lixian; Xie, Bin; Wang, Wei; Chen, Bingzhi; van Peer, Arend Frans; Li, Shaojie; Wu, Taju; Xie, Baogui

    2015-11-01

    The formation of fruiting body in Volvariella volvacea is affected by endogenous genes and environmental factors. However, its regulation at a molecular level is still poorly understood. To study the genes involved in the formation of fruiting body, we cloned a new regulator of the G protein signaling (RGS) encoding gene (rgs) from V. volvacea. Phylogenetic analysis showed that RGS in V. volvacea and other basidiomycete RGS proteins from Schizophyllum commune and Coprinus cinereus belong to the same clade. In addition, we assayed intracellular cAMP content in the three developmental stages (mycelium, fruiting body primordia, and button). We also found that the expression of rgs was highly positively correlated to the content of intracellular cAMP during fruiting body formation. The conserved protein sequences and expression of rgs, together with high concent of cAMP at primordia tissue, suggested that rgs gene and cAMP may play a crucial role in fruiting body formation in V. volvacea. PMID:26264785

  2. [Hemophilia camps.

    PubMed

    Juárez-Sierra, Julieta; Del Pilar Torres-Arreola, Laura; Marín-Palomares, Teresa; Dueñas-González, María Teresa; Monteros-Rincón, Martha Patricia; Osorio-Guzmán, Maricela

    2013-01-01

    We reported the experience of hemophilia camps which was accomplished with patients from hospitals of the Instituto Mexicano del Seguro Social. The aim was to prepare the families and patients regarding the disease treatment, in order to promote the self sufficiency and to know the impact of the program on the course of the disease. Surveys were applied about treatment items and personal opinions were collected. The results of the national hemophilia camp were: group of 56 patients, average 14 years, 2 % women, 51 % severe hemophilia and 43 % had hemophilic brothers. Benefits: patients increased their knowledge about earlier bleeding identification and the self-infusion method; they became aware on their responsibility in self care, timely treatment and duties at home. Hemophilia camps with patients are an option for attitude change before disease complications. Social network creation and the increase in self-sufficiency are other benefits. PMID:24290020

  3. Characterization of phosphodiesterase 2A in human malignant melanoma PMP cells

    PubMed Central

    MORITA, HIROSHI; MURATA, TAKU; SHIMIZU, KASUMI; OKUMURA, KENYA; INUI, MADOKA; TAGAWA, TOSHIRO

    2013-01-01

    The prognosis for malignant melanoma is poor; therefore, new diagnostic methods and treatment strategies are urgently needed. Phosphodiesterase 2 (PDE2) is one of 21 phosphodiesterases, which are divided into 11 families (PDE1-PDE11). PDE2 hydrolyzes cyclic AMP (cAMP) and cyclic GMP (cGMP), and its binding to cGMP enhances the hydrolysis of cAMP. We previously reported the expression of PDE1, PDE3 and PDE5 in human malignant melanoma cells. However, the expression of PDE2 in these cells has not been investigated. Herein, we examined the expression of PDE2A and its role in human oral malignant melanoma PMP cells. Sequencing of RT-PCR products revealed that PDE2A2 was the only variant expressed in PMP cells. Four point mutations were detected; one missense mutation at nucleotide position 734 (from C to T) resulted in the substitution of threonine with isoleucine at amino acid position 214. The other three were silent mutations. An in vitro migration assay and a terminal deoxynucleotidyl transferase-mediated dUTP nick end-labeling assay revealed that suppressing PDE2 activity with its specific inhibitor, erythro-9-(2-hydroxy-3-nonyl)-adenine (EHNA), had no impact on cell motility or apoptosis. Furthermore, the cytotoxicity of EHNA, assessed using a trypan blue exclusion assay, was negligible. On the other hand, assessment of cell proliferation by BrdU incorporation and cell cycle analysis by flow cytometry revealed that EHNA treatment inhibited DNA synthesis and increased the percentage of G2/M-arrested cells. Furthermore, cyclin A mRNA expression was downregulated, while cyclin E mRNA expression was upregulated in EHNA-treated cells. Our results demonstrated that the PDE2A2 variant carrying point mutations is expressed in PMP cells and may affect cell cycle progression by modulating cyclin A expression. Thus, PDE2A2 is a possible new molecular target for the treatment of malignant melanoma. PMID:23381931

  4. Exposure to an Extremely-Low-Frequency Magnetic Field Stimulates Adrenal Steroidogenesis via Inhibition of Phosphodiesterase Activity in a Mouse Adrenal Cell Line

    PubMed Central

    Kitaoka, Kazuyoshi; Kawata, Shiyori; Yoshida, Tomohiro; Kadoriku, Fumiya; Kitamura, Mitsuo

    2016-01-01

    Extremely low-frequency magnetic fields (ELF-MFs) are generated by power lines and household electrical devices. In the last several decades, some evidence has shown an association between ELF-MF exposure and depression and/or anxiety in epidemiological and animal studies. The mechanism underlying ELF-MF-induced depression is considered to involve adrenal steroidogenesis, which is triggered by ELF-MF exposure. However, how ELF-MFs stimulate adrenal steroidogenesis is controversial. In the current study, we investigated the effect of ELF-MF exposure on the mouse adrenal cortex-derived Y-1 cell line and the human adrenal cortex-derived H295R cell line to clarify whether the ELF-MF stimulates adrenal steroidogenesis directly. ELF-MF exposure was found to significantly stimulate adrenal steroidogenesis (p < 0.01–0.05) and the expression of adrenal steroid synthetic enzymes (p < 0.05) in Y-1 cells, but the effect was weak in H295R cells. Y-1 cells exposed to an ELF-MF showed significant decreases in phosphodiesterase activity (p < 0.05) and intracellular Ca2+ concentration (p < 0.01) and significant increases in intracellular cyclic adenosine monophosphate (cAMP) concentration (p < 0.001–0.05) and cAMP response element-binding protein phosphorylation (p < 0.05). The increase in cAMP was not inhibited by treatment with NF449, an inhibitor of the Gs alpha subunit of G protein. Our results suggest that ELF-MF exposure stimulates adrenal steroidogenesis via an increase in intracellular cAMP caused by the inhibition of phosphodiesterase activity in Y-1 cells. The same mechanism may trigger the increase in adrenal steroid secretion in mice observed in our previous study. PMID:27100201

  5. The expression of inducible cAMP early repressor (ICER) is altered in prostate cancer cells and reverses the transformed phenotype of the LNCaP prostate tumor cell line.

    PubMed

    Yehia, G; Razavi, R; Memin, E; Schlotter, F; Molina, C A

    2001-08-15

    Inducible cAMP early repressor (ICER) has been shown to be an important mediator of cAMP antiproliferative activity. In this report, it was found that cAMP retards LNCaP cell growth; in contrast, cAMP inhibits the growth of PC-3 and DU-145 cells. ICER protein levels were markedly reduced in prostate cancer epithelial cells and undetectable and uninducible by cAMP in LNCaP and DU 145 cells. Forced expression of ICER in LNCaP cells caused inhibition of cell growth and thymidine incorporation and halted cells at the G(1) phase of the cell cycle. These ICER-bearing LNCaP cells were rendered unable to grow in soft agar and unable to form tumors in nude mice. These results suggest that deregulation of ICER expression may be related to carcinogenesis of the prostate gland. PMID:11507053

  6. A phosphodiesterase 4B-dependent interplay between tumor cells and the microenvironment regulates angiogenesis in B-cell lymphoma

    PubMed Central

    Suhasini, Avvaru N.; Lin, An-Ping; Bhatnagar, Harshita; Kim, Sang-Woo; Moritz, August W.; Aguiar, Ricardo C. T.

    2015-01-01

    Angiogenesis associates with poor outcome in diffuse large B-cell lymphoma (DLBCL), but the contribution of the lymphoma cells to this process remains unclear. Addressing this knowledge gap may uncover unsuspecting proangiogenic signaling nodes and highlight alternative antiangiogenic therapies. Here we identify the second messenger cyclic-AMP (cAMP) and the enzyme that terminates its activity, phosphodiesterase 4B (PDE4B), as regulators of B-cell lymphoma angiogenesis. We first show that cAMP, in a PDE4B-dependent manner, suppresses PI3K/AKT signals to down-modulate VEGF secretion and vessel formation in vitro. Next, we create a novel mouse model that combines the lymphomagenic Myc transgene with germline deletion of Pde4b. We show that lymphomas developing in a Pde4b-null background display significantly lower microvessel density in association with lower VEGF levels and PI3K/AKT activity. We recapitulate these observations by treating lymphoma-bearing mice with the FDA-approved PDE4 inhibitor Roflumilast. Lastly, we show that primary human DLBCLs with high PDE4B expression display significantly higher microvessel density. Here, we defined an unsuspected signaling circuitry in which the cAMP generated in lymphoma cells downmodulates PI3K/AKT and VEGF secretion to negatively influence vessel development in the microenvironment. These data identify PDE4 as an actionable antiangiogenic target in DLBCL. PMID:26503641

  7. Does phosphodiesterase 11A (PDE11A) hold promise as a future therapeutic target?

    PubMed

    Kelly, Michy P

    2015-01-01

    Phosphodiesterase 11A (PDE11A) is the most recently discovered 3', 5'-cyclic nucleotide phosphodiesterase. By breaking down both cAMP and cGMP, PDE11A is a critical regulator of intracellular signaling. To date, PDE11A has been implicated to play a role in tumorigenesis, brain function, and inflammation. Here, we consolidate and, where necessary, reconcile the PDE11A literature to evaluate this enzyme as a potential therapeutic target. We compare the results and methodologies of numerous studies that report conflicting tissue expression profiles for PDE11A. We conclude that PDE11A expression is relatively restricted in the body, with reliable expression reported in tissues such as the brain (particularly the hippocampus), the prostate, and the adrenal gland. Each of the four PDE11A splice variants (PDE11A1-4) appears to exhibit a distinct tissue expression profile and has a unique N-terminal regulatory region, suggesting that each isoform could be individually targeted with a small molecule or biologic. Progress has been made in identifying a tool PDE11A inhibitor as well as an activator; however, the functional effects of these pharmacological tools remain to be determined. Importantly, PDE11A knockout mice do exist and appear healthy into late age, suggesting a potential safety window for targeting this enzyme. Considering the implication of PDE11A in disease-relevant biology, the potential to selectively target specific PDE11A variants, and the possibility of either activating or inhibiting the enzyme, we believe PDE11A holds promise as a potential future therapeutic target. PMID:25159071

  8. Null mutations of the Dictyostelium cyclic nucleotide phosphodiesterase gene block chemotactic cell movement in developing aggregates.

    PubMed

    Sucgang, R; Weijer, C J; Siegert, F; Franke, J; Kessin, R H

    1997-12-01

    Extracellular cAMP is a critical messenger in the multicellular development of the cellular slime mold Dictyostelium discoideum. The levels of cAMP are controlled by a cyclic nucleotide phosphodiesterase (PDE) that is secreted by the cells. The PDE gene (pdsA) is controlled by three promoters that permit expression during vegetative growth, during aggregation, and in prestalk cells of the older structures. Targeted disruption of the gene aborts development, and complementation with a modified pdsA restores development. Two distinct promoters must be used for full complementation, and an inhibitory domain of the PDE must be removed. We took advantage of newly isolated PDE-null cells and the natural chimerism of the organism to ask whether the absence of PDE affected individual cell behavior. PDE-null cells aggregated with isogenic wild-type cells in chimeric mixtures, but could not move in a coordinated manner in mounds. The wild-type cells move inward toward the center of the mound, leaving many of the PDE-null cells at the periphery of the aggregate. During the later stages of development, PDE-null cells in the chimera segregate to regions which correspond to the prestalk region and the rear of the slug. Participation in the prespore/spore population returns with the restoration of a modified pdsA to the null cells. PMID:9405107

  9. Dual specificity and novel structural folding of yeast phosphodiesterase-1 for hydrolysis of second messengers cyclic adenosine and guanosine 3',5'-Monophosphate

    DOE PAGESBeta

    Tian, Yuanyuan; Cui, Wenjun; Huang, Manna; Robinson, Howard; Wan, Yiqian; Wang, Yousheng; Ke, Hengming

    2014-08-05

    Cyclic nucleotide phosphodiesterases (PDEs) decompose second messengers cAMP and cGMP that play critical roles in many physiological processes. PDE1 of Saccharomyces cerevisiae has been subcloned and expressed in Escherichia coli. Recombinant yPDE1 has a KM of 110 μM and a kcat of 16.9 s⁻¹ for cAMP and a KM of 105 μM and a kcat of 11.8 s₅⁻¹ for cGMP. Thus, the specificity constant (kcat/KMcAMP)/(kcat/KMcGMP) of 1.4 indicates a dual specificity of yPDE1 for hydrolysis of both cAMP and cGMP. The crystal structures of unliganded yPDE1 and its complex with GMP at 1.31 Å resolution reveal a new structural foldingmore » that is different from those of human PDEs but is partially similar to that of some other metalloenzymes such as metallo-β-lactamase. In spite of their different structures and divalent metals, yPDE1 and human PDEs may share a common mechanism for hydrolysis of cAMP and cGMP.« less

  10. Expression of three mammalian cDNAs that interfere with RAS function in Saccharomyces cerevisiae.

    PubMed Central

    Colicelli, J; Nicolette, C; Birchmeier, C; Rodgers, L; Riggs, M; Wigler, M

    1991-01-01

    Saccharomyces cerevisiae strains expressing the activated RAS2Val19 gene or lacking both cAMP phosphodiesterase genes, PDE1 and PDE2, have impaired growth control and display an acute sensitivity to heat shock. We have isolated two classes of mammalian cDNAs from yeast expression libraries that suppress the heat shock-sensitive phenotype of RAS2Val19 strain. Members of the first class of cDNAs also suppress the heat shock-sensitive phenotype of pde1- pde2- strains and encode cAMP phosphodiesterases. Members of the second class fail to suppress the phenotype of pde1- pde2- strains and therefore are candidate cDNAs encoding proteins that interact with RAS proteins. We report the nucleotide sequence of three members of this class. Two of these cDNAs share considerable sequence similarity, but none are clearly similar to previously isolated genes. Images PMID:1849280

  11. High-throughput screening of phosphodiesterase activity in living cells.

    PubMed

    Rich, Thomas C; Karpen, Jeffrey W

    2005-01-01

    Phosphodiesterases (PDEs) hydrolyze the second messengers cyclic adenosine monophosphate (cAMP) and cyclic guanosine 5'-monophosphate (cGMP) and play a crucial role in the termination and spatial segregation of cyclic nucleotide signals. Despite a wealth of molecular information, very little is known about how PDEs regulate cAMP and cGMP signals in living cells because conventional methods lack the necessary spatial and temporal resolution. We present here a sensitive optical method for monitoring cAMP levels and PDE activity near the membrane, using cyclic nucleotide-gated (CNG) ion channels as sensors. These channels are directly opened by the binding of cyclic nucleotides and allow cations to cross the membrane. The olfactory channel A subunit (CNGA2) has been genetically modified to improve its cAMP sensitivity and specificity. Channel activity is assessed by measuring Ca2+ influx using standard fluorometric techniques. In addition to studying PDEs in their native setting, the approach should be particularly useful in high-throughput screening assays to test for compounds that affect PDE activity, as well as the activities of the many G protein-coupled receptors that cause changes in intracellular cAMP. PMID:15988054

  12. Phosphodiesterase-4 modulation as a potential therapeutic for cognitive loss in pathological and non-pathological aging: possibilities and pitfalls.

    PubMed

    Hansen, Rolf T; Zhang, Han-Ting

    2015-01-01

    Phosphodiesterases (PDEs) are a super family of 11 enzyme families responsible for the hydrolysis of the intracellular secondary messengers cyclic AMP (cAMP) and cyclic GMP (cGMP). PDE4, in particular, is highly expressed in brain regions involved with regulation of memory, anxiety, and depression, including the hippocampus, amygdala, and nucleus accumbens. Senescence has been shown to result in extreme dysregulation of the cAMP pathway in various brain regions. Thus, as a critical controller of intracellular cAMP levels, PDE4 may be a potential target for the treatment of senescence-related cognitive disorders, which could be pathological and/or non-pathological in origin. While there is great potential in the development of novel PDE4 inhibitors for treatment of senescent-cognition impairment, there are also currently many pitfalls that need to be overcome. PDE4 has four subfamilies (PDE4A, B, C, and D) that are differentially expressed throughout the brain and body, as well as at least 25 splice variants derived from alternative splicing and multiple promoter sites. PDE4 subtypes have been shown to have differential effects on behavior, and cAMP itself has also been shown to play a contrasting role in behavior in different brain regions. This review will focus on what is currently understood about PDE4 in aging, the potential for PDE4 modulation as a cognitive therapy, and current pitfalls and limitations that need to be overcome in the PDE4 field. Overall, furthering our understanding of this incredibly complex pathway may one day assist with the development of novel therapeutics for both pathological and non-pathological cognitive disorders associated with senescence. PMID:25159075

  13. The effect of resveratrol on beta amyloid-induced memory impairment involves inhibition of phosphodiesterase-4 related signaling.

    PubMed

    Wang, Gang; Chen, Ling; Pan, Xiaoyu; Chen, Jiechun; Wang, Liqun; Wang, Weijie; Cheng, Ruochuan; Wu, Fan; Feng, Xiaoqing; Yu, Yingcong; Zhang, Han-Ting; O'Donnell, James M; Xu, Ying

    2016-04-01

    Resveratrol, a natural polyphenol found in red wine, has wide spectrum of pharmacological properties including antioxidative and antiaging activities. Beta amyloid peptides (Aβ) are known to involve cognitive impairment, neuroinflammatory and apoptotic processes in Alzheimer's disease (AD). Activation of cAMP and/or cGMP activities can improve memory performance and decrease the neuroinflammation and apoptosis. However, it remains unknown whether the memory enhancing effect of resveratrol on AD associated cognitive disorders is related to the inhibition of phosphodiesterase 4 (PDE4) subtypes and subsequent increases in intracellular cAMP and/or cGMP activities. This study investigated the effect of resveratrol on Aβ1-42-induced cognitive impairment and the participation of PDE4 subtypes related cAMP or cGMP signaling. Mice microinfused with Aβ1-42 into bilateral CA1 subregions displayed learning and memory impairment, as evidenced by reduced memory acquisition and retrieval in the water maze and retention in the passive avoidance tasks; it was also significant that neuroinflammatory and pro-apoptotic factors were increased in Aβ1-42-treated mice. Aβ1-42-treated mice also increased in PDE4A, 4B and 4D expression, and decreased in PKA level. However, PKA inhibitor H89, but not PKG inhibitor KT5823, prevented resveratrol's effects on these parameters. Resveratrol also reversed Aβ1-42-induced decreases in phosphorylated cAMP response-element binding protein (pCREB), brain derived neurotrophic factor (BDNF) and anti-apoptotic factor BCl-2 expression, which were reversed by H89. These findings suggest that resveratrol reversing Aβ-induced learning and memory disorder may involve the regulation of neuronal inflammation and apoptosis via PDE4 subtypes related cAMP-CREB-BDNF signaling. PMID:26980711

  14. The effect of resveratrol on beta amyloid-induced memory impairment involves inhibition of phosphodiesterase-4 related signaling

    PubMed Central

    Wang, Gang; Chen, Ling; Pan, Xiaoyu; Chen, Jiechun; Wang, Liqun; Wang, Weijie; Cheng, Ruochuan; Wu, Fan; Feng, Xiaoqing; Yu, Yingcong; Zhang, Han-Ting; O'Donnell, James M.; Xu, Ying

    2016-01-01

    Resveratrol, a natural polyphenol found in red wine, has wide spectrum of pharmacological properties including antioxidative and antiaging activities. Beta amyloid peptides (Aβ) are known to involve cognitive impairment, neuroinflammatory and apoptotic processes in Alzheimer's disease (AD). Activation of cAMP and/or cGMP activities can improve memory performance and decrease the neuroinflammation and apoptosis. However, it remains unknown whether the memory enhancing effect of resveratrol on AD associated cognitive disorders is related to the inhibition of phosphodiesterase 4 (PDE4) subtypes and subsequent increases in intracellular cAMP and/or cGMP activities. This study investigated the effect of resveratrol on Aβ1-42-induced cognitive impairment and the participation of PDE4 subtypes related cAMP or cGMP signaling. Mice microinfused with Aβ1-42 into bilateral CA1 subregions displayed learning and memory impairment, as evidenced by reduced memory acquisition and retrieval in the water maze and retention in the passive avoidance tasks; it was also significant that neuroinflammatory and pro-apoptotic factors were increased in Aβ1-42-treated mice. Aβ1-42-treated mice also increased in PDE4A, 4B and 4D expression, and decreased in PKA level. However, PKA inhibitor H89, but not PKG inhibitor KT5823, prevented resveratrol's effects on these parameters. Resveratrol also reversed Aβ1-42-induced decreases in phosphorylated cAMP response-element binding protein (pCREB), brain derived neurotrophic factor (BDNF) and anti-apoptotic factor BCl-2 expression, which were reversed by H89. These findings suggest that resveratrol reversing Aβ-induced learning and memory disorder may involve the regulation of neuronal inflammation and apoptosis via PDE4 subtypes related cAMP-CREB-BDNF signaling. PMID:26980711

  15. A fluorescence polarization assay for cyclic nucleotide phosphodiesterases.

    PubMed

    Huang, Wei; Zhang, Yan; Sportsman, J Richard

    2002-06-01

    Cyclic nucleotide phosphodiesterases (PDEs) catalyze the hydrolysis of the 3'-ester bond of cyclic AMP (cAMP) and cyclic GMP (cGMP), important second messengers in the transduction of a variety of extracellular signals. There is growing interest in the study of PDEs as drug targets for novel therapeutics. We describe the development of a homogeneous fluorescence polarization assay for PDEs based on the strong binding of PDE reaction products (i.e., AMP or GMP) onto modified nanoparticles through interactions with immobilized trivalent metal cations. This assay technology (IMAP) is applicable to both cAMP- and cGMP-specific PDEs. Results of the assay in 384- and 1536-well microplates are presented. PMID:12097184

  16. Phosphodiesterase 4D Inhibitors Limit Prostate Cancer Growth Potential

    PubMed Central

    Powers, Ginny L.; Hammer, Kimberly D.P.; Domenech, Maribella; Frantskevich, Katsiaryna; Malinowski, Rita L.; Bushman, Wade; Beebe, David J.; Marker, Paul C.

    2014-01-01

    Phosphodiesterase 4D (PDE4D) has recently been implicated as a proliferation-promoting factor in prostate cancer and is over-expressed in human prostate carcinoma. However, the effects of PDE4D inhibition using pharmacological inhibitors have not been examined in prostate cancer. These studies examined the effects of selective PDE4D inhibitors, NVP-ABE171 and cilomilast, as anti-prostate cancer therapies in both in vitro and in vivo models. The effects of PDE4D inhibitors on pathways that are critical in prostate cancer and/or downstream of cyclic AMP (cAMP) were examined. Both NVP-ABE171 and cilomilast decreased cell growth. In vitro, PDE4D inhibitors lead to decreased signaling of the sonic hedgehog (SHH), Androgen Receptor (AR), and MAPK pathways, but growth inhibition was best correlated to the sonic hedgehog pathway. PDE4D inhibition also reduced proliferation of epithelial cells induced by paracrine signaling from co-cultured stromal cells that had activated hedgehog signaling. In addition, PDE4D inhibitors decreased the weight of the prostate in wild-type mice. Prostate cancer xenografts grown in nude mice that were treated with cilomilast or NVP-ABE171 had decreased wet weight and increased apoptosis compared to vehicle treated controls. These studies suggest the pharmacological inhibition of PDE4D using small molecule inhibitors is an effective option for prostate cancer therapy. Implications PDE4D inhibitors decrease the growth of prostate cancer cells in vivo and in vitro, and PDE4D inhibition has therapeutic potential in prostate cancer. PMID:25149359

  17. In vivo model with targeted cAMP biosensor reveals changes in receptor-microdomain communication in cardiac disease.

    PubMed

    Sprenger, Julia U; Perera, Ruwan K; Steinbrecher, Julia H; Lehnart, Stephan E; Maier, Lars S; Hasenfuss, Gerd; Nikolaev, Viacheslav O

    2015-01-01

    3',5'-cyclic adenosine monophosphate (cAMP) is an ubiquitous second messenger that regulates physiological functions by acting in distinct subcellular microdomains. Although several targeted cAMP biosensors are developed and used in single cells, it is unclear whether such biosensors can be successfully applied in vivo, especially in the context of disease. Here, we describe a transgenic mouse model expressing a targeted cAMP sensor and analyse microdomain-specific second messenger dynamics in the vicinity of the sarcoplasmic/endoplasmic reticulum calcium ATPase (SERCA). We demonstrate the biocompatibility of this targeted sensor and its potential for real-time monitoring of compartmentalized cAMP signalling in adult cardiomyocytes isolated from a healthy mouse heart and from an in vivo cardiac disease model. In particular, we uncover the existence of a phosphodiesterase-dependent receptor-microdomain communication, which is affected in hypertrophy, resulting in reduced β-adrenergic receptor-cAMP signalling to SERCA. PMID:25917898

  18. Marketing Your Day Camp.

    ERIC Educational Resources Information Center

    Coleman, George

    1997-01-01

    Marketing strategies for day camps include encouraging camp staff to get involved in organizations involving children, families, and communities; holding camp fairs; offering the use of camp facilities to outside groups; hosting sport leagues and local youth outings; planning community fairs; and otherwise involving the camp in the community. (LP)

  19. IGF-I–induced Differentiation of L6 Myogenic Cells Requires the Activity of cAMP-Phosphodiesterase

    PubMed Central

    De Arcangelis, Vania; Coletti, Dario; Conti, Marco; Lagarde, Michel; Molinaro, Mario; Adamo, Sergio; Nemoz, Georges; Naro, Fabio

    2003-01-01

    Inhibition of type 4 cAMP-specific phosphodiesterase (PDE4) activity in L6-C5 and L6-E9 abolished myogenic differentiation induced by low-serum medium and IGF-I. L6-C5 cells cultured in low-serum medium displayed a PDE4 activity higher than cells cultured in serum-free medium, a condition not sufficient to induce differentiation. In the presence of serum, PDE4D3, the major isoform natively expressed in L6-C5 cells, translocated to a Triton-insoluble fraction, which increased the PDE specific activity of the fraction, and exhibited a Mr shift typical of phosphorylation of this isoform. Furthermore, serum promoted the localization of PDE4D3 to a vesicular subcellular compartment. In L6-C5 cells, IGF-I is a stronger inducer of myogenic differentiation in the presence than in absence of serum. Its ability to trigger differentiation in the absence of serum was restored by overexpressing wild-type PDE4D3, but not a phosphorylation-insensitive mutant. This finding was confirmed in single cells overexpressing a GFP-PDE4D3 fusion protein by assessing nuclear accumulation of myogenin in both L6-C5 and L6-E9. Overexpression of other PDE isoforms was less efficient, confirming that PDE4D3 is the physiologically relevant phosphodiesterase isoform in the control of myogenesis. These results show that downregulation of cAMP signaling through cAMP-phosphodiesterase stimulation is a prerequisite for induction of myogenesis. PMID:12686596

  20. cAMP-Specific Phosphodiesterases 8A and 8B, Essential Regulators of Leydig Cell Steroidogenesis

    PubMed Central

    Shimizu-Albergine, Masami; Tsai, Li-Chun Lisa; Patrucco, Enrico

    2012-01-01

    Phosphodiesterase (PDE) 8A and PDE8B are high-affinity, cAMP-specific phosphodiesterases that are highly expressed in Leydig cells. PDE8A is largely associated with mitochondria, whereas PDE8B is broadly distributed in the cytosol. We used a new, PDE8-selective inhibitor, PF-04957325, and genetically ablated PDE8A(−/−), PDE8B(−/−) and PDE8A(−/−)/B(−/−) mice to determine roles for these PDEs in the regulation of testosterone production. PF-04957325 treatment of WT Leydig cells or MA10 cells increased steroid production but had no effect in PDE8A (−/−)/B(−/−) double-knockout cells, confirming the selectivity of the drug. Moreover, under basal conditions, cotreatment with PF-04957325 plus rolipram, a PDE4-selective inhibitor, synergistically potentiated steroid production. These results suggest that the pool(s) of cAMP regulating androgen production are controlled by PDE8s working in conjunction with PDE4. Likewise, PDE8A (−/−)/B(−/−) cells had higher testosterone production than cells from either PDE8A(−/−) or PDE8B(−/−) mice, suggesting that both PDE8s work in concert to regulate steroid production. We further demonstrate that combined inhibition of PDE8s and PDE4 greatly increased PKA activity including phosphorylation of cholesterol-ester hydrolase (CEH)/hormone-sensitive lipase (HSL). CEH/HSL phosphorylation also was increased in PDE8A(−/−)/B(−/−) cells compared with WT cells. Finally, combined inhibition of PDE8s and PDE4 increased the expression of steroidogenic acute regulatory (StAR) protein. Together these findings suggest that both PDE8A and PDE8B play essential roles to maintain low cAMP levels, thereby suppressing resting steroidogenesis by keeping CEH/HSL inactive and StAR protein expression low. They also suggest that in order for PDE inhibitor therapy to be an effective stimulator of steroidogenesis, both PDE8 isozymes and PDE4 need to be simultaneously targeted. PMID:22232524

  1. Transcriptional and post‐transcriptional regulation of monocyte chemoattractant protein‐3 gene expression in human endothelial cells by phorbol ester and cAMP signalling

    PubMed Central

    Kondo, A; Isaji, S; Nishimura, Y; Tanaka, T

    2000-01-01

    Monocyte chemoattractant protein‐3 (MCP‐3) is one of the most broadly active chemokines, potentially inducing chemotaxis of all leucocytic cells. In the present study, we examined the regulation of MCP‐3 mRNA and protein production in endothelial cells by protein kinase C (PKC) activator, phorbol 12‐myristate 13‐acetate (PMA) and cAMP signalling. On stimulation of endothelial cells with 10 nm PMA, MCP‐3 mRNA increased to 300‐fold the basal level at 3 hr and rapidly declined to 0·2‐fold the basal level at 24 hr. PMA‐induced MCP‐3 mRNA and protein production of human endothelial cells were partially inhibited by pretreatment with the adenylate cyclase activator, forskolin, or membrane‐permeable cAMP derivative. The PMA‐induced MCP‐3 mRNA increase was almost abrogated when cells were pretreated with cycloheximide (CHX). Forskolin inhibited the transcription of PMA‐induced MCP‐3 gene expression. Following PMA stimulation for 3 hr, subsequent addition of actinomycin D suppressed the rapid decay of PMA‐induced MCP‐3 mRNA. These results suggest that PMA induces the transcriptional activation of the MCP‐3 gene through de novo protein synthesis and the rapid decay of PMA‐induced MCP‐3 mRNA through de novo synthesis of adenosine/uridine (AU)‐rich element binding proteins and cAMP signalling inhibits the PMA‐induced transcriptional activation of the MCP‐3 gene expression. PMID:10792504

  2. Parallel Allostery by cAMP and PDE Coordinates Activation and Termination Phases in cAMP Signaling.

    PubMed

    Krishnamurthy, Srinath; Tulsian, Nikhil Kumar; Chandramohan, Arun; Anand, Ganesh S

    2015-09-15

    The second messenger molecule cAMP regulates the activation phase of the cAMP signaling pathway through high-affinity interactions with the cytosolic cAMP receptor, the protein kinase A regulatory subunit (PKAR). Phosphodiesterases (PDEs) are enzymes responsible for catalyzing hydrolysis of cAMP to 5' AMP. It was recently shown that PDEs interact with PKAR to initiate the termination phase of the cAMP signaling pathway. While the steps in the activation phase are well understood, steps in the termination pathway are unknown. Specifically, the binding and allosteric networks that regulate the dynamic interplay between PKAR, PDE, and cAMP are unclear. In this study, PKAR and PDE from Dictyostelium discoideum (RD and RegA, respectively) were used as a model system to monitor complex formation in the presence and absence of cAMP. Amide hydrogen/deuterium exchange mass spectrometry was used to monitor slow conformational transitions in RD, using disordered regions as conformational probes. Our results reveal that RD regulates its interactions with cAMP and RegA at distinct loci by undergoing slow conformational transitions between two metastable states. In the presence of cAMP, RD and RegA form a stable ternary complex, while in the absence of cAMP they maintain transient interactions. RegA and cAMP each bind at orthogonal sites on RD with resultant contrasting effects on its dynamics through parallel allosteric relays at multiple important loci. RD thus serves as an integrative node in cAMP termination by coordinating multiple allosteric relays and governing the output signal response. PMID:26276689

  3. Interprofessional Flight Camp.

    PubMed

    Alfes, Celeste M; Rowe, Amanda S

    2016-01-01

    The Dorothy Ebersbach Academic Center for Flight Nursing in Cleveland, OH, holds an annual flight camp designed for master's degree nursing students in the acute care nurse practitioner program, subspecializing in flight nursing at the Frances Payne Bolton School of Nursing at Case Western Reserve University. The weeklong interprofessional training is also open to any health care provider working in an acute care setting and focuses on critical care updates, trauma, and emergency care within the critical care transport environment. This year, 29 graduate nursing students enrolled in a master's degree program from Puerto Rico attended. Although the emergency department in Puerto Rico sees and cares for trauma patients, there is no formal trauma training program. Furthermore, the country only has 1 rotor wing air medical transport service located at the Puerto Rico Medical Center in San Juan. Flight faculty and graduate teaching assistants spent approximately 9 months planning for their participation in our 13th annual flight camp. Students from Puerto Rico were extremely pleased with the learning experiences at camp and expressed particular interest in having more training time within the helicopter flight simulator. PMID:27021671

  4. Phosphodiesterase types 3 and 4 regulate the phasic contraction of neonatal rat bladder smooth myocytes via distinct mechanisms.

    PubMed

    Zhai, Kui; Chang, Yan; Wei, Bin; Liu, Qinghua; Leblais, Véronique; Fischmeister, Rodolphe; Ji, Guangju

    2014-05-01

    Activation of the cyclic AMP (cAMP) pathway reduces bladder contractility. However, the role of phosphodiesterase (PDE) families in regulating this function is poorly understood. Here, we compared the contractile function of the cAMP hydrolyzing PDEs in neonatal rat bladder smooth myocytes. RT-PCR and Western blotting analysis revealed that several isoforms of PDE1-4 were expressed in neonatal rat bladder. While 8-methoxymethyl-3-isobutyl-1-methylxanthine (a PDE1 inhibitor) and BAY-60-7550 (a PDE2 inhibitor) had no effect on the carbachol-enhanced phasic contractions of bladder strips, cilostamide (Cil, a PDE3 inhibitor) and Ro-20-1724 (Ro, a PDE4 inhibitor) significantly reduced these contractions. This inhibitory effect of Ro was blunted by the PKA inhibitor H-89, while the inhibitory effect of Cil was strongly attenuated by the PKG inhibitor KT 5823. Application of Ro in single bladder smooth myocytes resulted in an increase in Ca(2+) spark frequency but a decrease both in Ca(2+) transients and in sarcoplasmic reticulum (SR) Ca(2+) content. In contrast, Cil had no effect on these events. Furthermore, Ro-induced inhibition of the phasic contractions was significantly blocked by ryanodine and iberiotoxin. Taken together, PDE3 and PDE4 are the main PDE isoforms in maintaining the phasic contractions of bladder smooth myocytes, with PDE4 being functionally more active than PDE3. However, their roles are mediated through different mechanisms. PMID:24463006

  5. Heterozygous mutations in cyclic AMP phosphodiesterase-4D (PDE4D) and protein kinase A (PKA) provide new insights into the molecular pathology of acrodysostosis.

    PubMed

    Kaname, Tadashi; Ki, Chang-Seok; Niikawa, Norio; Baillie, George S; Day, Jonathan P; Yamamura, Ken-Ichi; Ohta, Tohru; Nishimura, Gen; Mastuura, Nobuo; Kim, Ok-Hwa; Sohn, Young Bae; Kim, Hyun Woo; Cho, Sung Yoon; Ko, Ah-Ra; Lee, Jin Young; Kim, Hyun Wook; Ryu, Sung Ho; Rhee, Hwanseok; Yang, Kap-Seok; Joo, Keehyoung; Lee, Jooyoung; Kim, Chi Hwa; Cho, Kwang-Hyun; Kim, Dongsan; Yanagi, Kumiko; Naritomi, Kenji; Yoshiura, Ko-Ichiro; Kondoh, Tatsuro; Nii, Eiji; Tonoki, Hidefumi; Houslay, Miles D; Jin, Dong-Kyu

    2014-11-01

    Acrodysostosis without hormone resistance is a rare skeletal disorder characterized by brachydactyly, nasal hypoplasia, mental retardation and occasionally developmental delay. Recently, loss-of-function mutations in the gene encoding cAMP-hydrolyzing phosphodiesterase-4D (PDE4D) have been reported to cause this rare condition but the pathomechanism has not been fully elucidated. To understand the pathogenetic mechanism of PDE4D mutations, we conducted 3D modeling studies to predict changes in the binding efficacy of cAMP to the catalytic pocket in PDE4D mutants. Our results indicated diminished enzyme activity in the two mutants we analyzed (Gly673Asp and Ile678Thr; based on PDE4D4 residue numbering). Ectopic expression of PDE4D mutants in HEK293 cells demonstrated this reduction in activity, which was identified by increased cAMP levels. However, the cells from an acrodysostosis patient showed low cAMP accumulation, which resulted in a decrease in the phosphorylated cAMP Response Element-Binding Protein (pCREB)/CREB ratio. The reason for this discrepancy was due to a compensatory increase in expression levels of PDE4A and PDE4B isoforms, which accounted for the paradoxical decrease in cAMP levels in the patient cells expressing mutant isoforms with a lowered PDE4D activity. Skeletal radiographs of 10-week-old knockout (KO) rats showed that the distal part of the forelimb was shorter than in wild-type (WT) rats and that all the metacarpals and phalanges were also shorter in KO, as the name acrodysostosis implies. Like the G-protein α-stimulatory subunit and PRKAR1A, PDE4D critically regulates the cAMP signal transduction pathway and influences bone formation in a way that activity-compromising PDE4D mutations can result in skeletal dysplasia. We propose that specific inhibitory PDE4D mutations can lead to the molecular pathology of acrodysostosis without hormone resistance but that the pathological phenotype may well be dependent on an over-compensatory induction

  6. Victory Junction Gang Camp

    ERIC Educational Resources Information Center

    Shell, Ryan

    2007-01-01

    This article describes the Victory Junction Gang Camp, a not-for-profit, NASCAR-themed camp for children with chronic medical conditions that serves 24 different disease groups. The mission of the camp is to give children life-changing camping experiences that are exciting, fun, and empowering in a safe and medically sound environment. While doing…

  7. Camp through the Decades.

    ERIC Educational Resources Information Center

    Nicodemus, Teresa

    2003-01-01

    Camp pioneers relate how camping has grown to become more diverse, environmentally aware, safe, and conscious of its responsibility to promote healthy development of children. Changing trends in clothing, transportation, and food preparation at camp are described. The joys, discoveries, and teachable moments that camp offers children have endured.…

  8. Astro Camp Plus

    NASA Technical Reports Server (NTRS)

    2006-01-01

    Stennis Space Center's new Astro Camp Plus camp kicked off June 19 for teens ages 13-15. The new camp delves more deeply into the science, math and technology concepts introduced in the center's popular Astro Camp series. Campers including Jasmyne White (left) and Dana Yingst, both of Slidell, La., learn how NASA uses 'podcasting' to broadcast video, and made their own podcasts.

  9. Cyclic AMP phosphodiesterases in the zebra finch: distribution, cloning and characterization of a PDE4B homolog.

    PubMed

    Thompson, B E; Freking, F; Pho, V; Schlinger, B A; Cherry, J A

    2000-11-10

    Songbirds are important animal models for studying neural mechanisms underlying learning and memory. While evidence has emerged that cAMP plays a significant role in invertebrate and mammalian learning, little is known about the role of cAMP pathways in regulating neuronal function in birds. With the goal of identifying important components of this pathway, we report the first cloning of a cAMP-specific, Type IV phosphodiesterase (PDE4) in a non-mammalian vertebrate. A combination of PCR analysis and cDNA library screening was used to show that homologs of the four known mammalian PDE4 genes also exist in zebra finch. A full-length cDNA representing the zebra finch homolog of PDE4B1 was isolated from a telencephalic library. Expression of this cDNA in human embryonic kidney 293 (HEK) cells yielded an enzyme that hydrolyzed cAMP with a low K(m) and was inhibited by micromolar concentrations of rolipram; these properties are typical of all known mammalian PDE4s. In brain, northern blots revealed transcripts of 3.6 and 4.4 kb in adults, but only the 3.6 kb transcript in juveniles, suggesting that PDE4 expression is developmentally regulated. In situ hybridization of tissue sections demonstrated that PDE4 message was distributed widely throughout the adult zebra finch brain, including regions controlling the learning of songs and the acquisition of spatial memories. These data suggest that PDE4 enzymes may influence a variety of brain functions in these birds and play a role in learning. PMID:11072099

  10. A Computational Modeling and Simulation Approach to Investigate Mechanisms of Subcellular cAMP Compartmentation.

    PubMed

    Yang, Pei-Chi; Boras, Britton W; Jeng, Mao-Tsuen; Docken, Steffen S; Lewis, Timothy J; McCulloch, Andrew D; Harvey, Robert D; Clancy, Colleen E

    2016-07-01

    Subcellular compartmentation of the ubiquitous second messenger cAMP has been widely proposed as a mechanism to explain unique receptor-dependent functional responses. How exactly compartmentation is achieved, however, has remained a mystery for more than 40 years. In this study, we developed computational and mathematical models to represent a subcellular sarcomeric space in a cardiac myocyte with varying detail. We then used these models to predict the contributions of various mechanisms that establish subcellular cAMP microdomains. We used the models to test the hypothesis that phosphodiesterases act as functional barriers to diffusion, creating discrete cAMP signaling domains. We also used the models to predict the effect of a range of experimentally measured diffusion rates on cAMP compartmentation. Finally, we modeled the anatomical structures in a cardiac myocyte diad, to predict the effects of anatomical diffusion barriers on cAMP compartmentation. When we incorporated experimentally informed model parameters to reconstruct an in silico subcellular sarcomeric space with spatially distinct cAMP production sites linked to caveloar domains, the models predict that under realistic conditions phosphodiesterases alone were insufficient to generate significant cAMP gradients. This prediction persisted even when combined with slow cAMP diffusion. When we additionally considered the effects of anatomic barriers to diffusion that are expected in the cardiac myocyte dyadic space, cAMP compartmentation did occur, but only when diffusion was slow. Our model simulations suggest that additional mechanisms likely contribute to cAMP gradients occurring in submicroscopic domains. The difference between the physiological and pathological effects resulting from the production of cAMP may be a function of appropriate compartmentation of cAMP signaling. Therefore, understanding the contribution of factors that are responsible for coordinating the spatial and temporal

  11. A Computational Modeling and Simulation Approach to Investigate Mechanisms of Subcellular cAMP Compartmentation

    PubMed Central

    Yang, Pei-Chi; Boras, Britton W.; Jeng, Mao-Tsuen; Lewis, Timothy J.; McCulloch, Andrew D.; Harvey, Robert D.; Clancy, Colleen E.

    2016-01-01

    Subcellular compartmentation of the ubiquitous second messenger cAMP has been widely proposed as a mechanism to explain unique receptor-dependent functional responses. How exactly compartmentation is achieved, however, has remained a mystery for more than 40 years. In this study, we developed computational and mathematical models to represent a subcellular sarcomeric space in a cardiac myocyte with varying detail. We then used these models to predict the contributions of various mechanisms that establish subcellular cAMP microdomains. We used the models to test the hypothesis that phosphodiesterases act as functional barriers to diffusion, creating discrete cAMP signaling domains. We also used the models to predict the effect of a range of experimentally measured diffusion rates on cAMP compartmentation. Finally, we modeled the anatomical structures in a cardiac myocyte diad, to predict the effects of anatomical diffusion barriers on cAMP compartmentation. When we incorporated experimentally informed model parameters to reconstruct an in silico subcellular sarcomeric space with spatially distinct cAMP production sites linked to caveloar domains, the models predict that under realistic conditions phosphodiesterases alone were insufficient to generate significant cAMP gradients. This prediction persisted even when combined with slow cAMP diffusion. When we additionally considered the effects of anatomic barriers to diffusion that are expected in the cardiac myocyte dyadic space, cAMP compartmentation did occur, but only when diffusion was slow. Our model simulations suggest that additional mechanisms likely contribute to cAMP gradients occurring in submicroscopic domains. The difference between the physiological and pathological effects resulting from the production of cAMP may be a function of appropriate compartmentation of cAMP signaling. Therefore, understanding the contribution of factors that are responsible for coordinating the spatial and temporal

  12. Role of Membrane Microdomains in Compartmentation of cAMP Signaling

    PubMed Central

    Agarwal, Shailesh R.; Yang, Pei-Chi; Rice, Monica; Singer, Cherie A.; Nikolaev, Viacheslav O.; Lohse, Martin J.; Clancy, Colleen E.; Harvey, Robert D.

    2014-01-01

    Spatially restricting cAMP production to discrete subcellular locations permits selective regulation of specific functional responses. But exactly where and how cAMP signaling is confined is not fully understood. Different receptors and adenylyl cyclase isoforms responsible for cAMP production are not uniformly distributed between lipid raft and non-lipid raft domains of the plasma membrane. We sought to determine the role that these membrane domains play in organizing cAMP responses in HEK293 cells. The freely diffusible FRET-based biosensor Epac2-camps was used to measure global cAMP responses, while versions of the probe targeted to lipid raft (Epac2-MyrPalm) and non-raft (Epac2-CAAX) domains were used to monitor local cAMP production near the plasma membrane. Disruption of lipid rafts by cholesterol depletion selectively altered cAMP responses produced by raft-associated receptors. The results indicate that receptors associated with lipid raft as well as non-lipid raft domains can contribute to global cAMP responses. In addition, basal cAMP activity was found to be significantly higher in non-raft domains. This was supported by the fact that pharmacologic inhibition of adenylyl cyclase activity reduced basal cAMP activity detected by Epac2-CAAX but not Epac2-MyrPalm or Epac2-camps. Responses detected by Epac2-CAAX were also more sensitive to direct stimulation of adenylyl cyclase activity, but less sensitive to inhibition of phosphodiesterase activity. Quantitative modeling was used to demonstrate that differences in adenylyl cyclase and phosphodiesterase activities are necessary but not sufficient to explain compartmentation of cAMP associated with different microdomains of the plasma membrane. PMID:24752595

  13. Bidirectional promoters link cAMP signaling with irreversible differentiation through promoter-associated non-coding RNA (pancRNA) expression in PC12 cells

    PubMed Central

    Yamamoto, Naoki; Agata, Kiyokazu; Nakashima, Kinichi; Imamura, Takuya

    2016-01-01

    Bidirectional promoters are the major source of gene activation-associated noncoding RNA (ncRNA). PC12 cells offer an interesting model for understanding the mechanism underlying bidirectional promoter-mediated cell cycle control. Nerve growth factor (NGF)-stimulated PC12 cells elongate neurites, and are in a reversible cell-cycle-arrested state. In contrast, these cells irreversibly differentiate and cannot re-enter the normal cell cycle after NGF plus cAMP treatment. In this study, using directional RNA-seq, we found that bidirectional promoters for protein-coding genes with promoter-associated ncRNA (pancRNA) were enriched for cAMP response element consensus sequences, and were preferred targets for transcriptional regulation by the transcription factors in the cAMP-dependent pathway. A spindle-formation-associated gene, Nusap1 and pancNusap1 were among the most strictly co-transcribed pancRNA–mRNA pairs. This pancRNA–mRNA pair was specifically repressed in irreversibly differentiated PC12 cells. Knockdown (KD) and overexpression experiments showed that pancNusap1 positively regulated the Nusap1 expression in a sequence-specific manner, which was accompanied by histone acetylation at the Nusap1 promoter. Furthermore, pancNusap1 KD recapitulated the effects of cAMP on cell cycle arrest. Thus, we conclude that pancRNA-mediated histone acetylation contributes to the establishment of the cAMP-induced transcription state of the Nusap1 locus and contributes to the irreversible cell cycle exit for terminal differentiation of PC12 cells. PMID:26945044

  14. Bidirectional promoters link cAMP signaling with irreversible differentiation through promoter-associated non-coding RNA (pancRNA) expression in PC12 cells.

    PubMed

    Yamamoto, Naoki; Agata, Kiyokazu; Nakashima, Kinichi; Imamura, Takuya

    2016-06-20

    Bidirectional promoters are the major source of gene activation-associated noncoding RNA (ncRNA). PC12 cells offer an interesting model for understanding the mechanism underlying bidirectional promoter-mediated cell cycle control. Nerve growth factor (NGF)-stimulated PC12 cells elongate neurites, and are in a reversible cell-cycle-arrested state. In contrast, these cells irreversibly differentiate and cannot re-enter the normal cell cycle after NGF plus cAMP treatment. In this study, using directional RNA-seq, we found that bidirectional promoters for protein-coding genes with promoter-associated ncRNA (pancRNA) were enriched for cAMP response element consensus sequences, and were preferred targets for transcriptional regulation by the transcription factors in the cAMP-dependent pathway. A spindle-formation-associated gene, Nusap1 and pancNusap1 were among the most strictly co-transcribed pancRNA-mRNA pairs. This pancRNA-mRNA pair was specifically repressed in irreversibly differentiated PC12 cells. Knockdown (KD) and overexpression experiments showed that pancNusap1 positively regulated the Nusap1 expression in a sequence-specific manner, which was accompanied by histone acetylation at the Nusap1 promoter. Furthermore, pancNusap1 KD recapitulated the effects of cAMP on cell cycle arrest. Thus, we conclude that pancRNA-mediated histone acetylation contributes to the establishment of the cAMP-induced transcription state of the Nusap1 locus and contributes to the irreversible cell cycle exit for terminal differentiation of PC12 cells. PMID:26945044

  15. Histamine H2 receptor antagonism by T-593: studies on cAMP generation in Hepa cells expressing histamine H2 receptor.

    PubMed

    Tashiro, T; Ono, K; Watanabe, T; Inoie, M; Arai, H; Kimura, S; Kurokawa, K

    1999-07-01

    Histamine H2 receptor antagonism by T-593 was investigated in Hepa cells expressing canine histamine H2 receptors. T-593 inhibited generation of cAMP in Hepa cells stimulated by 10(-5) mol/l histamine with an IC50 value of 2.3 x 10(-6) mol/l, (S)-(-)-T-593, one of the enantiomers comprising racemic T-593, inhibited cAMP generation with an IC50 value of 6.1 x 10(-7) mol/l. On the other hand, the other enantiomer (R)-(+)-T-593 exhibited only a negligible effect. Incubation of the cell with (S)-(-)-T-593 for 60 min depressed the maximal response of the concentration-response curve of histamine with a nonparallel rightward shift. The slope of a Schild plot was 1.27. In contrast, (S)-(-)-T-593 caused a parallel rightward shift of the curve, with a Schild plot slope that did not significantly differ from unity, by treating the cells for 15 min. The H2 receptor-blocking action of (S)-(-)-T-593 remained almost unaffected after washing out the drug, whereas the effect of ranitidine was reversible after washing. These results suggest that T-593 possesses a time-dependent insurmountable antagonistic action against histamine H2 receptor. T-593 may interact with the histamine H2 receptor molecule in a slowly associable and dissociable manner. PMID:10352421

  16. Age-related changes of cyclic AMP phosphodiesterase activity in rat brain regions and a new phosphodiesterase inhibitor--nootropic agent adafenoxate.

    PubMed

    Stancheva, S L; Alova, L G

    1991-01-01

    1. The low- and high-KM cyclic AMP phosphodiesterase (cAMP PDE) activity in cerebral cortex, striatum, hypothalamus and hippocampus of young (4-5-month-old) and aged (22-month-old) rats has been studied. 2. A significant rise in the high-KM cAMP PDE activity in the cerebral cortex, hypothalamus and hippocampus in aged rats has been found. 3. The activity of the low-KM cAMP PDE does not change during senescence in all the brain structures studied. 4. In a series of increased concentrations (from 5 x 10(-4) to 1 x 10(-5) M) adafenoxate inhibits low- and high-KM cAMP PDE in most of the brain structures studied in both age groups. 5. The present results provide evidence for realization of the CNS effects of adafenoxate through inhibition of cAMP PDE activity and regulation of the intracellular level of cAMP. PMID:1662175

  17. Clinical and Molecular Genetics of the Phosphodiesterases (PDEs)

    PubMed Central

    Azevedo, Monalisa F.; Faucz, Fabio R.; Bimpaki, Eirini; Horvath, Anelia; Levy, Isaac; de Alexandre, Rodrigo B.; Ahmad, Faiyaz; Manganiello, Vincent

    2014-01-01

    Cyclic nucleotide phosphodiesterases (PDEs) are enzymes that have the unique function of terminating cyclic nucleotide signaling by catalyzing the hydrolysis of cAMP and GMP. They are critical regulators of the intracellular concentrations of cAMP and cGMP as well as of their signaling pathways and downstream biological effects. PDEs have been exploited pharmacologically for more than half a century, and some of the most successful drugs worldwide today affect PDE function. Recently, mutations in PDE genes have been identified as causative of certain human genetic diseases; even more recently, functional variants of PDE genes have been suggested to play a potential role in predisposition to tumors and/or cancer, especially in cAMP-sensitive tissues. Mouse models have been developed that point to wide developmental effects of PDEs from heart function to reproduction, to tumors, and beyond. This review brings together knowledge from a variety of disciplines (biochemistry and pharmacology, oncology, endocrinology, and reproductive sciences) with emphasis on recent research on PDEs, how PDEs affect cAMP and cGMP signaling in health and disease, and what pharmacological exploitations of PDEs may be useful in modulating cyclic nucleotide signaling in a way that prevents or treats certain human diseases. PMID:24311737

  18. Regulation of the pro-inflammatory cytokine osteopontin by GIP in adipocytes - A role for the transcription factor NFAT and phosphodiesterase 3B

    SciTech Connect

    Omar, Bilal; Banke, Elin; Guirguis, Emilia; Aakesson, Lina; Manganiello, Vincent; Lyssenko, Valeriya; Groop, Leif; Gomez, Maria F.; Degerman, Eva

    2012-09-07

    Highlights: Black-Right-Pointing-Pointer GIP stimulates lipogenesis and osteopontin expression in primary adipocytes. Black-Right-Pointing-Pointer GIP-induced osteopontin expression is NFAT-dependent. Black-Right-Pointing-Pointer Osteopontin expression is PDE3-dependent. Black-Right-Pointing-Pointer Osteopontin expression is increased in PDE3B KO mice. -- Abstract: The incretin - glucose-dependent insulinotropic polypeptide (GIP) - and the pro-inflammatory cytokine osteopontin are known to have important roles in the regulation of adipose tissue functions. In this work we show that GIP stimulates lipogenesis and osteopontin expression in primary adipocytes. The GIP-induced increase in osteopontin expression was inhibited by the NFAT (the transcription factor nuclear factor of activated T-cells) inhibitor A-285222. Also, the NFAT kinase glycogen synthase kinase (GSK) 3 was upregulated by GIP. To test whether cAMP might be involved in GIP-mediated effects on osteopontin a number of strategies were used. Thus, the {beta}3-adrenergic receptor agonist CL316,243 stimulated osteopontin expression, an effects which was mimicked by OPC3911, a specific inhibitor of phosphodiesterase 3. Furthermore, treatment of phosphodiesterase 3B knock-out mice with CL316,243 resulted in a dramatic upregulation of osteopontin in adipose tissue which was not the case in wild-type mice. In summary, we delineate mechanisms by which GIP stimulates osteopontin in adipocytes. Given the established link between osteopontin and insulin resistance, our data suggest that GIP by stimulating osteopontin expression, also could promote insulin resistance in adipocytes.

  19. Intracellular tortuosity underlies slow cAMP diffusion in adult ventricular myocytes

    PubMed Central

    Richards, Mark; Lomas, Oliver; Jalink, Kees; Ford, Kerrie L.; Vaughan-Jones, Richard D.; Lefkimmiatis, Konstantinos; Swietach, Pawel

    2016-01-01

    Aims 3′,5′-Cyclic adenosine monophosphate (cAMP) signals in the heart are often confined to concentration microdomains shaped by cAMP diffusion and enzymatic degradation. While the importance of phosphodiesterases (degradative enzymes) in sculpting cAMP microdomains is well established in cardiomyocytes, less is known about cAMP diffusivity (DcAMP) and factors affecting it. Many earlier studies have reported fast diffusivity, which argues against sharply defined microdomains. Methods and results [cAMP] dynamics in the cytoplasm of adult rat ventricular myocytes were imaged using a fourth generation genetically encoded FRET-based sensor. The [cAMP]-response to the addition and removal of isoproterenol (β-adrenoceptor agonist) quantified the rates of cAMP synthesis and degradation. To obtain a read out of DcAMP, a stable [cAMP] gradient was generated using a microfluidic device which delivered agonist to one half of the myocyte only. After accounting for phosphodiesterase activity, DcAMP was calculated to be 32 µm2/s; an order of magnitude lower than in water. Diffusivity was independent of the amount of cAMP produced. Saturating cAMP-binding sites with the analogue 6-Bnz-cAMP did not accelerate DcAMP, arguing against a role of buffering in restricting cAMP mobility. cAMP diffused at a comparable rate to chemically unrelated but similar sized molecules, arguing for a common physical cause of restricted diffusivity. Lower mitochondrial density and order in neonatal cardiac myocytes allowed for faster diffusion, demonstrating the importance of mitochondria as physical barriers to cAMP mobility. Conclusion In adult cardiac myocytes, tortuosity due to physical barriers, notably mitochondria, restricts cAMP diffusion to levels that are more compatible with microdomain signalling. PMID:27089919

  20. 1,3-Dichloro-2-propanol inhibits progesterone production through the expression of steroidogenic enzymes and cAMP concentration in Leydig cells.

    PubMed

    Sun, Jianxia; Bai, Shun; Bai, Weibin; Zou, Feiyan; Zhang, Lei; Li, Guoqiang; Hu, Yunfeng; Li, Mingwei; Yan, Rian; Su, Zhijian; Huang, Yadong

    2014-07-01

    1,3-Dichloro-2-propanol (1,3-DCP) is a well-known food processing contaminant that has been shown to impede male reproductive function. However, its mechanism of action remains elusive. In this study, the effects of 1,3-DCP on progesterone production were investigated using the R2C Leydig cell model. 1,3-DCP significantly reduced cell viability from 7.48% to 97.4% at doses comprised between 0.5 and 6mM. Single cell gel/comet assays and atomic force microscopy assays showed that 1,3-DCP induced early phase cell apoptosis. In addition, 1,3-DCP significantly reduced progesterone production detected by radioimmunoassay (RIA). The results from quantitative polymerase chain reaction and western blotting demonstrated that the mRNA expression levels of steroidogenic acute regulatory protein (StAR), cytochrome P450 side-chain cleavage enzyme and 3β-hydroxysteroid dehydrogenase were significantly down-regulated in R2C cells. Particularly, the change rhythm of Star expression was highly consistent with progesterone production. Furthermore, the cyclic adenosine monophosphate (cAMP) and the mitochondrial membrane potential mediated by ROS, which are involved in regulating progesterone synthesis were also decreased in response to the 1,3-DCP treatment. Overall, the data presented here suggested that 1,3-DCP interferes with the male steroidogenic capacity mainly by down-regulating the level of cAMP and the key enzymes involved in the androgen synthesis pathway. PMID:24518350

  1. Spilanthol from Acmella Oleracea Lowers the Intracellular Levels of cAMP Impairing NKCC2 Phosphorylation and Water Channel AQP2 Membrane Expression in Mouse Kidney

    PubMed Central

    Gerbino, Andrea; Schena, Giorgia; Milano, Serena; Milella, Luigi; Barbosa, Alan Franco; Armentano, Francesca; Procino, Giuseppe; Svelto, Maria; Carmosino, Monica

    2016-01-01

    Acmella oleracea is well recognized in Brazilian traditional medicine as diuretic, although few scientific data have been published to support this effect. Aim of this study was to determine the molecular effect of Acmella oleracea extract and its main alkylamide spilanthol on two major processes involved in the urine concentrating mechanism: Na-K-2Cl symporter (NKCC2) activity in the thick ascending limb and water channel aquaporin 2 accumulation at the apical plasma membrane of collecting duct cells. Phosphorylation of NKCC2 was evaluated as index of its activation by Western blotting. Rate of aquaporin 2 apical expression was analyzed by confocal laser microscopy. Spilanthol-induced intracellular signalling events were dissected by video-imaging experiments. Exposure to spilanthol reduced the basal phosphorylation level of NKCC2 both in freshly isolated mouse kidney slices and in NKCC2-expresing HEK293 cells. In addition, exposure to spilanthol strongly reduced both desmopressin and low Cl−-dependent increase in NKCC2 phosphorylation in mouse kidney slices and NKCC2-expressing HEK293 cells, respectively. Similarly, spilanthol reduced both desmopressin- and forskolin-stimulated aquaporin 2 accumulation at the apical plasma membrane of collecting duct in mouse kidney slice and MCD4 cells, respectively. Of note, when orally administered, spilanthol induced a significant increase in both urine output and salt urinary excretion associated with a markedly reduced urine osmolality compared with control mice. Finally, at cellular level, spilanthol rapidly reduced or reversed basal and agonist-increased cAMP levels through a mechanism involving increases in intracellular [Ca2+]. In conclusion, spilanthol-induced inhibition of cAMP production negatively modulates urine-concentrating mechanisms thus holding great promise for its use as diuretic. PMID:27213818

  2. Spilanthol from Acmella Oleracea Lowers the Intracellular Levels of cAMP Impairing NKCC2 Phosphorylation and Water Channel AQP2 Membrane Expression in Mouse Kidney.

    PubMed

    Gerbino, Andrea; Schena, Giorgia; Milano, Serena; Milella, Luigi; Barbosa, Alan Franco; Armentano, Francesca; Procino, Giuseppe; Svelto, Maria; Carmosino, Monica

    2016-01-01

    Acmella oleracea is well recognized in Brazilian traditional medicine as diuretic, although few scientific data have been published to support this effect. Aim of this study was to determine the molecular effect of Acmella oleracea extract and its main alkylamide spilanthol on two major processes involved in the urine concentrating mechanism: Na-K-2Cl symporter (NKCC2) activity in the thick ascending limb and water channel aquaporin 2 accumulation at the apical plasma membrane of collecting duct cells. Phosphorylation of NKCC2 was evaluated as index of its activation by Western blotting. Rate of aquaporin 2 apical expression was analyzed by confocal laser microscopy. Spilanthol-induced intracellular signalling events were dissected by video-imaging experiments. Exposure to spilanthol reduced the basal phosphorylation level of NKCC2 both in freshly isolated mouse kidney slices and in NKCC2-expresing HEK293 cells. In addition, exposure to spilanthol strongly reduced both desmopressin and low Cl--dependent increase in NKCC2 phosphorylation in mouse kidney slices and NKCC2-expressing HEK293 cells, respectively. Similarly, spilanthol reduced both desmopressin- and forskolin-stimulated aquaporin 2 accumulation at the apical plasma membrane of collecting duct in mouse kidney slice and MCD4 cells, respectively. Of note, when orally administered, spilanthol induced a significant increase in both urine output and salt urinary excretion associated with a markedly reduced urine osmolality compared with control mice. Finally, at cellular level, spilanthol rapidly reduced or reversed basal and agonist-increased cAMP levels through a mechanism involving increases in intracellular [Ca2+]. In conclusion, spilanthol-induced inhibition of cAMP production negatively modulates urine-concentrating mechanisms thus holding great promise for its use as diuretic. PMID:27213818

  3. Active site coupling in PDE:PKA complexes promotes resetting of mammalian cAMP signaling.

    PubMed

    Krishnamurthy, Srinath; Moorthy, Balakrishnan Shenbaga; Xin Xiang, Lim; Xin Shan, Lim; Bharatham, Kavitha; Tulsian, Nikhil Kumar; Mihalek, Ivana; Anand, Ganesh S

    2014-09-16

    Cyclic 3'5' adenosine monophosphate (cAMP)-dependent-protein kinase (PKA) signaling is a fundamental regulatory pathway for mediating cellular responses to hormonal stimuli. The pathway is activated by high-affinity association of cAMP with the regulatory subunit of PKA and signal termination is achieved upon cAMP dissociation from PKA. Although steps in the activation phase are well understood, little is known on how signal termination/resetting occurs. Due to the high affinity of cAMP to PKA (KD ∼ low nM), bound cAMP does not readily dissociate from PKA, thus begging the question of how tightly bound cAMP is released from PKA to reset its signaling state to respond to subsequent stimuli. It has been recently shown that phosphodiesterases (PDEs) can catalyze dissociation of bound cAMP and thereby play an active role in cAMP signal desensitization/termination. This is achieved through direct interactions with the regulatory subunit of PKA, thereby facilitating cAMP dissociation and hydrolysis. In this study, we have mapped direct interactions between a specific cyclic nucleotide phosphodiesterase (PDE8A) and a PKA regulatory subunit (RIα isoform) in mammalian cAMP signaling, by a combination of amide hydrogen/deuterium exchange mass spectrometry, peptide array, and computational docking. The interaction interface of the PDE8A:RIα complex, probed by peptide array and hydrogen/deuterium exchange mass spectrometry, brings together regions spanning the phosphodiesterase active site and cAMP-binding sites of RIα. Computational docking combined with amide hydrogen/deuterium exchange mass spectrometry provided a model for parallel dissociation of bound cAMP from the two tandem cAMP-binding domains of RIα. Active site coupling suggests a role for substrate channeling in the PDE-dependent dissociation and hydrolysis of cAMP bound to PKA. This is the first instance, to our knowledge, of PDEs directly interacting with a cAMP-receptor protein in a mammalian system, and

  4. Compartmentation of cAMP signalling in cardiomyocytes in health and disease.

    PubMed

    Perera, R K; Nikolaev, V O

    2013-04-01

    3',5'-cyclic adenosine monophosphate (cAMP) is a ubiquitous second messenger critically involved in the regulation of heart function. It has been shown to act in discrete subcellular signalling compartments formed by differentially localized receptors, phosphodiesterases and protein kinases. Cardiac diseases such as hypertrophy or heart failure are associated with structural and functional remodelling of these microdomains which leads to changes in cAMP compartmentation. In this review, we will discuss recent key findings which provided new insights into cAMP compartmentation in cardiomyocytes with a particular focus on its alterations in heart disease. PMID:23383621

  5. An Essential Role of cAMP Response Element Binding Protein in Ginsenoside Rg1-Mediated Inhibition of Na+/Glucose Cotransporter 1 Gene Expression.

    PubMed

    Wang, Chun-Wen; Su, Shih-Chieh; Huang, Shu-Fen; Huang, Yu-Chuan; Chan, Fang-Na; Kuo, Yu-Han; Hung, Mei-Whey; Lin, Hang-Chin; Chang, Wen-Liang; Chang, Tsu-Chung

    2015-12-01

    The Na(+)/glucose cotransporter 1 (SGLT1) is responsible for glucose uptake in intestinal epithelial cells. It has been shown that the intestinal SGLT1 level is significantly increased in diabetic individuals and positively correlated with the pathogenesis of diabetes. The development of targeted therapeutics that can reduce the intestinal SGLT1 expression level is, therefore, important. In this study, we showed that ginsenoside Rg1 effectively decreased intestinal glucose uptake through inhibition of SGLT1 gene expression in vivo and in vitro. Transient transfection analysis of the SGLT1 promoter revealed an essential cAMP response element (CRE) that confers the Rg1-mediated inhibition of SGLT1 gene expression. Chromatin immunoprecipitation assay and targeted CRE-binding protein (CREB) silencing demonstrated that Rg1 reduced the promoter binding of CREB and CREB binding protein associated with an inactivated chromatin status. In addition, further studies showed that the epidermal growth factor receptor (EGFR) signaling pathway also plays an essential role in the inhibitory effect of Rg1; taken together, our study demonstrates the involvement of the EGFR-CREB signaling pathway in the Rg1-mediated downregulation of SGLT1 expression, which offers a potential strategy in the development of antihyperglycemic and antidiabetic treatments. PMID:26429938

  6. A Summer Camp Assessment.

    ERIC Educational Resources Information Center

    Pratte, Janice L.; DiNardi, Salvatore R.

    1979-01-01

    Reported are the results of a project assessing the impact of a revised Massachusetts sanitary code on 500 summer camps for children. The study compared camp compliances with the proposed regulations to the level of compliance with existing regulations. (BT)

  7. Michelle: Growing Through Camping

    ERIC Educational Resources Information Center

    Ouellet, Annette M.

    1977-01-01

    A mother, whose physically handicapped 7-year-old daughter has prosthetic feet, describes how her child adjusted well first to a summer day camp and then to a week long camping program run by the Girl Scouts. (GW)

  8. Inhibitors of phosphodiesterases PDE2, PDE3, and PDE4 do not increase the sinoatrial tachycardia of noradrenaline and prostaglandin PGE₁ in mice.

    PubMed

    Galindo-Tovar, Alejandro; Vargas, María Luisa; Kaumann, Alberto J

    2016-02-01

    Phosphodiesterases PDE2, PDE3, and PDE4 are expressed in murine sinoatrial cells. PDE3 and/or PDE4 reduce heart rate but apparently do not influence the tachycardia mediated through sinoatrial β1- and β2-adrenoceptors despite the high content of sinoatrial cAMP. The function of PDE2 is, however, uncertain. Prostaglandin PGE1 elicits sinoatrial tachycardia through EP receptors, but the control by phosphodiesterases is unknown. We investigated on spontaneously beating right atria of mice the effects of the PDE2 inhibitors Bay 60-7550 and EHNA on basal beating and the tachycardia produced by noradrenaline (3 nM) and PGE1 (1 μM). Bay 60-7550 (1 μM), but not EHNA (10 μM), increased basal sinoatrial beating. EHNA also failed to produce tachycardia in the presence of the adenosine deaminase inhibitor 2'-deoxycoformycin (10 μM), remaining inconclusive whether PDE2 reduces basal sinoatrial beating. Rolipram (10 μM) and cilostamide (300 nM) caused moderate tachycardia. The tachycardia evoked by Bay 60-7550 was similar in the absence and presence of rolipram. Noradrenaline elicited stable tachycardia that was not increased by Bay 60-7550. A stable tachycardia caused by PGE1 was not increased by the inhibitors of PDE2, PDE3, and PDE4. Unlike PDE3 and PDE4 which reduce murine basal sinoatrial beating, a possible effect of PDE2 needs further research. The stable tachycardia produced by noradrenaline and PGE1, together with the lack potentiation by the inhibitors of PDE2, PDE3, and PDE4, suggests that cAMP generated at the receptor compartments is hardly hydrolyzed by these phophodiesterases. Evidence from human volunteers is consistent with this proposal. PMID:26531832

  9. Role of phosphodiesterase 5 in synaptic plasticity and memory

    PubMed Central

    Puzzo, Daniela; Sapienza, Salvatore; Arancio, Ottavio; Palmeri, Agostino

    2008-01-01

    Phosphodiesterases (PDEs) are enzymes that break down the phosphodiesteric bond of the cyclic nucleotides, cAMP and cGMP, second messengers that regulate many biological processes. PDEs participate in the regulation of signal transduction by means of a fine regulation of cyclic nucleotides so that the response to cell stimuli is both specific and activates the correct third messengers. Several PDE inhibitors have been developed and used as therapeutic agents because they increase cyclic nucleotide levels by blocking the PDE function. In particular, sildenafil, an inhibitor of PDE5, has been mainly used in the treatment of erectile dysfunction but is now also utilized against pulmonary hypertension. This review examines the physiological role of PDE5 in synaptic plasticity and memory and the use of PDE5 inhibitors as possible therapeutic agents against disorders of the central nervous system (CNS). PMID:18728748

  10. Camp is a Celebration.

    ERIC Educational Resources Information Center

    North Dakota Farmers Union, Jamestown. Dept. of Youth Activities.

    A camping workbook provides materials for use in discussion of 3 important aspects of farm living: rural power, communication, and conservation. It is intended that this material be used in class sessions held during a junior youth camp. A sample of the youth camp schedule reveals 3 forty-five minute time periods during the day designated as class…

  11. Cadmium-induced decrement of the LH receptor expression and cAMP levels in the testis of rats.

    PubMed

    Gunnarsson, David; Nordberg, Gunnar; Lundgren, Per; Selstam, Gunnar

    2003-02-01

    Cadmium (Cd) is a widespread environmental pollutant, characterized by its ability to affect various organs. Adverse effect of Cd on the testis including decreased testosterone production are well-known phenomena, but the cellular events explaining these effects have not yet been established. In the present study the initial steps of gonadotropin mediated testosterone biosynthesis were examined in vivo in rats, in relation to Cd dose and time after injection. In the dose-response experiment Male Sprague-Dawley rats received a single subcutaneous (s.c.) injection of CdCl(2) (1, 5 or 10 micromol/kg body weight) and were sacrificed 48 h after injection. A statistically significant decrease in luteinizing hormone (LH) receptor mRNA level in the testicular tissue was demonstrated at the highest dose (10 micromol/kg). In the temporal-response experiment rats were given 10 micromol/kg of CdCl(2) s.c. and sacrificed 0.48, 4.8, 48 or 144 h after injection. LH receptor mRNA levels as well as cyclic adenosine monophosphate (cAMP) levels were found to be significantly lowered at 48 and 144 h. These observations of the mechanisms whereby Cd exerts its effect on the initial steps of testosterone biosynthesis are the first from in vivo experiments. PMID:12504342

  12. Stimulation of StAR expression by cAMP is controlled by inhibition of highly inducible SIK1 via CRTC2, a co-activator of CREB.

    PubMed

    Lee, Jinwoo; Tong, Tiegang; Takemori, Hiroshi; Jefcoate, Colin

    2015-06-15

    In mouse steroidogenic cells the activation of cholesterol metabolism is mediated by steroidogenic acute regulatory protein (StAR). Here, we visualized a coordinated regulation of StAR transcription, splicing and post-transcriptional processing, which are synchronized by salt inducible kinase (SIK1) and CREB-regulated transcription coactivator (CRTC2). To detect primary RNA (pRNA), spliced primary RNA (Sp-RNA) and mRNA in single cells, we generated probe sets by using fluorescence in situ hybridization (FISH). These methods allowed us to address the nature of StAR gene expression and to visualize protein-nucleic acid interactions through direct detection. We show that SIK1 represses StAR expression in Y1 adrenal and MA10 testis cells through inhibition of processing mediated by CRTC2. Digital image analysis matches qPCR analyses of the total cell culture. Evidence is presented for spatially separate accumulation of StAR pRNA and Sp-RNA at the gene loci in the nucleus. These findings establish that cAMP, SIK and CRTC mediate StAR expression through activation of individual StAR gene loci. PMID:25662274

  13. Maternal exposure to secondhand cigarette smoke primes the lung for induction of phosphodiesterase-4D5 isozyme and exacerbated Th2 responses: rolipram attenuates the airway hyperreactivity and muscarinic receptor expression but not lung inflammation and atopy.

    PubMed

    Singh, Shashi P; Mishra, Neerad C; Rir-Sima-Ah, Jules; Campen, Mathew; Kurup, Viswanath; Razani-Boroujerdi, Seddigheh; Sopori, Mohan L

    2009-08-01

    Airway hyperreactivity (AHR), lung inflammation, and atopy are clinical signs of allergic asthma. Gestational exposure to cigarette smoke (CS) markedly increases the risk for childhood allergic asthma. Muscarinic receptors regulate airway smooth muscle tone, and asthmatics exhibit increased AHR to muscarinic agonists. We have previously reported that in a murine model of bronchopulmonary aspergillosis, maternal exposure to mainstream CS increases AHR after acute intratracheal administration of Aspergillus fumigatus extract. However, the mechanism by which gestational CS induces allergic asthma is unclear. We now show for the first time that, compared with controls, mice exposed prenatally to secondhand CS exhibit increased lung inflammation (predominant infiltration by eosinophils and polymorphs), atopy, and airway resistance, and produce proinflammatory cytokines (IL-4, IL-5, IL-6, and IL-13, but not IL-2 or IFN-gamma). These changes, which occur only after an allergen (A. fumigatus extract) treatment, are correlated with marked up-regulated lung expression of M1, M2, and M3 muscarinic receptors and phosphodiesterase (PDE)4D5 isozyme. Interestingly, the PDE4-selective inhibitor rolipram attenuates the increase in AHR, muscarinic receptors, and PDE4D5, but fails to down-regulate lung inflammation, Th2 cytokines, or serum IgE levels. Thus, the fetus is extraordinarily sensitive to CS, inducing allergic asthma after postnatal exposure to allergens. Although the increased AHR might reflect increased PDE4D5 and muscarinic receptor expression, the mechanisms underlying atopy and lung inflammation are unrelated to the PDE4 activity. Thus, PDE4 inhibitors might ease AHR, but are unlikely to attenuate lung inflammation and atopy associated with childhood allergic asthma. PMID:19596983

  14. Phosphodiesterase inhibitor-dependent inverse agonism of agouti-related protein on melanocortin 4 receptor in sea bass (Dicentrarchus labrax)

    PubMed Central

    Sánchez, Elisa; Rubio, Vera Cruz; Thompson, Darren; Metz, Juriaan; Flik, Gert; Millhauser, Glenn L.; Cerdá-Reverter, José Miguel

    2009-01-01

    The melanocortin 4 receptor (MC4R) is a G protein-coupled receptor mainly expressed in the central nervous system of vertebrates. Activation of the MC4R leads to a decrease in food intake, whereas inactivating mutations are a genetic cause of obesity. The binding of agouti-related protein (AGRP) reduces not only agonist-stimulated cAMP production (competitive antagonist) but also the basal activity of the receptor, as an inverse agonist. Transgenic zebrafish overexpressing AGRP display increased food intake and linear growth, indicative of a physiological role for the melanocortin system in the control of the energy balance in fish. We report on the cloning, pharmacological characterization, tissue distribution, and detailed brain mapping of a sea bass (Dicentrarchus labrax) MC4R ortholog. Sea bass MC4R is profusely expressed within food intake-controlling pathways of the fish brain. However, the activity of the melanocortin system during progressive fasting does not depend on the hypothalamic/pituitary proopiomelanocortin (POMC) and MC4R expression, which suggests that sea bass MC4R is constitutively activated and regulated by AGRP binding. We demonstrate that AGRP acts as competitive antagonist and reduces MTII-induced cAMP production. AGRP also decreases the basal activity of the receptor as an inverse agonist. This observation suggests that MC4R is constitutively active and supports the evolutionary conservation of the AGRP/MC4R interactions. The inverse agonism, but not the competitive antagonism, depends on the presence of a phosphodiesterase inhibitor (IBMX). This suggests that inverse agonism and competitive antagonism operate through different intracellular signaling pathways, a view that opens up new targets for the treatment of melanocortin-induced metabolic syndrome. PMID:19225141

  15. Cyclic nucleotide phosphodiesterase-1C (PDE1C) drives cell proliferation, migration and invasion in glioblastoma multiforme cells in vitro.

    PubMed

    Rowther, Farjana B; Wei, Weinbin; Dawson, Timothy P; Ashton, Katherine; Singh, Anushree; Madiesse-Timchou, Mylene P; Thomas, D G T; Darling, John L; Warr, Tracy

    2016-03-01

    Cyclic nucleotides (cAMP & cGMP) are critical intracellular second messengers involved in the transduction of a diverse array of stimuli and their catabolism is mediated by phosphodiesterases (PDEs). We previously detected focal genomic amplification of PDE1C in >90 glioblastoma multiforme (GBM) cells suggesting a potential as a novel therapeutic target in these cells. In this report, we show that genomic gain of PDE1C was associated with increased expression in low passage GBM-derived cell cultures. We demonstrate that PDE1C is essential in driving cell proliferation, migration and invasion in GBM cultures since silencing of this gene significantly mitigates these functions. We also define the mechanistic basis of this functional effect through whole genome expression analysis by identifying down-stream gene effectors of PDE1C which are involved in cell cycle and cell adhesion regulation. In addition, we also demonstrate that Vinpocetine, a general PDE1 inhibitor, can also attenuate proliferation with no effect on invasion/migration. Up-regulation of at least one of this gene set (IL8, CXCL2, FOSB, NFE2L3, SUB1, SORBS2, WNT5A, and MMP1) in TCGA GBM cohorts is associated with worse outcome and PDE1C silencing down-regulated their expression, thus also indicating potential to influence patient survival. Therefore we conclude that proliferation, migration, and invasion of GBM cells could also be regulated downstream of PDE1C. PMID:25620587

  16. Creating healthy camp experiences.

    PubMed

    Walton, Edward A; Tothy, Alison S

    2011-04-01

    The American Academy of Pediatrics has created recommendations for health appraisal and preparation of young people before participation in day or resident camps and to guide health and safety practices for children at camp. These recommendations are intended for parents, primary health care providers, and camp administration and health center staff. Although camps have diverse environments, there are general guidelines that apply to all situations and specific recommendations that are appropriate under special conditions. This policy statement has been reviewed and is supported by the American Camp Association. PMID:21444589

  17. Efficacy of B-Type Natriuretic Peptide Is Coupled to Phosphodiesterase 2A in Cardiac Sympathetic Neurons

    PubMed Central

    Li, Dan; Lu, Chieh-Ju; Hao, Guoliang; Wright, Hannah; Woodward, Lavinia; Liu, Kun; Vergari, Elisa; Surdo, Nicoletta C.; Herring, Neil; Zaccolo, Manuela; Paterson, David J.

    2015-01-01

    Elevated B-type natriuretic peptide (BNP) regulates cGMP-phosphodiesterase activity. Its elevation is regarded as an early compensatory response to cardiac failure where it can facilitate sympathovagal balance and cardiorenal homeostasis. However, recent reports suggest a paradoxical proadrenergic action of BNP. Because phosphodiesterase activity is altered in cardiovascular disease, we tested the hypothesis that BNP might lose its efficacy by minimizing the action of cGMP on downstream pathways coupled to neurotransmission. BNP decreased norepinephrine release from atrial preparations in response to field stimulation and also significantly reduced the heart rate responses to sympathetic nerve stimulation in vitro. Using electrophysiological recording and fluorescence imaging, BNP also reduced the depolarization evoked calcium current and intracellular calcium transient in isolated cardiac sympathetic neurons. Pharmacological manipulations suggested that the reduction in the calcium transient was regulated by a cGMP/protein kinase G pathway. Fluorescence resonance energy transfer measurements for cAMP, and an immunoassay for cGMP, showed that BNP increased cGMP, but not cAMP. In addition, overexpression of phosphodiesterase 2A after adenoviral gene transfer markedly decreased BNP stimulation of cGMP and abrogated the BNP responses to the calcium current, intracellular calcium transient, and neurotransmitter release. These effects were reversed on inhibition of phosphodiesterase 2A. Moreover, phosphodiesterase 2A activity was significantly elevated in stellate neurons from the prohypertensive rat compared with the normotensive control. Our data suggest that abnormally high levels of phosphodiesterase 2A may provide a brake against the inhibitory action of BNP on sympathetic transmission. PMID:25916722

  18. Effect of some polysaccharides in cyclic nucleotide level and phosphodiesterase activity in organs of mice with lewis' lung carcinoma

    SciTech Connect

    Veksler, I.G.; Antonenko, S.G.

    1986-04-01

    This paper describes the results of a study of the effect of the bacterial polysaccharide prodigiosan and the yeast cell membrane bipolymer zymosan on the absolute and relative levels of cAMP, cAMP, and cAMP-dependent phosphodiesterase (PDE) in the thymus, spleen, and lungs of healthy mice and of mice with metastasizing Lewis' lung carcinoma. The cyclic nucleotide concentration was determined by radioimmunoassay. Radioactivity of the samples was studied in an SL-30 liquid scintillation counter. PDE activity was determined by paper chromatography using 8-/sup 3/H-cAMP as the substrate.

  19. Regulation of cyclooxygenase-2 expression by cAMP response element and mRNA stability in a human airway epithelial cell line exposed to zinc

    SciTech Connect

    Wu Weidong Silbajoris, Robert A.; Cao Dongsun; Bromberg, Philip A.; Zhang Qiao; Peden, David B.; Samet, James M.

    2008-09-01

    Exposure to zinc-laden particulate matter in ambient and occupational settings has been associated with proinflammatory responses in the lung. Cyclooxygenase 2-derived eicosanoids are important modulators of airway inflammation. In this study, we characterized the transcriptional and posttranscriptional events that regulate COX-2 expression in a human bronchial epithelial cell line BEAS-2B exposed to Zn{sup 2+}. Zn{sup 2+} exposure resulted in pronounced increases in COX-2 mRNA and protein expression, which were prevented by pretreatment with the transcription inhibitor actinomycin D, implying the involvement of transcriptional regulation. This was supported by the observation of increased COX-2 promoter activity in Zn{sup 2+}-treated BEAS-2B cells. Mutation of the cAMP response element (CRE), but not the {kappa}B-binding sites in the COX-2 promoter markedly reduced COX-2 promoter activity induced by Zn{sup 2+}. Inhibition of NF{kappa}B activation did not block Zn{sup 2+}-induced COX-2 expression. Measurement of mRNA stability demonstrated that Zn{sup 2+} exposure impaired the degradation of COX-2 mRNA in BEAS-2B cells. This message stabilization effect of Zn{sup 2+} exposure was shown to be dependent on the integrity of the 3'-untranslated region found in the COX-2 transcript. Taken together, these data demonstrate that the CRE and mRNA stability regulates COX-2 expression induced in BEAS-2B cells exposed to extracellular Zn{sup 2+}.

  20. Inhibition of type 4 cyclic nucleotide phosphodiesterase blocks intracellular TLR signaling in chronic lymphocytic leukemia and normal hematopoietic cells.

    PubMed

    Tan, Ying; Watkins, Amanda A; Freeman, Benjamin B; Meyers, John A; Rifkin, Ian R; Lerner, Adam

    2015-01-01

    A subset of chronic lymphocytic leukemia (CLL) BCRs interacts with Ags expressed on apoptotic cells, suggesting that CLL BCRs have the potential to internalize apoptotic cell RNA- or DNA-containing fragments with resultant activation of TLR7 or TLR9, respectively. By blocking cAMP degradation, type 4 cAMP phosphodiesterase (PDE4) inhibitors activate cAMP-mediated signaling and induce apoptosis in CLL cells. In this study, we show that autologous irradiated leukemic cells induce proliferation in CLL cells and that such proliferation is blocked by a TLR7/8/9 inhibitor, by DNase, and by the PDE4 inhibitor rolipram. Rolipram also inhibited CLL cell proliferation induced by synthetic TLR7 and TLR9 agonists, as well as TLR agonist-induced costimulatory molecule expression and TNF-a (but not IL-6 or IL-10) production. Whereas treatment with a TLR9 agonist protected IgH V region unmutated, but not mutated, CLL cells from apoptosis, PDE4 inhibitors augmented apoptosis in both subtypes, suggesting that cAMP-mediated signaling may abrogate a TLR9-mediated survival signal in prognostically unfavorable IGHV unmutated CLL cells. Rolipram inhibited both TLR7/8- and TLR9-induced IFN regulatory factor 5 and NF-kB p65 nuclear translocation. PDE4 inhibitors also blocked TLR signaling in normal human immune cells. In PBMC and CD14-positive monocytes, PDE4 inhibitors blocked IFN-a or TNF-a (but not IL-6) production, respectively, following stimulation with synthetic TLR agonists or RNA-containing immune complexes. These results suggest that PDE4 inhibitors may be of clinical utility in CLL or autoimmune diseases that are driven by TLR-mediated signaling. PMID:25416804

  1. Phosphodiesterase-2 inhibitor reverses corticosterone-induced neurotoxicity and related behavioural changes via cGMP/PKG dependent pathway.

    PubMed

    Xu, Ying; Pan, Jianchun; Chen, Ling; Zhang, Chong; Sun, Jiao; Li, Jianxin; Nguyen, Linda; Nair, Neetu; Zhang, Hanting; O'Donnell, James M

    2013-05-01

    Phosphodiesterase 2 (PDE2) is an enzyme responsible for hydrolysis of cyclic adenosine monophosphate (cAMP) and cyclic guanosine monophosphate (cGMP) to restrict intracellular signalling of these second messenger molecules. This study investigated how PDE2 inhibitor Bay 60-7550 affects the dysregulated glucocorticoid signalling in neuronal cells and regulates depressive behaviours after chronic stress in mice. We found that exposure of hippocampal neurons to corticosterone resulted in time- and concentration-dependent increases in PDE2 expression. These intriguing findings were confirmed in the hippocampal cell line HT-22. After corticosterone exposure for 24 h, HT-22 cells showed a concentration-dependent increase in mRNA levels for PDE2 subtypes, PDE2A1 and 2A3, as well as for the total PDE2A protein expression. Bay 60-7550 was found to reverse the cell lesion induced by corticosterone (50 μm). This neuroprotective effect was blocked by pretreatment with protein kinase G inhibitor KT5823, but not protein kinase A inhibitor H89, suggesting the involvement of cGMP-dependent signalling. Although Bay 60-7550 treatment for 24 h did not change the levels of phosphorylated mitogen-activated protein kinases ERK1/2 (pERK) and phosphorylated cAMP response element-binding protein (pCREB), it down-regulated pERK at 2 h and up-regulated a CREB co-activator, CREB-binding protein, at 24 h. Both of these effects were blocked by KT 5823. Furthermore, Bay 60-7550 reversed corticosterone-induced down-regulation of brain-derived neurotrophic factor protein levels 24 h after corticosterone exposure. In behavioural testing, Bay 60-7550 produced antidepressant-like effects and reduced corticosterone levels in stressed mice, further supporting the involvement of a PDE2-dependent pathway in mediating Bay 60-7550's effect during stress hormone insults. PMID:22850435

  2. Isolation and characterization of human cDNAs encoding a cGMP-stimulated 3',5'-cyclic nucleotide phosphodiesterase.

    PubMed

    Rosman, G J; Martins, T J; Sonnenburg, W K; Beavo, J A; Ferguson, K; Loughney, K

    1997-05-20

    Human cyclic GMP-stimulated 3',5'-cyclic nucleotide phosphodiesterase (PDE2A3) cDNAs were cloned from hippocampus and fetal brain cDNA libraries. A 4.2-kb composite DNA sequence constructed from overlapping cDNA clones encodes a 941 amino acid protein with a predicted molecular mass of 105,715 Da. Extracts prepared from yeast expressing the human PDE2A3 hydrolyzed both cyclic AMP (cAMP) and cyclic GMP (cGMP). This activity was inhibited by EHNA, a selective PDE2 inhibitor, and was stimulated three-fold by cGMP. Human PDE2A is expressed in brain and to a lesser extent in heart, placenta, lung, skeletal muscle, kidney and pancreas. The human PDE2A3 differs from the bovine PDE2A1 and rat PDE2A2 proteins at the amino terminus but its amino-terminal sequence is identical to the bovine PDE2A3 sequence. The different amino termini probably arise from alternative exon splicing of the PDE2A mRNA. PMID:9210593

  3. Phosphodiesterase Inhibition to Target the Synaptic Dysfunction in Alzheimer's Disease

    NASA Astrophysics Data System (ADS)

    Bales, Kelly R.; Plath, Niels; Svenstrup, Niels; Menniti, Frank S.

    Alzheimer's Disease (AD) is a disease of synaptic dysfunction that ultimately proceeds to neuronal death. There is a wealth of evidence that indicates the final common mediator of this neurotoxic process is the formation and actions on synaptotoxic b-amyloid (Aβ). The premise in this review is that synaptic dysfunction may also be an initiating factor in for AD and promote synaptotoxic Aβ formation. This latter hypothesis is consistent with the fact that the most common risk factors for AD, apolipoprotein E (ApoE) allele status, age, education, and fitness, encompass suboptimal synaptic function. Thus, the synaptic dysfunction in AD may be both cause and effect, and remediating synaptic dysfunction in AD may have acute effects on the symptoms present at the initiation of therapy and also slow disease progression. The cyclic nucleotide (cAMP and cGMP) signaling systems are intimately involved in the regulation of synaptic homeostasis. The phosphodiesterases (PDEs) are a superfamily of enzymes that critically regulate spatial and temporal aspects of cyclic nucleotide signaling through metabolic inactivation of cAMP and cGMP. Thus, targeting the PDEs to promote improved synaptic function, or 'synaptic resilience', may be an effective and facile approach to new symptomatic and disease modifying therapies for AD. There continues to be a significant drug discovery effort aimed at discovering PDE inhibitors to treat a variety of neuropsychiatric disorders. Here we review the current status of those efforts as they relate to potential new therapies for AD.

  4. Activation of wild-type and deltaF508-CFTR by phosphodiesterase inhibitors through cAMP-dependent and -independent mechanisms.

    PubMed

    Al-Nakkash, L; Hwang, T C

    1999-03-01

    The cAMP-dependent activation of the cystic fibrosis transmembrane conductance regulator (CFTR) and its modulation through inhibition of phosphodiesterases (PDE) were studied with the cell-attached patch-clamp technique in Calu-3 cells (expressing endogenous CFTR) and NIH3T3 cells [expressing either wild-type (Wt)-CFTR or DeltaF508-CFTR]. In Calu-3 cells, CFTR current was augmented by increasing concentrations of 8-(4-chlorophenylthio)-adenosine 3', 5'-cyclic monophosphate (CPT-cAMP) and reached a saturating level at >/=60 microM. Varying the forskolin concentration also modulated CFTR activity; 10 microM was maximally effective since supplemental application of 200 microM CPT-cAMP had no additional effect. Activation of CFTR by increasing the cAMP concentration occurs through an increase of the NPo (product of the number of functional channels and the open probability) since the single-channel amplitude remains unchanged. In Calu-3 and NIH3T3-Wt cells, PDE inhibitors, milrinone (100 microM), 8-cyclopentyl-1, 3-dipropylxanthine (CPX, 25 microM), and 3-isobutyl-1-methylxanthine (IBMX, 200 microM), did not enhance CFTR current initially activated with 10 microM forskolin, but each potentiated CFTR activity elicited with a submaximal forskolin concentration (e.g., 100 nM) and prolonged the deactivation of CFTR channel current upon removal of forskolin. Millimolar IBMX increased the NPo of both Wt- and DeltaF508-CFTR even under maximal cAMP stimulation. Quantitatively, these effects of millimolar IBMX on NPo approximate those of genistein, which potentiates the cAMP-dependent CFTR activity via a mechanism that does not involve increases in cellular cAMP. Thus, depending on the concentration, PDE inhibitors may affect CFTR through different mechanisms. PMID:10089568

  5. Pharmacology of phosphodiesterase-5 inhibitors.

    PubMed

    Corbin, J D; Francis, S H

    2002-01-01

    The clinical properties (efficacy and safety profile) of a medicine are related not only to its mode of action, but also to its selectivity for its target (usually a receptor or enzyme) and are also influenced by its pharmacokinetic properties (absorption, distribution, metabolism and elimination). The growing number of phosphodiesterase inhibitors that are selective for phosphodiesterase-5 (PDE5) represent a promising new class of compounds that are useful for the treatment of erectile dysfunction and perhaps other disorders. Some of the basic pharmacodynamic and pharmacokinetic parameters that describe drug action are discussed with regard to the new PDE5 inhibitors. Central topics reviewed are the concentration that produces a given in vitro response, or potency (IC50), maximum plasma concentration (Cmax), time to Cmax (Tmax), half-life (t 1/2), area under the curve (AUC), bioavailability, onset and duration of action, and the balance to achieve optimum safety and efficacy. To illustrate these concepts, a group of inhibitors with varying selectivities and potencies for PDE5 (theophylline, IBMX, zaprinast, sildenafil, tadalafil and vardenafil) are discussed. Each drug has its own set of unique pharmacological characteristics based on its specific molecular structure, enzyme inhibition profile and pharmacokinetic properties. Each PDE5 inhibitor has a distinct selectivity that contributes to its safety profile. As with all new drugs, and especially those in a new class, careful evaluation will be necessary to ensure the optimal use of the PDE5 inhibitors. PMID:12166544

  6. Engineering of a red-light–activated human cAMP/cGMP-specific phosphodiesterase

    PubMed Central

    Gasser, Carlos; Taiber, Sandra; Yeh, Chen-Min; Wittig, Charlotte Helene; Hegemann, Peter; Ryu, Soojin; Wunder, Frank; Möglich, Andreas

    2014-01-01

    Sensory photoreceptors elicit vital physiological adaptations in response to incident light. As light-regulated actuators, photoreceptors underpin optogenetics, which denotes the noninvasive, reversible, and spatiotemporally precise perturbation by light of living cells and organisms. Of particular versatility, naturally occurring photoactivated adenylate cyclases promote the synthesis of the second messenger cAMP under blue light. Here, we have engineered a light-activated phosphodiesterase (LAPD) with complementary light sensitivity and catalytic activity by recombining the photosensor module of Deinococcus radiodurans bacterial phytochrome with the effector module of Homo sapiens phosphodiesterase 2A. Upon red-light absorption, LAPD up-regulates hydrolysis of cAMP and cGMP by up to sixfold, whereas far-red light can be used to down-regulate activity. LAPD also mediates light-activated cAMP and cGMP hydrolysis in eukaryotic cell cultures and in zebrafish embryos; crucially, the biliverdin chromophore of LAPD is available endogenously and does not need to be provided exogenously. LAPD thus establishes a new optogenetic modality that permits light control over diverse cAMP/cGMP-mediated physiological processes. Because red light penetrates tissue more deeply than light of shorter wavelengths, LAPD appears particularly attractive for studies in living organisms. PMID:24889611

  7. Auphen and dibutyryl cAMP suppress growth of hepatocellular carcinoma by regulating expression of aquaporins 3 and 9 in vivo

    PubMed Central

    Peng, Rui; Zhao, Guang-Xi; Li, Jing; Zhang, Yu; Shen, Xi-Zhong; Wang, Ji-Yao; Sun, Jian-Yong

    2016-01-01

    AIM: To investigate whether the regulation of aquaporin 3 (AQP3) and AQP9 induced by Auphen and dibutyryl cAMP (dbcAMP) inhibits hepatic tumorigenesis. METHODS: Expression of AQP3 and AQP9 was detected by Western blot, immunohistochemistry (IHC), and RT-PCR in HCC samples and paired non-cancerous liver tissue samples from 30 hepatocellular carcinoma (HCC) patients. A xenograft tumor model was used in vivo. Nine nude mice were divided into control, Auphen-treated, and dbcAMP-treated groups (n = 3 for each group). AQP3 and AQP9 protein expression after induction of xenograft tumors was detected by IHC and mRNA by RT-PCR analysis. The terminal deoxynucleotidyl transferase-mediated dUTP nick end labeling assay and histological evaluation were used to detect apoptosis of tumor cells, and the concentration of serum α-fetoprotein (AFP) was measured using RT-PCR and an ELISA kit. RESULTS: The volumes and weights of tumors decreased significantly in the Auphen- and dbcAMP-treated mice compared with the control mice (P < 0.01). The levels of AQP3 were significantly lower in the Auphen treatment group, and levels of AQP9 were significantly higher in thedbcAMP treatment mice than in the control mice (P < 0.01). The reduction of AQP3 by Auphen and increase of AQP9 by dbcAMP in nude mice suppressed tumor growth of HCC, which resulted in reduced AFP levels in serum and tissues, and apoptosis of tumor cells in the Auphen- and dbcAMP-treated mice, when compared with control mice (P < 0.01). Compared with para-carcinoma tissues, AQP3 expression increased in tumor tissues whereas the expression of AQP9 decreased. By correlating clinicopathological and expression levels, we demonstrated that the expression of AQP3 and AQP9 was correlated with clinical progression of HCC and disease outcomes. CONCLUSION: AQP3 increases in HCC while AQP9 decreases. Regulation of AQP3 and AQP9 expression by Auphen and dbcAMP inhibits the development and growth of HCC. PMID:27022216

  8. Pyrroloquinoline Quinone Stimulates Mitochondrial Biogenesis through cAMP Response Element-binding Protein Phosphorylation and Increased PGC-1α Expression*

    PubMed Central

    Chowanadisai, Winyoo; Bauerly, Kathryn A.; Tchaparian, Eskouhie; Wong, Alice; Cortopassi, Gino A.; Rucker, Robert B.

    2010-01-01

    Bioactive compounds reported to stimulate mitochondrial biogenesis are linked to many health benefits such increased longevity, improved energy utilization, and protection from reactive oxygen species. Previously studies have shown that mice and rats fed diets lacking in pyrroloquinoline quinone (PQQ) have reduced mitochondrial content. Therefore, we hypothesized that PQQ can induce mitochondrial biogenesis in mouse hepatocytes. Exposure of mouse Hepa1–6 cells to 10–30 μm PQQ for 24–48 h resulted in increased citrate synthase and cytochrome c oxidase activity, Mitotracker staining, mitochondrial DNA content, and cellular oxygen respiration. The induction of this process occurred through the activation of cAMP response element-binding protein (CREB) and peroxisome proliferator-activated receptor-γ coactivator-1α (PGC-1α), a pathway known to regulate mitochondrial biogenesis. PQQ exposure stimulated phosphorylation of CREB at serine 133, activated the promoter of PGC-1α, and increased PGC-1α mRNA and protein expression. PQQ did not stimulate mitochondrial biogenesis after small interfering RNA-mediated reduction in either PGC-1α or CREB expression. Consistent with activation of the PGC-1α pathway, PQQ increased nuclear respiratory factor activation (NRF-1 and NRF-2) and Tfam, TFB1M, and TFB2M mRNA expression. Moreover, PQQ protected cells from mitochondrial inhibition by rotenone, 3-nitropropionic acid, antimycin A, and sodium azide. The ability of PQQ to stimulate mitochondrial biogenesis accounts in part for action of this compound and suggests that PQQ may be beneficial in diseases associated with mitochondrial dysfunction. PMID:19861415

  9. PDE2-mediated cAMP hydrolysis accelerates cardiac fibroblast to myofibroblast conversion and is antagonized by exogenous activation of cGMP signaling pathways.

    PubMed

    Vettel, C; Lämmle, S; Ewens, S; Cervirgen, C; Emons, J; Ongherth, A; Dewenter, M; Lindner, D; Westermann, D; Nikolaev, V O; Lutz, S; Zimmermann, W H; El-Armouche, A

    2014-04-15

    Recent studies suggest that the signal molecules cAMP and cGMP have antifibrotic effects by negatively regulating pathways associated with fibroblast to myofibroblast (MyoCF) conversion. The phosphodiesterase 2 (PDE2) has the unique property to be stimulated by cGMP, which leads to a remarkable increase in cAMP hydrolysis and thus mediates a negative cross-talk between both pathways. PDE2 has been recently investigated in cardiomyocytes; here we specifically addressed its role in fibroblast conversion and cardiac fibrosis. PDE2 is abundantly expressed in both neonatal rat cardiac fibroblasts (CFs) and cardiomyocytes. The overexpression of PDE2 in CFs strongly reduced basal and isoprenaline-induced cAMP synthesis, and this decrease was sufficient to induce MyoCF conversion even in the absence of exogenous profibrotic stimuli. Functional stress-strain experiments with fibroblast-derived engineered connective tissue (ECT) demonstrated higher stiffness in ECTs overexpressing PDE2. In regard to cGMP, neither basal nor atrial natriuretic peptide-induced cGMP levels were affected by PDE2, whereas the response to nitric oxide donor sodium nitroprusside was slightly but significantly reduced. Interestingly, despite persistently depressed cAMP levels, both cGMP-elevating stimuli were able to completely prevent the PDE2-induced MyoCF phenotype, arguing for a double-tracked mechanism. In conclusion, PDE2 accelerates CF to MyoCF conversion, which leads to greater stiffness in ECTs. Atrial natriuretic peptide- and sodium nitroprusside-mediated cGMP synthesis completely reverses PDE2-induced fibroblast conversion. Thus PDE2 may augment cardiac remodeling, but this effect can also be overcome by enhanced cGMP. The redundant role of cAMP and cGMP as antifibrotic meditators may be viewed as a protective mechanism in heart failure. PMID:24531807

  10. Reversal of Oxidative Stress-Induced Anxiety by Inhibition of Phosphodiesterase-2 in Mice

    PubMed Central

    Masood, Anbrin; Nadeem, Ahmed; Mustafa, S. Jamal; O’Donnell, James M.

    2010-01-01

    The pathogenesis of several neuropsychiatric disorders, including anxiety and depression, has been linked to oxidative stress, in part via alterations in cyclic nucleotide signaling. Phosphodiesterase-2 (PDE2), which regulates cGMP and cAMP signaling, may affect anxiety-related behavior through reduction of oxidative stress. The present study evaluated the effects of oxidative stress on behavior and assessed the anxiolytic effects of the PDE2 inhibitor Bay 60-7550 [(2-(3,4-dimethoxybenzyl)-7-{(1R)-1-[(1R)-1-hydroxyethyl]-4-phenylbutyl}-5-methyl imidazo-[5,1-f][1,2,4]triazin-4(3H)-one)]. Treatment of mice with L-buthionine-(S,R)-sulfoximine (300 mg/kg), an inducer of oxidative stress, caused anxiety-like behavioral effects in elevated plus-maze, open-field, and hole-board tests through the NADPH oxidase pathway; these effects were antagonized by Bay 60-7550 (3 mg/kg) and apocynin (3 mg/kg), an inhibitor of NADPH oxidase. The Bay 60-7550-mediated decrease in oxidative stress (i.e., superoxide anion and reactive oxygen species generation in cultured neurons and total antioxidant capacity and lipid peroxides in amygdala and hypothalamus) and expression of NADPH oxidase subunits (i.e., p47 phox and gp91 phox expression in amygdala, hypothalamus, and cultured neurons) was associated with increased cGMP and phosphorylation of vasodilator-stimulated phosphoprotein at Ser239, suggesting an important role of cGMP-protein kinase G signaling in reduction of anxiety. Overall, the present results indicate that oxidative stress induces anxiety-like behavior in mice and that PDE2 inhibition reverses it through an increase in cGMP signaling. Thus, PDE2 may be a novel pharmacological target for treatment of anxiety in neuropsychiatric and neurodegenerative disorders that involve oxidative stress. PMID:18456873

  11. Reversal of oxidative stress-induced anxiety by inhibition of phosphodiesterase-2 in mice.

    PubMed

    Masood, Anbrin; Nadeem, Ahmed; Mustafa, S Jamal; O'Donnell, James M

    2008-08-01

    The pathogenesis of several neuropsychiatric disorders, including anxiety and depression, has been linked to oxidative stress, in part via alterations in cyclic nucleotide signaling. Phosphodiesterase-2 (PDE2), which regulates cGMP and cAMP signaling, may affect anxiety-related behavior through reduction of oxidative stress. The present study evaluated the effects of oxidative stress on behavior and assessed the anxiolytic effects of the PDE2 inhibitor Bay 60-7550 [(2-(3,4-dimethoxybenzyl)-7-{(1R)-1-[(1R)-1-hydroxyethyl]-4-phenylbutyl}-5-methyl imidazo-[5,1-f][1,2,4]triazin-4(3H)-one)]. Treatment of mice with L-buthionine-(S,R)-sulfoximine (300 mg/kg), an inducer of oxidative stress, caused anxiety-like behavioral effects in elevated plusmaze, open-field, and hole-board tests through the NADPH oxidase pathway; these effects were antagonized by Bay 60-7550 (3 mg/kg) and apocynin (3 mg/kg), an inhibitor of NADPH oxidase. The Bay 60-7550-mediated decrease in oxidative stress (i.e., superoxide anion and reactive oxygen species generation in cultured neurons and total antioxidant capacity and lipid peroxides in amygdala and hypothalamus) and expression of NADPH oxidase subunits (i.e., p47 phox and gp91 phox expression in amygdala, hypothalamus, and cultured neurons) was associated with increased cGMP and phosphorylation of vasodilator-stimulated phosphoprotein at Ser239, suggesting an important role of cGMP-protein kinase G signaling in reduction of anxiety. Overall, the present results indicate that oxidative stress induces anxiety-like behavior in mice and that PDE2 inhibition reverses it through an increase in cGMP signaling. Thus, PDE2 may be a novel pharmacological target for treatment of anxiety in neuropsychiatric and neurodegenerative disorders that involve oxidative stress. PMID:18456873

  12. Cyclic nucleotide phosphodiesterases in heart and vessels: A therapeutic perspective.

    PubMed

    Bobin, Pierre; Belacel-Ouari, Milia; Bedioune, Ibrahim; Zhang, Liang; Leroy, Jérôme; Leblais, Véronique; Fischmeister, Rodolphe; Vandecasteele, Grégoire

    2016-01-01

    Cyclic nucleotide phosphodiesterases (PDEs) degrade the second messengers cyclic adenosine monophosphate (cAMP) and cyclic guanosine monophosphate (cGMP), thereby regulating multiple aspects of cardiac and vascular muscle functions. This highly diverse class of enzymes encoded by 21 genes encompasses 11 families that are not only responsible for the termination of cyclic nucleotide signalling, but are also involved in the generation of dynamic microdomains of cAMP and cGMP, controlling specific cell functions in response to various neurohormonal stimuli. In the myocardium and vascular smooth muscle, the PDE3 and PDE4 families predominate, degrading cAMP and thereby regulating cardiac excitation-contraction coupling and smooth muscle contractile tone. PDE3 inhibitors are positive inotropes and vasodilators in humans, but their use is limited to acute heart failure and intermittent claudication. PDE5 is particularly important for the degradation of cGMP in vascular smooth muscle, and PDE5 inhibitors are used to treat erectile dysfunction and pulmonary hypertension. There is experimental evidence that these PDEs, as well as other PDE families, including PDE1, PDE2 and PDE9, may play important roles in cardiac diseases, such as hypertrophy and heart failure, as well as several vascular diseases. After a brief presentation of the cyclic nucleotide pathways in cardiac and vascular cells, and the major characteristics of the PDE superfamily, this review will focus on the current use of PDE inhibitors in cardiovascular diseases, and the recent research developments that could lead to better exploitation of the therapeutic potential of these enzymes in the future. PMID:27184830

  13. Phosphodiesterase 10A Upregulation Contributes to Pulmonary Vascular Remodeling

    PubMed Central

    Tian, Xia; Vroom, Christina; Ghofrani, Hossein Ardeschir; Weissmann, Norbert; Bieniek, Ewa; Grimminger, Friedrich; Seeger, Werner; Schermuly, Ralph Theo; Pullamsetti, Soni Savai

    2011-01-01

    Phosphodiesterases (PDEs) modulate the cellular proliferation involved in the pathophysiology of pulmonary hypertension (PH) by hydrolyzing cAMP and cGMP. The present study was designed to determine whether any of the recently identified PDEs (PDE7-PDE11) contribute to progressive pulmonary vascular remodeling in PH. All in vitro experiments were performed with lung tissue or pulmonary arterial smooth muscle cells (PASMCs) obtained from control rats or monocrotaline (MCT)-induced pulmonary hypertensive (MCT-PH) rats, and we examined the effects of the PDE10 inhibitor papaverine (Pap) and specific small interfering RNA (siRNA). In addition, papaverine was administrated to MCT-induced PH rats from day 21 to day 35 by continuous intravenous infusion to examine the in vivo effects of PDE10A inhibition. We found that PDE10A was predominantly present in the lung vasculature, and the mRNA, protein, and activity levels of PDE10A were all significantly increased in MCT PASMCs compared with control PASMCs. Papaverine and PDE10A siRNA induced an accumulation of intracellular cAMP, activated cAMP response element binding protein and attenuated PASMC proliferation. Intravenous infusion of papaverine in MCT-PH rats resulted in a 40%–50% attenuation of the effects on pulmonary hypertensive hemodynamic parameters and pulmonary vascular remodeling. The present study is the first to demonstrate a central role of PDE10A in progressive pulmonary vascular remodeling, and the results suggest a novel therapeutic approach for the treatment of PH. PMID:21494592

  14. Phosphodiesterase 10A upregulation contributes to pulmonary vascular remodeling.

    PubMed

    Tian, Xia; Vroom, Christina; Ghofrani, Hossein Ardeschir; Weissmann, Norbert; Bieniek, Ewa; Grimminger, Friedrich; Seeger, Werner; Schermuly, Ralph Theo; Pullamsetti, Soni Savai

    2011-01-01

    Phosphodiesterases (PDEs) modulate the cellular proliferation involved in the pathophysiology of pulmonary hypertension (PH) by hydrolyzing cAMP and cGMP. The present study was designed to determine whether any of the recently identified PDEs (PDE7-PDE11) contribute to progressive pulmonary vascular remodeling in PH. All in vitro experiments were performed with lung tissue or pulmonary arterial smooth muscle cells (PASMCs) obtained from control rats or monocrotaline (MCT)-induced pulmonary hypertensive (MCT-PH) rats, and we examined the effects of the PDE10 inhibitor papaverine (Pap) and specific small interfering RNA (siRNA). In addition, papaverine was administrated to MCT-induced PH rats from day 21 to day 35 by continuous intravenous infusion to examine the in vivo effects of PDE10A inhibition. We found that PDE10A was predominantly present in the lung vasculature, and the mRNA, protein, and activity levels of PDE10A were all significantly increased in MCT PASMCs compared with control PASMCs. Papaverine and PDE10A siRNA induced an accumulation of intracellular cAMP, activated cAMP response element binding protein and attenuated PASMC proliferation. Intravenous infusion of papaverine in MCT-PH rats resulted in a 40%-50% attenuation of the effects on pulmonary hypertensive hemodynamic parameters and pulmonary vascular remodeling. The present study is the first to demonstrate a central role of PDE10A in progressive pulmonary vascular remodeling, and the results suggest a novel therapeutic approach for the treatment of PH. PMID:21494592

  15. Specific Inhibition of Phosphodiesterase-4B Results in Anxiolysis and Facilitates Memory Acquisition.

    PubMed

    McGirr, Alexander; Lipina, Tatiana V; Mun, Ho-Suk; Georgiou, John; Al-Amri, Ahmed H; Ng, Enoch; Zhai, Dongxu; Elliott, Christina; Cameron, Ryan T; Mullins, Jonathan G L; Liu, Fang; Baillie, George S; Clapcote, Steven J; Roder, John C

    2016-03-01

    Cognitive dysfunction is a core feature of dementia and a prominent feature in psychiatric disease. As non-redundant regulators of intracellular cAMP gradients, phosphodiesterases (PDE) mediate fundamental aspects of brain function relevant to learning, memory, and higher cognitive functions. Phosphodiesterase-4B (PDE4B) is an important phosphodiesterase in the hippocampal formation, is a major Disrupted in Schizophrenia 1 (DISC1) binding partner and is itself a risk gene for psychiatric illness. To define the effects of specific inhibition of the PDE4B subtype, we generated mice with a catalytic domain mutant form of PDE4B (Y358C) that has decreased ability to hydrolyze cAMP. Structural modeling predictions of decreased function and impaired binding with DISC1 were confirmed in cell assays. Phenotypic characterization of the PDE4B(Y358C) mice revealed facilitated phosphorylation of CREB, decreased binding to DISC1, and upregulation of DISC1 and β-Arrestin in hippocampus and amygdala. In behavioral assays, PDE4B(Y358C) mice displayed decreased anxiety and increased exploration, as well as cognitive enhancement across several tests of learning and memory, consistent with synaptic changes including enhanced long-term potentiation and impaired depotentiation ex vivo. PDE4B(Y358C) mice also demonstrated enhanced neurogenesis. Contextual fear memory, though intact at 24 h, was decreased at 7 days in PDE4B(Y358C) mice, an effect replicated pharmacologically with a non-selective PDE4 inhibitor, implicating cAMP signaling by PDE4B in a very late phase of consolidation. No effect of the PDE4B(Y358C) mutation was observed in the prepulse inhibition and forced swim tests. Our data establish specific inhibition of PDE4B as a promising therapeutic approach for disorders of cognition and anxiety, and a putative target for pathological fear memory. PMID:26272049

  16. Phosphodiesterase inhibitors for the treatment of pulmonary hypertension.

    PubMed

    Wilkins, M R; Wharton, J; Grimminger, F; Ghofrani, H A

    2008-07-01

    The pulmonary vascular bed is both a source of and target for a number of vasoactive factors. Among the most important for pulmonary vascular homeostasis are factors that utilise cyclic guanosine monophosphate (cGMP) as an intracellular second messenger. These include nitric oxide and the natriuretic peptide family (atrial, brain and C-type natriuretic peptides). In the search for therapeutic strategies that engage the cGMP signalling pathway for the treatment of pulmonary arterial hypertension (PAH), inhibition of cGMP metabolism by phosphodiesterase type 5 (PDE5)-targeted compounds has proven most successful to date. One PDE5 inhibitor, sildenafil, has been shown to improve pulmonary haemodynamics and exercise capacity in patients with PAH and is now an approved treatment. Others are under investigation. An interesting, although still tentative, observation is the potential of sildenafil to reduce pulmonary vascular resistance without adversely affecting ventilation-perfusion matching. Another is the expression of phosphodiesterase type 5 in the hypertrophied right ventricle. These data suggest that phosphodiesterase type 5 inhibitors may have effects that distinguish them from other treatments for pulmonary hypertension and merit further study. PMID:18591337

  17. Molecular cloning and subcellular distribution of the novel PDE4B4 cAMP-specific phosphodiesterase isoform.

    PubMed Central

    Shepherd, Malcolm; McSorley, Theresa; Olsen, Aileen E; Johnston, Lee Ann; Thomson, Neil C; Baillie, George S; Houslay, Miles D; Bolger, Graeme B

    2003-01-01

    We have isolated cDNAs encoding PDE4B4, a new cAMP-specific phosphodiesterase (PDE4) isoform with novel properties. The amino acid sequence of PDE4B4 demonstrates that it is encoded by the PDE4B gene, but that it differs from the previously isolated PDE4B1, PDE4B2 and PDE4B3 isoforms by the presence of a novel N-terminal region of 17 amino acids. PDE4B4 contains both of the upstream conserved region 1 (UCR1) and UCR2 regulatory units that are characteristic of 'long' PDE4 isoforms. RNase protection demonstrated that PDE4B4 mRNA is expressed preferentially in liver, skeletal muscle and various regions of the brain, which differs from the pattern of tissue distribution of the other known PDE4B long forms, PDE4B1 and PDE4B3. Expression of PDE4B4 cDNA in COS7 cells produced a protein of 85 kDa under denaturing conditions. Subcellular fractionation of recombinant, COS7-cell expressed PDE4B4 showed that the protein was localized within the cytosol, which was confirmed by confocal microscopic analysis of living COS7 cells transfected with a green fluorescent protein-PDE4B4 chimaera. PDE4B4 exhibited a K(m) for cAMP of 5.4 microM and a V(max), relative to that of the long PDE4B1 isoform, of 2.1. PDE4B4 was inhibited by the prototypical PDE4 inhibitor rolipram [4-[3-(cyclopentoxyl)-4-methoxyphenyl]-2-pyrrolidinone] with an IC(50) of 83 nM. Treatment of COS7 cells with forskolin, to elevate cAMP levels, produced activation of PDE4B4, which was associated with the phosphorylation of PDE4B4 on Ser-56 within UCR1. The unique tissue distribution and intracellular targeting of PDE4B4 suggests that this isoform may have a distinct functional role in regulating cAMP levels in specific cell types. PMID:12441002

  18. Union Camp`s deinking evolution

    SciTech Connect

    White, K.M.

    1995-12-01

    When Union Camp Corp. (Wayne, NJ) opened its fiber recycling plant in Franklin, VA, in December 1994, it was on the cutting edge of a new type of deinking mill that has sprung up in the US during the past few years. Designed to handle a lower-quality feedstock that was once a threat to deinking facilities, the mill is processing 400 tpd of mixed office waste paper. Although mill operators eventually hope to switch to a higher-quality feedstock, the mill has successfully been processing all its incoming fiber into 100% recycled pulp. This pulp is used for the company`s various recycled-content papers, which are manufactured at Union Camp`s paper mill, also located in Franklin, or sold on the open market. Combined with modern technology and teamwork, these factors have led to a deinking facility that is exceeding plant operators` expectations for both capacity and throughput.

  19. Foreign Language Camps.

    ERIC Educational Resources Information Center

    Griswold, Jean S.

    The Colorado State University Foreign Language Weekend Camps (also called the "Poor Man's Study Abroad") are described in this report. Developed to provide an international component and a mini foreign experience for the university's students, the camps are designed to accomplish several purposes including: to offer both foreign and United States…

  20. Marketing for Camp Trends.

    ERIC Educational Resources Information Center

    Biddle, Alicia

    1998-01-01

    To effectively market a camp, current trends and issues must be considered: specialty programming, the Americans With Disabilities Act, competing recreational programs, changes in the school year, programming for seniors, and accountability. Camps should have a marketing strategy that includes public relations, a marketing plan, a pricing…

  1. Camp Joy: Embracing Diversity.

    ERIC Educational Resources Information Center

    Krehbiel, Amy

    2001-01-01

    Camp Joy (Ohio) offers a racially integrated program to disadvantaged inner-city foster children. To attract quality minority staff, the camp recruits through former campers, word of mouth, a leader-in-training program, job and internship fairs, and networking with nearby colleges and social agencies. Staff training and the intrinsic rewards of…

  2. Today's Child - Tomorrow's Camp.

    ERIC Educational Resources Information Center

    Ditter, Robert B.

    1988-01-01

    Are camps solely in the business of providing fun, or are they facilitators of crucial life skills? A social worker explores the fun versus character-building debate, concluding that though camp is not group therapy, it contributes to overall personal growth and to the social and emotional development of all children. (JMM)

  3. Physical Fitness at Camp.

    ERIC Educational Resources Information Center

    Steen, Thomas B.; And Others

    1990-01-01

    Describes decline in youth fitness, emphasizing role of camping programs in youth fitness education. Describes Michigan camp's fitness program, consisting of daily workouts, fitness education, and record keeping. Describes fitness consultants' role in program. Discusses program's highlights and problems, suggesting changes for future use. Shows…

  4. Maximizing Camp Property.

    ERIC Educational Resources Information Center

    Knapp, Clifford

    1987-01-01

    Provides selected 14-item outdoor-environmental education bibliography and 20-item checklist of factors weekend/summer camp directors should consider when pondering entry into the outdoor education market. Covers issues of camp philosophy, staff, facilities additions/alterations, equipment, food service, competition, environmental impact,…

  5. Camp Nursing: Student Internships.

    ERIC Educational Resources Information Center

    Harwood, Catherine Hoe; Van Hofwegen, Lynn

    2002-01-01

    Camps can meet or supplement their health care delivery needs by using student nurses. Three models for student nurse internships, basic information about nursing education, and tips for negotiating student nurse internships are described. Sidebars present resources for camp health centers, nursing student competence characteristics, types of…

  6. Friends' Discovery Camp

    ERIC Educational Resources Information Center

    Seymour, Seth

    2008-01-01

    This article features Friends' Discovery Camp, a program that allows children with and without autism spectrum disorder to learn and play together. In Friends' Discovery Camp, campers take part in sensory-rich experiences, ranging from hands-on activities and performing arts to science experiments and stories teaching social skills. Now in its 7th…

  7. Osteopenia in transgenic mice with osteoblast-targeted expression of the inducible cAMP early repressor

    PubMed Central

    Chandhoke, Taranpreet K.; Huang, Yu-Feng; Liu, Fei; Gronowicz, Gloria A.; Adams, Douglas J.; Harrison, John R.; Kream, Barbara E.

    2008-01-01

    ICER is a member of the CREM family of basic leucine zipper transcription factors that acts as a dominant negative regulator of gene transcription. Four different isoforms of ICER (I, Iγ, II and IIγ) are transcribed from the P2 promoter of the Crem gene. We previously found that each of the ICER isoforms is induced by parathyroid hormone in osteoblasts. The goal of the present study was to assess the function of ICER in bone by overexpressing ICER in osteoblasts of transgenic mice. ICER I and ICER II cDNAs, each containing an N-terminal FLAG epitope tag, were cloned downstream of a fragment containing 3.6 kb of the rat Col1a1 promoter and most of the rat Col1a1 first intron to produce pOBCol3.6-ICER I and pOBCol3.6-ICER II transgenes, respectively. Multiple lines of mice were generated bearing the ICER I and ICER II transgenes. At 8 weeks of age, ICER I and ICER II transgenic mice had lower body weights and decreased bone mineral density of femurs and vertebrae. Further studies were done with ICER I transgenic mice, which had had greatly reduced trabecular bone volume and a markedly decreased bone formation rate in femurs. Osteoblast differentiation and osteocalcin expression were reduced in ex vivo bone marrow cultures from ICER I transgenic mice. ICER I antagonized the activity of ATF4 at its consensus DNA binding site in the osteocalcin promoter in vitro. Thus, transgenic mice with osteoblast-targeted overexpression of ICER resulted in osteopenia caused primarily by reduced bone formation. We speculate that ICER regulates the activity and/or expression of ATF/CREB factors required for normal bone formation. PMID:18460422

  8. Facilitation of corticostriatal transmission following pharmacological inhibition of striatal phosphodiesterase 10A: role of nitric oxide-soluble guanylyl cyclase-cGMP signaling pathways.

    PubMed

    Padovan-Neto, Fernando E; Sammut, Stephen; Chakroborty, Shreaya; Dec, Alexander M; Threlfell, Sarah; Campbell, Peter W; Mudrakola, Vishnu; Harms, John F; Schmidt, Christopher J; West, Anthony R

    2015-04-01

    The striatum contains a rich variety of cyclic nucleotide phosphodiesterases (PDEs), which play a critical role in the regulation of cAMP and cGMP signaling. The dual-substrate enzyme PDE10A is the most highly expressed PDE in striatal medium-sized spiny neurons (MSNs) with low micromolar affinity for both cyclic nucleotides. Previously, we have shown that systemic and local administration of the selective PDE10A inhibitor TP-10 potently increased the responsiveness of MSNs to cortical stimulation. However, the signaling mechanisms underlying PDE10A inhibitor-induced changes in corticostriatal transmission are only partially understood. The current studies assessed the respective roles of cAMP and cGMP in the above effects using soluble guanylyl cyclase (sGC) or adenylate cyclase (AC) specific inhibitors. Cortically evoked spike activity was monitored in urethane-anesthetized rats using in vivo extracellular recordings performed proximal to a microdialysis probe during local infusion of vehicle, the selective sGC inhibitor ODQ, or the selective AC inhibitor SQ 22536. Systemic administration of TP-10 (3.2 mg/kg) robustly increased cortically evoked spike activity in a manner that was blocked following intrastriatal infusion of ODQ (50 μm). The effects of TP-10 on evoked activity were due to accumulation of cGMP, rather than cAMP, as the AC inhibitor SQ was without effect. Consistent with these observations, studies in neuronal NO synthase (nNOS) knock-out (KO) mice confirmed that PDE10A operates downstream of nNOS to limit cGMP production and excitatory corticostriatal transmission. Thus, stimulation of PDE10A acts to attenuate corticostriatal transmission in a manner largely dependent on effects directed at the NO-sGC-cGMP signaling cascade. PMID:25855188

  9. The human phosphodiesterase PDE10A gene genomic organization and evolutionary relatedness with other PDEs containing GAF domains.

    PubMed

    Fujishige, K; Kotera, J; Yuasa, K; Omori, K

    2000-10-01

    PDE10A is a cyclic nucleotide phosphodiesterase (PDE) exhibiting properties of a cAMP PDE and a cAMP-inhibited cGMP PDE. The transcripts are specifically expressed in the striatum. The human gene encoding PDE10A was cloned and investigated. The PDE10A gene spanned > 200 kb and contained 24 exons. The exon-intron organization of PDE10A was different from those of PDE5A and PDE6B, although these three PDEs include two GAF domains and have similar amino-acid sequences. The promoter sequence of PDE10A was highly GC-rich and did not contain a TATA motif and a CAAT box, suggesting it is a housekeeping gene. In Caenorhabditis elegans, the C32E12.2 gene encoding a probable PDE that is 48% identical to the human PDE10A protein showed similar exon organization to PDE10A but not PDE5A and PDE6B. This, together with the phylogenic tree analysis, suggested that the ancestral gene for PDE10A existed in a lower organism such as C. elegans. PMID:10998054

  10. Development of a New Radiofluorinated Quinoline Analog for PET Imaging of Phosphodiesterase 5 (PDE5) in Brain

    PubMed Central

    Liu, Jianrong; Wenzel, Barbara; Dukic-Stefanovic, Sladjana; Teodoro, Rodrigo; Ludwig, Friedrich-Alexander; Deuther-Conrad, Winnie; Schröder, Susann; Chezal, Jean-Michel; Moreau, Emmanuel; Brust, Peter; Maisonial-Besset, Aurélie

    2016-01-01

    Phosphodiesterases (PDEs) are enzymes that play a major role in cell signalling by hydrolysing the secondary messengers cyclic adenosine monophosphate (cAMP) and/or cyclic guanosine monophosphate (cGMP) throughout the body and brain. Altered cyclic nucleotide-mediated signalling has been associated with a wide array of disorders, including neurodegenerative disorders. Recently, PDE5 has been shown to be involved in neurodegenerative disorders such as Alzheimer’s disease, but its precise role has not been elucidated yet. To visualize and quantify the expression of this enzyme in brain, we developed a radiotracer for specific PET imaging of PDE5. A quinoline-based lead compound has been structurally modified resulting in the fluoroethoxymethyl derivative ICF24027 with high inhibitory activity towards PDE5 (IC50 = 1.86 nM). Radiolabelling with fluorine-18 was performed by a one-step nucleophilic substitution reaction using a tosylate precursor (RCY(EOB) = 12.9% ± 1.8%; RCP > 99%; SA(EOS) = 70–126 GBq/μmol). In vitro autoradiographic studies of [18F]ICF24027 on different mouse tissue as well as on porcine brain slices demonstrated a moderate specific binding to PDE5. In vivo studies in mice revealed that [18F]ICF24027 was metabolized under formation of brain penetrable radiometabolites making the radiotracer unsuitable for PET imaging of PDE5 in brain. PMID:27110797

  11. Stress history increases alcohol intake in relapse: Relation to phosphodiesterase 10A

    PubMed Central

    Logrip, Marian L.; Zorrilla, Eric P.

    2012-01-01

    Stressful experiences in humans can result in elevated alcohol drinking, as exemplified in many individuals with post-traumatic stress disorder. However, how stress history, rather than acute stressors, influences alcohol intake remains uncertain. To model the protracted effects of past stress, male Wistar rats were subjected to light-cued footshock stress (Stress History) or light cues alone (Control) prior to their acquisition of alcohol self-administration (1-h sessions, fixed ratio1–3, 100 µl of 10% v/v alcohol as reinforcer). Stress history did not alter mean alcohol intake during acquisition of self-administration, but it increased preference for the alcohol-paired lever over the inactive lever. Following an extinction period, rats with a history of stress exposure and low baseline alcohol intake showed a 2-fold elevation in alcohol self-administration, as compared to low-drinking rats with no stress history. Similar effects were not seen in rats self-administering 0.1% sucrose. Analysis of mRNA levels of phosphodiesterase 10A (PDE10A), a dual-specificity cAMP and cGMP hydrolyzing enzyme, showed that stress history increased Pde10a mRNA levels in the basolateral amygdala and, in low drinking rats, the prelimbic prefrontal cortex (plPFC). Pde10a mRNA levels in the plPFC correlated directly with greater alcohol self-administration during the relapse-like phase, and greater BLA Pde10a mRNA levels correlated with increased ethanol preference after acquisition. The data demonstrate that stress history sensitizes otherwise low alcohol drinkers to consume more alcohol in a relapse-like situation, and identify stress-induced neuroadaptations in amygdala and prefrontal cortical Pde10a expression as changes that may drive heightened alcohol intake and preference in susceptible individuals. PMID:22741603

  12. cAMP signalling in trypanosomatids: role in pathogenesis and as a drug target.

    PubMed

    Makin, Laura; Gluenz, Eva

    2015-08-01

    Despite recent research linking cAMP signalling to virulence in trypanosomatids and detailed studies of trypanosomatid adenylyl cyclases (ACs) and phosphodiesterases (PDEs) since their discoveries 40 years ago, downstream components of the pathway and their biological functions have remained remarkably elusive. However, in recent years, significant discoveries have been made: a role for parasite ACs has been proposed in cytokinesis, evasion of the host immune response, and social motility. cAMP phosphodiesterases PDEB1 and PDEB2 were found to be essential for survival and virulence of Trypanosoma brucei and, in Trypanosoma cruzi, PDEC2 was shown to be required for normal osmoregulation. As we discuss here, these breakthroughs have led to an ongoing surge in the development of PDE inhibitors as lead compounds for trypanocidal drugs. PMID:26004537

  13. cAMP signalling in trypanosomatids: role in pathogenesis and as a drug target

    PubMed Central

    Makin, Laura; Gluenz, Eva

    2015-01-01

    Despite recent research linking cAMP signalling to virulence in trypanosomatids and detailed studies of trypanosomatid adenylyl cyclases (ACs) and phosphodiesterases (PDEs) since their discoveries 40 years ago, downstream components of the pathway and their biological functions have remained remarkably elusive. However, in recent years, significant discoveries have been made: a role for parasite ACs has been proposed in cytokinesis, evasion of the host immune response, and social motility. cAMP phosphodiesterases PDEB1 and PDEB2 were found to be essential for survival and virulence of Trypanosoma brucei and, in Trypanosoma cruzi, PDEC2 was shown to be required for normal osmoregulation. As we discuss here, these breakthroughs have led to an ongoing surge in the development of PDE inhibitors as lead compounds for trypanocidal drugs. PMID:26004537

  14. Foreign Language Camps: Camp Waskowitz. Teacher's Guide and Planning Book.

    ERIC Educational Resources Information Center

    Baudin, Phil; And Others

    This guide to running a foreign language camp is intended to cover all aspects of camp administration and program planning. The philosophy of language camps is set forth. The chairperson's responsibilities regarding staff recruitment, staff assignments, and handling finances are outlined. Sample schedules for French, Spanish, and German camps are…

  15. cAMP enhances BMP2-signaling through PKA and MKP1-dependent mechanisms

    SciTech Connect

    Ghayor, Chafik; Ehrbar, Martin; Miguel, Blanca San; Graetz, Klaus W.; Weber, Franz E.

    2009-04-03

    Recent studies suggest that the elevation of intracellular cyclic adenosine monophosphate (cAMP) and the activation of the protein kinase A regulate BMP-induced osteogenesis. However, the precise mechanisms underlying the enhancing effect of cAMP on BMP2 signaling were not completely revealed. In this study we investigated the effect of elevated cAMP level and PKA activation on the BMP2-induced osteoblastic differentiation in pluripotent C2C12 cells. Alkaline phosphatase activity and its mRNA were consistently induced by BMP2 treatment. The pretreatment of C2C12 cells with Forskolin, a cAMP generating agent, dbcAMP, an analogue of cAMP, or IBMX (3-isobutyl 1-methyl xanthine), and a nonspecific inhibitor of phosphodiesterases elicited further activation of alkaline phosphatase. Furthermore, elevated intracellular cAMP level increased BMP2-induced MKP1. On the other hand, BMP2-induced Erk phosphorylation (p44/p42) and cell proliferation were suppressed in the presence of cAMP. Thus, cAMP might enhance BMP2-induced osteoblastic differentiation by a MKP1-Erk-dependent mechanism.

  16. Global and local missions of cAMP signaling in neural plasticity, learning, and memory

    PubMed Central

    Lee, Daewoo

    2015-01-01

    The fruit fly Drosophila melanogaster has been a popular model to study cAMP signaling and resultant behaviors due to its powerful genetic approaches. All molecular components (AC, PDE, PKA, CREB, etc) essential for cAMP signaling have been identified in the fly. Among them, adenylyl cyclase (AC) gene rutabaga and phosphodiesterase (PDE) gene dunce have been intensively studied to understand the role of cAMP signaling. Interestingly, these two mutant genes were originally identified on the basis of associative learning deficits. This commentary summarizes findings on the role of cAMP in Drosophila neuronal excitability, synaptic plasticity and memory. It mainly focuses on two distinct mechanisms (global versus local) regulating excitatory and inhibitory synaptic plasticity related to cAMP homeostasis. This dual regulatory role of cAMP is to increase the strength of excitatory neural circuits on one hand, but to act locally on postsynaptic GABA receptors to decrease inhibitory synaptic plasticity on the other. Thus the action of cAMP could result in a global increase in the neural circuit excitability and memory. Implications of this cAMP signaling related to drug discovery for neural diseases are also described. PMID:26300775

  17. The genetically encoded tool set for investigating cAMP: more than the sum of its parts

    PubMed Central

    Patel, Neha; Gold, Matthew G.

    2015-01-01

    Intracellular fluctuations of the second messenger cyclic AMP (cAMP) are regulated with spatial and temporal precision. This regulation is supported by the sophisticated arrangement of cyclases, phosphodiesterases, anchoring proteins, and receptors for cAMP. Discovery of these nuances to cAMP signaling has been facilitated by the development of genetically encodable tools for monitoring and manipulating cAMP and the proteins that support cAMP signaling. In this review, we discuss the state-of-the-art in development of different genetically encoded tools for sensing cAMP and the activity of its primary intracellular receptor protein kinase A (PKA). We introduce sequences for encoding adenylyl cyclases that enable cAMP levels to be artificially elevated within cells. We chart the evolution of sequences for selectively modifying protein–protein interactions that support cAMP signaling, and for driving cAMP sensors and manipulators to different subcellular locations. Importantly, these different genetically encoded tools can be applied synergistically, and we highlight notable instances that take advantage of this property. Finally, we consider prospects for extending the utility of the tool set to support further insights into the role of cAMP in health and disease. PMID:26300778

  18. Phosphodiesterase MoPdeH targets MoMck1 of the conserved mitogen-activated protein (MAP) kinase signalling pathway to regulate cell wall integrity in rice blast fungus Magnaporthe oryzae.

    PubMed

    Yin, Ziyi; Tang, Wei; Wang, Jingzhen; Liu, Xinyu; Yang, Lina; Gao, Chuyun; Zhang, Jinlong; Zhang, Haifeng; Zheng, Xiaobo; Wang, Ping; Zhang, Zhengguang

    2016-06-01

    In the rice blast fungus Magnaporthe oryzae, the high-affinity cyclic adenosine monophosphate (cAMP) phosphodiesterase MoPdeH is important not only for cAMP signalling and pathogenicity, but also for cell wall integrity (CWI) maintenance through an unknown mechanism. By utilizing affinity purification, we found that MoPdeH interacts with MoMck1, one of the components of the mitogen-activated protein (MAP) kinase cascade that regulates CWI. Overexpression of MoMCK1 suppressed defects in autolysis and pathogenicity of the ΔMopdeH mutant, although partially, suggesting that MoPdeH plays a critical role in CWI maintenance mediated by the MAP kinase pathway. We found that MoMck1 and two other MAP kinase cascade components, MoMkk1 and MoMps1, modulate intracellular cAMP levels by regulating the expression of MoPDEH through a feedback loop. In addition, disruption of MoMKK1 resulted in less aerial hyphal formation, defective asexual development and attenuated pathogenicity. Moreover, MoMkk1 plays a role in the response to osmotic stress via regulation of MoOsm1 phosphorylation levels, whereas endoplasmic reticulum (ER) stress enhances MoMps1 phosphorylation and loss of the MAP kinase cascade component affects the unfolded protein response (UPR) pathway. Taken together, our findings demonstrate that MoPdeH functions upstream of the MoMck1-MoMkk1-MoMps1 MAP kinase pathway to regulate CWI, and that MoPdeH also mediates crosstalk between the cAMP signalling pathway, the osmotic sensing high osmolarity glycerol (HOG) pathway and the dithiothreitol (DTT)-induced UPR pathway in M. oryzae. PMID:27193947

  19. Detailed characterization of a purified type 4 phosphodiesterase, HSPDE4B2B: differentiation of high- and low-affinity (R)-rolipram binding.

    PubMed

    Rocque, W J; Holmes, W D; Patel, I R; Dougherty, R W; Ittoop, O; Overton, L; Hoffman, C R; Wisely, G B; Willard, D H; Luther, M A

    1997-03-01

    We have overexpressed in a baculovirus expression system, and purified to > 95% homogeneity, milligram quantities of a human recombinant rolipram-sensitive cAMP phosphodiesterase, HSPDE4B2B (amino acid residues 81-564). The protein expression levels were approximately 8 mg of HSPDE4B2B (81-564) per liter of Sf9 cells. The Km of the purified enzyme for cAMP was 4 microM and the Ki for the Type 4 phosphodiesterase-specific inhibitor (R)-rolipram was 0.6 microM. The specific activity of the purified protein was 40 mumol/min/mg protein. A nonequilibrium filter binding assay revealed a high-affinity (R)-rolipram binding site on the purified enzyme with a Kd of 1.5 nM and a stoichiometry of 0.05-0.3 mol of (R)-rolipram per mol of HSPDE4B2B (81-564). Equilibrium dialysis experiments revealed a single binding constant of 140 nM with a stoichiometry of 0.75 mol of (R)-rolipram per mol of HSPDE4B2B (81-564). Size exclusion chromatography and analytical ultracentrifugation experiments suggest that the protein exists in multiple association states larger than a monomer. Proteolysis experiments revealed a 43-kDa fragment that contained catalytic and rolipram-inhibitable activities, but the fragment showed no high-affinity (R)-rolipram binding. Based on the proteolytic cleavage studies a 43-kDa protein was constructed, expressed, and purified. This protein, HSPDE4B2B (152-528), had Km and Vmax similar to those of the HSPDE4B2B (81-564) protein, but did not exhibit high-affinity (R)-rolipram binding. The protein did show low-affinity (R)-rolipram binding using the equilibrium binding assay. These results show that a low-affinity binding site for (R)-rolipram is solely contained within the catalytic domain of HSPDE4B2B, whereas high-affinity (R)-rolipram binding requires residues within the catalytic domain and residues flanking N- and/or C-terminal to the catalytic region. PMID:9056484

  20. Better Positioning Those Camp Jobs.

    ERIC Educational Resources Information Center

    Henderson, Karla A.

    1989-01-01

    Discusses summer camps' difficulties in recruiting college students as staff, suggesting camps have "image problem." Describes study of job descriptions to evaluate whether camps offer useful career experiences. Examines frequency and types of job tasks. Examines how camp directors might use job descriptions to recruit more effectively. (TES)

  1. Camp Invention Connects to Classrooms.

    ERIC Educational Resources Information Center

    Krebs, Danute V.; Clark, Barbara D.

    2000-01-01

    This article describes Camp Invention, a national creativity day camp that integrates science, math, social studies, and the arts. The one week camp for children entering grades 2-6 attracts many academically gifted children because of its hands-on curriculum. The camp's curriculum and activities are discussed. (Contains two references.) (CR)

  2. 7-Benzylamino-6-chloro-2-piperazino-4-pyrrolidino-pteridine, a potent inhibitor of cAMP-specific phosphodiesterase, enhancing nuclear protein binding to the CRE consensus sequence in human tumour cells.

    PubMed

    Wagner, Barbara; Jakobs, Sandra; Habermeyer, Michael; Hippe, Frankie; Cho-Chung, Yoon Sang; Eisenbrand, Gerhard; Marko, Doris

    2002-02-15

    The cAMP-specific phosphodiesterase isoenzyme family PDE4 represents the highest cAMP-hydrolysing activity in many human cancer cell lines including the human large cell lung carcinoma cell line LXFL529L. Treatment of LXFL529L cells with the potent PDE4 inhibitor 7-benzylamino-6-chloro-2-piperazino-4-pyrrolidino-pteridine (DC-TA-46) induces dose-dependent growth inhibition. Cells are arrested in the G(1)-phase of the cell cycle and the induction of apoptosis is observed. In this study, we investigated the effect of DC-TA-46 on downstream elements of the cAMP-pathway. DC-TA-46 mediated inhibition of PDE4 activity in LXFL529L cells resulted in an increase of the intracellular cAMP level and significant induction of the activity of protein kinase A (PKA). The regulatory PKA subunit RIalpha was predominantly expressed in LXFL529L cells. In contrast to effects induced by cAMP analogues like 8-Cl-cAMP, the expression of the regulatory subunits of PKA remained unaffected by DC-TA-46. Treatment of LXFL529L cells with DC-TA-46 enhanced the binding of nuclear proteins to the cAMP-responsive element (CRE) consensus sequence TGACGTCA in a time- and dose-dependent manner, indicating the activation of transcription factors by PKA phosphorylation. PMID:11992633

  3. Eukaryotic-Type Ser/Thr Protein Kinase Mediated Phosphorylation of Mycobacterial Phosphodiesterase Affects its Localization to the Cell Wall

    PubMed Central

    Malhotra, Neha; Chakraborti, Pradip K.

    2016-01-01

    Phosphodiesterase enzymes, involved in cAMP hydrolysis reaction, are present throughout phylogeny and their phosphorylation mediated regulation remains elusive in prokaryotes. In this context, we focused on this enzyme from Mycobacterium tuberculosis. The gene encoded by Rv0805 was PCR amplified and expressed as a histidine-tagged protein (mPDE) utilizing Escherichia coli based expression system. In kinase assays, upon incubation with mycobacterial Clade I eukaryotic-type Ser/Thr kinases (PknA, PknB, and PknL), Ni-NTA purified mPDE protein exhibited transphosphorylation ability albeit with varying degree. When mPDE was co-expressed one at a time with these kinases in E. coli, it was also recognized by an anti-phosphothreonine antibody, which further indicates its phosphorylating ability. Mass spectrometric analysis identified Thr-309 of mPDE as a phosphosite. In concordance with this observation, anti-phosphothreonine antibody marginally recognized mPDE-T309A mutant protein; however, such alteration did not affect the enzymatic activity. Interestingly, mPDE expressed in Mycobacterium smegmatis yielded a phosphorylated protein that preferentially localized to cell wall. In contrast, mPDE-T309A, the phosphoablative variant of mPDE, did not show such behavior. On the other hand, phosphomimics of mPDE (T309D or T309E), exhibited similar cell wall anchorage as was observed with the wild-type. Thus, our results provide credence to the fact that eukaryotic-type Ser/Thr kinase mediated phosphorylation of mPDE renders negative charge to the protein, promoting its localization on cell wall. Furthermore, multiple sequence alignment revealed that Thr-309 is conserved among mPDE orthologs of M. tuberculosis complex, which presumably emphasizes evolutionary significance of phosphorylation at this residue. PMID:26904001

  4. Camping in the Snow.

    ERIC Educational Resources Information Center

    Brown, Constance

    1979-01-01

    Describes the experience of winter snow camping. Provides suggestions for shelter, snow kitchens, fires and stoves, cooking, latrines, sleeping warm, dehydration prevention, and clothing. Illustrated with full color photographs. (MA)

  5. Hitler's Death Camps.

    ERIC Educational Resources Information Center

    Wieser, Paul

    1995-01-01

    Presents a high school lesson on Hitler's death camps and the widespread policy of brutality and oppression against European Jews. Includes student objectives, instructional procedures, and a chart listing the value of used clothing taken from the Jews. (CFR)

  6. Pre-Camp Checklists.

    ERIC Educational Resources Information Center

    Camping Magazine, 1995

    1995-01-01

    Provides checklists to prepare for camp opening. Covers such areas as health histories of all campers and staff, staff credentials, condition of facilities and equipment, staff training, horse stables and equipment, and safety of swimming areas. (LP)

  7. Astro Camp Counselors

    NASA Technical Reports Server (NTRS)

    2007-01-01

    Barbara Marino (left), Stennis Space Center education technology specialist, shows Astro Camp Counselor Beverly Fitzsimmons a LEGO model during a teambuilding exercise May 29 at SSC's North Gate computer lab as a part of the counselors' `new hire' orientation.

  8. Phosphodiesterase 4 in inflammatory diseases: Effects of apremilast in psoriatic blood and in dermal myofibroblasts through the PDE4/CD271 complex.

    PubMed

    Schafer, Peter H; Truzzi, Francesca; Parton, Anastasia; Wu, Lei; Kosek, Jolanta; Zhang, Ling-Hua; Horan, Gerald; Saltari, Annalisa; Quadri, Marika; Lotti, Roberta; Marconi, Alessandra; Pincelli, Carlo

    2016-07-01

    Phosphodiesterases 4 (PDE4) act as proinflammatory enzymes via degradation of cAMP, whereas PDE4 inhibitors play an anti-inflammatory role in vitro and in vivo. In particular, apremilast has been recently approved for the treatment of psoriasis and psoriatic arthritis. However, little is known on the expression pattern of PDE4 in psoriasis. We report that PDE4B and PDE4D mRNA are overexpressed in peripheral blood mononuclear cells (PBMC) from psoriasis, as compared with normal controls, while apremilast reduces PBMC production of a number of pro-inflammatory cytokines and increases the levels of anti-inflammatory mediators. PDE4 expression is up-regulated in psoriatic dermis as compared with normal skin, with particular regard to fibroblasts. This is confirmed in vitro, where both dermal fibroblasts (DF) and, to a greater extent, myofibroblasts (DM) express all PDE4 isoforms at the mRNA and protein level. Because PDE4 interacts with the nerve growth factor (NGF) receptor CD271 in lung fibroblasts, we evaluated the relationship and function of PDE4 and CD271 in normal human skin fibroblasts. All PDE4 isoforms co-immunoprecipitate with CD271 in DM, while apremilast inhibits apoptosis induced by β-amyloid, a CD271 ligand, in DM. Furthermore, apremilast significantly reduces NGF- and transforming growth factor-β1 (TGF-β1)-induced fibroblast migration, and inhibits DF differentiation into DM mediated by NGF or TGF-β1. Finally, in DM, apremilast significantly reduces cAMP degradation induced by treatment with β-amyloid. Taken together, these results indicate that PDE4 play an important role in psoriasis. In addition, the study reveals that the PDE4/CD271 complex could be important in modulating fibroblast functions. PMID:26806620

  9. Rescuing lesbian camp.

    PubMed

    Hemmings, Clare

    2007-01-01

    This paper explores the limits of lesbian camp as it is currently conceived within Lesbian Studies. I argue, in what I hope is a rather circuitous way, that a reliance on repudiative models of identity formation fixes gender as complementary and sexuality as oppositional, irrespective of intention. In this context, I imagine instead what it would take to theorize femininity itself as camp, and femme subjects as ideal for working this through at the level of praxis. PMID:17804378

  10. Turning Neighbors into Friends: The Los Angeles Camp Experience.

    ERIC Educational Resources Information Center

    Jenya, Judith

    2002-01-01

    A camp for Los Angeles (California) children traumatized by war, community violence, and hatred gives them time to heal and feel safe. The camp's theme of reconciliation and nonviolence is expressed by an all-volunteer staff through community, teamwork, and cooperation. Nature's diversity is used to show how diverse groups can interact, thrive,…

  11. "Don't Laugh at Me" Coming to Camps.

    ERIC Educational Resources Information Center

    Coleman, Marla

    2000-01-01

    Describes a multimedia resource developed for camps by the American Camping Association and Peter Yarrow, of Peter, Paul, and Mary, to help children express and manage their emotions, cooperate and appreciate the individual strengths that each child brings to a group, celebrate diversity, and practice negotiating and resolving conflict creatively.…

  12. Guidebook: InCamp Education for Migrant Farmworkers.

    ERIC Educational Resources Information Center

    Lynch, Robert; Smith, Mona

    An In-Camp Learning Program focuses on the specific needs of the out-of-school youth and adult migrant farmworker. Although its primary intent is that of education, other areas such as health and social services are addressed. In 1975, New York's In-Camp Learning Program focused on the assessed and expressed educational needs of approximately 400…

  13. A Role for Phosphodiesterase 3B in Acquisition of Brown Fat Characteristics by White Adipose Tissue in Male Mice

    PubMed Central

    Hockman, Steven; Chung, Youn Wook; Ahmad, Faiyaz; Gavrilova, Oksana; Raghavachari, Nalini; Yang, Yanqin; Niu, Gang; Chen, Xiaoyuan; Yu, Zu Xi; Liu, Shiwei; Degerman, Eva

    2013-01-01

    Obesity is linked to various diseases, including insulin resistance, diabetes, and cardiovascular disorders. The idea of inducing white adipose tissue (WAT) to assume characteristics of brown adipose tissue (BAT), and thus gearing it to fat burning instead of storage, is receiving serious consideration as potential treatment for obesity and related disorders. Phosphodiesterase 3B (PDE3B) links insulin- and cAMP-signaling networks in tissues associated with energy metabolism, including WAT. We used C57BL/6 PDE3B knockout (KO) mice to elucidate mechanisms involved in the formation of BAT in epididymal WAT (EWAT) depots. Examination of gene expression profiles in PDE3B KO EWAT revealed increased expression of several genes that block white and promote brown adipogenesis, such as C-terminal binding protein, bone morphogenetic protein 7, and PR domain containing 16, but a clear BAT-like phenotype was not completely induced. However, acute treatment of PDE3B KO mice with the β3-adrenergic agonist, CL316243, markedly increased the expression of cyclooxygenase-2, which catalyzes prostaglandin synthesis and is thought to be important in the formation of BAT in WAT and the elongation of very long-chain fatty acids 3, which is linked to BAT recruitment upon cold exposure, causing a clear shift toward fat burning and the induction of BAT in KO EWAT. These data provide insight into the mechanisms of BAT formation in mouse EWAT, suggesting that, in a C57BL/6 background, an increase in cAMP, caused by ablation of PDE3B and administration of CL316243, may promote differentiation of prostaglandin-responsive progenitor cells in the EWAT stromal vascular fraction into functional brown adipocytes. PMID:23766131

  14. Theophylline Represses IL-8 Secretion from Airway Smooth Muscle Cells Independently of Phosphodiesterase Inhibition. Novel Role as a Protein Phosphatase 2A Activator.

    PubMed

    Patel, Brijeshkumar S; Rahman, Md Mostafizur; Rumzhum, Nowshin N; Oliver, Brian G; Verrills, Nicole M; Ammit, Alaina J

    2016-06-01

    Theophylline is an old drug experiencing a renaissance owing to its beneficial antiinflammatory effects in chronic respiratory diseases, such as asthma and chronic obstructive pulmonary disease. Multiple modes of antiinflammatory action have been reported, including inhibition of the enzymes that degrade cAMP-phosphodiesterase (PDE). Using primary cultures of airway smooth muscle (ASM) cells, we recently revealed that PDE4 inhibitors can potentiate the antiinflammatory action of β2-agonists by augmenting cAMP-dependent expression of the phosphatase that deactivates mitogen-activated protein kinase (MAPK)-MAPK phosphatase (MKP)-1. Therefore, the aim of this study was to address whether theophylline repressed cytokine production in a similar, PDE-dependent, MKP-1-mediated manner. Notably, theophylline did not potentiate cAMP release from ASM cells treated with the long-acting β2-agonist formoterol. Moreover, theophylline (0.1-10 μM) did not increase formoterol-induced MKP-1 messenger RNA expression nor protein up-regulation, consistent with the lack of cAMP generation. However, theophylline (at 10 μM) was antiinflammatory and repressed secretion of the neutrophil chemoattractant cytokine IL-8, which is produced in response to TNF-α. Because theophylline's effects were independent of PDE4 inhibition or antiinflammatory MKP-1, we then wished to elucidate the novel mechanisms responsible. We investigated the impact of theophylline on protein phosphatase (PP) 2A, a master controller of multiple inflammatory signaling pathways, and show that theophylline increases TNF-α-induced PP2A activity in ASM cells. Confirmatory results were obtained in A549 lung epithelial cells. PP2A activators have beneficial effects in ex vivo and in vivo models of respiratory disease. Thus, our study is the first to link theophylline with PP2A activation as a novel mechanism to control respiratory inflammation. PMID:26574643

  15. Exchange protein directly activated by cAMP (epac): a multidomain cAMP mediator in the regulation of diverse biological functions.

    PubMed

    Schmidt, Martina; Dekker, Frank J; Maarsingh, Harm

    2013-04-01

    Since the discovery nearly 60 years ago, cAMP is envisioned as one of the most universal and versatile second messengers. The tremendous feature of cAMP to tightly control highly diverse physiologic processes, including calcium homeostasis, metabolism, secretion, muscle contraction, cell fate, and gene transcription, is reflected by the award of five Nobel prizes. The discovery of Epac (exchange protein directly activated by cAMP) has ignited a new surge of cAMP-related research and has depicted novel cAMP properties independent of protein kinase A and cyclic nucleotide-gated channels. The multidomain architecture of Epac determines its activity state and allows cell-type specific protein-protein and protein-lipid interactions that control fine-tuning of pivotal biologic responses through the "old" second messenger cAMP. Compartmentalization of cAMP in space and time, maintained by A-kinase anchoring proteins, phosphodiesterases, and β-arrestins, contributes to the Epac signalosome of small GTPases, phospholipases, mitogen- and lipid-activated kinases, and transcription factors. These novel cAMP sensors seem to implement certain unexpected signaling properties of cAMP and thereby to permit delicate adaptations of biologic responses. Agonists and antagonists selective for Epac are developed and will support further studies on the biologic net outcome of the activation of Epac. This will increase our current knowledge on the pathophysiology of devastating diseases, such as diabetes, cognitive impairment, renal and heart failure, (pulmonary) hypertension, asthma, and chronic obstructive pulmonary disease. Further insights into the cAMP dynamics executed by the Epac signalosome will help to optimize the pharmacological treatment of these diseases. PMID:23447132

  16. Cyclic nucleotide phosphodiesterase 1 and vascular aging.

    PubMed

    Yan, Chen

    2015-12-01

    VSMCs (vascular smooth muscle cells) play critical roles in arterial remodelling with aging, hypertension and atherosclerosis. VSMCs exist in diverse phenotypes and exhibit phenotypic plasticity, e.g. changing from a quiescent/contractile phenotype to an active myofibroblast-like, often called 'synthetic', phenotype. Synthetic VSMCs are able to proliferate, migrate and secrete ECM (extracellular matrix) proteinases and ECM proteins. In addition, they produce pro-inflammatory molecules, providing an inflammatory microenvironment for leucocyte penetration, accumulation and activation. The aging VSMCs have also shown changes in cellular phenotype, responsiveness to contracting and relaxing mediators, replicating potential, matrix synthesis, inflammatory mediators and intracellular signalling. VSMC dysfunction plays a key role in age-associated vascular remodelling. Cyclic nucleotide PDEs (phosphodiesterases), by catalysing cyclic nucleotide hydrolysis, play a critical role in regulating the amplitude, duration and compartmentalization of cyclic nucleotide signalling. Abnormal alterations of PDEs and subsequent changes in cyclic nucleotide homoeostasis have been implicated in a number of different diseases. In the study published in the latest issue of Clinical Science, Bautista Niño and colleagues have shown that, in cultured senescent human VSMCs, PDE1A and PDE1C mRNA levels are significantly up-regulated and inhibition of PDE1 activity with vinpocetine reduced cellular senescent makers in senescent VSMCs. Moreover, in the premature aging mice with genomic instability (Ercc1(d/-)), impaired aortic ring relaxation in response to SNP (sodium nitroprusside), an NO (nitric oxide) donor, was also largely improved by vinpocetine. More interestingly, using data from human GWAS (genome-wide association studies), it has been found that PDE1A single nucleotide polymorphisms is significantly associated with diastolic blood pressure and carotid intima-media thickening, two

  17. Inactivation of oncogenic cAMP-specific phosphodiesterase 4D by miR-139-5p in response to p53 activation

    PubMed Central

    Cao, Bo; Wang, Kebing; Liao, Jun-Ming; Zhou, Xiang; Liao, Peng; Zeng, Shelya X; He, Meifang; Chen, Lianzhou; He, Yulong; Li, Wen; Lu, Hua

    2016-01-01

    Increasing evidence highlights the important roles of microRNAs in mediating p53’s tumor suppression functions. Here, we report miR-139-5p as another new p53 microRNA target. p53 induced the transcription of miR-139-5p, which in turn suppressed the protein levels of phosphodiesterase 4D (PDE4D), an oncogenic protein involved in multiple tumor promoting processes. Knockdown of p53 reversed these effects. Also, overexpression of miR-139-5p decreased PDE4D levels and increased cellular cAMP levels, leading to BIM-mediated cell growth arrest. Furthermore, our analysis of human colorectal tumor specimens revealed significant inverse correlation between the expression of miR-139-5p and that of PDE4D. Finally, overexpression of miR-139-5p suppressed the growth of xenograft tumors, accompanied by decrease in PDE4D and increase in BIM. These results demonstrate that p53 inactivates oncogenic PDE4D by inducing the expression of miR-139-5p. DOI: http://dx.doi.org/10.7554/eLife.15978.001 PMID:27383270

  18. Zinc deficiency decreases the activity of calmodulin regulated cyclic nucleotide phosphodiesterases in vivo in selected rat tissues.

    PubMed

    Law, J S; McBride, S A; Graham, S; Nelson, N R; Slotnick, B M; Henkin, R I

    1988-08-01

    The effect of zinc deficiency on calmodulin function was investigated by assessing the in vivo activity of two calmodulin regulated enzymes, adenosine 3',5'-monophosphate (c-AMP) and guanosine 3',5'-monophosphate (c-GMP) phosphodiesterase (PDE) in several rat tissues. Enzymatic activities in brain, heart, and testis of rats fed a zinc deficient diet were compared with activities in these tissues from pair fed, zinc supplemented rats. In testis, a tissue in which zinc concentration decreased with zinc deficient diet, enzyme activities were significantly decreased over those in rats who were pair fed zinc supplemented diets. In brain and heart, tissues in which zinc concentrations did not change with either diet, enzymatic activities between the groups were not different. These results indicate that zinc deficiency influences the activity of calmodulin-regulated phosphodiesterases in vivo supporting the hypothesis that zinc plays a role in calmodulin function in vivo in zinc sensitive tissues. PMID:2484550

  19. Are diabetes camps effective?

    PubMed

    Barone, Mark Thomaz Ugliara; Vivolo, Marco Antonio; Madden, Paul B

    2016-04-01

    In the present article data about Diabetes Camps (DC) from all continents were reviewed in order to answer the title question "are diabetes camps effective?". Articles from peer reviewed journals and abstracts published in international conferences proceedings were raised. The effectiveness was considered in terms of knowledge acquisition, and psychosocial and physiological changes. Even though expected improvements were not found in all studies, in a deeper and wider analysis the aspects that influence the most toward gains are identified. Among them are: number of participations in a DC, post-camp educational opportunities, staff training, and program oriented toward campers' autonomy. To conclude, practical recommendations are addressed intending to amplify DC's potential. PMID:27103364

  20. How Green Is Camping? Environmental Stewardship in North Carolina Camps.

    ERIC Educational Resources Information Center

    Warren, Roger; Bingham, Cindy

    1994-01-01

    A survey of 47 residential camps in North Carolina revealed that most camps had written environmental objectives, practiced recycling, attempted to reduce water use and energy consumption, practiced low-impact camping, included environmental issues in staff training, and provided environmental education to campers. Includes survey questions. (LP)

  1. A Camping We Will Go.

    ERIC Educational Resources Information Center

    McAnaney, Kate Divine

    1989-01-01

    The mother of a physically disabled child encourages the participation of such children in mainstream camping programs. Suggestions to maximize the benefits of the camping experience are offered. (DB)

  2. Astro Camp 2000 Rocketry Exercise

    NASA Technical Reports Server (NTRS)

    2000-01-01

    Children at Astro Camp at the John C. Stennis Space Center in Hancock County, Miss., launch rockets as one of their activities in the weeklong camp. Each week during the summer, approximately 30 children ages 9-12 from across Mississippi and Louisiana spend a week learning about space flight. Astro Camp Saturday offers a condensed version of Astro Camp on the third Saturday of each month from January through May 2001.

  3. cAMP Regulation of the lactose operon.

    PubMed

    Szeberenyi, Jozsef

    2004-05-01

    Terms to be familiar with before you start to solve the test: lactose operon, adenylate cyclase, cAMP, catabolite activator protein (CAP), expression plasmid, lac operator, lac repressor, lactose, glucose, promoter, cis- and trans-acting factors. PMID:21706723

  4. CCI: A Worldwide Camping Community.

    ERIC Educational Resources Information Center

    Fawver, Gary Keith

    1992-01-01

    Describes the efforts of Christian Camping International (CCI), an alliance of Christian camping associations from Australia, Canada, the Far East, Latin America, New Zealand, United States, South Africa, Japan, and Brazil. The purpose of CCI is to help develop effective Christian camps, conferences, and retreat ministries. (LP)

  5. Children's Camps in the Adirondacks.

    ERIC Educational Resources Information Center

    Bond, Hallie E.

    2003-01-01

    In the late 19th century, camps in the Adirondacks responded to concerns that the American character was softening. Much camping philosophy came from the progressive movement in education. Aspects of Indian culture were adopted because they seemed to fit naturally in the Adirondacks, and children loved them. Adirondack camps have always been…

  6. Outdoor Education, Camp Casey.

    ERIC Educational Resources Information Center

    Stansberry, Steve; And Others

    A curriculum guide for the Camp Casey Outdoor Education program is contained in this document. Designed for fifth grade students and teachers in Northshore School District, Bothell, Washington, it emphasizes learning activities for use in the outdoors. General understandings relevant to the objectives of the program form the frame into which the…

  7. Saving the Camping Experience.

    ERIC Educational Resources Information Center

    Davies, Jean S.

    1989-01-01

    Discusses developmental encroachment on isolated caves near Pittsford, Vermont. Describes cooperative effort by recreational camps and other agencies to acquire land surrounding cave entrances for conservation. Details successes and hurdles of land acquisition. Describes ongoing work by local campers to maintain and enjoy caves and property. (TES)

  8. Summer Fish Camp.

    ERIC Educational Resources Information Center

    Remick, Dennis; Pulu, Tupou L.

    The booklet presents a description and illustrates, with photographs, the Eskimo lifestyle and the kinds of activities that occur at a summer fish camp on the Yukon River. Eleven suggested activities are listed for the teacher to present when using the booklet. Activities include studying the map of Alaska; tracing the life cycle of the fish;…

  9. A Flying Summer Camp

    ERIC Educational Resources Information Center

    Mercurio, Frank X.

    1975-01-01

    Describes a five-day summer camp which provided 12 children, ages 9-14, with a complete flying experience. The training consisted of ground school and one hour actual flying time, including the basics of aircraft control and a flight prepared and executed by the students. (MLH)

  10. Camp Animal Crackers.

    ERIC Educational Resources Information Center

    McMillan, Erin; And Others

    This guide contains profiles of 16 activities for young children to participate in while attending camp. Each profile contains the theme of the activity, a list of materials needs, a description of the activity itself, and an explanation of the teacher's role in the activity. The activities focus on: (1) songs and dances; (2) dramatic play; (3)…

  11. Recycling at Camp.

    ERIC Educational Resources Information Center

    Cummins, William M.

    1988-01-01

    Outlines a Michigan summer camp's efforts to reduce solid waste disposal by recycling cardboard, tin, glass, aluminum, and plastic milk containers. Points out variables affecting the success of such efforts. Discusses Michigan state funding for the development of recycling programs. (SV)

  12. Sphingomyelin Phosphodiesterase Acid-like 3A (SMPDL3A) Is a Novel Nucleotide Phosphodiesterase Regulated by Cholesterol in Human Macrophages*

    PubMed Central

    Traini, Mathew; Quinn, Carmel M.; Sandoval, Cecilia; Johansson, Erik; Schroder, Kate; Kockx, Maaike; Meikle, Peter J.; Jessup, Wendy; Kritharides, Leonard

    2014-01-01

    Cholesterol-loaded foam cell macrophages are prominent in atherosclerotic lesions and play complex roles in both inflammatory signaling and lipid metabolism, which are underpinned by large scale reprogramming of gene expression. We performed a microarray study of primary human macrophages that showed that transcription of the sphingomyelin phosphodiesterase acid-like 3A (SMPDL3A) gene is up-regulated after cholesterol loading. SMPDL3A protein expression in and secretion from primary macrophages are stimulated by cholesterol loading, liver X receptor ligands, and cyclic AMP, and N-glycosylated SMPDL3A protein is detectable in circulating blood. We demonstrate for the first time that SMPDL3A is a functional phosphodiesterase with an acidic pH optimum. We provide evidence that SMPDL3A is not an acid sphingomyelinase but unexpectedly is active against nucleotide diphosphate and triphosphate substrates at acidic and neutral pH. SMPDL3A is a major source of nucleotide phosphodiesterase activity secreted by liver X receptor-stimulated human macrophages. Extracellular nucleotides such as ATP may activate pro-inflammatory responses in immune cells. Increased expression and secretion of SMPDL3A by cholesterol-loaded macrophage foam cells in lesions may decrease local concentrations of pro-inflammatory nucleotides and potentially represent a novel anti-inflammatory axis linking lipid metabolism with purinergic signaling in atherosclerosis. PMID:25288789

  13. Spatiotemporal regulation of cAMP signaling controls the human trophoblast fusion

    PubMed Central

    Gerbaud, Pascale; Taskén, Kjetil; Pidoux, Guillaume

    2015-01-01

    During human placentation, mononuclear cytotrophoblasts fuse to form multinucleated syncytia ensuring hormonal production and nutrient exchanges between the maternal and fetal circulation. Syncytial formation is essential for the maintenance of pregnancy and for fetal growth. The cAMP signaling pathway is the major route to trigger trophoblast fusion and its activation results in phosphorylation of specific intracellular target proteins, in transcription of fusogenic genes and assembly of macromolecular protein complexes constituting the fusogenic machinery at the plasma membrane. Specificity in cAMP signaling is ensured by generation of localized pools of cAMP controlled by cAMP phosphodiesterases (PDEs) and by discrete spatial and temporal activation of protein kinase A (PKA) in supramolecular signaling clusters inside the cell organized by A-kinase-anchoring proteins (AKAPs) and by organization of signal termination by protein phosphatases (PPs). Here we present original observations on the available components of the cAMP signaling pathway in the human placenta including PKA, PDE, and PP isoforms as well as AKAPs. We continue to discuss the current knowledge of the spatiotemporal regulation of cAMP signaling triggering trophoblast fusion. PMID:26441659

  14. The phosphodiesterase 4 inhibitor roflumilast augments the Th17-promoting capability of dendritic cells by enhancing IL-23 production, and impairs their T cell stimulatory activity due to elevated IL-10.

    PubMed

    Bros, Matthias; Montermann, Evelyn; Cholaszczyńska, Anna; Reske-Kunz, Angelika B

    2016-06-01

    Phosphodiesterase 4 (PDE4) inhibitors serve to prevent degradation of the intracellular second messenger cAMP, resulting in broad anti-inflammatory effects on different cell types including immune cells. Agents that elevate cAMP levels via activation of adenylate cyclase have been shown to imprint a Th17-promoting capacity in dendritic cells (DCs). Therefore, we studied the potential of therapeutically relevant PDE inhibitors to induce a pronounced Th17-skewing capacity in DCs. Here we show that mouse bone marrow-derived (BM-) DCs when treated with the PDE4 inhibitor roflumilast (ROF, trade name: Daxas) in the course of stimulation with LPS (ROF-DCs) evoked elevated IL-17 levels in cocultured allogeneic T cells. In addition, as compared with control settings, levels of IFN-γ remained unaltered, while contents of Th2 cytokines (IL-5, IL-10) were diminished. ROF enhanced expression of the Th17-promoting factor IL-23 in BM-DCs. In line, neutralizing antibodies specific for IL-23 or IL-6 when applied to DC/T cell cocultures partially inhibited the IL17-promoting effect of ROF-DCs. Furthermore, ROF-DCs displayed a markedly diminished allogeneic T cell stimulatory capacity due to enhanced production of IL-10, which was restored upon application of IL-10 specific neutralizing antibody to DC/T cell cocultures. Both the IL-17-inducing and impaired T cell stimulatory capacity of BM-DCs were mimicked by a specific activator of protein kinase A, while stimulation of EPACs (exchange proteins of activated cAMP) did not yield such effects. Taken together, our findings suggest that PDE4 inhibitors aside from their broad overall anti-inflammatory effects may enhance the Th17-polarizing capacity in DCs as an unwanted side effect. PMID:27070502

  15. A short review on structure and role of cyclic-3’,5’-adenosine monophosphate-specific phosphodiesterase 4 as a treatment tool

    PubMed Central

    Eskandari, Nahid; Mirmosayyeb, Omid; Bordbari, Gazaleh; Bastan, Reza; Yousefi, Zahra; Andalib, Alireza

    2015-01-01

    Cyclic nucleotide phosphodiesterases (PDEs) are known as a super-family of enzymes which catalyze the metabolism of the intracellular cyclic nucleotides, cyclic-3’,5’-adenosine monophosphate (cAMP), and cyclic-3’,5’-guanosine monophosphate that are expressed in a variety of cell types that can exert various functions based on their cells distribution. The PDE4 family has been the focus of vast research efforts over recent years because this family is considered as a prime target for therapeutic intervention in a number of inflammatory diseases such as asthma, chronic obstructive pulmonary disease, and rheumatoid arthritis, and it should be used and researched by pharmacists. This is because the major isoform of PDE that regulates inflammatory cell activity is the cAMP-specific PDE, PDE4. This review discusses the relationship between PDE4 and its inhibitor drugs based on structures, cells distribution, and pharmacological properties of PDE4 which can be informative for all pharmacy specialists. PMID:26645022

  16. Comparative involvement of cyclic nucleotide phosphodiesterases and adenylyl cyclase on adrenocorticotropin-induced increase of cyclic adenosine monophosphate in rat and human glomerulosa cells.

    PubMed

    Côté, M; Payet, M D; Rousseau, E; Guillon, G; Gallo-Payet, N

    1999-08-01

    The present study investigated the role and identity of cyclic nucleotide phosphodiesterases (PDEs) in the regulation of basal and ACTH-stimulated levels of intracellular cAMP in human and rat adrenal glomerulosa cells. Comparative dose-response curves indicated that maximal hormone-stimulated cAMP accumulation was 11- and 24-fold higher in human and rat cells, compared with cAMP production obtained in corresponding membranes, respectively. Similarly to 3-isobutyl-1-methyl-xanthine, 25 microM erythro-9-[2-hydroxy-3-nonyl]adenine (EHNA, a specific PDE2 inhibitor), caused a large increase in ACTH-stimulated cAMP accumulation; by contrast, it did not change cAMP production in membranes. Moreover, in membrane fractions, addition of 10 microM cGMP inhibited ACTH-induced cAMP production, an effect completely reversed by addition of 25 microM EHNA. These results indicate that PDE2 activity is involved in the regulation of cAMP accumulation induced by ACTH, and suggest that ACTH inhibits this activity. Indeed, time-course studies indicated that ACTH induced a rapid decrease in cGMP production, resulting in PDE2 inhibition, which in turn, contributed [with adenylyl cyclase (AC) activation] to an accumulation in cAMP for 15 min. Thereafter, cAMP content decreased, because of cAMP-stimulated PDE2, as confirmed by measurement of PDE activity that was activated by ACTH, but only after a 10-min incubation. Hence, we demonstrate that the ACTH-induced increase in intracellular cAMP is the result of a balance between activation of AC and direct modulation of PDE2 activity, an effect mediated by cGMP content. Although similar results were observed in both models, PDE2 involvement is more important in rat than in human adrenal glomerulosa cells, whereas AC is more stimulated in human than in rat glomerulosa cells. PMID:10433216

  17. Osthol attenuates neutrophilic oxidative stress and hemorrhagic shock-induced lung injury via inhibition of phosphodiesterase 4.

    PubMed

    Tsai, Yung-Fong; Yu, Huang-Ping; Chung, Pei-Jen; Leu, Yann-Lii; Kuo, Liang-Mou; Chen, Chun-Yu; Hwang, Tsong-Long

    2015-12-01

    Oxidative stress caused by neutrophils is an important pathogenic factor in trauma/hemorrhagic (T/H)-induced acute lung injury (ALI). Osthol, a natural coumarin found in traditional medicinal plants, has therapeutic potential in various diseases. However, the pharmacological effects of osthol in human neutrophils and its molecular mechanism of action remain elusive. In this study, our data showed that osthol potently inhibited the production of superoxide anion (O2(•-)) and reactive oxidants derived therefrom as well as expression of CD11b in N-formylmethionylleucylphenylalanine (FMLP)-activated human neutrophils. However, osthol inhibited neutrophil degranulation only slightly and it failed to inhibit the activity of subcellular NADPH oxidase. FMLP-induced phosphorylation of extracellular signal-regulated kinase (ERK) and protein kinase B (Akt) was inhibited by osthol. Notably, osthol increased the cAMP concentration and protein kinase A (PKA) activity in activated neutrophils. PKA inhibitors reversed the inhibitory effects of osthol, suggesting that these are mediated through cAMP/PKA-dependent inhibition of ERK and Akt activation. Furthermore, the activity of cAMP-specific phosphodiesterase (PDE) 4, but not PDE3 or PDE7, was significantly reduced by osthol. In addition, osthol reduced myeloperoxidase activity and pulmonary edema in rats subjected to T/H shock. In conclusion, our data suggest that osthol has effective anti-inflammatory activity in human neutrophils through the suppression of PDE4 and protects significantly against T/H shock-induced ALI in rats. Osthol may have potential for future clinical application as a novel adjunct therapy to treat lung inflammation caused by adverse circulatory conditions. PMID:26432981

  18. Therapeutic Utility of Phosphodiesterase Type I Inhibitors in Neurological Conditions

    PubMed Central

    Medina, Alexandre E.

    2011-01-01

    Neuronal plasticity is an essential property of the brain that is impaired in different neurological conditions. Phosphodiesterase type 1 (PDE1) inhibitors can enhance levels of the second messengers cAMP/cGMP leading to the expression of neuronal plasticity-related genes, neurotrophic factors, and neuroprotective molecules. These neuronal plasticity enhancement properties make PDE1 inhibitors good candidates as therapeutic agents in many neurological conditions. However, the lack of specificity of the drugs currently available poses a challenge to the systematic evaluation of the beneficial effect of these agents. The development of more specific drugs may pave the way for the use of PDE1 inhibitors as therapeutic agents in cases of neurodevelopmental conditions such as fetal alcohol spectrum disorders and in degenerative disorders such as Alzheimer's and Parkinson's. PMID:21373359

  19. Phosphodiesterase inhibitors, pentoxifylline and rolipram, increase bone mass mainly by promoting bone formation in normal mice.

    PubMed

    Kinoshita, T; Kobayashi, S; Ebara, S; Yoshimura, Y; Horiuchi, H; Tsutsumimoto, T; Wakabayashi, S; Takaoka, K

    2000-12-01

    The administration of either Pentoxifylline (PTX), a methylxanthine derivative and an inhibitor of cyclic AMP (c-AMP) phosphodiesterases (PDEs), or Rolipram, an inhibitor specific to type-4 PDE (PDE4) in normal mice, significantly increased both cortical and cancellous bone mass. Vertebrae and tibiae from mice treated with PTX or Rolipram were analyzed by means of bone densitometry and histomorphometry. The results revealed that both PTX and Rolipram increased bone mass in normal mice mainly through the acceleration of bone formation. These findings suggest that both PTX and Rolipram can enhance physiological bone formation and thereby increase bone mass in normal mice. The possibility that these agents may be of value for the treatment of osteoporosis is discussed. PMID:11113392

  20. ABCD of the phosphodiesterase family: interaction and differential activity in COPD

    PubMed Central

    Halpin, David MG

    2008-01-01

    Phosphodiesterases (PDEs) are important enzymes that hydrolyze the cyclic nucleotides adenosine 3′5′-cyclic monophosphate (cAMP) and guanosine 3′5′-cyclic mono-phosphate (cGMP) to their inactive 5′ monophosphates. They are highly conserved across species and as well as their role in signal termination, they also have a vital role in intracellular localization of cyclic nucleotide signaling and integration of the cyclic nucleotide pathways with other signaling pathways. Because of their pivotal role in intracellular signaling, they are now of considerable interest as therapeutic targets in a wide variety diseases, including COPD where PDE inhibitors may have bronchodilator, anti-inflammatory and pulmonary vasodilator actions. This review examines the diversity and cellular localization of the isoforms of PDE, the known and speculative relevance of this to the treatment of COPD, and the range of PDE inhibitors in development together with a discussion of their possible role in treating COPD. PMID:19281073

  1. A Novel Glycerophosphodiester Phosphodiesterase, GDE5, Controls Skeletal Muscle Development via a Non-enzymatic Mechanism*

    PubMed Central

    Okazaki, Yuri; Ohshima, Noriyasu; Yoshizawa, Ikumi; Kamei, Yasutomi; Mariggiò, Stefania; Okamoto, Keiko; Maeda, Masahiro; Nogusa, Yoshihito; Fujioka, Yuichiro; Izumi, Takashi; Ogawa, Yoshihiro; Shiro, Yoshitsugu; Wada, Masanobu; Kato, Norihisa; Corda, Daniela; Yanaka, Noriyuki

    2010-01-01

    Mammalian glycerophosphodiester phosphodiesterases (GP-PDEs) have been identified recently and shown to be implicated in several physiological functions. This study isolated a novel GP-PDE, GDE5, and showed that GDE5 selectively hydrolyzes glycerophosphocholine (GroPCho) and controls skeletal muscle development. We show that GDE5 expression was reduced in atrophied skeletal muscles in mice and that decreasing GDE5 abundance promoted myoblastic differentiation, suggesting that decreased GDE5 expression has a counter-regulatory effect on the progression of skeletal muscle atrophy. Forced expression of full-length GDE5 in cultured myoblasts suppressed myogenic differentiation. Unexpectedly, a truncated GDE5 construct (GDE5ΔC471), which contained a GP-PDE sequence identified in other GP-PDEs but lacked GroPCho phosphodiesterase activity, showed a similar inhibitory effect. Furthermore, transgenic mice specifically expressing GDE5ΔC471 in skeletal muscle showed less skeletal muscle mass, especially type II fiber-rich muscle. These results indicate that GDE5 negatively regulates skeletal muscle development even without GroPCho phosphodiesterase activity, providing novel insight into the biological significance of mammalian GP-PDE function in a non-enzymatic mechanism. PMID:20576599

  2. Various phosphodiesterase activities in different regions of the heart alter the cardiac effects of nitric oxide.

    PubMed

    Demirel-Yilmaz, Emine; Cenik, Basar; Ozcan, Gulnihal; Derici, Mehmet Kursat

    2012-09-01

    The modulation of cardiac functions by nitric oxide (NO) was established. This study examined the influences of phosphodiesterase (PDE) inhibitors on the action of NO in the different regions of the rat heart. NO donor diethylamine nonoate (DEA/NO) (0.1-100 μM) decreased functions of the right atrium. DEA/NO-induced depression of the developed tension of the right atrium was inhibited by [erythro-9-(2-hydroxy-3-nonyl)adenine] (PDE2 inhibitor), augmented by milrinone (PDE3 inhibitor), and upturned by rolipram (PDE4 inhibitor). A DEA/NO-induced decrease in the resting tension was inhibited by vinpocetine (PDE1 inhibitor) and [erythro-9-(2-hydroxy-3-nonyl)adenine] but reversed by rolipram. The decreased sinus rate by DEA/NO was prevented by vinpocetine and rolipram. DEA/NO increased cyclic guanosine monophosphate and cyclic adenosine monophosphate (cAMP) concentrations in the right atrium, and rolipram enhanced increased cAMP level. DEA/NO had no effect on the contraction of the papillary muscle. However, unchanged contraction under DEA/NO stimulation was decreased by vinpocetine, milrinone, and rolipram. DEA/NO increased cyclic guanosine monophosphate concentration but has no effect on cAMP in the papillary muscle. However, in the presence of vinpocetine and milrinone, DEA/NO reduced cAMP level. The PDE5 inhibitor sildenafil has no effect on DEA/NO actions. This study indicates that a variety of PDE activities in different regions of the rat heart shapes the action of NO on the myocardium. PMID:22653417

  3. cAMP controls rod photoreceptor sensitivity via multiple targets in the phototransduction cascade

    PubMed Central

    Astakhova, Luba A.; Samoiliuk, Evgeniia V.; Govardovskii, Victor I.

    2012-01-01

    In early studies, both cyclic AMP (cAMP) and cGMP were considered as potential secondary messengers regulating the conductivity of the vertebrate photoreceptor plasma membrane. Later discovery of the cGMP specificity of cyclic nucleotide–gated channels has shifted attention to cGMP as the only secondary messenger in the phototransduction cascade, and cAMP is not considered in modern schemes of phototransduction. Here, we report evidence that cAMP may also be involved in regulation of the phototransduction cascade. Using a suction pipette technique, we recorded light responses of isolated solitary rods from the frog retina in normal solution and in the medium containing 2 µM of adenylate cyclase activator forskolin. Under forskolin action, flash sensitivity rose more than twofold because of a retarded photoresponse turn-off. The same concentration of forskolin lead to a 2.5-fold increase in the rod outer segment cAMP, which is close to earlier reported natural day/night cAMP variations. Detailed analysis of cAMP action on the phototransduction cascade suggests that several targets are affected by cAMP increase: (a) basal dark phosphodiesterase (PDE) activity decreases; (b) at the same intensity of light background, steady background-induced PDE activity increases; (c) at light backgrounds, guanylate cyclase activity at a given fraction of open channels is reduced; and (d) the magnitude of the Ca2+ exchanger current rises 1.6-fold, which would correspond to a 1.6-fold elevation of [Ca2+]in. Analysis by a complete model of rod phototransduction suggests that an increase of [Ca2+]in might also explain effects (b) and (c). The mechanism(s) by which cAMP could regulate [Ca2+]in and PDE basal activity is unclear. We suggest that these regulations may have adaptive significance and improve the performance of the visual system when it switches between day and night light conditions. PMID:23008435

  4. Immunomodulatory Effects of Lippia sidoides Extract: Induction of IL-10 Through cAMP and p38 MAPK-Dependent Mechanisms

    PubMed Central

    Rajgopal, Arun; Rebhun, John F.; Burns, Charlie R.; Scholten, Jeffrey D.; Balles, John A.

    2015-01-01

    Abstract Lippia sidoides is an aromatic shrub that grows wild in the northeastern region of Brazil. In local traditional medicine, the aerial portions of this species are used as anti-infectives, antiseptics, spasmolytics, sedatives, hypotensives, and anti-inflammatory agents. In this research, we evaluate the potential immunological properties of Lippia extract through in vitro analysis of its ability to modulate intracellular cyclic adenosine monophosphate (cAMP) levels and interleukin-10 (IL-10) production. These results show that Lippia extract increases intracellular cAMP through the inhibition of phosphodiesterase activity. They also demonstrate that Lippia extract increases IL-10 production in THP-1 monocytes through both an increase in intracellular cAMP and the activation of p38 MAPK. These results suggest that the Lippia-mediated inhibition of phosphodiesterase activity and the subsequent increase in intracellular cAMP may explain some of the biological activities associated with L. sidoides. In addition, the anti-inflammatory activity of L. sidoides may also be due, in part, to its ability to induce IL-10 production through the inhibition of cyclic nucleotide-dependent phosphodiesterase activity and by its activation of the p38 MAPK pathway. PMID:25599252

  5. Foreign Language Camps at the College Level.

    ERIC Educational Resources Information Center

    Griswold, Jean S.

    Information on Colorado State University's foreign language camps for college students is presented. Advantages of the following two models for camps are identified: a single language camp, and a combination multi-language camp with four languages (Spanish, French, German, and Chinese). Features of the camps include: speaking in the foreign…

  6. Astro Camp Goes to Florida

    NASA Technical Reports Server (NTRS)

    2007-01-01

    Katie Craig, daughter of former Stennis Space Center Deputy Director Mark Craig, launches a 'balloon rocket' with the help of Rebecca Compretta, Astro Camp coordinator at SSC. SSC took Astro Camp on the road to Florida this week to engage children and their parents during activities surrounding the Aug. 8 launch of Space Shuttle Endeavour on NASA's STS-118 mission to the International Space Station. Astro Camp is SSC's popular space camp program designed to inspire and educate students using science and math principles.

  7. No ordinary boot camp.

    PubMed

    Tichy, N M

    2001-04-01

    Many companies now run boot camps--comprehensive orientation programs designed to help new hires hit the ground running. They're intense and intimidating, and new employees emerge from them with strong bonds to other recruits and to the organization. But at Trilogy, organizational consultant Noel Tichy discovered one program that's a breed apart. In this article, Tichy gives us a detailed tour of Trilogy's boot camp, Trilogy University, to demonstrate why it's so different--and so effective. Like the best boot camps, it serves as an immersion in both the technical skills new recruits will need for their jobs and Trilogy's corporate culture, which emphasizes risk-taking, teamwork, humility, and a strong customer focus. But this is a new-employee orientation session that's so fundamental to the company as a whole that it's presided over by the CEO and top corporate executives for fully six months of the year. Why? In two three-month sessions, these top executives hone their own strategic thinking about the company as they decide what to teach the new recruits each session. They also find the company's next generation of new products as they judge the innovative ideas the recruits are tasked with developing--making the program Trilogy's main R&D engine. And they pull the company's rising technical stars into mentoring roles for the new recruits, helping to build the next generation of top leadership. After spending months on-site studying Trilogy University, Tichy came away highly impressed by the power of the virtuous teaching cycle the program has set in motion. Leaders of the organization are learning from recruits at the same time that the recruits are learning from the leaders. It's a model, he argues, that other companies would do well to emulate. PMID:11299694

  8. The Camp Health Manual. An Excellent Reference Written Especially for Organized Camps. Revised.

    ERIC Educational Resources Information Center

    Goldring, David; Middelkamp, J. Neal

    This book is a guide to the diagnosis and care of sick children in organized camping situations. This book presents health care information for the management of medical and surgical problems by the camp counselor, camp director, camp nurse, and camp physician. The chapters are: (1) Camp Standards; (2) The Infirmary; (3) Infirmary Supplies; (4)…

  9. Camp Greentop's Adventure Camp: We Ain't No Rudypoo's.

    ERIC Educational Resources Information Center

    Groff, Diane; Albright, Brian; Purvis, Katie; Creamer, Justin; Pease, Alicia

    2002-01-01

    A day-by-day account describes Camp Greentop's first 5-day adventure camping trip, which was attended by five individuals with disabilities and their counselors. The first day was spent in games and initiatives designed to develop communication, teamwork, and dependability. Other days were devoted to hiking, rock climbing, and whitewater rafting.…

  10. Blending Technology with Camp Tradition: Technology Can Simplify Camp Operations.

    ERIC Educational Resources Information Center

    Salzman, Jeff

    2000-01-01

    Discusses uses of technology appropriate for camps, which are service organizations based on building relationships. Describes relationship marketing and how it can be enhanced through use of Web sites, interactive brochures, and client databases. Outlines other technology uses at camp: automated dispensing of medications, satellite tracking of…

  11. Summer Camp as Therapeutic Context: The Camp Logan Program.

    ERIC Educational Resources Information Center

    McCammon, Susan; And Others

    These symposium papers describe various aspects of the Camp Logan, South Carolina, program, a therapeutic summer residential program for children, ages 8-14, who have significant behavior problems. The philosophy and advantages of the therapeutic camping model are discussed, e.g., structure during the summer, controlled though informal…

  12. Renal Epithelial Cyst Formation and Enlargement in vitro: Dependence on cAMP

    NASA Astrophysics Data System (ADS)

    Mangoo-Karim, Roberto; Uchic, Marie; Lechene, Claude; Grantham, Jared J.

    1989-08-01

    Cysts, a common abnormality of kidneys, are collections of urine-like fluid enclosed by a continuous layer of epithelial cells. Renal cysts derive from nephrons and collecting ducts and progressively enlarge as a consequence of epithelial proliferation and transepithelial fluid secretion. The initiation of cyst formation and the factors that control cyst enlargement are unknown. We used an in vitro model of renal cysts to explore the role of the cAMP signal transduction system in the formation and expansion of cysts. MDCK cells, cultured in hydrated-collagen gel, produced polarized monolayered epithelial cysts when intracellular cAMP was increased by prostaglandin E1, arginine vasopressin, cholera toxin, forskolin, or 8-bromoadenosine 3',5'-cyclic monophosphate. All agonists were potentiated by 3-isobutyl-1-methylxanthine, a nucleotide phosphodiesterase inhibitor. The cell proliferation component of cyst enlargement was accelerated by cAMP agonists, as shown by the increased growth of MDCK cells in subconfluent monolayers. The fluid secretion component, reflected by the transepithelial movement of fluid across polarized monolayers of MDCK cells grown on permeable supports, was stimulated by cAMP agonists in the basolateral medium. Chloride levels were higher in the cyst fluid and the secreted fluid than in the bathing medium. We conclude that the development of MDCK cysts is dependent on cAMP. This signal transduction system may be an important modulator of epithelial cell proliferation and transepithelial fluid secretion in the kidney.

  13. Cloning, chromosomal assignment, and regulation of the rat thyrotropin receptor: Expression of the gene is regulated by thyrotropin, agents that increase cAMP levels, and thyroid autoantibodies

    SciTech Connect

    Akamizu, Takashi; Ikuyama, Shoichiro; Saji, Motoyasu; Kosugi, Shinji; Kozak, C.; McBride, O.W.; Kohn, L.D. )

    1990-08-01

    A rat thyrotropin (thyroid-stimulating hormone, TSH) receptor cDNA was isolated that encoded a protein of 764 amino acids, M{sub r} 86,528. Transfection of the cDNA caused COS-7 cells to develop a TSH-sensitive adenylate cyclase response and the ability to bind {sup 125}I-labeled TSH; both activities were similar to those of rat FRTL-5 thyroid cells and not duplicated by lutropin. The gene represented by the cDNA was assigned to mouse chromosome 12 and human chromosome 14. Northern analyses identified two species of mRNA, 5.6 and 3.3 kilobases, in FRTL-5 thyroid cells; the transcripts appeared to differ only in the extent of their 3{prime} noncoding sequences. There were minimal amounts of the two mRNAs in rat ovary, and neither was detected in RNA preparations from rat testis, liver, lung, brain, spleen, and FRT thyroid cells, which do not have a functional TSH receptor. TSH decreased both mRNA species 3- to 4-fold within 8 hr in FRTL-5 thyroid cells; down-regulation was dependent on TSH concentration and duplicated by forskolin, cholera toxin, or 8-bromo-cAMP but not by a phorbol ester. Down-regulation was also duplicated by throid-stimulating autoantibodies, which increased cAMP levels, but not by thyrotropin binding-inhibiting autoantibodies, which actually increased TSH receptor mRNA levels.

  14. Cloning, chromosomal assignment, and regulation of the rat thyrotropin receptor: expression of the gene is regulated by thyrotropin, agents that increase cAMP levels, and thyroid autoantibodies.

    PubMed Central

    Akamizu, T; Ikuyama, S; Saji, M; Kosugi, S; Kozak, C; McBride, O W; Kohn, L D

    1990-01-01

    A rat thyrotropin (thyroid-stimulating hormone, TSH) receptor cDNA was isolated that encoded a protein of 764 amino acids, Mr 86,528. Transfection of the cDNA caused COS-7 cells to develop a TSH-sensitive adenylate cyclase response and the ability to bind 125I-labeled TSH; both activities were similar to those of rat FRTL-5 thyroid cells and not duplicated by lutropin. The gene represented by the cDNA was assigned to mouse chromosome 12 and human chromosome 14. Northern analyses identified two species of mRNA, 5.6 and 3.3 kilobases, in FRTL-5 thyroid cells; the transcripts appeared to differ only in the extent of their 3' noncoding sequences. There were minimal amounts of the two mRNAs in rat ovary, and neither was detected in RNA preparations from rat testis, liver, lung, brain, spleen, and FRT thyroid cells, which do not have a functional TSH receptor. TSH decreased both mRNA species 3- to 4-fold within 8 hr in FRTL-5 thyroid cells; down-regulation was dependent on TSH concentration and duplicated by forskolin, cholera toxin, or 8-bromo-cAMP but not by a phorbol ester. Down-regulation was also duplicated by thyroid-stimulating autoantibodies, which increased cAMP levels, but not by thyrotropin binding-inhibiting auto-antibodies, which actually increased TSH receptor mRNA levels. Images PMID:1696008

  15. FRET measurements of intracellular cAMP concentrations and cAMP analog permeability in intact cells.

    PubMed

    Börner, Sebastian; Schwede, Frank; Schlipp, Angela; Berisha, Filip; Calebiro, Davide; Lohse, Martin J; Nikolaev, Viacheslav O

    2011-04-01

    Real-time measurements of second messengers in living cells, such as cAMP, are usually performed by ratiometric fluorescence resonance energy transfer (FRET) imaging. However, correct calibration of FRET ratios, accurate calculations of absolute cAMP levels and actual permeabilities of different cAMP analogs have been challenging. Here we present a protocol that allows precise measurements of cAMP concentrations and kinetics by expressing FRET-based cAMP sensors in cells and modulating them with an inhibitor of adenylyl cyclase activity and a cell-permeable cAMP analog that fully inhibits and activates the sensors, respectively. Using this protocol, we observed different basal cAMP levels in primary mouse cardiomyocytes, thyroid cells and in 293A cells. The protocol can be generally applied for calibration of second messenger or metabolite concentrations measured by FRET, and for studying kinetics and pharmacological properties of their membrane-permeable analogs. The complete procedure, including cell preparation and FRET measurements, takes 3-6 d. PMID:21412271

  16. Phosphodiesterase 4-targeted treatments for autoimmune diseases

    PubMed Central

    2013-01-01

    Advancements in phosphodiesterase (PDE)-targeted therapies have shown promise in recent years for treating patients with a variety of autoimmune diseases. This review summarizes the development of PDE4 inhibitors and the associated literature with a focus on treatments for autoimmune diseases. After the initial investigations of the prototypic PDE inhibitor, rolipram, more selective inhibitors targeting the PDE4 isozyme have been developed. With phase II and phase III clinical trials currently underway to evaluate the safety and efficacy of the latest generation of PDE4 inhibitors, namely apremilast, a new class of treatments may be around the corner for patients suffering from chronic, autoimmune diseases. PMID:23557064

  17. Explaining the Value of Camp.

    ERIC Educational Resources Information Center

    Chenery, Mary Faeth

    1994-01-01

    Overviews the philosophy and theory of camp experiences and discusses the special benefits of camps, including experiences that can lead to significant life-changing outcomes, sound educational goals, a sense of psychological and physical safety, and helping children to deal with social problems such as the degradation of the environment and human…

  18. Camping and the Nuclear Drama.

    ERIC Educational Resources Information Center

    Henderson, Karla A.; And Others

    1986-01-01

    Outlines effects on children of nuclear war threat and suggests that camping experiences can give children reasons to hope for and ways to work toward a peaceful world. Recommends 11 ways for camp staff to involve campers in social changes necessary to address nuclear issues. Includes resources list. (LFL)

  19. Kids Camping Takes the Challenge.

    ERIC Educational Resources Information Center

    James, Vickie L.; Hohnbaum, Claudia

    2002-01-01

    A Wisconsin Girl Scout camp integrated The Healthy Kids Challenge into its program. The camp evaluated policies related to meals, snacks, physical activities, team building, and self-esteem. Staff inservice training resulted in healthier meals on the same budget and developed ownership of the program. Campers and families had opportunities to…

  20. Finding the Right Music Camp.

    ERIC Educational Resources Information Center

    Lockhart, Gene R.

    1984-01-01

    Many factors go into selecting a camp. Location, cost, and health facilities are all important considerations. The musical development and objectives of the child and the camp's particular instructional, emotional, recreational, physical, and geographical requirements are all important in the selection process. (CS)

  1. Encountering Child Abuse at Camp.

    ERIC Educational Resources Information Center

    Durall, John K.

    1997-01-01

    Defines child abuse, including the three categories: physical, sexual, and psychological. Presents characteristics and behaviors of each type of abuse, and long-term effects. Discusses how to handle abuse that occurs at camp, and the effects on the camp. Sidebars present abuse statistics, 15 activities that promote psychological wellness, and 8…

  2. Phosphodiesterase 10A inhibitors: analysis of US/EP patents granted since 2012.

    PubMed

    Bartolomé-Nebreda, José-Manuel; Conde-Ceide, Susana; García, María

    2015-01-01

    Phosphodiesterases are enzymes that metabolically inactivate the intracellular second messengers 3',5'-cyclic adenosine and guanosine monophosphate contributing to the control of multiple biological processes. Among them, PDE10A has the most restricted distribution with high expression in striatal medium spiny neurons. Dysfunction of this key brain circuit has been associated with different psychiatric and neurodegenerative disorders. The unique role of PDE10A, together with its increased pharmacological characterization, have prompted enormous interest in investigating the potential of inhibitors of this enzyme as potential novel therapeutic agents This article reviews PDE10A related patents issued in the period 2012-2014 in the USA and Europe offering also a perspective on potential avenues for the future clinical development of phosphodiesterase 10A inhibitors. PMID:26030079

  3. Phosphodiesterase 1C is dispensable for rapid response termination of olfactory sensory neurons

    PubMed Central

    Cygnar, Katherine D.; Zhao, Haiqing

    2009-01-01

    In the nose, odorants are detected on the cilia of olfactory sensory neurons (OSNs), where a cAMP-mediated signaling pathway transforms odor stimulation into electrical responses. Phosphodiesterase (PDE) activity in OSN cilia was long thought to account for rapid response termination by degrading odor-induced cAMP. Two PDEs with distinct cellular localization have been found in OSNs: PDE1C in cilia; PDE4A throughout the cell but absent from cilia. We disrupted both genes in mice and performed electroolfactogram analysis. Unexpectedly, eliminating PDE1C did not prolong response termination. Prolonged termination occurred only in mice lacking both PDEs, suggesting that cAMP degradation by PDE1C in cilia is not a rate-limiting factor for response termination in wildtype. Pde1c−/− OSNs instead displayed reduced sensitivity and attenuated adaptation to repeated stimulation, suggesting potential roles for PDE1C in regulating sensitivity and adaptation. These observations provide new perspectives in regulation of olfactory transduction. PMID:19305400

  4. Age-related alterations in cyclic nucleotide phosphodiesterase activity in dystrophic mouse leg muscle.

    PubMed

    Bloom, Timothy J

    2005-11-01

    Previous reports have described both increased and decreased cyclic nucleotide phosphodiesterase (PDE) activity in dystrophic muscle. Total PDE activity was measured in hind leg muscle from a mouse model of Duchenne muscular dystrophy (mdx) and a genetic control strain at 5, 8, 10, and 15 weeks of age. Total PDE activity declined in fractions isolated from mdx muscle over this time period, but was stable in fractions from control mice. Compared with age-matched controls, younger mdx muscle had higher cAMP and cGMP PDE activity. However, at 15 weeks, fractions from both strains had similar cGMP PDE activity and mdx fractions had lower cAMP PDE activity than controls. Particulate fractions from mdx muscle showed an age-related decline in sensitivity to the PDE4 inhibitor RO 20-1724. A similar loss of sensitivity to the PDE2 inhibitor erythro-9-(2-hydroxyl-3-nonyl)-adenine (EHNA) was seen in a particulate fraction from mdx muscle and to a lesser degree in control muscle. These results suggest that the earlier disagreement regarding altered cyclic nucleotide metabolism in dystrophic muscle may be due to changes with age in PDE activity of dystrophic tissue. The age-related decline in particulate PDE activity seen in dystrophic muscle appears to be isozyme-specific and not due to a generalized decrease in total PDE activity. PMID:16391714

  5. Perspectives on Camp Administration. Readings for Camp Director Education. Camp Administration Series.

    ERIC Educational Resources Information Center

    Farley, Elizabeth, Ed.

    The publication includes 47 selected readings for camp directors who are interested in reviewing the current status of the profession and who want to be a part of shaping its future. The articles, selected from periodicals directly related to camping and, where appropriate, from related journals, were selected and organized to support the American…

  6. cAMP-stimulated Protein Phosphatase 2A Activity Associated with Muscle A Kinase-anchoring Protein (mAKAP) Signaling Complexes Inhibits the Phosphorylation and Activity of the cAMP-specific Phosphodiesterase PDE4D3*

    PubMed Central

    Dodge-Kafka, Kimberly L.; Bauman, Andrea; Mayer, Nicole; Henson, Edward; Heredia, Lorena; Ahn, Jung; McAvoy, Thomas; Nairn, Angus C.; Kapiloff, Michael S.

    2010-01-01

    The concentration of the second messenger cAMP is tightly controlled in cells by the activity of phosphodiesterases. We have previously described how the protein kinase A-anchoring protein mAKAP serves as a scaffold for the cAMP-dependent protein kinase PKA and the cAMP-specific phosphodiesterase PDE4D3 in cardiac myocytes. PKA and PDE4D3 constitute a negative feedback loop whereby PKA-catalyzed phosphorylation and activation of PDE4D3 attenuate local cAMP levels. We now show that protein phosphatase 2A (PP2A) associated with mAKAP complexes is responsible for reversing the activation of PDE4D3 by catalyzing the dephosphorylation of PDE4D3 serine residue 54. Mapping studies reveal that a C-terminal mAKAP domain (residues 2085–2319) binds PP2A. Binding to mAKAP is required for PP2A function, such that deletion of the C-terminal domain enhances both base-line and forskolin-stimulated PDE4D3 activity. Interestingly, PP2A holoenzyme associated with mAKAP complexes in the heart contains the PP2A targeting subunit B56δ. Like PDE4D3, B56δ is a PKA substrate, and PKA phosphorylation of mAKAP-bound B56δ enhances phosphatase activity 2-fold in the complex. Accordingly, expression of a B56δ mutant that cannot be phosphorylated by PKA results in increased PDE4D3 phosphorylation. Taken together, our findings demonstrate that PP2A associated with mAKAP complexes promotes PDE4D3 dephosphorylation, serving both to inhibit PDE4D3 in unstimulated cells and also to mediate a cAMP-induced positive feedback loop following adenylyl cyclase activation and B56δ phosphorylation. In general, PKA·PP2A·mAKAP complexes exemplify how protein kinases and phosphatases may participate in molecular signaling complexes to dynamically regulate localized intracellular signaling. PMID:20106966

  7. cAMP-stimulated protein phosphatase 2A activity associated with muscle A kinase-anchoring protein (mAKAP) signaling complexes inhibits the phosphorylation and activity of the cAMP-specific phosphodiesterase PDE4D3.

    PubMed

    Dodge-Kafka, Kimberly L; Bauman, Andrea; Mayer, Nicole; Henson, Edward; Heredia, Lorena; Ahn, Jung; McAvoy, Thomas; Nairn, Angus C; Kapiloff, Michael S

    2010-04-01

    The concentration of the second messenger cAMP is tightly controlled in cells by the activity of phosphodiesterases. We have previously described how the protein kinase A-anchoring protein mAKAP serves as a scaffold for the cAMP-dependent protein kinase PKA and the cAMP-specific phosphodiesterase PDE4D3 in cardiac myocytes. PKA and PDE4D3 constitute a negative feedback loop whereby PKA-catalyzed phosphorylation and activation of PDE4D3 attenuate local cAMP levels. We now show that protein phosphatase 2A (PP2A) associated with mAKAP complexes is responsible for reversing the activation of PDE4D3 by catalyzing the dephosphorylation of PDE4D3 serine residue 54. Mapping studies reveal that a C-terminal mAKAP domain (residues 2085-2319) binds PP2A. Binding to mAKAP is required for PP2A function, such that deletion of the C-terminal domain enhances both base-line and forskolin-stimulated PDE4D3 activity. Interestingly, PP2A holoenzyme associated with mAKAP complexes in the heart contains the PP2A targeting subunit B56delta. Like PDE4D3, B56delta is a PKA substrate, and PKA phosphorylation of mAKAP-bound B56delta enhances phosphatase activity 2-fold in the complex. Accordingly, expression of a B56delta mutant that cannot be phosphorylated by PKA results in increased PDE4D3 phosphorylation. Taken together, our findings demonstrate that PP2A associated with mAKAP complexes promotes PDE4D3 dephosphorylation, serving both to inhibit PDE4D3 in unstimulated cells and also to mediate a cAMP-induced positive feedback loop following adenylyl cyclase activation and B56delta phosphorylation. In general, PKA.PP2A.mAKAP complexes exemplify how protein kinases and phosphatases may participate in molecular signaling complexes to dynamically regulate localized intracellular signaling. PMID:20106966

  8. Surface expression of GABAA receptors is transcriptionally controlled by the interplay of cAMP-response element-binding protein and its binding partner inducible cAMP early repressor.

    PubMed

    Hu, Yinghui; Lund, Ingrid V; Gravielle, Maria C; Farb, David H; Brooks-Kayal, Amy R; Russek, Shelley J

    2008-04-01

    The regulated expression of type A gamma-aminobutyric acid (GABA) receptor (GABA(A)R) subunit genes plays a critical role in neuronal maturation and synaptogenesis. It is also associated with a variety of neurological diseases. Changes in GABA(A) receptor alpha1 subunit gene (GABRA1) expression have been reported in animal models of epilepsy, alcohol abuse, withdrawal, and stress. Understanding the genetic mechanism behind such changes in alpha subunit expression will lead to a better understanding of the role that signal transduction plays in control over GABA(A)R function and brings with it the promise of providing new therapeutic tools for the prevention or cure of a variety of neurological disorders. Here we show that activation of protein kinase C increases alpha1 subunit levels via phosphorylation of CREB (pCREB) that is bound to the GABRA1 promoter (GABRA1p). In contrast, activation of protein kinase A decreases levels of alpha1 even in the presence of pCREB. Decrease of alpha1 is dependent upon the inducible cAMP early repressor (ICER) as directly demonstrated by ICER-induced down-regulation of endogenous alpha1-containing GABA(A)Rs at the cell surface of cortical neurons. Taken together with the fact that there are less alpha1gamma2-containing GABA(A)Rs in neurons after protein kinase A stimulation and that activation of endogenous dopamine receptors down-regulates alpha1 subunit mRNA levels subsequent to induction of ICER, our studies identify a transcriptional mechanism for regulating the cell surface expression of alpha1-containing GABA(A)Rs that is dependent upon the formation of CREB heterodimers. PMID:18180303

  9. Renal 2',3'-Cyclic Nucleotide 3'-Phosphodiesterase Is an Important Determinant of AKI Severity after Ischemia-Reperfusion.

    PubMed

    Jackson, Edwin K; Menshikova, Elizabeth V; Mi, Zaichuan; Verrier, Jonathan D; Bansal, Rashmi; Janesko-Feldman, Keri; Jackson, Travis C; Kochanek, Patrick M

    2016-07-01

    A positional isomer of 3',5'-cAMP, 2',3'-cAMP, is produced by kidneys in response to energy depletion, and renal 2',3'-cyclic nucleotide 3'-phosphodiesterase (CNPase) metabolizes 2',3'-cAMP to 2'-AMP; 2',3'-cAMP is a potent opener of mitochondrial permeability transition pores (mPTPs), which can stimulate autophagy. Because autophagy protects against AKI, it is conceivable that inhibition of CNPase protects against ischemia-reperfusion (IR) -induced AKI. Therefore, we investigated renal outcomes, mitochondrial function, number, area, and autophagy in CNPase-knockout (CNPase(-/-)) versus wild-type (WT) mice using a unique two-kidney, hanging-weight model of renal bilateral IR (20 minutes of ischemia followed by 48 hours of reperfusion). Analysis of urinary purines showed attenuated metabolism of 2',3'-cAMP to 2'-AMP in CNPase(-/-) mice. Neither genotype nor IR affected BP, heart rate, urine volume, or albumin excretion. In WT mice, renal IR reduced (14)C-inulin clearance (index of GFR) and increased renal vascular resistance (measured by transit time nanoprobes) and urinary excretion of kidney injury molecule-1 and neutrophil gelatinase-associated lipocalin. IR did not affect these parameters in CNPase(-/-) mice. Histologic analysis revealed that IR induced severe damage in kidneys from WT mice, whereas histologic changes were minimal after IR in CNPase(-/-) mice. Measurements of renal cardiolipin levels, citrate synthase activity, rotenone-sensitive NADH oxidase activity, and proximal tubular mitochondrial and autophagosome area and number (by transmission electron microscopy) indicted accelerated autophagy/mitophagy in injured CNPase(-/-) mice. We conclude that CNPase deletion attenuates IR-induced AKI, in part by accelerating autophagy with targeted removal of damaged mitochondria. PMID:26574047

  10. Inhibitors of cyclic nucleotide phosphodiesterase isozymes type-III and type-IV suppress mitogenesis of rat mesangial cells.

    PubMed Central

    Matousovic, K; Grande, J P; Chini, C C; Chini, E N; Dousa, T P

    1995-01-01

    We studied interactions between the mitogen-activated protein kinase (MAPK) signalling pathway and cAMP-protein kinase (PKA) signaling pathway in regulation of mitogenesis of mesangial cells (MC) determined by [3H]thymidine incorporation, with or without added EGF. Forskolin or dibutyryl cAMP strongly (by 60-70%) inhibited [3H]thymidine incorporation into MC. Cilostamide, lixazinone or cilostazol selective inhibitors of cAMP-phosphodiesterase (PDE) isozyme PDE-III, inhibited mitogenesis to similar extent as forskolin and DBcAMP and activated in situ PKA, but without detectable increase in cAMP levels. Cilostamide and cilostazol were more than three times more effective at inhibiting mesangial mitogenesis than rolipram and denbufylline, inhibitors of isozyme PDE-IV, even though PDE-IV was two times more abundant in MC than was PDE-III. On the other hand, when incubated with forskolin, rolipram-enhanced cAMP accumulation was far greater (10-100x) than with cilostamide. EGF increased MAPK activity (+300%); PDE isozyme inhibitors which suppressed mitogenesis also inhibited MAPK. PDE isozyme inhibitors also suppressed PDGF-stimulated MC proliferation. We conclude that cAMP inhibits the mitogen-dependent MAPK-signaling pathway probably by decreasing the activity of Raf-1 due to PKA-catalyzed phosphorylation. Further, we surmise that minor increase in the cAMP pool metabolized by PDE-III is intimately related to regulation of mesangial proliferation. Thus, PDE isozyme inhibitors have the potential to suppress MC proliferation by a focused effect upon signaling pathways. Images PMID:7615811

  11. Camp Courageous of Iowa Staff Manual.

    ERIC Educational Resources Information Center

    Camp Courageous of Iowa, Monticello.

    Designed as a useful and practical tool for the staff at Camp Courageous of Iowa, a year-round residential camp serving all handicapped individuals, the manual outlines safety rules for camp activities, characteristics of the mentally and physically handicapped, and a general description of the camp and its objectives. Contents of the manual…

  12. Self-Concept Change in Camp Staff.

    ERIC Educational Resources Information Center

    Henderson, Karla A.; Bialeschki, M. Deborah

    The 1981 study ascertained whether the self-concept of 66 camp staff from 2 Wisconsin camps changed more than a control group of 18 college students attending summer school; if differences in self-concept were based on a particular characteristic (age, gender, staff position, years at camp); and in what ways, if any, self-concept of camp staff…

  13. 1970s: Camping in Tough Times.

    ERIC Educational Resources Information Center

    Camping Magazine, 1999

    1999-01-01

    In the 1970s, camps were challenged by economic recession, growing administrative demands, and changing attitudes and interests of campers and staff. An illustrative article from 1972, "Facing the Camping Future with Confidence" (Michael F. Buynak), discusses hidden costs in camp operation and the need for camp administration to shift from…

  14. American Camping Association Annual Report, 1999.

    ERIC Educational Resources Information Center

    American Camping Association, Martinsville, IN.

    Founded in 1910 as the Camp Directors' Association of America, the American Camping Association (ACA) is the largest organization serving the organized camping industry. Over 5,500 members come from all segments of the camp profession. This annual report for 1999 describes ACA activities in support of organizational commitments. These commitments…

  15. Easter Seal Guide to Special Camping Programs.

    ERIC Educational Resources Information Center

    Crane, Helen B., Ed.

    Intended for organizations having or planning to establish resident resident camping programs for people with special needs, this guide supplements the American Camping Association's Standards. The philosophy, aims, and objectives of specialized camping programs are considered, and the following are discussed: administration, camp site selection,…

  16. Day Camp Manual: Administration. Book I.

    ERIC Educational Resources Information Center

    Babcock, William

    The first book in a 5-book manual on day camping focuses on summer day camp administration. The book defines day camps as organized group experiences in outdoor living on a day-by-day basis and under trained leadership. It includes a philosophy of day camping, noting benefits to the campers. The book is divided into further chapters that describe…

  17. Foreign Language Weekend Camp Administrative Guidelines.

    ERIC Educational Resources Information Center

    Le Bow, M. Denise

    A procedure for undertaking a foreign language weekend camp is presented. Responsibilities before and after the camp are identified for the foreign language council, the administrators in charge of the camp program for each language, the camp director, participating teachers, and assistants. It is suggested that a minimum of three planning…

  18. Basic Camp Management: An Introduction to Camp Administration. Revised 3rd Edition.

    ERIC Educational Resources Information Center

    Ball, Armand; Ball, Beverly

    This book is the primary text for the Certified Camp Director Program and the Basic Camp Directors Course sponsored by the American Camping Association (Indiana). It provides an orientation for new and prospective camp directors and a quick reference for experienced camp directors. The book covers the following topics: (1) an historical overview…

  19. New Members of the Mammalian Glycerophosphodiester Phosphodiesterase Family

    PubMed Central

    Ohshima, Noriyasu; Kudo, Takahiro; Yamashita, Yosuke; Mariggiò, Stefania; Araki, Mari; Honda, Ayako; Nagano, Tomomi; Isaji, Chiaki; Kato, Norihisa; Corda, Daniela; Izumi, Takashi; Yanaka, Noriyuki

    2015-01-01

    The known mammalian glycerophosphodiester phosphodiesterases (GP-PDEs) hydrolyze glycerophosphodiesters. In this study, two novel members of the mammalian GP-PDE family, GDE4 and GDE7, were isolated, and the molecular basis of mammalian GP-PDEs was further explored. The GDE4 and GDE7 sequences are highly homologous and evolutionarily close. GDE4 is expressed in intestinal epithelial cells, spermatids, and macrophages, whereas GDE7 is particularly expressed in gastro-esophageal epithelial cells. Unlike other mammalian GP-PDEs, GDE4 and GDE7 cannot hydrolyze either glycerophosphoinositol or glycerophosphocholine. Unexpectedly, both GDE4 and GDE7 show a lysophospholipase D activity toward lysophosphatidylcholine (lyso-PC). We purified the recombinant GDE4 and GDE7 proteins and show that these enzymes can hydrolyze lyso-PC to produce lysophosphatidic acid (LPA). Further characterization of purified recombinant GDE4 showed that it can also convert lyso-platelet-activating factor (1-O-alkyl-sn-glycero-3-phosphocholine; lyso-PAF) to alkyl-LPA. These data contribute to our current understanding of mammalian GP-PDEs and of their physiological roles via the control of lyso-PC and lyso-PAF metabolism in gastrointestinal epithelial cells and macrophages. PMID:25528375

  20. Von Braun's Dream: Space Camp.

    ERIC Educational Resources Information Center

    Coleman, C. C.

    1982-01-01

    Describes the "Space Camp" program for boys and girls at the Alabama Space and Rocket Center (Huntsville, Alabama), including typical activities. Includes address for obtaining information on participation in the program. (JN)

  1. Camping Out On An Asteroid

    NASA Video Gallery

    An astronaut and a geologist recently spent three days camping out as though they were on an asteroid. They were inside NASA's Space Exploration Vehicle prototype, flying it virtually in a digital ...

  2. Antidepressant- and anxiolytic-like effects of the phosphodiesterase-4 (PDE4) inhibitor rolipram on behavior depend on cyclic AMP-response element binding protein (CREB)-mediated neurogenesis in the hippocampus

    PubMed Central

    Li, Yun-Feng; Huang, Ying; Amsdell, Simon L.; Xiao, Lan; O'Donnell, James M.; Zhang, Han-Ting

    2009-01-01

    Inhibition of phosphodiesterase-4 (PDE4), an enzyme that catalyzes the hydrolysis of cyclic AMP (cAMP), increases phosphorylation of cAMP-response element binding protein (pCREB) and hippocampal neurogenesis, and produces antidepressant-like effects on behavior; however, causal links among these have not been established. In the present study, chronic administration of rolipram produced antidepressant- and anxiolytic-like effects on behavior in mice. It also increased cAMP and pCREB levels in the hippocampus and prefrontal cortex, but increased Sox2, a marker for mitotic progenitor cells, only in the hippocampus. Chronic rolipram treatment also increased hippocampal neurogenesis, as evidenced by increased bromodeoxyuridine (BrdU)-positive cells in the hippocampal dentate gyrus. Methylazoxymethanol (MAM), which is toxic to proliferating cells, reversed rolipram-induced increases in BrdU-positive cells and pCREB in the hippocampus and partially blocked its behavioral effects. Approximately 84% of BrdU-positive cells became newborn neurons, 93% of which co-expressed pCREB; these proportions were not altered by rolipram or MAM, either alone or in combination. Finally, three weeks following the end of MAM treatment, when neurogenesis was no longer inhibited, rolipram again increased hippocampal pCREB, with its antidepressant- and anxiolytic-like effects resumed. Overall, the present results suggest that rolipram produces its effects on behavior in a manner that at least partially depends on its neurogenic action in the hippocampus, targeting mitotic progenitor cells rather than newborn or mature neurons; cAMP/CREB signaling in hippocampal newborn neurons is critical for neurogenesis and contributes to the behavioral effects of rolipram. PMID:19516250

  3. The cAMP Response Element Binding protein (CREB) is activated by Insulin-like Growth Factor-1 (IGF-1) and regulates myostatin gene expression in skeletal myoblast

    SciTech Connect

    Zuloaga, R.; Fuentes, E.N.; Molina, A.; Valdés, J.A.

    2013-10-18

    Highlights: •IGF-1 induces the activation of CREB via IGF-1R/PI3K/PLC signaling pathway. •Calcium dependent signaling pathways regulate myostatin gene expression. •IGF-1 regulates myostatin gene expression via CREB transcription in skeletal myoblast. -- Abstract: Myostatin, a member of the Transforming Growth Factor beta (TGF-β) superfamily, plays an important role as a negative regulator of skeletal muscle growth and differentiation. We have previously reported that IGF-1 induces a transient myostatin mRNA expression, through the activation of the Nuclear Factor of Activated T cells (NFAT) in an IP{sub 3}/calcium-dependent manner. Here we examined the activation of CREB transcription factor as downstream targets of IGF-1 during myoblast differentiation and its role as a regulator of myostatin gene expression. In cultured skeletal myoblast, IGF-1 induced the phosphorylation and transcriptional activation of CREB via IGF-1 Receptor/Phosphatidylinositol 3-Kinase (PI3K)/Phospholipase C gamma (PLC γ), signaling pathways. Also, IGF-1 induced calcium-dependent molecules such as Calmodulin Kinase II (CaMK II), Extracellular signal-regulated Kinases (ERK), Protein Kinase C (PKC). Additionally, we examined myostatin mRNA levels and myostatin promoter activity in differentiated myoblasts stimulated with IGF-1. We found a significant increase in mRNA contents of myostatin and its reporter activity after treatment with IGF-1. The expression of myostatin in differentiated myoblast was downregulated by the transfection of siRNA–CREB and by pharmacological inhibitors of the signaling pathways involved in CREB activation. By using pharmacological and genetic approaches together these data demonstrate that IGF-1 regulates the myostatin gene expression via CREB transcription factor during muscle cell differentiation.

  4. Advances in targeting cyclic nucleotide phosphodiesterases

    PubMed Central

    Maurice, Donald H.; Ke, Hengming; Ahmad, Faiyaz; Wang, Yousheng; Chung, Jay; Manganiello, Vincent C.

    2014-01-01

    Cyclic nucleotide phosphodiesterases (PDEs) catalyse the hydrolysis of cyclic AMP and cyclic GMP, thereby regulating the intracellular concentrations of these cyclic nucleotides, their signalling pathways and, consequently, myriad biological responses in health and disease. Currently, a small number of PDE inhibitors are used clinically for treating the pathophysiological dysregulation of cyclic nucleotide signalling in several disorders, including erectile dysfunction, pulmonary hypertension, acute refractory cardiac failure, intermittent claudication and chronic obstructive pulmonary disease. However, pharmaceutical interest in PDEs has been reignited by the increasing understanding of the roles of individual PDEs in regulating the subcellular compartmentalization of specific cyclic nucleotide signalling pathways, by the structure-based design of novel specific inhibitors and by the development of more sophisticated strategies to target individual PDE variants. PMID:24687066

  5. Psychiatric aspects of phosphodiesterases: An overview

    PubMed Central

    Murthy, Vasantmeghna S.; Mangot, Ajish G.

    2015-01-01

    Phosphodiesterases (PDE) are exciting new targets in medical sciences. These enzymes are some of the key mediators of cellular functions in the body and hence are attractive sites for drug-induced modulations. With the finding that Tofisopam, a new anxiolytic, inhibits PDEs, the authors were inspired to look into the role of PDE and drugs acting on them in psychiatry. Hence, the review was undertaken. We found several research materials available highlighting the role of PDE in cellular functions and the possible newer etiological mechanisms of neuropsychiatric illnesses such as schizophrenia, depression/anxiety disorders, and cognitive dysfunction involving PDEs. We also found that there are many molecules acting on PDEs, which have the potential to alter the way we treat mental illnesses today. This article is intended to provide an in-depth look at these enzymes so that more cost-effective therapeutic molecules may be synthesized and marketed in India for managing mental illnesses. PMID:26729948

  6. Predictive QSAR modeling of phosphodiesterase 4 inhibitors.

    PubMed

    Kovalishyn, Vasyl; Tanchuk, Vsevolod; Charochkina, Larisa; Semenuta, Ivan; Prokopenko, Volodymyr

    2012-02-01

    A series of diverse organic compounds, phosphodiesterase type 4 (PDE-4) inhibitors, have been modeled using a QSAR-based approach. 48 QSAR models were compared by following the same procedure with different combinations of descriptors and machine learning methods. QSAR methodologies used random forests and associative neural networks. The predictive ability of the models was tested through leave-one-out cross-validation, giving a Q² = 0.66-0.78 for regression models and total accuracies Ac=0.85-0.91 for classification models. Predictions for the external evaluation sets obtained accuracies in the range of 0.82-0.88 (for active/inactive classifications) and Q² = 0.62-0.76 for regressions. The method showed itself to be a potential tool for estimation of IC₅₀ of new drug-like candidates at early stages of drug development. PMID:22023934

  7. Advances in targeting cyclic nucleotide phosphodiesterases.

    PubMed

    Maurice, Donald H; Ke, Hengming; Ahmad, Faiyaz; Wang, Yousheng; Chung, Jay; Manganiello, Vincent C

    2014-04-01

    Cyclic nucleotide phosphodiesterases (PDEs) catalyse the hydrolysis of cyclic AMP and cyclic GMP, thereby regulating the intracellular concentrations of these cyclic nucleotides, their signalling pathways and, consequently, myriad biological responses in health and disease. Currently, a small number of PDE inhibitors are used clinically for treating the pathophysiological dysregulation of cyclic nucleotide signalling in several disorders, including erectile dysfunction, pulmonary hypertension, acute refractory cardiac failure, intermittent claudication and chronic obstructive pulmonary disease. However, pharmaceutical interest in PDEs has been reignited by the increasing understanding of the roles of individual PDEs in regulating the subcellular compartmentalization of specific cyclic nucleotide signalling pathways, by the structure-based design of novel specific inhibitors and by the development of more sophisticated strategies to target individual PDE variants. PMID:24687066

  8. Cardiac uses of phosphodiesterase-5 inhibitors.

    PubMed

    Schwartz, Bryan G; Levine, Laurence A; Comstock, Gary; Stecher, Vera J; Kloner, Robert A

    2012-01-01

    Phosphodiesterase-5 inhibitors (PDE5Is) improve erectile function by enhancing nitric oxide availability in the penis and its supplying vasculature, resulting in vasodilation and increased blood flow. PDE5Is might benefit cardiovascular diseases because phosphodiesterase-5 is also located elsewhere in the body, including the pulmonary and systemic vasculature and in hypertrophied myocardium. PDE5Is are approved for pulmonary arterial hypertension, given that they improved several hemodynamic and clinical parameters in large randomized trials. Initial evidence suggests that PDE5Is benefit patients with congestive heart failure and secondary pulmonary hypertension. PDE5Is seem to improve hemodynamic and clinical parameters in patients with high-altitude pulmonary edema (HAPE) and high-altitude pulmonary hypertension. In climbers with prior episodes of HAPE, PDE5Is prevented HAPE in 2 small randomized trials. In small randomized trials of PDE5Is, patients with Raynaud's phenomenon demonstrated improved blood flow, fewer symptoms and frequency of attacks, and resolution of digital ulcers. In addition to enhancing vasodilation, PDE5Is seem to protect the myocardium through complex pathways that involve nitric oxide, cyclic guanosine monophosphate, protein kinase G, extracellular-signal-regulated kinase, B-cell lymphoma protein-2, and Rho kinase inhibition. In animal models of acute myocardial infarction, PDE5Is consistently reduced infarct size indicating cardioprotection and PDE5Is also promote reverse remodeling and reduce myocardial apoptosis, fibrosis, and hypertrophy. PDE5Is might also benefit patients with treatment-resistant hypertension, preeclampsia, or peripheral arterial disease. This review presents the pathophysiology and trial data with regard to the use of PDE5Is for cardiac diseases. PMID:22192662

  9. AMPK antagonizes hepatic glucagon-stimulated cyclic AMP signalling via phosphorylation-induced activation of cyclic nucleotide phosphodiesterase 4B

    PubMed Central

    Johanns, M.; Lai, Y.-C.; Hsu, M.-F.; Jacobs, R.; Vertommen, D.; Van Sande, J.; Dumont, J. E.; Woods, A.; Carling, D.; Hue, L.; Viollet, B.; Foretz, M; Rider, M H

    2016-01-01

    Biguanides such as metformin have previously been shown to antagonize hepatic glucagon-stimulated cyclic AMP (cAMP) signalling independently of AMP-activated protein kinase (AMPK) via direct inhibition of adenylate cyclase by AMP. Here we show that incubation of hepatocytes with the small-molecule AMPK activator 991 decreases glucagon-stimulated cAMP accumulation, cAMP-dependent protein kinase (PKA) activity and downstream PKA target phosphorylation. Moreover, incubation of hepatocytes with 991 increases the Vmax of cyclic nucleotide phosphodiesterase 4B (PDE4B) without affecting intracellular adenine nucleotide concentrations. The effects of 991 to decrease glucagon-stimulated cAMP concentrations and activate PDE4B are lost in hepatocytes deleted for both catalytic subunits of AMPK. PDE4B is phosphorylated by AMPK at three sites, and by site-directed mutagenesis, Ser304 phosphorylation is important for activation. In conclusion, we provide a new mechanism by which AMPK antagonizes hepatic glucagon signalling via phosphorylation-induced PDE4B activation. PMID:26952277

  10. The effect of phosphodiesterase inhibitors on the extinction of cocaine-induced conditioned place preference in mice.

    PubMed

    Liddie, Shervin; Anderson, Karen L; Paz, Andres; Itzhak, Yossef

    2012-10-01

    Several phosphodiesterase inhibitors (PDEis) improve cognition, suggesting that an increase in brain cAMP and cGMP facilitates learning and memory. Since extinction of drug-seeking behavior requires associative learning, consolidation and formation of new memory, the present study investigated the efficacy of three different PDEis in the extinction of cocaine-induced conditioned place preference (CPP) in B6129S mice. Mice were conditioned by escalating doses of cocaine which was resistant to extinction by free exploration. Immediately following each extinction session mice received (a) saline/vehicle, (b) rolipram (PDE4 inhibitor), (c) BAY-73-6691 (PDE9 inhibitor) or (d) papaverine (PDE10A inhibitor). Mice that received saline/vehicle during extinction training showed no reduction in CPP for >10 days. BAY-73-6691 (a) dose-dependently increased cGMP in hippocampus and amygdala, (b) significantly facilitated extinction and (c) diminished the reinstatement of cocaine CPP. Rolipram, which selectively increased brain cAMP levels, and papaverine which caused increases in both cAMP and cGMP levels, had no significant effect on the extinction of cocaine CPP. The results suggest that increase in hippocampal and amygdalar cGMP levels via blockade of PDE9 has a prominent role in the consolidation of extinction learning. PMID:22596207

  11. The cAMP response element binding protein (CREB) is activated by insulin-like growth factor-1 (IGF-1) and regulates myostatin gene expression in skeletal myoblast.

    PubMed

    Zuloaga, R; Fuentes, E N; Molina, A; Valdés, J A

    2013-10-18

    Myostatin, a member of the Transforming Growth Factor beta (TGF-β) superfamily, plays an important role as a negative regulator of skeletal muscle growth and differentiation. We have previously reported that IGF-1 induces a transient myostatin mRNA expression, through the activation of the Nuclear Factor of Activated T cells (NFAT) in an IP3/calcium-dependent manner. Here we examined the activation of CREB transcription factor as downstream targets of IGF-1 during myoblast differentiation and its role as a regulator of myostatin gene expression. In cultured skeletal myoblast, IGF-1 induced the phosphorylation and transcriptional activation of CREB via IGF-1 Receptor/Phosphatidylinositol 3-Kinase (PI3K)/Phospholipase C gamma (PLC γ), signaling pathways. Also, IGF-1 induced calcium-dependent molecules such as Calmodulin Kinase II (CaMK II), Extracellular signal-regulated Kinases (ERK), Protein Kinase C (PKC). Additionally, we examined myostatin mRNA levels and myostatin promoter activity in differentiated myoblasts stimulated with IGF-1. We found a significant increase in mRNA contents of myostatin and its reporter activity after treatment with IGF-1. The expression of myostatin in differentiated myoblast was downregulated by the transfection of siRNA-CREB and by pharmacological inhibitors of the signaling pathways involved in CREB activation. By using pharmacological and genetic approaches together these data demonstrate that IGF-1 regulates the myostatin gene expression via CREB transcription factor during muscle cell differentiation. PMID:24064350

  12. Self-incompatibility involved in the level of acetylcholine and cAMP.

    PubMed

    Tezuka, Takafumi; Akita, Isamu; Yoshino, Natsuko

    2007-11-01

    Elongation of pollen tubes in pistils after self-pollination of Lilium longiflorum cv. Hinomoto exhibiting strong gametophytic self-incompatibility was promoted by cAMP and also promoted by some metabolic modulators, namely, activators (forskolin and cholera toxin) of adenylate cyclase and inhibitors (3-isobutyl-1-methylxanthine and pertussis) of cyclic nucleotide phosphodiesterase. Moreover, the elongation was promoted by acetylcholine (ACh) and other choline derivatives, such as acetylthiocholine, L-alpha-phosphatidylcholine and chlorocholinechloride [CCC; (2-chloroethyl) trimethyl ammonium chloride]. A potent inhibitor (neostigmine) of acetylcholinesterase (AChE) as well as acetylcholine also promoted the elongation. cAMP enhanced choline acetyltransferase (ChAT) activity and suppressed AChE activity in the pistils, suggesting that the results are closely correlated with self-incompatibility in L. longiflorum. In short, it came to light that cAMP modulates ChAT (acetylcholine-forming enzyme) and AChE (acetylchoine-decomposing enzyme) activities to enhance the level of ACh in the pistils of L. logiflorum after self-incompatible pollination. These results indicate that the self-incompatibility on self-pollination is caused by low levels of ACh and/or cAMP. PMID:19704589

  13. The role of ventral striatal cAMP signaling in stress-induced behaviors.

    PubMed

    Plattner, Florian; Hayashi, Kanehiro; Hernández, Adan; Benavides, David R; Tassin, Tara C; Tan, Chunfeng; Day, Jonathan; Fina, Maggy W; Yuen, Eunice Y; Yan, Zhen; Goldberg, Matthew S; Nairn, Angus C; Greengard, Paul; Nestler, Eric J; Taussig, Ronald; Nishi, Akinori; Houslay, Miles D; Bibb, James A

    2015-08-01

    The cAMP and cAMP-dependent protein kinase A (PKA) signaling cascade is a ubiquitous pathway acting downstream of multiple neuromodulators. We found that the phosphorylation of phosphodiesterase-4 (PDE4) by cyclin-dependent protein kinase 5 (Cdk5) facilitated cAMP degradation and homeostasis of cAMP/PKA signaling. In mice, loss of Cdk5 throughout the forebrain elevated cAMP levels and increased PKA activity in striatal neurons, and altered behavioral responses to acute or chronic stressors. Ventral striatum- or D1 dopamine receptor-specific conditional knockout of Cdk5, or ventral striatum infusion of a small interfering peptide that selectively targeted the regulation of PDE4 by Cdk5, produced analogous effects on stress-induced behavioral responses. Together, our results demonstrate that altering cAMP signaling in medium spiny neurons of the ventral striatum can effectively modulate stress-induced behavioral states. We propose that targeting the Cdk5 regulation of PDE4 could be a new therapeutic approach for clinical conditions associated with stress, such as depression. PMID:26192746

  14. Project REACH: Field Guides 1-4. No. 1: Camp Secretary, No. 2: Camp Nurse, No. 3: Camp Food Services, No. 4: Camp Maintenance Supervisor. Appendices Q 1-4. Camp Staff Training Series.

    ERIC Educational Resources Information Center

    Robb, Gary; And Others

    Resulting from a 3 year project to develop competency based programs for camp personnel working with the physically handicapped, the four field guides task analyze the roles of the camp secretary, camp nurse, camp food services staff, and camp maintenance supervisor. All guides contain an introduction and information on general duties, job…

  15. Activated cAMP receptors switch encystation into sporulation

    PubMed Central

    Kawabe, Yoshinori; Morio, Takahiro; James, John L.; Prescott, Alan R.; Tanaka, Yoshimasa; Schaap, Pauline

    2009-01-01

    Metazoan embryogenesis is controlled by a limited number of signaling modules that are used repetitively at successive developmental stages. The development of social amoebas shows similar reiterated use of cAMP-mediated signaling. In the model Dictyostelium discoideum, secreted cAMP acting on 4 cAMP receptors (cARs1-4) coordinates cell movement during aggregation and fruiting body formation, and induces the expression of aggregation and sporulation genes at consecutive developmental stages. To identify hierarchy in the multiple roles of cAMP, we investigated cAR heterogeneity and function across the social amoeba phylogeny. The gene duplications that yielded cARs 2-4 occurred late in evolution. Many species have only a cAR1 ortholog that duplicated independently in the Polysphondylids and Acytostelids. Disruption of both cAR genes of Polysphondylium pallidum (Ppal) did not affect aggregation, but caused complete collapse of fruiting body morphogenesis. The stunted structures contained disorganized stalk cells, which supported a mass of cysts instead of spores; cAMP triggered spore gene expression in Ppal, but not in the cAR null mutant, explaining its sporulation defect. Encystation is the survival strategy of solitary amoebas, and lower taxa, like Ppal, can still encyst as single cells. Recent findings showed that intracellular cAMP accumulation suffices to trigger encystation, whereas it is a complementary requirement for sporulation. Combined, the data suggest that cAMP signaling in social amoebas evolved from cAMP-mediated encystation in solitary amoebas; cAMP secretion in aggregates prompted the starving cells to form spores and not cysts, and additionally organized fruiting body morphogenesis. cAMP-mediated aggregation was the most recent innovation. PMID:19369200

  16. 1985 Parents' Guide to Accredited Camps. Ninth Annual Edition.

    ERIC Educational Resources Information Center

    American Camping Association, Martinsville, IN.

    The ninth annual edition of the listing of American Camping Association camps provides information for parents on choosing a camp, brief descriptions of approximately 1,900 camps, and 5 indexes. A narrative section offers information on how to look for a camp, special needs campers, kinds of camps, accreditation, contacting a camp, budgeting,…

  17. Regulation of cAMP Intracellular Levels in Human Platelets Stimulated by 2-Arachidonoylglycerol.

    PubMed

    Signorello, Maria Grazia; Leoncini, Giuliana

    2016-05-01

    We demonstrated that in human platelets the endocannabinoid 2-arachidonoylglycerol (2-AG) decreased dose- and time-dependently cAMP intracellular levels. No effect on cAMP decrease induced by 2-AG was observed in the presence of the adenylate cyclase inhibitor SQ22536 as well in platelets pretreated with the thromboxane A2 receptor antagonist, SQ29548 or with aspirin, inhibitor of arachidonic acid metabolism through the cyclooxygenase pathway. An almost complete recovering of cAMP level was measured in platelets pretreated with the specific inhibitor of phosphodiesterase (PDE) 3A, milrinone. In platelets pretreated with LY294002 or MK2206, inhibitors of PI3K/AKT pathway, and with U73122, inhibitor of phospholipase C pathway, only a partial prevention was shown. cAMP intracellular level depends on synthesis by adenylate cyclase and hydrolysis by PDEs. In 2-AG-stimulated platelets adenylate cyclase activity seems to be unchanged. In contrast PDEs appear to be involved. In particular PDE3A was specifically activated, as milrinone reversed cAMP reduction by 2-AG. 2-AG enhanced PDE3A activity through its phosphorylation. The PI3K/AKT pathway and PKC participate to this PDE3A phosphorylation/activation mechanism as it was greatly inhibited by platelet pretreatment with LY294002, MK2206, U73122, or the PKC specific inhibitor GF109203X. Taken together these data suggest that 2-AG potentiates its power of platelet agonist reducing cAMP intracellular level. J. Cell. Biochem. 117: 1240-1249, 2016. © 2015 Wiley Periodicals, Inc. PMID:26460717

  18. Protein kinases A and C regulate receptor-mediated increases in cAMP in rabbit erythrocytes

    PubMed Central

    Sridharan, Meera; Bowles, Elizabeth A.; Stephenson, Alan H.; Ellsworth, Mary L.; Sprague, Randy S.

    2010-01-01

    Activation of the β-adrenergic receptor (β-AR) or the prostacyclin receptor (IPR) results in increases in cAMP and ATP release from erythrocytes. cAMP levels depend on a balance between synthesis via adenylyl cyclase and hydrolysis by phosphodiesterases (PDEs). Previously, we reported that cAMP increases associated with activation of the β-AR and IPR in rabbit and human erythrocytes are tightly regulated by distinct PDEs (1). Importantly, inhibitors of these PDEs potentiated both increases in cAMP and ATP release. It has been shown that increases in protein kinase (PK) activity can activate PDE3 and PDE4. Both PKA and PKC are present in the erythrocyte and can phosphorylate and activate these PDEs. Here we investigate the hypothesis that PKA regulates PDE activity associated with the β-AR and both PKA and PKC regulate the PDE activity associated with the IPR in rabbit erythrocytes. Pretreatment of erythrocytes with the PKA inhibitor, H89 (10 μM), in the presence of the PDE4 inhibitor, rolipram (10 μM), augmented isoproterenol (1 μM)-induced cAMP increases. In contrast, in the presence of the PDE3 inhibitor, cilostazol (10 μM), pretreatment of erythrocytes with either H89 (1 μM) or two chemically dissimilar inhibitors of PKC, calphostin C (1 μM) or GFX109203X (1 μM), potentiated iloprost (1 μM)-induced cAMP increases. Furthermore, pretreatment of erythrocytes with both H89 and GFX109203X in the presence of cilostazol augmented the iloprost-induced increases in cAMP to a greater extent than either PK inhibitor individually. These results support the hypothesis that PDEs associated with receptor-mediated increases in cAMP in rabbit erythrocytes are regulated by kinases specific to the receptor's signaling pathway. PMID:20008267

  19. gRICH68 and gRICH70 are 2',3'-cyclic-nucleotide 3'-phosphodiesterases induced during goldfish optic nerve regeneration.

    PubMed

    Ballestero, R P; Wilmot, G R; Agranoff, B W; Uhler, M D

    1997-04-25

    Biochemical characterization of changes in gene expression that accompany optic nerve regeneration has led to the identification of proteins that may play key roles in the regeneration process. In this report, a cDNA encoding gRICH70, a novel isoform of the regeneration-induced gRICH68 protein, has been identified and characterized in goldfish. Both gRICH68 and gRICH70 show significant homology (34-36%) to mammalian 2',3'-cyclic-nucleotide 3'-phosphodiesterases (CNPases), hence the name goldfish regeneration-induced CNPase homolog (gRICH). The predicted 431-amino acid gRICH70 protein is 88% homologous to gRICH68, and the retinal mRNA for gRICH70 is coordinately induced with gRICH68 mRNA during optic nerve regeneration. Enzymatic analysis of recombinant proteins confirms that both gRICH proteins possess CNPase activity. Despite the relatively limited sequence homology, the kinetic constants obtained suggest that both gRICH proteins are at least as efficient as recombinant mouse CNP1 in catalyzing the hydrolysis of 2',3'-cAMP. Immunoprecipitation studies indicate that gRICH proteins are responsible for the majority of the CNPase activity detected in regenerating goldfish retinas. The evidence presented demonstrates that gRICH68 and gRICH70 correspond to a previously described doublet of acidic proteins that are selectively induced in the goldfish retina during optic nerve regeneration. Thus, CNPase enzyme activity is implicated for the first time in the process of nerve regeneration. PMID:9111061

  20. Adaptive phenotypic modulation of human arterial endothelial cells to fluid shear stress-encoded signals: modulation by phosphodiesterase 4D-VE-cadherin signalling.

    PubMed

    Rampersad, Sarah N; Wudwud, Alie; Hubert, Fabien; Maurice, Donald H

    2016-07-01

    Although cAMP-signalling regulates numerous functions of vascular endothelial cells (VECs), including their ability to impact vascular resistance in response to changes in blood flow dynamics, few of the mechanisms underlying these effects have yet to be described. In addition to forming stable adherens junctions (AJs) in static VEC cultures, VE-cadherin (VECAD) has emerged as a critical component in a key mechanosensor responsible for linking altered blood flow dynamics and the VEC-mediated control of vascular resistance. Previously, a cAMP phosphodiesterase, PDE4D, was shown to coordinate the VEC permeability limiting effects of cAMP-elevating agents in human arterial VECs (HAECs). Herein, we report that PDE4D acts to allow cAMP-elevating agents to regulate VECADs' role as a sensor of flow-associated fluid shear stress (FSS)-encoded information in HAECs. Thus, we report that PDE4 activity is increased in HAECs exposed to laminar FSS and that this effect contributes to controlling how FSS impacts the morphological and gene expression changes in HAECs exposed to flow. More specifically, we report that PDE4D regulates the efficiency with which VECAD, within its mechanosensor, controls VEGFR2 and Akt activities. Indeed, we show that PDE4D knockdown (KD) significantly blunts responses of HAECs to levels of FSS characteristically found in areas of the vasculature in which stenosis is prevalent. We propose that this effect may provide a new therapeutic avenue in modulating VEC behaviour at these sites by promoting an adaptive and vasculo-protective phenotype. PMID:26658094

  1. The novel phosphodiesterase 10A inhibitor THPP-1 has antipsychotic-like effects in rat and improves cognition in rat and rhesus monkey.

    PubMed

    Smith, Sean M; Uslaner, Jason M; Cox, Christopher D; Huszar, Sarah L; Cannon, Christopher E; Vardigan, Joshua D; Eddins, Donnie; Toolan, Dawn M; Kandebo, Monika; Yao, Lihang; Raheem, Izzat T; Schreier, John D; Breslin, Michael J; Coleman, Paul J; Renger, John J

    2013-01-01

    Phosphodiesterase 10A (PDE10A) is a novel target for the treatment of schizophrenia that may address multiple symptomatic domains associated with this disorder. PDE10A is highly expressed in the brain and functions to metabolically inactivate the important second messengers cAMP and cGMP. Here we describe effects of a potent and orally bioavailable PDE10A inhibitor [2-(6-chloropyridin-3-yl)-4-(2-methoxyethoxy)-7,8-dihydropyrido[4,3-d]pyrimidin-6(5H)-yl](imidazo[1,5-a]pyridin-1-yl)methanone] (THPP-1) on striatal signaling pathways, in behavioral tests that predict antipsychotic potential, and assays that measure episodic-like memory in rat and executive function in rhesus monkey. THPP-1 exhibits nanomolar potency on the PDE10A enzyme, demonstrates excellent pharmacokinetic properties in multiple preclinical animal species, and is selective for PDE10A over other PDE families of enzymes. THPP-1 significantly increased phosphorylation of proteins in the striatum involved in synaptic plasticity, including the a-amino-3-hydroxy-5-methylisoxazole-4-proprionic acid receptor (AMPA) GluR1 subunit, extracellular receptor kinase (ERK), and cAMP-response element binding protein (CREB). THPP-1 produced dose-dependent effects in preclinical assays predictive of antipsychotic activity including attenuation of MK-801-induced psychomotor activation and condition avoidance responding in rats. At similar plasma exposures, THPP-1 significantly increased object recognition memory in rat and attenuated a ketamine-induced deficit in the object retrieval detour task in rhesus monkey. These findings suggest that PDE10A inhibitors have the potential to impact multiple symptomatic domains of schizophrenia including positive symptoms and cognitive impairment. This article is part of a Special Issue entitled 'Cognitive Enhancers'. PMID:22750078

  2. Development of a high throughput screen for allosteric modulators of melanocortin-4 receptor signaling using a real time cAMP assay.

    PubMed

    Pantel, Jacques; Williams, Savannah Y; Mi, Dehui; Sebag, Julien; Corbin, Jackie D; Weaver, C David; Cone, Roger D

    2011-06-11

    The melanocortin MC(4) receptor is a potential target for the development of drugs for both obesity and cachexia. Melanocortin MC(4) receptor ligands known thus far are orthosteric agonists or antagonists, however the agonists, in particular, have generally exhibited unwanted side effects. For some receptors, allosteric modulators are expected to reduce side-effect profiles. To identify allosteric modulators of the melanocortin MC(4) receptor, we created HEK293 cell lines coexpressing the human melanocortin MC(4) receptor and a modified luciferase-based cAMP sensor. Monitoring luminescence as a readout of real-time intracellular cAMP concentration, we demonstrate that this cell line is able to report melanocortin agonist responses, as well as inverse agonist response to the physiological AgRP peptide. Based on the MC4R-GLO cell line, we developed an assay that was shown to meet HTS standards (Z'=0.50). A pilot screen run on the Microsource Spectrum compound library (n=2000) successfully identified 62 positive modulators. This screen identified predicted families of compounds: β(2)AR agonists - the β(2)AR being endogenously expressed in HEK293 cells, an adenylyl cyclase activator and finally a distribution of phosphodiesterase (PDE) inhibitors well characterized or recently identified. In this last category, we identified a structural family of coumarin-derived compounds (imperatorin, osthol and prenyletin), along with deracoxib, a drug in veterinary use for its COX2 inhibitory properties. This latter finding unveiled a new off-target mechanism of action for deracoxib as a PDE inhibitor. Overall, these data are the first report of a HTS for allosteric modulators for a Gs protein coupled receptor. PMID:21296065

  3. Development of a high throughput screen for allosteric modulators of melanocortin-4 receptor signaling using a real time cAMP assay

    PubMed Central

    Pantel, Jacques; Williams, Savannah Y.; Mi, Dehui; Sebag, Julien; Corbin, Jackie D.; Weaver, C. David; Cone, Roger D.

    2011-01-01

    The melanocortin MC4 receptor is a potential target for the development of drugs for both obesity and cachexia. Melanocortin MC4 receptor ligands known thus far are orthosteric agonists or antagonists, however the agonists, in particular, have generally exhibited unwanted side effects. For some receptors, allosteric modulators are expected to reduce side-effect profiles. To identify allosteric modulators of the melanocortin MC4 receptor, we created HEK293 cell lines coexpressing the human melanocortin MC4 receptor and a modified luciferase-based cAMP sensor. Monitoring luminescence as a readout of real-time intracellular cAMP concentration, we demonstrate this cell line is able to report melanocortin agonist responses, as well as inverse agonist response to the physiological AgRP peptide. Based on the MC4R-GLO cell line, we developed an assay that was shown to meet HTS standards (Z’=0.50). A pilot screen run on the Microsource Spectrum compound library (n= 2,000) successfully identified 62 positive modulators. This screen identified predicted families of compounds: β2AR agonists –the β2AR being endogenously expressed in HEK293 cells-, an adenylyl cyclase activator and finally a distribution of phosphodiesterase (PDE) inhibitors well characterized or recently identified. In this last category, we identified a structural family of coumarin-derived compounds (imperatorin, osthol and prenyletin), along with deracoxib, a drug in veterinary use for its COX2 inhibitory properties. This latter finding unveiled a new off-target mechanism of action for deracoxib as a PDE inhibitor. Overall, these data are the first report of an HTS for allosteric modulators for a Gs protein coupled receptor. PMID:21296065

  4. Human phosphodiesterase 4D7 (PDE4D7) expression is increased in TMPRSS2-ERG-positive primary prostate cancer and independently adds to a reduced risk of post-surgical disease progression

    PubMed Central

    Böttcher, R; Henderson, D J P; Dulla, K; van Strijp, D; Waanders, L F; Tevz, G; Lehman, M L; Merkle, D; van Leenders, G J L H; Baillie, G S; Jenster, G; Houslay, M D; Hoffmann, R

    2015-01-01

    Background: There is an acute need to uncover biomarkers that reflect the molecular pathologies, underpinning prostate cancer progression and poor patient outcome. We have previously demonstrated that in prostate cancer cell lines PDE4D7 is downregulated in advanced cases of the disease. To investigate further the prognostic power of PDE4D7 expression during prostate cancer progression and assess how downregulation of this PDE isoform may affect disease outcome, we have examined PDE4D7 expression in physiologically relevant primary human samples. Methods: About 1405 patient samples across 8 publically available qPCR, Affymetrix Exon 1.0 ST arrays and RNA sequencing data sets were screened for PDE4D7 expression. The TMPRSS2-ERG gene rearrangement status of patient samples was determined by transformation of the exon array and RNA seq expression data to robust z-scores followed by the application of a threshold >3 to define a positive TMPRSS2-ERG gene fusion event in a tumour sample. Results: We demonstrate that PDE4D7 expression positively correlates with primary tumour development. We also show a positive association with the highly prostate cancer-specific gene rearrangement between TMPRSS2 and the ETS transcription factor family member ERG. In addition, we find that in primary TMPRSS2-ERG-positive tumours PDE4D7 expression is significantly positively correlated with low-grade disease and a reduced likelihood of progression after primary treatment. Conversely, PDE4D7 transcript levels become significantly decreased in castration resistant prostate cancer (CRPC). Conclusions: We further characterise and add physiological relevance to PDE4D7 as a novel marker that is associated with the development and progression of prostate tumours. We propose that the assessment of PDE4D7 levels may provide a novel, independent predictor of post-surgical disease progression. PMID:26575822

  5. Camp for Youth With Type 1 Diabetes.

    PubMed

    Fegan-Bohm, Kelly; Weissberg-Benchell, Jill; DeSalvo, Daniel; Gunn, Sheila; Hilliard, Marisa

    2016-08-01

    Camps for youth with type 1 diabetes (T1D) have grown in size and scope since they first emerged in the 1920s. Anecdotal evidence suggests that attending camp with other youth with T1D is beneficial, largely attributed to sharing fun, active experiences and removing the isolation of living with diabetes. However, few studies have evaluated the psychosocial and medical impacts of T1D camp attendance during and after camp sessions. In addition, T1D camps have been a setting for numerous studies on a variety of T1D-related research questions not related to camp itself, such as testing novel diabetes management technologies in an active, non-laboratory setting. This paper reviews the evidence of psychosocial and medical outcomes associated with T1D camp attendance across the globe, provides an overview of other research conducted at camp, and offers recommendations for future research conducted at T1D camp. PMID:27292106

  6. Identification of two splice variant forms of type-IVB cyclic AMP phosphodiesterase, DPD (rPDE-IVB1) and PDE-4 (rPDE-IVB2) in brain: selective localization in membrane and cytosolic compartments and differential expression in various brain regions.

    PubMed

    Lobban, M; Shakur, Y; Beattie, J; Houslay, M D

    1994-12-01

    In order to detect the two splice variant forms of type-IVB cyclic AMP phosphodiesterase (PDE) activity, DPD (type-IVB1) and PDE-4 (type-IVB2), anti-peptide antisera were generated. One set ('DPD/PDE-4-common'), generated against a peptide sequence found at the common C-terminus of these two PDEs, detected both PDEs. A second set was PDE-4 specific, being directed against a peptide sequence found within the unique N-terminal region of PDE-4. In brain, DPD was found exclusively in the cytosol and PDE-4 exclusively associated with membranes. Both brain DPD and PDE-4 activities, isolated by immunoprecipitation, were cyclic AMP-specific (KmcyclicAMP: approximately 5 microM for DPD; approximately 4 microM for PDE-4) and were inhibited by low rolipram concentrations (K1rolipram approximately 1 microM for both). Transient expression of DPD in COS-1 cells allowed identification of an approx. 64 kDa species which co-migrated on SDS/PAGE with the immunoreactive species identified in both brain cytosol and membrane fractions using the DPD/PDE-4-common antisera. The subunit size observed for PDE-4 (approx. 64 kDa) in brain membranes was similar to that predicted from the cDNA sequence, but that observed for DPD was approx. 4 kDa greater. Type-IV, rolipram-inhibited PDE activity was found in all brain regions except the pituitary, where it formed between 30 and 70% of the PDE activity in membrane and cytosolic fractions when assayed with 1 microM cyclic AMP, PDE-4 formed 40-50% of the membrane type-IV activity in all brain regions save the midbrain (approx. 20%). DPD distribution was highly restricted to certain regions, providing approx. 35% of the type-IV cytosolic activity in hippocampus and 13-21% in cortex, hypothalamus and striatum with no presence in brain stem, cerebellum, midbrain and pituitary. The combined type-IVB PDE activities of DPD and PDE-4 contributed approx. 10% of the total PDE activity in most brain regions except for the pituitary (zero) and the mid

  7. Treatment of Skin Inflammation with Benzoxaborole Phosphodiesterase Inhibitors: Selectivity, Cellular Activity, and Effect on Cytokines Associated with Skin Inflammation and Skin Architecture Changes.

    PubMed

    Dong, Chen; Virtucio, Charlotte; Zemska, Olga; Baltazar, Grober; Zhou, Yasheen; Baia, Diogo; Jones-Iatauro, Shannon; Sexton, Holly; Martin, Shamra; Dee, Joshua; Mak, Yvonne; Meewan, Maliwan; Rock, Fernando; Akama, Tsutomu; Jarnagin, Kurt

    2016-09-01

    Psoriasis and atopic dermatitis are skin diseases affecting millions of patients. Here, we characterize benzoxaborole phosphodiesterase (PDE)-4 inhibitors, a new topical class that has demonstrated therapeutic benefit for psoriasis and atopic dermatitis in phase 2 or phase 3 studies. Crisaborole [AN2728, 4-((1-hydroxy-1,3-dihydrobenzo[c][1,2]oxaborol-5-yl)oxy)benzonitrile], compd2 [2-ethoxy-6-((1-hydroxy-1,3-dihydrobenzo[c][1,2]oxaborol-5-yl)oxy)nicotinonitrile], compd3 [6-((1-hydroxy-1,3-dihydrobenzo[c][1,2]oxaborol-5-yl)oxy)-2-(2-isopropoxyethoxy)nicotinonitrile], and compd4 [5-chloro-6-((1-hydroxy-1,3-dihydrobenzo[c][1,2]oxaborol-5-yl)oxy)-2-((4-oxopentyl)oxy)nicotinonitrile] are potent PDE4 inhibitors with similar affinity for PDE4 isoforms and equivalent inhibition on the catalytic domain and the full-length enzyme. These benzoxaboroles are less active on other PDE isozymes. Compd4 binds to the catalytic domain of PDE4B2 with the oxaborole group chelating the catalytic bimetal and overlapping with the phosphate in cAMP during substrate hydrolysis, and the interaction extends into the adenine pocket. In cell culture, benzoxaborole PDE4 inhibitors suppress the release of tumor necrosis factor-α, interleukin (IL)-23, IL-17, interferon-γ, IL-4, IL-5, IL-13, and IL-22, and these cytokines contribute to the pathologic changes in skin structure and barrier functions as well as immune dysregulation in atopic dermatitis and psoriasis. Treatment with compd3 or N(6),2'-O-dibutyryladenosine 3',5'-cyclic monophosphate increases cAMP response element binding protein phosphorylation in human monocytes and decreases extracellular signal-regulated kinase phosphorylation in human T cells; these changes lead to reduced cytokine production and are among the mechanisms by which compd3 blocks cytokine release. Topical compd3 penetrates the skin and suppresses phorbol myristate acetate-induced IL-13, IL-22, IL-17F, and IL-23 transcription and calcipotriol-induced thymic stromal

  8. Hydromania: Summer Science Camp Curriculum.

    SciTech Connect

    Moura, Joan

    1995-07-01

    In 1992, Bonneville Power Administration (BPA) and the US Department of Energy (DOE) began a collaborative pilot project with the Portland Parks and Recreation Community Schools Program and others to provide summer science camps to children in Grades 4--6. Camps run two weeks in duration between late June and mid-August. Sessions are five days per week, from 9 a.m. to 3 p.m. In addition to hands-on science and math curriculum, at least three field trips are incorporated into the educational learning experience. The purpose of the BPA/DOE summer camps is to make available opportunities for fun, motivating experiences in science to students who otherwise would have difficulty accessing them. This includes inner city, minority, rural and low income students. Public law 101-510, which Congress passed in 1990, authorizes DOE facilities to establish collaborative inner-city and rural partnership programs in science and math. A primary goal of the BPA summer hands on science camps is to bring affordable science camp experiences to students where they live. It uses everyday materials to engage students` minds and to give them a sense that they have succeeded through a fun hands-on learning environment.

  9. Cultural behaviour and the invention of traditions: music and musical practices in the early concentration camps, 1933-6/7.

    PubMed

    Fackler, Guido

    2010-01-01

    This article investigates music in the concentration camps before the second world war. For the camp authorities, ordering prisoners to sing songs or play in orchestras was an instrument of domination. But for the prisoners, music could also be an expression of solidarity and survival: inmates could retain a degree of their own agency in the pre-war camps, despite the often unbearable living conditions and harsh treatment by guards. The present article emphasizes this ambiguity of music in the early camps. It illustrates the emergence of musical traditions in the pre-war camps which came to have a significant impact on everyday life in the camps. It helps to overcome the view that concentration camp prisoners were simply passive victims. PMID:20845575

  10. He Sapa Bloketu Waecun: 2008 Summer Science and Cultural Camps

    NASA Astrophysics Data System (ADS)

    Kliche, D. V.; Sanovia, J.; Decker, R.; Bolman, J.

    2008-12-01

    The South Dakota School of Mines, Humboldt State University and Sinte Gleska University with support from the National Science Foundation, sponsored four camps for South Dakota Lakota youth to nurture a geosciences learning community linked to culturally significant sites in the Black Hills. These camps utilized outdoor, experiential learning to integrate indigenous knowledge with contemporary western science. The project resulted in increased awareness among Native and non-Native Americans, young and adult, about the importance of geosciences in their connection and interpretation of nature. The project also motivated participants in learning and becoming active in land and resources protection and the importance of becoming knowledgeable and active in regulatory policies (both Tribal and State). The four camps were scheduled during the month of June, 2008, which is the month of the summer solstice, a sacred time for the Lakota people which signal the Lakota Sundance Ceremony. The timing of the camps was chosen to give the Native American participants the framework to express their connection to Native lands through the understanding of their oral history. For the first time in such camps, middle and high school students were encouraged to have a parent or relative attending with them. The camps proved to be a great success among students and their families. The curriculum and activities helped participants immerse themselves mentally, physically and spiritually into an experience of a life time. We plan to show our results from these camps and emphasize the usefulness of this new approach in teaching science and encouraging the new generation to pursue careers in geosciences.

  11. Academic Boot Camp for the Writing of Psychology Research Reports

    ERIC Educational Resources Information Center

    Skues, Jason L.; Wise, Lisa

    2014-01-01

    Herein, we describe the implementation of, and responses to, a structured writing workshop in the form of an academic boot camp. Participants were 42 undergraduate psychology students from a medium-sized Australian university who were completing their major assignment for the semester. A majority of the students expressed satisfaction with the…

  12. Cytochemical demonstration of cyclic 3', 5' AMP phosphodiesterase in different tissue of migratory locust (Locusta migratoria migratorioides R.F.).

    PubMed

    Benedeczky, I; Rózsa, K S

    1981-01-01

    The localization of cyclic 3', 5'-AMP phosphodiesterase was studied by ultrastructural cytochemical methods in the various tissues of Locusta migratoria. Large amounts of electron dense, granular reaction products were detectable on the surface of the corpora allata and the corpus cardiacum, bound to the basal lamina. In the protocerebral neuropile rather large amounts of reaction products were observed in the processes of the glial cells. A significant number of lead phosphate deposit was found to occur on the membrane of certain large axons, the microtubules of the axons, furthermore on the membrane of several terminals. Reaction product was also observable in certain terminals, bound to the synaptic vesicles and the mitochondria. At the same time, electron dense deposits were not detectable at all on the surface of cerebral neurons. In the case of myocardium, reaction product was only found on the basal lamina and the extracellular surface of the sarcolemma. On the basis of our results it can be stated that the cyclic 3', 5'-AMP phosphodiesterase is detectable by cytochemical methods in different tissues of Locusta migratoria and presumably it fulfils the task of the extracellular cAMP level regulation. PMID:6260713

  13. Youth development and the camp experience.

    PubMed

    Garst, Barry A; Browne, Laurie P; Bialeschki, M Deborah

    2011-01-01

    The organized camp experience has been an important part of the lives of children, youth, and adults for over 150 years. The camp experience is a way for young people to explore and search for an authenticity often missing in other parts of their lives that contributes to their healthy transition into adulthood. Over the past decade, tremendous growth in the volume and rigor of camp-related research has occurred, facilitated by a targeted research agenda conducted by the American Camp Association. This agenda was founded on three national research projects conducted between 2003 and 2007: a study to identify the developmental outcomes of the camp experience, a benchmarking study of the youth development supports and opportunities provided through camp experiences, and a program improvement project directed toward enhancing supports and opportunities provided by camps. The findings from these research projects suggest that camp experiences promote developmental outcomes in both campers and staff and that camps provide the supports and opportunities needed for positive youth development. This article explores the developmental outcomes of the camp experience and the characteristics of the supports and opportunities afforded by camp experiences, including settings, structures, and programs and activities, as a way to provide a clearer understanding of camp as a positive youth development setting. Innovations and opportunities in research related to the provision of quality camp experiences are also considered. PMID:21786411

  14. Current use of phosphodiesterase inhibitors in urology

    PubMed Central

    Hakky, Tariq Said; Jain, Lakshay

    2015-01-01

    The causes of male erectile dysfunction (ED) are quite variable and are now commonly divided into etiologies such as ischemia, smooth muscle damage, or altered blood flow. Although varying rates of ED have been reported in literature, the number of men with ED is projected to increase worldwide by 2025 to approximately 322 million. Since the introduction of phosphodiesterase 5 (PDE5) inhibitors, there has been a paradigm shift in the treatment of ED because PDE5 inhibitors address a broad spectrum of etiologies for ED. Today, the American Urological Association recommends the use of three PDE5 inhibitors (sildenafil, tadalafil, and vardenafil) as a first-line therapy for the treatment of ED. This review evaluates the pharmacological mechanism of PDE5 inhibitors along with the impact and use of sildenafil, vardenafil, tadalafil, and avanafil. By increasing intracellular cGMP levels, PDE5 inhibitors have been shown to be effective in the treatment of ED. Through their effects on other cellular signaling pathways, PDE5 inhibitors have the potential for treating other urologic conditions as well. The use of PDE5 inhibitors can also be combined to produce a synergistic effect in conditions such as male hypogonadism and benign prostatic hyperplasia in addition to ED. PMID:26328208

  15. Phosphodiesterase-5 Inhibitors: Action on the Signaling Pathways of Neuroinflammation, Neurodegeneration, and Cognition

    PubMed Central

    Peixoto, Christina Alves; Nunes, Ana Karolina Santana; Garcia-Osta, Ana

    2015-01-01

    Phosphodiesterase type 5 inhibitors (PDE5-Is) have recently emerged as a potential therapeutic strategy for neuroinflammatory, neurodegenerative, and memory loss diseases. Mechanistically, PDE5-Is produce an anti-inflammatory and neuroprotection effect by increasing expression of nitric oxide synthases and accumulation of cGMP and activating protein kinase G (PKG), the signaling pathway of which is thought to play an important role in the development of several neurodiseases, such as Alzheimer's disease (AD), Parkinson's disease (PD), and multiple sclerosis (MS). The aim of this paper was to review present knowledge of the signaling pathways that underlie the use of PDE5-Is in neuroinflammation, neurogenesis, learning, and memory. PMID:26770022

  16. Phosphodiesterase 4 Inhibitors Attenuate the Asthma Phenotype Produced by β2-Adrenoceptor Agonists in Phenylethanolamine N-Methyltransferase-Knockout Mice.

    PubMed

    Forkuo, Gloria S; Kim, Hosu; Thanawala, Vaidehi J; Al-Sawalha, Nour; Valdez, Daniel; Joshi, Radhika; Parra, Sergio; Pera, Tonio; Gonnella, Patricia A; Knoll, Brian J; Walker, Julia K L; Penn, Raymond B; Bond, Richard A

    2016-08-01

    Mice lacking the endogenous β2-adrenoceptor (β2AR) agonist epinephrine (phenylethanolamine N-methyltransferase [PNMT]-knockout mice) are resistant to developing an "asthma-like" phenotype in an ovalbumin sensitization and challenge (Ova S/C) model, and chronic administration of β2AR agonists to PNMT-KO mice restores the phenotype. Based on these and other studies showing differential effects of various β2AR ligands on the asthma phenotype, we have speculated that the permissive effect of endogenous epinephrine and exogenous β2AR agonists on allergic lung inflammation can be explained by qualitative β2AR signaling. The β2AR can signal through at least two pathways: the canonical Gαs-cAMP pathway and a β-arrestin-dependent pathway. Previous studies suggest that β-arrestin-2 is required for allergic lung inflammation. On the other hand, cell-based assays suggest antiinflammatory effects of Gαs-cAMP signaling. This study was designed to test whether the in vitro antiinflammatory effects of phosphodiesterase 4 inhibitors, known to increase intracellular cAMP in multiple airway cell types, attenuate the asthma-like phenotype produced by the β2AR agonists formoterol and salmeterol in vivo in PNMT-KO mice, based on the hypothesis that skewing β2AR signaling toward Gαs-cAMP pathway is beneficial. Airway inflammatory cells, epithelial mucus production, and airway hyperresponsiveness were quantified. In Ova S/C PNMT-KO mice, formoterol and salmeterol restored the asthma-like phenotype comparable to Ova S/C wild-type mice. However, coadministration of either roflumilast or rolipram attenuated this formoterol- or salmeterol-driven phenotype in Ova S/C PNMT-KO. These findings suggest that amplification of β2AR-mediated cAMP by phosphodiesterase 4 inhibitors attenuates the asthma-like phenotype promoted by β-agonists. PMID:26909542

  17. The Popeye Domain Containing Genes and cAMP Signaling

    PubMed Central

    Brand, Thomas; Poon, Kar Lai; Simrick, Subreena; Schindler, Roland F.R.

    2016-01-01

    3'-5'-cyclic adenosine monophosphate (cAMP) is a second messenger, which plays an important role in the heart. It is generated in response to activation of G-protein-coupled receptors (GPCRs). Initially, it was thought that protein kinase A (PKA) exclusively mediates cAMP-induced cellular responses such as an increase in cardiac contractility, relaxation, and heart rate. With the identification of the exchange factor directly activated by cAMP (EPAC) and hyperpolarizing cyclic nucleotide-gated (HCN) channels as cAMP effector proteins it became clear that a protein network is involved in cAMP signaling. The Popeye domain containing (Popdc) genes encode yet another family of cAMP-binding proteins, which are prominently expressed in the heart. Loss-of-function mutations in mice are associated with cardiac arrhythmia and impaired skeletal muscle regeneration. Interestingly, the cardiac phenotype, which is present in both, Popdc1 and Popdc2 null mutants, is characterized by a stress-induced sinus bradycardia, suggesting that Popdc proteins participate in cAMP signaling in the sinuatrial node. The identification of the two-pore channel TREK-1 and Caveolin 3 as Popdc-interacting proteins represents a first step into understanding the mechanisms of heart rate modulation triggered by Popdc proteins. PMID:27500161

  18. Going Camping? A Basic Guide to Camping with Students.

    ERIC Educational Resources Information Center

    1977

    Beginning with the "often overlooked areas", this guide to camping trips for secondary students presents initial planning procedures such as: site selection; number of leaders per number of students (2 leaders for each group of 10-12); parental permission forms; transportation arrangements; public school announcements; medical aid expertise and…

  19. Making Camp Environmentally Friendly: How Two Camps Did It.

    ERIC Educational Resources Information Center

    Westerman, Martin; Griner-Johnson, Russ

    1991-01-01

    Describes the efforts of two camps administered by the Brandeis-Bardin Institute (Brandeis, California) in implementing water and energy conservation programs, involving recycling, composting, and landfill savings. Programs were successful in eliminating excess waste and teaching campers to care more about their environments at home and at work.…

  20. Starting a Technology Camp for Children.

    ERIC Educational Resources Information Center

    Litowitz, Len S.; Baylor, Steven C.

    1997-01-01

    Presents information for starting and maintaining a technology camp for students. Includes selecting topics, identifying participants, marketing, managing the camp, budgeting, and benefits to students and host institutions. (JOW)

  1. Reading Camps for Teenagers in Hungary.

    ERIC Educational Resources Information Center

    Nagy, Attila

    1991-01-01

    Describes reading camps in Hungary, which numbered around 140 in the late 1980s and lasted from 10 to 14 days each summer. Notes that these camps arouse interest through a series of conversations between equal partners. (RS)

  2. Upregulation of Phosphodiesterase type 5 in the Hyperplastic Prostate

    PubMed Central

    Zhang, Wenhao; Zang, Ning; Jiang, Yaoming; Chen, Ping; Wang, Xinghuan; Zhang, Xinhua

    2015-01-01

    Both erectile dysfunction (ED) and lower urinary tract symptoms (LUTS)/benign prostatic hyperplasia (BPH) are common in the aging male. Numerous clinical trials have demonstrated the efficacy and safety of phosphodiesterase type 5 inhibitors (PDE5-Is) for treating LUTS/BPH with/without ED. However, the influence of BPH on prostatic PDE5 expression has never been studied. A testosterone-induced rat model of BPH was developed and human hyperplastic prostate specimens were harvested during cystoprostatectomy. PDE5, nNOS, eNOS and α1-adrenoreceptor subtypes (α1aARs, α1bARs and α1dARs) were determined with real-time RT-PCR for rat tissues whilst PDE5 and α1-adrenoreceptor subtypes were determined in human samples. PDE5 was further analyzed with Western-blot and histological examination. Serum testosterone was measured with ELISA. The rat BPH model was validated as having a significantly enlarged prostate. PDE5 localized mainly in fibromuscular stroma in prostate. Our data showed a significant and previously undocumented upregulation of PDE5 in both rat and human BPH, along with increased expression of nNOS and α1dARs for rat tissues and α1aARs for human BPH. The upregulation of PDE5 in the hyperplastic prostate could explain the mechanism and contribute to the high effectiveness of PDE5-Is for treating LUTS/BPH. Fibromuscular stroma could be the main target for PDE5-Is within prostate. PMID:26657792

  3. Including People with Disabilities in Camp Programs: A Resource for Camp Directors.

    ERIC Educational Resources Information Center

    Roswal, Glenn M., Ed.; Dowd, Karen J., Ed.; Bynum, Jerry W., Ed.

    Written primarily by camp administrators affiliated with the National Easter Seal Society, this publication is designed to help camp directors meet the challenges of including campers of all abilities in their camp programs. The first section provides an overview of the inclusion concept in general and at camp, and discusses legal and medical…

  4. Function and Protective Capacity of Treponema pallidum subsp. pallidum Glycerophosphodiester Phosphodiesterase

    PubMed Central

    Cameron, Caroline E.; Castro, Christa; Lukehart, Sheila A.; Van Voorhis, Wesley C.

    1998-01-01

    Infectious syphilis, caused by the spirochete bacterium Treponema pallidum subsp. pallidum, remains a public health concern worldwide. The immune-response evasion mechanisms employed by T. pallidum are poorly understood, and prior attempts to identify immunoprotective antigens for subsequent vaccine design have been unsuccessful. Previous investigations conducted in our laboratory identified the T. pallidum glycerophosphodiester phosphodiesterase as a potential immunoprotective antigen by using a differential immunologic expression library screen. In studies reported here, heterologous expression of the T. pallidum glycerophosphodiester phosphodiesterase in Escherichia coli yielded a full-length, enzymatically active protein. Characterization of the recombinant molecule showed it to be bifunctional, in that it exhibited specific binding to human immunoglobulin A (IgA), IgD, and IgG in addition to possessing enzymatic activity. IgG fractionation studies revealed specific binding of the recombinant enzyme to the Fc fragment of human IgG, a characteristic that may play a role in enabling the syphilis spirochete to evade the host immune response. In further investigations, immunization with the recombinant enzyme significantly protected rabbits from subsequent T. pallidum challenge, altering lesion development at the sites of challenge. In all cases, animals immunized with the recombinant molecule developed atypical pale, flat, slightly indurated, and nonulcerative reactions at the challenge sites that resolved before lesions appeared in the control animals. Although protection in the immunized rabbits was incomplete, as demonstrated by the presence of T. pallidum in the rabbit infectivity test, glycerophosphodiester phosphodiesterase nevertheless represents a significantly immunoprotective T. pallidum antigen and thus may be useful for inclusion in an antigen cocktail vaccine for syphilis. PMID:9826352

  5. Function and protective capacity of Treponema pallidum subsp. pallidum glycerophosphodiester phosphodiesterase.

    PubMed

    Cameron, C E; Castro, C; Lukehart, S A; Van Voorhis, W C

    1998-12-01

    Infectious syphilis, caused by the spirochete bacterium Treponema pallidum subsp. pallidum, remains a public health concern worldwide. The immune-response evasion mechanisms employed by T. pallidum are poorly understood, and prior attempts to identify immunoprotective antigens for subsequent vaccine design have been unsuccessful. Previous investigations conducted in our laboratory identified the T. pallidum glycerophosphodiester phosphodiesterase as a potential immunoprotective antigen by using a differential immunologic expression library screen. In studies reported here, heterologous expression of the T. pallidum glycerophosphodiester phosphodiesterase in Escherichia coli yielded a full-length, enzymatically active protein. Characterization of the recombinant molecule showed it to be bifunctional, in that it exhibited specific binding to human immunoglobulin A (IgA), IgD, and IgG in addition to possessing enzymatic activity. IgG fractionation studies revealed specific binding of the recombinant enzyme to the Fc fragment of human IgG, a characteristic that may play a role in enabling the syphilis spirochete to evade the host immune response. In further investigations, immunization with the recombinant enzyme significantly protected rabbits from subsequent T. pallidum challenge, altering lesion development at the sites of challenge. In all cases, animals immunized with the recombinant molecule developed atypical pale, flat, slightly indurated, and nonulcerative reactions at the challenge sites that resolved before lesions appeared in the control animals. Although protection in the immunized rabbits was incomplete, as demonstrated by the presence of T. pallidum in the rabbit infectivity test, glycerophosphodiester phosphodiesterase nevertheless represents a significantly immunoprotective T. pallidum antigen and thus may be useful for inclusion in an antigen cocktail vaccine for syphilis. PMID:9826352

  6. Inhibition of phosphodiesterase 2 reverses impaired cognition and neuronal remodeling caused by chronic stress.

    PubMed

    Xu, Ying; Pan, Jianchun; Sun, Jiao; Ding, Lianshu; Ruan, Lina; Reed, Miranda; Yu, Xuefeng; Klabnik, Jonathan; Lin, Dan; Li, Jianxin; Chen, Ling; Zhang, Chong; Zhang, Hanting; O'Donnell, James M

    2015-02-01

    Chronic stress and neuronal vulnerability have recently been recognized as factors contributing to cognitive disorders. One way to modify neuronal vulnerability is through mediation of phosphodiesterase 2 (PDE2), an enzyme that exerts its action on cognitive processes via the control of intracellular second messengers, cGMP and, to a lesser extent, cAMP. This study explored the effects of a PDE2 inhibitor, Bay 60-7550, on stress-induced learning and memory dysfunction in terms of its ramification on behavioral, morphologic, and molecular changes. Bay 60-7550 reversed stress-induced cognitive impairment in the Morris water maze, novel object recognition, and location tasks (object recognition test and/or object location test), effects prevented by treatment with 7-NI, a selective inhibitor of neuronal nitric oxide synthase; MK801, a glutamate receptor (NMDAR) inhibitor; myr-AIP, a CaMKII inhibitor; and KT5823, a protein kinase G inhibitor. Bay 60-7550 also ameliorated stress-induced structural remodeling in the CA1 of the hippocampus, leading to increases in dendritic branching, length, and spine density. However, the neuroplasticity initiated by Bay 60-7550 was not seen in the presence of 7-NI, MK801, myr-AIP, or KT5823. PDE2 inhibition reduced stress-induced extracellular-regulated protein kinase activation and attenuated stress-induced decreases in transcription factors (e.g., Elk-1, TORC1, and CREB phosphorylation) and plasticity-related proteins (e.g., Egr-1 and brain-derived neurotrophic factor). Pretreatment with inhibitors of NMDA, CaMKII, neuronal nitric oxide synthase, and protein kinase G (or protein kinase A) blocked the effects of Bay 60-7550 on cGMP or cAMP signaling. These findings indicate that the effect of PDE2 inhibition on stress-induced memory impairment is potentially mediated via modulation of neuroplasticity-related NMDAR-CaMKII-cGMP/cAMP signaling. PMID:25442113

  7. Attenuation by phosphodiesterase inhibitors of lipopolysaccharide-induced thromboxane release and bronchoconstriction in rat lungs.

    PubMed

    Uhlig, S; Featherstone, R L; Held, H D; Nüsing, R; Schudt, C; Wendel, A

    1997-12-01

    Exposure of perfused rat lungs to lipopolysaccharides (LPS) causes induction of cyclooxygenase-2 followed by thromboxane (TX)-mediated bronchoconstriction (BC). Recently, phosphodiesterase (PDE) inhibitors have received much interest because they not only are bronchodilators but also can suppress release of proinflammatory mediators. In the present study, we investigated the effect of three different PDE inhibitors on TX release and BC in LPS-exposed perfused rat lungs. The PDE inhibitors used were motapizone (PDE III specific), rolipram (PDE IV specific), and zardaverine (mixed PDE III and IV specific). At 5 microM, a concentration at which all three compounds selectively block their respective PDE isoenzyme, rolipram (IC50 = 0.04 microM) and zardaverine (IC50 = 1.8 microM) largely attenuated the LPS-induced BC, whereas motapizone was almost ineffective (IC50 = 40 microM). In contrast to LPS, BC induced by the TX-mimetic U46619 was prevented with comparable strength by motapizone and rolipram. In LPS-treated lungs, the TX release was reduced to 50% of controls by rolipram and zardaverine but was unaltered in the presence of 5 microM motapizone. Increasing intracellular cAMP through perfusion of db-cAMP or forskolin (activates adenylate cyclase) also reduced TX release and BC. We conclude that PDE inhibitors act via elevation of intracellular cAMP. Although both PDE III and PDE IV inhibitors can relax airway smooth muscle, in the model of LPS-induced BC, PDE IV inhibitors are more effective because (in contrast to PDE III inhibitors) they also attenuate TX release. PMID:9400021

  8. Phosphodiesterase 2 and 5 inhibition attenuates the object memory deficit induced by acute tryptophan depletion.

    PubMed

    van Donkelaar, Eva L; Rutten, Kris; Blokland, Arjan; Akkerman, Sven; Steinbusch, Harry W M; Prickaerts, Jos

    2008-12-14

    The underlying mechanism of short-term memory improvement after inhibition of specific phosphodiesterases (PDEs) is still poorly understood. The present study aimed to reveal the ability of PDE5 and PDE2 inhibitors, that increase cyclic guanosine monophosphate (cGMP) and both cyclic adenosine monophosphate (cAMP) and cGMP, respectively, to reverse an object recognition deficit induced by acute tryptophan depletion. Acute tryptophan depletion is a pharmacological challenge tool to lower central serotonin (5-hydroxytryptamine; 5-HT) levels by depleting the availability of its dietary precursor tryptophan. Short-term object memory was tested in male Wistar rats by exposing them to the object recognition task. First, the effects of acute tryptophan depletion upon object recognition 2 h after administration of the nutritional mixture were established. Subsequently, acute tryptophan depletion was combined with the PDE5 inhibitor vardenafil (1, 3 and 10 mg/kg) or with the PDE2 inhibitor BAY 60-7550 (0.3, 1 and 3 mg/kg), 30 min prior to testing. Acute tryptophan depletion significantly lowered plasma tryptophan levels and impaired object recognition performance. Vardenafil (3 and 10 mg/kg) and BAY 60-7550 (3 mg/kg) were able to attenuate the acute tryptophan depletion induced object recognition impairment. Thus, both PDE5 and PDE2 inhibition improved short-term object recognition performance after an acute tryptophan depletion induced deficit. The underlying mechanisms, however, remain poorly understood and further studies are needed to determine whether the present findings can be explained by a direct effect of enhanced cAMP and cGMP levels upon 5-HT activity, or even other neurotransmitter systems, and possibly an interaction with synthesis of nitric oxide or effects upon cerebral blood flow function. PMID:18957291

  9. Inhibition of phosphodiesterase 2 reverses impaired cognition and neuronal remodeling caused by chronic stress

    PubMed Central

    Xu, Ying; Pan, Jianchun; Sun, Jiao; Ding, Lianshu; Ruan, Lina; Reed, Miranda; Yu, Xuefeng; Klabni, Jonathan; Lin, Dan; Li, Jianxin; Chen, Ling; Zhang, Chong; Zhang, Hanting; O’Donnell, James M.

    2014-01-01

    Chronic stress and neuronal vulnerability have recently been recognized as factors contributing to cognitive disorders. One way to modify neuronal vulnerability is through mediation of phosphodiesterase 2 (PDE2), an enzyme that exerts its action on cognitive processes via the control of intracellular second messengers, cGMP and, to a lesser extent, cAMP. This study explored the effects of a PDE2 inhibitor, Bay 60-7550, on stress-induced learning and memory dysfunction in terms of its ramification on behavioral, morphological and molecular changes. Bay 60-7550 reversed stress-induced cognitive impairment in the Morris water maze (MWM), novel object recognition and location tasks (ORT/OLT), effects prevented by treatment with 7-NI, a selective inhibitor of neuronal nitric oxide synthase (nNOS); MK801, a glutamate receptor (NMDAR) inhibitor; myr-AIP, a CaMKII inhibitor; and KT5823, a PKG inhibitor. Bay 60-7550 also ameliorated stress-induced structural remodeling in the CA1 of the hippocampus, leading to increases in dendritic branching, length, and spine density. However, the neuroplasticity initiated by Bay 60-7550 was not seen in the presence of 7-NI, MK801, myr-AIP or KT5823. PDE2 inhibition reduced stress-induced ERK activation and attenuated stress-induced decreases in transcription factors (e.g., Elk-1, TORC1, and pCREB) and plasticity-related proteins (e.g, Egr-1 and BDNF). Pre-treatment with inhibitors of NMDA, CaMKII, nNOS, PKG (or PKA), blocked the effects of Bay 60-7550 on cGMP or cAMP signaling. These findings indicate that the effect of PDE2 inhibition on stress-induced memory impairment is potentially mediated via modulation of neuroplasticity-related, NMDAR-CaMKII-cGMP/cAMP signaling. PMID:25442113

  10. Camping & the Whirl of Insurance Cycles.

    ERIC Educational Resources Information Center

    Milgrim, Darrow

    1988-01-01

    Suggests possible responses for summer camp operators facing insurance rate increases and other insurance industry changes. Examines areas of risk in summer camping and suggests general ways that camps can become more desirable to the insurance industry as "insurable groups." (TES)

  11. DIRECTORY OF CAMPS FOR THE HANDICAPPED.

    ERIC Educational Resources Information Center

    National Easter Seal Society for Crippled Children and Adults, Chicago, IL.

    ONE HUNDRED AND SEVENTY-SEVEN RESIDENT CAMPS IN THE UNITED STATES AND CANADA AND 77 DAY CAMPS IN THE UNITED STATES WHICH SERVE CHILDREN OR ADULTS WITH PHYSICAL, MENTAL, SOCIAL, AND EMOTIONAL HANDICAPS ARE LISTED ALPHABETICALLY BY STATE. FOR EACH CAMP, INFORMATION ON TYPES OF THE HANDICAPPED WHO ARE ACCEPTED, SPECIFIC EXCLUSIONS, AGE RANGE, NUMBER…

  12. 22 CFR 62.30 - Camp counselors.

    Code of Federal Regulations, 2014 CFR

    2014-04-01

    ... 22 Foreign Relations 1 2014-04-01 2014-04-01 false Camp counselors. 62.30 Section 62.30 Foreign... Provisions § 62.30 Camp counselors. (a) Introduction. In order to promote diverse opportunities for... sponsors to facilitate the entry of foreign nationals to serve as counselors in U.S. summer camps....

  13. 22 CFR 62.30 - Camp counselors.

    Code of Federal Regulations, 2013 CFR

    2013-04-01

    ... 22 Foreign Relations 1 2013-04-01 2013-04-01 false Camp counselors. 62.30 Section 62.30 Foreign... Provisions § 62.30 Camp counselors. (a) Introduction. In order to promote diverse opportunities for... sponsors to facilitate the entry of foreign nationals to serve as counselors in U.S. summer camps....

  14. The Elementary Camp Experience: An ARC Partnership

    ERIC Educational Resources Information Center

    Krzemien, Marta; Krotzer, Kate

    2012-01-01

    Foreign Language Elementary Summer Camp is a fantastic way to spend part of the summer. For the past ten years in Glastonbury, Connecticut, the authors have offered a variety of foreign language elementary camps that meet daily for three hours for three to four weeks during July. Currently, they offer camps for Chinese, English Language Learners…

  15. 22 CFR 62.30 - Camp counselors.

    Code of Federal Regulations, 2012 CFR

    2012-04-01

    ... 22 Foreign Relations 1 2012-04-01 2012-04-01 false Camp counselors. 62.30 Section 62.30 Foreign... Provisions § 62.30 Camp counselors. (a) Introduction. In order to promote diverse opportunities for... sponsors to facilitate the entry of foreign nationals to serve as counselors in U.S. summer camps....

  16. The Economic Impact of Organized Camps.

    ERIC Educational Resources Information Center

    Henderson, Karla A.; Bialeschki, M. Deborah

    1982-01-01

    Organized camping for youth and adults has demonstrated economic benefits for many communities. A questionnaire was sent to various agencies and private individuals to collect information on the economic feasibility of organized camps. The data indicated that 300 organized camps in Wisconsin contributed approximately $51 million to the state's…

  17. Language Camps: Thirteen Years of Minor Miracles.

    ERIC Educational Resources Information Center

    Peterson, Jean

    A language camp program that began with a small group of 10- to 12-year-olds whose faculty parents wanted them to retain the German learned on sabbaticals abroad has developed into a program of annual week-long day and resident camps for 150 children, aged 9 to 14 years, learning German, French, Spanish, and Norwegian. The camp was originally…

  18. 36 CFR 13.1208 - Lake Camp.

    Code of Federal Regulations, 2010 CFR

    2010-07-01

    ... 36 Parks, Forests, and Public Property 1 2010-07-01 2010-07-01 false Lake Camp. 13.1208 Section 13... § 13.1208 Lake Camp. Leaving a boat, trailer, or vehicle unattended for more than 72 hours at the facilities associated with the Lake Camp launching ramp is prohibited without authorization from...

  19. Extension Sustainability Camp: Design, Implementation, and Evaluation

    ERIC Educational Resources Information Center

    Brain, Roslynn; Upton, Sally; Tingey, Brett

    2015-01-01

    Sustainability Camps provide an opportunity for Extension educators to be in the forefront of sustainability outreach and to meet the growing demand for sustainability education. This article shares development, implementation, and evaluation of an Extension Sustainability Camp for youth, grades 4-6. Camp impact was measured via daily pre-and…

  20. 36 CFR 13.1208 - Lake Camp.

    Code of Federal Regulations, 2011 CFR

    2011-07-01

    ... 36 Parks, Forests, and Public Property 1 2011-07-01 2011-07-01 false Lake Camp. 13.1208 Section 13... § 13.1208 Lake Camp. Leaving a boat, trailer, or vehicle unattended for more than 72 hours at the facilities associated with the Lake Camp launching ramp is prohibited without authorization from...

  1. Day Camp Manual: Program. Book IV.

    ERIC Educational Resources Information Center

    Babcock, William

    Book IV in a 5-book day camp manual discusses the camp program. Section I describes the organization, definition, and elements essential to successful day camp programs. Section II, which addresses the benefits and special considerations of mass programs, includes rainy day contingencies, materials to have on hand, and activity suggestions.…

  2. Camping for Youth with Chronic Illnesses.

    ERIC Educational Resources Information Center

    Burns, Joanna L.; Keller, M. Jean

    1994-01-01

    Camp Fortnight brought 25 British children with cystic fibrosis to experience a 2-week camping program in Texas. Campers (ages 11-15) participated in wilderness experiences, a challenge course, fishing, horseback riding, creative arts, cooking, hiking, outdoor camping, and field trips. Profiles campers and their experiences. (LP)

  3. Winter Wilderness Travel and Camping.

    ERIC Educational Resources Information Center

    Gilchrest, Norman

    Knowledge and skill are needed for safe and enjoyable travel and camping in the wilderness in winter. The beauty of snow and ice, reduced human use, and higher tolerance of animals toward humans make the wilderness attractive during winter. The uniqueness of winter travel presents several challenges that are not present in other seasons. Safety is…

  4. Applications of Research to Camping.

    ERIC Educational Resources Information Center

    Henderson, Karla A.

    1987-01-01

    Considers contributions of basic/theoretical, applied/practical, and marketing research to the field of camping. Outlines research concerns: application of qualitative methods, practical application of marketing research, effective research dissemination, and focus on longitudinal studies using larger samples. Affirms role of research to document…

  5. Lyme Disease Comes to Camp.

    ERIC Educational Resources Information Center

    Peterson, Michael

    1989-01-01

    Describes one summer camp's plan for dealing with Lyme disease. Describes the disease and the deer tick. Recommends avoiding tick exposure through clothing, frequent examination, showers, and avoiding high grass and brushy areas, and using chemical insect repellents and chemicals to kill ticks in deer mouse nests. (DHP)

  6. AVTC Hosts TechnoCamp

    ERIC Educational Resources Information Center

    Miner, Brenda

    2006-01-01

    The Area Vo-Tech Center (AVTC) in Russellville, Arkansas, recently hosted its first TechnoCamp to encourage enrollment based on the aptitude and interest level of the students enrolling in the various programs. The center currently offers student enrollment in auto technology, computer engineering, cosmetology, construction technology, drafting…

  7. A Successful French Weekend Camp.

    ERIC Educational Resources Information Center

    Baughin, Judith A.

    A weekend immersion French language camp was organized by Raymond Walters College of the University of Cincinnati with the cooperation of a group of interested high school French teachers. The extensive planning and the activities of the weekend are described in detail. The most important aspects of the planning process are noted. These were (1)…

  8. Sleep At Camp: A Survey.

    ERIC Educational Resources Information Center

    Pravda, Myra

    1997-01-01

    Among 40 camp directors surveyed, the majority believed that campers get enough sleep, but that staff members and directors do not get enough sleep. Addresses how sleep deprivation can affect job performance and offers strategies for helping staff understand the importance of sleep to keep them alert and functioning in their job. Includes…

  9. cAMP induction by ouabain promotes endothelin-1 secretion via MAPK/ERK signaling in beating rabbit atria.

    PubMed

    Peng, Li-Qun; Li, Ping; Zhang, Qiu-Li; Hong, Lan; Liu, Li-Ping; Cui, Xun; Cui, Bai-Ri

    2016-01-01

    Adenosine 3',5'-cyclic monophosphate (cAMP) participates in the regulation of numerous cellular functions, including the Na(+)-K(+)-ATPase (sodium pump). Ouabain, used in the treatment of several heart diseases, is known to increase cAMP levels but its effects on the atrium are not understood. The aim of the present study was to examine the effect of ouabain on the regulation of atrial cAMP production and its roles in atrial endothelin-1 (ET-1) secretion in isolated perfused beating rabbit atria. Our results showed that ouabain (3.0 µmol/L) significantly increased atrial dynamics and cAMP levels during recovery period. The ouabain-increased atrial dynamics was blocked by KB-R7943 (3.0 µmol/L), an inhibitor for reverse mode of Na(+)-Ca(2+) exchangers (NCX), but did not by L-type Ca(2+) channel blocker nifedipine (1.0 µmol/L) or protein kinase A (PKA) selective inhibitor H-89 (3.0 µmol/L). Ouabain also enhanced atrial intracellular cAMP production in response to forskolin and theophyline (100.0 µmol/L), an inhibitor of phosphodiesterase, potentiated the ouabain-induced increase in cAMP. Ouabain and 8-Bromo-cAMP (0.5 µmol/L) markedly increased atrial ET-1 secretion, which was blocked by H-89 and by PD98059 (30 µmol/L), an inhibitor of extracellular-signal-regulated kinase (ERK) without changing ouabain-induced atrial dynamics. Our results demonstrated that ouabain increases atrial cAMP levels and promotes atrial ET-1 secretion via the mitogen-activated protein kinase (MAPK)/ERK signaling pathway. These findings may explain the development of cardiac hypertrophy in response to digitalis-like compounds. PMID:26807018

  10. cAMP induction by ouabain promotes endothelin-1 secretion via MAPK/ERK signaling in beating rabbit atria

    PubMed Central

    Peng, Li-qun; Li, Ping; Zhang, Qiu-li; Hong, Lan; Liu, Li-ping

    2016-01-01

    Adenosine 3',5'-cyclic monophosphate (cAMP) participates in the regulation of numerous cellular functions, including the Na+-K+-ATPase (sodium pump). Ouabain, used in the treatment of several heart diseases, is known to increase cAMP levels but its effects on the atrium are not understood. The aim of the present study was to examine the effect of ouabain on the regulation of atrial cAMP production and its roles in atrial endothelin-1 (ET-1) secretion in isolated perfused beating rabbit atria. Our results showed that ouabain (3.0 µmol/L) significantly increased atrial dynamics and cAMP levels during recovery period. The ouabain-increased atrial dynamics was blocked by KB-R7943 (3.0 µmol/L), an inhibitor for reverse mode of Na+-Ca2+ exchangers (NCX), but did not by L-type Ca2+ channel blocker nifedipine (1.0 µmol/L) or protein kinase A (PKA) selective inhibitor H-89 (3.0 µmol/L). Ouabain also enhanced atrial intracellular cAMP production in response to forskolin and theophyline (100.0 µmol/L), an inhibitor of phosphodiesterase, potentiated the ouabain-induced increase in cAMP. Ouabain and 8-Bromo-cAMP (0.5 µmol/L) markedly increased atrial ET-1 secretion, which was blocked by H-89 and by PD98059 (30 µmol/L), an inhibitor of extracellular-signal-regulated kinase (ERK) without changing ouabain-induced atrial dynamics. Our results demonstrated that ouabain increases atrial cAMP levels and promotes atrial ET-1 secretion via the mitogen-activated protein kinase (MAPK)/ERK signaling pathway. These findings may explain the development of cardiac hypertrophy in response to digitalis-like compounds. PMID:26807018

  11. The role of ventral striatal cAMP signaling in stress-induced behaviors

    PubMed Central

    Plattner, Florian; Hayashi, Kanehiro; Hernandez, Adan; Benavides, David R.; Tassin, Tara C.; Tan, Chunfeng; Day, Jonathan; Fina, Maggy W.; Yuen, Eunice Y.; Yan, Zhen; Goldberg, Matthew S.; Nairn, Angus C.; Greengard, Paul; Nestler, Eric J.; Taussig, Ronald; Nishi, Akinori; Houslay, Miles D.; Bibb, James A.

    2015-01-01

    The cAMP/PKA signaling cascade is a ubiquitous pathway acting downstream of multiple neuromodulators. We found that the phosphorylation of phosphodiesterase-4 (PDE4) by cyclin-dependent protein kinase 5 (Cdk5) facilitates cAMP degradation and homeostasis of cAMP/PKA signaling. In mice, loss of Cdk5 throughout the forebrain elevated cAMP levels and increased PKA activity in striatal neurons, and altered behavioral responses to acute or chronic stressors. Ventral striatum- or D1 dopamine receptor-specific conditional knockout of Cdk5, or ventral striatum infusion of a small interfering peptide that selectively targets the regulation of PDE4 by Cdk5, all produced analogical effects on stress-induced behavioral responses. Together, our results demonstrate that altering cAMP signaling in medium spiny neurons of the ventral striatum can effectively modulate stress-induced behavioral states. We propose that targeting the Cdk5 regulation of PDE4 could be a new therapeutic approach for clinical conditions associated with stress, such as depression. PMID:26192746

  12. Guanylyl cyclase and cGMP-specific phosphodiesterase participate in the acrosome reaction of starfish sperm.

    PubMed

    Kawase, Osamu; Ueno, Seiichi; Minakata, Hiroyuki; Hoshi, Motonori; Matsumoto, Midori

    2004-11-01

    In the starfish, Asterias amurensis, the cooperation of three components of the egg jelly, i.e. ARIS (acrosome reaction-inducing substance), Co-ARIS and asterosap, is responsible for inducing the acrosome reaction. Experimentally, ARIS and asterosap are sufficient for the induction. However, when sperm are treated only with asterosap, they become unresponsive to the egg jelly to undergo the reaction. In this study, we analysed the mechanism of the acrosome reaction, using sperm inactivation by asterosap as a clue. Asterosap causes a rapid and transient increase in intracellular cGMP through the activation of the asterosap receptor, a guanylyl cyclase, and causes an increase in intracellular Ca(2+). When sperm were pretreated with asterosap, the guanylyl cyclase seemed to be inactivated irreversibly by dephosphorylation. They were still responsive to ARIS but no longer to asterosap. However, in the presence of IBMX or zaprinast, inhibitors against phosphodiesterases (PDEs), they retained their capacity to undergo the acrosome reaction in response to the egg jelly or ARIS alone. IBMX and zaprinast suppressed the intracellular catabolism of cGMP, but not of cAMP. These results suggest that guanylyl cyclase and cGMP-specific, IBMX- and zaprinast-susceptible PDEs are involved in the regulation of the acrosome reaction. PMID:15751545

  13. Structures of the Four Subfamilies of Phosphodiesterase-4 Provide Insight into the Selectivity of Their Inhibitors

    SciTech Connect

    Wang, H.; Peng, M; Chen , Y; Geng, J; Robinson, H; Houslay , M; Cai, J; Ke, H

    2007-01-01

    PDE4 (phosphodiesterase-4)-selective inhibitors have attracted much attention as potential therapeutics for the treatment of both depression and major inflammatory diseases, but their practical application has been compromised by side effects. A possible cause for the side effects is that current PDE4-selective inhibitors similarly inhibit isoforms from all four PDE4 subfamilies. The development of PDE4 subfamily-selective inhibitors has been hampered by a lack of structural information. In the present study, we rectify this by providing the crystal structures of the catalytic domains of PDE4A, PDE4B and PDE4D in complex with the PDE4 inhibitor NVP 4-[8-(3-nitrophenyl)-[1,7]naphthyridin-6-yl]benzoic acid as well as the unliganded PDE4C structure. NVP binds in the same conformation to the deep cAMP substrate pocket and interacts with the same residues in each instance. However, detailed structural comparison reveals significant conformational differences. Although the active sites of PDE4B and PDE4D are mostly comparable, PDE4A shows significant displacements of the residues next to the invariant glutamine residue that is critical for substrate and inhibitor binding. PDE4C appears to be more distal from other PDE4 subfamilies, with certain key residues being disordered. Our analyses provide the first structural basis for the development of PDE4 subfamily-selective inhibitors.

  14. Pharmacology of a new cyclic nucleotide phosphodiesterase type 4 inhibitor, V11294.

    PubMed

    Gale, Donna D; Hofer, Peter; Spina, Domenico; Seeds, Esther A; Banner, Katherine H; Harrison, Selina; Douglas, Gary; Matsumoto, Tatsui; Page, Clive P; Wong, Richard H; Jordan, Stephan; Smith, Forrest; Banik, Nandini; Halushka, Perry V; Cavalla, David; Rotshteyn, Yakov; Kyle, Donald J; Burch, Ronald M; Chasin, Mark

    2003-01-01

    V11294 is a new cyclic nucleotide phosphodiesterase type 4 (PDE4) inhibitor of the rolipram class. In this report we present the pharmacological profile of V11294. V11294 inhibited PDE4 isolated from human lung with IC(50) 405 nM, compared to 3700 nM for rolipram. In contrast, V11294 inhibition of human PDE3 and PDE5 occurred only at concentrations greater than 100,000 nM. Like rolipram, V11294 inhibited PDE4D more potently than other PDE4 subtypes. V11294, when incubated with human anticoagulated whole blood in vitro, or administered to mice, caused increased cAMP concentration, consistent with inhibition of PDE4. V11294 inhibited lectin-induced proliferation and lipopolysaccharide-induced TNFalpha synthesis by human adherent monocytes in vitro and inhibited lipopolysaccharide-induced TNFalpha synthesis in mice. V11294 caused relaxation of guinea pig isolated trachea and inhibited allergen-induced bronchoconstriction and eosinophilia in guinea pigs at doses of 1 and 3 mg/kg, p.o. In ferrets, V11294 was not emetogenic at doses up to 30 mg/kg, p.o., despite plasma concentration reaching 10-fold the IC(50) for PDE4. In contrast, rolipram induced severe retching and vomiting at 10 mg/kg, p.o. In conclusion, V11294 is an orally active PDE4 inhibitor that exhibits antiinflammatory activity in vitro, and in vivo at doses that are not emetogenic. PMID:12670778

  15. Structural basis for the design of selective phosphodiesterase 4B inhibitors.

    PubMed

    Fox, David; Burgin, Alex B; Gurney, Mark E

    2014-03-01

    Phosphodiesterase-4B (PDE4B) regulates the pro-inflammatory Toll Receptor -Tumor Necrosis Factor α (TNFα) pathway in monocytes, macrophages and microglial cells. As such, it is an important, although under-exploited molecular target for anti-inflammatory drugs. This is due in part to the difficulty of developing selective PDE4B inhibitors as the amino acid sequence of the PDE4 active site is identical in all PDE4 subtypes (PDE4A-D). We show that highly selective PDE4B inhibitors can be designed by exploiting sequence differences outside the active site. Specifically, PDE4B selectivity can be achieved by capture of a C-terminal regulatory helix, now termed CR3 (Control Region 3), across the active site in a conformation that closes access by cAMP. PDE4B selectivity is driven by a single amino acid polymorphism in CR3 (Leu674 in PDE4B1 versus Gln594 in PDE4D). The reciprocal mutations in PDE4B and PDE4D cause a 70-80 fold shift in selectivity. Our structural studies show that CR3 is flexible and can adopt multiple orientations and multiple registries in the closed conformation. The new co-crystal structure with bound ligand provides a guide map for the design of PDE4B selective anti-inflammatory drugs. PMID:24361374

  16. Inhibition of phosphodiesterase-4 promotes oligodendrocyte precursor cell differentiation and enhances CNS remyelination.

    PubMed

    Syed, Yasir A; Baer, Alexandra; Hofer, Matthias P; González, Ginez A; Rundle, Jon; Myrta, Szymon; Huang, Jeffrey K; Zhao, Chao; Rossner, Moritz J; Trotter, Matthew W B; Lubec, Gert; Franklin, Robin J M; Kotter, Mark R

    2013-12-01

    The increasing effectiveness of new disease-modifying drugs that suppress disease activity in multiple sclerosis has opened up opportunities for regenerative medicines that enhance remyelination and potentially slow disease progression. Although several new targets for therapeutic enhancement of remyelination have emerged, few lend themselves readily to conventional drug development. Here, we used transcription profiling to identify mitogen-activated protein kinase (Mapk) signalling as an important regulator involved in the differentiation of oligodendrocyte progenitor cells (OPCs) into oligodendrocytes. We show in tissue culture that activation of Mapk signalling by elevation of intracellular levels of cyclic adenosine monophosphate (cAMP) using administration of either dibutyryl-cAMP or inhibitors of the cAMP-hydrolysing enzyme phosphodiesterase-4 (Pde4) enhances OPC differentiation. Finally, we demonstrate that systemic delivery of a Pde4 inhibitor leads to enhanced differentiation of OPCs within focal areas of toxin-induced demyelination and a consequent acceleration of remyelination. These data reveal a novel approach to therapeutic enhancement of remyelination amenable to pharmacological intervention and hence with significant potential for translation. PMID:24293318

  17. Inhibition of phosphodiesterase-4 promotes oligodendrocyte precursor cell differentiation and enhances CNS remyelination

    PubMed Central

    Syed, Yasir A; Baer, Alexandra; Hofer, Matthias P; González, Ginez A; Rundle, Jon; Myrta, Szymon; Huang, Jeffrey K; Zhao, Chao; Rossner, Moritz J; Trotter, Matthew W B; Lubec, Gert; Franklin, Robin J M; Kotter, Mark R

    2013-01-01

    The increasing effectiveness of new disease-modifying drugs that suppress disease activity in multiple sclerosis has opened up opportunities for regenerative medicines that enhance remyelination and potentially slow disease progression. Although several new targets for therapeutic enhancement of remyelination have emerged, few lend themselves readily to conventional drug development. Here, we used transcription profiling to identify mitogen-activated protein kinase (Mapk) signalling as an important regulator involved in the differentiation of oligodendrocyte progenitor cells (OPCs) into oligodendrocytes. We show in tissue culture that activation of Mapk signalling by elevation of intracellular levels of cyclic adenosine monophosphate (cAMP) using administration of either dibutyryl-cAMP or inhibitors of the cAMP-hydrolysing enzyme phosphodiesterase-4 (Pde4) enhances OPC differentiation. Finally, we demonstrate that systemic delivery of a Pde4 inhibitor leads to enhanced differentiation of OPCs within focal areas of toxin-induced demyelination and a consequent acceleration of remyelination. These data reveal a novel approach to therapeutic enhancement of remyelination amenable to pharmacological intervention and hence with significant potential for translation. PMID:24293318

  18. Cyclic adenosine monophosphate phosphodiesterase in brain: effect on anxiety.

    PubMed

    Beer, B; Chasin, M; Clody, D E; Vogel, J R

    1972-04-28

    Drugs that reduce anxiety may be mediated by cyclic adenosine monophosphate in the brain because (i) potent anxiety-reducing drugs are also potent inhibitors of brain phosphodiesterase activity; (ii) dibutyryl cyclic adenosine monophosphate has the ability to reduce anxiety; (iii) the methylxanthines show significant anxiety-reducing effects; (iv) theophylline and chlordiazepoxide produce additive anxiety-reducing activity; and (v) there is a significant correlation between the anxiety-reducing property of drugs and their ability to inhibit phosphodiesterase activity in the brain. PMID:4402069

  19. Phosphodiesterase type 5 inhibitors in pulmonary arterial hypertension.

    PubMed

    Montani, David; Chaumais, Marie-Camille; Savale, Laurent; Natali, Delphine; Price, Laura C; Jaïs, Xavier; Humbert, Marc; Simonneau, Gérald; Sitbon, Olivier

    2009-09-01

    Pulmonary arterial hypertension (PAH) is a rare disease characterized by vascular proliferation and remodeling, resulting in a progressive increase in pulmonary arterial resistance, right heart failure, and death. The pathogenesis of PAH is multifactorial, with endothelial cell dysfunction playing an integral role. This endothelial dysfunction is characterized by an overproduction of vasoconstrictors and proliferative factors, such as endothelin-1, and a reduction of vasodilators and antiproliferative factors, such prostacyclin and nitric oxide. Phosphodiesterase type 5 (PDE-5) is implicated in this process by inactivating cyclic guanosine monophosphate, the nitric oxide pathway second messenger. PDE-5 is abundantly expressed in lung tissue, and appears to be upregulated in PAH. Three oral PDE-5 inhibitors are available (sildenafil, tadalafil, and vardenafil) and are the recommended first-line treatment for erectile dysfunction. Experimental studies have shown the beneficial effects of PDE-5 inhibitors on pulmonary vascular remodeling and vasodilatation, justifying their investigation in PAH. Randomized clinical trials in monotherapy or combination therapy have been conducted in PAH with sildenafil and tadalafil, which are therefore currently the approved PDE-5 inhibitors in PAH treatment. Sildenafil and tadalafil significantly improve clinical status, exercise capacity, and hemodynamics of PAH patients. Combination therapy of PDE-5 inhibitors with prostacyclin analogs and endothelin receptor antagonists may be helpful in the management of PAH although further studies are needed in this area. The third PDE-5 inhibitor, vardenafil, is currently being investigated in PAH. Side effects are usually mild and transient and include headache, flushing, nasal congestion, digestive disorders, and myalgia. Mild and moderate renal or hepatic failure does not significantly affect the metabolism of PDE-5 inhibitors, whereas coadministration of bosentan decreases sildenafil and

  20. cAMP biosensors applied in molecular pharmacological studies of G protein-coupled receptors.

    PubMed

    Mathiesen, Jesper Mosolff; Vedel, Line; Bräuner-Osborne, Hans

    2013-01-01

    Cyclic adenosine monophosphate (cAMP) is a common second messenger that mediates numerous biological responses. Intracellular cAMP levels are increased by activation of G(s)-coupled G protein-coupled receptors (GPCRs) and decreased by activation of G(i)-coupled GPCRs via the adenylyl cyclase. Many end-point assays for quantifying GPCR-mediated changes in intracellular cAMP levels exist. More recently, fluorescence resonance energy transfer (FRET)-based cAMP biosensors that can quantify intracellular cAMP levels in real time have been developed. These FRET-based cAMP biosensors have been used primarily in single cell FRET microscopy to monitor and visualize changes in cAMP upon GPCR activation. Here, a similar cAMP biosensor with a more efficient mCerulean/mCitrine FRET pair is described for use in the 384-well plate format. After cloning and expression in HEK293 cells, the biosensor is characterized in the 384-well plate format and used for measuring the signaling of the G(s)-coupled β(2)-adrenergic receptor. The procedures described may be applied for other FRET-based biosensors in terms of characterization and conversion to the 384-well plate format. PMID:23374187

  1. The ACA Standards Organizer. A Tool for the Preparation of Camps for Visitation for American Camping Association Camp Accreditation and Site Approval.

    ERIC Educational Resources Information Center

    Chenery, Mary Faeth

    This comprehensive guide helps camp administrators prepare for camp accreditation and site approval visits by the American Camping Association (ACA). The introductory sections outline the purpose and administration of the standards program and provide definitions of camping terms. Standards are organized under one section for camp accreditation…

  2. The dual phosphodiesterase 3 and 4 inhibitor RPL554 stimulates CFTR and ciliary beating in primary cultures of bronchial epithelia.

    PubMed

    Turner, Mark J; Matthes, Elizabeth; Billet, Arnaud; Ferguson, Amy J; Thomas, David Y; Randell, Scott H; Ostrowski, Lawrence E; Abbott-Banner, Kathy; Hanrahan, John W

    2016-01-01

    Cystic fibrosis (CF), a genetic disease caused by mutations in the CFTR gene, is a life-limiting disease characterized by chronic bacterial airway infection and severe inflammation. Some CFTR mutants have reduced responsiveness to cAMP/PKA signaling; hence, pharmacological agents that elevate intracellular cAMP are potentially useful for the treatment of CF. By inhibiting cAMP breakdown, phosphodiesterase (PDE) inhibitors stimulate CFTR in vitro and in vivo. Here, we demonstrate that PDE inhibition by RPL554, a drug that has been shown to cause bronchodilation in asthma and chronic obstructive pulmonary disease (COPD) patients, stimulates CFTR-dependent ion secretion across bronchial epithelial cells isolated from patients carrying the R117H/F508del CF genotype. RPL554-induced CFTR activity was further increased by the potentiator VX-770, suggesting an additional benefit by the drug combination. RPL554 also increased cilia beat frequency in primary human bronchial epithelial cells. The results indicate RPL554 may increase mucociliary clearance through stimulation of CFTR and increasing ciliary beat frequency and thus could provide a novel therapeutic option for CF. PMID:26545902

  3. The Pseudomonas aeruginosa Chp Chemosensory System Regulates Intracellular cAMP Levels by Modulating Adenylate Cyclase Activity

    PubMed Central

    Fulcher, Nanette B.; Holliday, Phillip M.; Klem, Erich; Cann, Martin J.; Wolfgang, Matthew C.

    2010-01-01

    Summary Multiple virulence systems in the opportunistic pathogen Pseudomonas aeruginosa are regulated by the second messenger signaling molecule adenosine 3’, 5’-cyclic monophosphate (cAMP). Production of cAMP by the putative adenylate cyclase enzyme CyaB represents a critical control point for virulence gene regulation. To identify regulators of CyaB, we screened a transposon insertion library for mutants with reduced intracellular cAMP. The majority of insertions resulting in reduced cAMP mapped to the Chp gene cluster encoding a putative chemotaxis-like chemosensory system. Further genetic analysis of the Chp system revealed that it has both positive and negative effects on intracellular cAMP and that it regulates cAMP levels by modulating CyaB activity. The Chp system was previously implicated in the production and function of type IV pili (TFP). Given that cAMP and the cAMP-dependent transcriptional regulator Vfr control TFP biogenesis gene expression, we explored the relationship between cAMP, the Chp system and TFP regulation. We discovered that the Chp system controls TFP production through modulation of cAMP while control of TFP-dependent twitching motility is cAMP-independent. Overall, our data define a novel function for a chemotaxis-like system in controlling cAMP production and establish a regulatory link between the Chp system, TFP and other cAMP-dependent virulence systems. PMID:20345659

  4. Mechanisms of Cyclic Nucleotide Phosphodiesterases in Modulating T Cell Responses in Murine Graft-versus-Host Disease

    PubMed Central

    Weber, Michael; Lupp, Corinna; Stein, Pamela; Kreft, Andreas; Bopp, Tobias; Wehler, Thomas C.; Schmitt, Edgar; Schild, Hansjörg; Radsak, Markus P.

    2013-01-01

    Graft-versus-host disease (GvHD) is a key contributor to the morbidity and mortality after allogeneic hematopoetic stem cell transplantation (HSCT). Regulatory Foxp3+ CD4+ T cells (Treg) suppress conventional T cell activation and can control GvHD. In our previous work, we demonstrate that a basic mechanism of Treg mediated suppression occurs by the transfer of cyclic adenosine monophosphate (cAMP) to responder cells. Whether this mechanism is relevant for Treg mediated suppression of GvHD is currently unknown. To address this question, bone marrow and T cells from C57BL/6 mice were transferred into lethally irradiated BALB/c recipients, and the course of GvHD and survival were monitored. Transplanted recipients developed severe GvHD that was strongly ameliorated by the transfer of donor Treg cells. Towards the underlying mechanisms, in vitro studies revealed that Treg communicated with DCs via gap junctions, resulting in functional inactivation of DC by a metabolic pathway involving cAMP that is modulated by the phosphodiesterase (PDE) 4 inhibitor rolipram. PDE2 or PDE3 inhibitors as well as rolipram suppressed allogeneic T cell activation, indirectly by enhancing Treg mediated suppression of DC activation and directly by inhibiting responder T cell proliferation. In line with this, we observed a cooperative suppression of GvHD upon Treg transfer and additional rolipram treatment. In conclusion, we propose that an important pathway of Treg mediated control of GvHD is based on a cAMP dependent mechanism. These data provide the basis for future concepts to manipulate allogeneic T cell responses to prevent GvHD. PMID:23483980

  5. Mechanisms of cyclic nucleotide phosphodiesterases in modulating T cell responses in murine graft-versus-host disease.

    PubMed

    Weber, Michael; Lupp, Corinna; Stein, Pamela; Kreft, Andreas; Bopp, Tobias; Wehler, Thomas C; Schmitt, Edgar; Schild, Hansjörg; Radsak, Markus P

    2013-01-01

    Graft-versus-host disease (GvHD) is a key contributor to the morbidity and mortality after allogeneic hematopoetic stem cell transplantation (HSCT). Regulatory Foxp3(+) CD4(+) T cells (Treg) suppress conventional T cell activation and can control GvHD. In our previous work, we demonstrate that a basic mechanism of Treg mediated suppression occurs by the transfer of cyclic adenosine monophosphate (cAMP) to responder cells. Whether this mechanism is relevant for Treg mediated suppression of GvHD is currently unknown. To address this question, bone marrow and T cells from C57BL/6 mice were transferred into lethally irradiated BALB/c recipients, and the course of GvHD and survival were monitored. Transplanted recipients developed severe GvHD that was strongly ameliorated by the transfer of donor Treg cells. Towards the underlying mechanisms, in vitro studies revealed that Treg communicated with DCs via gap junctions, resulting in functional inactivation of DC by a metabolic pathway involving cAMP that is modulated by the phosphodiesterase (PDE) 4 inhibitor rolipram. PDE2 or PDE3 inhibitors as well as rolipram suppressed allogeneic T cell activation, indirectly by enhancing Treg mediated suppression of DC activation and directly by inhibiting responder T cell proliferation. In line with this, we observed a cooperative suppression of GvHD upon Treg transfer and additional rolipram treatment. In conclusion, we propose that an important pathway of Treg mediated control of GvHD is based on a cAMP dependent mechanism. These data provide the basis for future concepts to manipulate allogeneic T cell responses to prevent GvHD. PMID:23483980

  6. "Store-operated" cAMP signaling contributes to Ca2+-activated Cl- secretion in T84 colonic cells.

    PubMed

    Nichols, Jonathan M; Maiellaro, Isabella; Abi-Jaoude, Joanne; Curci, Silvana; Hofer, Aldebaran M

    2015-10-15

    Apical cAMP-dependent CFTR Cl(-) channels are essential for efficient vectorial movement of ions and fluid into the lumen of the colon. It is well known that Ca(2+)-mobilizing agonists also stimulate colonic anion secretion. However, CFTR is apparently not activated directly by Ca(2+), and the existence of apical Ca(2+)-dependent Cl(-) channels in the native colonic epithelium is controversial, leaving the identity of the Ca(2+)-activated component unresolved. We recently showed that decreasing free Ca(2+) concentration ([Ca(2+)]) within the endoplasmic reticulum (ER) lumen elicits a rise in intracellular cAMP. This process, which we termed "store-operated cAMP signaling" (SOcAMPS), requires the luminal ER Ca(2+) sensor STIM1 and does not depend on changes in cytosolic Ca(2+). Here we assessed the degree to which SOcAMPS participates in Ca(2+)-activated Cl(-) transport as measured by transepithelial short-circuit current (Isc) in polarized T84 monolayers in parallel with imaging of cAMP and PKA activity using fluorescence resonance energy transfer (FRET)-based reporters in single cells. In Ca(2+)-free conditions, the Ca(2+)-releasing agonist carbachol and Ca(2+) ionophore increased Isc, cAMP, and PKA activity. These responses persisted in cells loaded with the Ca(2+) chelator BAPTA-AM. The effect on Isc was enhanced in the presence of the phosphodiesterase (PDE) inhibitor 3-isobutyl-1-methylxanthine (IBMX), inhibited by the CFTR inhibitor CFTRinh-172 and the PKA inhibitor H-89, and unaffected by Ba(2+) or flufenamic acid. We propose that a discrete component of the "Ca(2+)-dependent" secretory activity in the colon derives from cAMP generated through SOcAMPS. This alternative mode of cAMP production could contribute to the actions of diverse xenobiotic agents that disrupt ER Ca(2+) homeostasis, leading to diarrhea. PMID:26316590

  7. Discovery of a cAMP Deaminase That Quenches Cyclic AMP-Dependent Regulation

    PubMed Central

    Goble, Alissa M.; Feng, Youjun; Raushel, Frank M.; Cronan, John E.

    2013-01-01

    An enzyme of unknown function within the amidohydrolase superfamily was discovered to catalyze the hydrolysis of the universal second messenger, cyclic-3’, 5’-adenosine monophosphate (cAMP). The enzyme, which we have named CadD, is encoded by the human pathogenic bacterium Leptospira interrogans. Although CadD is annotated as an adenosine deaminase, the protein specifically deaminates cAMP to cyclic-3’, 5’-inosine monophosphate (cIMP) with a kcat/Km of 2.7 ± 0.4 × 105 M−1 s−1 and has no activity on adenosine, adenine, or 5’-adenosine monophosphate (AMP). This is the first identification of a deaminase specific for cAMP. Expression of CadD in Escherichia coli mimics the loss of adenylate cyclase in that it blocks growth on carbon sources that require the cAMP-CRP transcriptional activator complex for expression of the cognate genes. The cIMP reaction product cannot replace cAMP as the ligand for CRP binding to DNA in vitro and cIMP is a very poor competitor of cAMP activation of CRP for DNA binding. Transcriptional analyses indicate that CadD expression represses expression of several cAMP-CRP dependent genes. CadD adds a new activity to the cAMP metabolic network and may be a useful tool in intracellular study of cAMP-dependent processes. PMID:24074367

  8. [cAMP cascade in regulation of protein glycosylation].

    PubMed

    Surman, Magdalena; Janik, Marcelina

    2014-01-01

    O- and N-glycosylation are the most common and complex of the post-translational modifications. Both are enzymatic processes and it was suggested that both could be regulated by cAMP cascade at the early stages. N-glycosylation starts with the formation of lipid-linked oligosaccharides and this process is catalysed by crucial glycosyltransferase - dolichol phosphate mannose synthase. The results of several studies strongly suggest that the cAMP acting through a cAMP-dependent protein kinase A-mediated protein phosphorylation/dephosphorylation cycle may modulate activation of this enzyme. It was shown that cAMP can also up regulate another enzyme involved in phosphodolichole synthesis - cis-prenyltransferase. The mechanism acting here is the alteration of the rate of its gene expression. cAMP cascade is also involved in regulation of O-glycosylation since phosphorylation of human glutamine:fructose-6-phosphate amidotransferase results in depletion of O-GlcNAc structure formation. These observation suggested an important role of GPCRs and their ligand in regulation of N- and O-glycan synthesis. PMID:26263760

  9. cAMP/PKA/CREB/GLT1 signaling involved in the antidepressant-like effects of phosphodiesterase 4D inhibitor (GEBR-7b) in rats

    PubMed Central

    Liu, Xu; Guo, Haibiao; Sayed, Mohammad Daud SOM; Lu, Yang; Yang, Ting; Zhou, Dongsheng; Chen, Zhongming; Wang, Haitao; Wang, Chuang; Xu, Jiangping

    2016-01-01

    Objectives GEBR-7b, a potential phosphodiesterase 4D inhibitor, has been shown to have memory-enhancing effects in rodents. However, it is still unknown whether GEBR-7b also has the antidepressant-like effects in rats. Herein, we examined the potential of GEBR-7b to attenuate depression-like behaviors in the rat model of depression induced by chronic unpredictable stress (CUS). Next, we also investigated the alterations of cyclic adenosine monophosphate (cAMP), protein kinase A (PKA) catalytic subunit (PKAca), cAMP response element-binding (CREB), and glutamate transporter 1 (GLT1) levels produced by GEBR-7b in the rats model of depression. Methods Effects of GEBR-7b on CUS (35 days)-induced depression-like behaviors were examined by measuring immobility time in the forced swimming test (FST). Hippocampal cAMP levels were examined by enzyme-linked immunosorbent assay, whereas PKAca, phosphorylation of CREB (pCREB), CREB, and GLT1 in the hippocampus of rats were subjected to Western blot analysis. Results CUS exposure caused a depression-like behavior evidenced by the increased immobility time in FST. Depression-like behavior induced by CUS was accompanied by a significant increased GLT, decreased cAMP, PKAca, pCREB activities in hippocampus. However, repeated GEBR-7b administration significantly reversed CUS-induced depression-like behavior and changes of cAMP/PKA/CREB/GLT1 signaling. No alteration was observed in locomotor activity in open field test. Conclusion These findings indicate that GEBR-7b reversed the depression-like behaviors induced by CUS in rats, which is at least in part mediated by modulating cAMP, PKAca, pCREB, and GLT1 levels in the hippocampus of rats, supporting its neuroprotective potential against behavioral and biochemical dysfunctions induced by CUS. PMID:26855578

  10. Inhibition of Phosphodiesterase-4 during Pneumococcal Pneumonia Reduces Inflammation and Lung Injury in Mice.

    PubMed

    Tavares, Luciana P; Garcia, Cristiana C; Vago, Juliana P; Queiroz-Junior, Celso M; Galvão, Izabela; David, Bruna A; Rachid, Milene A; Silva, Patrícia M R; Russo, Remo C; Teixeira, Mauro M; Sousa, Lirlândia P

    2016-07-01

    Pneumococcal pneumonia is a leading cause of mortality worldwide. The inflammatory response to bacteria is necessary to control infection, but it may also contribute to tissue damage. Phosphodiesterase-4 inhibitors, such as rolipram (ROL), effectively reduce inflammation. Here, we examined the impact of ROL in a pneumococcal pneumonia murine model. Mice were infected intranasally with 10(5)-10(6) CFU of Streptococcus pneumoniae, treated with ROL in a prophylactic or therapeutic schedule in combination, or not, with the antibiotic ceftriaxone. Inflammation and bacteria counts were assessed, and ex vivo phagocytosis assays were performed. ROL treatment during S. pneumoniae infection decreased neutrophil recruitment into lungs and airways and reduced lung injury. Prophylactic ROL treatment also decreased cytokine levels in the airways. Although modulation of inflammation by ROL ameliorated pneumonia, bacteria burden was not reduced. On the other hand, antibiotic therapy reduced bacteria without reducing neutrophil infiltration, cytokine level, or lung injury. Combined ROL and ceftriaxone treatment decreased lethality rates and was more efficient in reducing inflammation, by increasing proresolving protein annexin A1 (AnxA1) expression, and bacterial burden by enhancing phagocytosis. Lack of AnxA1 increased inflammation and lethality induced by pneumococcal infection. These data show that immunomodulatory effects of phosphodiesterase-4 inhibitors are useful during severe pneumococcal pneumonia and suggest their potential benefit as adjunctive therapy during infectious diseases. PMID:26677751

  11. The YmdB Phosphodiesterase Is a Global Regulator of Late Adaptive Responses in Bacillus subtilis

    PubMed Central

    Diethmaier, Christine; Newman, Joseph A.; Kovács, Ákos T.; Kaever, Volkhard; Herzberg, Christina; Rodrigues, Cecilia; Boonstra, Mirjam; Kuipers, Oscar P.

    2014-01-01

    Bacillus subtilis mutants lacking ymdB are unable to form biofilms, exhibit a strong overexpression of the flagellin gene hag, and are deficient in SlrR, a SinR antagonist. Here, we report the functional and structural characterization of YmdB, and we find that YmdB is a phosphodiesterase with activity against 2′,3′- and 3′,5′-cyclic nucleotide monophosphates. The structure of YmdB reveals that the enzyme adopts a conserved phosphodiesterase fold with a binuclear metal center. Mutagenesis of a catalytically crucial residue demonstrates that the enzymatic activity of YmdB is essential for biofilm formation. The deletion of ymdB affects the expression of more than 800 genes; the levels of the σD-dependent motility regulon and several sporulation genes are increased, and the levels of the SinR-repressed biofilm genes are decreased, confirming the role of YmdB in regulating late adaptive responses of B. subtilis. PMID:24163345

  12. Central Atlantic Magmatic Province (CAMP): The Palisade Sill Connection

    NASA Astrophysics Data System (ADS)

    Ghatak, A.; Basu, A. R.

    2012-12-01

    The extensively studied 200Ma Central Atlantic Magmatic Province (CAMP) is considered to be the world's largest continental Large Igneous Province (LIP) covering up to 7 X 106 km2. This igneous province has been linked to the ~200Ma Mesozoic opening of the Central Atlantic Ocean. This opening fragmented the CAMP into several segments that occur on four different tectonic plates today. The CAMP related LIP is different from others in that it constitutes almost entirely of dikes and sills with sparse volcanic outflows. The 200 Ma Palisade Sill, exposed along the Hudson River in northeastern North America is an expression of the CAMP magmatism. On the basis of similar ages of eruption, Palisade Sill tholeiites have been correlated to other CAMP exposures in four continents. We provide an isotopic tracer study of the Palisade Sill basalts and relate them to low-Ti (<2 wt %) CAMP related tholeiites from North and South America, western Europe, and West Africa. We report Nd-Sr-Pb isotopic and multiple trace element data of 19 basalts and gabbros, 3 chilled margin basalts, and 4 sandstones spanning the entire length and thickness of the Palisade Sill in New York and New Jersey. These geochemical data are essential to understand the relationship between mantle geodynamic processes involved in the generation of the CAMP tholeiites prior to the formation of the of the Atlantic Ocean crust. The Palisade Sill basalts of this study yield the typical composition of low-Ti CAMP tholeiites with small LREE enrichments (LaN/SmN = 1.7 to 2.3), radiogenic Sr and negative ɛNd(I) values (87Sr/87Sr(I) = 0.70668 to 0.71037; ɛNd(I) = -0.64 to -3.8), and Pb-isotopic ratios (e.g. 206Pb/204Pb = 18.11 to 18.69) above the NHRL and subparallel to it. These geochemical data indicate the Palisade Sill basalts were derived from a slightly enriched OIB-like mantle source. Further, these rocks were derived by ~15% melting of a slightly depleted spinel peridotite with up to 20% contamination by the

  13. US SPACE CAMP CALIFORNIA - DAY CAMP GRAND OPENING WITH KEVIN JONES (WHISMAN SCHOOL) AND LUCRETIA

    NASA Technical Reports Server (NTRS)

    1996-01-01

    US SPACE CAMP CALIFORNIA - DAY CAMP GRAND OPENING WITH KEVIN JONES (WHISMAN SCHOOL) AND LUCRETIA SUTHERLIN (MCNAIR SCHOOL). AMES SPONSORED STUDENTS AND RACHAEL QUIRING (STAFF) - AUTOGRAPH SIGNING BY Astronaut Wally Schirra

  14. A Day in the Life of Three Special Needs Camps

    ERIC Educational Resources Information Center

    Winbaum, Stephen

    2006-01-01

    In this article, the author talks about three special needs camps--Camp Kirk, Camp Talisman and Camp Caglewood. Camp Kirk's philosophy is to encourage their children to take risks in a structured setting, like high ropes courses, rock climbing wall, martial arts, and traditional activities like swimming, arts and craft, drama, and others. Once…

  15. College Camp Fairs: Tips for Hosts, Directors, and Students.

    ERIC Educational Resources Information Center

    Gaskill, Paul L.; Campbell, Don

    1995-01-01

    Camp fairs are held on college campuses to recruit students for summer camp positions. Provides marketing and promotion strategies, recruiting suggestions for camp directors, tips for students helping to organize the fair, and suggestions for students who want to secure a summer camp position. Includes a sample camp fair evaluation. (LP)

  16. Stories from Camp: Understanding the Impact of What We Do.

    ERIC Educational Resources Information Center

    DeGraaf, Don; Glover, Jessie

    2002-01-01

    A study examining the impacts of camp on staff interviewed 29 former seasonal camp staff. All respondents reported positive benefits in their personal and professional lives and the strong influence of camp in shaping career choices. Reflections on camp fell into three categories: uniqueness of camp, making memories for kids, and freedom. (TD)

  17. A putative antipruritic mechanism of the phosphodiesterase-4 inhibitor E6005 by attenuating capsaicin-induced depolarization of C-fibre nerves.

    PubMed

    Wakita, Hisashi; Ohkuro, Masayoshi; Ishii, Naoto; Hishinuma, Ieharu; Shirato, Manabu

    2015-03-01

    E6005, a potent, selective phosphodiesterase (PDE) 4 inhibitor, has been developed as a novel topical agent of atopic dermatitis (AD). It has been shown to inhibit itching in patients with AD as well in mouse models. To study the mechanism underlying the anti-pruritic effect of E6005, we examined its effect on the activation of dorsal root ganglion (DRG) neurons associated with the itch sensation. Depolarization of DRG neurons by a transient receptor potential vanilloid 1 (TRPV 1) activator, capsaicin was attenuated by E6005 as well as by a 3',5'-cyclic adenosine monophosphate (cAMP) elevator, forskolin. E6005 elevated intracellular levels of cAMP in DRG cells. Taken together, these results suggest that E6005 suppresses TRPV1-mediated C-fibre depolarization through elevation of cAMP levels, thereby exerting an anti-pruritic effect. Thus, E6005 shows the potential to be a new agent for managing pruritus in various skin disorders, including AD. PMID:25431172

  18. Program evaluation of Protovation Camp

    NASA Astrophysics Data System (ADS)

    Healy, Laurel Lynell Martin

    The purpose of this program evaluation was to determine the extent to which Protovation Camp utilized the combined resources of multiple institutions to impact student learning in science, technology, engineering, and math. The partnership consisted of multiple institutions: the university, providing graduate students to facilitate inquiry-based lessons; the science center, allowing the use of their facilities and resources; and the elementary school, contributing rising third through fifth grade campers. All of these components were examined. The mixed-methods approach used post hoc quantitative data for campers, which consisted of pre-test and post-test scores on the Test of Science-Related Attitudes (TOSRA), the Draw-A-Scientist Test, and content tests based on the camp activities. Additionally, TOSRA scores and current survey results for the graduate students were used along with qualitative data collected from plusdelta charts to determine the impact of participation in Protovation Camp on teachers and students. Results of the program evaluation indicated that when students were taught inquiry-based lessons that ignite wonder, both their attitudes toward science and their knowledge about science improved. An implication for teacher preparation programs was that practicing inquiry-based lessons on actual students (campers) was an important component for teachers (graduate students) as they prepare to positively impact student learning in their own classrooms. Immediate feedback from the campers in the form of pre-test and post-test scores and from peers on plusdelta charts allowed the graduate students the opportunity to make needed adjustments to improve effectiveness before using the lesson with a new set of campers or later in their own classrooms. Keywords. Teacher preparation, Inquiry-based instruction, STEM instructions, University and museum partnerships

  19. Camp Programs Provide Community Opportunities: Camp Henry Helps Community with Youth Diversion Program.

    ERIC Educational Resources Information Center

    Vander Kooi, Gregory; Astle, Judy Hughes; Jacobs, Jeff

    2001-01-01

    A Michigan program gives juvenile first-offenders in alcohol, drug, or tobacco crimes the option of completing a youth diversion program at a local camp as an alternative to traditional juvenile justice. Considerations for the camp included board support, compatibility with camp philosophy, and staff competence. The program has lower recidivism…

  20. Themes from a Camp Maintenance Network: Camp Maintenance and Property Personnel Share Their Insights and Challenges.

    ERIC Educational Resources Information Center

    Whyman, Wynne

    2003-01-01

    A camp maintenance survey was completed by maintenance personnel from 99 camps. Results highlighted several important considerations: ensuring sufficient maintenance funds for aging infrastructure, including camp/property personnel in decision making, publicizing completed maintenance projects, examining long-term needs of the land, and adopting…

  1. AKAP79, PKC, PKA and PDE4 participate in a Gq-linked muscarinic receptor and adenylate cyclase 2 cAMP signalling complex

    PubMed Central

    Shen, Jia X.; Cooper, Dermot M. F.

    2014-01-01

    AC2 (adenylate cyclase 2) is stimulated by activation of Gq-coupled muscarinic receptors through PKC (protein kinase C) to generate localized cAMP in HEK (human embryonic kidney)-293 cells. In the present study, we utilized a sensitive live-cell imaging technique to unravel the proteins that play essential roles in a Gq-coupled muscarinic receptor-mediated cAMP signalling complex. We reveal that, upon agonist binding to the Gq-coupled muscarinic receptor, AKAP79 (A-kinase-anchoring protein 79) recruits PKC to activate AC2 to produce cAMP. The cAMP formed is degraded by PDE4 (phosphodiesterase 4) activated by an AKAP-anchored PKA (protein kinase A). Calcineurin, a phosphatase bound to AKAP79, is not involved in this regulation. Overall, a transient cAMP increase is generated from AC2 by Gq-coupled muscarinic receptor activation, subject to sophisticated regulation through AKAP79, PKC, PDE4 and PKA, which significantly enhances acetylcholine-mediated signalling. PMID:23889134

  2. cGMP-PDE3-cAMP signal pathway involved in the inhibitory effect of CNP on gastric motility in rat.

    PubMed

    Cai, Ying-Lan; Sun, Qian; Huang, Xu; Jiang, Jing-Zhi; Zhang, Mo-Han; Piao, Li-Hua; Jin, Zheng; Xu, Wen-Xie

    2013-01-10

    In the present study, we investigated the mechanism of C-type natriuretic peptide (CNP)-induced inhibitory effect on spontaneous contraction of gastric antral smooth muscle to clarify CNP-NPR-B/pGC-cGMP downstream signal transduction pathway using organ bath and ELISA methods in rat. CNP significantly reduced the amplitude of the spontaneous contraction and increased the contents of cGMP and cAMP in the gastric antral smooth muscle tissue. In the presence of IBMX, a non-selective phosphodiesterase (PDE) inhibitor, the inhibitory effect of CNP on spontaneous contraction was significantly suppressed; however, the production of cGMP but not cAMP was still increased by CNP. EHNA, a PDE2 inhibitor, did not affect both CNP-induced inhibition of the contraction and CNP-induced increase of cGMP and cAMP generations in gastric smooth muscle tissue, while milrinone, a PDE3 inhibitor, similar to IBMX, attenuated the CNP-induced inhibitory effect on spontaneous contraction and increased the content of cGMP but not cAMP. The results suggest that cGMP-PDE3-cAMP signal pathway is also involved in the CNP-induced inhibition of gastric motility in rat. PMID:23186653

  3. Cigarette Smoke-Induced Emphysema and Pulmonary Hypertension Can Be Prevented by Phosphodiesterase 4 and 5 Inhibition in Mice

    PubMed Central

    Pichl, Alexandra; Bednorz, Mariola; Ghofrani, Hossein Ardeschir; Schermuly, Ralph Theo; Seeger, Werner; Grimminger, Friedrich; Weissmann, Norbert

    2015-01-01

    Rationale Chronic obstructive pulmonary disease (COPD) is a widespread disease, with no curative therapies available. Recent findings suggest a key role of NO and sGC-cGMP signaling for the pathogenesis of the disease. Previous data suggest a downregulation/inactivation of the cGMP producing soluble guanylate cyclase, and sGC stimulation prevented cigarette smoke-induced emphysema and pulmonary hypertension (PH) in mice. We thus aimed to investigate if the inhibition of the cGMP degrading phosphodiesterase (PDE)5 has similar effects. Results were compared to the effects of a PDE 4 inhibitor (cAMP elevating) and a combination of both. Methods C57BL6/J mice were chronically exposed to cigarette smoke and in parallel either treated with Tadalafil (PDE5 inhibitor), Piclamilast (PDE4 inhibitor) or both. Functional measurements (lung compliance, hemodynamics) and structural investigations (alveolar and vascular morphometry) as well as the heart ratio were determined after 6 months of tobacco smoke exposure. In addition, the number of alveolar macrophages in the respective lungs was counted. Results Preventive treatment with Tadalafil, Piclamilast or a combination of both almost completely prevented the development of emphysema, the increase in lung compliance, tidal volume, structural remodeling of the lung vasculature, right ventricular systolic pressure, and right ventricular hypertrophy induced by cigarette smoke exposure. Single, but not combination treatment prevented or reduced smoke-induced increase in alveolar macrophages. Conclusion Cigarette smoke-induced emphysema and PH could be prevented by inhibition of the phosphodiesterases 4 and 5 in mice. PMID:26058042

  4. Cis-Lunar Base Camp

    NASA Technical Reports Server (NTRS)

    Merrill, Raymond G.; Goodliff, Kandyce E.; Mazanek, Daniel D.; Reeves, John D., Jr.

    2012-01-01

    Historically, when mounting expeditions into uncharted territories, explorers have established strategically positioned base camps to pre-position required equipment and consumables. These base camps are secure, safe positions from which expeditions can depart when conditions are favorable, at which technology and operations can be tested and validated, and facilitate timely access to more robust facilities in the event of an emergency. For human exploration missions into deep space, cis-lunar space is well suited to serve as such a base camp. The outer regions of cis-lunar space, such as the Earth-Moon Lagrange points, lie near the edge of Earth s gravity well, allowing equipment and consumables to be aggregated with easy access to deep space and to the lunar surface, as well as more distant destinations, such as near-Earth Asteroids (NEAs) and Mars and its moons. Several approaches to utilizing a cis-lunar base camp for sustainable human exploration, as well as some possible future applications are identified. The primary objective of the analysis presented in this paper is to identify options, show the macro trends, and provide information that can be used as a basis for more detailed mission development. Compared within are the high-level performance and cost of 15 preliminary cis-lunar exploration campaigns that establish the capability to conduct crewed missions of up to one year in duration, and then aggregate mass in cis-lunar space to facilitate an expedition from Cis-Lunar Base Camp. Launch vehicles, chemical propulsion stages, and electric propulsion stages are discussed and parametric sizing values are used to create architectures of in-space transportation elements that extend the existing in-space supply chain to cis-lunar space. The transportation options to cis-lunar space assessed vary in efficiency by almost 50%; from 0.16 to 0.68 kg of cargo in cis-lunar space for every kilogram of mass in Low Earth Orbit (LEO). For the 15 cases, 5-year campaign

  5. A cardiac mitochondrial cAMP signaling pathway regulates calcium accumulation, permeability transition and cell death

    PubMed Central

    Wang, Z; Liu, D; Varin, A; Nicolas, V; Courilleau, D; Mateo, P; Caubere, C; Rouet, P; Gomez, A-M; Vandecasteele, G; Fischmeister, R; Brenner, C

    2016-01-01

    Although cardiac cytosolic cyclic 3′,5′-adenosine monophosphate (cAMP) regulates multiple processes, such as beating, contractility, metabolism and apoptosis, little is known yet on the role of this second messenger within cardiac mitochondria. Using cellular and subcellular approaches, we demonstrate here the local expression of several actors of cAMP signaling within cardiac mitochondria, namely a truncated form of soluble AC (sACt) and the exchange protein directly activated by cAMP 1 (Epac1), and show a protective role for sACt against cell death, apoptosis as well as necrosis in primary cardiomyocytes. Upon stimulation with bicarbonate (HCO3−) and Ca2+, sACt produces cAMP, which in turn stimulates oxygen consumption, increases the mitochondrial membrane potential (ΔΨm) and ATP production. cAMP is rate limiting for matrix Ca2+ entry via Epac1 and the mitochondrial calcium uniporter and, as a consequence, prevents mitochondrial permeability transition (MPT). The mitochondrial cAMP effects involve neither protein kinase A, Epac2 nor the mitochondrial Na+/Ca2+ exchanger. In addition, in mitochondria isolated from failing rat hearts, stimulation of the mitochondrial cAMP pathway by HCO3− rescued the sensitization of mitochondria to Ca2+-induced MPT. Thus, our study identifies a link between mitochondrial cAMP, mitochondrial metabolism and cell death in the heart, which is independent of cytosolic cAMP signaling. Our results might have implications for therapeutic prevention of cell death in cardiac pathologies. PMID:27100892

  6. A Kid's Guide to Family Camping.

    ERIC Educational Resources Information Center

    Regimbal, Celia; Kasakow, Nancy

    Fifth and sixth grade students of the University School at the University of Wyoming participated in a successful week's overnight outdoor experience in September 1983. Because results of a pre-camp questionnaire indicated that most students had minimal knowledge of the planning necessary for camping, activities and classes were organized to…

  7. Cedar Ridge Camp: Using the Local Environment

    ERIC Educational Resources Information Center

    Burke, Grayson

    2007-01-01

    In 2007 Cedar Ridge Camp opened for its first season as a traditional co-ed summer camp and year-round outdoor education and recreation centre. The mission would centre on creating a program that would encourage personal development and growth through a shared outdoor experience. Cedar Ridge's main goals were to promote the formation of close…

  8. Camping Safety--Bring 'Em Back Alive.

    ERIC Educational Resources Information Center

    Schmidt, Ernest F.

    1980-01-01

    A "prioritized" list of dangers of the woods is discussed and suggestions for safety in organized camping are listed. Available from: Center for Environmental, Camping and Outdoor Education; University of North Carolina at Greensboro; Pine Lake Field Campus; 4016 Blumenthal Road; Greensboro, NC, 27406. (AN)

  9. 36 CFR 327.7 - Camping.

    Code of Federal Regulations, 2013 CFR

    2013-07-01

    ... 36 Parks, Forests, and Public Property 3 2013-07-01 2012-07-01 true Camping. 327.7 Section 327.7... GOVERNING PUBLIC USE OF WATER RESOURCE DEVELOPMENT PROJECTS ADMINISTERED BY THE CHIEF OF ENGINEERS § 327.7...) Camping at one or more campsites at any one water resource project for a period longer than 14 days...

  10. American Camping Association. Annual Report, 1985.

    ERIC Educational Resources Information Center

    American Camping Association, Martinsville, IN.

    The 1985 American Camping Association (ACA) annual report reviews trends and successes of the year. The document is in the format of a calendar covering the period October 1985 through September 1986. Calendar pages, on which relevant camping events are noted, alternate with pages of text. An introductory message from ACA President Jean McMullan…

  11. 1940s: Camping in the War Years.

    ERIC Educational Resources Information Center

    Camping Magazine, 1999

    1999-01-01

    Camps continued to operate during World War II, but young male counselors, food, and supplies were difficult to obtain. An illustrative article from 1943, "Meal Planning for Summer Camps in Wartime" (Agnes B. Peterson), presents a guide to planning nutritious meals for campers despite shortages caused by wartime rationing, increased food costs,…

  12. Celebrations: American Camping Association Annual Report 1986.

    ERIC Educational Resources Information Center

    American Camping Association, Martinsville, IN.

    The 1986 American Camping Association (ACA) annual report reviews the year's achievements and outlines goals for the future. An introductory message from ACA President Jean McMullan notes successful fund raising to improve the association's national headquarters, passage of federal legislation exempting camps from paying federal unemployment…

  13. Creating a Sun-Safe Camp.

    ERIC Educational Resources Information Center

    Landrey, Ann

    1996-01-01

    Strategies for minimizing sun exposure of campers and staff include educating campers about the sun's effect on their skin, scheduling activities when the sun is less intense, creating shade at the camp site, incorporating sun protection into camp dress code, and training staff regarding sun protection. Addresses OSHA and liability issues. (LP)

  14. Science Camp: Just for the Girls

    ERIC Educational Resources Information Center

    Cavanagh, Sean

    2007-01-01

    Research shows that girls tend to lose interest in science and math as they move through the education pipeline--a retreat that often begins during middle school. Summer science camps can be part of reversing that trend, some say. Academic camps are on the rise across the country, including ones to get adolescent girls excited about the…

  15. Suicides in the Nazi Concentration Camps.

    ERIC Educational Resources Information Center

    Ryn, Zdzislaw

    1986-01-01

    On the basis of psychiatric interviews with 69 former prisoners of the Auschwitz-Birkenau concentration camp, this paper describes the circumstances, motives, and ways of committing suicide in the camp. The interviews made it clear that thousands of prisoners perished by suicide. The number of committed suicides was larger than that of attempted…

  16. Incontinent Children Discover Hope at Camp Oakhurst.

    ERIC Educational Resources Information Center

    Raps, Marvin A.

    1991-01-01

    Describes summer-camp experiences of young people who are incontinent because of spina bifida. Focuses on residential camp program as encouragement for youth to learn independent incontinence management. Describes self-catheterization program and process as minor inconvenience and positive alternative to soiled clothing and accompanying…

  17. Passenger Vans: A Transportation Concern for Camps.

    ERIC Educational Resources Information Center

    Gentle, Don; Gordon, Harold; Schainman, Steve

    2002-01-01

    In April 2002, the National Transportation Safety Board reissued a cautionary warning about the high accident potential of 15-passenger vans; some states have prohibited their use for transporting children. Three camp directors discuss legal and cost considerations for camps, possible alternative transportation, additional staff training needs,…

  18. Measuring Mindfulness in Summer Camp Staff

    ERIC Educational Resources Information Center

    Gillard, Ann; Roark, Mark F.; Nyaga, Lewis Ramsey Kanyiba; Bialeschki, M. Deborah

    2011-01-01

    Examining mindfulness in a non-clinical and non-therapeutic setting such as a summer camp is an area of growing interest. Our study tested three mindfulness scales with staff in a summer camp setting, and we conducted preliminary reliability and validity analyses for any modifications needed in the scales. Results indicated two major findings: (a)…

  19. Teen Moms and Babies Benefit from Camping.

    ERIC Educational Resources Information Center

    Goode, Marsha; Broesamle, Barbara

    1987-01-01

    Describes nine-day residential camp for Michigan teenage mothers/babies to enhance personal growth and develop responsible social skills. Outlines goals, pre-camp planning, staff, activities, evaluation. Reports 31 teen moms (ages 13-21) and 35 babies attended in 1986. Indicates participants were in therapy, experienced abuse, had low self-esteem,…

  20. Children with Cancer: Positive Benefits of Camp.

    ERIC Educational Resources Information Center

    Winfree, Christy; Williams, Richard; Powell, Gwynn M.

    2002-01-01

    A relatively new method of helping pediatric cancer patients cope with their illness is specially designed summer camps. Camp helps children with cancer address psychological effects of the disease, bodily changes, and self-concept, and helps parents and siblings cope. Sidebars present resources and tips on incorporating children with cancer into…

  1. Foreign Language Camps: A Success Story.

    ERIC Educational Resources Information Center

    Shrum, Judith

    The foreign language camps, operated by Virginia Tech since 1981, offer high school students a one-week foreign language experience in French, German, or Spanish. The camps bring together native speakers, high school students and teachers, and university students and faculty to experience learning situations similar to those encountered in a…

  2. LANGUAGE CAMPS IN THE UNITED STATES.

    ERIC Educational Resources Information Center

    BOUNIOL, EILEEN C.

    A SUMMARY IS GIVEN OF AN INFORMAL SURVEY MADE OF STATE FOREIGN LANGUAGE SUPERVISORS TO DETERMINE THE EXTENT AND NATURE OF LANGUAGE CAMPS IN THE UNITED STATES. DATA FROM QUESTIONNAIRES RETURNED BY THE SUPERVISORS REVEALED INFORMATION ABOUT THE NAMES AND LOCATIONS OF 12 SUMMER CAMPS, PERTINENT DETAILS OF EACH PROGRAM, SPONSORING INSTITUTIONS, DATES…

  3. Smart Kids at CAMP-US

    ERIC Educational Resources Information Center

    Sinatra, Richard

    2004-01-01

    This article describes the summer literacy, athletic, and computer program know as CAMP-US. Each year, CAMP-US serves approximately 500 children in 10-day sessions. The first half of each day is devoted to literacy and computer instruction, while the second half is spent engaging in recreational activities such as swimming, soccer, softball,…

  4. Dealing with World Issues in Camp.

    ERIC Educational Resources Information Center

    Kujawa, Charles

    1986-01-01

    Discusses dealing with global issues in the camp setting in a way that broadens young people's world views. Topics include the educational advantages of the camp setting, desired outcomes for campers, guidelines for staff, and program ideas for dealing with issues such as environmental awareness, racism, and economic justice. (JHZ)

  5. Using the Internet To Promote Your Camp.

    ERIC Educational Resources Information Center

    Schenk, Tom

    1997-01-01

    Discusses the benefits of camps having a World Wide Web site and tips for choosing a Web provider. Offers recommendations for creating a Web page, including determining a goal, outlining content, choosing a graphic style and colors, testing the Web design, and promoting the Web site in camp brochures. Includes sample Web sites and Web resources.…

  6. Prevention of Potential Juvenile Delinquency Through Camping.

    ERIC Educational Resources Information Center

    Kiddie Kamp Corp., Boston, MA.

    A two-year demonstration project, to determine whether a short-term (30 days) summer camp experience could contribute to prevention of juvenile delinquency, found that camping induced positive attitudinal change and improved adjustment to school. The project involved 200 adolescent boys from varied social backgrounds who were selected on evidence…

  7. 1980s: Camp in a Computer Age.

    ERIC Educational Resources Information Center

    Camping Magazine, 1999

    1999-01-01

    Important issues for camps in the 1980s included soaring land values, high insurance rates, the spread of computers, and growing demand for specialized services. An illustrative article from 1985, "Yuppies Are Coming" (Tim Duffield), discusses the willingness of baby-boomer parents to pay for high-quality enrichment experiences at camp and other…

  8. Youth Development and the Camp Experience

    ERIC Educational Resources Information Center

    Garst, Barry A.; Browne, Laurie P.; Bialeschki, M. Deborah

    2011-01-01

    The organized camp experience has been an important part of the lives of children, youth, and adults for over 150 years and is a social institution that touches more lives than any other except for schools. Camp is more than a location or a program; it encompasses the affective, cognitive, behavioral, physical, social, and spiritual benefits that…

  9. What Happens to Campers at Camp?

    ERIC Educational Resources Information Center

    Powell, Gwynn M.

    2003-01-01

    A study of 66 children with cancer and 43 siblings attending the Ronald McDonald Camp found that disease-specific camps allow children membership in a community of peers, which enhances self-esteem and social acceptance. A separate, longitudinal study of 38 beginning and experienced campers found that campers' intrapersonal and interpersonal…

  10. Summer Camp of Mathematical Modeling in China

    ERIC Educational Resources Information Center

    Tian, Xiaoxi; Xie, Jinxing

    2013-01-01

    The Summer Camp of Mathematical Modeling in China is a recently created experience designed to further Chinese students' academic pursuits in mathematical modeling. Students are given more than three months to research on a mathematical modeling project. Researchers and teams with outstanding projects are invited to the Summer Camp to present…

  11. Forest Fire: A Crisis Reality for Camp.

    ERIC Educational Resources Information Center

    Brown, Don; Mickelson, Rhonda

    2002-01-01

    Two camp directors were interviewed about evacuations from their camps due to forest fires. Topics covered include descriptions of the events; actions taken; aspects of advance planning that proved helpful; unexpected portions of the experience and resultant changes made in plans; relations with outside agencies, the media, and parents; working…

  12. Tim Horton Camps: Spotlight on Onondaga Farms.

    ERIC Educational Resources Information Center

    Evanson, Jason

    2003-01-01

    Tim Horton Children's Foundation operates camps for disadvantaged children that build their self-confidence. The newest camp, Onondaga Farms in Ontario, is also a year-round outdoor education center. Its five program streams--environmental education, creative arts, agriculture, adventure, and recreation--all have direct links to Ontario's…

  13. Tech Camp Unleashes Creativity and Collaboration

    ERIC Educational Resources Information Center

    Bardin, Joe

    2008-01-01

    Each August, teachers from around the state gather for the Arizona K-12 Center's Tech Camp, a week-long immersion in technology for the classroom. The Arizona K-12 Center's mission is to improve teaching and learning in Arizona's schools through high-quality professional development and teacher leadership. The formula Tech Camp follows is a simple…

  14. Teenagers and Risk-Taking at Camp.

    ERIC Educational Resources Information Center

    Woods, Ann

    2002-01-01

    Teen risk-taking is normal, healthy developmental behavior. Teens act out their fantasies--good and bad--at camp because it is a safe place away from parents. Signs of unhealthy risk-taking, camp staff responses, and how the September 11 tragedy might affect risk-taking are discussed. Sidebars describe tips for understanding adolescent behavior…

  15. Why It's Good to Go to Camp

    ERIC Educational Resources Information Center

    Howell, Matthew

    2008-01-01

    In the author's fourth year of undergraduate studies at the University of Waterloo he had the opportunity to explore the benefits attained by children attending a summer camp by way of an academic literature review. The author worked with Dr. Troy Glover who has been commissioned by a group of camping associations to perform a study on the…

  16. Residential Camping for At Risk Preschoolers.

    ERIC Educational Resources Information Center

    Kosberg, Bernard; Mellins, Claude A.; Pubentz, Jennifer L.

    A combination of experiential education and residential camping can have positive effects on preschool children with emotional, behavioral, and learning problems. Preschool Prep at Ramapo Anchorage Camp (Rhinebeck, New York) was developed to meet the specific needs of high-risk children aged 4-6. Many of the children come from unsafe urban…

  17. Accounting Boot Camp for College Juniors

    ERIC Educational Resources Information Center

    Myring, Mark; Wrege, William; Van Alst, Lucinda

    2008-01-01

    We describe a day-long introduction to new accounting majors, which we call a boot camp. Boot camp it is an effort to make juniors more aware of their identity, career purposes and learning resources that are now parts of their world, much of which is not covered explicitly in the accounting curriculum. This paper provides an overview of the…

  18. [Medicine in the Soviet labor camps].

    PubMed

    Supady, J

    1998-01-01

    The existence of medical attention in Soviet labour camps was dictated not by humane reasons but by economical motives. It was just a wish to regenerate physical strength of some labour camp's slaves in order to exploit them to work out definite production as well as ideological plans. However, despite the lack of medicines, equipment, proper housing conditions and qualified staff, employed in labour camp's medical service people--doctors, hospital attendants, nurses, etc.--that were mostly recruited from amongst prisoners, showed a great deal of kindness and did their best to come to unwell and ill patients' assistance. Prisoners' lives were not uncommonly saved through exemption from work by a doctor's decision sending to hospital or transfer to another health category. In comparison with gehenna of labour camp's slave everyday life a sojourn in camp hospital seemed a paradise and a rescue from death to prisoners. PMID:10216383

  19. Phosphodiesterase isoenzyme families in human osteoarthritis chondrocytes – functional importance of phosphodiesterase 4

    PubMed Central

    Tenor, Hermann; Hedbom, Erik; Häuselmann, Hans-Jörg; Schudt, Christian; Hatzelmann, Armin

    2002-01-01

    We studied whether selective inhibitors of cyclic nucleotide hydrolysing phosphodiesterase (PDE) isoenzymes influence IL-1β-induced nitric oxide (NO) release from human articular chondrocytes. In addition, the pattern of PDE isoenzymes contributing to cyclic nucleotide hydrolysis in human chondrocytes was characterized.Chondrocytes were isolated from human osteoarthritic cartilage and cultured in alginate beads. IL-1β-induced chondrocyte products (nitric oxide and prostaglandin E2) were measured in culture supernatants after 48 h incubation time. PDE activities were assessed in chondrocyte lysates. Inducible nitric oxide synthase (iNOS) and PDE4A-D proteins were detected by immunoblotting.The selective PDE4 inhibitors Piclamilast and Roflumilast partially attenuated IL-1β-induced NO production whereas selective inhibitors of PDE2 (EHNA), PDE3 (Motapizone) or PDE5 (Sildenafil) were inactive. Indomethacin reversed the reduction of IL-1β-induced NO by PDE4 inhibitors. It was shown that autocrine prostaglandin E2 (PGE2) enabled PDE4 inhibitors to reduce IL-1β-induced NO in this experimental setting.Major PDE4 and PDE1 activities were identified in chondrocyte lysates whereas only minor activities of PDE2, 3 and 5 were found. IL-1β and cyclic AMP-mimetics upregulated PDE4 activity and this was associated with an augmentation of PDE4B2 protein.Based on the view that nitric oxide contributes to cartilage degradation in osteoarthritis our study suggests that PDE4 inhibitors may have chondroprotective effects. PMID:11834608

  20. The extracellular signal-regulated kinase mitogen-activated protein kinase/ribosomal S6 protein kinase 1 cascade phosphorylates cAMP response element-binding protein to induce MUC5B gene expression via D-prostanoid receptor signaling.

    PubMed

    Choi, Yeon Ho; Lee, Sang-Nam; Aoyagi, Hiroki; Yamasaki, Yasundo; Yoo, Jung-Yoon; Park, Boryung; Shin, Dong Min; Yoon, Ho-Geun; Yoon, Joo-Heon

    2011-09-30

    Mucus hypersecretion is a prominent feature of respiratory diseases, and MUC5B is a major airway mucin. Mucin gene expression can be affected by inflammatory mediators, including prostaglandin (PG) D(2,) an inflammatory mediator synthesized by hematopoietic PGD synthase (H-PGDS). PGD(2) binds to either D-prostanoid receptor (DP1) or chemoattractant receptor homologous molecule expressed on T-helper type 2 cells (CRTH2). We investigated the mechanisms by which PGD(2) induces MUC5B gene expression in airway epithelial cells. Western blot analysis showed that H-PGDS was highly expressed in nasal polyps. Similar results were obtained for PGD(2) expression. In addition, we could clearly detect the expressions of both H-PGDS and DP1 in nasal epithelial cells but not CRTH2. We demonstrated that PGD(2) increased MUC5B gene expression in normal human nasal epithelial cells as well as in NCI-H292 cells in vitro. S5751, a DP1 antagonist, inhibited PGD(2)-induced MUC5B expression, whereas a CRTH2 antagonist (OC0459) did not. These data suggest that PGD(2) induced MUC5B expression via DP1. Pretreatment with extracellular signal-regulated kinase (ERK) inhibitor (PD98059) blocked both PGD(2)-induced ERK mitogen-activated protein kinase (MAPK) activation and MUC5B expression. Proximity ligation assays showed direct interaction between RSK1 and cAMP response element-binding protein (CREB). Stimulation with PGD(2) caused an increase in intracellular cAMP levels, whereas intracellular Ca(2+) did not have such an effect. PGD(2)-induced MUC5B mRNA levels were regulated by CREB via direct interaction with two cAMP-response element sites (-921/-914 and -900/-893). Finally, we demonstrated that PGD(2) can induce MUC5B overproduction via ERK MAPK/RSK1/CREB signaling and that DP1 receptor may have suppressive effects in controlling MUC5B overproduction in the airway. PMID:21832046

  1. Kinetic and Structural Studies of Phosphodiesterase-8A and Implication on the Inhibitor Selectivity

    SciTech Connect

    Wang, H.; Yan, Z; Yang, S; Cai, J; Robinson, H; Ke, H

    2008-01-01

    Cyclic nucleotide phosphodiesterase-8 (PDE8) is a family of cAMP-specific enzymes and plays important roles in many biological processes, including T-cell activation, testosterone production, adrenocortical hyperplasia, and thyroid function. However, no PDE8 selective inhibitors are available for trial treatment of human diseases. Here we report kinetic properties of the highly active PDE8A1 catalytic domain prepared from refolding and its crystal structures in the unliganded and 3-isobutyl-1-methylxanthine (IBMX) bound forms at 1.9 and 2.1 Angstroms resolutions, respectively. The PDE8A1 catalytic domain has a KM of 1.8 eM, Vmax of 6.1 emol/min/mg, a kcat of 4.0 s-1 for cAMP, and a KM of 1.6 mM, Vmax of 2.5 emol/min/mg, a kcat of 1.6 s-1 for cGMP, thus indicating that the substrate specificity of PDE8 is dominated by KM. The structure of the PDE8A1 catalytic domain has similar topology as those of other PDE families but contains two extra helices around Asn685-Thr710. Since this fragment is distant from the active site of the enzyme, its impact on the catalysis is unclear. The PDE8A1 catalytic domain is insensitive to the IBMX inhibition (IC50 = 700 eM). The unfavorable interaction of IBMX in the PDE8A1-IBMX structure suggests an important role of Tyr748 in the inhibitor binding. Indeed, the mutation of Tyr748 to phenylalanine increases the PDE8A1 sensitivity to several nonselective or family selective PDE inhibitors. Thus, the structural and mutagenesis studies provide not only insight into the enzymatic properties but also guidelines for design of PDE8 selective inhibitors.

  2. Kinetic and structural studies of phosphodiesterase-8A and implication on the inhibitor selectivity

    PubMed Central

    Wang, Huanchen; Yan, Zier; Yang, Serena; Cai, Jiwen; Robinson, Howard; Ke, Hengming

    2009-01-01

    Cyclic nucleotide phosphodiesterase-8 (PDE8) is a family of cAMP-specific enzymes and plays important roles in many biological processes, including T-cell activation, testosterone production, adrenocortical hyperplasia, and thyroid function. However, no PDE8 selective inhibitors are available for trial treatment of human diseases. Here we report kinetic properties of the highly active PDE8A1 catalytic domain prepared from refolding and its crystal structures in the unliganded and 3-isobutyl-1-methylxanthine (IBMX) bound forms at 1.9 and 2.1 Å resolutions, respectively. The PDE8A1 catalytic domain has KM of 1.8 μM, Vmax of 6.1 μmol/min/mg, kcat of 4.0 s−1 for cAMP, and KM of 1.6 mM, Vmax of 2.5 μmol/min/mg, kcat of 1.6 s−1 for cGMP, thus indicating that the substrate specificity of PDE8 is dominated by KM. The structure of the PDE8A1 catalytic domain has similar topology as those of other PDE families, but contains two extra helices around Asn685-Thr710. Since this fragment is distant from the active site of the enzyme, its impact on the catalysis is unclear. The PDE8A1 catalytic domain is insensitive to the IBMX inhibition (IC50 = 700 μM). The unfavorable interaction of IBMX in the PDE8A1-IBMX structure suggests an important role of Tyr748 in the inhibitor binding. Indeed, the mutation of Tyr748 to phenylalanine increases the PDE8A1 sensitivity to several non-selective or family-selective PDE inhibitors. Thus, the structural and mutagenesis studies provide not only insight into the enzymatic properties, but also guidelines for design of PDE8 selective inhibitors. PMID:18983167

  3. The role of cilostazol, a phosphodiesterase 3 inhibitor, on oocyte maturation and subsequent pregnancy in mice.

    PubMed

    Li, Min; Yu, Yang; Yan, Jie; Yan, Li-Ying; Zhao, Yue; Li, Rong; Liu, Ping; Hsueh, Aaron J; Qiao, Jie

    2012-01-01

    It is important to identify effective contraceptive drugs that cause minimal disruption to physiological processes. Phosphodiesterase 3 (PDE3) inhibitors suppress meiosis in oocytes by decreasing the level of cAMP and blocking the extrusion of the first polar body. In this study, we tested the PDE3 inhibitor, cilostazol, as a potential contraceptive agent. The effects of cilostazol treatment in vitro and in vivo on the suppression of oocyte maturation in a mouse model were investigated. The results indicated that treatment with increasing concentrations of cilostazol led to a dose-dependent arrest in meiosis progression. The effective in vitro concentration was 1 µM and was 300 mg/kg in vivo. The effect of cilostazol was reversible. After removal of the drug, meiosis resumed and mouse oocytes matured in vitro, and showed normal chromosome alignment and spindle organization. After fertilization using an ICSI method, the oocytes showed normal morphology, fertilization rate, embryo cleavage, blastocyst formation, and number of viable pups when compared with controls. The offspring showed similar body weight and fertility. In vivo, the mice became infertile if the drug was injected sequentially, and became pregnant following discontinuation of cilostazol. More importantly, no side effects of cilostazol were observed in treated female mice as demonstrated by blood pressure and heart rate monitoring. It is concluded that cilostazol, a drug routinely used for intermittent claudication, can effectively inhibit oocyte maturation in vitro and in vivo, does not affect the developmental potential of oocytes following drug removal and has few side effects in female mice treated with this drug. These findings suggest that cilostazol may be a potential new contraceptive agent that may facilitate an efficacy and safety study of this drug. PMID:22292006

  4. Cyclic nucleotide phosphodiesterase (PDE) isozymes as targets of the intracellular signalling network: benefits of PDE inhibitors in various diseases and perspectives for future therapeutic developments

    PubMed Central

    Keravis, Thérèse; Lugnier, Claire

    2012-01-01

    Cyclic nucleotide phosphodiesterases (PDEs) that specifically inactivate the intracellular messengers cAMP and cGMP in a compartmentalized manner represent an important enzyme class constituted by 11 gene-related families of isozymes (PDE1 to PDE11). Downstream receptors, PDEs play a major role in controlling the signalosome at various levels of phosphorylations and protein/protein interactions. Due to the multiplicity of isozymes, their various intracellular regulations and their different cellular and subcellular distributions, PDEs represent interesting targets in intracellular pathways. Therefore, the investigation of PDE isozyme alterations related to various pathologies and the design of specific PDE inhibitors might lead to the development of new specific therapeutic strategies in numerous pathologies. This manuscript (i) overviews the different PDEs including their endogenous regulations and their specific inhibitors; (ii) analyses the intracellular implications of PDEs in regulating signalling cascades in pathogenesis, exemplified by two diseases affecting cell cycle and proliferation; and (iii) discusses perspectives for future therapeutic developments. PMID:22014080

  5. 2',3'-cyclic nucleotide 3'-phosphodiesterases inhibit hepatitis B virus replication.

    PubMed

    Ma, Hui; Zhao, Xing-Liang; Wang, Xue-Yan; Xie, Xing-Wang; Han, Jin-Chao; Guan, Wen-Li; Wang, Qin; Zhu, Lin; Pan, Xiao-Ben; Wei, Lai

    2013-01-01

    2',3'-cyclic nucleotide 3'-phosphodiesterase (CNP) is a member of the interferon-stimulated genes, which includes isoforms CNP1 and CNP2. CNP1 is locally expressed in the myelin sheath but CNP2 is additionally expressed at low levels outside the nervous system. CNPs regulate multiple cellular functions and suppress protein production by association with polyadenylation of mRNA. Polyadenylation of Hepatitis B virus (HBV) RNAs is crucial for HBV replication. Whether CNPs interact with polyadenylation signal of HBV RNAs and interfere HBV replication is unknown. In this study, we evaluated expressions of CNP isoforms in hepatoma cell lines and their effects on HBV replication. We found that CNP2 is moderately expressed and gently responded to interferon treatment in HepG2, but not in Huh7 cells. The CNP1 and CNP2 potently inhibited HBV production by blocking viral proteins synthesis and reducing viral RNAs, respectively. In chronic hepatitis B patients, CNP was expressed in most of HBV-infected hepatocytes of liver specimens. Knockdown of CNP expression moderately improved viral production in the HepG2.2.15 cells treated with IFN-α. In conclusion, CNP might be a mediator of interferon-induced response against HBV. PMID:24260477

  6. Engineering of Bacillus subtilis Strains To Allow Rapid Characterization of Heterologous Diguanylate Cyclases and Phosphodiesterases

    PubMed Central

    Gao, Xiaohui; Dong, Xiao; Subramanian, Sundharraman; Matthews, Paige M.; Cooper, Caleb A.; Kearns, Daniel B.

    2014-01-01

    Microbial processes, including biofilm formation, motility, and virulence, are often regulated by changes in the available concentration of cyclic dimeric guanosine monophosphate (c-di-GMP). Generally, high c-di-GMP concentrations are correlated with decreased motility and increased biofilm formation and low c-di-GMP concentrations are correlated with an increase in motility and activation of virulence pathways. The study of c-di-GMP is complicated, however, by the fact that organisms often encode dozens of redundant enzymes that synthesize and hydrolyze c-di-GMP, diguanylate cyclases (DGCs), and c-di-GMP phosphodiesterases (PDEs); thus, determining the contribution of any one particular enzyme is challenging. In an effort to develop a facile system to study c-di-GMP metabolic enzymes, we have engineered a suite of Bacillus subtilis strains to assess the effect of individual heterologously expressed proteins on c-di-GMP levels. As a proof of principle, we characterized all 37 known genes encoding predicted DGCs and PDEs in Clostridium difficile using parallel readouts of swarming motility and fluorescence from green fluorescent protein (GFP) expressed under the control of a c-di-GMP-controlled riboswitch. We found that 27 of the 37 putative C. difficile 630 c-di-GMP metabolic enzymes had either active cyclase or phosphodiesterase activity, with agreement between our motility phenotypes and fluorescence-based c-di-GMP reporter. Finally, we show that there appears to be a threshold level of c-di-GMP needed to inhibit motility in Bacillus subtilis. PMID:25085482

  7. cAMP prevents TNF-induced apoptosis through inhibiting DISC complex formation in rat hepatocytes

    SciTech Connect

    Bhattacharjee, Rajesh; Xiang, Wenpei; Wang, Yinna; Zhang, Xiaoying

    2012-06-22

    Highlights: Black-Right-Pointing-Pointer cAMP blocks cell death induced by TNF and actinomycin D in cultured hepatocytes. Black-Right-Pointing-Pointer cAMP blocks NF-{kappa}B activation induced by TNF and actinomycin D. Black-Right-Pointing-Pointer cAMP blocks DISC formation following TNF and actinomycin D exposure. Black-Right-Pointing-Pointer cAMP blocks TNF signaling at a proximal step. -- Abstract: Tumor necrosis factor {alpha} (TNF) is a pleiotropic proinflammatory cytokine that plays a role in immunity and the control of cell proliferation, cell differentiation, and apoptosis. The pleiotropic nature of TNF is due to the formation of different signaling complexes upon the binding of TNF to its receptor, TNF receptor type 1 (TNFR1). TNF induces apoptosis in various mammalian cells when the cells are co-treated with a transcription inhibitor like actinomycin D (ActD). When TNFR1 is activated, it recruits an adaptor protein, TNF receptor-associated protein with death domain (TRADD), through its cytoplasmic death effector domain (DED). TRADD, in turn, recruits other signaling proteins, including TNF receptor-associated protein 2 (TRAF2) and receptor-associated protein kinase (RIPK) 1, to form a complex. Subsequently, this complex combines with FADD and procaspase-8, converts into a death-inducing signaling complex (DISC) to induce apoptosis. Cyclic AMP (cAMP) is a second messenger that regulates various cellular processes such as cell proliferation, gene expression, and apoptosis. cAMP analogues are reported to act as anti-apoptotic agents in various cell types, including hepatocytes. We found that a cAMP analogue, dibutyryl cAMP (db-cAMP), inhibits TNF + ActD-induced apoptosis in rat hepatocytes. The protein kinase A (PKA) inhibitor KT-5720 reverses this inhibitory effect of cAMP on apoptosis. Cytoprotection by cAMP involves down-regulation of various apoptotic signal regulators like TRADD and FADD and inhibition of caspase-8 and caspase-3 cleavage. We also found

  8. Camp Blanding Lightning Mapping Array

    NASA Technical Reports Server (NTRS)

    Blakeslee,Richard; Christian, Hugh; Bailey, Jeffrey; Hall, John; Uman, Martin; Jordan, Doug; Krehbiel, Paul; Rison, William; Edens, Harald

    2011-01-01

    A seven station, short base-line Lightning Mapping Array was installed at the Camp Blanding International Center for Lightning Research and Testing (ICLRT) during April 2011. This network will support science investigations of Terrestrial Gamma-Ray Flashes (TGFs) and lightning initiation using rocket triggered lightning at the ICLRT. The network operations and data processing will be carried out through a close collaboration between several organizations, including the NASA Marshall Space Flight Center, University of Alabama in Huntsville, University of Florida, and New Mexico Tech. The deployment was sponsored by the Defense Advanced Research Projects Agency (DARPA). The network does not have real-time data dissemination. Description, status and plans will be discussed.

  9. The effect of Sildenafil on human platelet secretory function is controlled by a complex interplay between phosphodiesterases 2, 3 and 5.

    PubMed

    Dunkern, Torsten R; Hatzelmann, Armin

    2005-03-01

    Human platelets contain the cyclic nucleotide-hydrolyzing phosphodiesterases (PDEs) 2, 3 and 5. The cGMP-PDE5 inhibitors Sildenafil and Zaprinast have been demonstrated to potentiate the anti-platelet aggregatory effect of NO donors like sodium nitroprusside (SNP) in vitro but the mechanisms of Sildenafil's action on the secretory function of human platelets have not been analysed in detail. In the present paper, we show (1) that both compounds potentiate the SNP-induced increase in cGMP in human platelets concentration-dependently. (2) However, whereas Sildenafil plus SNP treatment only partially inhibits thrombin-induced release of serotonin, the less selective Zaprinast plus SNP cause a complete inhibition. (3) The inhibition mediated by Sildenafil plus SNP is limited to low compound concentrations at which cAMP levels are increased, probably due to cGMP-mediated inhibition of PDE3. (4) High concentrations of Sildenafil (plus SNP) neither affect cAMP levels, likely due to the activation of PDE2, nor inhibits the release of serotonin. Thus, increases in both cyclic nucleotides seem to control platelet function. (5) Accordingly, treatment with increasing concentrations of Sildenafil plus SNP and a selective PDE2 inhibitor, which by its own has no effect, induced a concentration-dependent increase in cAMP and complete inhibition of platelet activation. In summary, our data indicate that Sildenafil inhibits secretory function of human platelets at least in part due to the cGMP-mediated effects on intracellular cAMP and that entire inhibition of serotonin release from thrombin-activated platelets is controlled by both cyclic nucleotides. PMID:15567064

  10. Vasoactive intestinal peptide synergistically stimulates DNA synthesis in mouse 3T3 cells: Role of cAMP, Ca sup 2+ , and protein kinase C

    SciTech Connect

    Zurier, B.B.; Kozma, M.; Sinnett-Smith, J.; Rozengurt, E. )

    1988-05-01

    Vasoactive intestinal peptide synergistically stimulated initiation of DNA synthesis in Swiss 3T3 cells. The peptide stimulated ({sup 3}H)thymidine incorporation in the presence of insulin and either forskolin or an inhibitor of cAMP phosphodiesterase in a concentration-dependent manner. Half-maximal effect was obtained at 1 nM. At mitogenic concentrations, VIP stimulated a marked accumulation (eightfold) of cAMP. In contrast to other growth-promoting neuropeptides, VIP did not induce an increase in cytosolic free Ca{sup 2+} or the activation of protein kinase C. The authors conclude that neuropeptides can modulate long-term cell proliferation through multiple signaling pathways.

  11. Sequence analysis and characterization of a 40-kilodalton Borrelia hermsii glycerophosphodiester phosphodiesterase homolog.

    PubMed Central

    Shang, E S; Skare, J T; Erdjument-Bromage, H; Blanco, D R; Tempst, P; Miller, J N; Lovett, M A

    1997-01-01

    We report the purification, molecular cloning, and characterization of a 40-kDa glycerophosphodiester phosphodiesterase homolog from Borrelia hermsii. The 40-kDa protein was solubilized from whole organisms with 0.1% Triton X-100, phase partitioned into the Triton X-114 detergent phase, and purified by fast-performance liquid chromatography (FPLC). The gene encoding the 40-kDa protein was cloned from a B. hermsii chromosomal DNA lambda EXlox expression library and identified by using affinity antibodies generated against the purified native protein. The deduced amino acid sequence included a 20-amino-acid signal peptide encoding a putative leader peptidase II cleavage site, indicating that the 40-kDa protein was a lipoprotein. Based on significant homology (31 to 52% identity) of the 40-kDa protein to glycerophosphodiester phosphodiesterases of Escherichia coli (GlpQ), Bacillus subtilis (GlpQ), and Haemophilus influenzae (Hpd; protein D), we have designated this B. hermsii 40-kDa lipoprotein a glycerophosphodiester phosphodiesterase (Gpd) homolog, the first B. hermsii lipoprotein to have a putative functional assignment. A nonlipidated form of the Gpd homolog was overproduced as a fusion protein in E. coli BL21(DE3)(pLysE) and was used to immunize rabbits to generate specific antiserum. Immunoblot analysis with anti-Gpd serum recognized recombinant H. influenzae protein D, and conversely, antiserum to H. influenzae protein D recognized recombinant B. hermsii Gpd (rGpd), indicating antigenic conservation between these proteins. Antiserum to rGpd also identified native Gpd as a constituent of purified outer membrane vesicles prepared from B. hermsii. Screening of other pathogenic spirochetes with anti-rGpd serum revealed the presence of antigenically related proteins in Borrelia burgdorferi, Treponema pallidum, and Leptospira kirschneri. Further sequence analysis both upstream and downstream of the Gpd homolog showed additional homologs of glycerol metabolism

  12. cAMP signaling in neurons: patterns of neuronal expression and intracellular localization for a novel protein, AKAP 150, that anchors the regulatory subunit of cAMP-dependent protein kinase II beta.

    PubMed Central

    Glantz, S B; Amat, J A; Rubin, C S

    1992-01-01

    In mammalian brain, physiological signals carried by cyclic AMP (cAMP) seem to be targeted to effector sites via the tethering of cAMP-dependent protein kinase II beta (PKAII beta) to intracellular structures. Recently characterized A kinase anchor proteins (AKAPs) are probable mediators of the sequestration of PKAII beta because they contain a high-affinity binding site for the regulatory subunit (RII beta) of the kinase and a distinct intracellular targeting domain. To establish a cellular basis for this targeting mechanism, we have employed immunocytochemistry to 1) identify the types of neurons that are enriched in AKAPs, 2) determine the primary intracellular location of the anchor protein, and 3) demonstrate that an AKAP and RII beta are coenriched and colocalized in neurons that utilize the adenylate cyclase-cyclic AMP-dependent protein kinase (PKA) signaling pathway. Antibodies directed against rat brain AKAP 150 were used to elucidate the regional, cellular and intracellular distribution of a prototypic anchor protein in the CNS. AKAP 150 is abundant in Purkinje cells and in neurons of the olfactory bulb, basal ganglia, cerebral cortex, and other forebrain regions. In contrast, little AKAP 150 is detected in neurons of the thalamus, hypothalamus, midbrain, and hindbrain. A high proportion of total AKAP 150 is concentrated in primary branches of dendrites, where it is associated with microtubules. We also discovered that the patterns of accumulation and localization of RII beta (and PKAII beta) in brain are similar to those of AKAP 150. The results suggest that bifunctional AKAP 150 tethers PKAII beta to the dendritic cytoskeleton, thereby creating a discrete target site for the reception and propagation of signals carried by cAMP. Images PMID:1333841

  13. Characterization of particulate cyclic nucleotide phosphodiesterases from bovine brain: Purification of a distinct cGMP-stimulated isoenzyme

    SciTech Connect

    Murashima, Seiko; Tanaka, Takayuki; Hockman, S.; Manganiello, V. )

    1990-06-05

    In the absence of detergent, {approx}80-85% of the total cGMP-stimulated phosphodiesterase (PDE) activity in bovine brain was associated with washed particulate fractions; {approx}85-90% of the calmodulin-sensitive PDE was soluble. Particulate cGMP-stimulated PDE was higher in cerebral cortical gray matter than in other regions. Homogenization of the brain particulate fraction in 1% Lubrol increased cGMP-stimulated activity {approx}100% and calmodulin-stimulated {approx}400-500%. Although 1% Lubrol readily solubilized these PDE activities, {approx}75% of the cAMP PDE activity (0.5 {mu}M ({sup 3}H)cAMP) that was not affected by cGMP was not solubilized. This cAMP PDE activity was very sensitive to inhibition by Rolipram but not cilostamide. Thus, three different PDE types, i.e., cGMP stimulated, calmodulin sensitive, and Rolipram inhibited, are associated in different ways with crude bovine brain particulate fractions. The brain enzyme exhibited a slightly greater subunit M{sub r} than did soluble forms from calf liver or bovine brain, as evidenced by protein staining or immunoblotting after polyacrylamide gel electrophoresis under denaturing conditions. Incubation of brain particulate and liver soluble cGMP-stimulated PDEs with V{sub 8} protease produced several peptides of similar size, as well as at least two distinct fragments of {approx}27 kDa from the brain and {approx}23 kDa from the liver enzyme. After photolabeling in the presence of ({sup 32}P)cGMP and digestion with V{sub 8} protease, ({sup 32}P)cGMP in each PDE was predominantly recovered with a peptide of {approx}14 kDa. All of these observations are consistent with the existence of at least two discrete forms (isoenzymes) of cGMP-stimulated PDE.

  14. Depletion of tyrosyl DNA phosphodiesterase 2 activity enhances etoposide-mediated double-strand break formation and cell killing.

    PubMed

    Kont, Yasemin Saygideger; Dutta, Arijit; Mallisetty, Apurva; Mathew, Jeena; Minas, Tsion; Kraus, Christina; Dhopeshwarkar, Priyanka; Kallakury, Bhaskar; Mitra, Sankar; Üren, Aykut; Adhikari, Sanjay

    2016-07-01

    DNA topoisomerase 2 (Top2) poisons, including common anticancer drugs etoposide and doxorubicin kill cancer cells by stabilizing covalent Top2-tyrosyl-DNA 5'-phosphodiester adducts and DNA double-strand breaks (DSBs). Proteolytic degradation of the covalently attached Top2 leaves a 5'-tyrosylated blocked termini which is removed by tyrosyl DNA phosphodiesterase 2 (TDP2), prior to DSB repair through non-homologous end joining (NHEJ). Thus, TDP2 confers resistance of tumor cells to Top2-poisons by repairing such covalent DNA-protein adducts, and its pharmacological inhibition could enhance the efficacy of Top2-poisons. We discovered NSC111041, a selective inhibitor of TDP2, by optimizing a high throughput screening (HTS) assay for TDP2's 5'-tyrosyl phosphodiesterase activity and subsequent validation studies. We found that NSC111041 inhibits TDP2's binding to DNA without getting intercalated into DNA and enhanced etoposide's cytotoxicity synergistically in TDP2-expressing cells but not in TDP2 depleted cells. Furthermore, NSC111041 enhanced formation of etoposide-induced γ-H2AX foci presumably by affecting DSB repair. Immuno-histochemical analysis showed higher TDP2 expression in a sub-set of different type of tumor tissues. These findings underscore the feasibility of clinical use of suitable TDP2 inhibitors in adjuvant therapy with Top2-poisons for a sub-set of cancer patients with high TDP2 expression. PMID:27235629

  15. "It's not Just Camp!": Understanding the Meaning of Children's Cancer Camps for Children and Families.

    PubMed

    Laing, Catherine M; Moules, Nancy J

    2016-01-01

    The purpose of this philosophical hermeneutic inquiry was to understand the meaning of children's cancer camps for the child with cancer and the family. Six childhood cancer families and 5 cancer camp counselors were interviewed, in order to bring understanding to this topic. Findings from this research revealed that camp means different things for different families, and that much is at play in the cancer camp experience: the healing and developmental power of play, finding acceptance and fit, grief as something to live with versus "get over," storytelling as a means of reshaping and understanding traumatic experiences, and the solidarity of the community as one that creates intense, healing bonds. Children's cancer camps, we conclude, should be considered a necessity, versus a luxury, and could even be thought of as a psychosocial intervention for some children and families. Barriers such as structure of funding and access to resources are present and likely due to the separateness of camps from hospital programs. PMID:25643975

  16. Why Do Counselors Return to Work at Camp?

    ERIC Educational Resources Information Center

    Becker, William A.

    The reasons that counselors in resident summer camps return to work are explored, taking into account the differences between private and agency camps, and differing attitudes of male and female camp counselors. A random sample of returning counselors at 15 private and 15 agency camps in the Northeast were selected for the study. Six attitudinal…

  17. Whatever Happened to School Camping? An Occasional Paper.

    ERIC Educational Resources Information Center

    Donaldson, George W.; Donaldson, Louise E.

    1982-01-01

    School camping began with three movements in American society: children's camping, which has been traced back as far as 1823; the Nature Study Movement, which is thought to have begun as early as 1839; and the holistic "new education." In 1940, L. B. Sharp opened National Camp in New Jersey and Julian W. Smith initiated the Clear Lake camp near…

  18. Institutionalized Adolescents' Perceptions of a Summer Camp Program

    ERIC Educational Resources Information Center

    Herr, David E.

    1977-01-01

    Describes the use of the facilities of Camp Easter Seal, Virginia, for institutionalized adolescents from different hospitals in Virginia. Also includes the attitudes of the patients toward their camping experience, their camp counselors, and what they learned from their camping experience. (Author/RK)

  19. Summer Camps: A Fun Way to Reinforce Math Skills

    ERIC Educational Resources Information Center

    Tichenor, Mercedes; Plavchan, Joan

    2010-01-01

    Faculty members from a university teacher education department partnered with a local school district to develop a summer camp program for children at-risk. This four week summer camp for elementary students provides reading and math intervention to rising first graders. This article discusses the math aspects of the camp, including camp lessons,…

  20. Alabama State Lodging Tax: A Lesson for All Camps.

    ERIC Educational Resources Information Center

    Sieckmann, Amy

    2003-01-01

    A dispute between an Alabama business conference center and a nearby camp prompted the state's revenue department to charge camps an old lodging tax that had never been applied to them before. The state camping association members worked together to have the tax law and regulations amended so that nonprofit camps were exempt from the tax in most…

  1. Site and Facilities: A Resource Book for Camps.

    ERIC Educational Resources Information Center

    Ball, Armand, Ed.; Ball, Beverly, Ed.

    This resource book draws together articles on the development and maintenance of camp sites and facilities. The articles, previously published by "Camping Magazine" and "Journal of Christian Camping," cover (1) site planning and long-range development, including redesigning multiple camp facilities for year-round programs, remodeling and…

  2. Accreditation Standards for Camp Programs and Services. Revised 1998 Edition.

    ERIC Educational Resources Information Center

    American Camping Association, Martinsville, IN.

    The American Camping Association (ACA) accreditation program aims to educate camp owners and directors in the administration of key aspects of camp operation, particularly those related to program quality and the health and safety of campers and staff, and to assist the public in selecting camps that meet industry-accepted and…

  3. (Compendium of State Laws and Regulations for Youth Camps).

    ERIC Educational Resources Information Center

    Brookhiser, Judy, Comp.; van der Smissen, Betty, Comp.

    State laws and regulations applicable to youth camp operations provided by state agencies are organized in this Compendium under ten major headings; personnel; program safety; personal health, first aid, and medical services; site and facilities; sanitation; food service; transportation; primitive camping and out-of-camp trips; day camping; and…

  4. Design and Development Issues for Educational Robotics Training Camps

    ERIC Educational Resources Information Center

    Ucgul, Memet; Cagiltay, Kursat

    2014-01-01

    The aim of this study is to explore critical design issues for educational robotics training camps and to describe how these factors should be implemented in the development of such camps. For this purpose, two robotics training camps were organized for elementary school students. The first camp had 30 children attendees, and the second had 22. As…

  5. cAMP prevents TNF-induced apoptosis through inhibiting DISC complex formation in rat hepatocytes

    PubMed Central

    Bhattacharjee, Rajesh; Xiang, Wenpei; Wang, Yinna; Zhang, Xiaoying; Billiar, Timothy R.

    2013-01-01

    Tumor necrosis factor α (TNF) is a pleiotropic proinflammatory cytokine that plays a role in immunity and the control of cell proliferation, cell differentiation, and apoptosis. The pleiotropic nature of TNF is due to the formation of different signaling complexes upon the binding of TNF to its receptor, TNF receptor type 1 (TNFR1). TNF induces apoptosis in various mammalian cells when the cells are co-treated with a transcription inhibitor like actinomycin D (ActD). When TNFR1 is activated, it recruits an adaptor protein, TNF receptor-associated protein with death domain (TRADD), through its cytoplasmic death effector domain (DED). TRADD, in turn, recruits other signaling proteins, including TNF receptor-associated protein 2 (TRAF2) and receptor-associated protein kinase (RIPK) 1, to form a complex. Subsequently, this complex combines with FADD and procaspase-8, converts into a death-inducing signaling complex (DISC) to induce apoptosis. Cyclic AMP (cAMP) is a second messenger that regulates various cellular processes such as cell proliferation, gene expression, and apoptosis. cAMP analogues are reported to act as anti-apoptotic agents in various cell types, including hepatocytes. We found that a cAMP analogue, dibutyryl cAMP (db-cAMP), inhibits TNF + ActD-induced apoptosis in rat hepatocytes. The protein kinase A (PKA) inhibitor KT-5720 reverses this inhibitory effect of cAMP on apoptosis. Cytoprotection by cAMP involves down-regulation of various apoptotic signal regulators like TRADD and FADD and inhibition of caspase-8 and caspase-3 cleavage. We also found that cAMP exerts its affect at the proximal level of TNF signaling by inhibiting the formation of the DISC complex upon the binding of TNF to TNFR1. In conclusion, our study shows that cAMP prevents TNF + ActD-induced apoptosis in rat hepatocytes by inhibiting DISC complex formation. PMID:22634003

  6. A Lightning Safety Primer for Camps.

    ERIC Educational Resources Information Center

    Attarian, Aram

    1992-01-01

    Provides the following information about lightning, which is necessary for camp administrators and staff: (1) warning signs of lightning; (2) dangers of lightning; (3) types of lightning injuries; (4) prevention of lightning injury; and (5) helpful training tips. (KS)

  7. Retarded Children at Camp with Normal Children

    ERIC Educational Resources Information Center

    Flax, Norman; Peters, Edward N.

    1969-01-01

    Statistical analysis of data from written forms and scales (designed to measure children's behavior in groups), observations, and interviews indicated that many educalble mentally retarded children can participate successfully in camp activities with normal children. (DR)

  8. An asymmetric dizinc phosphodiesterase model with phenolate and carboxylate bridges.

    PubMed

    Chen, Jingwen; Wang, Xiaoyong; Zhu, Yangguang; Lin, Jun; Yang, Xiaoliang; Li, Yizhi; Lu, Yi; Guo, Zijian

    2005-05-16

    A phosphodiesterase model with two zinc centers has been synthesized and characterized. The compound, [Zn(2)(L(-)(2H))(AcO)(H(2)O)](PF(6)).2H(2)O (Zn(2)L'), was formed using an "end-off" type compartmental ligand, 2,6-bis{[(2-pyridylmethyl)(2-hydroxyethyl)amino]methyl}-4-methylphenol (L), and zinc acetate dihydrate. The X-ray crystallographic analysis shows that Zn(2)L' contains a mu-acetato-mu-cresolato-dizinc(II) core comprised of a quasi-trigonal bipyramidal Zn and a distorted octahedral Zn, and the distance between them is 3.421 Angstroms which is close to the dizinc distance in related natural metalloenzymes. Phosphodiesterase activity of Zn(2)L' was investigated using bis(4-nitrophenyl) phosphate (BNPP) as the substrate. The pH dependence of the BNPP cleavage in aqueous buffer media shows a sigmoid-shaped pH-k(obs) profile with an inflection point around pH 7.13 which is close to the first pK(a) value of 7.20 for Zn(2)L' obtained from the potentiometric titration. The catalytic rate constant (k(cat)) is 4.60 x 10(-6) s(-1) at pH 7.20 and 50 degrees C which is ca. 10(5)-fold higher than that of the uncatalyzed reaction. The deprotonated alcoholic group appended on Zn(2)L' is responsible for the cleavage reaction. The possible mechanism for the BNPP cleavage promoted by Zn(2)L' is proposed on the basis of kinetic and spectral analysis. The dizinc complex formed in situ in anhydrous DMSO exhibits a similar ability to cleave BNPP. This study provides a less common example for the phosphodiesterase model in which the metal-bound alkoxide is the nucleophile. PMID:15877422

  9. Thromboangiitis obliterans successfully treated with phosphodiesterase type 5 inhibitors.

    PubMed

    Abeles, Aryeh M; Nicolescu, Mirela; Pinchover, Zachary; Abeles, Micha

    2014-08-01

    Thromboangiitis obliterans, or Buerger's disease, is a non-atherosclerotic, segmental, inflammatory disease affecting the small- and medium-sized vessels of the distal extremities. Other than discontinuation of tobacco, there is no standard-of-care treatment. Although two randomized trials have demonstrated a role for intravenous iloprost, no oral drug has yet been demonstrated to be effective in treating thromboangiitis obliterans. We present the first three reported cases of thromboangiitis obliterans successfully treated with phosphodiesterase type 5 inhibitors, followed by a discussion of the rationale for the use of these agents in thromboangiitis obliterans. PMID:24000082

  10. Standards for Day and Resident Camps: The Accreditation Programs of the American Camping Association. 1990 Edition, Revised 1993.

    ERIC Educational Resources Information Center

    American Camping Association, Martinsville, IN.

    This book outlines standards that provide a basis for camp accreditation and site approval by the American Camping Association (ACA), Inc. The purpose of the ACA accreditation programs is to educate camp owners and directors in the administration of key aspects of camp operation, particularly those related to program quality and the health and…

  11. Phosphodiesterase 4D acts downstream of Neuropilin to control Hedgehog signal transduction and the growth of medulloblastoma.

    PubMed

    Ge, Xuecai; Milenkovic, Ljiljana; Suyama, Kaye; Hartl, Tom; Purzner, Teresa; Winans, Amy; Meyer, Tobias; Scott, Matthew P

    2015-01-01

    Alterations in Hedgehog (Hh) signaling lead to birth defects and cancers including medulloblastoma, the most common pediatric brain tumor. Although inhibitors targeting the membrane protein Smoothened suppress Hh signaling, acquired drug resistance and tumor relapse call for additional therapeutic targets. Here we show that phosphodiesterase 4D (PDE4D) acts downstream of Neuropilins to control Hh transduction and medulloblastoma growth. PDE4D interacts directly with Neuropilins, positive regulators of Hh pathway. The Neuropilin ligand Semaphorin3 enhances this interaction, promoting PDE4D translocation to the plasma membrane and cAMP degradation. The consequent inhibition of protein kinase A (PKA) enhances Hh transduction. In the developing cerebellum, genetic removal of Neuropilins reduces Hh signaling activity and suppresses proliferation of granule neuron precursors. In mouse medulloblastoma allografts, PDE4D inhibitors suppress Hh transduction and inhibit tumor growth. Our findings reveal a new regulatory mechanism of Hh transduction, and highlight PDE4D as a promising target to treat Hh-related tumors. PMID:26371509

  12. Phosphodiesterase 4D acts downstream of Neuropilin to control Hedgehog signal transduction and the growth of medulloblastoma

    PubMed Central

    Ge, Xuecai; Milenkovic, Ljiljana; Suyama, Kaye; Hartl, Tom; Purzner, Teresa; Winans, Amy; Meyer, Tobias; Scott, Matthew P

    2015-01-01

    Alterations in Hedgehog (Hh) signaling lead to birth defects and cancers including medulloblastoma, the most common pediatric brain tumor. Although inhibitors targeting the membrane protein Smoothened suppress Hh signaling, acquired drug resistance and tumor relapse call for additional therapeutic targets. Here we show that phosphodiesterase 4D (PDE4D) acts downstream of Neuropilins to control Hh transduction and medulloblastoma growth. PDE4D interacts directly with Neuropilins, positive regulators of Hh pathway. The Neuropilin ligand Semaphorin3 enhances this interaction, promoting PDE4D translocation to the plasma membrane and cAMP degradation. The consequent inhibition of protein kinase A (PKA) enhances Hh transduction. In the developing cerebellum, genetic removal of Neuropilins reduces Hh signaling activity and suppresses proliferation of granule neuron precursors. In mouse medulloblastoma allografts, PDE4D inhibitors suppress Hh transduction and inhibit tumor growth. Our findings reveal a new regulatory mechanism of Hh transduction, and highlight PDE4D as a promising target to treat Hh-related tumors. DOI: http://dx.doi.org/10.7554/eLife.07068.001 PMID:26371509

  13. Inhibition of human cAMP-phosphodiesterase as a mechanism of the spasmolytic effect of Matricaria recutita L.

    PubMed

    Maschi, Omar; Cero, Esther Dal; Galli, Germana V; Caruso, Donatella; Bosisio, Enrica; Dell'Agli, Mario

    2008-07-01

    Mechanisms underlying the spasmolytic activity of chamomile still remain unclear. Inhibition of cAMP- and cGMP-phosphodiesterases (PDE) is one of the mechanisms operated by spasmolytic drugs. In this study, the effect of chamomile on PDE was investigated. Human platelet cAMP-PDE and recombinant PDE5A1 were assayed in the presence of infusions prepared from sifted flowers and capitula. LC-ESI-MS/MS analysis showed different compositions in infusions made with sifted flowers and capitula. Chamomile inhibited cAMP-PDE activity (IC50 = 17.9-40.5 microg/mL), while cGMP-PDE5 was less affected (-15% at 50 microg/mL). Among the individual compounds tested, only flavonoids showed an inhibitory effect (IC50 = 1.3-14.9 microM), contributing to around 39% of the infusion inhibition; other compounds responsible for cAMP-PDE inhibition still remain unknown. Although experimental evidence supporting the use of chamomile for gastrointestinal minor spasms dates back to the fifties, cAMP-PDE inhibition as a likely mechanism underlying the spasmolytic activity is reported for the first time. PMID:18553893

  14. Surface plasmon resonance biosensor assay for the analysis of small-molecule inhibitor binding to human and parasitic phosphodiesterases.

    PubMed

    Siderius, Marco; Shanmugham, Anitha; England, Paul; van der Meer, Tiffany; Bebelman, Jan Paul; Blaazer, Antoni R; de Esch, Iwan J P; Leurs, Rob

    2016-06-15

    In the past decade, surface plasmon resonance (SPR) biosensor-based technology has been exploited more and more to characterize the interaction between drug targets and small-molecule modulators. Here, we report the successful application of SPR methodology for the analysis of small-molecule binding to two therapeutically relevant cAMP phosphodiesterases (PDEs), Trypanosoma brucei PDEB1 which is implicated in African sleeping sickness and human PDE4D which is implicated in a plethora of disease conditions including inflammatory pulmonary disorders such as asthma, chronic obstructive pulmonary disease and central nervous system (CNS) disorders. A protocol combining the use of directed capture using His-tagged PDE_CDs with covalent attachment to the SPR surface was developed. This methodology allows the determination of the binding kinetics of small-molecule PDE inhibitors and also allows testing their specificity for the two PDEs. The SPR-based assay could serve as a technology platform for the development of highly specific and high-affinity PDE inhibitors, accelerating drug discovery processes. PMID:27033007

  15. Inhibition of phosphodiesterase-4 reverses the cognitive dysfunction and oxidative stress induced by Aβ25-35 in rats.

    PubMed

    Zhuo, Yeye; Guo, Haibiao; Cheng, Yufang; Wang, Chuang; Wang, Canmao; Wu, Jingang; Zou, Zhengqiang; Gan, Danna; Li, Yiwen; Xu, Jiangping

    2016-08-01

    Phosphodiesterase-4 (PDE4) inhibitors prevent the breakdown of the second messenger cAMP and have been demonstrated to improve learning in several animal models of cognition. In this study, we explored the antioxidative effects of rolipram in Alzheimer's disease (AD) by using bilateral Aβ25-35 injection into the hippocampus of rats, which were used as an AD model. Rats received 3 intraperitoneal (i.p.) doses of rolipram (0.1, 0.5 and 1.25 mg/kg) daily after the injection of Aβ25-35 for 25 days. Chronic administration of rolipram prevented the memory impairments induced by Aβ25-35, as assessed using the passive avoidance test and the Morris water maze test. Furthermore, rolipram significantly reduced the oxidative stress induced by Aβ25-35, as evidenced by the decrease in the levels of reactive oxygen species (ROS) and malondialdehyde (MDA), and restored the reduced GSH levels and superoxide dismutase (SOD) activity. Moreover, western blotting and real-time reverse transcription polymerase chain reaction (RT-PCR) analysis showed that rolipram remarkably upregulated thioredoxin (Trx) and inhibited the inducible nitric oxide synthase/nitric oxide (iNOS/NO) pathway in the hippocampus. These results demonstrated that rolipram improved the learning and memory abilities in an Aβ25-35-induced AD rat model. The mechanism underlying these effects may be due to the noticeable antioxidative effects of rolipram. PMID:26920899

  16. The Characterization of Escherichia coli CpdB as a Recombinant Protein Reveals that, besides Having the Expected 3´-Nucleotidase and 2´,3´-Cyclic Mononucleotide Phosphodiesterase Activities, It Is Also Active as Cyclic Dinucleotide Phosphodiesterase

    PubMed Central

    López-Villamizar, Iralis; Cabezas, Alicia; Pinto, Rosa María; Canales, José; Ribeiro, João Meireles; Cameselle, José Carlos; Costas, María Jesús

    2016-01-01

    Endogenous cyclic diadenylate phosphodiesterase activity was accidentally detected in lysates of Escherichia coli BL21. Since this kind of activity is uncommon in Gram-negative bacteria, its identification was undertaken. After partial purification and analysis by denaturing gel electrophoresis, renatured activity correlated with a protein identified by fingerprinting as CpdB (cpdB gene product), which is annotated as 3´-nucleotidase / 2´,3´-cyclic-mononucleotide phosphodiesterase, and it is synthesized as a precursor protein with a signal sequence removable upon export to the periplasm. It has never been studied as a recombinant protein. The coding sequence of mature CpdB was cloned and expressed as a GST fusion protein. The study of the purified recombinant protein, separated from GST, confirmed CpdB annotation. The assay of catalytic efficiencies (kcat/Km) for a large substrate set revealed novel CpdB features, including very high efficiencies for 3´-AMP and 2´,3´-cyclic mononucleotides, and previously unknown activities on cyclic and linear dinucleotides. The catalytic efficiencies of the latter activities, though low in relative terms when compared to the major ones, are far from negligible. Actually, they are perfectly comparable to those of the ‘average’ enzyme and the known, bona fide cyclic dinucleotide phosphodiesterases. On the other hand, CpdB differs from these enzymes in its extracytoplasmic location and in the absence of EAL, HD and DHH domains. Instead, it contains the domains of the 5´-nucleotidase family pertaining to the metallophosphoesterase superfamily, although CpdB lacks 5´-nucleotidase activity. The possibility that the extracytoplasmic activity of CpdB on cyclic dinucleotides could have physiological meaning is discussed. PMID:27294396

  17. PKA and PDE4D3 anchoring to AKAP9 provides distinct regulation of cAMP signals at the centrosome

    PubMed Central

    Terrin, Anna; Monterisi, Stefania; Stangherlin, Alessandra; Zoccarato, Anna; Koschinski, Andreas; Surdo, Nicoletta C.; Mongillo, Marco; Sawa, Akira; Jordanides, Niove E.; Mountford, Joanne C.

    2012-01-01

    Previous work has shown that the protein kinase A (PKA)–regulated phosphodiesterase (PDE) 4D3 binds to A kinase–anchoring proteins (AKAPs). One such protein, AKAP9, localizes to the centrosome. In this paper, we investigate whether a PKA–PDE4D3–AKAP9 complex can generate spatial compartmentalization of cyclic adenosine monophosphate (cAMP) signaling at the centrosome. Real-time imaging of fluorescence resonance energy transfer reporters shows that centrosomal PDE4D3 modulated a dynamic microdomain within which cAMP concentration selectively changed over the cell cycle. AKAP9-anchored, centrosomal PKA showed a reduced activation threshold as a consequence of increased autophosphorylation of its regulatory subunit at S114. Finally, disruption of the centrosomal cAMP microdomain by local displacement of PDE4D3 impaired cell cycle progression as a result of accumulation of cells in prophase. Our findings describe a novel mechanism of PKA activity regulation that relies on binding to AKAPs and consequent modulation of the enzyme activation threshold rather than on overall changes in cAMP levels. Further, we provide for the first time direct evidence that control of cell cycle progression relies on unique regulation of centrosomal cAMP/PKA signals. PMID:22908311

  18. 2',3'-Cyclic nucleotide 3'-phosphodiesterase: a novel RNA-binding protein that inhibits protein synthesis.

    PubMed

    Gravel, Michel; Robert, Francis; Kottis, Vicky; Gallouzi, Imed-Eddine; Pelletier, Jerry; Braun, Peter E

    2009-04-01

    2',3'-Cyclic nucleotide 3'-phosphodiesterase (CNP) is one of the earliest myelin-related proteins to be specifically expressed in differentiating oligodendrocytes (ODCs) in the central nervous system (CNS) and is implicated in myelin biogenesis. CNP possesses an in vitro enzymatic activity, whose in vivo relevance remains to be defined, because substrates with 2',3,-cyclic termini have not yet been identified. To characterize CNP function better, we previously determined the structure of the CNP catalytic domain by NMR. Interestingly, the structure is remarkably similar to the plant cyclic nucleotide phosphodiesterase (CPDase) from A. thaliana and the bacterial 2'-5' RNA ligase from T. thermophilus, which are known to play roles in RNA metabolism. Here we show that CNP is an RNA-binding protein. Furthermore, by using precipitation analyses, we demonstrate that CNP associates with poly(A)(+) mRNAs in vivo and suppresses translation in vitro in a dose-dependent manner. With SELEX, we isolated RNA aptamers that can suppress the inhibitory effect of CNP on translation. We also demonstrate that CNP1 can bridge an association between tubulin and RNA. These results suggest that CNP1 may regulate expression of mRNAs in ODCs of the CNS. PMID:19021295

  19. A PKG inhibitor increases Ca(2+)-regulated exocytosis in guinea pig antral mucous cells: cAMP accumulation via PDE2A inhibition.

    PubMed

    Tanaka, Saori; Tanaka, Rina; Harada, Saeko; Kohda, Yuka; Matsumura, Hitoshi; Shimamoto, Chikao; Sawabe, Yukinori; Marunaka, Yoshinori; Kuwabara, Hiroko; Takahashi, Yuko; Ito, Shigenori; Nakahari, Takashi

    2013-05-01

    In antral mucous cells, acetylcholine (ACh, 1 μM) activates Ca(2+)-regulated exocytosis, consisting of an initial peak that declines rapidly (initial transient phase) followed by a second slower decline (late phase) lasting during ACh stimulation. The addition of 8-bromo-cGMP (8-BrcGMP) enhanced the initial phase, which was inhibited by the protein kinase G (PKG) inhibitor guanosine 3',5'-cyclic monophosphorothoiate, β-phenyl-1,N(2)-etheno-8-bromo, Rp-isomer, sodium salt (Rp-8-BrPETcGMPS, 100 nM). However, Rp-8-BrPETcGMPS produced a delayed, but transient, increase in the exocytotic frequency during the late phase that was abolished by a protein kinase A (PKA) inhibitor (PKI-amide), suggesting that Rp-8-BrPETcGMPS accumulates cAMP. The cGMP-dependent phosphodiesterase 2 (PDE2), which degrades cAMP, may exist in antral mucous cells. The PDE2 inhibitor BAY-60-7550 (250 nM) mimicked the effect of Rp-8-BrPETcGMPS on ACh-stimulated exocytosis. Measurement of the cGMP and cAMP contents in antral mucosae revealed that ACh stimulates the accumulation of cGMP and that BAY-60-7550 accumulates cAMP similarly to Rp-8-BrPETcGMPS during ACh stimulation. Analyses of Western blot and immunohistochemistry demonstrated that PDE2A exists in antral mucous cells. In conclusion, Rp-8-BrPETcGMPS accumulates cAMP by inhibiting PDE2 in ACh-stimulated antral mucous cells, leading to the delayed, but transient, increase in the frequency of Ca(2+)-regulated exocytosis. PDE2 may prevent antral mucous cells from excessive mucin secretion caused by the cAMP accumulation. PMID:23449671

  20. Control of βAR- and N-methyl-D-aspartate (NMDA) Receptor-Dependent cAMP Dynamics in Hippocampal Neurons

    PubMed Central

    Chay, Andrew; Zamparo, Ilaria; Koschinski, Andreas; Zaccolo, Manuela; Blackwell, Kim T.

    2016-01-01

    Norepinephrine, a neuromodulator that activates β-adrenergic receptors (βARs), facilitates learning and memory as well as the induction of synaptic plasticity in the hippocampus. Several forms of long-term potentiation (LTP) at the Schaffer collateral CA1 synapse require stimulation of both βARs and N-methyl-D-aspartate receptors (NMDARs). To understand the mechanisms mediating the interactions between βAR and NMDAR signaling pathways, we combined FRET imaging of cAMP in hippocampal neuron cultures with spatial mechanistic modeling of signaling pathways in the CA1 pyramidal neuron. Previous work implied that cAMP is synergistically produced in the presence of the βAR agonist isoproterenol and intracellular calcium. In contrast, we show that when application of isoproterenol precedes application of NMDA by several minutes, as is typical of βAR-facilitated LTP experiments, the average amplitude of the cAMP response to NMDA is attenuated compared with the response to NMDA alone. Models simulations suggest that, although the negative feedback loop formed by cAMP, cAMP-dependent protein kinase (PKA), and type 4 phosphodiesterase may be involved in attenuating the cAMP response to NMDA, it is insufficient to explain the range of experimental observations. Instead, attenuation of the cAMP response requires mechanisms upstream of adenylyl cyclase. Our model demonstrates that Gs-to-Gi switching due to PKA phosphorylation of βARs as well as Gi inhibition of type 1 adenylyl cyclase may underlie the experimental observations. This suggests that signaling by β-adrenergic receptors depends on temporal pattern of stimulation, and that switching may represent a novel mechanism for recruiting kinases involved in synaptic plasticity and memory. PMID:26901880

  1. Control of βAR- and N-methyl-D-aspartate (NMDA) Receptor-Dependent cAMP Dynamics in Hippocampal Neurons.

    PubMed

    Chay, Andrew; Zamparo, Ilaria; Koschinski, Andreas; Zaccolo, Manuela; Blackwell, Kim T

    2016-02-01

    Norepinephrine, a neuromodulator that activates β-adrenergic receptors (βARs), facilitates learning and memory as well as the induction of synaptic plasticity in the hippocampus. Several forms of long-term potentiation (LTP) at the Schaffer collateral CA1 synapse require stimulation of both βARs and N-methyl-D-aspartate receptors (NMDARs). To understand the mechanisms mediating the interactions between βAR and NMDAR signaling pathways, we combined FRET imaging of cAMP in hippocampal neuron cultures with spatial mechanistic modeling of signaling pathways in the CA1 pyramidal neuron. Previous work implied that cAMP is synergistically produced in the presence of the βAR agonist isoproterenol and intracellular calcium. In contrast, we show that when application of isoproterenol precedes application of NMDA by several minutes, as is typical of βAR-facilitated LTP experiments, the average amplitude of the cAMP response to NMDA is attenuated compared with the response to NMDA alone. Models simulations suggest that, although the negative feedback loop formed by cAMP, cAMP-dependent protein kinase (PKA), and type 4 phosphodiesterase may be involved in attenuating the cAMP response to NMDA, it is insufficient to explain the range of experimental observations. Instead, attenuation of the cAMP response requires mechanisms upstream of adenylyl cyclase. Our model demonstrates that Gs-to-Gi switching due to PKA phosphorylation of βARs as well as Gi inhibition of type 1 adenylyl cyclase may underlie the experimental observations. This suggests that signaling by β-adrenergic receptors depends on temporal pattern of stimulation, and that switching may represent a novel mechanism for recruiting kinases involved in synaptic plasticity and memory. PMID:26901880

  2. Determination of phosphodiesterase I activity in human blood serum.

    PubMed

    Hynie, I; Meuffels, M; Poznanski, W J

    1975-09-01

    Phosphodiesterase I (EC 3.1.4.1) activity was detected in normal human blood serum. The enzyme is stable at laboratory temperature for three days, but is inactivated at pH less than 7. The pH for optimum activity increases with the substrate concentration (under the conditions used, from pH 9.0 to 10.2) and, conversely, the Km increases with pH and buffer concentration. The enzyme is inhibited by ethylenediaminetetraacetate but not by phosphate (0.1 mol/liter). We developed a simple quantitative method for its determination, based on hydrolysis of the p-nitrophenyl ester of thymidine 5'-monophosphate and subsequent measurement of the liberated p-nitrophenol at 400 nm in NaOH (0.1 mol/liter). Normal values (mean +/- 2 SD) were determined to be 33 +/- 6.4 U/liter. Preliminary studies indicate that phosphodiesterase I activity is greater than normal in serum of patients with necrotic changes in the liver or kidney or in cases of breast cancer, but not in that of patients with myocardial infarction, bone cancer, lung cancer, or chronic liver cirrhosis. PMID:168991

  3. [Phosphodiesterase-5 inhibitors for the treatment of pulmonary arterial hypertension].

    PubMed

    Beltrán-Gámez, Miguel E; Sandoval-Zárate, Julio; Pulido, Tomás

    2015-01-01

    In experimental and clinical cardiology, phosphodiesterase type 5 (PDE-5) inhibitors have brought scientific interest as a therapeutic tool in pulmonary arterial hypertension (PAH) management in recent years. Phosphodiesterases are a superfamily of enzymes that inactivate cyclic adenosine monophosphate and cyclic guanosine monophosphate, the second messengers of prostacyclin and nitric oxide. The rationale for the use of PDE-5 inhibitors in PAH is based on their capacity to overexpresss the nitric oxide pathway pursued inhibition of cyclic guanosine monophosphate hydrolysis. By increasing cyclic guanosine monophosphate levels it promotes vasodilation, antiproliferative and pro-apoptotic effects that may reverse pulmonary vascular remodeling. There is also evidence that these drugs may directly enhance right ventricular contractility through an increase in cyclic adenosine monophosphate mediated by the inhibition of the cyclic guanosine monophosphate -sensitive PDE-3. Sildenafil, tadalafil and vardenafil are 3 specific PDE-5 inhibitors in current clinical use, which share similar mechanisms of action but present some significant differences regarding potency, selectivity for PDE-5 and pharmacokinetic properties. Sildenafil received approval in 2005 by the Food and Drug Administration and the European Medicines Agency and tadalafil in 2009 by the Food and Drug Administration and the European Medicines Agency for the treatment of PAH in patients classified as NYHA/WHO functional class II and III. In Mexico, sildenafil and tadalafil were approved by Comisión Federal de Protección contra Riesgos Sanitarios for this indication in 2010 and 2011, respectively. PMID:26047999

  4. A Novel Conditional Genetic System Reveals That Increasing Neuronal cAMP Enhances Memory and Retrieval

    PubMed Central

    Isiegas, Carolina; McDonough, Conor; Huang, Ted; Havekes, Robbert; Fabian, Sara; Wu, Long-Jun; Xu, Hui; Zhao, Ming-Gao; Kim, Jae-Ick; Lee, Yong-Seok; Lee, Hye-Ryeon; Ko, Hyoung-Gon; Lee, Nuribalhae; Choi, Sun-Lim; Lee, Jeong-Sik; Son, Hyeon; Zhuo, Min; Kaang, Bong-Kiun; Abel, Ted

    2010-01-01

    Consistent evidence from pharmacological and genetic studies shows that cAMP is a critical modulator of synaptic plasticity and memory formation. However, the potential of the cAMP signaling pathway as a target for memory enhancement remains unclear because of contradictory findings from pharmacological and genetic approaches. To address these issues, we have developed a novel conditional genetic system in mice based on the heterologous expression of an Aplysia octopamine receptor, a G-protein-coupled receptor whose activation by its natural ligand octopamine leads to rapid and transient increases in cAMP. We find that activation of this receptor transgenically expressed in mouse forebrain neurons induces a rapid elevation of hippocampal cAMP levels, facilitates hippocampus synaptic plasticity, and enhances the consolidation and retrieval of fear memory. Our findings clearly demonstrate that acute increases in cAMP levels selectively in neurons facilitate synaptic plasticity and memory, and illustrate the potential of this heterologous system to study cAMP-mediated processes in mammalian systems. PMID:18550764

  5. Persistent cAMP Signaling by Internalized LH Receptors in Ovarian Follicles.

    PubMed

    Lyga, Sandra; Volpe, Silvia; Werthmann, Ruth C; Götz, Konrad; Sungkaworn, Titiwat; Lohse, Martin J; Calebiro, Davide

    2016-04-01

    A crucial event in female reproduction occurs at midcycle, when a LH peak induces the final maturation of ovarian follicles. LH signals via a G protein-coupled receptor selectively expressed in the outermost follicular cell layers. However, how LH signals are relayed inside these cells and finally to the oocyte is incompletely understood. Here, we monitored LH signaling in intact ovarian follicles of transgenic mice expressing a fluorescent cAMP sensor. We found that LH stimulation induces 2 phases of cAMP signaling in all cell layers surrounding the oocyte. Interfering with LH receptor internalization abolished the second, persistent cAMP phase and partially inhibited oocyte meiosis resumption. These data suggest that persistent cAMP signals from internalized LH receptors contribute to transmitting LH effects inside follicle cells and ultimately to the oocyte. Thus, this study indicates that the recently proposed paradigm of cAMP signaling by internalized G protein-coupled receptors is implicated in receptor function and is physiologically relevant. PMID:26828746

  6. Business and Finance: A Resource Book for Camps. Camp Administration Series.

    ERIC Educational Resources Information Center

    Ball, Armand, Ed.; Ball, Beverly, Ed.

    this resource book contains 35 articles designed to present general information concerning camp business and financial management to directors or administrators. Typically a specialist is employed to supervise camp business and finance, but the administrator needs to have a broad understanding of five specific business topics: (1) liability and…

  7. Catch the Dream of Camping: Albuquerque Keynoters Share Insight into the Future of Camping.

    ERIC Educational Resources Information Center

    Coleman, Marla

    2000-01-01

    Highlights the views of three keynote speakers who will share their insights into the future of the camp experience at the next American Camping Association National Conference. Environmentalist Bill McKibben, educator Cynthia Tobias, and futurist Ira Blumenthal focus, respectively, on connecting children to the natural world, understanding…

  8. Allegheny National Forest, CCC Camp ANF1 , The camp’s main ...

    Library of Congress Historic Buildings Survey, Historic Engineering Record, Historic Landscapes Survey

    Allegheny National Forest, CCC Camp ANF-1 , The camp’s main entrance is located at the intersection of Duhring Road (ANF 131) and ANF 124, Pennsylvania, with the interior site road known as Trail Ride Drive., Marienville, Forest County, PA

  9. Efficacy and Mechanism of a Glycoside Compound Inhibiting Abnormal Prion Protein Formation in Prion-Infected Cells: Implications of Interferon and Phosphodiesterase 4D-Interacting Protein

    PubMed Central

    Nishizawa, Keiko; Oguma, Ayumi; Kawata, Maki; Sakasegawa, Yuji; Teruya, Kenta

    2014-01-01

    ABSTRACT A new type of antiprion compound, Gly-9, was found to inhibit abnormal prion protein formation in prion-infected neuroblastoma cells, in a prion strain-independent manner, when the cells were treated for more than 1 day. It reduced the intracellular prion protein level and significantly modified mRNA expression levels of genes of two types: interferon-stimulated genes were downregulated after more than 2 days of treatment, and the phosphodiesterase 4D-interacting protein gene, a gene involved in microtubule growth, was upregulated after more than 1 day of treatment. A supplement of interferon given to the cells partly restored the abnormal prion protein level but did not alter the normal prion protein level. This interferon action was independent of the Janus activated kinase-signal transducer and activator of transcription signaling pathway. Therefore, the changes in interferon-stimulated genes might be a secondary effect of Gly-9 treatment. However, gene knockdown of phosphodiesterase 4D-interacting protein restored or increased both the abnormal prion protein level and the normal prion protein level, without transcriptional alteration of the prion protein gene. It also altered the localization of abnormal prion protein accumulation in the cells, indicating that phosphodiesterase 4D-interacting protein might affect prion protein levels by altering the trafficking of prion protein-containing structures. Interferon and phosphodiesterase 4D-interacting protein had no direct mutual link, demonstrating that they regulate abnormal prion protein levels independently. Although the in vivo efficacy of Gly-9 was limited, the findings for Gly-9 provide insights into the regulation of abnormal prion protein in cells and suggest new targets for antiprion compounds. IMPORTANCE This report describes our study of the efficacy and potential mechanism underlying the antiprion action of a new antiprion compound with a glycoside structure in prion-infected cells, as well as

  10. Action of methyl-, propyl- and butylparaben on GPR30 gene and protein expression, cAMP levels and activation of ERK1/2 and PI3K/Akt signaling pathways in MCF-7 breast cancer cells and MCF-10A non-transformed breast epithelial cells.

    PubMed

    Wróbel, Anna Maria; Gregoraszczuk, Ewa Łucja

    2015-10-14

    In the present study, we examined cAMP levels and activation of the MAPK/ERK1/2 and PI3K/Akt signaling pathways in response to the actions of parabens on GPR30 in MCF-7 and MCF-10A cells. Cells were exposed to methyl-, propyl- or butylparaben at a concentration of 20nM; 17-β-estradiol (10nM) was used as a positive control. 17β-estradiol and all tested parabens increased GPR30 gene and protein expression in MCF-7 and MCF-10A cells. No parabens affected cAMP levels in either cell line, with the exception of propylparaben in MCF-10A cells. 17β-estradiol, propylparaben, and butylparaben increased phosphorylation of ERK1/2 in MCF-7 cells, whereas 17β-estradiol, methyl- and butylparaben, but not propylparaben, increased phosphorylation of ERK1/2 in MCF-10A cells. Akt activation was noted only in MCF-7 cells and only with propylparaben treatment. Collectively, the data presented here point to a nongenomic mechanism of action of parabens in activation GPR30 in both cancer and non-cancer breast cell lines through βγ dimer-mediated activation of the ERK1/2 pathway, but not the cAMP/PKA pathway. Moreover, among investigated parabens, propylparaben appears to inhibit apoptosis in cancer cells through activation of Akt kinases, confirming conclusions suggested by our previously published data. Nevertheless, continuing research on the carcinogenic action of parabens is warranted. PMID:26253279

  11. Parents' Perceptions of Benefits of Summer Camp for Campers with Mental Retardation.

    ERIC Educational Resources Information Center

    McCormick, Bryan; And Others

    1992-01-01

    Study explored the perception of parents of campers with mental retardation regarding the benefits of summer camps. Parents rated the relative importance of 22 possible benefits and indicated 6 dimensions of benefits in an ideal program: social skill development, social competence, respite care, cognitive development, expressive development, and…

  12. Positive genetic feedback governs cAMP spiral wave formation in Dictyostelium.

    PubMed Central

    Levine, H; Aranson, I; Tsimring, L; Truong, T V

    1996-01-01

    The aggregation stage of the life cycle of Dictyostelium discoideum is governed by the chemotactic response of individual amoebae to excitable waves of cAMP. We modeled this process through a recently introduced hybrid automata-continuum scheme and used computer simulation to unravel the role of specific components of this complex developmental process. Our results indicated an essential role for positive feedback between the cAMP signaling and the expression of the genes encoding the signal transduction and response machinery. PMID:8692824

  13. Anxiolytic Effects of Phosphodiesterase-2 Inhibitors Associated with Increased cGMP Signaling

    PubMed Central

    Masood, Anbrin; Huang, Ying; Hajjhussein, Hassan; Xiao, Lan; Li, Hao; Wang, Wei; Hamza, Adel; Zhan, Chang-Guo

    2009-01-01

    Phosphodiesterase (PDE)-2 is a component of the nitric-oxide synthase (NOS)/guanylyl cyclase signaling pathway in the brain. Given recent evidence that pharmacologically induced changes in NO-cGMP signaling can affect anxiety-related behaviors, the effects of the PDE2 inhibitors (2-(3,4-dimethoxybenzyl)-7-det-5-methylimidazo-[5,1-f][1,2,4]triazin-4(3H)-one) (Bay 60-7550) and 3-(8-methoxy-1-methyl-2-oxo-7-phenyl-2,3-dihydro-1H-benzo[e][1,4]diazepin-5-yl)benzamide (ND7001), as well as modulators of NO, were assessed on cGMP signaling in neurons and on the behavior of mice in the elevated plus-maze, hole-board, and open-field tests, well established procedures for the evaluation of anxiolytics. Bay 60-7550 (1 μM) and ND7001 (10 μM) increased basal and N-methyl-d-aspartate- or detanonoate-stimulated cGMP in primary cultures of rat cerebral cortical neurons; Bay 60-7550, but not ND7001, also increased cAMP. Increased cGMP signaling, either by administration of the PDE2 inhibitors Bay 60-7550 (0.5, 1, and 3 mg/kg) or ND7001 (1 mg/kg), or the NO donor detanonoate (0.5 mg/kg), antagonized the anxiogenic effects of restraint stress on behavior in the three tests. These drugs also produced anxiolytic effects on behavior in nonstressed mice in the elevated plus-maze and hole-board tests; these effects were antagonized by the guanylyl cyclase inhibitor 1H-[1,2,4]oxadiazolo[4,3-a]quinoxalin-1-one (20 mg/kg). By contrast, the NOS inhibitor Nω-nitro-l-arginine methyl ester (50 mg/kg), which reduces cGMP signaling, produced anxiogenic effects similar to restraint stress. Overall, the present behavioral and neurochemical data suggest that PDE2 may be a novel pharmacological target for the development of drugs for the treatment of anxiety disorders. PMID:19684253

  14. A specialized program for children with developmental disabilities within a "typical" overnight summer camp: Camp Ramah's Tikvah Program.

    PubMed

    Blas, Howard I

    2007-10-01

    The Tikvah Program is an overnight camping program at Camp Ramah in New England that serves campers with a range of developmental disabilities. The program has evolved over its 37-year history and includes a camping program, vocational training program, and inclusion program. Select graduates are hired by the camp for summer employment. The Tikvah Program offers a model for serving campers with special needs within a larger "typical" summer camp. Although serving the needs of such campers offers unique challenges, the presence of such a program in a regular summer camp offers tremendous opportunities and benefits for campers with special needs and more typically developing campers. PMID:17823062

  15. The EAL domain containing protein STM2215 (rtn) is needed during Salmonella infection and has cyclic di-GMP phosphodiesterase activity.

    PubMed

    Zheng, Yi; Sambou, Tounkang; Bogomolnaya, Lydia M; Cirillo, Jeffrey D; McClelland, Michael; Andrews-Polymenis, Helene

    2013-08-01

    Salmonella Typhimurium gene STM2215 (rtn) is conserved among many enterobacteriaceae. Mutants lacking STM2215 poorly colonized the liver and spleen in intraperitoneal infection of mice and poorly colonized the intestine and deeper tissues in oral infection. These phenotypes were complemented by a wild-type copy of STM2215 provided in trans. STM2215 deletion mutants grew normally in J774A.1 murine macrophages but were unable to invade Caco-2 colonic epithelial cells. Consistent with this finding, mutants in STM2215 produced lower levels of effectors of the TTSS-1. STM2215 is a predicted c-di-GMP phosphodiesterase, but lacks identifiable sensor domains. Biochemical analysis of STM2215 determined that it is located in the inner membrane and has c-di-GMP phosphodiesterase activity in vitro dependent on an intact EAL motif. Unlike some previously identified members of this family, STM2215 did not affect motility, was expressed on plates, and in liquid media at late exponential and early stationary phase during growth. Defined mutations in STM2215 revealed that neither the predicted periplasmic domain nor the anchoring of the protein to the inner membrane is necessary for the activity of this protein during infection. However, the EAL domain of STM2215 is required during infection, suggesting that its phosphodiesterase activity is necessary during infection. PMID:23734719

  16. Icariside II, a novel phosphodiesterase-5 inhibitor, attenuates streptozotocin-induced cognitive deficits in rats.

    PubMed

    Yin, Caixia; Deng, Yuanyuan; Gao, Jianmei; Li, Xiaohui; Liu, Yuangui; Gong, Qihai

    2016-07-22

    Beta-amyloid (Aβ) deposition and neuroinflammation are involved in Alzheimer's disease (AD)-type neurodegeneration with cognitive deficits. Phosphodiesterase-5 (PDE5) inhibitors have recently been studied as a potential target for cognitive enhancement by reducing inflammatory responses and Aβ levels. The present study was designed to investigate the effects of icariside II (ICS II), a novel PDE5 inhibitor derived from the traditional Chinese herb Epimedium brevicornum, on cognitive deficits, Aβ levels and neuroinflammation induced by intracerebroventricular-streptozotocin (ICV-STZ) in rats. The results demonstrated that ICV-STZ exhibited cognitive deficits and neuronal morphological damage, along with Aβ increase and neuroinflammation in the rat hippocampus. ICS II improved cognitive deficits, attenuated neuronal death, and decreased the levels of Aβ1-40, Aβ1-42 and PDE5 in the hippocampus of STZ rats. Furthermore, administration of ICS II at the dose of 10mg/kg for 21days significantly suppressed the expression of beta-amyloid precursor protein (APP), beta-secretase1 (BACE1) and increased the expressions of neprilysin (NEP) together with inhibited interleukin-1β (IL-1β), tumor necrosis factor (TNF)-α, cyclooxygenase-2 (COX-2) and transforming growth factor-β1 (TGF-β1) levels. In addition, ICS II exerted a beneficial effect on inhibition of IκB-α degradation and NF-κB activation induced by STZ. Taken together, the present study demonstrated that ICS II was a potential therapeutic agent for AD treatment. PMID:27109920

  17. Phosphodiesterase as a new therapeutic target for the treatment of spinal cord injury and neurodegenerative diseases.

    PubMed

    Paterniti, Irene; Esposito, Emanuela; Cuzzocrea, Salvatore

    2014-01-01

    Interest in Central Nervous System (CNS) inflammation has rapidly grown over the past decade driven by the increasing evidence indicating that chronic inflammation and neuroinflammation in the brain may play an important role in the progressive neuronal cell death in many chronic CNS diseases, such as Alzheimer and Parkinson's diseases, traumatic brain injury, spinal cord injury (SCI), as well as pathologies associated with CNS infections. In peripheral tissues, generally inflammation has a protective role limiting the survival and proliferation of invading pathogens, promoting tissue repair and recovery. This innate response normally resolves over a few weeks, accompanyied by tissue repair aided by macrophages recruited to the site. However, when the inflammatory response does not undergo resolution, it might turn into chronic inflammation. Any chronic inflammatory process can damage healthy tissue and the brain may be particularly vulnerable, since destroyed neurons can not be replaced. Recently, several reports have suggested that phosphodiesterases (PDEs) are new targets for central nervous system (CNS) diseases. All the PDEs are expressed in the CNS, making this gene family a particularly attractive source of new targets for the treatment of psychiatric and neurodegenerative disorders. Significantly, all neurons express multiple PDEs, which differ in cyclic nucleotide specificity, affinity, regulatory control and subcellular compartmentalization. Therefore, PDEs inhibition represents a mechanism through which it could be possible to precisely modulate neuronal activity. In this article, we review the current state of art of PDEs in the CNS diseases associated with neuroinflammation. PMID:24533816

  18. Phosphodiesterase 5/protein kinase G signal governs stemness of prostate cancer stem cells through Hippo pathway.

    PubMed

    Liu, Naihua; Mei, Liu; Fan, Xueying; Tang, Chao; Ji, Xing; Hu, Xinhua; Shi, Wei; Qian, Yu; Hussain, Musaddique; Wu, Junsong; Wang, Chaojun; Lin, Shaoqiang; Wu, Ximei

    2016-08-01

    Cancer stem cells (CSC) are critical for initiation, metastasis, and relapse of cancers, however, the underlying mechanism governing stemness of CSC remains unknown. Herein, we have investigated the roles of phosphodiesterase 5 (PDE5) in stemness of prostate cancer cells. Both PDE5 and WW domain-containing transcription regulator protein-1 (TAZ), a core effector of Hippo pathway, are highly expressed in the PC3-derived cancer stem cells (PCSC). Either TAZ knockdown or inhibition of PDE5 activity attenuated colony formation, altered expression patterns of stem cell markers, and enhanced cisplatin cytotoxicity, resulting in attenuation of stemness in PCSC. In addition, inhibition of PDE5 activity by its specific inhibitors activates cGMP-dependent protein kinase G (PKG), which in turn induces MST/LATS kinases, resulting in cytosolic degradation of TAZ and activation of Hippo pathway. Accordingly, knockdown of TAZ almost completely abolished PDE5 inhibitor-induced attenuation in stemness in cultured PCSC, whereas knockdown of TAZ not only abolished PDE5 inhibitor-induced attenuation in stemness but also facilitated PDE5 inhibitor-induced trans-differentiation in PCSC xenografts. Together, the present study has uncovered that PDE/cGMP/PKG signal targets to Hippo/TAZ pathway in maintaining stemness of PCSC, and suggested that PDE5 inhibitors in combination with chemotherapeutic agents could effectively prevent initiation, metastasis, and relapse of prostate cancer. PMID:27179930

  19. [Late psychological problems after concentration camp incarceration].

    PubMed

    Eitinger, L

    1991-01-01

    After the Second World War most studies showed that concentration camp prisoners suffered from serious somatic and psychological damage. This survey documents that Norwegian ex-prisoners from concentration camps were examined and diagnosed, and received adequate somatic therapy; moreover that the majority also attained an acceptable, financially--more or less--secure existence. But eventually it could be observed that, apart from a few who received individual psychotherapy, too little was done to help ex-prisoners with their psychological problems. PMID:2038563

  20. Summer camp course in nuclear operations

    SciTech Connect

    Peterson, P.F.; James, J.Z. ); Terrell, B.E. )

    1993-01-01

    This paper describes a new kind of nuclear engineering curriculum that echoes an old method of professional training - the intensive summer camp. For many years a staple of the training of civil engineers and foresters, summer camp courses immerse the student in an intensive, focused experience, isolated from the familiar campus and resembling the actual work environment for which the student is being trained. With financial support from the U.S. Department of Energy, University of California-Berkeley (UCB) and Pacific Gas Electric (PG E) have launched such a course for UCB nuclear engineering undergraduates.

  1. Characterization of Binding and Inhibitory Properties of TAK-063, a Novel Phosphodiesterase 10A Inhibitor

    PubMed Central

    Harada, Akina; Suzuki, Kazunori; Kamiguchi, Naomi; Miyamoto, Maki; Tohyama, Kimio; Nakashima, Kosuke; Taniguchi, Takahiko; Kimura, Haruhide

    2015-01-01

    Phosphodiesterase 10A (PDE10A) inhibition is a novel and promising approach for the treatment of central nervous system disorders such as schizophrenia and Huntington’s disease. A novel PDE10A inhibitor, TAK-063 [1-[2-fluoro-4-(1H-pyrazol-1-yl)phenyl]-5-methoxy-3-(1-phenyl-1H-pyrazol-5-yl)-pyridazin-4(1H)-one] has shown high inhibitory activity and selectivity for human recombinant PDE10A2 in vitro; the half-maximal inhibitory concentration was 0.30 nM, and selectivity over other phosphodiesterases (PDEs) was more than 15000-fold. TAK-063 at 10 µM did not show more than 50% inhibition or stimulation of 91 enzymes or receptors except for PDEs. In vitro autoradiography (ARG) studies using rat brain sections revealed that [3H]TAK-063 selectively accumulated in the caudate putamen (CPu), nucleus accumbens (NAc), globus pallidus, substantia nigra, and striatonigral projection, where PDE10A is highly expressed. This [3H]TAK-063 accumulation was almost entirely blocked by an excess amount of MP-10, a PDE10A selective inhibitor, and the accumulation was not observed in brain slices of Pde10a-knockout mice. In rat brain sections, [3H]TAK-063 bound to a single high-affinity site with mean ± SEM dissociation constants of 7.2 ± 1.2 and 2.6 ± 0.5 nM for the CPu and NAc shell, respectively. Orally administered [14C]TAK-063 selectively accumulated in PDE10A expressing brain regions in an in vivo ARG study in rats. Striatal PDE10A occupancy by TAK-063 in vivo was measured using T-773 as a tracer and a dose of 0.88 mg/kg (p.o.) was calculated to produce 50% occupancy in rats. Translational studies with TAK-063 and other PDE10A inhibitors such as those presented here will help us better understand the pharmacological profile of this class of potential central nervous system drugs. PMID:25815469

  2. Camp Hope: Integrating Tai Chi and Humanbecoming in a Camp for Children.

    PubMed

    Delis, Pamela Coombs; Leger, Robin R; Longton, Fran; Chandler, David Lynn

    2015-10-01

    This article is about Camp Hope, a two-week summer day program for children age 6-12 from low income families and violent neighborhoods and for children who are struggling academically or identified as having behavioral health issues. The camp is operated by House of Peace and Education (HOPE) a 501(c)(3) nonprofit company that began in 1996. The program is designed to offer enrichment activities in a caring community for young children. The theoretical framework behind the vision and policies is Parse's humanbecoming paradigm. In particular, the camp administrators and staff try to see all persons as co-creating of their everchanging humanuniverse process. PMID:26396217

  3. Phosphodiesterase 8a Supports HIV-1 Replication in Macrophages at the Level of Reverse Transcription

    PubMed Central

    Booiman, Thijs; Cobos Jiménez, Viviana; van Dort, Karel A.; van 't Wout, Angélique B.; Kootstra, Neeltje A.

    2014-01-01

    Background HIV-1 infected macrophages play a key role in HIV-1 infection. Even during anti-retroviral treatment, macrophages keep producing virus due to suboptimal tissue penetration and reduced efficacy of antiretrovirals. It is therefore of major importance to understand which host factors are involved in HIV-1 replication in macrophages. Previously, we have shown that genetic polymorphisms in phosphodiesterase 8a (PDE8A) are strongly associated with HIV-1 replication in these cells. Here we analyzed the mechanism and regulation of PDE8A in HIV-1 replication in macrophages. Results PDE8A mRNA expression strongly increases upon differentiation of monocytes into macrophages, which corresponds to the increased susceptibility of mature macrophages to HIV-1. In parallel, expression of microRNA miR-145-5p, predicted to target PDE8A mRNA, strongly decreased. The interaction of miR-145-5p with the 3′ UTR of PDE8A mRNA could be experimentally validated, suggesting that indeed miR-145-5p can regulate PDE8A expression levels. Knockdown of PDE8A in macrophages resulted in a decrease in total HIV-1 replication and proviral DNA levels. These observations confirm that PDE8A regulates HIV-1 replication in macrophages and that this effect is mediated through early steps in the viral replication cycle. Conclusions PDE8A is highly expressed in macrophages, and its expression is regulated by miR-145-5p. Our findings strongly suggest that PDE8A supports HIV-1 replication in macrophages and that this effect is mediated at the level of reverse transcription. PMID:25295610

  4. STC1 induction by PACAP is mediated through cAMP and ERK1/2 but not PKA in cultured cortical neurons

    PubMed Central

    Holighaus, Yvonne; Weihe, Eberhard; Eiden, Lee E.

    2011-01-01

    The neuroprotective actions of PACAP (pituitary adenylate cyclase-activating polypeptide) in vitro and in vivo suggest that activation of its cognate G protein-coupled receptor PAC1 or downstream signaling molecules, and thus activation of PACAP target genes, could be of therapeutic benefit. Here we show, that cultured rat cortical neurons predominantly expressed the PAC1hop and null variants, activation of which resulted in elevation of the two second messengers cAMP and Ca2+ and expression of the putative neuroprotectant stanniocalcin 1 (STC1). PACAP signaling to the STC1 gene proceeded through the extracellular signal-regulated kinases 1 and 2 (ERK1/2), but not through the cAMP dependent protein kinase (PKA), and was mimicked by the adenylate cyclase activator forskolin. PACAP- and forskolin-mediated activation of ERK1/2 occurred through cAMP, but not PKA. These results suggest that STC1 gene induction proceeds through cAMP and ERK1/2, independently of PKA, the canonical cAMP effector. In contrast, PACAP signaling to the BDNF gene proceeded through PKA, suggesting that two different neuroprotective cAMP pathways co-exist in differentiated cortical neurons. The selective activation of a potentially neuroprotective cAMP dependent pathway different from the canonical cAMP pathway used in many physiological processes, such as memory storage, has implications for pharmacological activation of neuroprotection in vivo. PMID:21975601

  5. 8. VIEW EAST, CIVILIAN CONSERVATION CORPS CAMP PARSONS GARAGE, MODERN ...

    Library of Congress Historic Buildings Survey, Historic Engineering Record, Historic Landscapes Survey

    8. VIEW EAST, CIVILIAN CONSERVATION CORPS CAMP PARSONS GARAGE, MODERN SHED, CIVILIAN CONSERVATION CORPS CAMP PARSONS BLACKSMITH SHOP, AND MODERN SHED. - Parsons Nursery, South side of U.S. Route 219, Parsons, Tucker County, WV

  6. CPSC Warns of Carbon Monoxide Poisoning with Camping Equipment

    MedlinePlus

    ... Warns of Carbon Monoxide Poisoning Hazard with Camping Equipment The U.S. Consumer Product Safety Commission (CPSC) warns ... about the carbon monoxide (CO) hazard with camping equipment. CO can kill you! From 2002–2006, CPSC ...

  7. Effects of metal ions and disulfide bonds on the activity of phosphodiesterase from Trimeresurus stejnegeri venom.

    PubMed

    Peng, Lili; Xu, Xiaolong; Guo, Mingchun; Yan, Xincheng; Wang, Shasha; Gao, Shang; Zhu, Shanshan

    2013-06-01

    Obviously different from the other known phosphodiesterases, the phosphodiesterase from Trimeresurus stejnegeri venom (TS-PDE) consists of two different chains linked with disulfide bonds and contains both endogenous Cu(2+) and Zn(2+). Cu(2+) and Zn(2+) are important for its phosphodiesterase activity. In this study, the effects of metal ions and small-molecule reductants on its structure and activity have been investigated by polyacrylamide gel electrophoresis, high performance liquid chromatography, fluorescence and electron paramagnetic resonance spectroscopy. The results show that TS-PDE has one class of Zn(2+) binding site and two classes of Cu(2+) binding site, including the high affinity activator sites and the low affinity sites. Cu(2+) ions function as a switch for its phosphodiesterase activity. The catalytic activity of TS-PDE does not have an absolute requirement for Cu(2+) and Zn(2+). Mg(2+), Mn(2+), Ni(2+), Co(2+) and Ca(2+) are all effective for its phosphodiesterase activity. TS-PDE has seven disulfide bonds and ten free cysteine residues. l-Ascorbate inhibits the phosphodiesterase activity of TS-PDE through reduction of the Cu(2+), while dithiothreitol, glutathione and tris(2-carboxyethyl)phosphine inhibit the phosphodiesterase activity of TS-PDE by reducing both the Cu(2+) and disulfide bonds. The catalytic activity of TS-PDE relies on its disulfide bonds and bimetallic cluster. In addition, biologically-relevant reductants, glutathione and l-ascorbate, have been found to be endogenous inhibitors to the phosphodiesterase activity of TS-PDE. PMID:23775423

  8. Camp Health Aide Manual = Manual para trabajadores de salud.

    ERIC Educational Resources Information Center

    Robinson, June Grube; And Others

    This bilingual manual serves as a textbook for migrant Camp Health Aides. Camp Health Aides are members of migrant labor camps enlisted to provide information about health and social services to migrant workers and their families. The manual is divided into 12 tabbed sections representing lessons. Teaching notes printed on contrasting paper…

  9. 49 CFR 218.80 - Movement of occupied camp cars.

    Code of Federal Regulations, 2014 CFR

    2014-10-01

    ... 49 Transportation 4 2014-10-01 2014-10-01 false Movement of occupied camp cars. 218.80 Section 218... ADMINISTRATION, DEPARTMENT OF TRANSPORTATION RAILROAD OPERATING PRACTICES Protection of Occupied Camp Cars § 218.80 Movement of occupied camp cars. Occupied cars may not be humped or flat switched unless coupled...

  10. 49 CFR 218.80 - Movement of occupied camp cars.

    Code of Federal Regulations, 2012 CFR

    2012-10-01

    ... 49 Transportation 4 2012-10-01 2012-10-01 false Movement of occupied camp cars. 218.80 Section 218... ADMINISTRATION, DEPARTMENT OF TRANSPORTATION RAILROAD OPERATING PRACTICES Protection of Occupied Camp Cars § 218.80 Movement of occupied camp cars. Occupied cars may not be humped or flat switched unless coupled...

  11. 49 CFR 218.80 - Movement of occupied camp cars.

    Code of Federal Regulations, 2013 CFR

    2013-10-01

    ... 49 Transportation 4 2013-10-01 2013-10-01 false Movement of occupied camp cars. 218.80 Section 218... ADMINISTRATION, DEPARTMENT OF TRANSPORTATION RAILROAD OPERATING PRACTICES Protection of Occupied Camp Cars § 218.80 Movement of occupied camp cars. Occupied cars may not be humped or flat switched unless coupled...

  12. Trainer Guide: Business and Finance Managerial. Camp Administration Series.

    ERIC Educational Resources Information Center

    Farley, Elizabeth, Ed.

    Suggested ideas on conducting a managerial workshop for camp directors are offered in this trainer's guide. Workshops must be at least one full day of training (6 hours) on each topic to be counted toward the American Camping Association (ACA) Camp Director Certification Program. Suggested topics to be addressed are: (1) basic principles and…

  13. 28 CFR 523.15 - Camp or farm good time.

    Code of Federal Regulations, 2010 CFR

    2010-07-01

    ... 28 Judicial Administration 2 2010-07-01 2010-07-01 false Camp or farm good time. 523.15 Section..., CLASSIFICATION, AND TRANSFER COMPUTATION OF SENTENCE Extra Good Time § 523.15 Camp or farm good time. An inmate assigned to a farm or camp is automatically awarded extra good time, beginning on the date of commitment...

  14. 28 CFR 523.15 - Camp or farm good time.

    Code of Federal Regulations, 2013 CFR

    2013-07-01

    ... 28 Judicial Administration 2 2013-07-01 2013-07-01 false Camp or farm good time. 523.15 Section..., CLASSIFICATION, AND TRANSFER COMPUTATION OF SENTENCE Extra Good Time § 523.15 Camp or farm good time. An inmate assigned to a farm or camp is automatically awarded extra good time, beginning on the date of commitment...

  15. 28 CFR 523.15 - Camp or farm good time.

    Code of Federal Regulations, 2012 CFR

    2012-07-01

    ... 28 Judicial Administration 2 2012-07-01 2012-07-01 false Camp or farm good time. 523.15 Section..., CLASSIFICATION, AND TRANSFER COMPUTATION OF SENTENCE Extra Good Time § 523.15 Camp or farm good time. An inmate assigned to a farm or camp is automatically awarded extra good time, beginning on the date of commitment...

  16. 28 CFR 523.15 - Camp or farm good time.

    Code of Federal Regulations, 2014 CFR

    2014-07-01

    ... 28 Judicial Administration 2 2014-07-01 2014-07-01 false Camp or farm good time. 523.15 Section..., CLASSIFICATION, AND TRANSFER COMPUTATION OF SENTENCE Extra Good Time § 523.15 Camp or farm good time. An inmate assigned to a farm or camp is automatically awarded extra good time, beginning on the date of commitment...

  17. 28 CFR 523.15 - Camp or farm good time.

    Code of Federal Regulations, 2011 CFR

    2011-07-01

    ... 28 Judicial Administration 2 2011-07-01 2011-07-01 false Camp or farm good time. 523.15 Section..., CLASSIFICATION, AND TRANSFER COMPUTATION OF SENTENCE Extra Good Time § 523.15 Camp or farm good time. An inmate assigned to a farm or camp is automatically awarded extra good time, beginning on the date of commitment...

  18. Summer Camp and Positive Youth Development: Program with Romanian Youth

    ERIC Educational Resources Information Center

    Feenstra, Jennifer S.

    2015-01-01

    A variety of activities are used in camps to help promote positive youth development, improving social skills and self-esteem in campers. I expanded on previous camp research in this study to address the influence camps have on trust, belief in the honesty of others, empowerment, and care for others in youth in Eastern Europe. Since 1999, New…

  19. 49 CFR 218.80 - Movement of occupied camp cars.

    Code of Federal Regulations, 2010 CFR

    2010-10-01

    ... 49 Transportation 4 2010-10-01 2010-10-01 false Movement of occupied camp cars. 218.80 Section 218.80 Transportation Other Regulations Relating to Transportation (Continued) FEDERAL RAILROAD ADMINISTRATION, DEPARTMENT OF TRANSPORTATION RAILROAD OPERATING PRACTICES Protection of Occupied Camp Cars § 218.80 Movement of occupied camp cars....

  20. 49 CFR 218.80 - Movement of occupied camp cars.

    Code of Federal Regulations, 2011 CFR

    2011-10-01

    ... 49 Transportation 4 2011-10-01 2011-10-01 false Movement of occupied camp cars. 218.80 Section 218... ADMINISTRATION, DEPARTMENT OF TRANSPORTATION RAILROAD OPERATING PRACTICES Protection of Occupied Camp Cars § 218.80 Movement of occupied camp cars. Occupied cars may not be humped or flat switched unless coupled...