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Sample records for lysophosphatidic acid-3 receptor-mediated

  1. Lysophosphatidic Acid (LPA Signaling in Human and Ruminant Reproductive Tract

    Directory of Open Access Journals (Sweden)

    Izabela Wocławek-Potocka

    2014-01-01

    Full Text Available Lysophosphatidic acid (LPA through activating its G protein-coupled receptors (LPAR 1–6 exerts diverse cellular effects that in turn influence several physiological processes including reproductive function of the female. Studies in various species of animals and also in humans have identified important roles for the receptor-mediated LPA signaling in multiple aspects of human and animal reproductive tract function. These aspects range from ovarian and uterine function, estrous cycle regulation, early embryo development, embryo implantation, decidualization to pregnancy maintenance and parturition. LPA signaling can also have pathological consequences, influencing aspects of endometriosis and reproductive tissue associated tumors. The review describes recent progress in LPA signaling research relevant to human and ruminant reproduction, pointing at the cow as a relevant model to study LPA influence on the human reproductive performance.

  2. Ectopic expression of Crambe abyssinica lysophosphatidic acid ...

    African Journals Online (AJOL)

    USER

    2010-06-21

    Jun 21, 2010 ... lysophosphatidic acid acyltransferase in transgenic rapeseed increases its oil .... pathway [fatty acid desaturase-2 (BnFAD2, AY577313), fatty acid desaturase-3 ..... Acyltransferases from basic science to modified seed oils.

  3. In vitro effect of lysophosphatidic acid on proliferation, invasion and ...

    African Journals Online (AJOL)

    Purpose: To evaluate the effect of lysophosphatidic acid (LPA) on the proliferation, invasion and migration ... reproduction in any medium, provided the original work is properly credited. Tropical .... air dried at room temperature overnight.

  4. Induction of aryl hydrocarbon receptor-mediated and estrogen receptor-mediated activities, and modulation of cell proliferation by dinaphthofurans

    Czech Academy of Sciences Publication Activity Database

    Vondráček, Jan; Chramostová, Kateřina; Plíšková, M.; Bláha, L.; Brack, W.; Kozubík, Alois; Machala, M.

    2004-01-01

    Roč. 23, č. 9 (2004), s. 2214-2220 ISSN 0730-7268 R&D Projects: GA ČR GA525/03/1527 Institutional research plan: CEZ:AV0Z5004920 Keywords : aryl hydrocarbon receptor-mediated activity * estrogenicity * intercellular communication inhibition Subject RIV: BO - Biophysics Impact factor: 2.121, year: 2004

  5. Receptor-mediated gene delivery using chemically modified chitosan

    International Nuclear Information System (INIS)

    Kim, T H; Jiang, H L; Nah, J W; Cho, M H; Akaike, T; Cho, C S

    2007-01-01

    Chitosan has been investigated as a non-viral vector because it has several advantages such as biocompatibility, biodegradability and low toxicity with high cationic potential. However, the low specificity and low transfection efficiency of chitosan need to be solved prior to clinical application. In this paper, we focused on the galactose or mannose ligand modification of chitosan for enhancement of cell specificity and transfection efficiency via receptor-mediated endocytosis in vitro and in vivo

  6. Characterization of GABA/sub A/ receptor-mediated 36chloride uptake in rat brain synaptoneurosomes

    International Nuclear Information System (INIS)

    Luu, M.D.; Morrow, A.L.; Paul, S.M.; Schwartz, R.D.

    1987-01-01

    γ-Aminobutyric acid (GABA) receptor-mediated 36 chloride ( 36 Cl - ) uptake was measured in synaptoneurosomes from rat brain. GABA and GABA agonists stimulated 36 Cl - uptake in a concentration-dependent manner with the following order of potency: Muscimol>GABA>piperidine-4-sulfonic acid (P4S)>4,5,6,7-tetrahydroisoxazolo-[5,4-c]pyridin-3-ol (THIP)=3-aminopropanesulfonic acid (3APS)>>taurine. Both P4S and 3APS behaved as partial agonists, while the GABA/sub B/ agonist, baclofen, was ineffective. The response to muscimol was inhibited by bicuculline and picrotoxin in a mixed competitive/non-competitive manner. Other inhibitors of GABA receptor-opened channels or non-neuronal anion channels such as penicillin, picrate, furosemide and disulfonic acid stilbenes also inhibited the response to muscimol. A regional variation in muscimol-stimulated 36 Cl - uptake was observed; the largest responses were observed in the cerebral cortex, cerebellum and hippocampus, moderate responses were obtained in the striatum and hypothalamus and the smallest response was observed in the pons-medulla. GABA receptor-mediated 36 Cl - uptake was also dependent on the anion present in the media. The muscinol response varied in media containing the following anions: Br - >Cl - ≥NO 3 - >I - ≥SCN - >>C 3 H 5 OO - ≥ClO 4 - >F - , consistent with the relative anion permeability through GABA receptor-gated anion channels and the enhancement of convulsant binding to the GABA receptor-gated Cl - channel. 43 references, 4 figures, 3 tables

  7. Spiroguanidine rhodamines as fluorogenic probes for lysophosphatidic acid

    Science.gov (United States)

    Wang, Lei; Sibrian-Vazquez, Martha; Escobedo, Jorge O.; Wang, Jialu; Moore, Richard G.

    2015-01-01

    Direct determination of total lysophosphatidic acid (LPA) was accomplished using newly developed spiroguanidines derived from rhodamine B as universal fluorogenic probes. Optimum conditions for the quantitative analysis of total LPA were investigated. The linear range for the determination of total LPA is up to 5 μM with a limit of detection of 0.512 μM. PMID:25516957

  8. In vitro effect of lysophosphatidic acid on proliferation, invasion and ...

    African Journals Online (AJOL)

    Purpose: To evaluate the effect of lysophosphatidic acid (LPA) on the proliferation, invasion and migration ability of 3AO, SKOV3 and CAOV3 human ovarian cancer cell lines. Methods: SKOV3, 3AO and CAOV3 cell lines were respectively treated with LPA. Changes in the proliferation rate of these cell lines were observed ...

  9. The ins and outs of lysophosphatidic acid signaling

    NARCIS (Netherlands)

    Moolenaar, Wouter H; van Meeteren, Laurens A; Giepmans, Ben N G

    Lysophosphatidic acid (LPA) is a lipid mediator with a wide variety of biological actions, particularly as an inducer of cell proliferation, migration and survival. LPA binds to specific G-protein-coupled receptors and thereby activates multiple signal transduction pathways, including those

  10. The liver taxis of receptor mediated lactosaminated human growth hormone

    International Nuclear Information System (INIS)

    Chen Zelian; Shi Lin; Li Tongling; Pang Qijie; He Juying; Guan Changtian

    2002-01-01

    Radiography imaging is used to assess liver taxis mechanism of anti-dwarfism drug lactosaminated human growth hormone (L-rhGH). Both L-rhGH and rhGH labelled with 131 I are used to study their biodistribution in animals (including rabbits, cocks and rats). The results show that L-rhGH is of specific hepatic targeting property, and the maximum hepatic concentration rate is 76.8%, which is two times of rhGH. Its hepatic binding is receptor mediated

  11. Stronger Dopamine D1 Receptor-Mediated Neurotransmission in Dyskinesia.

    Science.gov (United States)

    Farré, Daniel; Muñoz, Ana; Moreno, Estefanía; Reyes-Resina, Irene; Canet-Pons, Júlia; Dopeso-Reyes, Iria G; Rico, Alberto J; Lluís, Carme; Mallol, Josefa; Navarro, Gemma; Canela, Enric I; Cortés, Antonio; Labandeira-García, José L; Casadó, Vicent; Lanciego, José L; Franco, Rafael

    2015-12-01

    Radioligand binding assays to rat striatal dopamine D1 receptors showed that brain lateralization of the dopaminergic system were not due to changes in expression but in agonist affinity. D1 receptor-mediated striatal imbalance resulted from a significantly higher agonist affinity in the left striatum. D1 receptors heteromerize with dopamine D3 receptors, which are considered therapeutic targets for dyskinesia in parkinsonian patients. Expression of both D3 and D1-D3 receptor heteromers were increased in samples from 6-hydroxy-dopamine-hemilesioned rats rendered dyskinetic by treatment with 3, 4-dihydroxyphenyl-L-alanine (L-DOPA). Similar findings were obtained using striatal samples from primates. Radioligand binding studies in the presence of a D3 agonist led in dyskinetic, but not in lesioned or L-DOPA-treated rats, to a higher dopamine sensitivity. Upon D3-receptor activation, the affinity of agonists for binding to the right striatal D1 receptor increased. Excess dopamine coming from L-DOPA medication likely activates D3 receptors thus making right and left striatal D1 receptors equally responsive to dopamine. These results show that dyskinesia occurs concurrently with a right/left striatal balance in D1 receptor-mediated neurotransmission.

  12. Receptor-mediated endocytosis of trichosanthin in choriocarcinoma cells

    International Nuclear Information System (INIS)

    Chan, W.Y.; Huang Hai; Tam, S.-C.

    2003-01-01

    Trichosanthin (TCS) is a ribosome inactivating protein (RIP). It is generally believed that its many biological activities act through inhibition of ribosomes resulting in a decrease in protein synthesis. It has been hypothesized that the rate of entry of TCS into cells to reach ribosomes is an important factor in determining its biological activity. To prove this hypothesis, we have mapped out and compared the intracellular routing of TCS in two cell lines, namely the choriocarcinoma JAR cell line, which is known to be highly sensitive to the toxic effects of TCS, and the hepatoma H35 cell line, to which TCS shows minimal toxicity. Results from laser scanning confocal microscopy indicated that fluorescein isothiocyanate labeled TCS quickly accumulated inside JAR cells within 4 h of incubation while only a low level of fluorescent signals was detected in H35 cells during the same period of time. When TCS was conjugated with gold particles (Au) and its intracellular locations were traced with a transmission electron microscope, it was found that most of TCS were bound to coated pits on the JAR cell surface and were rapidly internalized within an hour. By 4 h, TCS reached almost every cytoplasmic region including ribosomes, and the JAR cell began to degenerate. In H35 cells, however, the binding of TCS to coated pits was not observed, but instead, a small amount of TCS was found to penetrate the cell non-specifically by direct diffusion across the cell membrane. Our observations suggest that most of TCS enter JAR cells via a specific receptor mediated pathway, which allows a swift transport of TCS across the membrane and a rapid accumulation of intracellular TCS, while in H35 cells, TCS takes a slow and non-specific route. The receptor-mediated uptake together with the specific intracellular routing of TCS may partly account for the differential vulnerability of the choriocarcinoma cell line towards the toxicity of TCS

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

    Lifescience Database Archive (English)

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

  14. Receptor-Mediated Endocytosis and Brain Delivery of Therapeutic Biologics

    Directory of Open Access Journals (Sweden)

    Guangqing Xiao

    2013-01-01

    Full Text Available Transport of macromolecules across the blood-brain-barrier (BBB requires both specific and nonspecific interactions between macromolecules and proteins/receptors expressed on the luminal and/or the abluminal surfaces of the brain capillary endothelial cells. Endocytosis and transcytosis play important roles in the distribution of macromolecules. Due to the tight junction of BBB, brain delivery of traditional therapeutic proteins with large molecular weight is generally not possible. There are multiple pathways through which macromolecules can be taken up into cells through both specific and nonspecific interactions with proteins/receptors on the cell surface. This review is focused on the current knowledge of receptor-mediated endocytosis/transcytosis and brain delivery using the Angiopep-2-conjugated system and the molecular Trojan horses. In addition, the role of neonatal Fc receptor (FcRn in regulating the efflux of Immunoglobulin G (IgG from brain to blood, and approaches to improve the pharmacokinetics of therapeutic biologics by generating Fc fusion proteins, and increasing the pH dependent binding affinity between Fc and FcRn, are discussed.

  15. Somatostatin receptor-mediated imaging and therapy: basic science, current knowledge, limitations and future perspectives

    Energy Technology Data Exchange (ETDEWEB)

    Breeman, W.A.P.; Jong, M. de; Kwekkeboom, D.J.; Valkema, R.; Bakker, W.H.; Kooij, P.P.M. [Dept. of Nuclear Medicine, Erasmus Medical Centre Rotterdam (Netherlands); Visser, T.J. [Dept. of Internal Medicine, Erasmus Medical Centre Rotterdam (Netherlands); Krenning, E.P. [Dept. of Nuclear Medicine, Erasmus Medical Centre Rotterdam (Netherlands); Dept. of Internal Medicine, Erasmus Medical Centre Rotterdam (Netherlands)

    2001-09-01

    In vivo somatostatin receptor-mediated scintigraphy has proven to be a valuable method for the visualisation of neuroendocrine tumours and their metastases. A new application is the use of radiolabelled analogues for somatostatin receptor-mediated therapy. This paper presents a review on the basic science, historical background and current knowledge of somatostatin receptor subtypes and their expression in neuroendocrine tumours. New somatostatin analogues, new chelators, ''new'' radionuclides and combinations thereof are also discussed. Due attention is given to limitations and future perspectives of somatostatin receptor-mediated imaging and therapy. (orig.)

  16. Somatostatin receptor-mediated imaging and therapy: basic science, current knowledge, limitations and future perspectives

    International Nuclear Information System (INIS)

    Breeman, W.A.P.; Jong, M. de; Kwekkeboom, D.J.; Valkema, R.; Bakker, W.H.; Kooij, P.P.M.; Visser, T.J.; Krenning, E.P.

    2001-01-01

    In vivo somatostatin receptor-mediated scintigraphy has proven to be a valuable method for the visualisation of neuroendocrine tumours and their metastases. A new application is the use of radiolabelled analogues for somatostatin receptor-mediated therapy. This paper presents a review on the basic science, historical background and current knowledge of somatostatin receptor subtypes and their expression in neuroendocrine tumours. New somatostatin analogues, new chelators, ''new'' radionuclides and combinations thereof are also discussed. Due attention is given to limitations and future perspectives of somatostatin receptor-mediated imaging and therapy. (orig.)

  17. Fcγ receptor-mediated inflammation inhibits axon regeneration.

    Directory of Open Access Journals (Sweden)

    Gang Zhang

    Full Text Available Anti-glycan/ganglioside antibodies are the most common immune effectors found in patients with Guillain-Barré Syndrome, which is a peripheral autoimmune neuropathy. We previously reported that disease-relevant anti-glycan autoantibodies inhibited axon regeneration, which echo the clinical association of these antibodies and poor recovery in Guillain-Barré Syndrome. However, the specific molecular and cellular elements involved in this antibody-mediated inhibition of axon regeneration are not previously defined. This study examined the role of Fcγ receptors and macrophages in the antibody-mediated inhibition of axon regeneration. A well characterized antibody passive transfer sciatic nerve crush and transplant models were used to study the anti-ganglioside antibody-mediated inhibition of axon regeneration in wild type and various mutant and transgenic mice with altered expression of specific Fcγ receptors and macrophage/microglia populations. Outcome measures included behavior, electrophysiology, morphometry, immunocytochemistry, quantitative real-time PCR, and western blotting. We demonstrate that the presence of autoantibodies, directed against neuronal/axonal cell surface gangliosides, in the injured mammalian peripheral nerves switch the proregenerative inflammatory environment to growth inhibitory milieu by engaging specific activating Fcγ receptors on recruited monocyte-derived macrophages to cause severe inhibition of axon regeneration. Our data demonstrate that the antibody orchestrated Fcγ receptor-mediated switch in inflammation is one mechanism underlying inhibition of axon regeneration. These findings have clinical implications for nerve repair and recovery in antibody-mediated immune neuropathies. Our results add to the complexity of axon regeneration in injured peripheral and central nervous systems as adverse effects of B cells and autoantibodies on neural injury and repair are increasingly recognized.

  18. Lysophosphatidic acid triggers mast cell-driven atherosclerotic plaque destabilization by increasing vascular inflammation.

    NARCIS (Netherlands)

    Bot, M.; , van, Berkel T.J.C.

    2013-01-01

    Lysophosphatidic acid (LPA), a bioactive lysophospholipid, accumulates in the atherosclerotic plaque. It has the capacity to activate mast cells, which potentially exacerbates plaque progression. In this study, we thus aimed to investigate whether LPA contributes to plaque destabilization by

  19. Lysophosphatidic acid metabolism and elimination in cardiovascular disease

    Science.gov (United States)

    Salous, Abdelghaffar Kamal

    The bioactive lipids lysophosphatidic acid (LPA) and sphingosine 1-phosphate (S1P) are present in human and mouse plasma at a concentration of ~0.1-1 microM and regulate physiological and pathophysiological processes in the cardiovascular system including atherothrombosis, intimal hyperplasia, and immune function, edema formation, and permeability. PPAP2B, the gene encoding LPP3, a broad activity integral membrane enzyme that terminates LPA actions in the vasculature, has a single nucleotide polymorphism that been recently associated with coronary artery disease risk. The synthesis and signaling of LPA and S1P in the cardiovascular system have been extensively studied but the mechanisms responsible for their elimination are less well understood. The broad goal of this research was to examine the role of LPP3 in the termination of LPA signaling in models of cardiovascular disease involving vascular wall cells, investigate the role of LPP3 in the elimination of plasma LPA, and further characterize the elimination of plasma LPA. The central hypothesis is that LPP3 plays an important role in attenuating the pathological responses to LPA signaling and that it mediates the elimination of exogenously applied bioactive lipids from the plasma. These hypotheses were tested using molecular biological approaches, in vitro studies, synthetic lysophospholipid mimetics, modified surgical procedures, and mass spectrometry assays. My results indicated that LPP3 played a critical role in attenuating LPA signaling mediating the pathological processes of intimal hyperplasia and vascular leak in mouse models of disease. Additionally, enzymatic inactivation of lysophospholipids by LPP and PLA enzymes in the plasma was not a primary mechanism for the rapid elimination of plasma LPA and S1P. Instead, evidence strongly suggested a transcellular uptake mechanism by hepatic non-parenchymal cells as the predominant mechanism for elimination of these molecules. These results support a model in

  20. Lysophosphatidic acid as a lipid mediator with multiple biological actions.

    Science.gov (United States)

    Aikawa, Shizu; Hashimoto, Takafumi; Kano, Kuniyuki; Aoki, Junken

    2015-02-01

    Lysophosphatidic acid (LPA) is one of the simplest glycerophospholipids with one fatty acid chain and a phosphate group as a polar head. Although LPA had been viewed just as a metabolic intermediate in de novo lipid synthetic pathways, it has recently been paid much attention as a lipid mediator. LPA exerts many kinds of cellular processes, such as cell proliferation and smooth muscle contraction, through cognate G protein-coupled receptors. Because lipids are not coded by the genome directly, it is difficult to know their patho- and physiological roles. However, recent studies have identified several key factors mediating the biological roles of LPA, such as receptors and producing enzymes. In addition, studies of transgenic and gene knockout animals for these LPA-related genes, have revealed the biological significance of LPA. In this review we will summarize recent advances in the studies of LPA production and its roles in both physiological and pathological conditions. © The Authors 2014. Published by Oxford University Press on behalf of the Japanese Biochemical Society. All rights reserved.

  1. Lysophosphatidic acid mediates pleiotropic responses in skeletal muscle cells

    International Nuclear Information System (INIS)

    Jean-Baptiste, Gael; Yang Zhao; Khoury, Chamel; Greenwood, Michael T.

    2005-01-01

    Lysophosphatidic acid (LPA) is a potent modulator of growth, cell survival, and apoptosis. Although all four LPA receptors are expressed in skeletal muscle, very little is known regarding the role they play in this tissue. We used RT-PCR to demonstrate that cultured skeletal muscle C2C12 cells endogenously express multiple LPA receptor subtypes. The demonstration that LPA mediates the activation of ERK1/2 MAP kinase and Akt/PKB in C2C12 cells is consistent with the widely observed mitogenic properties of LPA. In spite of these observations, LPA did not induce proliferation in C2C12 cells. Paradoxically, we found that prolonged treatment of C2C12 cells with LPA led to caspase 3 and PARP cleavage as well as the activation of stress-associated MAP kinases JNK and p38. In spite of these typically pro-apoptotic responses, LPA did not induce cell death. Blocking ERK1/2 and Akt/PKB activation with specific pharmacological inhibitors, nevertheless, stimulated LPA-mediated apoptosis. Taken together, these results suggest that both mitogenic and apoptotic responses serve to counterbalance the effects of LPA in cultured C2C12 cells

  2. Lysophosphatidic acid acyltransferase beta regulates mTOR signaling.

    Directory of Open Access Journals (Sweden)

    Michelle A Blaskovich

    Full Text Available Lysophosphatidic acid acyltransferase (LPAAT-β is a phosphatidic acid (PA generating enzyme that plays an essential role in triglyceride synthesis. However, LPAAT-β is now being studied as an important regulator of cell growth and differentiation and as a potential therapeutic target in cancer since PA is necessary for the activity of key proteins such as Raf, PKC-ζ and mTOR. In this report we determine the effect of LPAAT-β silencing with siRNA in pancreatic adenocarcinoma cell lines. We show for the first time that LPAAT-β knockdown inhibits proliferation and anchorage-independent growth of pancreatic cancer cells. This is associated with inhibition of signaling by mTOR as determined by levels of mTORC1- and mTORC2-specific phosphorylation sites on 4E-BP1, S6K and Akt. Since PA regulates the activity of mTOR by modulating its binding to FKBP38, we explored the possibility that LPAAT-β might regulate mTOR by affecting its association with FKBP38. Coimmunoprecipitation studies of FKBP38 with mTOR show increased levels of FKBP38 associated with mTOR when LPAAT-β protein levels are knocked down. Furthermore, depletion of LPAAT-β results in increased Lipin 1 nuclear localization which is associated with increased nuclear eccentricity, a nuclear shape change that is dependent on mTOR, further confirming the ability of LPAAT-β to regulate mTOR function. Our results provide support for the hypothesis that PA generated by LPAAT-β regulates mTOR signaling. We discuss the implications of these findings for using LPAAT-β as a therapeutic target.

  3. Upregulation of endothelin ETB receptor-mediated vasoconstriction in rat coronary artery after organ culture

    DEFF Research Database (Denmark)

    Eskesen, Karen; Edvinsson, Lars

    2006-01-01

    The aim of this study was to examine if endothelin ET(B) receptor-mediated contraction occurred in isolated segments of rat coronary arteries during organ culture. Presence of contractile endothelin ET(B) receptors was studied by measuring the change in isometric tension in rings of left anterior......(+)-solution was not modified after 1 day in culture medium. The experiments indicate that organ culture of rat coronary arteries upregulate endothelin ET(B) receptor-mediated contraction by inducing synthesis of new protein....... descending coronary arteries isolated from hearts of rats as response to application of the selective endothelin ET(B) receptor agonist, Sarafotoxin 6c and endothelin-1. In segments cultured 1 day in serum free Dulbecco's Modified Eagle's Medium, Sarafotoxin 6c induced a concentration dependent contraction...

  4. THIP, a hypnotic and antinociceptive drug, enhances a tonic GABAA receptor mediated conductance in mouse neocortex

    DEFF Research Database (Denmark)

    Drasbek, Kim Ryun; Jensen, Kimmo

    2006-01-01

    its cellular actions in the neocortex are uncertain, we studied the effects of THIP on neurons in slices of frontoparietal neocortex of 13- to 19-day-old (P13-19) mice. Using whole-cell patch-clamp recordings, we found that the clinically relevant THIP concentration of 1 μM induced a robust tonic GABA...... suggest that THIP activates an extrasynaptic GABA(A) receptor-mediated conductance in the neocortex, which may alter the cortical network activity....

  5. The Influence of Receptor-Mediated Interactions on Reaction-Diffusion Mechanisms of Cellular Self-organisation

    KAUST Repository

    Klika, Vá clav; Baker, Ruth E.; Headon, Denis; Gaffney, Eamonn A.

    2011-01-01

    formation, motivating numerous theoretical and experimental studies, though its verification at the molecular level in biological systems has remained elusive. In this work, we consider the influence of receptor-mediated dynamics within the framework

  6. Neuropeptide Y restores non-receptor-mediated vasoconstrictive action in superior mesenteric arteries in portal hypertension.

    Science.gov (United States)

    Hartl, Johannes; Dietrich, Peter; Moleda, Lukas; Müller-Schilling, Martina; Wiest, Reiner

    2015-12-01

    Vascular hyporeactivity to vasoconstrictors contributes to splanchnic arterial vasodilatation and hemodynamic dysregulation in portal hypertension. Neuropeptide Y (NPY), a sympathetic cotransmitter, has been shown to improve adrenergic vascular contractility in portal hypertensive rats and markedly attenuate hyperdynamic circulation. To further characterize the NPY-effects in portal hypertension, we investigated its role for non-receptor-mediated vasoconstriction in the superior mesenteric artery (SMA) of portal vein ligated (PVL) and sham-operated rats. Ex vivo SMA perfusion of PVL and sham rats was used to analyse the effects of NPY on pressure response to non-receptor-mediated vasoconstriction. Dose-response curves to KCl (30-300 mM) were used to bypass G protein-coupled receptor mechanisms. Potential involvement of the cyclooxygenase-pathway was tested by non-selective cyclooxygenase-inhibition using indomethacin. KCl-induced vascular contractility but not vascular sensitivity was significantly attenuated in PVL rats as compared with sham rats. Administration of NPY resulted in an augmentation of KCl-evoked vascular sensitivity being not different between study groups. However, KCl-induced vascular contractility was markedly more enhanced in PVL rats, thus, vascular response was no more significantly different between PVL and sham rats after addition of NPY. Administration of indomethacin abolished the NPY-induced enhancement of vasoconstriction. Receptor-independent vascular contractility is impaired in mesenteric arteries in portal hypertension. NPY improves non-receptor mediated mesenteric vasoconstriction more effective in portal hypertension than in healthy conditions correcting splanchnic vascular hyporesponsiveness. This beneficial vasoactive action of NPY adds to its well known more pronounced effects on adrenergic vasoconstriction in portal hypertension making it a promising therapeutic agent in portal hypertension. © 2015 John Wiley & Sons A

  7. Receptor-Mediated Drug Delivery to Macrophages in Chemotherapy of Leishmaniasis

    Science.gov (United States)

    Mukhopadhyay, Amitabha; Chaudhuri, Gautam; Arora, Sunil K.; Sehgal, Shobha; Basu, Sandip K.

    1989-05-01

    Methotrexate coupled to maleylated bovine serum albumin was taken up efficiently through the ``scavenger'' receptors present on macrophages and led to selective killing of intracellular Leishmania mexicana amazonensis amastigotes in cultured hamster peritoneal macrophages. The drug conjugate was nearly 100 times as effective as free methotrexate in eliminating the intracellular parasites. Furthermore, in a model of experimental cutaneous leishmaniasis in hamsters, the drug conjugate brought about more than 90% reduction in the size of footpad lesions within 11 days. In contrast, the free drug at a similar concentration did not significantly affect lesion size. These studies demonstrate the potential of receptor-mediated drug delivery in the therapy of macrophage-associated diseases.

  8. Lysophosphatidic acids are new substrates for the phosphatase domain of soluble epoxide hydrolase[S

    Science.gov (United States)

    Oguro, Ami; Imaoka, Susumu

    2012-01-01

    Soluble epoxide hydrolase (sEH) is a bifunctional enzyme that has a C-terminus epoxide hydrolase domain and an N-terminus phosphatase domain. The endogenous substrates of epoxide hydrolase are known to be epoxyeicosatrienoic acids, but the endogenous substrates of the phosphatase activity are not well understood. In this study, to explore the substrates of sEH, we investigated the inhibition of the phosphatase activity of sEH toward 4-methylumbelliferyl phosphate by using lecithin and its hydrolyzed products. Although lecithin itself did not inhibit the phosphatase activity, the hydrolyzed lecithin significantly inhibited it, suggesting that lysophospholipid or fatty acid can inhibit it. Next, we investigated the inhibition of phosphatase activity by lysophosphatidyl choline, palmitoyl lysophosphatidic acid, monopalmitoyl glycerol, and palmitic acid. Palmitoyl lysophosphatidic acid and fatty acid efficiently inhibited phosphatase activity, suggesting that lysophosphatidic acids (LPAs) are substrates for the phosphatase activity of sEH. As expected, palmitoyl, stearoyl, oleoyl, and arachidonoyl LPAs were efficiently dephosphorylated by sEH (Km, 3–7 μM; Vmax, 150–193 nmol/min/mg). These results suggest that LPAs are substrates of sEH, which may regulate physiological functions of cells via their metabolism. PMID:22217705

  9. Lysophosphatidic acids are new substrates for the phosphatase domain of soluble epoxide hydrolase.

    Science.gov (United States)

    Oguro, Ami; Imaoka, Susumu

    2012-03-01

    Soluble epoxide hydrolase (sEH) is a bifunctional enzyme that has a C-terminus epoxide hydrolase domain and an N-terminus phosphatase domain. The endogenous substrates of epoxide hydrolase are known to be epoxyeicosatrienoic acids, but the endogenous substrates of the phosphatase activity are not well understood. In this study, to explore the substrates of sEH, we investigated the inhibition of the phosphatase activity of sEH toward 4-methylumbelliferyl phosphate by using lecithin and its hydrolyzed products. Although lecithin itself did not inhibit the phosphatase activity, the hydrolyzed lecithin significantly inhibited it, suggesting that lysophospholipid or fatty acid can inhibit it. Next, we investigated the inhibition of phosphatase activity by lysophosphatidyl choline, palmitoyl lysophosphatidic acid, monopalmitoyl glycerol, and palmitic acid. Palmitoyl lysophosphatidic acid and fatty acid efficiently inhibited phosphatase activity, suggesting that lysophosphatidic acids (LPAs) are substrates for the phosphatase activity of sEH. As expected, palmitoyl, stearoyl, oleoyl, and arachidonoyl LPAs were efficiently dephosphorylated by sEH (Km, 3-7 μM; Vmax, 150-193 nmol/min/mg). These results suggest that LPAs are substrates of sEH, which may regulate physiological functions of cells via their metabolism.

  10. Understanding magnetic nanoparticle osteoblast receptor-mediated endocytosis using experiments and modeling

    International Nuclear Information System (INIS)

    Tran, Nhiem; Webster, Thomas J

    2013-01-01

    Iron oxide nanoparticles are promising candidates for controlling drug delivery through an external magnetic force to treat a wide range of diseases, including osteoporosis. Previous studies have demonstrated that in the presence of hydroxyapatite coated magnetite (Fe 3 O 4 ) nanoparticles, osteoblast (or bone forming cell) proliferation and long-term functions (such as calcium deposition) were significantly enhanced. Hydroxyapatite is the major inorganic component of bone. As a further attempt to understand why, in the current study, the uptake of such nanoparticles into osteoblasts was experimentally investigated and mathematically modeled. Magnetite nanoparticles were synthesized using a co-precipitation method and were coated with hydroxyapatite. A cellular uptake experiment at low temperatures indicated that receptor-mediated endocytosis contributed to the internalization of the magnetic nanoparticles into osteoblasts. A model was further developed to explain the uptake of magnetic nanoparticles into osteoblasts using receptor-mediated endocytosis. This model may explain the internalization of hydroxyapatite into osteoblasts to elevate intracellular calcium levels necessary to promote osteoblast functions to treat a wide range of orthopedic problems, including osteoporosis. (paper)

  11. Melanocortin MC(4) receptor-mediated feeding and grooming in rodents.

    Science.gov (United States)

    Mul, Joram D; Spruijt, Berry M; Brakkee, Jan H; Adan, Roger A H

    2013-11-05

    Decades ago it was recognized that the pharmacological profile of melanocortin ligands that stimulated grooming behavior in rats was strikingly similar to that of Xenopus laevis melanophore pigment dispersion. After cloning of the melanocortin MC1 receptor, expressed in melanocytes, and the melanocortin MC4 receptor, expressed mainly in brain, the pharmacological profiles of these receptors appeared to be very similar and it was demonstrated that these receptors mediate melanocortin-induced pigmentation and grooming respectively. Grooming is a low priority behavior that is concerned with care of body surface. Activation of central melanocortin MC4 receptors is also associated with meal termination, and continued postprandial stimulation of melanocortin MC4 receptors may stimulate natural postprandial grooming behavior as part of the behavioral satiety sequence. Indeed, melanocortins fail to suppress food intake or induce grooming behavior in melanocortin MC4 receptor-deficient rats. This review will focus on how melanocortins affect grooming behavior through the melanocortin MC4 receptor, and how melanocortin MC4 receptors mediate feeding behavior. This review also illustrates how melanocortins were the most likely candidates to mediate grooming and feeding based on the natural behaviors they induced. Copyright © 2013 Elsevier B.V. All rights reserved.

  12. Receptor-mediated photo-cytotoxicity: synthesis of a photoactivatable psoralen derivative conjugated to insulin.

    Science.gov (United States)

    Gasparro, F P; Knobler, R M; Yemul, S S; Bisaccia, E; Edelson, R L

    1986-12-15

    4'-Aminomethyl-4,5',8-trimethylpsoralen has been chemically conjugated to insulin using a carbodiimide derivative. The psoralen moiety retains its photochemical reactivity as evidenced by its ability to crosslink DNA after exposure to long wavelength ultraviolet light (UVA, 320-400 nm). This chimeric molecule has been used to selectively kill a population of lymphocytes whose expression of insulin receptors has been stimulated with phytohemagglutinin. Insulin carries the psoralen into the cell via receptor-mediated endocytosis, where it is subsequently activated by exposure to UVA light. The UVA induced activity of AMT-insulin can be blocked by the presence of native insulin. The viability of unstimulated lymphocytes was not affected by AMT-insulin and UVA light. The hybrid insulin-psoralen molecule may be a prototype for a family of phototoxic drugs which can be selectively delivered to subsets of lymphocytes.

  13. Platelet alpha-2 adrenergic receptor-mediated phosphoinositide responses in endogenous depression

    International Nuclear Information System (INIS)

    Mori, Hideki; Koyama, Tsukasa; Yamashita, Itaru

    1991-01-01

    We have previously indicated that epinephrine stimulates phosphoinositide (PI) hydrolysis by activating alpha-2 adrenergic receptors in human platelets. This method involves the measurement of the accumulation of [ 3 H]-inositol-1-phosphate (IP-1) as an index of Pl hydrolysis; lithium is added to inhibit the metabolism of IP-1, thus giving an enhanced signal. In the present study, we assessed the platelet alpha-2 adrenergic receptor-mediated PI responses in samples from 15 unmedicated patients with endogenous depression and 15 age- and sex-matched control subjects. The responses to epinephrine in the depressed patients were significantly higher than those of the controls, whereas the basal values did not differ significantly. These results support the hypothesis that platelet alpha-2 adrenergic receptors may be supersensitive in patients with endogenous depression

  14. Bicarbonate Contributes to GABAA Receptor-Mediated Neuronal Excitation in Surgically-Resected Human Hypothalamic Hamartomas

    Science.gov (United States)

    Do-Young, Kim; Fenoglio, Kristina A.; Kerrigan, John F.; Rho, Jong M.

    2009-01-01

    SUMMARY The role of bicarbonate (HCO3-) in GABAA receptor-mediated depolarization of human hypothalamic hamartoma (HH) neurons was investigated using cellular electrophysiological and calcium imaging techniques. Activation of GABAA receptors with muscimol (30 μM) provoked neuronal excitation in over 70% of large (18-22 μM) HH neurons in HCO3- buffer. Subsequent perfusion of HCO3--free HEPES buffer produced partial suppression of muscimol-induced excitation. Additionally, 53% of large HH neurons under HCO3--free conditions exhibited reduced intracellular calcium accumulation by muscimol. These results suggest that HCO3- efflux through GABAA receptors on a subpopulation of large HH neurons may contribute to membrane depolarization and subsequent activation of L-type calcium channels. PMID:19022626

  15. Gelidium amansii promotes dendritic spine morphology and synaptogenesis, and modulates NMDA receptor-mediated postsynaptic current.

    Science.gov (United States)

    Hannan, Md Abdul; Mohibbullah, Md; Hong, Yong-Ki; Nam, Joo Hyun; Moon, Il Soo

    2014-01-01

    Neurotrophic factors are essential for the differentiation and maturation of developing neurons as well as providing survival support to the mature neurons. Moreover, therapeutically neurotrophic factors are promising to reconstruct partially damaged neuronal networks in neurodegenerative diseases. In the previous study, we reported that the ethanol extract of an edible marine alga, Gelidium amansii (GAE) had shown promising effects in the development and maturation of both axon and dendrites of hippocampal neurons. Here, we demonstrate that in primary culture of hippocampal neurons (1) GAE promotes a significant increase in the number of filopodia and dendritic spines; (2) promotes synaptogenesis; (3) enhances N-methyl-D-aspartic acid (NMDA) receptor recruitment; and (4) modulates NMDA-receptor-mediated postsynaptic current. Taken together these findings that GAE might be involved in both morphological and functional maturation of neurons suggest the possibility that GAE may constitute a promising candidate for novel compounds for the prevention and treatment of neurodegenerative diseases.

  16. Multifunctional pH-Responsive Folate Receptor Mediated Polymer Nanoparticles for Drug Delivery.

    Science.gov (United States)

    Cai, Xiaoqing; Yang, Xiaoye; Wang, Fang; Zhang, Chen; Sun, Deqing; Zhai, Guangxi

    2016-07-01

    Multifunctional pH-responsive folate receptor mediated targeted polymer nanoparticles (TPNps) were developed for docetaxel (DTX) delivery based on poly(ethylene glycol)-block-poly(propylene glycol)-block-poly(ethylene glycol)poly (β-amino ester) (P123-PAE) and poly(ethylene glycol)-block-poly(propylene glycol)-block-poly(ethylene glycol)-folate (P123-FA) copolymers. The DTX was loaded into the TPNps with a decent drug loading content of 15.02 ± 0.14 wt%. In vitro drug release results showed that the DTX was released from the TPNps at a pH-dependent manner. Tetrazolium dye (MTT) assay revealed that the bland polymer nanoparticles displayed almost nontoxicity at 200 μg/mL concentration. However, the DTX-loaded TPNps showed high anti-tumor activity at low IC50 (0.72 μg/mL) for MCF-7 cells following 48 h incubation. Cellular uptake experiments revealed that the TPNps had higher degree of cellular uptake than nontargeted polymer nanoparticles, indicating that the nanoparticles were internalized into the cells via FA receptor-mediated endocytosis. Moreover, the cellular uptake pathways for the FA grafted polymer were involved in energy-dependent, clathrin-mediated and caveolae-mediated endocytosis. The cell killing effect and cellular uptake of the DTX-TPNps by the MCF-7 cells were all enhanced by about two folds at pH 5.5 when compared with pH 7.4. The TPNps also significantly prolonged the in vivo retention time for the DTX. These results suggest that the biocompatible pH responsive folate-modified polymer nanoparticles present a promising safe nanosystem for intracellular targeted delivery of DTX.

  17. Receptor-mediated uptake of low density lipoprotein stimulates bile acid synthesis by cultured rat hepatocytes

    International Nuclear Information System (INIS)

    Junker, L.H.; Davis, R.A.

    1989-01-01

    The cellular mechanisms responsible for the lipoprotein-mediated stimulation of bile acid synthesis in cultured rat hepatocytes were investigated. Adding 280 micrograms/ml of cholesterol in the form of human or rat low density lipoprotein (LDL) to the culture medium increased bile acid synthesis by 1.8- and 1.6-fold, respectively. As a result of the uptake of LDL, the synthesis of [14C]cholesterol from [2-14C]acetate was decreased and cellular cholesteryl ester mass was increased. Further studies demonstrated that rat apoE-free LDL and apoE-rich high density lipoprotein (HDL) both stimulated bile acid synthesis 1.5-fold, as well as inhibited the formation of [14C]cholesterol from [2-14C]acetate. Reductive methylation of LDL blocked the inhibition of cholesterol synthesis, as well as the stimulation of bile acid synthesis, suggesting that these processes require receptor-mediated uptake. To identify the receptors responsible, competitive binding studies using 125I-labeled apoE-free LDL and 125I-labeled apoE-rich HDL were performed. Both apoE-free LDL and apoE-rich HDL displayed an equal ability to compete for binding of the other, suggesting that a receptor or a group of receptors that recognizes both apolipoproteins is involved. Additional studies show that hepatocytes from cholestyramine-treated rats displayed 2.2- and 3.4-fold increases in the binding of apoE-free LDL and apoE-rich HDL, respectively. These data show for the first time that receptor-mediated uptake of LDL by the liver is intimately linked to processes activating bile acid synthesis

  18. The role of receptor-mediated T-cells activation disorders in pulmonary tuberculosis

    Directory of Open Access Journals (Sweden)

    Irina E. Esimova

    2017-01-01

    Full Text Available Aim. To analyze the peculiarities and mechanisms of receptor-mediated T-lymphocytes disorders in different clinical forms of pulmonary tuberculosis.Materials and мethods. The study involved 116 patients with first diagnosed infiltrative and disseminated drug-sensitive and drug-resistant pulmonary tuberculosis. The key stages in receptor-mediated activation of T-lymphocytes, isolated from blood, after their CD3/CD28-induction in vitro with addition of intracellular transport blocker were analyzed. Their immunotyping was carried out with the method of two- and threecolor flow cytofluorometry. The obtained results were statistically analyzed.Results. The breach of extracellular and intracellular stages of T-lymphocytes activation, shown by reduction in total number of CD3- and CD28-positive cells, and CD3+CD28+IL2+, CD3+CD28+IL2–, CD3+NF-kB+, CD3+NFAT2+ lymphocytes, and increase in number of CD3+CTLA4+ cells, was identified with most of their manifestations in disseminated drug-resistant pulmonary tuberculosis. It was shown that the content of CD3+AP-1+ lymphocytes is variable in drug-resistant pulmonary tuberculosis: it increases in the infiltrative form and decreases in the disseminated form.Conclusion. The results showed different mechanisms leading to a deficiency of IL-2-positive lymphocytes and T-lymphocytopenia: from “functional reserve” exhaustion of T-cells in drug-sensitive pulmonary tuberculosis to immunosuppression under the influence of suppressive cytokines (in case of the infiltrative form and inhibitory protein CTLA4 (in case of the disseminated form in drug-resistant pulmonary tuberculosis. 

  19. Effects of sarcosine and N, N-dimethylglycine on NMDA receptor-mediated excitatory field potentials.

    Science.gov (United States)

    Lee, Mei-Yi; Lin, Yi-Ruu; Tu, Yi-Shu; Tseng, Yufeng Jane; Chan, Ming-Huan; Chen, Hwei-Hsien

    2017-02-28

    Sarcosine, a glycine transporter type 1 inhibitor and an N-methyl-D-aspartate (NMDA) receptor co-agonist at the glycine binding site, potentiates NMDA receptor function. Structurally similar to sarcosine, N,N-dimethylglycine (DMG) is also N-methyl glycine-derivative amino acid and commonly used as a dietary supplement. The present study compared the effects of sarcosine and DMG on NMDA receptor-mediated excitatory field potentials (EFPs) in mouse medial prefrontal cortex brain slices using a multi-electrode array system. Glycine, sarcosine and DMG alone did not alter the NMDA receptor-mediated EFPs, but in combination with glutamate, glycine and its N-methyl derivatives significantly increased the frequency and amplitude of EFPs. The enhancing effects of glycine analogs in combination with glutamate on EFPs were remarkably reduced by the glycine binding site antagonist 7-chlorokynurenate (7-CK). However, DMG, but not sarcosine, reduced the frequency and amplitude of EFPs elicited by co-application of glutamate plus glycine. D-cycloserine, a partial agonist at the glycine binding site on NMDA receptors, affected EFPs in a similar manner to DMG. Furthermore, DMG, but not sarcosine, reduced the frequencies and amplitudes of EFPs elicited by glutamate plus D-serine, another endogenous ligand for glycine binding site. These findings suggest that sarcosine acts as a full agonist, yet DMG is a partial agonist at glycine binding site of NMDA receptors. The molecular docking analysis indicated that the interactions of glycine, sarcosine, and DMG to NMDA receptors are highly similar, supporting that the glycine binding site of NMDA receptors is a critical target site for sarcosine and DMG.

  20. Fc-receptor-mediated phagocytosis is regulated by mechanical properties of the target

    Science.gov (United States)

    Beningo, Karen A.; Wang, Yu-li

    2002-01-01

    Phagocytosis is an actin-based process used by macrophages to clear particles greater than 0.5 microm in diameter. In addition to its role in immunological responses, phagocytosis is also necessary for tissue remodeling and repair. To prevent catastrophic autoimmune reactions, phagocytosis must be tightly regulated. It is commonly assumed that the recognition/selection of phagocytic targets is based solely upon receptor-ligand binding. Here we report an important new criterion, that mechanical parameters of the target can dramatically affect the efficiency of phagocytosis. When presented with particles of identical chemical properties but different rigidity, macrophages showed a strong preference to engulf rigid objects. Furthermore, phagocytosis of soft particles can be stimulated with the microinjection of constitutively active Rac1 but not RhoA, and with lysophosphatidic acid, an agent known to activate the small GTP-binding proteins of the Rho family. These data suggest a Rac1-dependent mechanosensory mechanism for phagocytosis, which probably plays an important role in a number of physiological and pathological processes from embryonic development to autoimmune diseases.

  1. The Influence of Receptor-Mediated Interactions on Reaction-Diffusion Mechanisms of Cellular Self-organisation

    Czech Academy of Sciences Publication Activity Database

    Klika, Václav; Baker, R. E.; Headon, D.; Gaffney, E. A.

    2012-01-01

    Roč. 74, č. 4 (2012), s. 935-957 ISSN 0092-8240 Institutional research plan: CEZ:AV0Z20760514 Keywords : reaction-diffusion * receptor-mediated patterning * turing models Subject RIV: BO - Biophysics Impact factor: 2.023, year: 2012 http://www.springerlink.com/content/9713544x6871w4n6/?MUD=MP

  2. Vascular endothelin ET(B) receptor-mediated contraction requires phosphorylation of ERK1/2 proteins

    DEFF Research Database (Denmark)

    Luo, Guogang; Jamali, Roya; Cao, Yong-Xiao

    2006-01-01

    In cardiovascular diseases, endothelin type B (ET(B)) receptors in arterial smooth muscle cells are upregulated. The present study revealed that organ culture of rat mesenteric artery segments enhanced endothelin ET(B) receptor-mediated contraction paralleled with increase in the receptor mRNA an...

  3. Lysophosphatidic acid generation by pulmonary NKT cell ENPP-2/autotaxin exacerbates hyperoxic lung injury.

    Science.gov (United States)

    Nowak-Machen, Martina; Lange, Martin; Exley, Mark; Wu, Sherry; Usheva, Anny; Robson, Simon C

    2015-12-01

    Hyperoxia is still broadly used in clinical practice in order to assure organ oxygenation in critically ill patients, albeit known toxic effects. In this present study, we hypothesize that lysophosphatidic acid (LPA) mediates NKT cell activation in a mouse model of hyperoxic lung injury. In vitro, pulmonary NKT cells were exposed to hyperoxia for 72 h, and the induction of the ectonucleotide pyrophosphatase/phosphodiesterase 2 (ENPP-2) was examined and production of lysophosphatidic acid (LPA) was measured. In vivo, animals were exposed to 100 % oxygen for 72 h and lungs and serum were harvested. Pulmonary NKT cells were then incubated with the LPA antagonist Brp-LPA. Animals received BrP-LPA prior to oxygen exposure. Autotaxin (ATX, ENPP-2) was significantly up-regulated on pulmonary NKT cells after hyperoxia (p NKT cells. LPA levels were significantly reduced by incubating NKT cells with LPA-BrP during oxygen exposure (p NKT cell numbers in vivo. BrP-LPA injection significantly improved survival as well as significantly decreased lung injury and lowered pulmonary NKT cell numbers. We conclude that NKT cell-induced hyperoxic lung injury is mediated by pro-inflammatory LPA generation, at least in part, secondary to ENPP-2 up-regulation on pulmonary NKT cells. Being a potent LPA antagonist, BrP-LPA prevents hyperoxia-induced lung injury in vitro and in vivo.

  4. Nicotine impairs cyclooxygenase-2-dependent kinin-receptor-mediated murine airway relaxations

    International Nuclear Information System (INIS)

    Xu, Yuan; Cardell, Lars-Olaf

    2014-01-01

    Introduction: Cigarette smoke induces local inflammation and airway hyperreactivity. In asthmatics, it worsens the symptoms and increases the risk for exacerbation. The present study investigates the effects of nicotine on airway relaxations in isolated murine tracheal segments. Methods: Segments were cultured for 24 h in the presence of vehicle, nicotine (10 μM) and/or dexamethasone (1 μM). Airway relaxations were assessed in myographs after pre-contraction with carbachol (1 μM). Kinin receptors, cyclooxygenase (COX) and inflammatory mediator expressions were assessed by real-time PCR and confocal-microscopy-based immunohistochemistry. Results: The organ culture procedure markedly increased bradykinin- (selective B 2 receptor agonist) and des-Arg 9 -bradykinin- (selective B 1 receptor agonist) induced relaxations, and slightly increased relaxation induced by isoprenaline, but not that induced by PGE 2 . The kinin receptor mediated relaxations were epithelium-, COX-2- and EP2-receptor-dependent and accompanied by drastically enhanced mRNA levels of kinin receptors, as well as inflammatory mediators MCP-1 and iNOS. Increase in COX-2 and mPGES-1 was verified both at mRNA and protein levels. Nicotine selectively suppressed the organ-culture-enhanced relaxations induced by des-Arg 9 -bradykinin and bradykinin, at the same time reducing mPGES-1 mRNA and protein expressions. α7-nicotinic acetylcholine receptor inhibitors α-bungarotoxin and MG624 both blocked the nicotine effects on kinin B 2 receptors, but not those on B 1 . Dexamethasone completely abolished kinin-induced relaxations. Conclusion: It is tempting to conclude that a local inflammatory process per se could have a bronchoprotective component by increasing COX-2 mediated airway relaxations and that nicotine could impede this safety mechanism. Dexamethasone further reduced airway inflammation together with relaxations. This might contribute to the steroid resistance seen in some patients with asthma

  5. Nicotine impairs cyclooxygenase-2-dependent kinin-receptor-mediated murine airway relaxations

    Energy Technology Data Exchange (ETDEWEB)

    Xu, Yuan, E-mail: yuan.xu@ki.se; Cardell, Lars-Olaf

    2014-02-15

    Introduction: Cigarette smoke induces local inflammation and airway hyperreactivity. In asthmatics, it worsens the symptoms and increases the risk for exacerbation. The present study investigates the effects of nicotine on airway relaxations in isolated murine tracheal segments. Methods: Segments were cultured for 24 h in the presence of vehicle, nicotine (10 μM) and/or dexamethasone (1 μM). Airway relaxations were assessed in myographs after pre-contraction with carbachol (1 μM). Kinin receptors, cyclooxygenase (COX) and inflammatory mediator expressions were assessed by real-time PCR and confocal-microscopy-based immunohistochemistry. Results: The organ culture procedure markedly increased bradykinin- (selective B{sub 2} receptor agonist) and des-Arg{sup 9}-bradykinin- (selective B{sub 1} receptor agonist) induced relaxations, and slightly increased relaxation induced by isoprenaline, but not that induced by PGE{sub 2}. The kinin receptor mediated relaxations were epithelium-, COX-2- and EP2-receptor-dependent and accompanied by drastically enhanced mRNA levels of kinin receptors, as well as inflammatory mediators MCP-1 and iNOS. Increase in COX-2 and mPGES-1 was verified both at mRNA and protein levels. Nicotine selectively suppressed the organ-culture-enhanced relaxations induced by des-Arg{sup 9}-bradykinin and bradykinin, at the same time reducing mPGES-1 mRNA and protein expressions. α7-nicotinic acetylcholine receptor inhibitors α-bungarotoxin and MG624 both blocked the nicotine effects on kinin B{sub 2} receptors, but not those on B{sub 1}. Dexamethasone completely abolished kinin-induced relaxations. Conclusion: It is tempting to conclude that a local inflammatory process per se could have a bronchoprotective component by increasing COX-2 mediated airway relaxations and that nicotine could impede this safety mechanism. Dexamethasone further reduced airway inflammation together with relaxations. This might contribute to the steroid resistance seen in

  6. Enhanced NMDA receptor-mediated intracellular calcium signaling in magnocellular neurosecretory neurons in heart failure rats.

    Science.gov (United States)

    Stern, Javier E; Potapenko, Evgeniy S

    2013-08-15

    An enhanced glutamate excitatory function within the hypothalamic supraoptic and paraventricluar nuclei is known to contribute to increased neurosecretory and presympathetic neuronal activity, and hence, neurohumoral activation, during heart failure (HF). Still, the precise mechanisms underlying enhanced glutamate-driven neuronal activity in HF remain to be elucidated. Here, we performed simultaneous electrophysiology and fast confocal Ca²⁺ imaging to determine whether altered N-methyl-d-aspartate (NMDA) receptor-mediated changes in intracellular Ca²⁺ levels (NMDA-ΔCa²⁺) occurred in hypothalamic magnocellular neurosecretory cells (MNCs) in HF rats. We found that activation of NMDA receptors resulted in a larger ΔCa²⁺ in MNCs from HF when compared with sham rats. The enhanced NMDA-ΔCa²⁺ was neither dependent on the magnitude of the NMDA-mediated current (voltage clamp) nor on the degree of membrane depolarization or firing activity evoked by NMDA (current clamp). Differently from NMDA receptor activation, firing activity evoked by direct membrane depolarization resulted in similar changes in intracellular Ca²⁺ in sham and HF rats. Taken together, our results support a relatively selective alteration of intracellular Ca²⁺ homeostasis and signaling following activation of NMDA receptors in MNCs during HF. The downstream functional consequences of such altered ΔCa²⁺ signaling during HF are discussed.

  7. Novel isoforms of the TFIID subunit TAF4 modulate nuclear receptor-mediated transcriptional activity

    International Nuclear Information System (INIS)

    Brunkhorst, Adrian; Neuman, Toomas; Hall, Anita; Arenas, Ernest; Bartfai, Tamas; Hermanson, Ola; Metsis, Madis

    2004-01-01

    The transcription factor TFIID consists of TATA-binding protein (TBP) and TBP-associated factors (TAFs). TAFs are essential for modulation of transcriptional activity but the regulation of TAFs is complex and many important aspects remain unclear. In this study, we have identified and characterized five novel truncated forms of the TFIID subunit TAF4 (TAF II 135). Analysis of the mouse gene structure revealed that all truncations were the results of alternative splicing and resulted in the loss of domains or parts of domains implicated in TAF4 functional interactions. Results from transcriptional assays showed that several of the TAF4 isoforms exerted dominant negative effects on TAF4 activity in nuclear receptor-mediated transcriptional activation. In addition, alternative TAF4 isoforms could be detected in specific cell types. Our results indicate an additional level of complexity in TAF4-mediated regulation of transcription and suggest context-specific roles for these new TAF4 isoforms in transcriptional regulation in vivo

  8. Plant cell surface receptor-mediated signaling - a common theme amid diversity.

    Science.gov (United States)

    He, Yunxia; Zhou, Jinggeng; Shan, Libo; Meng, Xiangzong

    2018-01-29

    Sessile plants employ a diverse array of plasma membrane-bound receptors to perceive endogenous and exogenous signals for regulation of plant growth, development and immunity. These cell surface receptors include receptor-like kinases (RLKs) and receptor-like proteins (RLPs) that harbor different extracellular domains for perception of distinct ligands. Several RLK and RLP signaling pathways converge at the somatic embryogenesis receptor kinases (SERKs), which function as shared co-receptors. A repertoire of receptor-like cytoplasmic kinases (RLCKs) associate with the receptor complexes to relay intracellular signaling. Downstream of the receptor complexes, mitogen-activated protein kinase (MAPK) cascades are among the key signaling modules at which the signals converge, and these cascades regulate diverse cellular and physiological responses through phosphorylation of different downstream substrates. In this Review, we summarize the emerging common theme that underlies cell surface receptor-mediated signaling pathways in Arabidopsis thaliana : the dynamic association of RLKs and RLPs with specific co-receptors and RLCKs for signal transduction. We further discuss how signaling specificities are maintained through modules at which signals converge, with a focus on SERK-mediated receptor signaling. © 2018. Published by The Company of Biologists Ltd.

  9. Muscarinic receptor-mediated inositol tetrakisphosphate response in bovine adrenal chromaffin cells

    International Nuclear Information System (INIS)

    Sanborn, B.B.; Schneider, A.S.

    1990-01-01

    Inositol trisphosphate (IP 3 ), a product of the phosphoinositide cycle, mobilizes intracellular Ca 2+ in many cell types. New evidence suggests that inositol tetrakisphosphate (IP 4 ), an IP 3 derivative, may act as another second messenger to further alter calcium homeostasis. However, the function and mechanism of action of IP 4 are presently unresolved. We now report evidence of muscarinic receptor-mediated accumulation of IP 4 in bovine adrenal chromaffin cells, a classic neurosecretory system in which calcium movements have been well studied. Muscarine stimulated an increase in [ 3 H]IP 4 and [ 3 H]IP 3 accumulation in chromaffin cells and this effect was completely blocked by atropine. [ 3 H]IP 4 accumulation was detectable within 15 sec, increased to a maximum by 30 sec and thereafter declined. 2,3-diphosphoglycerate, an inhibitor of IP 3 and IP 4 hydrolysis, enhanced accumulation of these inositol polyphosphates. The results provide the first evidence of a rapid inositol tetrakisphosphate response in adrenal chromaffin cells, which should facilitate the future resolution of the relationship between IP 4 and calcium homeostasis

  10. Electroacupuncture Potentiates Cannabinoid Receptor-Mediated Descending Inhibitory Control in a Mouse Model of Knee Osteoarthritis

    Directory of Open Access Journals (Sweden)

    Xiao-Cui Yuan

    2018-04-01

    Full Text Available Knee osteoarthritis (KOA is a highly prevalent, chronic joint disorder, which can lead to chronic pain. Although electroacupuncture (EA is effective in relieving chronic pain in the clinic, the involved mechanisms remain unclear. Reduced diffuse noxius inhibitory controls (DNIC function is associated with chronic pain and may be related to the action of endocannabinoids. In the present study, we determined whether EA may potentiate cannabinoid receptor-mediated descending inhibitory control and inhibit chronic pain in a mouse model of KOA. We found that the optimized parameters of EA inhibiting chronic pain were the low frequency and high intensity (2 Hz + 1 mA. EA reversed the reduced expression of CB1 receptors and the 2-arachidonoylglycerol (2-AG level in the midbrain in chronic pain. Microinjection of the CB1 receptor antagonist AM251 into the ventrolateral periaqueductal gray (vlPAG can reversed the EA effect on pain hypersensitivity and DNIC function. In addition, CB1 receptors on GABAergic but not glutamatergic neurons are involved in the EA effect on DNIC function and descending inhibitory control of 5-HT in the medulla, thus inhibiting chronic pain. Our data suggest that endocannabinoid (2-AG-CB1R-GABA-5-HT may be a novel signaling pathway involved in the effect of EA improving DNIC function and inhibiting chronic pain.

  11. Receptor-mediated endocytosis of lysozyme in renal proximal tubules of the frog Rana temporaria

    Directory of Open Access Journals (Sweden)

    E.V. Seliverstova

    2015-04-01

    Full Text Available The mechanism of protein reabsorption in the kidney of lower vertebrates remains insufficiently investigated in spite of raising interest to the amphibian and fish kidneys as a useful model for physiological and pathophysiological examinations. In the present study, we examined the renal tubular uptake and the internalization rote of lysozyme after its intravenous injection in the wintering frog Rana temporaria using immunohisto- and immunocytochemistry and specific markers for some endocytic compartments. The distinct expression of megalin and cubilin in the proximal tubule cells of lysozyme-injected frogs was revealed whereas kidney tissue of control animals showed no positive immunoreactivity. Lysozyme was detected in the apical endocytic compartment of the tubular cells and colocalized with clathrin 10 min after injection. After 20 min, lysozyme was located in the subapical compartment negative to clathrin (endosomes, and intracellular trafficking of lysozyme was coincided with the distribution of megalin and cubilin. However, internalized protein was retained in the endosomes and did not reach lysosomes within 30 min after treatment that may indicate the inhibition of intracellular trafficking in hibernating frogs. For the first time, we provided the evidence that lysozyme is filtered through the glomeruli and absorbed by receptor-mediated clathrin-dependent endocytosis in the frog proximal tubule cells. Thus, the protein uptake in the amphibian mesonephros is mediated by megalin and cubilin that confirms a critical role of endocytic receptors in the renal reabsorption of proteins in amphibians as in mammals.

  12. Killing of intracellular Mycobacterium tuberculosis by receptor-mediated drug delivery

    International Nuclear Information System (INIS)

    Majumdar, S.; Basu, S.K.

    1991-01-01

    p-Aminosalicylic acid (PAS) conjugated to maleylated bovine serum albumin (MBSA) was taken up efficiently through high-affinity MBSA-binding sites on macrophages. Binding of the radiolabeled conjugate to cultured mouse peritoneal macrophages at 4 degrees C was competed for by MBSA but not by PAS. At 37 degrees C, the radiolabeled conjugate was rapidly degraded by the macrophages, leading to release of acid-soluble degradation products in the medium. The drug conjugate was nearly 100 times as effective as free PAS in killing the intracellular mycobacteria in mouse peritoneal macrophages infected in culture with Mycobacterium tuberculosis. The killing of intracellular mycobacteria mediated by the drug conjugate was effectively prevented by simultaneous addition of excess MBSA (100 micrograms/ml) or chloroquine (3 microM) to the medium, whereas these agents did not affect the microbicidal action of free PAS. These results suggest that (i) uptake of the PAS-MBSA conjugate was mediated by cell surface receptors on macrophages which recognize MBSA and (ii) lysosomal hydrolysis of the internalized conjugate resulted in intracellular release of a pharmacologically active form of the drug, which led to selective killing of the M. tuberculosis harbored by mouse macrophages infected in culture. This receptor-mediated modality of delivering drugs to macrophages could contribute to greater therapeutic efficacy and minimization of toxic side effects in the management of tuberculosis and other intracellular mycobacterial infections

  13. GABA-A Receptors Mediate Tonic Inhibition and Neurosteroid Sensitivity in the Brain.

    Science.gov (United States)

    Reddy, Doodipala Samba

    2018-01-01

    Neurosteroids like allopregnanolone (AP) are positive allosteric modulators of synaptic and extrasynaptic GABA-A receptors. AP and related neurosteroids exhibit a greater potency for δ-containing extrasynaptic receptors. The δGABA-A receptors, which are expressed extrasynaptically in the dentate gyrus and other regions, contribute to tonic inhibition, promoting network shunting as well as reducing seizure susceptibility. Levels of endogenous neurosteroids fluctuate with ovarian cycle. Natural and synthetic neurosteroids maximally potentiate tonic inhibition in the hippocampus and provide robust protection against a variety of limbic seizures and status epilepticus. Recently, a consensus neurosteroid pharmacophore model has been proposed at extrasynaptic δGABA-A receptors based on structure-activity relationship for functional activation of tonic currents and seizure protection. Aside from anticonvulsant actions, neurosteroids have been found to be powerful anxiolytic and anesthetic agents. Neurosteroids and Zn 2+ have preferential affinity for δ-containing receptors. Thus, Zn 2+ can prevent neurosteroid activation of extrasynaptic δGABA-A receptor-mediated tonic inhibition. Recently, we demonstrated that Zn 2+ selectively inhibits extrasynaptic δGABA-A receptors and thereby fully prevents AP activation of tonic inhibition and seizure protection. We confirmed that neurosteroids exhibit greater sensitivity at extrasynaptic δGABA-A receptors. Overall, extrasynaptic GABA-A receptors are primary mediators of tonic inhibition in the brain and play a key role in the pathophysiology of epilepsy and other neurological disorders. © 2018 Elsevier Inc. All rights reserved.

  14. Beta receptor-mediated modulation of the late positive potential in humans.

    Science.gov (United States)

    de Rover, Mischa; Brown, Stephen B R E; Boot, Nathalie; Hajcak, Greg; van Noorden, Martijn S; van der Wee, Nic J A; Nieuwenhuis, Sander

    2012-02-01

    Electrophysiological studies have identified a scalp potential, the late positive potential (LPP), which is modulated by the emotional intensity of observed stimuli. Previous work has shown that the LPP reflects the modulation of activity in extrastriate visual cortical structures, but little is known about the source of that modulation. The present study investigated whether beta-adrenergic receptors are involved in the generation of the LPP. We used a genetic individual differences approach (experiment 1) and a pharmacological manipulation (experiment 2) to test the hypothesis that the LPP is modulated by the activation of β-adrenergic receptors. In experiment 1, we found that LPP amplitude depends on allelic variation in the β1-receptor gene polymorphism. In experiment 2, we found that LPP amplitude was modulated by the β-blocker propranolol in a direction dependent on subjects' level of trait anxiety: In participants with lower trait anxiety, propranolol led to a (nonsignificant) decrease in the LPP modulation; in participants with higher trait anxiety, propranolol increased the emotion-related LPP modulation. These results provide initial support for the hypothesis that the LPP reflects the downstream effects, in visual cortical areas, of β-receptor-mediated activation of the amygdala.

  15. The effect of vanadate on receptor-mediated endocytosis of asialoorosomucoid in rat liver parenchymal cells

    International Nuclear Information System (INIS)

    Kindberg, G.M.; Gudmundsen, O.; Berg, T.

    1990-01-01

    Vanadate is a phosphate analogue that inhibits enzymes involved in phosphate release and transfer reactions. Since such reactions may play important roles in endocytosis, we studied the effects of vanadate on various steps in receptor-mediated endocytosis of asialoorosomucoid labeled with 125I-tyramine-cellobiose (125I-TC-AOM). The labeled degradation products formed from 125I-TC-AOM are trapped in the lysosomes and may therefore serve as lysosomal markers in subcellular fractionation studies. Vanadate reduced the amount of active surface asialoglycoprotein receptors approximately 70%, but had no effect on the rate of internalization and retroendocytosis of ligand. The amount of surface asialoglycoprotein receptors can be reduced by lowering the incubation temperature gradually from 37 to 15 degrees C; vanadate affected only the temperature--sensitive receptors. Vanadate inhibited degradation of 125I-TC-AOM 70-80%. Degradation was much more sensitive to vanadate than binding; half-maximal effects were seen at approximately 1 mM vanadate for binding and approximately 0.1 mM vanadate for degradation. By subcellular fractionation in sucrose and Nycodenz gradients, it was shown that vanadate completely prevented the transfer of 125I-TC-AOM from endosomes to lysosomes. Therefore, the inhibition of degradation by vanadate was indirect; in the presence of vanadate, ligand did not gain access to the lysosomes. The limited degradation in the presence of vanadate took place in a prelysosomal compartment. Vanadate did not affect cell viability and ATP content

  16. A pp32-retinoblastoma protein complex modulates androgen receptor-mediated transcription and associates with components of the splicing machinery

    International Nuclear Information System (INIS)

    Adegbola, Onikepe; Pasternack, Gary R.

    2005-01-01

    We have previously shown pp32 and the retinoblastoma protein interact. pp32 and the retinoblastoma protein are nuclear receptor transcriptional coregulators: the retinoblastoma protein is a coactivator for androgen receptor, the major regulator of prostate cancer growth, while pp32, which is highly expressed in prostate cancer, is a corepressor of the estrogen receptor. We now show pp32 increases androgen receptor-mediated transcription and the retinoblastoma protein modulates this activity. Using affinity purification and mass spectrometry, we identify members of the pp32-retinoblastoma protein complex as PSF and nonO/p54nrb, proteins implicated in coordinate regulation of nuclear receptor-mediated transcription and splicing. We show that the pp32-retinoblastoma protein complex is modulated during TPA-induced K562 differentiation. Present evidence suggests that nuclear receptors assemble multiprotein complexes to coordinately regulate transcription and mRNA processing. Our results suggest that pp32 and the retinoblastoma protein may be part of a multiprotein complex that coordinately regulates nuclear receptor-mediated transcription and mRNA processing

  17. Influence of ER leak on resting cytoplasmic Ca2+ and receptor-mediated Ca2+ signalling in human macrophage.

    Science.gov (United States)

    Layhadi, Janice A; Fountain, Samuel J

    2017-06-03

    Mechanisms controlling endoplasmic reticulum (ER) Ca 2+ homeostasis are important regulators of resting cytoplasmic Ca 2+ concentration ([Ca 2+ ] cyto ) and receptor-mediated Ca 2+ signalling. Here we investigate channels responsible for ER Ca 2+ leak in THP-1 macrophage and human primary macrophage. In the absence of extracellular Ca 2+ we employ ionomycin action at the plasma membrane to stimulate ER Ca 2+ leak. Under these conditions ionomycin elevates [Ca 2+ ] cyto revealing a Ca 2+ leak response which is abolished by thapsigargin. IP 3 receptors (Xestospongin C, 2-APB), ryanodine receptors (dantrolene), and translocon (anisomycin) inhibition facilitated ER Ca 2+ leak in model macrophage, with translocon inhibition also reducing resting [Ca 2+ ] cyto . In primary macrophage, translocon inhibition blocks Ca 2+ leak but does not influence resting [Ca 2+ ] cyto . We identify a role for translocon-mediated ER Ca 2+ leak in receptor-mediated Ca 2+ signalling in both model and primary human macrophage, whereby the Ca 2+ response to ADP (P2Y receptor agonist) is augmented following anisomycin treatment. In conclusion, we demonstrate a role of ER Ca 2+ leak via the translocon in controlling resting cytoplasmic Ca 2+ in model macrophage and receptor-mediated Ca 2+ signalling in model macrophage and primary macrophage. Copyright © 2017 Elsevier Inc. All rights reserved.

  18. Receptor-mediated mechanism for the transport of prolactin from blood to cerebrospinal fluid

    International Nuclear Information System (INIS)

    Walsh, R.J.; Slaby, F.J.; Posner, B.I.

    1987-01-01

    Prolactin (PRL) interacts with areas of the central nervous system which reside behind the blood-brain barrier. While vascular PRL does not cross this barrier, it is readily accessible to the cerebrospinal fluid (CSF) from which it may gain access to the PRL-responsive areas of the brain. Studies were undertaken to characterize the mechanism responsible for the translocation of PRL from blood to CSF. Rats were given external jugular vein injections of [ 125 -I]iodo-PRL in the presence or absence of an excess of unlabeled ovine PRL (oPRL), human GH, bovine GH, or porcine insulin. CSF and choroid plexus were removed 60 min later. CSF samples were electrophoresed on sodium dodecyl sulfate-polyacrylamide slab gels and resultant autoradiographs were analyzed with quantitative microdensitometry. The data revealed that unlabeled lactogenic hormones, viz. oPRL and human GH, caused a statistically significant inhibition of [ 125 I]iodo-PRL transport from blood to CSF. In contrast, nonlactogenic hormones, viz bovine GH and insulin, had no effect on [ 125 I]iodo-PRL transport into the CSF. An identical pattern of competition was observed in the binding of hormone to the choroid plexus. Furthermore, vascular injections of [ 125 I]iodo-PRL administered with a range of concentrations of unlabeled oPRL revealed a dose-response inhibition in the transport of [ 125 I]iodo-PRL from blood to CSF. The study demonstrates that PRL enters the CSF by a specific, PRL receptor-mediated transport mechanism. The data is consistent with the hypothesis that the transport mechanism resides at the choroid plexus. The existence of this transport mechanism reflects the importance of the cerebroventricular system in PRL-brain interactions

  19. Decreased α1-adrenergic receptor-mediated inositide hydrolysis in neurons from hypertensive rat brain

    International Nuclear Information System (INIS)

    Feldstein, J.B.; Gonzales, R.A.; Baker, S.P.; Sumners, C.; Crews, F.T.; Raizada, M.K.

    1986-01-01

    The expression of α 1 -adrenergic receptors and norepinephrine (NE)-stimulated hydrolysis of inositol phospholipid has been studied in neuronal cultures from the brains of normotensive (Wistar-Kyoto, WKY) and spontaneously hypertensive (SH) rats. Binding of 125 I-1-[β-(4-hydroxyphenyl)-ethyl-aminomethyl] tetralone (HEAT) to neuronal membranes was 68-85% specific and was rapid. Competition-inhibition experiments with various agonists and antagonists suggested that 125 I-HEAT bound selectively to α 1 -adrenergic receptors. Specific binding of 125 I-HEAT to neuronal membranes from SH rat brain cultures was 30-45% higher compared with binding in WKY normotensive controls. This increase was attributed to an increase in the number of α 1 -adrenergic receptors on SH rat brain neurons. Incubation of neuronal cultures of rat brain from both strains with NE resulted in a concentration-dependent stimulation of release of inositol phosphates, although neurons from SH rat brains were 40% less responsive compared with WKY controls. The decrease in responsiveness of SH rat brain neurons to NE, even though the α 1 -adrenergic receptors are increased, does not appear to be due to a general defect in membrane receptors and postreceptor signal transduction mechanisms. This is because neither the number of muscarinic-cholinergic receptors nor the carbachol-stimulated release of inositol phosphates is different in neuronal cultures from the brains of SH rats compared with neuronal cultures from the brains of WKY rats. These observations suggest that the increased expression of α 1 -adrenergic receptors does not parallel the receptor-mediated inositol phosphate hydrolysis in neuronal cultures from SH rat brain

  20. Milrinone attenuates thromboxane receptor-mediated hyperresponsiveness in hypoxic pulmonary arterial myocytes.

    Science.gov (United States)

    Santhosh, K T; Elkhateeb, O; Nolette, N; Outbih, O; Halayko, A J; Dakshinamurti, S

    2011-07-01

    Neonatal pulmonary hypertension (PPHN) is characterized by pulmonary vasoconstriction, due in part to dysregulation of the thromboxane prostanoid (TP) receptor. Hypoxia induces TP receptor-mediated hyperresponsiveness, whereas serine phosphorylation mediates desensitization of TP receptors. We hypothesized that prostacyclin (IP) receptor activity induces TP receptor phosphorylation and decreases ligand affinity; that TP receptor sensitization in hypoxic myocytes is due to IP receptor inactivation; and that this would be reversible by the cAMP-specific phosphodiesterase inhibitor milrinone. We examined functional regulation of TP receptors by serine phosphorylation and effects of IP receptor stimulation and protein kinase A (PKA) activity on TP receptor sensitivity in myocytes from neonatal porcine resistance pulmonary arteries after 72 h hypoxia in vitro. Ca(2+) response curves to U46619 (TP receptor agonist) were determined in hypoxic and normoxic myocytes incubated with or without iloprost (IP receptor agonist), forskolin (adenylyl cyclase activator), H8 (PKA inhibitor) or milrinone. TP and IP receptor saturation binding kinetics were measured in presence of iloprost or 8-bromo-cAMP. Ligand affinity for TP receptors was normalized in vitro by IP receptor signalling intermediates. However, IP receptor affinity was compromised in hypoxic myocytes, decreasing cAMP production. Milrinone normalized TP receptor sensitivity in hypoxic myocytes by restoring PKA-mediated regulatory TP receptor phosphorylation. TP receptor sensitivity and EC(50) for TP receptor agonists was regulated by PKA, as TP receptor serine phosphorylation by PKA down-regulated Ca(2+) mobilization. Hypoxia decreased IP receptor activity and cAMP generation, inducing TP receptor hyperresponsiveness, which was reversed by milrinone. © 2011 The Authors. British Journal of Pharmacology © 2011 The British Pharmacological Society.

  1. PKCɛ mediates substance P inhibition of GABAA receptors-mediated current in rat dorsal root ganglion.

    Science.gov (United States)

    Li, Li; Zhao, Lei; Wang, Yang; Ma, Ke-tao; Shi, Wen-yan; Wang, Ying-zi; Si, Jun-qiang

    2015-02-01

    The mechanism underlying the modulatory effect of substance P (SP) on GABA-activated response in rat dorsal root ganglion (DRG) neurons was investigated. In freshly dissociated rat DRG neurons, whole-cell patch-clamp technique was used to record GABA-activated current and sharp electrode intracellular recording technique was used to record GABA-induced membrane depolarization. Application of GABA (1-1000 μmol/L) induced an inward current in a concentration-dependent manner in 114 out of 127 DRG neurons (89.8 %) examined with whole-cell patch-clamp recordings. Bath application of GABA (1-1000 μmol/L) evoked a depolarizing response in 236 out of 257 (91.8%) DRG neurons examined with intracellular recordings. Application of SP (0.001-1 μmol/L) suppressed the GABA-activated inward current and membrane depolarization. The inhibitory effects were concentration-dependent and could be blocked by the selective neurokinin 1 (NK1) receptors antagonist spantide but not by L659187 and SR142801 (1 μmol/L, n=7), selective antagonists of NK2 and NK3. The inhibitory effect of SP was significantly reduced by the calcium chelator BAPTA-AM, phospholipase C (PLC) inhibitor U73122, and PKC inhibitor chelerythrine, respectively. The PKA inhibitor H-89 did not affect the SP effect. Remarkably, the inhibitory effect of SP on GABA-activated current was nearly completely removed by a selective PKCε inhibitor epilon-V1-2 but not by safingol and LY333531, selective inhibitors of PKCα and PKCβ. Our results suggest that NK1 receptor mediates SP-induced inhibition of GABA-activated current and membrane depolarization by activating intracellular PLC-Ca²⁺-PKCε cascade. SP might regulate the excitability of peripheral nociceptors through inhibition of the "pre-synaptic inhibition" evoked by GABA, which may explain its role in pain and neurogenic inflammation.

  2. Receptor-mediated endocytosis generates nanomechanical force reflective of ligand identity and cellular property.

    Science.gov (United States)

    Zhang, Xiao; Ren, Juan; Wang, Jingren; Li, Shixie; Zou, Qingze; Gao, Nan

    2018-08-01

    Whether environmental (thermal, chemical, and nutrient) signals generate quantifiable, nanoscale, mechanophysical changes in the cellular plasma membrane has not been well elucidated. Assessment of such mechanophysical properties of plasma membrane may shed lights on fundamental cellular process. Atomic force microscopic (AFM) measurement of the mechanical properties of live cells was hampered by the difficulty in accounting for the effects of the cantilever motion and the associated hydrodynamic force on the mechanical measurement. These challenges have been addressed in our recently developed control-based AFM nanomechanical measurement protocol, which enables a fast, noninvasive, broadband measurement of the real-time changes in plasma membrane elasticity in live cells. Here we show using this newly developed AFM platform that the plasma membrane of live mammalian cells exhibits a constant and quantifiable nanomechanical property, the membrane elasticity. This mechanical property sensitively changes in response to environmental factors, such as the thermal, chemical, and growth factor stimuli. We demonstrate that different chemical inhibitors of endocytosis elicit distinct changes in plasma membrane elastic modulus reflecting their specific molecular actions on the lipid configuration or the endocytic machinery. Interestingly, two different growth factors, EGF and Wnt3a, elicited distinct elastic force profiles revealed by AFM at the plasma membrane during receptor-mediated endocytosis. By applying this platform to genetically modified cells, we uncovered a previously unknown contribution of Cdc42, a key component of the cellular trafficking network, to EGF-stimulated endocytosis at plasma membrane. Together, this nanomechanical AFM study establishes an important foundation that is expandable and adaptable for investigation of cellular membrane evolution in response to various key extracellular signals. © 2017 Wiley Periodicals, Inc.

  3. Decrement of GABAA receptor-mediated inhibitory postsynaptic currents in dentate granule cells in epileptic hippocampus.

    Science.gov (United States)

    Isokawa, M

    1996-05-01

    1. Inhibitory postsynaptic currents (IPSCs) were studied in hippocampal dentate granule cells (DGCs) in the pilocarpine model and human temporal lobe epilepsy, with the use of the whole cell patch-clamp recording technique in slice preparations. 2. In the pilocarpine model, hippocampal slices were prepared from rats that were allowed to experience spontaneous seizures for 2 mo. Human hippocampal specimens were obtained from epileptic patients who underwent surgical treatment for medically intractable seizures. 3. IPSCs were generated by single perforant path stimulation and recorded at a membrane potential (Vm) of 0 mV near the reversal potential of glutamate excitatory postsynaptic currents in the voltage-clamp recording. IPSCs were pharmacologically identified as gamma-aminobutyric acid-A (GABAA) IPSCs by 10 microM bicuculline methiodide. 4. During low-frequency stimulation, IPSCs were not different in amplitude among non-seizure-experienced rat hippocampi, human nonsclerotic hippocampi, seizure-experienced rat hippocampi, and human sclerotic hippocampi. In the last two groups of DGCs, current-clamp recordings indicated the presence of prolonged excitatory postsynaptic potentials (EPSPs) mediated by the N-methyl-D-aspartate (NMDA) receptor. 5. High-frequency stimulation, administered at Vm = -30 mV to activate NMDA currents, reduced GABAA IPSC amplitude specifically in seizure-experienced rat hippocampi (t = 2.5, P < 0.03) and human sclerotic hippocampi (t = 7.7, P < 0.01). This reduction was blocked by an NMDA receptor antagonist, 2-amino-5-phosphonovaleric acid (APV) (50 microM). The time for GABAA IPSCs to recover to their original amplitude was also shortened by the application of APV. 6. I conclude that, when intensively activated, NMDA receptor-mediated excitatory transmission may interact with GABAergic synaptic inhibition in DGCs in seizure-experienced hippocampus to transiently reduce GABA(A) receptor-channel function. Such interactions may contribute to

  4. Receptor-mediated transcytosis of cyclophilin B through the blood-brain barrier.

    Science.gov (United States)

    Carpentier, M; Descamps, L; Allain, F; Denys, A; Durieux, S; Fenart, L; Kieda, C; Cecchelli, R; Spik, G

    1999-07-01

    Cyclophilin B (CyPB) is a cyclosporin A (CsA)-binding protein mainly located in intracellular vesicles and secreted in biological fluids. In previous works, we demonstrated that CyPB interacts with T lymphocytes and enhances in vitro cellular incorporation and activity of CsA. In addition to its immunosuppressive activity, CsA is able to promote regeneration of damaged peripheral nerves. However, the crossing of the drug from plasma to neural tissue is restricted by the relative impermeability of the blood-brain barrier. To know whether CyPB might also participate in the delivery of CsA into the brain, we have analyzed the interactions of CyPB with brain capillary endothelial cells. First, we demonstrated that CyPB binds to two types of binding sites present at the surface of capillary endothelial cells from various species of tissues. The first type of binding sites (K(D) = 300 nM; number of sites = 3 x 10(6)) is related to interactions with negatively charged compounds such as proteoglycans. The second type of binding sites, approximately 50,000 per cell, exhibits a higher affinity for CyPB (K(D) = 15 nM) and is involved in an endocytosis process, indicating it might correspond to a functional receptor. Finally, the use of an in vitro model of blood-brain barrier allowed us to demonstrate that CyPB is transcytosed by a receptor-mediated pathway (flux = 16.5 fmol/cm2/h). In these conditions, CyPB did not significantly modify the passage of CsA, indicating that it is unlikely to provide a pathway for CsA brain delivery.

  5. NMDA receptors mediate neuron-to-glia signaling in mouse cortical astrocytes.

    Science.gov (United States)

    Lalo, Ulyana; Pankratov, Yuri; Kirchhoff, Frank; North, R Alan; Verkhratsky, Alexei

    2006-03-08

    Chemical transmission between neurons and glial cells is an important element of integration in the CNS. Here, we describe currents activated by NMDA in cortical astrocytes, identified in transgenic mice that express enhanced green fluorescent protein under control of the human glial fibrillary acidic protein promoter. Astrocytes were studied by whole-cell voltage clamp either in slices or after gentle nonenzymatic mechanical dissociation. Acutely isolated astrocytes showed a three-component response to glutamate. The initial rapid component was blocked by 1,2,3,4-tetrahydro-6-nitro-2,3-dioxo-benzo[f]quinoxaline-7-sulfonamide (NBQX), which is an antagonist of AMPA receptors (IC50, 2 microM), and the NMDA receptor antagonist D-AP-5 blocked the later sustained component (IC50, 0.6 microM). The third component of glutamate application response was sensitive to D,L-threo-beta-benzyloxyaspartate, a glutamate transporter blocker. Fast application of NMDA evoked concentration-dependent inward currents (EC50, 0.3 microM); these showed use-dependent block by (+)-5-methyl-10,11-dihydro-5H-dibenzo [a,d] cyclohepten-5,10-imine maleate (MK-801). These NMDA-evoked currents were linearly dependent on membrane potential and were not affected by extracellular magnesium at concentrations up to 10 mM. Electrical stimulation of axons in layer IV-VI induced a complex inward current in astrocytes situated in the cortical layer II, part of which was sensitive to MK-801 at holding potential -80 mV and was not affected by the AMPA glutamate receptor antagonist NBQX. The fast miniature spontaneous currents were observed in cortical astrocytes in slices as well. These currents exhibited both AMPA and NMDA receptor-mediated components. We conclude that cortical astrocytes express functional NMDA receptors that are devoid of Mg2+ block, and these receptors are involved in neuronal-glial signal transmission.

  6. Scavenger receptors mediate the role of SUMO and Ftz-f1 in Drosophila steroidogenesis.

    Directory of Open Access Journals (Sweden)

    Ana Talamillo

    2013-04-01

    Full Text Available SUMOylation participates in ecdysteroid biosynthesis at the onset of metamorphosis in Drosophila melanogaster. Silencing the Drosophila SUMO homologue smt3 in the prothoracic gland leads to reduced lipid content, low ecdysone titers, and a block in the larval-pupal transition. Here we show that the SR-BI family of Scavenger Receptors mediates SUMO functions. Reduced levels of Snmp1 compromise lipid uptake in the prothoracic gland. In addition, overexpression of Snmp1 is able to recover lipid droplet levels in the smt3 knockdown prothoracic gland cells. Snmp1 expression depends on Ftz-f1 (an NR5A-type orphan nuclear receptor, the expression of which, in turn, depends on SUMO. Furthermore, we show by in vitro and in vivo experiments that Ftz-f1 is SUMOylated. RNAi-mediated knockdown of ftz-f1 phenocopies that of smt3 at the larval to pupal transition, thus Ftz-f1 is an interesting candidate to mediate some of the functions of SUMO at the onset of metamorphosis. Additionally, we demonstrate that the role of SUMOylation, Ftz-f1, and the Scavenger Receptors in lipid capture and mobilization is conserved in other steroidogenic tissues such as the follicle cells of the ovary. smt3 knockdown, as well as ftz-f1 or Scavenger knockdown, depleted the lipid content of the follicle cells, which could be rescued by Snmp1 overexpression. Therefore, our data provide new insights into the regulation of metamorphosis via lipid homeostasis, showing that Drosophila Smt3, Ftz-f1, and SR-BIs are part of a general mechanism for uptake of lipids such as cholesterol, required during development in steroidogenic tissues.

  7. P2X receptor-mediated ATP purinergic signaling in health and disease

    Directory of Open Access Journals (Sweden)

    Jiang LH

    2012-09-01

    Full Text Available Lin-Hua JiangSchool of Biomedical Sciences, Faculty of Biological Sciences, University of Leeds, Leeds, United KingdomAbstract: Purinergic P2X receptors are plasma membrane proteins present in a wide range of mammalian cells where they act as a cellular sensor, enabling cells to detect and respond to extracellular adenosine triphosphate (ATP, an important signaling molecule. P2X receptors function as ligand-gated Ca2+-permeable cationic channels that open upon ATP binding to elevate intracellular Ca2+ concentrations and cause membrane depolarization. In response to sustained activation, P2X receptors induce formation of a pore permeable to large molecules. P2X receptors also interact with distinct functional proteins and membrane lipids to form specialized signaling complexes. Studies have provided compelling evidence to show that such P2X receptor-mediated ATP-signaling mechanisms determine and regulate a growing number and diversity of important physiological processes, including neurotransmission, muscle contraction, and cytokine release. There is accumulating evidence to support strong causative relationships of altered receptor expression and function with chronic pain, inflammatory diseases, cancers, and other pathologies or diseases. Numerous high throughput screening drug discovery programs and preclinical studies have thus far demonstrated the proof of concepts that the P2X receptors are druggable targets and selective receptor antagonism is a promising therapeutics approach. This review will discuss the recent progress in understanding the mammalian P2X receptors with respect to the ATP-signaling mechanisms, physiological and pathophysiological roles, and development and preclinical studies of receptor antagonists.Keywords: extracellular ATP, ion channel, large pore, signaling complex, chronic pain, inflammatory diseases

  8. Receptor-mediated endocytosis of low density lipoproteins in aortic endothelial cells

    International Nuclear Information System (INIS)

    Sanan, D.A.

    1986-04-01

    Lipoprotein binding and metabolism in actively-dividing (subconfluent) and quiescent (postconfluent) bovine aortic endothelial cells (ECs) were qualitatively investigated by fluorescence microscopy using dioctadecylindocarbocyanine-labelled lipoproteins and by indirect immunofluorescence microscopy. LDL and acetylated-LDL (AcLDL) were seen bound to the surfaces of subconfluent ECs (at 4 degrees C or at 37 degrees C), as a random distribution of punctate foci. ECs therefore closely resembled fibroblasts in the distribution of LDL receptors on their surfaces. No binding of LDL was seen on postconfluent EC surfaces by either direct or indirect fluorescence microscopy. The patterns of AcLDL binding on postconfluent ECs resembled those on subconfluent ECs. Intracellular LDL and AcLDL occurred as perinuclear accumulations of large fluorescent disc-shaped profiles in subconfluent ECs. These accumulations were shown to arise from surface-bound material by pulse-chase experiments. Intracellular LDL was absent in the majority of postconfluent ECs, while AcLDL accumulation was massive. 'Wounding' of cultures allowed simultaneous assessment of lipoprotein metabolism in quiescent and actively-dividing areas of the same culture. It is concluded that postconfluent quiescent bovine aortic ECs in vitro metabolise virtually no LDL via the LDL-receptor pathway due to a vanishingly low number of LDL receptors. This contrasts with the ability of postconfluent cells to metabolise relatively large amounts of AcLDL via a receptor-mediated mechanism. The significance of these conclusions is discussed with respect to the interaction of plasma lipoproteins with the endothelium in vivo. 301 refs

  9. Proliferation of mouse endometrial stromal cells in culture is highly sensitive to lysophosphatidic acid signaling

    International Nuclear Information System (INIS)

    Aikawa, Shizu; Kano, Kuniyuki; Inoue, Asuka; Aoki, Junken

    2017-01-01

    Endometrial stromal cells (ESCs) proliferate rapidly both in vivo and in vitro. Here we show that proliferation of ESCs in vitro is strongly dependent on lysophosphatidic acid (LPA) signaling. LPA is produced by autotaxin (ATX) and induces various kinds of cellular processes including migration, proliferation and inhibition of cell death possibly through six G protein-coupled receptors (LPA 1-6 ). We found that ESCs proliferated rapidly in vitro in an autocrine manner and that the proliferation was prominently suppressed by either an ATX inhibitor (ONO-8430506) or an LPA 1/3 antagonist (Ki16425). Among the cells lines tested, mouse ESCs were the most sensitive to these inhibitors. Proliferation of ESCs isolated from either LPA 1 - or LPA 3 -deficient mice was comparable to proliferation of ESCs isolated from control mice. An LPA receptor antagonist (AM095), which was revealed to be a dual LPA 1 /LPA 3 antagonist, also suppressed the proliferation of ESCs. The present results show that LPA signaling has a critical role in the proliferation of ESCs, and that this role is possibly mediated redundantly by LPA 1 and LPA 3 . - Highlights: • Uterine endometrial stromal cells (ESCs) proliferate rapidly both in vivo and in vitro. • ESCs proliferated in vitro in an autocrine fashion. • Proliferation of mouse ESCs was prominently suppressed by inhibitors of lysophosphatidic acid (LPA) signaling. • LPA receptors, LPA 1 and LPA 3 , had redundant role in supporting the proliferation of ESCs.

  10. Phosphorylation and Internalization of Lysophosphatidic Acid Receptors LPA1, LPA2, and LPA3.

    Directory of Open Access Journals (Sweden)

    Rocío Alcántara-Hernández

    Full Text Available The lysophosphatidic acid receptors LPA1, LPA2, and LPA3 were individually expressed in C9 cells and their signaling and regulation were studied. Agonist-activation increases intracellular calcium concentration in a concentration-dependent fashion. Phorbol myristate acetate markedly inhibited LPA1- and LPA3-mediated effect, whereas that mediated by LPA2 was only partially diminished; the actions of the phorbol ester were inhibited by bisindolylmaleimide I and by overnight incubation with the protein kinase C activator, which leads to down regulation of this protein kinase. Homologous desensitization was also observed for the three LPA receptors studied, with that of LPA2 receptors being consistently of lesser magnitude; neither inhibition nor down-regulation of protein kinase C exerted any effect on homologous desensitization. Activation of LPA1-3 receptors induced ERK 1/2 phosphorylation; this effect was markedly attenuated by inhibition of epidermal growth factor receptor tyrosine kinase activity, suggesting growth factor receptor transactivation in this effect. Lysophosphatidic acid and phorbol myristate acetate were able to induce LPA1-3 phosphorylation, in time- and concentration-dependent fashions. It was also clearly observed that agonists and protein kinase C activation induced internalization of these receptors. Phosphorylation of the LPA2 subtype required larger concentrations of these agents and its internalization was less intense than that of the other subtypes.Our data show that these three LPA receptors are phosphoproteins whose phosphorylation state is modulated by agonist-stimulation and protein kinase C-activation and that differences in regulation and cellular localization exist, among the subtypes.

  11. Histamine H2 Receptor-Mediated Suppression of Intestinal Inflammation by Probiotic Lactobacillus reuteri.

    Science.gov (United States)

    Gao, Chunxu; Major, Angela; Rendon, David; Lugo, Monica; Jackson, Vanessa; Shi, Zhongcheng; Mori-Akiyama, Yuko; Versalovic, James

    2015-12-15

    -mediated suppression of colonic inflammation. The effective combination of diet, gut bacteria, and host receptor-mediated signaling may result in opportunities for therapeutic microbiology and provide clues for discovery and development of next-generation probiotics. Copyright © 2015 Gao et al.

  12. Astrocytes protect neurons against methylmercury via ATP/P2Y(1) receptor-mediated pathways in astrocytes.

    Science.gov (United States)

    Noguchi, Yusuke; Shinozaki, Youichi; Fujishita, Kayoko; Shibata, Keisuke; Imura, Yoshio; Morizawa, Yosuke; Gachet, Christian; Koizumi, Schuichi

    2013-01-01

    Methylmercury (MeHg) is a well known environmental pollutant that induces serious neuronal damage. Although MeHg readily crosses the blood-brain barrier, and should affect both neurons and glial cells, how it affects glia or neuron-to-glia interactions has received only limited attention. Here, we report that MeHg triggers ATP/P2Y1 receptor signals in astrocytes, thereby protecting neurons against MeHg via interleukin-6 (IL-6)-mediated pathways. MeHg increased several mRNAs in astrocytes, among which IL-6 was the highest. For this, ATP/P2Y1 receptor-mediated mechanisms were required because the IL-6 production was (i) inhibited by a P2Y1 receptor antagonist, MRS2179, (ii) abolished in astrocytes obtained from P2Y1 receptor-knockout mice, and (iii) mimicked by exogenously applied ATP. In addition, (iv) MeHg released ATP by exocytosis from astrocytes. As for the intracellular mechanisms responsible for IL-6 production, p38 MAP kinase was involved. MeHg-treated astrocyte-conditioned medium (ACM) showed neuro-protective effects against MeHg, which was blocked by anti-IL-6 antibody and was mimicked by the application of recombinant IL-6. As for the mechanism of neuro-protection by IL-6, an adenosine A1 receptor-mediated pathway in neurons seems to be involved. Taken together, when astrocytes sense MeHg, they release ATP that autostimulates P2Y1 receptors to upregulate IL-6, thereby leading to A1 receptor-mediated neuro-protection against MeHg.

  13. Blockade of lysophosphatidic acid receptors LPAR1/3 ameliorates lung fibrosis induced by irradiation

    International Nuclear Information System (INIS)

    Gan, Lu; Xue, Jian-Xin; Li, Xin; Liu, De-Song; Ge, Yan; Ni, Pei-Yan; Deng, Lin; Lu, You; Jiang, Wei

    2011-01-01

    Highlights: → Lysophosphatidic acid (LPA) levels and its receptors LPAR1/3 transcripts were elevated during the development of radiation-induced lung fibrosis. → Lung fibrosis was obviously alleviated in mice treated with the dual LPAR1/3 antagonist, VPC12249. → VPC12249 administration effectively inhibited radiation-induced fibroblast accumulation in vivo, and suppressed LPA-induced fibroblast proliferation in vitro. → LPA-LPAR1/3 signaling regulated TGFβ1 and CTGF expressions in radiation-challenged lungs, but only influenced CTGF expression in cultured fibroblasts. → LPA-LPAR1/3 signaling induced fibroblast proliferation through a CTGF-dependent pathway, rather than through TGFβ1 activation. -- Abstract: Lung fibrosis is a common and serious complication of radiation therapy for lung cancer, for which there are no efficient treatments. Emerging evidence indicates that lysophosphatidic acid (LPA) and its receptors (LPARs) are involved in the pathogenesis of fibrosis. Here, we reported that thoracic radiation with 16 Gy in mice induced development of radiation lung fibrosis (RLF) accompanied by obvious increases in LPA release and LPAR1 and LPAR3 (LPAR1/3) transcripts. RLF was significantly alleviated in mice treated with the dual LPAR1/3 antagonist, VPC12249. VPC12249 administration effectively prolonged animal survival, restored lung structure, inhibited fibroblast accumulation and reduced collagen deposition. Moreover, profibrotic cytokines in radiation-challenged lungs obviously decreased following administration of VPC12249, including transforming growth factor β1 (TGFβ1) and connective tissue growth factor (CTGF). In vitro, LPA induced both fibroblast proliferation and CTGF expression in a dose-dependent manner, and both were suppressed by blockade of LPAR1/3. The pro-proliferative activity of LPA on fibroblasts was inhibited by siRNA directed against CTGF. Together, our data suggest that the LPA-LPAR1/3 signaling system is involved in the

  14. Blockade of lysophosphatidic acid receptors LPAR1/3 ameliorates lung fibrosis induced by irradiation

    Energy Technology Data Exchange (ETDEWEB)

    Gan, Lu [State Key Laboratory of Biotherapy, West China Hospital, Sichuan University, Chengdu (China); Xue, Jian-Xin [Department of Thoracic Oncology, Cancer Center, West China Hospital, Sichuan University, Chengdu (China); Laboratory of Stem Cell Biology, West China Hospital, Sichuan University, Chengdu (China); Li, Xin [Department of Thoracic Oncology, Cancer Center, West China Hospital, Sichuan University, Chengdu (China); Liu, De-Song [Department of Pediatrics, Sichuan Provincial Hospital of Women and Children, Chengdu (China); Ge, Yan; Ni, Pei-Yan; Deng, Lin [State Key Laboratory of Biotherapy, West China Hospital, Sichuan University, Chengdu (China); Lu, You, E-mail: radyoulu@hotmail.com [State Key Laboratory of Biotherapy, West China Hospital, Sichuan University, Chengdu (China); Department of Thoracic Oncology, Cancer Center, West China Hospital, Sichuan University, Chengdu (China); Jiang, Wei, E-mail: wcumsjw72@hotmail.com [State Key Laboratory of Biotherapy, West China Hospital, Sichuan University, Chengdu (China); Molecular Medicine Research Center, West China Hospital, Sichuan University, Chengdu (China)

    2011-05-27

    Highlights: {yields} Lysophosphatidic acid (LPA) levels and its receptors LPAR1/3 transcripts were elevated during the development of radiation-induced lung fibrosis. {yields} Lung fibrosis was obviously alleviated in mice treated with the dual LPAR1/3 antagonist, VPC12249. {yields} VPC12249 administration effectively inhibited radiation-induced fibroblast accumulation in vivo, and suppressed LPA-induced fibroblast proliferation in vitro. {yields} LPA-LPAR1/3 signaling regulated TGF{beta}1 and CTGF expressions in radiation-challenged lungs, but only influenced CTGF expression in cultured fibroblasts. {yields} LPA-LPAR1/3 signaling induced fibroblast proliferation through a CTGF-dependent pathway, rather than through TGF{beta}1 activation. -- Abstract: Lung fibrosis is a common and serious complication of radiation therapy for lung cancer, for which there are no efficient treatments. Emerging evidence indicates that lysophosphatidic acid (LPA) and its receptors (LPARs) are involved in the pathogenesis of fibrosis. Here, we reported that thoracic radiation with 16 Gy in mice induced development of radiation lung fibrosis (RLF) accompanied by obvious increases in LPA release and LPAR1 and LPAR3 (LPAR1/3) transcripts. RLF was significantly alleviated in mice treated with the dual LPAR1/3 antagonist, VPC12249. VPC12249 administration effectively prolonged animal survival, restored lung structure, inhibited fibroblast accumulation and reduced collagen deposition. Moreover, profibrotic cytokines in radiation-challenged lungs obviously decreased following administration of VPC12249, including transforming growth factor {beta}1 (TGF{beta}1) and connective tissue growth factor (CTGF). In vitro, LPA induced both fibroblast proliferation and CTGF expression in a dose-dependent manner, and both were suppressed by blockade of LPAR1/3. The pro-proliferative activity of LPA on fibroblasts was inhibited by siRNA directed against CTGF. Together, our data suggest that the LPA-LPAR1

  15. Toluene diisocyanate: Induction of the autotaxin-lysophosphatidic acid axis and its association with airways symptoms

    Energy Technology Data Exchange (ETDEWEB)

    Broström, Julia M. [Division of Occupational and Environmental Medicine, Lund University, SE 221 85 Lund (Sweden); Ye, Zhi-wei [Institute of Environmental Medicine, Karolinska Institutet, Box 210, SE 171 77 Stockholm (Sweden); Axmon, Anna; Littorin, Margareta; Tinnerberg, Håkan; Lindh, Christian H. [Division of Occupational and Environmental Medicine, Lund University, SE 221 85 Lund (Sweden); Zheng, Huiyuan; Ghalali, Aram; Stenius, Ulla [Institute of Environmental Medicine, Karolinska Institutet, Box 210, SE 171 77 Stockholm (Sweden); Jönsson, Bo A.G. [Division of Occupational and Environmental Medicine, Lund University, SE 221 85 Lund (Sweden); Högberg, Johan, E-mail: johan.hogberg@ki.se [Institute of Environmental Medicine, Karolinska Institutet, Box 210, SE 171 77 Stockholm (Sweden)

    2015-09-15

    Diisocyanates are industrial chemicals which have a wide range of applications in developed and developing countries. They are notorious lung toxicants and respiratory sensitizers. However, the mechanisms behind their adverse effects are not adequately characterized. Autotaxin (ATX) is an enzyme producing lysophosphatidic acid (LPA), and the ATX-LPA axis has been implicated in lung related inflammatory conditions and diseases, including allergic asthma, but not to toxicity of environmental low-molecular-weight chemicals. We investigated effects of toluene diisocyanate (TDI) on ATX induction in human lung epithelial cell models, and we correlated LPA-levels in plasma to biomarkers of TDI exposure in urine collected from workers exposed to < 5 ppb (parts per billion). Information on workers' symptoms was collected through interviews. One nanomolar TDI robustly induced ATX release within 10 min in vitro. A P2X7- and P2X4-dependent microvesicle formation was implicated in a rapid ATX release and a subsequent protein synthesis. Co-localization between purinergic receptors and ATX was documented by immunofluorescence and confocal microscopy. The release was modulated by monocyte chemoattractant protein-1 (MCP-1) and by extracellular ATP. In workers, we found a dose–response relationship between TDI exposure biomarkers in urine and LPA levels in plasma. Among symptomatic workers reporting “sneezing”, the LPA levels were higher than among non-symptomatic workers. This is the first report indicating induction of the ATX-LPA axis by an environmental low-molecular-weight chemical, and our data suggest a role for the ATX-LPA axis in TDI toxicity. - Highlights: • Human epithelial cells release autotaxin in response to 1 nM toluene diisocyanate (TDI). • The release involves P2X4 and P2X7 receptors and is modulated by ATP and MCP-1. • Lysophosphatidic acid (LPA) was measured in workers exposed to < 5 ppb TDI. • LPA in plasma correlated to TDI exposure

  16. In vitro binding and receptor-mediated activity of terlipressin at vasopressin receptors V1 and V2.

    Science.gov (United States)

    Jamil, Khurram; Pappas, Stephen Chris; Devarakonda, Krishna R

    2018-01-01

    Terlipressin, a synthetic, systemic vasoconstrictor with selective activity at vasopressin-1 (V 1 ) receptors, is a pro-drug for the endogenous/natural porcine hormone [Lys 8 ]-vasopressin (LVP). We investigated binding and receptor-mediated cellular activities of terlipressin, LVP, and endogenous human hormone [Arg 8 ]-vasopressin (AVP) at V 1 and vasopressin-2 (V 2 ) receptors. Cell membrane homogenates of Chinese hamster ovary cells expressing human V 1 and V 2 receptors were used in competitive binding assays to measure receptor-binding activity. These cells were used in functional assays to measure receptor-mediated cellular activity of terlipressin, LVP, and AVP. Binding was measured by [ 3 H]AVP counts, and the activity was measured by fluorometric detection of intracellular calcium mobilization (V 1 ) and cyclic adenosine monophosphate (V 2 ). Binding potency at V 1 and V 2 was AVP>LVP>terlipressin. LVP and terlipressin had approximately sixfold higher affinity for V 1 than for V 2 . Cellular activity potency was also AVP>LVP>terlipressin. Terlipressin was a partial agonist at V 1 and a full agonist at V 2 ; LVP was a full agonist at both V 1 and V 2 . The in vivo response to terlipressin is likely due to the partial V 1 agonist activity of terlipressin and full V 1 agonist activity of its metabolite, LVP. These results provide supportive evidence for previous findings and further establish terlipressin pharmacology for vasopressin receptors.

  17. Receptor-mediated oral delivery of a bioencapsulated green fluorescent protein expressed in transgenic chloroplasts into the mouse circulatory system.

    Science.gov (United States)

    Limaye, Arati; Koya, Vijay; Samsam, Mohtashem; Daniell, Henry

    2006-05-01

    Oral delivery of biopharmaceutical proteins expressed in plant cells should reduce their cost of production, purification, processing, cold storage, transportation, and delivery. However, poor intestinal absorption of intact proteins is a major challenge. To overcome this limitation, we investigate here the concept of receptor-mediated oral delivery of chloroplast-expressed foreign proteins. Therefore, the transmucosal carrier cholera toxin B-subunit and green fluorescent protein (CTB-GFP), separated by a furin cleavage site, was expressed via the tobacco chloroplast genome. Polymerase chain reaction (PCR) and Southern blot analyses confirmed site-specific transgene integration and homoplasmy. Immunoblot analysis and ELISA confirmed expression of monomeric and pentameric forms of CTB-GFP, up to 21.3% of total soluble proteins. An in vitro furin cleavage assay confirmed integrity of the engineered furin cleavage site, and a GM1 binding assay confirmed the functionality of CTB-GFP pentamers. Following oral administration of CTB-GFP expressing leaf material to mice, GFP was observed in the mice intestinal mucosa, liver, and spleen in fluorescence and immunohistochemical studies, while CTB remained in the intestinal cell. This report of receptor-mediated oral delivery of a foreign protein into the circulatory system opens the door for low-cost production and delivery of human therapeutic proteins.

  18. Involvement of the lysophosphatidic acid-generating enzyme autotaxin in lymphocyte-endothelial cell interactions.

    Science.gov (United States)

    Nakasaki, Tae; Tanaka, Toshiyuki; Okudaira, Shinichi; Hirosawa, Michi; Umemoto, Eiji; Otani, Kazuhiro; Jin, Soojung; Bai, Zhongbin; Hayasaka, Haruko; Fukui, Yoshinori; Aozasa, Katsuyuki; Fujita, Naoya; Tsuruo, Takashi; Ozono, Keiichi; Aoki, Junken; Miyasaka, Masayuki

    2008-11-01

    Autotaxin (ATX) is a secreted protein with lysophospholipase D activity that generates lysophosphatidic acid (LPA) from lysophosphatidylcholine. Here we report that functional ATX is selectively expressed in high endothelial venules (HEVs) of both lymph nodes and Peyer's patches. ATX expression was developmentally regulated and coincided with lymphocyte recruitment to the lymph nodes. In adults, ATX expression was independent of HEV-expressed chemokines such as CCL21 and CXCL13, innate immunity signals including those via TLR4 or MyD88, and of the extent of lymphocyte trafficking across the HEVs. ATX expression was induced in venules at sites of chronic inflammation. Receptors for the ATX enzyme product LPA were constitutively expressed in HEV endothelial cells (ECs). In vitro, LPA induced strong morphological changes in HEV ECs. Forced ATX expression caused cultured ECs to respond to lysophosphatidylcholine, up-regulating lymphocyte binding to the ECs in a LPA receptor-dependent manner under both static and flow conditions. Although in vivo depletion of circulating ATX did not affect lymphocyte trafficking into the lymph nodes, we surmise, based on the above data, that ATX expressed by HEVs acts on HEVs in situ to facilitate lymphocyte binding to ECs and that ATX in the general circulation does not play a major role in this process. Tissue-specific inactivation of ATX will verify this hypothesis in future studies of its mechanism of action.

  19. Lysophosphatidic Acid Upregulates Laminin-332 Expression during A431 Cell Colony Dispersal

    Directory of Open Access Journals (Sweden)

    Hironobu Yamashita

    2010-01-01

    Full Text Available Lysophosphatidic acid (LPA is a bioactive phospholipid that affects various biological functions, such as cell proliferation, migration, survival, wound healing, and tumor invasion through LPA receptors. Previously, we reported that LPA induces A431 colony dispersal, accompanied by disruption of cell-cell contacts and cell migration. However, it remains unclear how LPA affects cell migration and gene expression during A431 colony dispersal. In this paper, we performed cDNA microarray analysis to investigate this question by comparing gene expression between untreated and LPA-treated A431 cells. Interestingly, these results revealed that LPA treatment upregulates several TGF-β1 target genes, including laminin-332 (Ln-332 components (α3, β3, and γ2 chains. Western blot analysis also showed that LPA increased phosphorylation of Smad2, an event that is carried out by TGF-β1 interactions. Among the genes upregulated, we further addressed the role of Ln-332. Real-time PCR analysis confirmed the transcriptional upregulation of all α3, β3, and γ2 chains of Ln-332 by LPA, corresponding to the protein level increases revealed by western blot. Further, the addition of anti-Ln-332 antibody prevented LPA-treated A431 colonies from dispersing. Taken together, our results suggest that LPA-induced Ln-332 plays a significant role in migration of individual cells from A431 colonies.

  20. 1-Oleoyl lysophosphatidic acid: a new mediator of emotional behavior in rats.

    Directory of Open Access Journals (Sweden)

    Estela Castilla-Ortega

    Full Text Available The role of lysophosphatidic acid (LPA in the control of emotional behavior remains to be determined. We analyzed the effects of the central administration of 1-oleoyl-LPA (LPA 18∶1 in rats tested for food consumption and anxiety-like and depression-like behaviors. For this purpose, the elevated plus-maze, open field, Y maze, forced swimming and food intake tests were performed. In addition, c-Fos expression in the dorsal periaqueductal gray matter (DPAG was also determined. The results revealed that the administration of LPA 18∶1 reduced the time in the open arms of the elevated plus-maze and induced hypolocomotion in the open field, suggesting an anxiogenic-like phenotype. Interestingly, these effects were present following LPA 18∶1 infusion under conditions of novelty but not under habituation conditions. In the forced swimming test, the administration of LPA 18∶1 dose-dependently increased depression-like behavior, as evaluated according to immobility time. LPA treatment induced no effects on feeding. However, the immunohistochemical analysis revealed that LPA 18∶1 increased c-Fos expression in the DPAG. The abundant expression of the LPA1 receptor, one of the main targets for LPA 18∶1, was detected in this brain area, which participates in the control of emotional behavior, using immunocytochemistry. These findings indicate that LPA is a relevant transmitter potentially involved in normal and pathological emotional responses, including anxiety and depression.

  1. Biofunctionalized Lysophosphatidic Acid/Silk Fibroin Film for Cornea Endothelial Cell Regeneration

    Science.gov (United States)

    Jeon, Hayan; Oliveira, Joaquim Miguel; Reis, Rui Luis; Khang, Gilson

    2018-01-01

    Cornea endothelial cells (CEnCs) tissue engineering is a great challenge to repair diseased or damaged CEnCs and require an appropriate biomaterial to support cell proliferation and differentiation. Biomaterials for CEnCs tissue engineering require biocompatibility, tunable biodegradability, transparency, and suitable mechanical properties. Silk fibroin-based film (SF) is known to meet these factors, but construction of functionalized graft for bioengineering of cornea is still a challenge. Herein, lysophosphatidic acid (LPA) is used to maintain and increase the specific function of CEnCs. The LPA and SF composite film (LPA/SF) was fabricated in this study. Mechanical properties and in vitro studies were performed using a rabbit model to demonstrate the characters of LPA/SF. ATR-FTIR was characterized to identify chemical composition of the films. The morphological and physical properties were performed by SEM, AFM, transparency, and contact angle. Initial cell density and MTT were performed for adhesion and cell viability in the SF and LPA/SF film. Reverse transcription polymerase chain reactions (RT-PCR) and immunofluorescence were performed to examine gene and protein expression. The results showed that films were designed appropriately for CEnCs delivery. Compared to pristine SF, LPA/SF showed higher biocompatibility, cell viability, and expression of CEnCs specific genes and proteins. These indicate that LPA/SF, a new biomaterial, offers potential benefits for CEnCs tissue engineering for regeneration. PMID:29710848

  2. Lysophosphatidic Acid Disrupts Junctional Integrity and Epithelial Cohesion in Ovarian Cancer Cells

    Directory of Open Access Journals (Sweden)

    Yueying Liu

    2012-01-01

    Full Text Available Ovarian cancer metastasizes via exfoliation of free-floating cells and multicellular aggregates from the primary tumor to the peritoneal cavity. A key event in EOC metastasis is disruption of cell-cell contacts via modulation of intercellular junctional components including cadherins. Ascites is rich in lysophosphatidic acid (LPA, a bioactive lipid that may promote early events in ovarian cancer dissemination. The objective of this paper was to assess the effect of LPA on E-cadherin junctional integrity. We report a loss of junctional E-cadherin in OVCAR3, OVCA429, and OVCA433 cells exposed to LPA. LPA-induced loss of E-cadherin was concentration and time dependent. LPA increased MMP-9 expression and promoted MMP-9-catalyzed E-cadherin ectodomain shedding. Blocking LPA receptor signaling inhibited MMP-9 expression and restored junctional E-cadherin staining. LPA-treated cells demonstrated a significant decrease in epithelial cohesion. Together these data support a model wherein LPA induces MMP-9 expression and MMP-9-catalyzed E-cadherin ectodomain shedding, resulting in loss of E-cadherin junctional integrity and epithelial cohesion, facilitating metastatic dissemination of ovarian cancer cells.

  3. Lysophosphatidic acid-functionalised titanium as a superior surface for supporting human osteoblast (MG63 maturation

    Directory of Open Access Journals (Sweden)

    JP Mansell

    2012-05-01

    Full Text Available Covalent modifications of titanium with small molecules known to promote human osteoblast maturation are especially attractive in developing superior biomaterials. An important step in securing competent bone formation at implant sites is promoting the formation of mature osteoblasts, either from committed pre-osteoblasts or from their mesenchymal progenitors. To this end our research has focussed on identifying molecules that enhance human osteoblast formation and maturation and to develop ways of covalently attaching these molecules to implant surfaces so that they are more likely to withstand the rigors of the implantation process whilst still retaining their bioactivity. Herein we report the novel production of lipid-functionalised titanium using lysophosphatidic acid or a related compound, (3S 1-fluoro-3-hydroxy-4-butyl-1-phosphonate. Both lipids were especially effective at co-operating with calcitriol to promote human osteoblast maturation at these modified Ti surfaces in vitro. The novel findings presented offer enticing new developments towards the fabrication of next-generation implant devices with the potential to significantly enhance the osseointegration process and with it improvements in future prosthesis performance and longevity.

  4. Lysophosphatidic acid stimulates thrombomodulin lectin-like domain shedding in human endothelial cells

    International Nuclear Information System (INIS)

    Wu Hualin; Lin ChiIou; Huang Yuanli; Chen, Pin-Shern; Kuo, Cheng-Hsiang; Chen, Mei-Shing; Wu, G.C.-C.; Shi, G.-Y.; Yang, H.-Y.; Lee Hsinyu

    2008-01-01

    Thrombomodulin (TM) is an anticoagulant glycoprotein highly expressed on endothelial cell surfaces. Increased levels of soluble TM in circulation have been widely accepted as an indicator of endothelial damage or dysfunction. Previous studies indicated that various proinflammatory factors stimulate TM shedding in various cell types such as smooth muscle cells and epithelial cells. Lysophosphatidic acid (LPA) is a bioactive lipid mediator present in biological fluids during endothelial damage or injury. In the present study, we first observed that LPA triggered TM shedding in human umbilical vein endothelial cells (HUVECs). By Cyflow analysis, we showed that the LPA-induced accessibility of antibodies to the endothelial growth factor (EGF)-like domain of TM is independent of matrix metalloproteinases (MMPs), while LPA-induced TM lectin-like domain shedding is MMP-dependent. Furthermore, a stable cell line expressing TM without its lectin-like domain exhibited a higher cell proliferation rate than a stable cell line expressing full-length TM. These results imply that LPA induces TM lectin-like domain shedding, which might contribute to the exposure of its EGF-like domain for EGF receptor (EGFR) binding, thereby stimulating subsequent cell proliferation. Based on our findings, we propose a novel mechanism for the exposure of TM EGF-like domain, which possibly mediates LPA-induced EGFR transactivation

  5. Halothane inhibits the cholinergic-receptor-mediated influx of calcium in primary culture of bovine adrenal medulla cells

    International Nuclear Information System (INIS)

    Yashima, N.; Wada, A.; Izumi, F.

    1986-01-01

    Adrenal medulla cells are cholinoceptive cells. Stimulation of the acetylcholine receptor causes the influx of Ca to the cells, and Ca acts as the coupler of the stimulus-secretion coupling. In this study, the authors investigated the effects of halothane on the receptor-mediated influx of 45 Ca using cultured bovine adrenal medulla cells. Halothane at clinical concentrations (0.5-2%) inhibited the influx of 45 Ca caused by carbachol, with simultaneous inhibition of catecholamine secretion. The influx of 45 Ca and the secretion of catecholamines caused by K depolarization were inhibited by a large concentration of Mg, which competes with Ca at Ca channels, but not inhibited by halothane. Inhibition of the 45 Ca influx by halothane was not overcome by increase in the carbachol concentration. Inhibition of the 45 Ca influx by halothane was examined in comparison with that caused by a large concentration of Mg by the application of Scatchard analysis as the function of the external Ca concentration. Halothane decreased the maximal influx of 45 Ca without altering the apparent kinetic constant of Ca to Ca channels. On the contrary, a large concentration of Mg increased the apparent kinetic constant without altering the maximal influx of 45 Ca. Based on these findings, the authors suggest that inhibition of the 45 Ca influx by halothane was not due to the direct competitive inhibition of Ca channels, nor to the competitive antagonism of agonist-receptor interaction. As a possibility, halothane seems to inhibit the receptor-mediated activation of Ca channels through the interference of coupling between the receptor and Ca channels

  6. Reboxetine Enhances the Olanzapine-Induced Antipsychotic-Like Effect, Cortical Dopamine Outflow and NMDA Receptor-Mediated Transmission

    Science.gov (United States)

    Marcus, Monica M; Jardemark, Kent; Malmerfelt, Anna; Björkholm, Carl; Svensson, Torgny H

    2010-01-01

    Preclinical data have shown that addition of the selective norepinephrine transporter (NET) inhibitor reboxetine increases the antipsychotic-like effect of the D2/3 antagonist raclopride and, in parallel, enhances cortical dopamine output. Subsequent clinical results suggested that adding reboxetine to stable treatments with various antipsychotic drugs (APDs) may improve positive, negative and depressive symptoms in schizophrenia. In this study, we investigated in rats the effects of adding reboxetine to the second-generation APD olanzapine on: (i) antipsychotic efficacy, using the conditioned avoidance response (CAR) test, (ii) extrapyramidal side effect (EPS) liability, using a catalepsy test, (iii) dopamine efflux in the medial prefrontal cortex and the nucleus accumbens, using in vivo microdialysis in freely moving animals and (iv) cortical N-methyl--aspartate (NMDA) receptor-mediated transmission, using intracellular electrophysiological recording in vitro. Reboxetine (6 mg/kg) enhanced the suppression of CAR induced by a suboptimal dose (1.25 mg/kg), but not an optimal (2.5 mg/kg) dose of olanzapine without any concomitant catalepsy. Addition of reboxetine to the low dose of olanzapine also markedly increased cortical dopamine outflow and facilitated prefrontal NMDA receptor-mediated transmission. Our data suggest that adjunctive treatment with a NET inhibitor may enhance the therapeutic effect of low-dose olanzapine in schizophrenia without increasing EPS liability and add an antidepressant action, thus in principle allowing for a dose reduction of olanzapine with a concomitant reduction of dose-related side effects, such as EPS and weight gain. PMID:20463659

  7. Prostaglandin E2 potentiation of P2X3 receptor mediated currents in dorsal root ganglion neurons

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    Huang Li-Yen

    2007-08-01

    Full Text Available Abstract Prostaglandin E2 (PGE2 is a well-known inflammatory mediator that enhances the excitability of DRG neurons. Homomeric P2X3 and heteromeric P2X2/3 receptors are abundantly expressed in dorsal root ganglia (DRG neurons and participate in the transmission of nociceptive signals. The interaction between PGE2 and P2X3 receptors has not been well delineated. We studied the actions of PGE2 on ATP-activated currents in dissociated DRG neurons under voltage-clamp conditions. PGE2 had no effects on P2X2/3 receptor-mediated responses, but significantly potentiated fast-inactivating ATP currents mediated by homomeric P2X3 receptors. PGE2 exerted its action by activating EP3 receptors. To study the mechanism underlying the action of PGE2, we found that the adenylyl cyclase activator, forskolin and the membrane-permeable cAMP analogue, 8-Br-cAMP increased ATP currents, mimicking the effect of PGE2. In addition, forskolin occluded the enhancement produced by PGE2. The protein kinase A (PKA inhibitors, H89 and PKA-I blocked the PGE2 effect. In contrast, the PKC inhibitor, bisindolymaleimide (Bis did not change the potentiating action of PGE2. We further showed that PGE2 enhanced α,β-meATP-induced allodynia and hyperalgesia and the enhancement was blocked by H89. These observations suggest that PGE2 binds to EP3 receptors, resulting in the activation of cAMP/PKA signaling pathway and leading to an enhancement of P2X3 homomeric receptor-mediated ATP responses in DRG neurons.

  8. Electroacupuncture improves cerebral blood flow and attenuates moderate ischemic injury via Angiotensin II its receptors-mediated mechanism in rats.

    Science.gov (United States)

    Li, Jing; He, Jiaojun; Du, Yuanhao; Cui, Jingjun; Ma, Ying; Zhang, Xuezhu

    2014-11-11

    To investigate the effects and potential mechanism of electroacupuncture intervention on expressions of Angiotensin II and its receptors-mediated signaling pathway in experimentally induced cerebral ischemia. Totally 126 male Wistar rats were randomly divided into control group, model group and EA group. The latter two were further divided into ten subgroups (n = 6) following Middle Cerebral Artery Occlusion (MCAO). Changes in regional cerebral blood flow (rCBF) and expressions of Angiotensin II and its receptors (AT1R, AT2R), as well as effector proteins in phosphatidyl inositol signal pathway were monitored before and at different times after MCAO. MCAO-induced decline of ipsilateral rCBF was partially suppressed by electroacupuncture, and contralateral blood flow was also superior to that of model group. Angiotensin II level was remarkably elevated immediately after MCAO, while electroacupuncture group exhibited significantly lower levels at 1 to 3 h and the value was significantly increased thereafter. The enhanced expression of AT1R was partially inhibited by electroacupuncture, while increased AT2R level was further induced. Electroacupuncture stimulation attenuated and postponed the upregulated-expressions of Gq and CaM these upregulations. ELISA results showed sharply increased expressions of DAG and IP3, which were remarkably neutralized by electroacupuncture. MCAO induced significant increases in expression of Angiotensin II and its receptor-mediated signal pathway. These enhanced expressions were significantly attenuated by electroacupuncture intervention, followed by reduced vasoconstriction and improved blood supply in ischemic region, and ultimately conferred beneficial effects on cerebral ischemia.

  9. Loss of Progesterone Receptor-Mediated Actions Induce Preterm Cellular and Structural Remodeling of the Cervix and Premature Birth

    Science.gov (United States)

    Yellon, Steven M.; Dobyns, Abigail E.; Beck, Hailey L.; Kurtzman, James T.; Garfield, Robert E.; Kirby, Michael A.

    2013-01-01

    A decline in serum progesterone or antagonism of progesterone receptor function results in preterm labor and birth. Whether characteristics of premature remodeling of the cervix after antiprogestins or ovariectomy are similar to that at term was the focus of the present study. Groups of pregnant rats were treated with vehicle, a progesterone receptor antagonist (onapristone or mifepristone), or ovariectomized on day 17 postbreeding. As expected, controls given vehicle delivered at term while rats delivered preterm after progesterone receptor antagonist treatment or ovariectomy. Similar to the cervix before term, the preterm cervix of progesterone receptor antagonist-treated rats was characterized by reduced cell nuclei density, decreased collagen content and structure, as well as a greater presence of macrophages per unit area. Thus, loss of nuclear progesterone receptor-mediated actions promoted structural remodeling of the cervix, increased census of resident macrophages, and preterm birth much like that found in the cervix at term. In contrast to the progesterone receptor antagonist-induced advance in characteristics associated with remodeling, ovariectomy-induced loss of systemic progesterone did not affect hypertrophy, extracellular collagen, or macrophage numbers in the cervix. Thus, the structure and macrophage census in the cervix appear sufficient for premature ripening and birth to occur well before term. With progesterone receptors predominantly localized on cells other than macrophages, the findings suggest that interactions between cells may facilitate the loss of progesterone receptor-mediated actions as part of a final common mechanism that remodels the cervix in certain etiologies of preterm and with parturition at term. PMID:24339918

  10. Ginseng pharmacology: a new paradigm based on gintonin-lysophosphatidic acid receptor interactions

    Directory of Open Access Journals (Sweden)

    Seung-Yeol eNah

    2015-10-01

    Full Text Available Ginseng, the root of Panax ginseng, is used as a traditional medicine. Despite the long history of the use of ginseng, there is no specific scientific or clinical rationale for ginseng pharmacology besides its application as a general tonic. The ambiguous description of ginseng pharmacology might be due to the absence of a predominant active ingredient that represents ginseng pharmacology. Recent studies show that ginseng abundantly contains lysophosphatidic acids (LPAs, which are phospholipid-derived growth factor with diverse biological functions including those claimed to be exhibited by ginseng. LPAs in ginseng form a complex with ginseng proteins, which can bind and deliver LPA to its cognate receptors with a high affinity. As a first messenger, gintonin produces second messenger Ca2+ via G protein-coupled LPA receptors. Ca2+ is an intracellular mediator of gintonin and initiates a cascade of amplifications for further intercellular communications by activation of Ca2+-dependent kinases, receptors, gliotransmitter and neurotransmitter release. Ginsenosides, which have been regarded as primary ingredients of ginseng, cannot elicit intracellular [Ca2+]i transients, since they lack specific cell surface receptor. However, ginsenosides exhibit non-specific ion channel and receptor regulations. This is the key characteristic that distinguishes gintonin from ginsenosides. Although the current discourse on ginseng pharmacology is focused on ginsenosides, gintonin can definitely provide a mode of action for ginseng pharmacology that ginsenosides cannot. This review article introduces a novel concept of ginseng ligand-LPA receptor interaction and proposes to establish a paradigm that shifts the focus from ginsenosides to gintonin as a major ingredient representing ginseng pharmacology.

  11. Autotaxin-lysophosphatidic acid axis is a novel molecular target for lowering intraocular pressure.

    Directory of Open Access Journals (Sweden)

    Padma Iyer

    Full Text Available Primary open-angle glaucoma is the second leading cause of blindness in the United States and is commonly associated with elevated intraocular pressure (IOP resulting from diminished aqueous humor (AH drainage through the trabecular pathway. Developing effective therapies for increased IOP in glaucoma patients requires identification and characterization of molecular mechanisms that regulate IOP and AH outflow. This study describes the identification and role of autotaxin (ATX, a secretory protein and a major source for extracellular lysophosphatidic acid (LPA, in regulation of IOP in a rabbit model. Quantitative proteomics analysis identified ATX as an abundant protein in both human AH derived from non-glaucoma subjects and in AH from different animal species. The lysophospholipase D (LysoPLD activity of ATX was found to be significantly elevated (by ∼1.8 fold; n=20 in AH derived from human primary open angle glaucoma patients as compared to AH derived from age-matched cataract control patients. Immunoblotting analysis of conditioned media derived from primary cultures of human trabecular meshwork (HTM cells has confirmed secretion of ATX and the ability of cyclic mechanical stretch of TM cells to increase the levels of secreted ATX. Topical application of a small molecular chemical inhibitor of ATX (S32826, which inhibited AH LysoPLD activity in vitro (by >90%, led to a dose-dependent and significant decrease of IOP in Dutch-Belted rabbits. Single intracameral injection of S32826 (∼2 µM led to significant reduction of IOP in rabbits, with the ocular hypotensive response lasting for more than 48 hrs. Suppression of ATX expression in HTM cells using small-interfering RNA (siRNA caused a decrease in actin stress fibers and myosin light chain phosphorylation. Collectively, these observations indicate that the ATX-LPA axis represents a potential therapeutic target for lowering IOP in glaucoma patients.

  12. Lysophosphatidic acid rescues bone mesenchymal stem cells from hydrogen peroxide-induced apoptosis.

    Science.gov (United States)

    Wang, Xian-Yun; Fan, Xue-Song; Cai, Lin; Liu, Si; Cong, Xiang-Feng; Chen, Xi

    2015-03-01

    The increase of reactive oxygen species in infracted heart significantly reduces the survival of donor mesenchymal stem cells, thereby attenuating the therapeutic efficacy for myocardial infarction. In our previous study, we demonstrated that lysophosphatidic acid (LPA) protects bone marrow-derived mesenchymal stem cells (BMSCs) against hypoxia and serum deprivation-induced apoptosis. However, whether LPA protects BMSCs from H2O2-induced apoptosis was not examined. In this study, we report that H2O2 induces rat BMSC apoptosis whereas LPA pre-treatment effectively protects BMSCs from H2O2-induced apoptosis. LPA protection of BMSC from the induced apoptosis is mediated mostly through LPA3 receptor. Furthermore, we found that membrane G protein Gi2 and Gi3 are involved in LPA-elicited anti-apoptotic effects through activation of ERK1/2- and PI3 K-pathways. Additionally, H2O2 increases levels of type II of light chain 3B (LC3B II), an autophagy marker, and H2O2-induced autophagy thus protected BMSCs from apoptosis. LPA further increases the expression of LC3B II in the presence of H2O2. In contrast, autophagy flux inhibitor bafilomycin A1 has no effect on LPA's protection of BMSC from H2O2-induced apoptosis. Taken together, our data suggest that LPA rescues H2O2-induced apoptosis mainly by interacting with Gi-coupled LPA3, resulting activation of the ERK1/2- and PI3 K/AKT-pathways and inhibition caspase-3 cleavage, and LPA protection of BMSCs against the apoptosis is independent of it induced autophagy.

  13. Lysophosphatidic acid induces expression of genes in human oral keratinocytes involved in wound healing.

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    Thorlakson, Hong Huynh; Engen, Stian Andre; Schreurs, Olav; Schenck, Karl; Blix, Inger Johanne Schytte

    2017-08-01

    Epithelial cells participate in wound healing by covering wounds, but also as important mediators of wound healing processes. Topical application of the phospholipid growth factor lysophosphatidic acid (LPA) accelerates dermal wound healing and we hypothesized that LPA can play a role in human oral wound healing through its effects on human oral keratinocytes (HOK). HOK were isolated from gingival biopsies and exposed to LPA. The LPA receptor profile, signal transduction pathways, gene expression and secretion of selected cytokines were analyzed. HOK expressed the receptors LPA 1 , LPA 5 and LPA 6 and LPA activated the ERK1/2, JNK and p38 intracellular pathways, substantiated by secretion of IL-6 and IL-8. The early (2h) and intermediate (6h) gene expression profiles of HOK after LPA treatment showed a wide array of regulated genes. The majority of the strongest upregulated genes were related to chemotaxis and inflammation, and became downregulated after 6h. At 6h, genes coding for factors involved in extracellular matrix remodeling and re-epithelialization became highly expressed. IL-36γ, not earlier known to be regulated by LPA, was strongly transcribed and translated but not secreted. After stimulation with LPA, HOK responded by regulating factors and genes that are essential in wound healing processes. As LPA is found in saliva and is released by activated cells after wounding, our results indicate that LPA has a favorable physiological role in oral wound healing. This may further point towards a beneficial role for application of LPA on oral surgical or chronic wounds. Copyright © 2017 Elsevier Ltd. All rights reserved.

  14. Lysophosphatidic Acid Level and the Incidence of Silent Brain Infarction in Patients with Nonvalvular Atrial Fibrillation

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    Li Zhou

    2010-10-01

    Full Text Available Lysophosphatidic acid (LPA, which is proposed to play an important role in normal physiological situations such as wound healing, vascular tone, vascular integrity and reproduction, may be involved in the etiology of some diseases such as atherosclerosis, cancer, obesity or myocardial infarction. Abnormal findings, including silent brain infarction (SBI, are frequently observed by magnetic resonance imaging (MRI in patients with nonvalvular atrial fibrillation (NVAF. However, whether there is a relationship between LPA level and the prevalence of SBI has not been extensively studied. In the present study, the association between them was investigated. 235 patients with NVAF, 116 cases of SBI without NVAF and 120 cases of healthy volunteers (control group, who did not receive any antithrombotic therapy, were enrolled in this study. Plasma LPA levels in the NVAF with SBI group were significantly higher than that in the control group (p < 0.01, NVAF without SBI group (p < 0.01 and SBI without NVAF group (p < 0.01. The LPA levels are lower in the control group than in the NVAF without SBI and SBI without NVAF groups (p < 0.01, however, the latter two groups did not significantly differ from each other for LPA levels (p > 0.05 There were significant differences in the positive rate of platelet activation between each of the groups (p < 0.01. The positive rate of platelet activation was significantly higher in the NVAF with SBI group. We suggest that LPA might be a novel marker for estimation of the status of platelet activation and the risk factor for SBI onset in NVAF patients. We expected that plasma LPA levels could predict the occurrence of SBI in NVAF patients.

  15. Lysophosphatidic acid acyltransferase β (LPAATβ promotes the tumor growth of human osteosarcoma.

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    Farbod Rastegar

    2010-12-01

    Full Text Available Osteosarcoma is the most common primary malignancy of bone with poorly characterized molecular pathways important in its pathogenesis. Increasing evidence indicates that elevated lipid biosynthesis is a characteristic feature of cancer. We sought to investigate the role of lysophosphatidic acid acyltransferase β (LPAATβ, aka, AGPAT2 in regulating the proliferation and growth of human osteosarcoma cells. LPAATβ can generate phosphatidic acid, which plays a key role in lipid biosynthesis as well as in cell proliferation and survival. Although elevated expression of LPAATβ has been reported in several types of human tumors, the role of LPAATβ in osteosarcoma progression has yet to be elucidated.Endogenous expression of LPAATβ in osteosarcoma cell lines is analyzed by using semi-quantitative PCR and immunohistochemical staining. Adenovirus-mediated overexpression of LPAATβ and silencing LPAATβ expression is employed to determine the effect of LPAATβ on osteosarcoma cell proliferation and migration in vitro and osteosarcoma tumor growth in vivo. We have found that expression of LPAATβ is readily detected in 8 of the 10 analyzed human osteosarcoma lines. Exogenous expression of LPAATβ promotes osteosarcoma cell proliferation and migration, while silencing LPAATβ expression inhibits these cellular characteristics. We further demonstrate that exogenous expression of LPAATβ effectively promotes tumor growth, while knockdown of LPAATβ expression inhibits tumor growth in an orthotopic xenograft model of human osteosarcoma.Our results strongly suggest that LPAATβ expression may be associated with the aggressive phenotypes of human osteosarcoma and that LPAATβ may play an important role in regulating osteosarcoma cell proliferation and tumor growth. Thus, targeting LPAATβ may be exploited as a novel therapeutic strategy for the clinical management of osteosarcoma. This is especially attractive given the availability of selective

  16. Lysophosphatidic acid directly induces macrophage-derived foam cell formation by blocking the expression of SRBI.

    Science.gov (United States)

    Chen, Linmu; Zhang, Jun; Deng, Xiao; Liu, Yan; Yang, Xi; Wu, Qiong; Yu, Chao

    2017-09-23

    The leading cause of morbidity and mortality is the result of cardiovascular disease, mainly atherosclerosis. The formation of macrophage foam cells by ingesting ox-LDL and focal retention in the subendothelial space are the hallmarks of the early atherosclerotic lesion. Lysophosphatidic acid (LPA), which is a low-molecular weight lysophospholipid enriched in oxidized LDL, exerts a range of effects on the cardiovascular system. Previous reports show that LPA increases the uptake of ox-LDL to promote the formation of foam cells. However, as the most active component of ox-LDL, there is no report showing whether LPA directly affects foam cell formation. The aim of this study was to investigate the effects of LPA on foam cell formation, as well as to elucidate the underlying mechanism. Oil red O staining and a Cholesterol/cholesteryl ester quantitation assay were used to evaluate foam cell formation in Raw264.7 macrophage cells. We utilized a Western blot and RT-PCR to investigate the relationship between LPA receptors and lipid transport related proteins. We found that LPA promoted foam cell formation, using 200 μM for 24 h. Meanwhile, the expression of the Scavenger receptor BI (SRBI), which promotes the efflux of free cholesterol, was decreased. Furthermore, the LPA 1/3 receptor antagonist Ki16425 significantly abolished the LPA effects, indicating that LPA 1/3 was involved in the foam cell formation and SRBI expression induced by LPA. Additionally, the LPA-induced foam cell formation was blocked with an AKT inhibitor. Our results suggest that LPA-enhanced foam cell formation is mediated by LPA 1/3 -AKT activation and subsequent SRBI expression. Copyright © 2017. Published by Elsevier Inc.

  17. Lysophosphatidic acid (LPA) effects on endometrial carcinoma in vitro proliferation, invasion, and matrix metalloproteinase activity.

    Science.gov (United States)

    Wang, Feng-qiang; Ariztia, Edgardo V; Boyd, Leslie R; Horton, Faith R; Smicun, Yoel; Hetherington, Jessica A; Smith, Phillip J; Fishman, David A

    2010-04-01

    Lysophosphatidic acid (LPA) has potent growth-regulatory effect in many cell types and has been linked to the in vivo tumor growth and metastasis in several malignancies. The goal of this study was to assess the regulation of (EC) microenvironment by LPA through the examination of its effect on cell proliferation, migration, invasion, uPA activity, and matrix metalloproteinase (MMP) secretion/activation. All experiments were performed in vitro using an EC cell line, HEC-1A. Cell proliferation was determined using the Promega MTS proliferation assay following 48 h of exposures to different concentrations of LPA (0.1, 1.0 and 10.0 microM). Cell invasion was assessed using a modified Boyden chamber assay with collagen I coated on the membrane. HEC-1A motility was examined by Boyden chamber migration assay as well as the scratch wound closure assay on type I collagen. MMP secretion/activation in HEC-1A conditioned medium was detected by gelatin zymography. MMP-7 mRNA expression was determined using real-time PCR. uPA activity was measured using a coupled colorimetric assay. LPA, at the concentrations of 0.1 and 1.0 microM, significantly induced the proliferation of HEC-1A cells (p0.05). Gelatin zymogram showed that HEC-1A cells secreted high levels of MMP-7, while MMP-2 and MMP-9 are barely detectable. In addition, LPA significantly enhanced uPA activity in HEC-1A conditioned medium in a concentration-dependent manner. LPA is a potent modulator of cellular proliferation and invasion for EC cells. It also has the capacity to stimulate the secretion/activity of uPA and MMP-7. Those results suggest that LPA is a bioactive modulator of EC microenvironment and may have a distinct regulation mechanism as observed in epithelial ovarian cancer. Copyright 2009. Published by Elsevier Inc.

  18. Lysophosphatidic acid induces chemotaxis in MC3T3-E1 osteoblastic cells

    Energy Technology Data Exchange (ETDEWEB)

    Masiello, Lisa M.; Fotos, Joseph S.; Galileo, Deni S.; Karin, Norm J.

    2006-07-01

    Lysophosphatidic acid (LPA) is a bioactive lipid that has pleiotropic effects on a variety of cell types and enhances the migration of endothelial and cancer cells, but it is not known if this lipid can alter osteoblast motility. We performed transwell migration assays using MC3T3-E1 osteoblastic cells and found LPA to be a potent chemotactic agent. Quantitative time-lapse video analysis of osteoblast migration after wounds were introduced into cell monolayers indicated that LPA stimulated both migration velocity and the average migration distance per cell. LPA also elicited substantial changes in cell shape and actin cytoskeletal structure; lipid-treated cells contained fewer stress fibers and displayed long membrane processes that were enriched in F-actin. Quantitative RT-PCR analysis showed that MC3T3-E1 cells express all four known LPA-specific G protein-coupled receptors (LPA1-LPA4) with a relative mRNA abundance of LPA1 > LPA4 > LPA2 >> LPA3. LPA-induced changes in osteoblast motility and morphology were antagonized by both pertussis toxin and Ki16425, a subtype-specific blocker of LPA1 and LPA3 receptor function. Cell migration in many cell types is linked to changes in intracellular Ca2+. Ki16425 also inhibited LPA-induced Ca2+ signaling in a dose-dependent manner, suggesting a link between LPA-induced Ca2+ transients and osteoblast chemotaxis. Our data show that LPA stimulates MC3T3-E1 osteoblast motility via a mechanism that is linked primarily to the G protein-coupled receptor LPA1.

  19. Bovine ovarian follicular growth and development correlate with lysophosphatidic acid expression.

    Science.gov (United States)

    Sinderewicz, Emilia; Grycmacher, Katarzyna; Boruszewska, Dorota; Kowalczyk-Zięba, Ilona; Staszkiewicz, Joanna; Ślężak, Tomasz; Woclawek-Potocka, Izabela

    2018-01-15

    The basis of successful reproduction is proper ovarian follicular growth and development. In addition to prostaglandins and vascular endothelial growth factor, a number of novel factors are suggested as important regulators of follicular growth and development: PGES, TFG, CD36, RABGAP1, DBI and BTC. This study focuses on examining the expression of these factors in granulosa and thecal cells that originate from different ovarian follicle types and their link with the expression of lysophosphatidic acid (LPA), known local regulator of reproductive functions in the cow. Ovarian follicles were divided into healthy, transitional, and atretic categories. The mRNA expression levels for PGES, TFG, CD36, RABGAP1, DBI and BTC in granulosa and thecal cells in different follicle types were measured by real-time PCR. The correlations among expression of enzymes synthesizing LPA (autotaxin, phospholipase A2), receptors for LPA and examined factors were measured. Immunolocalization of PGES, TFG, CD36, RABGAP1, DBI and BTC was examined by immunohistochemistry. We investigated follicle-type dependent mRNA expression of factors potentially involved in ovarian follicular growth and development, both in granulosa and thecal cells of bovine ovarian follicles. Strong correlations among receptors for LPA, enzymes synthesizing LPA, and the examined factors in healthy and transitional follicles were observed, with its strongest interconnection with TFG, DBI and RABGAP1 in granulosa cells, and TFG in thecal cells; whereas no correlations in atretic follicles were detected. A greater number of correlations were found in thecal cells than in granulosa cells as well as in healthy follicles than in transitional follicles. These data indicate the role of LPA in the growth, development and physiology of the bovine ovarian follicle. Copyright © 2017 Elsevier Inc. All rights reserved.

  20. Phenobarbital but not diazepam reduces AMPA/Kainate receptor mediated currents and exerts opposite actions on initial seizures in the neonatal rat hippocampus

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    Romain eNardou

    2011-07-01

    Full Text Available Diazepam (DZP and phenobarbital (PB are extensively used as first and second line drugs to treat acute seizures in neonates and their actions are thought to be mediated by increasing the actions of GABAergic signals. Yet, their efficacy is variable with occasional failure or even aggravation of recurrent seizures questioning whether other mechanisms are not involved in their actions. We have now compared the effects of DZP and PB on ictal-like events (ILEs in an in vitro model of mirror focus (MF. Using the three-compartment chamber with the two immature hippocampi and their commissural fibers placed in 3 different compartments, kainate was applied to one hippocampus and PB or DZP to the contralateral one, either after one ILE or after many recurrent ILEs that produce an epileptogenic MF. We report that in contrast to PB, DZP aggravated propagating ILEs from the start and did not prevent the formation of MF. PB reduced and DZP increased the network driven Giant Depolarising Potentials suggesting that PB may exert additional actions that are not mediated by GABA signalling. In keeping with this, PB but not DZP reduced field potentials recorded in the presence of GABA and NMDA receptor antagonists. These effects are mediated by a direct action on AMPA/Kainate receptors since PB: i reduced AMPA/Kainate receptor mediated currents induced by focal applications of glutamate ; ii reduced the amplitude and the frequency of AMPA but not NMDA receptor mediated miniature EPSCs; iii augmented the number of AMPA receptor mediated EPSCs failures evoked by minimal stimulation. These effects persisted in MF. Therefore, PB exerts its anticonvulsive actions partly by reducing AMPA/Kainate receptors mediated EPSCs in addition to the pro-GABA effects. We suggest that PB may have advantage over DZP in the treatment of initial neonatal seizures since the additional reduction of glutamate receptors mediated signals may reduce the severity of neonatal seizures.

  1. Prenatal Alcohol Exposure Increases Histamine H3 Receptor-Mediated Inhibition of Glutamatergic Neurotransmission in Rat Dentate Gyrus.

    Science.gov (United States)

    Varaschin, Rafael K; Allen, Nyika A; Rosenberg, Martina J; Valenzuela, C Fernando; Savage, Daniel D

    2018-02-01

    We have reported that prenatal alcohol exposure (PAE)-induced deficits in dentate gyrus, long-term potentiation (LTP), and memory are ameliorated by the histamine H 3 receptor inverse agonist ABT-239. Curiously, ABT-239 did not enhance LTP or memory in control offspring. Here, we initiated an investigation of how PAE alters histaminergic neurotransmission in the dentate gyrus and other brain regions employing combined radiohistochemical and electrophysiological approaches in vitro to examine histamine H 3 receptor number and function. Long-Evans rat dams voluntarily consumed either a 0% or 5% ethanol solution 4 hours each day throughout gestation. This pattern of drinking, which produces a mean peak maternal serum ethanol concentration of 60.8 ± 5.8 mg/dl, did not affect maternal weight gain, litter size, or offspring birthweight. Radiohistochemical studies in adult offspring revealed that specific [ 3 H]-A349821 binding to histamine H 3 receptors was not different in PAE rats compared to controls. However, H 3 receptor-mediated G i /G o protein-effector coupling, as measured by methimepip-stimulated [ 35 S]-GTPγS binding, was significantly increased in cerebral cortex, cerebellum, and dentate gyrus of PAE rats compared to control. A LIGAND analysis of detailed methimepip concentration-response curves in dentate gyrus indicated that PAE significantly elevates receptor-effector coupling by a lower affinity H 3 receptor population without significantly altering the affinities of H 3 receptor subpopulations. In agreement with the [ 35 S]-GTPγS studies, a similar range of methimepip concentrations also inhibited electrically evoked field excitatory postsynaptic potential responses and increased paired-pulse ratio, a measure of decreased glutamate release, to a significantly greater extent in dentate gyrus slices from PAE rats than in controls. These results suggest that a PAE-induced elevation in H 3 receptor-mediated inhibition of glutamate release from

  2. Lysophosphatidic acid receptor-5 negatively regulates cellular responses in mouse fibroblast 3T3 cells

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    Dong, Yan; Hirane, Miku; Araki, Mutsumi [Division of Cancer Biology and Bioinformatics, Department of Life Science, Faculty of Science and Engineering, Kinki University, 3-4-1, Kowakae, Higashiosaka, Osaka 577-8502 (Japan); Fukushima, Nobuyuki [Division of Molecular Neurobiology, Department of Life Science, Faculty of Science and Engineering, Kinki University, 3-4-1, Kowakae, Higashiosaka, Osaka 577-8502 (Japan); Tsujiuchi, Toshifumi, E-mail: ttujiuch@life.kindai.ac.jp [Division of Cancer Biology and Bioinformatics, Department of Life Science, Faculty of Science and Engineering, Kinki University, 3-4-1, Kowakae, Higashiosaka, Osaka 577-8502 (Japan)

    2014-04-04

    Highlights: • LPA{sub 5} inhibits the cell growth and motile activities of 3T3 cells. • LPA{sub 5} suppresses the cell motile activities stimulated by hydrogen peroxide in 3T3 cells. • Enhancement of LPA{sub 5} on the cell motile activities inhibited by LPA{sub 1} in 3T3 cells. • The expression and activation of Mmp-9 were inhibited by LPA{sub 5} in 3T3 cells. • LPA signaling via LPA{sub 5} acts as a negative regulator of cellular responses in 3T3 cells. - Abstract: Lysophosphatidic acid (LPA) signaling via G protein-coupled LPA receptors (LPA{sub 1}–LPA{sub 6}) mediates a variety of biological functions, including cell migration. Recently, we have reported that LPA{sub 1} inhibited the cell motile activities of mouse fibroblast 3T3 cells. In the present study, to evaluate a role of LPA{sub 5} in cellular responses, Lpar5 knockdown (3T3-L5) cells were generated from 3T3 cells. In cell proliferation assays, LPA markedly stimulated the cell proliferation activities of 3T3-L5 cells, compared with control cells. In cell motility assays with Cell Culture Inserts, the cell motile activities of 3T3-L5 cells were significantly higher than those of control cells. The activity levels of matrix metalloproteinases (MMPs) were measured by gelatin zymography. 3T3-L5 cells stimulated the activation of Mmp-2, correlating with the expression levels of Mmp-2 gene. Moreover, to assess the co-effects of LPA{sub 1} and LPA{sub 5} on cell motile activities, Lpar5 knockdown (3T3a1-L5) cells were also established from Lpar1 over-expressing (3T3a1) cells. 3T3a1-L5 cells increased the cell motile activities of 3T3a1 cells, while the cell motile activities of 3T3a1 cells were significantly lower than those of control cells. These results suggest that LPA{sub 5} may act as a negative regulator of cellular responses in mouse fibroblast 3T3 cells, similar to the case for LPA{sub 1}.

  3. Lysophosphatidic acid receptor-5 negatively regulates cellular responses in mouse fibroblast 3T3 cells

    International Nuclear Information System (INIS)

    Dong, Yan; Hirane, Miku; Araki, Mutsumi; Fukushima, Nobuyuki; Tsujiuchi, Toshifumi

    2014-01-01

    Highlights: • LPA 5 inhibits the cell growth and motile activities of 3T3 cells. • LPA 5 suppresses the cell motile activities stimulated by hydrogen peroxide in 3T3 cells. • Enhancement of LPA 5 on the cell motile activities inhibited by LPA 1 in 3T3 cells. • The expression and activation of Mmp-9 were inhibited by LPA 5 in 3T3 cells. • LPA signaling via LPA 5 acts as a negative regulator of cellular responses in 3T3 cells. - Abstract: Lysophosphatidic acid (LPA) signaling via G protein-coupled LPA receptors (LPA 1 –LPA 6 ) mediates a variety of biological functions, including cell migration. Recently, we have reported that LPA 1 inhibited the cell motile activities of mouse fibroblast 3T3 cells. In the present study, to evaluate a role of LPA 5 in cellular responses, Lpar5 knockdown (3T3-L5) cells were generated from 3T3 cells. In cell proliferation assays, LPA markedly stimulated the cell proliferation activities of 3T3-L5 cells, compared with control cells. In cell motility assays with Cell Culture Inserts, the cell motile activities of 3T3-L5 cells were significantly higher than those of control cells. The activity levels of matrix metalloproteinases (MMPs) were measured by gelatin zymography. 3T3-L5 cells stimulated the activation of Mmp-2, correlating with the expression levels of Mmp-2 gene. Moreover, to assess the co-effects of LPA 1 and LPA 5 on cell motile activities, Lpar5 knockdown (3T3a1-L5) cells were also established from Lpar1 over-expressing (3T3a1) cells. 3T3a1-L5 cells increased the cell motile activities of 3T3a1 cells, while the cell motile activities of 3T3a1 cells were significantly lower than those of control cells. These results suggest that LPA 5 may act as a negative regulator of cellular responses in mouse fibroblast 3T3 cells, similar to the case for LPA 1

  4. Clinical significance of plasma lysophosphatidic acid levels in the differential diagnosis of ovarian cancer

    Directory of Open Access Journals (Sweden)

    Yun-Jie Zhang

    2015-01-01

    Full Text Available Objective: To investigate the value of lysophosphatidic acid (LPA in the diagnosis of ovarian cancer. Materials and Methods: We first performed a hospital-based, case-control study involving 123 ovarian cancer patients and 101 benign ovarian tumor patients, and then conducted a meta-analysis with 19 case-control studies to assess the correlation between ovarian cancer and plasma LPA levels. Results: The case-control study results demonstrated that ovarian cancer patients have increased LPA and cancer antigen (CA-125 levels compared to patients with benign ovarian tumor (LPA: Ovarian cancer vs benign ovarian tumor: 5.28 ± 1.52 vs 1.82 ± 0.77 μmol/L; CA-125: Ovarian cancer vs benign ovarian tumor: 87.17 ± 45.81 vs. 14.03 ± 10.14 U/mL, which showed statistically significant differences (both P < 0.05. LPA with advanced sensitivity, specificity, positive predictive value, negative predictive value, and accuracy rate of diagnosis excelled CA-125 in the diagnosis of ovarian cancer (both P < 0.05. The areas under the receiver operating characteristic (ROC curve in the diagnosis of ovarian cancer (LPA: 0.983; CA-125: 0.910 were statistically significant compared with the reference (both P < 0.001 and the difference of the areas of ROC curve between LPA and CA-125 in the diagnosis of ovarian cancer showed statistically significant difference (P < 0.05. The meta-analysis results suggested that plasma LPA levels were higher in ovarian cancer tissues than in benign tissues (standardized mean difference (SMD =2.36, 95% confidence interval (CI: 1.61-3.11, P < 0.001 and normal tissues (SMD = 2.32, 95% CI: 1.77-2.87, P < 0.001. Conclusion: LPA shows greater value in the diagnosis of ovarian cancer compared to CA-125 and may be employed as a biological index to diagnose ovarian cancer.

  5. The overexpressed human 46-kDa mannose 6-phosphate receptor mediates endocytosis and sorting of β-glucuronidase

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    Watanabe, H.; Grubb, J.H.; Sly, W.S.

    1990-01-01

    The authors studied the function of the human small (46-kDa) mannose 6-phosphate receptor (SMPR) in transfected mouse L cells that do not express the larger insulin-like growth factor II/mannose 6-phosphate receptor. Cells overexpressing human SMPR were studied for enzyme binding to cell surface receptors, for binding to intracellular receptors in permeabilized cells, and for receptor-mediated endocytosis of recombinant human β-glucuronidase. Specific binding to human SMPR in permeabilized cells showed a pH optimum between pH 6.0 and pH 6.5. Binding was significant in the present of EDTA but was enhanced by added divalent cations. Up to 2.3% of the total functional receptor could be detected on the cell surface by enzyme binding. They present experiments showing that at very high levels of overexpression, and at pH 6.5, human SMPR mediated the endocytosis of β-glucuronidase. At pH 7.5, the rate of endocytosis was only 14% the rate seen at pH 6.5. Cells overexpressing human SMPR also showed reduced secretion of newly synthesized β-glucuronidase when compared to cells transfected with vector only, suggesting that overexpressed human SMPR can participate in sorting of newly synthesized β-glucuronidase and partially correct the sorting defect in mouse L cells that do not express the insulin-like growth factor II/mannose 6-phosphate receptor

  6. The Influence of Receptor-Mediated Interactions on Reaction-Diffusion Mechanisms of Cellular Self-organisation

    KAUST Repository

    Klika, Václav

    2011-11-10

    Understanding the mechanisms governing and regulating self-organisation in the developing embryo is a key challenge that has puzzled and fascinated scientists for decades. Since its conception in 1952 the Turing model has been a paradigm for pattern formation, motivating numerous theoretical and experimental studies, though its verification at the molecular level in biological systems has remained elusive. In this work, we consider the influence of receptor-mediated dynamics within the framework of Turing models, showing how non-diffusing species impact the conditions for the emergence of self-organisation. We illustrate our results within the framework of hair follicle pre-patterning, showing how receptor interaction structures can be constrained by the requirement for patterning, without the need for detailed knowledge of the network dynamics. Finally, in the light of our results, we discuss the ability of such systems to pattern outside the classical limits of the Turing model, and the inherent dangers involved in model reduction. © 2011 Society for Mathematical Biology.

  7. Rac1 switching at the right time and location is essential for Fcγ receptor-mediated phagosome formation.

    Science.gov (United States)

    Ikeda, Yuka; Kawai, Katsuhisa; Ikawa, Akira; Kawamoto, Kyoko; Egami, Youhei; Araki, Nobukazu

    2017-08-01

    Lamellipodia are sheet-like cell protrusions driven by actin polymerization mainly through Rac1, a GTPase molecular switch. In Fcγ receptor-mediated phagocytosis of IgG-opsonized erythrocytes (IgG-Es), Rac1 activation is required for lamellipodial extension along the surface of IgG-Es. However, the significance of Rac1 deactivation in phagosome formation is poorly understood. Our live-cell imaging and electron microscopy revealed that RAW264 macrophages expressing a constitutively active Rac1 mutant showed defects in phagocytic cup formation, while lamellipodia were formed around IgG-Es. Because activated Rac1 reduced the phosphorylation levels of myosin light chains, failure of the cup formation is probably due to inhibition of actin/myosin II contractility. Reversible photo-manipulation of the Rac1 switch in macrophages fed with IgG-Es could phenocopy two lamellipodial motilities: outward-extension and cup-constriction by Rac1 ON and OFF, respectively. In conjunction with fluorescence resonance energy transfer imaging of Rac1 activity, we provide a novel mechanistic model of phagosome formation spatiotemporally controlled by Rac1 switching within a phagocytic cup. © 2017. Published by The Company of Biologists Ltd.

  8. Ruthenium complexes with phenylterpyridine derivatives target cell membrane and trigger death receptors-mediated apoptosis in cancer cells.

    Science.gov (United States)

    Deng, Zhiqin; Gao, Pan; Yu, Lianling; Ma, Bin; You, Yuanyuan; Chan, Leung; Mei, Chaoming; Chen, Tianfeng

    2017-06-01

    Elucidation of the communication between metal complexes and cell membrane may provide useful information for rational design of metal-based anticancer drugs. Herein we synthesized a novel class of ruthenium (Ru) complexes containing phtpy derivatives (phtpy = phenylterpyridine), analyzed their structure-activity relationship and revealed their action mechanisms. The result showed that, the increase in the planarity of hydrophobic Ru complexes significantly enhanced their lipophilicity and cellular uptake. Meanwhile, the introduction of nitro group effectively improved their anticancer efficacy. Further mechanism studies revealed that, complex (2c), firstly accumulated on cell membrane and interacted with death receptors to activate extrinsic apoptosis signaling pathway. The complex was then transported into cell cytoplasm through transferrin receptor-mediated endocytosis. Most of the intracellular 2c accumulated in cell plasma, decreasing the level of cellular ROS, inducing the activation of caspase-9 and thus intensifying the apoptosis. At the same time, the residual 2c can translocate into cell nucleus to interact with DNA, induce DNA damage, activate p53 pathway and enhance apoptosis. Comparing with cisplatin, 2c possesses prolonged circulation time in blood, comparable antitumor ability and importantly, much lower toxicity in vivo. Taken together, this study uncovers the role of membrane receptors in the anticancer actions of Ru complexes, and provides fundamental information for rational design of membrane receptor targeting anticancer drugs. Copyright © 2017 Elsevier Ltd. All rights reserved.

  9. ZFAT plays critical roles in peripheral T cell homeostasis and its T cell receptor-mediated response

    International Nuclear Information System (INIS)

    Doi, Keiko; Fujimoto, Takahiro; Okamura, Tadashi; Ogawa, Masahiro; Tanaka, Yoko; Mototani, Yasumasa; Goto, Motohito; Ota, Takeharu; Matsuzaki, Hiroshi; Kuroki, Masahide; Tsunoda, Toshiyuki; Sasazuki, Takehiko; Shirasawa, Senji

    2012-01-01

    Highlights: ► We generated Cd4-Cre-mediated T cell-specific Zfat-deficient mice. ► Zfat-deficiency leads to reduction in the number of the peripheral T cells. ► Impaired T cell receptor-mediated response in Zfat-deficient peripheral T cells. ► Decreased expression of IL-7Rα, IL-2Rα and IL-2 in Zfat-deficient peripheral T cells. ► Zfat plays critical roles in peripheral T cell homeostasis. -- Abstract: ZFAT, originally identified as a candidate susceptibility gene for autoimmune thyroid disease, has been reported to be involved in apoptosis, development and primitive hematopoiesis. Zfat is highly expressed in T- and B-cells in the lymphoid tissues, however, its physiological function in the immune system remains totally unknown. Here, we generated the T cell-specific Zfat-deficient mice and demonstrated that Zfat-deficiency leads to a remarkable reduction in the number of the peripheral T cells. Intriguingly, a reduced expression of IL-7Rα and the impaired responsiveness to IL-7 for the survival were observed in the Zfat-deficient T cells. Furthermore, a severe defect in proliferation and increased apoptosis in the Zfat-deficient T cells following T cell receptor (TCR) stimulation was observed with a reduced IL-2Rα expression as well as a reduced IL-2 production. Thus, our findings reveal that Zfat is a critical regulator in peripheral T cell homeostasis and its TCR-mediated response.

  10. Cryptococcus neoformans Is Internalized by Receptor-Mediated or ‘Triggered’ Phagocytosis, Dependent on Actin Recruitment

    Science.gov (United States)

    Guerra, Caroline Rezende; Seabra, Sergio Henrique; de Souza, Wanderley; Rozental, Sonia

    2014-01-01

    Cryptococcosis by the encapsulated yeast Cryptococcus neoformans affects mostly immunocompromised individuals and is a frequent neurological complication in AIDS patients. Recent studies support the idea that intracellular survival of Cryptococcus yeast cells is important for the pathogenesis of cryptococcosis. However, the initial steps of Cryptococcus internalization by host cells remain poorly understood. Here, we investigate the mechanism of Cryptococcus neoformans phagocytosis by peritoneal macrophages using confocal and electron microscopy techniques, as well as flow cytometry quantification, evaluating the importance of fungal capsule production and of host cell cytoskeletal elements for fungal phagocytosis. Electron microscopy analyses revealed that capsular and acapsular strains of C. neoformans are internalized by macrophages via both ‘zipper’ (receptor-mediated) and ‘trigger’ (membrane ruffle-dependent) phagocytosis mechanisms. Actin filaments surrounded phagosomes of capsular and acapsular yeasts, and the actin depolymerizing drugs cytochalasin D and latrunculin B inhibited yeast internalization and actin recruitment to the phagosome area. In contrast, nocodazole and paclitaxel, inhibitors of microtubule dynamics decreased internalization but did not prevent actin recruitment to the site of phagocytosis. Our results show that different uptake mechanisms, dependent on both actin and tubulin dynamics occur during yeast internalization by macrophages, and that capsule production does not affect the mode of Cryptococcus uptake by host cells. PMID:24586631

  11. Design and development of hyaluronan-functionalized polybenzofulvene nanoparticles as CD44 receptor mediated drug delivery system

    Science.gov (United States)

    Licciardi, Mariano; Scialabba, Cinzia; Giammona, Gaetano; Paolino, Marco; Razzano, Vincenzo; Grisci, Giorgio; Giuliani, Germano; Makovec, Francesco; Cappelli, Andrea

    2017-06-01

    A tri-component polymer brush (TCPB ), composed of a polybenzofulvene copolymer bearing low molecular weight hyaluronic acid (HA) on the surface of its cylindrical brush-like backbone and oligo-PEG fractions, was employed in the preparation of 350 nm nanostructured drug delivery systems capable of delivering the anticancer drug doxorubicin. The obtained drug delivery systems were characterized on the basis of drug loading and release, dimensions and zeta potential, morphology and in vitro cell activity, and uptake on three different human cell lines, namely the bronchial epithelial 16HBE, the breast adenocarcinoma MCF-7, and the colon cancer HCT116 cells. Finally, the ability of doxorubicin-loaded TCPB nanoparticles (DOXO-TCPB) to be internalized into cancer cells by CD44 receptor mediated uptake was assessed by means of uptake studies in HCT cells. These data were supported by anti-CD44-FITC staining assay. The proposed TCPB nanostructured drug delivery systems have many potential applications in nanomedicine, including cancer targeted drug delivery.

  12. Stimulation of accumbal GABAA receptors inhibits delta2-, but not delta1-, opioid receptor-mediated dopamine efflux in the nucleus accumbens of freely moving rats.

    Science.gov (United States)

    Aono, Yuri; Kiguchi, Yuri; Watanabe, Yuriko; Waddington, John L; Saigusa, Tadashi

    2017-11-15

    The nucleus accumbens contains delta-opioid receptors that may reduce inhibitory neurotransmission. Reduction in GABA A receptor-mediated inhibition of accumbal dopamine release due to delta-opioid receptor activation should be suppressed by stimulating accumbal GABA A receptors. As delta-opioid receptors are divided into delta2- and delta1-opioid receptors, we analysed the effects of the GABA A receptor agonist muscimol on delta2- and delta1-opioid receptor-mediated accumbal dopamine efflux in freely moving rats using in vivo microdialysis. Drugs were administered intracerebrally through the dialysis probe. Doses of compounds indicate total amount administered (mol) during 25-50min infusions. The delta2-opioid receptor agonist deltorphin II (25.0nmol)- and delta1-opioid receptor agonist DPDPE (5.0nmol)-induced increases in dopamine efflux were inhibited by the delta2-opioid receptor antagonist naltriben (1.5nmol) and the delta1-opioid receptor antagonist BNTX (150.0pmol), respectively. Muscimol (250.0pmol) inhibited deltorphin II (25.0nmol)-induced dopamine efflux. The GABA A receptor antagonist bicuculline (50.0pmol), which failed to affect deltorphin II (25.0nmol)-induced dopamine efflux, counteracted the inhibitory effect of muscimol on deltorphin II-induced dopamine efflux. Neither muscimol (250.0pmol) nor bicuculline (50.0 and 500.0pmol) altered DPDPE (5.0nmol)-induced dopamine efflux. The present results show that reduction in accumbal GABA A receptor-mediated inhibition of dopaminergic activity is necessary to produce delta2-opioid receptor-induced increase in accumbal dopamine efflux. This study indicates that activation of delta2- but not delta1-opioid receptors on the cell bodies and/or terminals of accumbal GABAergic interneurons inhibits GABA release and, accordingly, decreases GABA A receptor-mediated inhibition of dopaminergic terminals, resulting in enhanced accumbal dopamine efflux. Copyright © 2017 Elsevier B.V. All rights reserved.

  13. Ah receptor mediated suppression of the antibody response in mice is primarily dependent on the Ah phenotype of lymphoid tissue

    International Nuclear Information System (INIS)

    Silkworth, J.B.; Antrim, L.A.; Sack, G.

    1986-01-01

    Halogenated aromatic hydrocarbons act through the aromatic hydrocarbon (Ah) receptor in mice to produce a series of toxic effects of the immune system. The receptor protein is a product of the Ah gene locus. Ah responsive (Ahb/Ahb) mice express a high affinity receptor in both lymphoid and nonlymphoid tissues whereas nonresponsive Ahd/Ahd mice express a poor affinity receptor. To determine the role of the Ah receptor of lymphoid tissue relative to that of nonlymphoid tissue in the induction of immune impairment, bone marrow was used to reconstitute lethally irradiated mice of the same or opposite Ah phenotype. All mice were given 3,3',4,4'-tetrachlorobiphenyl (35 and 350 mumol/kg) ip 2 days before immunization with sheep erythrocytes (SRBC). The immune response to this T dependent antigen and organ weights were determined 5 or 7 days later in normal or chimeric mice, respectively. Monoclonal Lyt 1.1 and Lyt 1.2 antibodies were used to establish the origin of the cells which repopulated the chimeric thymuses. The immune responses of both BALB/cBy (Ahb/Ahb) and the BALB/cBy X DBA/2 hybrid, CByD2F1 (Ahb/Ahd), were significantly suppressed but DBA/2 mice were unaffected. The immune responses of chimeric BALB/cBy----BALB/cBy and BALB/cBy----DBA/2 (donor----recipient) mice were also significantly suppressed and thymic atrophy was observed in both cases. The serum anti-SRBC antibody titers of DBA/2----BALB/cBy chimeras were also significantly decreased although not to the same extent as in BALB/cBy----DBA/2 mice. Chimeric DBA/2----DBA/2 mice were not affected. These results indicate that the sensitivity to Ah receptor mediated suppression of the antibody response is primarily determined by the Ah phenotype of the lymphoid tissue

  14. Biological functionalization of drug delivery carriers to bypass size restrictions of receptor-mediated endocytosis independently from receptor targeting.

    Science.gov (United States)

    Ansar, Maria; Serrano, Daniel; Papademetriou, Iason; Bhowmick, Tridib Kumar; Muro, Silvia

    2013-12-23

    Targeting of drug carriers to cell-surface receptors involved in endocytosis is commonly used for intracellular drug delivery. However, most endocytic receptors mediate uptake via clathrin or caveolar pathways associated with ≤200-nm vesicles, restricting carrier design. We recently showed that endocytosis mediated by intercellular adhesion molecule 1 (ICAM-1), which differs from clathrin- and caveolae-mediated pathways, allows uptake of nano- and microcarriers in cell culture and in vivo due to recruitment of cellular sphingomyelinases to the plasmalemma. This leads to ceramide generation at carrier binding sites and formation of actin stress-fibers, enabling engulfment and uptake of a wide size-range of carriers. Here we adapted this paradigm to enhance uptake of drug carriers targeted to receptors associated with size-restricted pathways. We coated sphingomyelinase onto model (polystyrene) submicro- and microcarriers targeted to clathrin-associated mannose-6-phosphate receptor. In endothelial cells, this provided ceramide enrichment at the cell surface and actin stress-fiber formation, modifying the uptake pathway and enhancing carrier endocytosis without affecting targeting, endosomal transport, cell-associated degradation, or cell viability. This improvement depended on the carrier size and enzyme dose, and similar results were observed for other receptors (transferrin receptor) and cell types (epithelial cells). This phenomenon also enhanced tissue accumulation of carriers after intravenous injection in mice. Hence, it is possible to maintain targeting toward a selected receptor while bypassing natural size restrictions of its associated endocytic route by functionalization of drug carriers with biological elements mimicking the ICAM-1 pathway. This strategy holds considerable promise to enhance flexibility of design of targeted drug delivery systems.

  15. The brain cytoplasmic RNA BC1 regulates dopamine D2 receptor-mediated transmission in the striatum.

    Science.gov (United States)

    Centonze, Diego; Rossi, Silvia; Napoli, Ilaria; Mercaldo, Valentina; Lacoux, Caroline; Ferrari, Francesca; Ciotti, Maria Teresa; De Chiara, Valentina; Prosperetti, Chiara; Maccarrone, Mauro; Fezza, Filomena; Calabresi, Paolo; Bernardi, Giorgio; Bagni, Claudia

    2007-08-15

    Dopamine D(2) receptor (D(2)DR)-mediated transmission in the striatum is remarkably flexible, and changes in its efficacy have been heavily implicated in a variety of physiological and pathological conditions. Although receptor-associated proteins are clearly involved in specific forms of synaptic plasticity, the molecular mechanisms regulating the sensitivity of D(2) receptors in this brain area are essentially obscure. We have studied the physiological responses of the D(2)DR stimulations in mice lacking the brain cytoplasmic RNA BC1, a small noncoding dendritically localized RNA that is supposed to play a role in mRNA translation. We show that the efficiency of D(2)-mediated transmission regulating striatal GABA synapses is under the control of BC1 RNA, through a negative influence on D(2) receptor protein level affecting the functional pool of receptors. Ablation of the BC1 gene did not result in widespread dysregulation of synaptic transmission, because the sensitivity of cannabinoid CB(1) receptors was intact in the striatum of BC1 knock-out (KO) mice despite D(2) and CB(1) receptors mediated similar electrophysiological actions. Interestingly, the fragile X mental retardation protein FMRP, one of the multiple BC1 partners, is not involved in the BC1 effects on the D(2)-mediated transmission. Because D(2)DR mRNA is apparently equally translated in the BC1-KO and wild-type mice, whereas the protein level is higher in BC1-KO mice, we suggest that BC1 RNA controls D(2)DR indirectly, probably regulating translation of molecules involved in D(2)DR turnover and/or stability.

  16. Testin, a novel binding partner of the calcium-sensing receptor, enhances receptor-mediated Rho-kinase signalling

    International Nuclear Information System (INIS)

    Magno, Aaron L.; Ingley, Evan; Brown, Suzanne J.; Conigrave, Arthur D.; Ratajczak, Thomas; Ward, Bryan K.

    2011-01-01

    Highlights: → A yeast two-hybrid screen revealed testin bound to the calcium-sensing receptor. → The second zinc finger of LIM domain 1 of testin is critical for interaction. → Testin bound to a region of the receptor tail important for cell signalling. → Testin and receptor interaction was confirmed in mammalian (HEK293) cells. → Overexpression of testin enhanced receptor-mediated Rho signalling in HEK293 cells. -- Abstract: The calcium-sensing receptor (CaR) plays an integral role in calcium homeostasis and the regulation of other cellular functions including cell proliferation and cytoskeletal organisation. The multifunctional nature of the CaR is manifested through ligand-dependent stimulation of different signalling pathways that are also regulated by partner binding proteins. Following a yeast two-hybrid library screen using the intracellular tail of the CaR as bait, we identified several novel binding partners including the focal adhesion protein, testin. Testin has not previously been shown to interact with cell surface receptors. The sites of interaction between the CaR and testin were mapped to the membrane proximal region of the receptor tail and the second zinc-finger of LIM domain 1 of testin, the integrity of which was found to be critical for the CaR-testin interaction. The CaR-testin association was confirmed in HEK293 cells by coimmunoprecipitation and confocal microscopy studies. Ectopic expression of testin in HEK293 cells stably expressing the CaR enhanced CaR-stimulated Rho activity but had no effect on CaR-stimulated ERK signalling. These results suggest an interplay between the CaR and testin in the regulation of CaR-mediated Rho signalling with possible effects on the cytoskeleton.

  17. ZFAT plays critical roles in peripheral T cell homeostasis and its T cell receptor-mediated response

    Energy Technology Data Exchange (ETDEWEB)

    Doi, Keiko [Department of Cell Biology, Faculty of Medicine, Fukuoka University, Fukuoka (Japan); Central Research Institute for Advanced Molecular Medicine, Fukuoka University, Fukuoka (Japan); Central Research Institute of Life Sciences for the Next Generation of Women Scientists, Fukuoka University, Fukuoka (Japan); Fujimoto, Takahiro [Department of Cell Biology, Faculty of Medicine, Fukuoka University, Fukuoka (Japan); Central Research Institute for Advanced Molecular Medicine, Fukuoka University, Fukuoka (Japan); Okamura, Tadashi [Division of Animal Models, Department of Infectious Diseases, Research Institute, National Center for Global Health and Medicine, Tokyo (Japan); Ogawa, Masahiro [Central Research Institute for Advanced Molecular Medicine, Fukuoka University, Fukuoka (Japan); Tanaka, Yoko [Department of Cell Biology, Faculty of Medicine, Fukuoka University, Fukuoka (Japan); Mototani, Yasumasa; Goto, Motohito [Division of Animal Models, Department of Infectious Diseases, Research Institute, National Center for Global Health and Medicine, Tokyo (Japan); Ota, Takeharu; Matsuzaki, Hiroshi [Department of Cell Biology, Faculty of Medicine, Fukuoka University, Fukuoka (Japan); Kuroki, Masahide [Central Research Institute for Advanced Molecular Medicine, Fukuoka University, Fukuoka (Japan); Tsunoda, Toshiyuki [Department of Cell Biology, Faculty of Medicine, Fukuoka University, Fukuoka (Japan); Central Research Institute for Advanced Molecular Medicine, Fukuoka University, Fukuoka (Japan); Sasazuki, Takehiko [Institute for Advanced Study, Kyushu University, Fukuoka (Japan); Shirasawa, Senji, E-mail: sshirasa@fukuoka-u.ac.jp [Department of Cell Biology, Faculty of Medicine, Fukuoka University, Fukuoka (Japan); Central Research Institute for Advanced Molecular Medicine, Fukuoka University, Fukuoka (Japan)

    2012-08-17

    Highlights: Black-Right-Pointing-Pointer We generated Cd4-Cre-mediated T cell-specific Zfat-deficient mice. Black-Right-Pointing-Pointer Zfat-deficiency leads to reduction in the number of the peripheral T cells. Black-Right-Pointing-Pointer Impaired T cell receptor-mediated response in Zfat-deficient peripheral T cells. Black-Right-Pointing-Pointer Decreased expression of IL-7R{alpha}, IL-2R{alpha} and IL-2 in Zfat-deficient peripheral T cells. Black-Right-Pointing-Pointer Zfat plays critical roles in peripheral T cell homeostasis. -- Abstract: ZFAT, originally identified as a candidate susceptibility gene for autoimmune thyroid disease, has been reported to be involved in apoptosis, development and primitive hematopoiesis. Zfat is highly expressed in T- and B-cells in the lymphoid tissues, however, its physiological function in the immune system remains totally unknown. Here, we generated the T cell-specific Zfat-deficient mice and demonstrated that Zfat-deficiency leads to a remarkable reduction in the number of the peripheral T cells. Intriguingly, a reduced expression of IL-7R{alpha} and the impaired responsiveness to IL-7 for the survival were observed in the Zfat-deficient T cells. Furthermore, a severe defect in proliferation and increased apoptosis in the Zfat-deficient T cells following T cell receptor (TCR) stimulation was observed with a reduced IL-2R{alpha} expression as well as a reduced IL-2 production. Thus, our findings reveal that Zfat is a critical regulator in peripheral T cell homeostasis and its TCR-mediated response.

  18. Role of receptor-mediated endocytosis in the antiangiogenic effects of human T lymphoblastic cell-derived microparticles.

    Science.gov (United States)

    Yang, Chun; Xiong, Wei; Qiu, Qian; Shao, Zhuo; Shao, Zuo; Hamel, David; Tahiri, Houda; Leclair, Grégoire; Lachapelle, Pierre; Chemtob, Sylvain; Hardy, Pierre

    2012-04-15

    Microparticles possess therapeutic potential regarding angiogenesis. We have demonstrated the contribution of apoptotic human CEM T lymphocyte-derived microparticles (LMPs) as inhibitors of angiogenic responses in animal models of inflammation and tumor growth. In the present study, we characterized the antivascular endothelial growth factor (VEGF) effects of LMPs on pathological angiogenesis in an animal model of oxygen-induced retinopathy and explored the role of receptor-mediated endocytosis in the effects of LMPs on human retinal endothelial cells (HRECs). LMPs dramatically inhibited cell growth of HRECs, suppressed VEGF-induced cell migration in vitro experiments, and attenuated VEGF-induced retinal vascular leakage in vivo. Intravitreal injections of fluorescently labeled LMPs revealed accumulation of LMPs in retinal tissue, with more than 60% reductions of the vascular density in retinas of rats with oxygen-induced neovascularization. LMP uptake experiments demonstrated that the interaction between LMPs and HRECs is dependent on temperature. In addition, endocytosis is partially dependent on extracellular calcium. RNAi-mediated knockdown of low-density lipoprotein receptor (LDLR) reduced the uptake of LMPs and attenuated the inhibitory effects of LMPs on VEGF-A protein expression and HRECs cell growth. Intravitreal injection of lentivirus-mediated RNA interference reduced LDLR protein expression in retina by 53% and significantly blocked the antiangiogenic effects of LMPs on pathological vascularization. In summary, the potent antiangiogenic LMPs lead to a significant reduction of pathological retinal angiogenesis through modulation of VEGF signaling, whereas LDLR-mediated endocytosis plays a partial, but pivotal, role in the uptake of LMPs in HRECs.

  19. Receptor-mediated endocytosis and intracellular trafficking of insulin and low-density lipoprotein by retinal vascular endothelial cells.

    Science.gov (United States)

    Stitt, A W; Anderson, H R; Gardiner, T A; Bailie, J R; Archer, D B

    1994-08-01

    The authors investigated the receptor-mediated endocytosis (RME) and intracellular trafficking of insulin and low-density lipoprotein (LDL) in cultured retinal vascular endothelial cells (RVECs). Low-density lipoprotein and insulin were conjugated to 10 nm colloidal gold, and these ligands were added to cultured bovine RVECs for 20 minutes at 4 degrees C. The cultures were then warmed to 37 degrees C and fixed after incubation times between 30 seconds and 1 hour. Control cells were incubated with unconjugated gold colloid at times and concentrations similar to those of the ligands. Additional control cells were exposed to several concentrations of anti-insulin receptor antibody or a saturating solution of unconjugated insulin before incubation with gold insulin. Using transmission electron microscopy, insulin gold and LDL gold were both observed at various stages of RME. Insulin-gold particles were first seen to bind to the apical plasma membrane (PM) before clustering in clathrin-coated pits and internalization in coated vesicles. Gold was later visualized in uncoated cytoplasmic vesicles, corresponding to early endosomes and multivesicular bodies (MVBs) or late endosomes. In several instances, localized regions of the limiting membrane of the MVBs appeared coated, a feature of endosomal membranes not previously described. After RME at the apical PM and passage through the endosomal system, the greater part of both insulin- and LDL-gold conjugates was seen to accumulate in large lysosome-like compartments. However, a small but significant proportion of the internalized ligands was transcytosed and released as discrete membrane-associated quanta at the basal cell surface. The uptake of LDL gold was greatly increased in highly vacuolated, late-passage RVECs. In controls, anti-insulin receptor antibody and excess unconjugated insulin caused up to 89% inhibition in gold-insulin binding and internalization. These results illustrate the internalization and intracellular

  20. Dopamine D2 receptors mediate two-odor discrimination and reversal learning in C57BL/6 mice

    Directory of Open Access Journals (Sweden)

    Grandy David K

    2004-04-01

    Full Text Available Abstract Background Dopamine modulation of neuronal signaling in the frontal cortex, midbrain, and striatum is essential for processing and integrating diverse external sensory stimuli and attaching salience to environmental cues that signal causal relationships, thereby guiding goal-directed, adaptable behaviors. At the cellular level, dopamine signaling is mediated through D1-like or D2-like receptors. Although a role for D1-like receptors in a variety of goal-directed behaviors has been identified, an explicit involvement of D2 receptors has not been clearly established. To determine whether dopamine D2 receptor-mediated signaling contributes to associative and reversal learning, we compared C57Bl/6J mice that completely lack functional dopamine D2 receptors to wild-type mice with respect to their ability to attach appropriate salience to external stimuli (stimulus discrimination and disengage from inappropriate behavioral strategies when reinforcement contingencies change (e.g. reversal learning. Results Mildly food-deprived female wild-type and dopamine D2 receptor deficient mice rapidly learned to retrieve and consume visible food reinforcers from a small plastic dish. Furthermore, both genotypes readily learned to dig through the same dish filled with sterile sand in order to locate a buried food pellet. However, the dopamine D2 receptor deficient mice required significantly more trials than wild-type mice to discriminate between two dishes, each filled with a different scented sand, and to associate one of the two odors with the presence of a reinforcer (food. In addition, the dopamine D2 receptor deficient mice repeatedly fail to alter their response patterns during reversal trials where the reinforcement rules were inverted. Conclusions Inbred C57Bl/6J mice that develop in the complete absence of functional dopamine D2 receptors are capable of olfaction but display an impaired ability to acquire odor-driven reinforcement contingencies

  1. Hypersensitivity to thromboxane receptor mediated cerebral vasomotion and CBF oscillations during acute NO-deficiency in rats.

    Directory of Open Access Journals (Sweden)

    Béla Horváth

    Full Text Available BACKGROUND: Low frequency (4-12 cpm spontaneous fluctuations of the cerebrovascular tone (vasomotion and oscillations of the cerebral blood flow (CBF have been reported in diseases associated with endothelial dysfunction. Since endothelium-derived nitric oxide (NO suppresses constitutively the release and vascular effects of thromboxane A(2 (TXA(2, NO-deficiency is often associated with activation of thromboxane receptors (TP. In the present study we hypothesized that in the absence of NO, overactivation of the TP-receptor mediated cerebrovascular signaling pathway contributes to the development of vasomotion and CBF oscillations. METHODOLOGY/PRINCIPAL FINDINGS: Effects of pharmacological modulation of TP-receptor activation and its downstream signaling pathway have been investigated on CBF oscillations (measured by laser-Doppler flowmetry in anesthetized rats and vasomotion (measured by isometric tension recording in isolated rat middle cerebral arteries, MCAs both under physiological conditions and after acute inhibition of NO synthesis. Administration of the TP-receptor agonist U-46619 (1 µg/kg i.v. to control animals failed to induce any changes of the systemic or cerebral circulatory parameters. Inhibition of the NO synthesis by nitro-L-arginine methyl ester (L-NAME, 100 mg/kg i.v. resulted in increased mean arterial blood pressure and a decreased CBF accompanied by appearance of CBF-oscillations with a dominant frequency of 148±2 mHz. U-46619 significantly augmented the CBF-oscillations induced by L-NAME while inhibition of endogenous TXA(2 synthesis by ozagrel (10 mg/kg i.v. attenuated it. In isolated MCAs U-46619 in a concentration of 100 nM, which induced weak and stable contraction under physiological conditions, evoked sustained vasomotion in the absence of NO, which effect could be completely reversed by inhibition of Rho-kinase by 10 µM Y-27632. CONCLUSION/SIGNIFICANCE: These results suggest that hypersensitivity of the TP

  2. Involvement of aberrant DNA methylation on reduced expression of lysophosphatidic acid receptor-1 gene in rat tumor cell lines

    International Nuclear Information System (INIS)

    Tsujiuchi, Toshifumi; Shimizu, Kyoko; Onishi, Mariko; Sugata, Eriko; Fujii, Hiromasa; Mori, Toshio; Honoki, Kanya; Fukushima, Nobuyuki

    2006-01-01

    Lysophosphatidic acid (LPA) is a bioactive phospholipid that stimulates cell proliferation, migration, and protects cells from apoptosis. It interacts with specific G protein-coupled transmembrane receptors. Recently, it has been reported that alterations of LPA receptor expression might be important in the malignant transformation of tumor cells. Therefore, to assess an involvement of DNA methylation in reduced expression of the LPA receptor-1 (lpa1) gene, we investigated the expression of the lpa1 gene and its DNA methylation patterns in rat tumor cell lines. Both rat brain-derived neuroblastoma B103 and liver-derived hepatoma RH7777 cells used in this study indicated no expression of lpa1. For the analysis of methylation status, bisulfite sequencing was performed with B103 and RH7777 cells, comparing with other lpa1 expressed cells and normal tissues of brain and liver. The lpa1 expressed cells and tissues were all unmethylated in this region of lpa1. In contrast, both B103 and RH7777 cells were highly methylated, correlating with reduced expression of the lpa1. Treatment with 5-aza 2'-deoxycytidine induced expression of lpa1 gene in B103 and RH7777 cells after 24 h. In RH7777 cells treated with 5-aza 2'-deoxycytidine, stress fiber formation was also observed in response to LPA in RH7777 cells, but not in untreated RH7777 cells. These results suggest that aberrant DNA methylation of the lpa1 gene may be involved in its reduced expression in rat tumor cells

  3. Clinical significance and correlation of the level change of plasma lysophosphatidic acid in patients before and during radiotherapy

    International Nuclear Information System (INIS)

    Yu Liang; Li Hong; Zhu Shengjie

    2011-01-01

    Objective: To investigate the effects of radiotherapy on the level of lysophosphatidic acid (LPA) in different patients. Methods: Three groups of patients (metastatic brain tumor group, non-brain tumor group and primary brain tumor group) were given external irradiation (by linear accelerator). LPA level, white blood cell count and platelet count in the blood plasma were evaluated pre-irradiation and after irradiation with 40 Gy and 60 Gy respectively. Results: The LPA level decreased gradually as irradiation doses increase in metastatic brain tumor group non-brain tumor group (after irradiation with 60 Gy 0.05), and neoplasm volume changed little. White blood cell count and platelet count gradually decreased with the increasing irradiation doses in metastatic brain tumor group and primary brain tumor group, but there was no significant correlation to LPA level. However, there was a negative correlation to LPA level in non-brain tumor group (r=-0.285 and r=-0.237, both P<0.05 ). Conclusions: There is a negative correlation between radiotherapy dose and LPA level in metastatic brain tumor patients and non-brain tumor patients. LPA level could be used as a predictor of the effect of the radiotherapy in tumor treatment. (authors)

  4. Dual Action of Lysophosphatidate-Functionalised Titanium: Interactions with Human (MG63 Osteoblasts and Methicillin Resistant Staphylococcus aureus.

    Directory of Open Access Journals (Sweden)

    Mette Elena Skindersoe

    Full Text Available Titanium (Ti is a widely used material for surgical implants; total joint replacements (TJRs, screws and plates for fixing bones and dental implants are forged from Ti. Whilst Ti integrates well into host tissue approximately 10% of TJRs will fail in the lifetime of the patient through a process known as aseptic loosening. These failures necessitate revision arthroplasties which are more complicated and costly than the initial procedure. Finding ways of enhancing early (osseointegration of TJRs is therefore highly desirable and continues to represent a research priority in current biomaterial design. One way of realising improvements in implant quality is to coat the Ti surface with small biological agents known to support human osteoblast formation and maturation at Ti surfaces. Lysophosphatidic acid (LPA and certain LPA analogues offer potential solutions as Ti coatings in reducing aseptic loosening. Herein we present evidence for the successful bio-functionalisation of Ti using LPA. This modified Ti surface heightened the maturation of human osteoblasts, as supported by increased expression of alkaline phosphatase. These functionalised surfaces also deterred the attachment and growth of Staphylococcus aureus, a bacterium often associated with implant failures through sepsis. Collectively we provide evidence for the fabrication of a dual-action Ti surface finish, a highly desirable feature towards the development of next-generation implantable devices.

  5. Lysophosphatidic acid induces reactive oxygen species generation by activating protein kinase C in PC-3 human prostate cancer cells

    International Nuclear Information System (INIS)

    Lin, Chu-Cheng; Lin, Chuan-En; Lin, Yueh-Chien; Ju, Tsai-Kai; Huang, Yuan-Li; Lee, Ming-Shyue; Chen, Jiun-Hong; Lee, Hsinyu

    2013-01-01

    Highlights: •LPA induces ROS generation through LPA 1 and LPA 3 . •LPA induces ROS generation by activating PLC. •PKCζ mediates LPA-induced ROS generation. -- Abstract: Prostate cancer is one of the most frequently diagnosed cancers in males, and PC-3 is a cell model popularly used for investigating the behavior of late stage prostate cancer. Lysophosphatidic acid (LPA) is a lysophospholipid that mediates multiple behaviors in cancer cells, such as proliferation, migration and adhesion. We have previously demonstrated that LPA enhances vascular endothelial growth factor (VEGF)-C expression in PC-3 cells by activating the generation of reactive oxygen species (ROS), which is known to be an important mediator in cancer progression. Using flow cytometry, we showed that LPA triggers ROS generation within 10 min and that the generated ROS can be suppressed by pretreatment with the NADPH oxidase (Nox) inhibitor diphenylene iodonium. In addition, transfection with LPA 1 and LPA 3 siRNA efficiently blocked LPA-induced ROS production, suggesting that both receptors are involved in this pathway. Using specific inhibitors and siRNA, phospholipase C (PLC) and protein kinase C (PKC) were also suggested to participate in LPA-induced ROS generation. Overall, we demonstrated that LPA induces ROS generation in PC-3 prostate cancer cells and this is mediated through the PLC/PKC/Nox pathway

  6. Endogenous lysophosphatidic acid participates in vascularisation and decidualisation at the maternal-fetal interface in the rat.

    Science.gov (United States)

    Sordelli, Micaela S; Beltrame, Jimena S; Zotta, Elsa; Gomez, Natalia; Dmytrenko, Ganna; Sales, María Elena; Blois, Sandra M; Davio, Carlos; Martinez, Silvina Perez; Franchi, Ana M; Ribeiro, María L

    2017-10-01

    Lysophosphatidic acid (LPA) affects several female reproductive functions through G-protein-coupled receptors. LPA contributes to embryo implantation via the lysophospholipid LPA 3 receptor. In the present study we investigated the participation of endogenous LPA signalling through the LPA 3 receptor in vascularisation and decidualisation, two crucial events at the maternal-fetal interface. Pregnant rats were treated with diacylglycerol pyrophosphate (DGPP), a highly selective antagonist of LPA 3 receptors, on Day 5 of gestation. Pregnant rats received intrauterine (i.u.) injections of single doses of DGPP (0.1mgkg -1 ) in a total volume of 2μL in the left horn (treated horn) in the morning of GD5. DGPP treatment produced aberrant embryo spacing and increased embryo resorption. The LPA 3 receptor antagonist decreased the cross-sectional length of the uterine and arcuate arteries and induced histological anomalies in the decidua and placentas. Marked haemorrhagic processes, infiltration of immune cells and tissue disorganisation were observed in decidual and placental tissues from sites of resorption. The mRNA expression of three vascularisation markers, namely interleukin 10 (Il10), vascular endothelial growth factor (Vegfa) and vascular endothelial growth factor receptor 1 (Vegfr1), was reduced at sites of resorption from Day 8. The results show that the disruption of endogenous LPA signalling by blocking the LPA 3 receptor modified the development of uterine vessels with consequences in the formation of the decidua and placenta and in the growth of embryos.

  7. The effects of aspirin on platelet function and lysophosphatidic acids depend on plasma concentrations of EPA and DHA.

    Science.gov (United States)

    Block, Robert C; Abdolahi, Amir; Tu, Xin; Georas, Steve N; Brenna, J Thomas; Phipps, Richard P; Lawrence, Peter; Mousa, Shaker A

    2015-05-01

    Aspirin's prevention of cardiovascular disease (CVD) events in individuals with type 2 diabetes mellitus is controversial. Eicosapentaenoic acid (EPA) and docosahexaenoic acid (DHA) and aspirin all affect the cyclooxygenase enzyme. The relationship between plasma EPA and DHA and aspirin's effects has not been determined. Thirty adults with type 2 diabetes mellitus ingested aspirin (81 mg/day) for 7 days, then EPA+DHA (2.6g/day) for 28 days, then both for another 7 days. Lysophosphatidic acid (LPA) species and more classic platelet function outcomes were determined. Plasma concentrations of total EPA+DHA were associated with 7-day aspirin reduction effects on these outcomes in a "V"-shaped manner for all 11 LPA species and ADP-induced platelet aggregation. This EPA+DHA concentration was quite consistent for each of the LPA species and ADP. These results support aspirin effects on lysolipid metabolism and platelet aggregation depending on plasma EPA+DHA concentrations in individuals with a disturbed lipid milieu. Copyright © 2014 Elsevier Ltd. All rights reserved.

  8. Co-ordinate regulation of growth factor receptors and lipid phosphate phosphatase-1 controls cell activation by exogenous lysophosphatidate.

    Science.gov (United States)

    Pilquil, C; Ling, Z C; Singh, I; Buri, K; Zhang, Q X; Brindley, D N

    2001-11-01

    The serum-derived lipid growth factors, lysophosphatidate (LPA) and sphingosine 1-phosphate (S1P), activate cells selectively through different members of a family of endothelial differentiation gene (EDG) receptors. Activation of EDG receptors by LPA and S1P provides a variety of signalling cascades depending upon the G-protein coupling of the different EDG receptors. This leads to chemotactic and mitogenic responses, which are important in wound healing. For example, LPA stimulates fibroblast division and S1P stimulates the chemotaxis and division of endothelial cells leading to angiogenesis. Counteracting these effects of LPA and S1P, are the actions of lipid phosphate phosphatases (LPP, or phosphatidate phosphohydrolases, Type 2). The isoform LPP-1 is expressed in the plasma membrane with its active site outside the cell. This enzyme is responsible for 'ecto-phosphatase' activity leading to the degradation of exogenous lipid phosphate mediators, particularly LPA. Expression of LPP-1 decreases cell activation by exogenous LPA. The mechanism for this is controversial and several mechanisms have been proposed. Evidence will be presented that the LPPs cross-talk with EDG and other growth factor receptors, thus, regulating the responses of the cells to lipid phosphate mediators of signal transduction.

  9. Lysophosphatidic acid induces reactive oxygen species generation by activating protein kinase C in PC-3 human prostate cancer cells

    Energy Technology Data Exchange (ETDEWEB)

    Lin, Chu-Cheng; Lin, Chuan-En; Lin, Yueh-Chien [Institute of Zoology, College of Life Science, National Taiwan University, Taipei, Taiwan, ROC (China); Ju, Tsai-Kai [Instrumentation Center, National Taiwan University, Taipei, Taiwan, ROC (China); Technology Commons, College of Life Science, National Taiwan University, Taipei, Taiwan, ROC (China); Huang, Yuan-Li [Department of Biotechnology, Asia University, Taichung, Taiwan, ROC (China); Lee, Ming-Shyue [Institute of Biochemistry and Molecular Biology, College of Medicine, National Taiwan University, Taipei, Taiwan, ROC (China); Chen, Jiun-Hong [Institute of Zoology, College of Life Science, National Taiwan University, Taipei, Taiwan, ROC (China); Department of Life Science, College of Life Science, National Taiwan University, Taipei, Taiwan, ROC (China); Lee, Hsinyu, E-mail: hsinyu@ntu.edu.tw [Institute of Zoology, College of Life Science, National Taiwan University, Taipei, Taiwan, ROC (China); Department of Life Science, College of Life Science, National Taiwan University, Taipei, Taiwan, ROC (China); Center for Biotechnology, National Taiwan University, Taipei, Taiwan, ROC (China); Research Center for Developmental Biology and Regenerative Medicine, National Taiwan University, Taipei, Taiwan, ROC (China)

    2013-11-01

    Highlights: •LPA induces ROS generation through LPA{sub 1} and LPA{sub 3}. •LPA induces ROS generation by activating PLC. •PKCζ mediates LPA-induced ROS generation. -- Abstract: Prostate cancer is one of the most frequently diagnosed cancers in males, and PC-3 is a cell model popularly used for investigating the behavior of late stage prostate cancer. Lysophosphatidic acid (LPA) is a lysophospholipid that mediates multiple behaviors in cancer cells, such as proliferation, migration and adhesion. We have previously demonstrated that LPA enhances vascular endothelial growth factor (VEGF)-C expression in PC-3 cells by activating the generation of reactive oxygen species (ROS), which is known to be an important mediator in cancer progression. Using flow cytometry, we showed that LPA triggers ROS generation within 10 min and that the generated ROS can be suppressed by pretreatment with the NADPH oxidase (Nox) inhibitor diphenylene iodonium. In addition, transfection with LPA{sub 1} and LPA{sub 3} siRNA efficiently blocked LPA-induced ROS production, suggesting that both receptors are involved in this pathway. Using specific inhibitors and siRNA, phospholipase C (PLC) and protein kinase C (PKC) were also suggested to participate in LPA-induced ROS generation. Overall, we demonstrated that LPA induces ROS generation in PC-3 prostate cancer cells and this is mediated through the PLC/PKC/Nox pathway.

  10. Mice Deficient in lysophosphatidic acid acyltransferase delta (Lpaatδ)/acylglycerophosphate acyltransferase 4 (Agpat4) Have Impaired Learning and Memory.

    Science.gov (United States)

    Bradley, Ryan M; Mardian, Emily B; Bloemberg, Darin; Aristizabal Henao, Juan J; Mitchell, Andrew S; Marvyn, Phillip M; Moes, Katherine A; Stark, Ken D; Quadrilatero, Joe; Duncan, Robin E

    2017-11-15

    We previously characterized LPAATδ/AGPAT4 as a mitochondrial lysophosphatidic acid acyltransferase that regulates brain levels of phosphatidylcholine (PC), phosphatidylethanolamine (PE), and phosphatidylinositol (PI). Here, we report that Lpaat δ -/- mice display impaired spatial learning and memory compared to wild-type littermates in the Morris water maze and our investigation of potential mechanisms associated with brain phospholipid changes. Marker protein immunoblotting suggested that the relative brain content of neurons, glia, and oligodendrocytes was unchanged. Relative abundance of the important brain fatty acid docosahexaenoic acid was also unchanged in phosphatidylserine, phosphatidylglycerol, and cardiolipin, in agreement with prior data on PC, PE and PI. In phosphatidic acid, it was increased. Specific decreases in ethanolamine-containing phospholipids were detected in mitochondrial lipids, but the function of brain mitochondria in Lpaat δ -/- mice was unchanged. Importantly, we found that Lpaat δ -/- mice have a significantly and drastically lower brain content of the N -methyl-d-asparate (NMDA) receptor subunits NR1, NR2A, and NR2B, as well as the α-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid (AMPA) receptor subunit GluR1, compared to wild-type mice. However, general dysregulation of PI-mediated signaling is not likely responsible, since phospho-AKT and phospho-mTOR pathway regulation was unaffected. Our findings indicate that Lpaat δ deficiency causes deficits in learning and memory associated with reduced NMDA and AMPA receptors. Copyright © 2017 American Society for Microbiology.

  11. Alpha 1-adrenergic receptor-mediated phosphoinositide hydrolysis and prostaglandin E2 formation in Madin-Darby canine kidney cells. Possible parallel activation of phospholipase C and phospholipase A2

    International Nuclear Information System (INIS)

    Slivka, S.R.; Insel, P.A.

    1987-01-01

    alpha 1-Adrenergic receptors mediate two effects on phospholipid metabolism in Madin-Darby canine kidney (MDCK-D1) cells: hydrolysis of phosphoinositides and arachidonic acid release with generation of prostaglandin E2 (PGE2). The similarity in concentration dependence for the agonist (-)-epinephrine in eliciting these two responses implies that they are mediated by a single population of alpha 1-adrenergic receptors. However, we find that the kinetics of the two responses are quite different, PGE2 production occurring more rapidly and transiently than the hydrolysis of phosphoinositides. The antibiotic neomycin selectively decreases alpha 1-receptor-mediated phosphatidylinositol 4,5-bisphosphate hydrolysis without decreasing alpha 1-receptor-mediated arachidonic acid release and PGE2 generation. In addition, receptor-mediated inositol trisphosphate formation is independent of extracellular calcium, whereas release of labeled arachidonic acid is largely calcium-dependent. Moreover, based on studies obtained with labeled arachidonic acid, receptor-mediated generation of arachidonic acid cannot be accounted for by breakdown of phosphatidylinositol monophosphate, phosphatidylinositol bisphosphate, or phosphatidic acid. Further studies indicate that epinephrine produces changes in formation or turnover of several classes of membrane phospholipids in MDCK cells. We conclude that alpha 1-adrenergic receptors in MDCK cells appear to regulate phospholipid metabolism by the parallel activation of phospholipase C and phospholipase A2. This parallel activation of phospholipases contrasts with models described in other systems which imply sequential activation of phospholipase C and diacylglycerol lipase or phospholipase A2

  12. Receptor-mediated endocytosis of polypeptide hormones is a regulated process: inhibition of [125I]iodoinsulin internalization in hypoinsulinemic diabetes of rat and man

    International Nuclear Information System (INIS)

    Carpentier, J.L.; Robert, A.; Grunberger, G.; van Obberghen, E.; Freychet, P.; Orci, L.; Gorden, P.

    1986-01-01

    Much data suggest that receptor-mediated endocytosis is regulated in states of hormone excess. Thus, in hyperinsulinemic states there is an accelerated loss of cell surface insulin receptors. In the present experiments we addressed this question in hypoinsulinemic states, in which insulin binding to cell surface receptors is generally increased. In hepatocytes obtained from hypoinsulinemic streptozotocin-induced diabetic rats, [ 125 I]iodoglucagon internalization was increased, while at the same time [ 125 I]iodoinsulin internalization was decreased. The defect in [ 125 I]iodoinsulin internalization was corrected by insulin treatment of the animal. In peripheral blood monocytes from patients with type I insulinopenic diabetes, internalization of [ 125 I]iodoinsulin was impaired; this defect was not present in insulin-treated patients. These data in the hypoinsulinemic rat and human diabetes suggest that receptor-mediated endocytosis is regulated in states of insulin deficiency as well as insulin excess. Delayed or reduced internalization of the insulin-receptor complex could amplify the muted signal caused by deficient hormone secretion

  13. Nitric oxide inhibits the bradykinin B2 receptor-mediated adrenomedullary catecholamine release but has no effect on adrenal blood flow response in vivo.

    Science.gov (United States)

    Bouallegue, Ali; Yamaguchi, Nobuharu

    2005-06-01

    The role of nitric oxide (NO) in bradykinin (BK)-induced adrenal catecholamine secretion still remains obscure. The present study was to investigate whether an inhibition of NO synthase with N(omega)-nitro-L-arginine methyl ester (L-NAME) would modulate BK-induced adrenal catecholamine secretion (ACS) and adrenal vasodilating response (AVR) in anesthetized dogs. Plasma catecholamine concentrations were determined with an HPLC coupled with an electrochemical detector. All drugs were locally administered to the left adrenal gland via intra-arterial infusion. BK dose-dependently increased both ACS and AVR. Hoe-140, a selective B(2) antagonist, significantly blocked the BK-induced increases in both ACS and AVR. In the presence of L-NAME, the BK-induced ACS was significantly enhanced, while the simultaneous AVR remained unaffected. These results suggest that the both BK-induced ACS and AVR are primarily mediated by B(2) receptors in the canine adrenal gland. Our results also suggest that the enhanced ACS in response to BK in the presence of L-NAME may have resulted from a specific inhibition of NO formation in the adrenal gland. It is concluded that the BK-induced NO may play an inhibitory role in the B(2)-receptor-mediated mechanisms regulating ACS, while it may not be implicated in the B(2)-receptor-mediated AVR under in vivo conditions.

  14. Differential modulation of expression of nuclear receptor mediated genes by tris(2-butoxyethyl) phosphate (TBOEP) on early life stages of zebrafish (Danio rerio)

    Energy Technology Data Exchange (ETDEWEB)

    Ma, Zhiyuan, E-mail: zhiyuan_nju@163.com [State Key Laboratory of Pollution Control and Resource Reuse, School of the Environment, Nanjing University, Nanjing, Jiangsu 210023 (China); Yu, Yijun, E-mail: yjun.yu@gmail.com [State Key Laboratory of Pollution Control and Resource Reuse, School of the Environment, Nanjing University, Nanjing, Jiangsu 210023 (China); Tang, Song [School of Environment and Sustainability, University of Saskatchewan, Saskatoon, SK S7N 5B3 (Canada); Liu, Hongling, E-mail: hlliu@nju.edu.cn [State Key Laboratory of Pollution Control and Resource Reuse, School of the Environment, Nanjing University, Nanjing, Jiangsu 210023 (China); Su, Guanyong; Xie, Yuwei [State Key Laboratory of Pollution Control and Resource Reuse, School of the Environment, Nanjing University, Nanjing, Jiangsu 210023 (China); Giesy, John P. [State Key Laboratory of Pollution Control and Resource Reuse, School of the Environment, Nanjing University, Nanjing, Jiangsu 210023 (China); Toxicology Centre, University of Saskatchewan, Saskatoon, SK S7N 5B3 (Canada); Department of Veterinary Biomedical Sciences, University of Saskatchewan, Saskatoon, SK S7N 5B3 (Canada); Department of Biology and Chemistry, City University of Hong Kong, Kowloon, Hong Kong Special Administrative Region (Hong Kong); Hecker, Markus [School of Environment and Sustainability, University of Saskatchewan, Saskatoon, SK S7N 5B3 (Canada); Toxicology Centre, University of Saskatchewan, Saskatoon, SK S7N 5B3 (Canada); Yu, Hongxia [State Key Laboratory of Pollution Control and Resource Reuse, School of the Environment, Nanjing University, Nanjing, Jiangsu 210023 (China)

    2015-12-15

    Highlights: • Effects of TBOEP on expression of genes of several nuclear hormone receptors and their relationship with adverse effect pathways in zebrafish. • TBOEP was neither an agonist nor antagonist of AR or AhR as determined by use of in vitro mammalian cell-based receptor transactivation assays. • Modulation of ER- and MR-dependent pathways allowed for development of feasible receptor-mediated, critical mechanisms of toxic action. - Abstract: As one substitute for phased-out brominated flame retardants (BFRs), tris(2-butoxyethyl) phosphate (TBOEP) is frequently detected in aquatic organisms. However, knowledge about endocrine disrupting mechanisms associated with nuclear receptors caused by TBOEP remained restricted to results from in vitro studies with mammalian cells. In the study, results of which are presented here, embryos/larvae of zebrafish (Danio rerio) were exposed to 0.02, 0.1 or 0.5 μM TBOEP to investigate expression of genes under control of several nuclear hormone receptors (estrogen receptors (ERs), androgen receptor (AR), thyroid hormone receptor alpha (TRα), mineralocorticoid receptor (MR), glucocorticoid receptor (GR), aryl hydrocarbon (AhR), peroxisome proliferator-activated receptor alpha (PPARα), and pregnane × receptor (P × R)) pathways at 120 hpf. Exposure to 0.5 μM TBOEP significantly (p < 0.05, one-way analysis of variance) up-regulated expression of estrogen receptors (ERs, er1, er2a, and er2b) genes and ER-associated genes (vtg4, vtg5, pgr, ncor, and ncoa3), indicating TBOEP modulates the ER pathway. In contrast, expression of most genes (mr, 11βhsd, ube2i,and adrb2b) associated with the mineralocorticoid receptor (MR) pathway were significantly down-regulated. Furthermore, in vitro mammalian cell-based (MDA-kb2 and H4IIE-luc) receptor transactivation assays, were also conducted to investigate possible agonistic or antagonistic effects on AR- and AhR-mediated pathways. In mammalian cells, none of these pathways were

  15. Role of lysophosphatidic acid receptor LPA2 in the development of allergic airway inflammation in a murine model of asthma

    Directory of Open Access Journals (Sweden)

    Chun Jerold

    2009-11-01

    Full Text Available Abstract Background Lysophosphatidic acid (LPA plays a critical role in airway inflammation through G protein-coupled LPA receptors (LPA1-3. We have demonstrated that LPA induced cytokine and lipid mediator release in human bronchial epithelial cells. Here we provide evidence for the role of LPA and LPA receptors in Th2-dominant airway inflammation. Methods Wild type, LPA1 heterozygous knockout mice (LPA1+/-, and LPA2 heterozygous knockout mice (LPA2+/- were sensitized with inactivated Schistosoma mansoni eggs and local antigenic challenge with Schistosoma mansoni soluble egg Ag (SEA in the lungs. Bronchoalveolar larvage (BAL fluids and lung tissues were collected for analysis of inflammatory responses. Further, tracheal epithelial cells were isolated and challenged with LPA. Results BAL fluids from Schistosoma mansoni egg-sensitized and challenged wild type mice (4 days of challenge showed increase of LPA level (~2.8 fold, compared to control mice. LPA2+/- mice, but not LPA1+/- mice, exposed to Schistosoma mansoni egg revealed significantly reduced cell numbers and eosinophils in BAL fluids, compared to challenged wild type mice. Both LPA2+/- and LPA1+/- mice showed decreases in bronchial goblet cells. LPA2+/- mice, but not LPA1+/- mice showed the decreases in prostaglandin E2 (PGE2 and LPA levels in BAL fluids after SEA challenge. The PGE2 production by LPA was reduced in isolated tracheal epithelial cells from LPA2+/- mice. These results suggest that LPA and LPA receptors are involved in Schistosoma mansoni egg-mediated inflammation and further studies are proposed to understand the role of LPA and LPA receptors in the inflammatory process.

  16. CD14 is a key mediator of both lysophosphatidic acid and lipopolysaccharide induction of foam cell formation.

    Science.gov (United States)

    An, Dong; Hao, Feng; Zhang, Fuqiang; Kong, Wei; Chun, Jerold; Xu, Xuemin; Cui, Mei-Zhen

    2017-09-01

    Macrophage uptake of oxidized low-density lipoprotein (oxLDL) plays an important role in foam cell formation and the pathogenesis of atherosclerosis. We report here that lysophosphatidic acid (LPA) enhances lipopolysaccharide (LPS)-induced oxLDL uptake in macrophages. Our data revealed that both LPA and LPS highly induce the CD14 expression at messenger RNA and protein levels in macrophages. The role of CD14, one component of the LPS receptor cluster, in LPA-induced biological functions has been unknown. We took several steps to examine the role of CD14 in LPA signaling pathways. Knockdown of CD14 expression nearly completely blocked LPA/LPS-induced oxLDL uptake in macrophages, demonstrating for the first time that CD14 is a key mediator responsible for both LPA- and LPS-induced oxLDL uptake/foam cell formation. To determine the molecular mechanism mediating CD14 function, we demonstrated that both LPA and LPS significantly induce the expression of scavenger receptor class A type I (SR-AI), which has been implicated in lipid uptake process, and depletion of CD14 levels blocked LPA/LPS-induced SR-AI expression. We further showed that the SR-AI-specific antibody, which quenches SR-AI function, blocked LPA- and LPS-induced foam cell formation. Thus, SR-AI is the downstream mediator of CD14 in regulating LPA-, LPS-, and LPA/LPS-induced foam cell formation. Taken together, our results provide the first experimental evidence that CD14 is a novel connecting molecule linking both LPA and LPS pathways and is a key mediator responsible for LPA/LPS-induced foam cell formation. The LPA/LPS-CD14-SR-AI nexus might be the new convergent pathway, contributing to the worsening of atherosclerosis. © 2017 by The American Society for Biochemistry and Molecular Biology, Inc.

  17. Spinal cord compression injury in lysophosphatidic acid 1 receptor-null mice promotes maladaptive pronociceptive descending control.

    Science.gov (United States)

    Suardíaz, M; Galan-Arriero, I; Avila-Martin, G; Estivill-Torrús, G; de Fonseca, F R; Chun, J; Gómez-Soriano, J; Bravo-Esteban, E; Taylor, J

    2016-02-01

    Although activation of the lysophosphatidic acid receptor 1 (LPA1) is known to mediate pronociceptive effects in peripheral pain models, the role of this receptor in the modulation of spinal nociception following spinal cord injury (SCI) is unknown. In this study, LPA1 regulation of spinal excitability mediated by supraspinal descending antinociceptive control systems was assessed following SCI in both wild-type (WT) and maLPA1-null receptor mice. The effect of a T8 spinal compression in WT and maLPA1-null mice was assessed up to 1 month after SCI using histological, immunohistochemical and behavioural techniques analysis including electrophysiological recording of noxious toes-Tibialis Anterior (TA) stimulus-response reflex activity. The effect of a T3 paraspinal transcutaneous electrical conditioning stimulus on TA noxious reflex temporal summation was also assessed. Histological analysis demonstrated greater dorsolateral funiculus damage after SCI in maLPA1-null mice, without a change in the stimulus-response function of the TA noxious reflex when compared to WT mice. While T3 conditioning stimulation in the WT group inhibited noxious TA reflex temporal summation after SCI, this stimulus strongly excited TA reflex temporal summation in maLPA1-null mice. The functional switch from descending inhibition to maladaptive facilitation of central excitability of spinal nociception demonstrated in maLPA1-null mice after SCI was unrelated to a general change in reflex activity. These data suggest that the LPA1 receptor is necessary for inhibition of temporal summation of noxious reflex activity, partly mediated via long-tract descending modulatory systems acting at the spinal level. © 2015 European Pain Federation - EFIC®

  18. Permanent Distal Occlusion of Middle Cerebral Artery in Rat Causes Local Increased ETB, 5-HT1B and AT1 Receptor-Mediated Contractility Downstream of Occlusion

    DEFF Research Database (Denmark)

    Rasmussen, Marianne N P; Hornbak, Malene; Larsen, Stine S

    2013-01-01

    Background/Aims: In response to experimental stroke, a characteristic functional and expressional upregulation of contractile G-protein-coupled receptors has been uncovered in the affected cerebral vasculature; however, the mechanism initiating this phenomenon remains unknown. Methods: Using...... a model of permanent distal occlusion of rat middle cerebral arteries, we investigated whether there was a regional difference in receptor-mediated contractility of segments located upstream and downstream of the occlusion site. The contractile response to endothelin, angiotensin and 5-hydroxytryptamine...... receptor stimulation was studied by sensitive wire myograph. Results: Only downstream segments exhibited an augmented contractile response to stimulation with each of the three ligands, with the response towards sarafotoxin 6c being especially augmented compared to sham, upstream and contralateral controls...

  19. Lysophosphatidic acid signaling via LPA_1 and LPA_3 regulates cellular functions during tumor progression in pancreatic cancer cells

    International Nuclear Information System (INIS)

    Fukushima, Kaori; Takahashi, Kaede; Yamasaki, Eri; Onishi, Yuka; Fukushima, Nobuyuki; Honoki, Kanya; Tsujiuchi, Toshifumi

    2017-01-01

    Lysophosphatidic acid (LPA) signaling via G protein-coupled LPA receptors exhibits a variety of biological effects, such as cell proliferation, motility and differentiation. The aim of this study was to evaluate the roles of LPA_1 and LPA_3 in cellular functions during tumor progression in pancreatic cancer cells. LPA_1 and LPA_3 knockdown cells were generated from PANC-1 cells. The cell motile and invasive activities of PANC-1 cells were inhibited by LPA_1 and LPA_3 knockdown. In gelatin zymography, LPA_1 and LPA_3 knockdown cells indicated the low activation of matrix metalloproteinase-2 (MMP-2) in the presence of LPA. Next, to assess whether LPA_1 and LPA_3 regulate cellular functions induced by anticancer drug, PANC-1 cells were treated with cisplatin (CDDP) for approximately 6 months. The cell motile and invasive activities of long-term CDDP treated cells were markedly higher than those of PANC-1 cells, correlating with the expression levels of LPAR1 and LPAR3 genes. In soft agar assay, the long-term CDDP treated cells formed markedly large sized colonies. In addition, the cell motile and invasive activities enhanced by CDDP were significantly suppressed by LPA_1 and LPA_3 knockdown as well as colony formation. These results suggest that LPA signaling via LPA_1 and LPA_3 play an important role in the regulation of cellular functions during tumor progression in PANC-1 cells. - Highlights: • The cell motile and invasive activities of PANC-1 cells were stimulated by LPA_1 and LPA_3. • LPA_1 and LPA_3 enhanced MMP-2 activation in PANC-1 cells. • The expressions of LPAR1 and LPAR3 genes were elevated in PANC-1 cells treated with cisplatin. • The cell motile and invasive activities of PANC-1 cells treated with cisplatin were suppressed by LPA_1 and LPA_3 knockdown. • LPA_1 and LPA_3 are involved in the regulation of cellular functions during tumor progression in PANC-1 cells.

  20. Derivatives of Dictyostelium differentiation-inducing factors inhibit lysophosphatidic acid–stimulated migration of murine osteosarcoma LM8 cells

    Energy Technology Data Exchange (ETDEWEB)

    Kubohara, Yuzuru, E-mail: ykuboha@juntendo.ac.jp [Department of Molecular and Cellular Biology, Institute for Molecular and Cellular Regulation (IMCR), Gunma University, Maebashi 371-8512 (Japan); Department of Health Science, Juntendo University Graduate School of Health and Sports Science, Inzai 270-1695 (Japan); Komachi, Mayumi [Department of Molecular and Cellular Biology, Institute for Molecular and Cellular Regulation (IMCR), Gunma University, Maebashi 371-8512 (Japan); Department of Radiation Oncology, Gunma University Graduate School of Medicine, Maebashi 371-8511 (Japan); Homma, Yoshimi [Department of Biomolecular Science, Institute of Biomedical Sciences, Fukushima Medical University School of Medicine, Fukushima 960-1295 (Japan); Kikuchi, Haruhisa; Oshima, Yoshiteru [Laboratory of Natural Product Chemistry, Tohoku University Graduate School of Pharmaceutical Sciences, Aoba-yama, Aoba-ku, Sendai 980-8578 (Japan)

    2015-08-07

    Osteosarcoma is a common metastatic bone cancer that predominantly develops in children and adolescents. Metastatic osteosarcoma remains associated with a poor prognosis; therefore, more effective anti-metastatic drugs are needed. Differentiation-inducing factor-1 (DIF-1), −2, and −3 are novel lead anti-tumor agents that were originally isolated from the cellular slime mold Dictyostelium discoideum. Here we investigated the effects of a panel of DIF derivatives on lysophosphatidic acid (LPA)-induced migration of mouse osteosarcoma LM8 cells by using a Boyden chamber assay. Some DIF derivatives such as Br-DIF-1, DIF-3(+2), and Bu-DIF-3 (5–20 μM) dose-dependently suppressed LPA-induced cell migration with associated IC{sub 50} values of 5.5, 4.6, and 4.2 μM, respectively. On the other hand, the IC{sub 50} values of Br-DIF-1, DIF-3(+2), and Bu-DIF-3 versus cell proliferation were 18.5, 7.2, and 2.0 μM, respectively, in LM8 cells, and >20, 14.8, and 4.3 μM, respectively, in mouse 3T3-L1 fibroblasts (non-transformed). Together, our results demonstrate that Br-DIF-1 in particular may be a valuable tool for the analysis of cancer cell migration, and that DIF derivatives such as DIF-3(+2) and Bu-DIF-3 are promising lead anti-tumor agents for the development of therapies that suppress osteosarcoma cell proliferation, migration, and metastasis. - Highlights: • LPA induces cell migration (invasion) in murine osteosarcoma LM8 cells. • DIFs are novel lead anti-tumor agents found in Dictyostelium discoideum. • We examined the effects of DIF derivatives on LPA-induced LM8 cell migration in vitro. • Some of the DIF derivatives inhibited LPA-induced LM8 cell migration.

  1. Depression of NMDA receptor-mediated synaptic transmission by four α2 adrenoceptor agonists on the in vitro rat spinal cord preparation

    Science.gov (United States)

    Faber, E S L; Chambers, J P; Evans, R H

    1998-01-01

    α2-Adrenoceptor agonists have a spinal site of analgesic action. In the current study the synaptic depressant actions of xylazine, detomidine, romifidine and dexmedetomidine have been compared on segmental reflexes containing NMDA receptor-mediated components in the neonatal rat hemisected spinal cord preparation in vitro.Reflexes were evoked in the ventral root following either supramaximal electrical stimulation of the corresponding ipsilateral lumbar dorsal root to evoke the high intensity excitatory postsynaptic potential (e.p.s.p.) involving all primary afferent fibres, or low intensity stimulation to evoke the solely A fibre-mediated low intensity e.p.s.p. The high intensity e.p.s.p. contains a greater NMDA receptor-mediated component.Xylazine, romifidine, detomidine and dexmedetomidine all depressed both the high intensity e.p.s.p. and the low intensity e.p.s.p. giving respective EC50 values of 0.91±0.2 μM (n=12), 23.4±3 nM (n=12), 37.7±7 nM (n=8) and 0.84±0.1 nM (n=4) for depression of the high intensity e.p.s.p. and 0.76±0.1 μM (n=12), 22.0±3 nM (n=12), 24.9±6 nM (n=4) and 2.7±0.6 nM (n=4) for depression of the low intensity e.p.s.p., respectively. Unlike the other three drugs, the two values for dexmedetomidine, showing a greater selectivity for the high intensity e.p.s.p., are significantly different.Each of these depressant actions was reversed by the selective α2-adrenoceptor antagonist atipamezole (1 μM).In contrast to previous reports of the actions of α2-adrenoceptor agonists on the in vitro spinal cord preparation, at concentrations ten fold higher than the above EC50 values xylazine, romifidine, detomidine and dexmedetomidine depressed the initial population spike of motoneurons (MSR). This depression was not reversed by atipamezole.Comparison of the rank order of the present EC50 values for depression of the high intensity e.p.s.p. with potency ratios from in vivo analgesic tests in previous studies show a close

  2. GABA(A receptor-mediated acceleration of aging-associated memory decline in APP/PS1 mice and its pharmacological treatment by picrotoxin.

    Directory of Open Access Journals (Sweden)

    Yuji Yoshiike

    Full Text Available Advanced age and mutations in the genes encoding amyloid precursor protein (APP and presenilin (PS1 are two serious risk factors for Alzheimer's disease (AD. Finding common pathogenic changes originating from these risks may lead to a new therapeutic strategy. We observed a decline in memory performance and reduction in hippocampal long-term potentiation (LTP in both mature adult (9-15 months transgenic APP/PS1 mice and old (19-25 months non-transgenic (nonTg mice. By contrast, in the presence of bicuculline, a GABA(A receptor antagonist, LTP in adult APP/PS1 mice and old nonTg mice was larger than that in adult nonTg mice. The increased LTP levels in bicuculline-treated slices suggested that GABA(A receptor-mediated inhibition in adult APP/PS1 and old nonTg mice was upregulated. Assuming that enhanced inhibition of LTP mediates memory decline in APP/PS1 mice, we rescued memory deficits in adult APP/PS1 mice by treating them with another GABA(A receptor antagonist, picrotoxin (PTX, at a non-epileptic dose for 10 days. Among the saline vehicle-treated groups, substantially higher levels of synaptic proteins such as GABA(A receptor alpha1 subunit, PSD95, and NR2B were observed in APP/PS1 mice than in nonTg control mice. This difference was insignificant among PTX-treated groups, suggesting that memory decline in APP/PS1 mice may result from changes in synaptic protein levels through homeostatic mechanisms. Several independent studies reported previously in aged rodents both an increased level of GABA(A receptor alpha1 subunit and improvement of cognitive functions by long term GABA(A receptor antagonist treatment. Therefore, reduced LTP linked to enhanced GABA(A receptor-mediated inhibition may be triggered by aging and may be accelerated by familial AD-linked gene products like Abeta and mutant PS1, leading to cognitive decline that is pharmacologically treatable at least at this stage of disease progression in mice.

  3. Escitalopram attenuates β-amyloid-induced tau hyperphosphorylation in primary hippocampal neurons through the 5-HT1A receptor mediated Akt/GSK-3β pathway.

    Science.gov (United States)

    Wang, Yan-Juan; Ren, Qing-Guo; Gong, Wei-Gang; Wu, Di; Tang, Xiang; Li, Xiao-Li; Wu, Fang-Fang; Bai, Feng; Xu, Lin; Zhang, Zhi-Jun

    2016-03-22

    Tau hyperphosphorylation is an important pathological feature of Alzheimer's disease (AD). To investigate whether escitalopram could inhibit amyloid-β (Aβ)-induced tau hyperphosphorylation and the underlying mechanisms, we treated the rat primary hippocampal neurons with Aβ1-42 and examined the effect of escitalopram on tau hyperphosphorylation. Results showed that escitalopram decreased Aβ1-42-induced tau hyperphosphorylation. In addition, escitalopram activated the Akt/GSK-3β pathway, and the PI3K inhibitor LY294002 blocked the attenuation of tau hyperphosphorylation induced by escitalopram. Moreover, the 5-HT1A receptor agonist 8-OH-DPAT also activated the Akt/GSK-3β pathway and decreased Aβ1-42-induced tau hyperphosphorylation. Furthermore, the 5-HT1A receptor antagonist WAY-100635 blocked the activation of Akt/GSK-3β pathway and the attenuation of tau hyperphosphorylation induced by escitalopram. Finally, escitalopram improved Aβ1-42 induced impairment of neurite outgrowth and spine density, and reversed Aβ1-42 induced reduction of synaptic proteins. Our results demonstrated that escitalopram attenuated Aβ1-42-induced tau hyperphosphorylation in primary hippocampal neurons through the 5-HT1A receptor mediated Akt/GSK-3β pathway.

  4. Chronic restraint stress causes a delayed increase in responding for palatable food cues during forced abstinence via a dopamine D1-like receptor-mediated mechanism.

    Science.gov (United States)

    Ball, Kevin T; Best, Olivia; Luo, Jonathan; Miller, Leah R

    2017-02-15

    Relapse to unhealthy eating habits in dieters is often triggered by stress. Animal models, moreover, have confirmed a causal role for acute stress in relapse. The role of chronic stress in relapse vulnerability, however, has received relatively little attention. Therefore, in the present study, we used an abstinence-based relapse model in rats to test the hypothesis that exposure to chronic stress increases subsequent relapse vulnerability. Rats were trained to press a lever for highly palatable food reinforcers in daily 3-h sessions and then tested for food seeking (i.e., responding for food associated cues) both before and after an acute or chronic restraint stress procedure (3h/day×1day or 10days, respectively) or control procedure (unstressed). The second food seeking test was conducted either 1day or 7days after the last restraint. Because chronic stress causes dopamine D1-like receptor-mediated alterations in prefrontal cortex (a relapse node), we also assessed dopaminergic involvement by administering either SCH-23390 (10.0μg/kg; i.p.), a dopamine D1-like receptor antagonist, or vehicle prior to daily treatments. Results showed that chronically, but not acutely, stressed rats displayed increased food seeking 7days, but not 1day, after the last restraint. Importantly, SCH-23390 combined with chronic stress reversed this effect. These results suggest that drugs targeting D 1 -like receptors during chronic stress may help to prevent future relapse in dieters. Copyright © 2016 Elsevier B.V. All rights reserved.

  5. Role of a guanine nucleotide-binding protein in α1-adrenergic receptor-mediated Ca2+ mobilization in DDT1 MF-2 cells

    International Nuclear Information System (INIS)

    Cornett, L.E.; Norris, J.S.

    1987-01-01

    In this study the mechanisms involved in α 1 -adrenergic receptor-mediated Ca 2+ mobilization at the level of the plasma membrane were investigated. Stimulation of 45 Ca 2+ efflux from saponin-permeabilized DDT 1 MF-2 cells was observed with the addition of either the α 1 -adrenergic agonist phenylephrine and guanosine-5'-triphosphate or the nonhydrolyzable guanine nucleotide guanylyl-imidodiphosphate. In the presence of [ 32 P] NAD, pertussis toxin was found to catalyze ADP-ribosylation of a M/sub r/ = 40,500 (n = 8) peptide in membranes prepared from DDT 1 , MF-2 cells, possibly the α-subunit of N/sub i/. However, stimulation of unidirectional 45 Ca 2+ efflux by phenylephrine was not affected by previous treatment of cells with 100 ng/ml pertussis toxin. These data suggest that the putative guanine nucleotide-binding protein which couples the α 1 -adrenergic receptor to Ca 2+ mobilization in DDT 1 MF-2 cells is not a pertussis toxin substrate and may possibly be an additional member of guanine nucleotide binding protein family

  6. CB1 receptor-mediated signaling underlies the hippocampal synaptic, learning, and memory deficits following treatment with JWH-081, a new component of spice/K2 preparations.

    Science.gov (United States)

    Basavarajappa, Balapal S; Subbanna, Shivakumar

    2014-02-01

    Recently, synthetic cannabinoids have been sprayed onto plant material, which is subsequently packaged and sold as "Spice" or "K2" to mimic the effects of marijuana. A recent report identified several synthetic additives in samples of "Spice/K2", including JWH-081, a synthetic ligand for the cannabinoid receptor 1 (CB1). The deleterious effects of JWH-081 on brain function are not known, particularly on CB1 signaling, synaptic plasticity, learning and memory. Here, we evaluated the effects of JWH-081 on pCaMKIV, pCREB, and pERK1/2 signaling events followed by long-term potentiation (LTP), hippocampal-dependent learning and memory tasks using CB1 receptor wild-type (WT) and knockout (KO) mice. Acute administration of JWH-081 impaired CaMKIV phosphorylation in a dose-dependent manner, whereas inhibition of CREB phosphorylation in CB1 receptor WT mice was observed only at higher dose of JWH-081 (1.25 mg/kg). JWH-081 at higher dose impaired CaMKIV and CREB phosphorylation in a time-dependent manner in CB1 receptor WT mice but not in KO mice and failed to alter ERK1/2 phosphorylation. In addition, SR treated or CB1 receptor KO mice have a lower pCaMKIV/CaMKIV ratio and higher pCREB/CREB ratio compared with vehicle or WT littermates. In hippocampal slices, JWH-081 impaired LTP in CB1 receptor WT but not in KO littermates. Furthermore, JWH-081 at higher dose impaired object recognition, spontaneous alternation and spatial memory on the Y-maze in CB1 receptor WT mice but not in KO mice. Collectively our findings suggest that deleterious effects of JWH-081 on hippocampal function involves CB1 receptor mediated impairments in CaMKIV and CREB phosphorylation, LTP, learning and memory in mice. © 2013 Wiley Periodicals, Inc.

  7. Ultrastructural evidence for the accumulation of insulin in nuclei of intact 3T3-L1 adipocytes by an insulin-receptor mediated process

    International Nuclear Information System (INIS)

    Smith, R.M.; Jarett, L.

    1987-01-01

    Monomeric ferritin-labeled insulin (F/sub m/-Ins), a biologically active, electron-dense marker of occupied insulin receptors, was used to characterize the internalization of insulin in 3T3-L1 adipocytes. F/sub m/-Ins bound specifically to insulin receptors and was internalized in a time- and temperature-dependent manner. In the nucleus, several F/sub m/-Ins particles usually were found in the same general location-near nuclear pores, associated with the periphery of the condensed chromatin. Addition of a 250-fold excess of unlabeled insulin or incubation at 15 0 C reduced the number of F/sub m/-Ins particles found in nuclei after 90 min by 99% or 92%, respectively. Nuclear accumulation of unlabeled ferritin was only 2% of that found with F/sub m/-Ins after 90 min at 37 0 C. Biochemical experiments utilizing 125 I-labeled insulin and subcellular fractionation indicated that intact 3T3-L1 adipocytes internalized insulin rapidly and that ≅ 3% of the internalized ligand accumulated in nuclei after 1 hr. These data provide biochemical and high-resolution ultrastructural evidence that 3T3-L1 adipocytes accumulate potentially significant amounts of insulin in nuclei by an insulin receptor-mediated process. The transport of insulin or the insulin-receptor complex to nuclei in this cell or in others may be directly involved in the long-term biological effects of insulin - in particular, in the control of DNA and RNA synthesis

  8. Receptor-mediated inhibition of adenylate cyclase and stimulation of arachidonic acid release in 3T3 fibroblasts. Selective susceptibility to islet-activating protein, pertussis toxin

    International Nuclear Information System (INIS)

    Murayama, T.; Ui, M.

    1985-01-01

    Thrombin exhibited diverse effects on mouse 3T3 fibroblasts. It (a) decreased cAMP in the cell suspension, (b) inhibited adenylate cyclase in the Lubrol-permeabilized cell suspension in a GTP-dependent manner, increased releases of (c) arachidonic acid and (d) inositol from the cell monolayer prelabeled with these labeled compounds, (e) increased 45 Ca 2+ uptake into the cell monolayer, and (f) increased 86 Rb + uptake into the cell monolayer in a ouabain-sensitive manner. Most of the effects were reproduced by bradykinin, platelet-activating factor, and angiotensin II. The receptors for these agonists are thus likely to be linked to three separate effector systems: the adenylate cyclase inhibition, the phosphoinositide breakdown leading to Ca 2+ mobilization and phospholipase A2 activation, and the Na,K-ATPase activation. Among the effects of these agonists, (a), (b), (c), and (e) were abolished, but (d) and (f) were not, by prior treatment of the cells with islet-activating protein (IAP), pertussis toxin, which ADP-ribosylates the Mr = 41,000 protein, the alpha-subunit of the inhibitory guanine nucleotide regulatory protein (Ni), thereby abolishing receptor-mediated inhibition of adenylate cyclase. The effects (a), (c), (d), and (e) of thrombin, but not (b), were mimicked by A23187, a calcium ionophore. The effects of A23187, in contrast to those of receptor agonists, were not affected by the treatment of cells with IAP. Thus, the IAP substrate, the alpha-subunit of Ni, or the protein alike, may play an additional role in signal transduction arising from the Ca 2+ -mobilizing receptors, probably mediating process(es) distal to phosphoinositide breakdown and proximal to Ca 2+ gating

  9. Striatal adenosine A2A receptor-mediated positron emission tomographic imaging in 6-hydroxydopamine-lesioned rats using [18F]-MRS5425

    International Nuclear Information System (INIS)

    Bhattacharjee, Abesh Kumar; Lang Lixin; Jacobson, Orit; Shinkre, Bidhan; Ma Ying; Niu Gang; Trenkle, William C.; Jacobson, Kenneth A.; Chen Xiaoyuan; Kiesewetter, Dale O.

    2011-01-01

    Introduction: A 2A receptors are expressed in the basal ganglia, specifically in striatopallidal GABAergic neurons in the striatum (caudate-putamen). This brain region undergoes degeneration of presynaptic dopamine projections and depletion of dopamine in Parkinson's disease. We developed an 18 F-labeled A 2A analog radiotracer ([ 18 F]-MRS5425) for A 2A receptor imaging using positron emission tomography (PET). We hypothesized that this tracer could image A 2A receptor changes in the rat model for Parkinson's disease, which is created following unilateral injection of the monoaminergic toxin 6-hydroxydopamine (6-OHDA) into the substantia nigra. Methods: [ 18 F]-MRS5425 was injected intravenously in anesthetized rats, and PET imaging data were collected. Image-derived percentage injected doses per gram (%ID/g) in regions of interest was measured in the striatum of normal rats and in rats unilaterally lesioned with 6-OHDA after intravenous administration of saline (baseline), D 2 agonist quinpirole (1.0 mg/kg) or D 2 antagonist raclopride (6.0 mg/kg). Results: Baseline %ID/g reached a maximum at 90 s and maintained plateau for 3.5 min, and then declined slowly thereafter. In 6-OHDA-lesioned rats, %ID/g was significantly higher in the lesioned side compared to the intact side, and the baseline total %ID/g (data from both hemispheres were combined) was significantly higher compared to quinpirole stimulation starting from 4.5 min until the end of acquisition at 30 min. Raclopride did not produce any change in uptake compared to baseline or between the hemispheres. Conclusion: Thus, increase of A 2A receptor-mediated uptake of radioactive MRS5425 could be a superior molecular target for Parkinson's imaging.

  10. Striatal adenosine A{sub 2A} receptor-mediated positron emission tomographic imaging in 6-hydroxydopamine-lesioned rats using [{sup 18}F]-MRS5425

    Energy Technology Data Exchange (ETDEWEB)

    Bhattacharjee, Abesh Kumar; Lang Lixin; Jacobson, Orit [Laboratory of Molecular Imaging and Nanomedicine, National Institute of Biomedical Imaging and Bioengineering, National Institutes of Health, Bethesda, MD 20892 (United States); Shinkre, Bidhan [Chemical Biology Unit, Laboratory of Cell Biochemistry and Biology, National Institute of Diabetes and Digestive and Kidney Diseases, National Institutes of Health, Bethesda, MD 20892 (United States); Ma Ying [Laboratory of Molecular Imaging and Nanomedicine, National Institute of Biomedical Imaging and Bioengineering, National Institutes of Health, Bethesda, MD 20892 (United States); Niu Gang [Laboratory of Molecular Imaging and Nanomedicine, National Institute of Biomedical Imaging and Bioengineering, National Institutes of Health, Bethesda, MD 20892 (United States); Department of Radiology and Imaging Sciences, Warren Grant Magnuson Clinical Center, National Institutes of Health, Bethesda, MD 20892 (United States); Trenkle, William C. [Chemical Biology Unit, Laboratory of Cell Biochemistry and Biology, National Institute of Diabetes and Digestive and Kidney Diseases, National Institutes of Health, Bethesda, MD 20892 (United States); Jacobson, Kenneth A. [Molecular Recognition Section, Laboratory of Bioorganic Chemistry, National Institute of Diabetes and Digestive and Kidney Diseases, National Institutes of Health, Bethesda, MD 20892 (United States); Chen Xiaoyuan [Laboratory of Molecular Imaging and Nanomedicine, National Institute of Biomedical Imaging and Bioengineering, National Institutes of Health, Bethesda, MD 20892 (United States); Kiesewetter, Dale O., E-mail: dk7k@nih.gov [Laboratory of Molecular Imaging and Nanomedicine, National Institute of Biomedical Imaging and Bioengineering, National Institutes of Health, Bethesda, MD 20892 (United States)

    2011-08-15

    Introduction: A{sub 2A} receptors are expressed in the basal ganglia, specifically in striatopallidal GABAergic neurons in the striatum (caudate-putamen). This brain region undergoes degeneration of presynaptic dopamine projections and depletion of dopamine in Parkinson's disease. We developed an {sup 18}F-labeled A{sub 2A} analog radiotracer ([{sup 18}F]-MRS5425) for A{sub 2A} receptor imaging using positron emission tomography (PET). We hypothesized that this tracer could image A{sub 2A} receptor changes in the rat model for Parkinson's disease, which is created following unilateral injection of the monoaminergic toxin 6-hydroxydopamine (6-OHDA) into the substantia nigra. Methods: [{sup 18}F]-MRS5425 was injected intravenously in anesthetized rats, and PET imaging data were collected. Image-derived percentage injected doses per gram (%ID/g) in regions of interest was measured in the striatum of normal rats and in rats unilaterally lesioned with 6-OHDA after intravenous administration of saline (baseline), D{sub 2} agonist quinpirole (1.0 mg/kg) or D{sub 2} antagonist raclopride (6.0 mg/kg). Results: Baseline %ID/g reached a maximum at 90 s and maintained plateau for 3.5 min, and then declined slowly thereafter. In 6-OHDA-lesioned rats, %ID/g was significantly higher in the lesioned side compared to the intact side, and the baseline total %ID/g (data from both hemispheres were combined) was significantly higher compared to quinpirole stimulation starting from 4.5 min until the end of acquisition at 30 min. Raclopride did not produce any change in uptake compared to baseline or between the hemispheres. Conclusion: Thus, increase of A{sub 2A} receptor-mediated uptake of radioactive MRS5425 could be a superior molecular target for Parkinson's imaging.

  11. P2Y receptor-mediated transient relaxation of rat longitudinal ileum preparations involves phospholipase C activation, intracellular Ca(2+) release and SK channel activation.

    Science.gov (United States)

    Mader, Felix; Krause, Ludwig; Tokay, Tursonjan; Hakenberg, Oliver W; Köhling, Rüdiger; Kirschstein, Timo

    2016-05-01

    Purinergic signaling plays a major role in the enteric nervous system, where it governs gut motility through a number of P2X and P2Y receptors. The aim of this study was to investigate the P2Y receptor-mediated motility in rat longitudinal ileum preparations. Ileum smooth muscle strips were prepared from rats, and fixed in an organ bath. Isometric contraction and relaxation responses of the muscle strips were measured with force transducers. Drugs were applied by adding of stock solutions to the organ bath to yield the individual final concentrations. Application of the non-hydrolyzable P2 receptor agonists α,β-Me-ATP or 2-Me-S-ADP (10, 100 μmol/L) dose-dependently elicited a transient relaxation response followed by a sustained contraction. The relaxation response was largely blocked by SK channel blockers apamin (500 nmol/L) and UCL1684 (10 μmol/L), PLC inhibitor U73122 (100 μmol/L), IP3 receptor blocker 2-APB (100 μmol/L) or sarcoendoplasmic Ca(2+) ATPase inhibitor thapsigargin (1 μmol/L), but not affected by atropine, NO synthase blocker L-NAME or tetrodotoxin. Furthermore, α,β-Me-ATP-induced relaxation was suppressed by P2Y1 receptor antagonist MRS2179 (50 μmol/L) or P2Y13 receptor antagonist MRS2211 (100 μmol/L), and was abolished by co-application of the two antagonists, whereas 2-Me-S-ADP-induced relaxation was abolished by P2Y6 receptor antagonist MRS2578 (50 μmol/L). In addition, P2Y1 receptor antagonist MRS2500 (1 μmol/L) not only abolished α,β-Me-ATP-induced relaxation, but also suppressed 2-Me-S-ADP-induced relaxation. P2Y receptor agonist-induced transient relaxation of rat ileum smooth muscle strips is mediated predominantly by P2Y1 receptor, but also by P2Y6 and P2Y13 receptors, and involves PLC, IP3, Ca(2+) release and SK channel activation, but is independent of acetylcholine and NO release.

  12. Dopamine D2 receptor-mediated G-protein activation in rat striatum: functional autoradiography and influence of unilateral 6-hydroxydopamine lesions of the substantia nigra.

    Science.gov (United States)

    Newman-Tancredi, A; Cussac, D; Brocco, M; Rivet, J M; Chaput, C; Touzard, M; Pasteau, V; Millan, M J

    2001-11-30

    , the present data indicate that, in rat striatum, the actions of quinelorane are mediated primarily by D2 receptors, and suggest that behavioural hypersensitivity to this agonist, induced by unilateral SNPC lesions, is associated with an increase in D2, but not D3 or D4, receptor-mediated G-protein activation.

  13. Enhanced root growth in phosphate-starved Arabidopsis by stimulating de novo phospholipid biosynthesis through the overexpression of LYSOPHOSPHATIDIC ACID ACYLTRANSFERASE 2 (LPAT2).

    Science.gov (United States)

    Angkawijaya, Artik Elisa; Nguyen, Van Cam; Nakamura, Yuki

    2017-09-01

    Upon phosphate starvation, plants retard shoot growth but promote root development presumably to enhance phosphate assimilation from the ground. Membrane lipid remodelling is a metabolic adaptation that replaces membrane phospholipids by non-phosphorous galactolipids, thereby allowing plants to obtain scarce phosphate yet maintain the membrane structure. However, stoichiometry of this phospholipid-to-galactolipid conversion may not account for the massive demand of membrane lipids that enables active growth of roots under phosphate starvation, thereby suggesting the involvement of de novo phospholipid biosynthesis, which is not represented in the current model. We overexpressed an endoplasmic reticulum-localized lysophosphatidic acid acyltransferase, LPAT2, a key enzyme that catalyses the last step of de novo phospholipid biosynthesis. Two independent LPAT2 overexpression lines showed no visible phenotype under normal conditions but showed increased root length under phosphate starvation, with no effect on phosphate starvation response including marker gene expression, root hair development and anthocyanin accumulation. Accompanying membrane glycerolipid profiling of LPAT2-overexpressing plants revealed an increased content of major phospholipid classes and distinct responses to phosphate starvation between shoot and root. The findings propose a revised model of membrane lipid remodelling, in which de novo phospholipid biosynthesis mediated by LPAT2 contributes significantly to root development under phosphate starvation. © 2017 John Wiley & Sons Ltd.

  14. Mechanism of sphingosine 1-phosphate- and lysophosphatidic Acid-induced up-regulation of adhesion molecules and eosinophil chemoattractant in nerve cells.

    LENUS (Irish Health Repository)

    Costello, Richard W

    2012-02-01

    The lysophospholipids sphingosine 1-phosphate (S1P) and lysophosphatidic acid (LPA) act via G-protein coupled receptors S1P(1-5) and LPA(1-3) respectively, and are implicated in allergy. Eosinophils accumulate at innervating cholinergic nerves in asthma and adhere to nerve cells via intercellular adhesion molecule-1 (ICAM-1). IMR-32 neuroblastoma cells were used as an in vitro cholinergic nerve cell model. The G(i) coupled receptors S1P(1), S1P(3), LPA(1), LPA(2) and LPA(3) were expressed on IMR-32 cells. Both S1P and LPA induced ERK phosphorylation and ERK- and G(i)-dependent up-regulation of ICAM-1 expression, with differing time courses. LPA also induced ERK- and G(i)-dependent up-regulation of the eosinophil chemoattractant, CCL-26. The eosinophil granule protein eosinophil peroxidase (EPO) induced ERK-dependent up-regulation of transcription of S1P(1), LPA(1), LPA(2) and LPA(3), providing the situation whereby eosinophil granule proteins may enhance S1P- and\\/or LPA- induced eosinophil accumulation at nerve cells in allergic conditions.

  15. Mechanism of sphingosine 1-phosphate- and lysophosphatidic Acid-induced up-regulation of adhesion molecules and eosinophil chemoattractant in nerve cells.

    LENUS (Irish Health Repository)

    Costello, Richard W

    2011-05-01

    The lysophospholipids sphingosine 1-phosphate (S1P) and lysophosphatidic acid (LPA) act via G-protein coupled receptors S1P(1-5) and LPA(1-3) respectively, and are implicated in allergy. Eosinophils accumulate at innervating cholinergic nerves in asthma and adhere to nerve cells via intercellular adhesion molecule-1 (ICAM-1). IMR-32 neuroblastoma cells were used as an in vitro cholinergic nerve cell model. The G(i) coupled receptors S1P(1), S1P(3), LPA(1), LPA(2) and LPA(3) were expressed on IMR-32 cells. Both S1P and LPA induced ERK phosphorylation and ERK- and G(i)-dependent up-regulation of ICAM-1 expression, with differing time courses. LPA also induced ERK- and G(i)-dependent up-regulation of the eosinophil chemoattractant, CCL-26. The eosinophil granule protein eosinophil peroxidase (EPO) induced ERK-dependent up-regulation of transcription of S1P(1), LPA(1), LPA(2) and LPA(3), providing the situation whereby eosinophil granule proteins may enhance S1P- and\\/or LPA- induced eosinophil accumulation at nerve cells in allergic conditions.

  16. Expression of lysophosphatidic acid receptor 1 and relation with cell proliferation, apoptosis, and angiogenesis on preneoplastic changes induced by cadmium chloride in the rat ventral prostate.

    Directory of Open Access Journals (Sweden)

    Riánsares Arriazu

    Full Text Available BACKGROUND: Lysophosphatidic acid (LPA is a phospholipid growth factor involved in cell proliferation, differentiation, migration, inflammation, angiogenesis, wound healing, cancer invasion, and survival. This study was directed to evaluate the immunoexpression of LPA-1, cell proliferation, apoptosis, and angiogenesis markers in preneoplastic lesions induced with cadmium chloride in rat prostate. METHODS: The following parameters were calculated in ventral prostate of normal rats and rats that received Cd in drinking water during 24 months: percentages of cells immunoreactive to LPA-1 (LILPA1, PCNA (LIPCNA, MCM7 (LIMCM7, ubiquitin (LIUBI, apoptotic cells (LIAPO, and p53 (LIp53; volume fraction of Bcl-2 (VFBcl-2; and length of microvessels per unit of volume (LVMV/mm3. Data were analyzed using Student's t-test and Pearson correlation test. RESULTS: The LILPA1 in dysplastic lesions and normal epithelium of Cd-treated rats was significantly higher than those in the control group. Markers of proliferation were significantly increased in dysplastic lesions, whereas some apoptotic markers were significantly decreased. No significant differences between groups were found in VFBcl-2. Dysplastic lesions showed a significant increase of LIp53. The length of microvessels per unit of volume was elevated in dysplastic acini. Statistically significant correlations were found only between LILPA1 and LIUBI. CONCLUSIONS: Our results suggest that LPA-1 might be implicated in dysplastic lesions induced by cadmium chloride development. More studies are needed to confirm its potential contribution to the disease.

  17. Lysophosphatidic acid activates peroxisome proliferator activated receptor-γ in CHO cells that over-express glycerol 3-phosphate acyltransferase-1.

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    Cliona M Stapleton

    2011-04-01

    Full Text Available Lysophosphatidic acid (LPA is an agonist for peroxisome proliferator activated receptor-γ (PPARγ. Although glycerol-3-phosphate acyltransferase-1 (GPAT1 esterifies glycerol-3-phosphate to form LPA, an intermediate in the de novo synthesis of glycerolipids, it has been assumed that LPA synthesized by this route does not have a signaling role. The availability of Chinese Hamster Ovary (CHO cells that stably overexpress GPAT1, allowed us to analyze PPARγ activation in the presence of LPA produced as an intracellular intermediate. LPA levels in CHO-GPAT1 cells were 6-fold higher than in wild-type CHO cells, and the mRNA abundance of CD36, a PPARγ target, was 2-fold higher. Transactivation assays showed that PPARγ activity was higher in the cells that overexpressed GPAT1. PPARγ activity was enhanced further in CHO-GPAT1 cells treated with the PPARγ ligand troglitazone. Extracellular LPA, phosphatidic acid (PA or a membrane-permeable diacylglycerol had no effect, showing that PPARγ had been activated by LPA generated intracellularly. Transient transfection of a vector expressing 1-acylglycerol-3-phosphate acyltransferase-2, which converts endogenous LPA to PA, markedly reduced PPARγ activity, as did over-expressing diacylglycerol kinase, which converts DAG to PA, indicating that PA could be a potent inhibitor of PPARγ. These data suggest that LPA synthesized via the glycerol-3-phosphate pathway can activate PPARγ and that intermediates of de novo glycerolipid synthesis regulate gene expression.

  18. Heart-type fatty-acid-binding protein (FABP3 is a lysophosphatidic acid-binding protein in human coronary artery endothelial cells

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    Ryoko Tsukahara

    2014-01-01

    Full Text Available Fatty-acid-binding protein 3, muscle and heart (FABP3, also known as heart-type FABP, is a member of the family of intracellular lipid-binding proteins. It is a small cytoplasmic protein with a molecular mass of about 15 kDa. FABPs are known to be carrier proteins for transporting fatty acids and other lipophilic substances from the cytoplasm to the nucleus, where these lipids are released to a group of nuclear receptors such as peroxisome proliferator-activated receptors (PPARs. In this study, using lysophosphatidic acid (LPA-coated agarose beads, we have identified FABP3 as an LPA carrier protein in human coronary artery endothelial cells (HCAECs. Administration of LPA to HCAECs resulted in a dose-dependent increase in PPARγ activation. Furthermore, the LPA-induced PPARγ activation was abolished when the FABP3 expression was reduced using small interfering RNA (siRNA. We further show that the nuclear fraction of control HCAECs contained a significant amount of exogenously added LPA, whereas FABP3 siRNA-transfected HCAECs had a decreased level of LPA in the nucleus. Taken together, these results suggest that FABP3 governs the transcriptional activities of LPA by targeting them to cognate PPARγ in the nucleus.

  19. Lysophosphatidic acid receptor activation affects the C13NJ microglia cell line proteome leading to alterations in glycolysis, motility, and cytoskeletal architecture

    Science.gov (United States)

    Bernhart, Eva; Kollroser, Manfred; Rechberger, Gerald; Reicher, Helga; Heinemann, Akos; Schratl, Petra; Hallström, Seth; Wintersperger, Andrea; Nusshold, Christoph; DeVaney, Trevor; Zorn-Pauly, Klaus; Malli, Roland; Graier, Wolfgang; Malle, Ernst; Sattler, Wolfgang

    2014-01-01

    Microglia, the immunocompetent cells of the CNS, are rapidly activated in response to injury and microglia migration towards and homing at damaged tissue plays a key role in CNS regeneration. Lysophosphatidic acid (LPA) is involved in signaling events evoking microglia responses through cognate G protein-coupled receptors. Here we show that human immortalized C13NJ microglia express LPA receptor subtypes LPA1, LPA2, and LPA3 on mRNA and protein level. LPA activation of C13NJ cells induced Rho and extracellular signal-regulated kinase activation and enhanced cellular ATP production. In addition, LPA induced process retraction, cell spreading, led to pronounced changes of the actin cytoskeleton and reduced cell motility, which could be reversed by inhibition of Rho activity. To get an indication about LPA-induced global alterations in protein expression patterns a 2-D DIGE/LC-ESI-MS proteomic approach was applied. On the proteome level the most prominent changes in response to LPA were observed for glycolytic enzymes and proteins regulating cell motility and/or cytoskeletal dynamics. The present findings suggest that naturally occurring LPA is a potent regulator of microglia biology. This might be of particular relevance in the pathophysiological context of neurodegenerative disorders where LPA concentrations can be significantly elevated in the CNS. PMID:19899077

  20. Lysophosphatidic Acid Up-regulates MT1-MMP Expression through a Gi –dependent Pathway in Human Umbilical Vein Endothelial Cells

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    Po-Wei Lin

    2009-11-01

    Full Text Available Lysophosphatidic acid (LPA is a low molecular weight lysophospholipid (LPL. Through binding to its specific G protein-coupled receptor family, LPA regulates various cellular functions, including proliferation, migration, invasion, and differentiation. Matrix-metalloproteinases (MMPs are zinc-dependent protease and play important roles in regulating the interaction between cells and extracellular matrix (ECM. Among these MMPs, membrane type 1-metalloproteinase (MT1-MMP not only degrades ECM protein but also activates metalloproteinase-2 (MMP-2, Gelatinase A, which are important to endothelial cell migration. Our previous study showed that LPA enhances MMP-2 expression and activity in human umbilical vein endothelial cells (HUVECs. In this study, we further revealed that LPA also induce MT1-MMP mRNA and protein expressions in HUVECs through real-time PCR and Western blotting, respectively. Furthermore, by applying chemical inhibitors, we found that LPA-induced MT1-MMP expression is mainly through a Gi- and partially through a Gq-dependent pathway. Our results provide new evidence that LPA might modulate ECM through regulating the expression of MT1-MMP.

  1. Enhancement of Human Endothelial Cell Adhesion to Type I Collagen by Lysophosphatidic Acid (LPA and Sphingosine-1-Phosphate (S1P

    Directory of Open Access Journals (Sweden)

    Hsinyu Lee

    2004-06-01

    Full Text Available The diverse cellular effects of lysophosphatidic acid (LPA and sphingosine-1-phosphate (S1P are transduced by two structurally homologous subfamilies of G protein-coupled receptors, which are encoded by endothelial differentiation genes (Edg Rs. Human umbilical cord vein endothelial cells (HUVECs express Edg Rs for LPA (Edg2 and S1P (Edg1 and 3, which transduce signals for migration of HUVECs through micropore filters coated with type I collagen. Since activation of integrins is essential for optimal migration of endothelial cells, we now examine the capacity of LPA and S1P to augment integrin mediation of endothelial cell binding to type I collagen. Lysophospholipid enhancement of HUVEC adhesion to type I collagen is detectable within 20 minutes. Enhancement of adhesion by both LPA and S1P is significant at 50 nM and optimal at 5µM. Pertussis toxin (PTx, a specific inhibitor of Gi, and C3 exotoxin, a specific inhibitor of Rho, both suppress LPA and S1P enhancement of HUVEC adhesion. In contrast, PD98059, which blocks MAP kinase kinase (MEK, and wortmannin, which inhibits phosphatidylinositol 3-kinase (PI3K, had no effect on LPA- or S1P-enhancement of HUVEC adhesion. Neutralizing monoclonal antibodies specific for α2 and β1 integrin chains, concomitantly decrease LPA and S1P enhancement of HUVEC adhesion to type I collagen. LPA and S1P thus promote type I collagen-dependent adhesion and migration of HUVECs by recruiting α2 and β1 integrin through both Gi and Rho pathways. Integrin α2/β1 therefore appears to be critical on the effects of LPA and S1P on endothelial cell physiology.

  2. Lysophosphatidic acid modulates the association of PTP1B with N-cadherin/catenin complex in SKOV3 ovarian cancer cells.

    Science.gov (United States)

    Huang, Ruby Yun-Ju; Wen, Chen-Chen; Liao, Chih-Kai; Wang, Shu-Huei; Chou, Liang-Yin; Wu, Jiahn-Chun

    2012-09-01

    LPA (lysophosphatidic acid) is a natural phospholipid that plays important roles in promoting cancer cell proliferation, invasion and metastases. We previously reported that LPA induces ovarian cancer cell dispersal and disruption of AJ (adherens junction) through the activation of SFK (Src family kinases). In this study, we have investigated the regulatory mechanisms during the early phase of LPA-induced cell dispersal. An in vitro model of the ovarian cancer cell line SKOV3 for cell dispersal was used. LPA induces rapid AJ disruption by increasing the internalization of N-cadherin-β-catenin. By using immunoprecipitations, LPA was shown to induce increased tyrosine phosphorylation of β-catenin and alter the balance of β-catenin-bound SFK and PTP1B (phosphotyrosine phosphatase 1B). The altered balance of tyrosine kinase/phosphatase correlated with a concomitant disintegration of the β-catenin-α-catenin, but not the β-catenin-N-cadherin complex. This disintegration of β-catenin from α-catenin and the cell dispersal caused by LPA can be rescued by blocking SFK activity with the chemical inhibitor, PP2. More importantly, PP2 also restores the level of PTP1B bound to β-catenin. We propose that LPA signalling alters AJ stability by changing the dynamics of tyrosine kinase/phosphatase bound to AJ proteins. This work provides further understanding of the early signalling events regulating ovarian cancer cell dispersal and AJ disruption induced by LPA. © The Author(s) Journal compilation © 2012 International Federation for Cell Biology.

  3. Aromatic hydrocarbon receptor inhibits lysophosphatidic acid-induced vascular endothelial growth factor-A expression in PC-3 prostate cancer cells

    International Nuclear Information System (INIS)

    Wu, Pei-Yi; Lin, Yueh-Chien; Lan, Shun-Yan; Huang, Yuan-Li; Lee, Hsinyu

    2013-01-01

    Highlights: •LPA-induced VEGF-A expression was regulated by HIF-1α and ARNT. •PI3K mediated LPA-induced VEGF-A expression. •AHR signaling inhibited LPA-induced VEGF-A expression in PC-3 cells. -- Abstract: Lysophosphatidic acid (LPA) is a lipid growth factor with multiple biological functions and has been shown to stimulate cancer cell secretion of vascular endothelial growth factor-A (VEGF-A) and trigger angiogenesis. Hypoxia-inducible factor-1 (HIF-1), a heterodimer consisting of HIF-1α and HIF-1β (also known as aromatic hydrocarbon receptor nuclear translocator (ARNT)) subunits, is an important regulator of angiogenesis in prostate cancer (PC) through the enhancement of VEGF-A expression. In this study, we first confirmed the ability of LPA to induce VEGF-A expression in PC-3 cells and then validated that LPA-induced VEGF-A expression was regulated by HIF-1α and ARNT through phosphatidylinositol 3-kinase activation. Aromatic hydrocarbon receptor (AHR), a receptor for dioxin-like compounds, functions as a transcription factor through dimerization with ARNT and was found to inhibit prostate carcinogenesis and vanadate-induced VEGF-A production. Since ARNT is a common dimerization partner of AHR and HIF-1α, we hypothesized that AHR might suppress LPA-induced VEGF-A expression in PC-3 cells by competing with HIF-1α for ARNT. Here we demonstrated that overexpression and ligand activation of AHR inhibited HIF-1-mediated VEGF-A induction by LPA treatment of PC-3 cells. In conclusion, our results suggested that AHR activation may inhibit LPA-induced VEGF-A expression in PC-3 cells by attenuating HIF-1α signaling, and subsequently, suppressing angiogenesis and metastasis of PC. These results suggested that AHR presents a potential therapeutic target for the prevention of PC metastasis

  4. Lysophosphatidic acid signaling through its receptor initiates profibrotic epithelial cell fibroblast communication mediated by epithelial cell derived connective tissue growth factor.

    Science.gov (United States)

    Sakai, Norihiko; Chun, Jerold; Duffield, Jeremy S; Lagares, David; Wada, Takashi; Luster, Andrew D; Tager, Andrew M

    2017-03-01

    The expansion of the fibroblast pool is a critical step in organ fibrosis, but the mechanisms driving expansion remain to be fully clarified. We previously showed that lysophosphatidic acid (LPA) signaling through its receptor LPA 1 expressed on fibroblasts directly induces the recruitment of these cells. Here we tested whether LPA-LPA 1 signaling drives fibroblast proliferation and activation during the development of renal fibrosis. LPA 1 -deficient (LPA 1 -/- ) or -sufficient (LPA 1 +/+ ) mice were crossed to mice with green fluorescent protein expression (GFP) driven by the type I procollagen promoter (Col-GFP) to identify fibroblasts. Unilateral ureteral obstruction-induced increases in renal collagen were significantly, though not completely, attenuated in LPA 1 -/- Col-GFP mice, as were the accumulations of both fibroblasts and myofibroblasts. Connective tissue growth factor was detected mainly in tubular epithelial cells, and its levels were suppressed in LPA 1 -/- Col-GFP mice. LPA-LPA 1 signaling directly induced connective tissue growth factor expression in primary proximal tubular epithelial cells, through a myocardin-related transcription factor-serum response factor pathway. Proximal tubular epithelial cell-derived connective tissue growth factor mediated renal fibroblast proliferation and myofibroblast differentiation. Administration of an inhibitor of myocardin-related transcription factor/serum response factor suppressed obstruction-induced renal fibrosis. Thus, targeting LPA-LPA 1 signaling and/or myocardin-related transcription factor/serum response factor-induced transcription could be promising therapeutic strategies for renal fibrosis. Copyright © 2016 International Society of Nephrology. Published by Elsevier Inc. All rights reserved.

  5. Lysophosphatidic acid-induced RhoA signaling and prolonged macrophage infiltration worsens fibrosis and fatty infiltration following rotator cuff tears.

    Science.gov (United States)

    Davies, Michael R; Lee, Lawrence; Feeley, Brian T; Kim, Hubert T; Liu, Xuhui

    2017-07-01

    Previous studies have suggested that macrophage-mediated chronic inflammation is involved in the development of rotator cuff muscle atrophy and degeneration following massive tendon tears. Increased RhoA signaling has been reported in chronic muscle degeneration, such as muscular dystrophy. However, the role of RhoA signaling in macrophage infiltration and rotator muscle degeneration remains unknown. Using a previously established rat model of massive rotator cuff tears, we found RhoA signaling is upregulated in rotator cuff muscle following a massive tendon-nerve injury. This increase in RhoA expression is greatly potentiated by the administration of a potent RhoA activator, lysophosphatidic acid (LPA), and is accompanied by increased TNFα and TGF-β1 expression in rotator cuff muscle. Boosting RhoA signaling with LPA significantly worsened rotator cuff muscle atrophy, fibrosis, and fatty infiltration, accompanied with massive monocytic infiltration of rotator cuff muscles. Co-staining of RhoA and the tissue macrophage marker CD68 showed that CD68+ tissue macrophages are the dominant cell source of increased RhoA signaling in rotator cuff muscles after tendon tears. Taken together, our findings suggest that LPA-mediated RhoA signaling in injured muscle worsens the outcomes of atrophy, fibrosis, and fatty infiltration by increasing macrophage infiltraion in rotator cuff muscle. Clinically, inhibiting RhoA signaling may represent a future direction for developing new treatments to improve muscle quality following massive rotator cuff tears. © 2016 Orthopaedic Research Society. Published by Wiley Periodicals, Inc. J Orthop Res 35:1539-1547, 2017. © 2016 Orthopaedic Research Society. Published by Wiley Periodicals, Inc.

  6. Lysophosphatidic acid signaling via LPA{sub 1} and LPA{sub 3} regulates cellular functions during tumor progression in pancreatic cancer cells

    Energy Technology Data Exchange (ETDEWEB)

    Fukushima, Kaori; Takahashi, Kaede; Yamasaki, Eri; Onishi, Yuka [Division of Molecular Oncology, Department of Life Science, Faculty of Science and Engineering, Kindai University, 3-4-1, Kowakae, Higashiosaka, Osaka 577-8502 (Japan); Fukushima, Nobuyuki [Division of Molecular Neurobiology, Department of Life Science, Faculty of Science and Engineering, Kindai University, 3-4-1, Kowakae, Higashiosaka, Osaka 577-8502 (Japan); Honoki, Kanya [Department of Orthopedic Surgery, Nara Medical University, 840 Shijo-cho, Kashihara, Nara 634-8521 (Japan); Tsujiuchi, Toshifumi, E-mail: ttujiuch@life.kindai.ac.jp [Division of Molecular Oncology, Department of Life Science, Faculty of Science and Engineering, Kindai University, 3-4-1, Kowakae, Higashiosaka, Osaka 577-8502 (Japan)

    2017-03-01

    Lysophosphatidic acid (LPA) signaling via G protein-coupled LPA receptors exhibits a variety of biological effects, such as cell proliferation, motility and differentiation. The aim of this study was to evaluate the roles of LPA{sub 1} and LPA{sub 3} in cellular functions during tumor progression in pancreatic cancer cells. LPA{sub 1} and LPA{sub 3} knockdown cells were generated from PANC-1 cells. The cell motile and invasive activities of PANC-1 cells were inhibited by LPA{sub 1} and LPA{sub 3} knockdown. In gelatin zymography, LPA{sub 1} and LPA{sub 3} knockdown cells indicated the low activation of matrix metalloproteinase-2 (MMP-2) in the presence of LPA. Next, to assess whether LPA{sub 1} and LPA{sub 3} regulate cellular functions induced by anticancer drug, PANC-1 cells were treated with cisplatin (CDDP) for approximately 6 months. The cell motile and invasive activities of long-term CDDP treated cells were markedly higher than those of PANC-1 cells, correlating with the expression levels of LPAR1 and LPAR3 genes. In soft agar assay, the long-term CDDP treated cells formed markedly large sized colonies. In addition, the cell motile and invasive activities enhanced by CDDP were significantly suppressed by LPA{sub 1} and LPA{sub 3} knockdown as well as colony formation. These results suggest that LPA signaling via LPA{sub 1} and LPA{sub 3} play an important role in the regulation of cellular functions during tumor progression in PANC-1 cells. - Highlights: • The cell motile and invasive activities of PANC-1 cells were stimulated by LPA{sub 1} and LPA{sub 3}. • LPA{sub 1} and LPA{sub 3} enhanced MMP-2 activation in PANC-1 cells. • The expressions of LPAR1 and LPAR3 genes were elevated in PANC-1 cells treated with cisplatin. • The cell motile and invasive activities of PANC-1 cells treated with cisplatin were suppressed by LPA{sub 1} and LPA{sub 3} knockdown. • LPA{sub 1} and LPA{sub 3} are involved in the regulation of cellular functions during tumor

  7. The influence of using anticoagulants (EDTA and citrate acid 3.8% toward the quantity of Platelet Rich Plasma (PRP

    Directory of Open Access Journals (Sweden)

    Lilies Anggarwati Astuti

    2016-06-01

    Full Text Available Platelet Rich Plasma (PRP is a blood concentrate that has a thrombocytes concentration several time higher than normal concentration of thrombocytes in normal human blood. PRP is a promising alternative to surgery with a safe and natural healing. The standard protocol for PRP preparation must be determined to get the right quantity and quality of the matrix of fibrin, leukocytes, platelets and growth factors. It could not be separated from the number of PRP produced. The use of PRP in the success of periodontal treatment would not be separated from methods to obtain it. To detect the influence of using anticoagulants (EDTA and citrate acid 3.8% toward the quantity of PRP. There are 41 subjects studied by taking 21 ml of venous blood in each of the seven tubes. Centrifugation performed twice with different speed, duration, use of anticoagulants then analyzed. This quantity between the two groups differed significantly between the PRP in EDTA group is higher 322.2 ml rather than citrate acid 3.8% group, then control group is higher 329.5 ml rather than citrate acid 3.8% group, while there is no difference between EDTA and control group. There is effect of the use of anticoagulants EDTA compared with citrate acid 3.8% in the quantity of PRP, and there was no effect using citrate acid 3.8% as anticoagulants in quantity of PRP.

  8. Monoclonal antibody to the type I insulin-like growth factor (IGF-I) receptor blocks IGF-I receptor-mediated DNA synthesis: clarification of the mitogenic mechanisms of IGF-I and insulin in human skin fibroblasts

    International Nuclear Information System (INIS)

    Flier, J.S.; Usher, P.; Moses, A.C.

    1986-01-01

    Insulin and insulin-like growth factor type I (IGF-I) stimulate an overlapping spectrum of biological responses in human skin fibroblasts. Although insulin and IGF-I are known to stimulate the incorporation of [ 3 H]thymidine into DNA in these cells, the identify of the receptor(s) that mediates this effect has not been fully clarified. The mouse anti-human IGF-I receptor antibody αIR-3 binds with specificity to IGF-I but not to insulin receptors in human placental membranes; it also specifically inhibits the binding of 125 I-labeled IGF-I but not 125 I-labeled insulin to suspensions of human skin fibroblasts in a dose-dependent manner. αIR-3 competitively inhibits IGF-I-mediated stimulation of [ 3 H]thymidine incorporation into DNA. This inhibition is dependent on the concentration of αIR-3 and in the presence of a fixed antibody concentration can be partially overcome by high concentrations of IGF-I. In contrast, at concentrations of 3 H]thymidine incorporation is not inhibited by αIR-3. However, the incremental effects of higher concentrations (> 1 μg/ml) of insulin on [ 3 H]thymidine incorporation are inhibited by αIR-3. αIR-3 is a highly specific antagonist of IGF-I receptor-mediated mitogenesis in human skin fibroblasts. By using this antibody, it is shown directly that insulin can act through the IGF-I receptor to stimulate DNA synthesis but can also activate this effect through the insulin receptor itself

  9. Modelling zwitterions in solution: 3-fluoro-γ-aminobutyric acid (3F-GABA).

    Science.gov (United States)

    Cao, Jie; Bjornsson, Ragnar; Bühl, Michael; Thiel, Walter; van Mourik, Tanja

    2012-01-02

    The conformations and relative stabilities of folded and extended 3-fluoro-γ-aminobutyric acid (3F-GABA) conformers were studied using explicit solvation models. Geometry optimisations in the gas phase with one or two explicit water molecules favour folded and neutral structures containing intramolecular NH···O-C hydrogen bonds. With three or five explicit water molecules zwitterionic minima are obtained, with folded structures being preferred over extended conformers. The stability of folded versus extended zwitterionic conformers increases on going from a PCM continuum solvation model to the microsolvated complexes, though extended structures become less disfavoured with the inclusion of more water molecules. Full explicit solvation was studied with a hybrid quantum-mechanical/molecular-mechanical (QM/MM) scheme and molecular dynamics simulations, including more than 6000 TIP3P water molecules. According to free energies obtained from thermodynamic integration at the PM3/MM level and corrected for B3LYP/MM total energies, the fully extended conformer is more stable than folded ones by about -4.5 kJ mol(-1). B3LYP-computed (3)J(F,H) NMR spin-spin coupling constants, averaged over PM3/MM-MD trajectories, agree best with experiment for this fully extended form, in accordance with the original NMR analysis. The seeming discrepancy between static PCM calculations and experiment noted previously is now resolved. That the inexpensive semiempirical PM3 method performs so well for this archetypical zwitterion is encouraging for further QM/MM studies of biomolecular systems. Copyright © 2012 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  10. Opioid receptor mediated anticonvulsant effect of pentazocine.

    Science.gov (United States)

    Khanna, N; Khosla, R; Kohli, J

    1998-01-01

    Intraperitoneal (i.p.) administration of (+/-) pentazocine (10, 30 & 50 mg/kg), a Sigma opioid agonist, resulted in a dose dependent anticonvulsant action against maximal electroshock seizures in mice. This anticonvulsant effect of pentazocine was not antagonized by both the doses of naloxone (1 and 10 mg/kg) suggesting thereby that its anticonvulsant action is probably mediated by Sigma opiate binding sites. Its anticonvulsant effect was potentiated by both the anticonvulsant drugs viz. diazepam and diphenylhydantoin. Morphine, mu opioid agonist, on the other hand, failed to protect the animals against maximal electroshock seizures when it was given in doses of 10-40 mg/kg body wt.

  11. Molecular Mechanisms of Dopamine Receptor Mediated Neuroprotection

    National Research Council Canada - National Science Library

    Sealfon, Stuart

    2000-01-01

    ... of the cellular changes characteristic of this process. Evidence from our laboratory and others suggest that activation of dopamine receptors can oppose the induction of apoptosis in dopamine neurons...

  12. Inositol trisphosphate receptor mediated spatiotemporal calcium signalling.

    Science.gov (United States)

    Miyazaki, S

    1995-04-01

    Spatiotemporal Ca2+ signalling in the cytoplasm is currently understood as an excitation phenomenon by analogy with electrical excitation in the plasma membrane. In many cell types, Ca2+ waves and Ca2+ oscillations are mediated by inositol 1,4,5-trisphosphate (IP3) receptor/Ca2+ channels in the endoplasmic reticulum membrane, with positive feedback between cytosolic Ca2+ and IP3-induced Ca2+ release creating a regenerative process. Remarkable advances have been made in the past year in the analysis of subcellular Ca2+ microdomains using confocal microscopy and of Ca2+ influx pathways that are functionally coupled to IP3-induced Ca2+ release. Ca2+ signals can be conveyed into the nucleus and mitochondria. Ca2+ entry from outside the cell allows repetitive Ca2+ release by providing Ca2+ to refill the endoplasmic reticulum stores, thus giving rise to frequency-encoded Ca2+ signals.

  13. Ga-66 labeled somatostatin analogue DOTA-DPhe1-Tyr3-octreotide as a potential agent for positron emission tomography imaging and receptor mediated internal radiotherapy of somatostatin receptor positive tumors

    International Nuclear Information System (INIS)

    Ugur, Oemer; Kothari, Paresh J.; Finn, Ronald D.; Zanzonico, Pat; Ruan, Shutian; Guenther, Ilonka; Maecke, Helmut R.; Larson, Steven M.

    2002-01-01

    Radionuclide labeled somatostatin analogues selectively target somatostatin receptor (SSTR)-expressing tumors as a basis for diagnosis and treatment of these tumors. Recently, a DOTA-functionalized somatostatin analogue, DOTATOC (DOTA-DPhe 1 -Tyr 3 -octreotide) has been developed. This compound has been shown to be superior to the other somatostatin analogues as indicated by its uniquely high tumor-to-non-target tissue ratio. DOTATOC can be labeled with a variety of radiometals including gallium radioisotopes. Gallium-66 is a positron emitting radionuclide (T 1/2 =9.5 hr; β + =56%), that can be produced in carrier free form by a low-beam energy cyclotron. In this study we investigated SSTR targeting characteristics of 66 Ga-DOTATOC in AR42J rat pancreas tumor implanted nude mice as a potential agent for diagnosis and receptor-mediated internal radiotherapy of SSTR-expressing tumors. We compared our results with 67 Ga- and 68 Ga- labeled DOTATOC. The radiolabeling procedure gave labeling yield ranged from 85-95% and radiochemical and chemical purity was >95%. In-vitro competitive binding curves and in-vivo competitive displacement studies with an excess of unlabeled peptide indicates that there is specific binding of the radioligand to SSTR. Animal biodistribution data and serial microPET TM images demonstrated rapid tumor uptake and rapid clearance from the blood and all tissues except kidney. Maximum % ID/g values for tumor were 10.0±0.7, 13.2±2.1 and 9.8±1.5 for 66 Ga-, 67 Ga-, and 68 Ga-DOTATOC, respectively. Calculated tumor, kidney and bone marrow doses for 66 Ga-DOTATOC based on biodistribution data were 178, 109 and 1.2 cGy/MBq, respectively. We conclude that 66 Ga labeled DOTATOC can be used for PET diagnosis and quantitative imaging-based dosimetry of SSTR positive tumors. 66 Ga-DOTATOC may also be used in higher doses for ablation of these tumors. However, kidney is the critical organ for toxicity (tumor/kidney ratio 1.64), and high kidney uptake must

  14. Ga-66 labeled somatostatin analogue DOTA-DPhe{sup 1}-Tyr{sup 3}-octreotide as a potential agent for positron emission tomography imaging and receptor mediated internal radiotherapy of somatostatin receptor positive tumors

    Energy Technology Data Exchange (ETDEWEB)

    Ugur, Oemer E-mail: ougur@hacettepe.edu.tr; Kothari, Paresh J.; Finn, Ronald D.; Zanzonico, Pat; Ruan, Shutian; Guenther, Ilonka; Maecke, Helmut R.; Larson, Steven M

    2002-02-01

    Radionuclide labeled somatostatin analogues selectively target somatostatin receptor (SSTR)-expressing tumors as a basis for diagnosis and treatment of these tumors. Recently, a DOTA-functionalized somatostatin analogue, DOTATOC (DOTA-DPhe{sup 1}-Tyr{sup 3}-octreotide) has been developed. This compound has been shown to be superior to the other somatostatin analogues as indicated by its uniquely high tumor-to-non-target tissue ratio. DOTATOC can be labeled with a variety of radiometals including gallium radioisotopes. Gallium-66 is a positron emitting radionuclide (T{sub 1/2} =9.5 hr; {beta}{sup +}=56%), that can be produced in carrier free form by a low-beam energy cyclotron. In this study we investigated SSTR targeting characteristics of {sup 66}Ga-DOTATOC in AR42J rat pancreas tumor implanted nude mice as a potential agent for diagnosis and receptor-mediated internal radiotherapy of SSTR-expressing tumors. We compared our results with {sup 67}Ga- and {sup 68}Ga- labeled DOTATOC. The radiolabeling procedure gave labeling yield ranged from 85-95% and radiochemical and chemical purity was >95%. In-vitro competitive binding curves and in-vivo competitive displacement studies with an excess of unlabeled peptide indicates that there is specific binding of the radioligand to SSTR. Animal biodistribution data and serial microPET{sup TM} images demonstrated rapid tumor uptake and rapid clearance from the blood and all tissues except kidney. Maximum % ID/g values for tumor were 10.0{+-}0.7, 13.2{+-}2.1 and 9.8{+-}1.5 for {sup 66}Ga-, {sup 67}Ga-, and {sup 68}Ga-DOTATOC, respectively. Calculated tumor, kidney and bone marrow doses for {sup 66}Ga-DOTATOC based on biodistribution data were 178, 109 and 1.2 cGy/MBq, respectively. We conclude that {sup 66}Ga labeled DOTATOC can be used for PET diagnosis and quantitative imaging-based dosimetry of SSTR positive tumors. {sup 66}Ga-DOTATOC may also be used in higher doses for ablation of these tumors. However, kidney is the

  15. h5-HT1B receptor-mediated constitutive Gαi3-protein activation in stably transfected Chinese hamster ovary cells: an antibody capture assay reveals protean efficacy of 5-HT

    Science.gov (United States)

    Newman-Tancredi, Adrian; Cussac, Didier; Marini, Laetitia; Touzard, Manuelle; Millan, Mark J

    2003-01-01

    Serotonin 5-HT1B receptors couple to G-proteins of the Gi/o family. However, their activation of specific G-protein subtypes is poorly characterised. Using an innovative antibody capture/guanosine-5′-0-(3-[35S]thio)-triphosphate ([35S]GTPγS) binding strategy, we characterised Gαi3 subunit activation by h5-HT1B receptors stably expressed in Chinese hamster ovary (CHO) cells. The agonists, 5-HT, alniditan and BMS181,101, stimulated Gαi3, whereas methiothepin and SB224,289 behaved as inverse agonists. The selective 5-HT1B receptor ligand, S18127, modestly stimulated Gαi3 and reversed the actions of both 5-HT and methiothepin. S18127 (1 μM) also produced parallel, dextral shifts of the 5-HT and methiothepin isotherms. Isotopic dilution experiments ([35S]GTPγS versus GTPγS) revealed high-affinity [35S]GTPγS binding to Gαi3 subunits in the absence of receptor ligands indicating constitutive activity. High-affinity [35S]GTPγS binding was increased 2.8-fold by 5-HT with an increase in the affinity of GTPγS for Gαi3 subunits. In contrast, methiothepin halved the number of high-affinity binding sites and decreased their affinity. h5-HT1B receptor-mediated Gαi3 subunit activation was dependent on the concentration of NaCl. At 300 mM, 5-HT stimulated [35S]GTPγS binding, basal Gαi3 activation was low and methiothepin was inactive. In contrast, at 10 mM NaCl, basal activity was enhanced and the inverse agonist activity of methiothepin was accentuated. Under these conditions, 5-HT decreased Gαi3 activation. In conclusion, at h5-HT1B receptors expressed in CHO cells: (i) inverse agonist induced inhibition of Gαi3, and its reversal by S18127, reveals constitutive activation of this Gα subunit; (ii) constitutive Gαi3 activation can be quantified by isotopic dilution [35S]GTPγS binding and (iii) decreasing NaCl concentrations enhances Gαi3 activation and leads to protean agonist properties of 5-HT: that is a switch to inhibition of Gαi3. PMID:12684263

  16. h5-HT(1B) receptor-mediated constitutive Galphai3-protein activation in stably transfected Chinese hamster ovary cells: an antibody capture assay reveals protean efficacy of 5-HT.

    Science.gov (United States)

    Newman-Tancredi, Adrian; Cussac, Didier; Marini, Laetitia; Touzard, Manuelle; Millan, Mark J

    2003-03-01

    1. Serotonin 5-HT(1B) receptors couple to G-proteins of the Gi/o family. However, their activation of specific G-protein subtypes is poorly characterised. Using an innovative antibody capture/guanosine-5'-0-(3-[(35)S]thio)-triphosphate ([(35)S]GTPgammaS) binding strategy, we characterised Galpha(i3) subunit activation by h5-HT(1B) receptors stably expressed in Chinese hamster ovary (CHO) cells. 2. The agonists, 5-HT, alniditan and BMS181,101, stimulated Galpha(i3), whereas methiothepin and SB224,289 behaved as inverse agonists. The selective 5-HT(1B) receptor ligand, S18127, modestly stimulated Galpha(i3) and reversed the actions of both 5-HT and methiothepin. S18127 (1 micro M) also produced parallel, dextral shifts of the 5-HT and methiothepin isotherms. 3. Isotopic dilution experiments ([(35)S]GTPgammaS versus GTPgammaS) revealed high-affinity [(35)S]GTPgammaS binding to Galpha(i3) subunits in the absence of receptor ligands indicating constitutive activity. High-affinity [(35)S]GTPgammaS binding was increased 2.8-fold by 5-HT with an increase in the affinity of GTPgammaS for Galpha(i3) subunits. In contrast, methiothepin halved the number of high-affinity binding sites and decreased their affinity. 4. h5-HT(1B) receptor-mediated Galpha(i3) subunit activation was dependent on the concentration of NaCl. At 300 mM, 5-HT stimulated [(35)S]GTPgammaS binding, basal Galpha(i3) activation was low and methiothepin was inactive. In contrast, at 10 mM NaCl, basal activity was enhanced and the inverse agonist activity of methiothepin was accentuated. Under these conditions, 5-HT decreased Galpha(i3) activation. 5. In conclusion, at h5-HT(1B) receptors expressed in CHO cells: (i) inverse agonist induced inhibition of Galpha(i3), and its reversal by S18127, reveals constitutive activation of this Galpha subunit; (ii) constitutive Galpha(i3) activation can be quantified by isotopic dilution [(35)S]GTPgammaS binding and (iii) decreasing NaCl concentrations enhances Galpha(i3

  17. Effects of Exogenous Gibberellic Acid3 on Iron and Manganese Plaque Amounts and Iron and Manganese Uptake in Rice

    Science.gov (United States)

    Guo, Yue; Zhu, Changhua; Gan, Lijun; Ng, Denny; Xia, Kai

    2015-01-01

    Gibberellins (GA) regulate various components of plant development. Iron and Mn plaque result from oxiding and hydroxiding Fe and Mn, respectively, on the roots of aquatic plant species such as rice (Oryza sativa L.). In this study, we found that exogenous gibberellic acid3 (GA3) spray decreased Fe plaque, but increased Mn plaque, with applications of Kimura B nutrient solution. Similar effects from GA3, leading to reduced Fe plaque and increased Mn plaque, were also found by scanning electron microscopy and energy dispersive X-ray spectrometric microanalysis. Reduced Fe plaque was observed after applying GA3 to the groups containing added Fe2+ (17 and 42 mg•L-1) and an increasing trend was detected in Mn plaques of the Mn2+ (34 and 84 mg•L-1) added treatments. In contrast, an inhibitor of GA3, uniconazole, reversed the effects of GA3. The uptake of Fe or Mn in rice plants was enhanced after GA3 application and Fe or Mn plaque production. Strong synergetic effects of GA3 application on Fe plaque production were detected. However, no synergetic effects on Mn plaque production were detected. PMID:25710173

  18. Screening-level Biomonitoring Equivalents for tiered interpretation of urinary 3-phenoxybenzoic acid (3-PBA) in a risk assessment context.

    Science.gov (United States)

    Aylward, Lesa L; Irwin, Kim; St-Amand, Annie; Nong, Andy; Hays, Sean M

    2018-02-01

    3-Phenoxybenzoic acid (3-PBA) is a common metabolite of several pyrethroid pesticides of differing potency and also occurs as a residue in foods resulting from environmental degradation of parent pyrethroid compounds. Thus, 3-PBA in urine is not a specific biomarker of exposure to a particular pyrethroid. However, an approach derived from the use of Biomonitoring Equivalents (BEs) can be used to estimate a conservative initial screening value for a tiered assessment of population data on 3-PBA in urine. A conservative generic urinary excretion fraction for 3-PBA was estimated from data for five pyrethroid compounds with human data. Estimated steady-state urinary 3-PBA concentrations associated with reference doses and acceptable daily intakes for each of the nine compounds ranged from 1.7 μg/L for cyhalothrin and deltamethrin to 520 μg/L for permethrin. The lower value can be used as a highly conservative Tier 1 screening value for assessment of population urinary 3-PBA data. A second tier screening value of 87 μg/L was derived based on weighting by relative exposure estimates for the different pyrethroid compounds, to be applied as part of the data evaluation process if biomonitoring data exceed the Tier 1 value. These BE values are most appropriately used to evaluate the central tendency of population biomarker concentration data in a risk assessment context. The provisional BEs were compared to available national biomonitoring data from the US and Canada. Copyright © 2017. Published by Elsevier Inc.

  19. Induced resistance by cresotic acid (3-hydroxy-4-methyl methylbenzoic acid) against wilt disease of melon and cotton

    International Nuclear Information System (INIS)

    Dong, H.; Li, Z.; Zhang, D.; Li, W.; Tang, W.

    2004-01-01

    Cresotic acid (3-hydroxy-4-methylbenzoic acid) was proved be active in controlling wilt diseases of melon and cotton plants grown in the house. Soil drench with 200-1000 ppm cresotic acid induced 62-77 %, 69-79 % and 50-60 % protection against Fusarium oxysporum f.sp melonis (FOM) in melon, Fusarium oxysporum f.sp vasinfectum (FOV) and Verticillium dahliae in cotton, respectively. Since no inhibitory effect of cresotic acid on mycelial growth of these three fungual pathogens was observed in vitro, it is suggested that control of these wilt diseases with cresotic acid resulted from induced resistance. Cresotic acid induced resistance in melon plants not only against race 0, race 1, race 2 and race 1,2, but also against a mixture of these four races of FOM, suggesting a non-race- specific resistance. Level of induced resistance by cresotic acid against FOM depended on inoculum pressure applied to melon plants. At 25 day after inoculation with FOM, percentage protection induced by cresotic acid under low inoculum pressure retained a level of 51 %, while under high inoculum pressure percentage protection decreased to only 10 %. High concentrations of cresotic acid significantly reduced plant growth. Reduction in fresh weight of melon (36-51%) and cotton (42-71%) was obtained with 500-1000 ppm cresotic acid, while only less than 8% reduction occurred with 100-200 ppm. (author)

  20. An Inter-Laboratory Comparison for the Urinary Acrolein Biomarker 3-Hydroxypropyl-Mercapturic Acid (3-HPMA

    Directory of Open Access Journals (Sweden)

    Scherer Gerhard

    2017-01-01

    Full Text Available An inter-laboratory comparison study on the acrolein biomarker of exposure 3-hydroxypropyl-mercapturic acid (3-HPMA with 12 laboratories from 7 globally distributed countries was performed. The laboratories received coded triplicates of 4 spiked and lyophilized urine samples (LU, 12 samples as well as 5 authentic urine pool samples (PU, 15 samples covering the 3-HPMA concentration range from background (non-smoking to heavy smoking levels for analysis by using their own (in-house analytical method. All laboratories applied liquid chromatography with tandem mass spectrometry (LC-MS/MS, with most of them (10 of 12 using solid phase extraction (SPE as sample work-up procedure. The intra-laboratory variation (indicating repeatability was determined by calculating the standard deviation (sr and the coefficient of variation (CVr of the triplicates, whereas the inter-laboratory variation (indicating reproducibility was determined by calculating the standard deviation between laboratories (sR and the corresponding coefficient of variation (CVR. After removal of outlier samples or laboratories, the mean CVr values for LU and PU test samples ranged from 2.1–3.6% (mean: 2.8% and 2.4–3.7% (mean: 3.3%, respectively, indicating good repeatability for the determination of 3-HPMA in both sample types. CVR for LU and PU test samples ranged from 9.1–31.9% (mean: 18.8% and 13.9–27.0% (mean: 18.5%, respectively, indicating limited reproducibility in 3-HPMA analysis for both sample types. Re-calculation of the PU results by applying an embedded calibration (EC, derived from the reported peak areas for the LU test samples, somewhat improved the CVR values (range: 9.6–28.8%, mean: 16.7%.

  1. Suppression of Thyroid Hormone Receptor-Mediated Transcription ...

    African Journals Online (AJOL)

    TH)-induced TR-mediated transcription. We further examined the effects of methamidophos on TR-thyroid hormone response element (TRE) binding using the liquid chemiluminescent DNA pull-down assay (LCDPA), and found no dissociation of ...

  2. Adrenergic receptor-mediated modulation of striatal firing patterns.

    Science.gov (United States)

    Ohta, Hiroyuki; Kohno, Yu; Arake, Masashi; Tamura, Risa; Yukawa, Suguru; Sato, Yoshiaki; Morimoto, Yuji; Nishida, Yasuhiro; Yawo, Hiromu

    2016-11-01

    Although noradrenaline and adrenaline are some of the most important neurotransmitters in the central nervous system, the effects of noradrenergic/adrenergic modulation on the striatum have not been determined. In order to explore the effects of adrenergic receptor (AR) agonists on the striatal firing patterns, we used optogenetic methods which can induce continuous firings. We employed transgenic rats expressing channelrhodopsin-2 (ChR2) in neurons. The medium spiny neuron showed a slow rising depolarization during the 1-s long optogenetic striatal photostimulation and a residual potential with 8.6-s half-life decay after the photostimulation. As a result of the residual potential, five repetitive 1-sec long photostimulations with 20-s onset intervals cumulatively increased the number of spikes. This 'firing increment', possibly relating to the timing control function of the striatum, was used to evaluate the AR modulation. The β-AR agonist isoproterenol decreased the firing increment between the 1st and 5th stimulation cycles, while the α 1 -AR agonist phenylephrine enhanced the firing increment. Isoproterenol and adrenaline increased the early phase (0-0.5s of the photostimulation) firing response. This adrenergic modulation was inhibited by the β-antagonist propranolol. Conversely, phenylephrine and noradrenaline reduced the early phase response. β-ARs and α 1 -ARs work in opposition controlling the striatal firing initiation and the firing increment. Copyright © 2016 Elsevier Ireland Ltd and Japan Neuroscience Society. All rights reserved.

  3. Outline of therapeutic interventions with muscarinic receptor-mediated transmission

    Czech Academy of Sciences Publication Activity Database

    Jakubík, Jan; Šantrůčková, Eva; Randáková, Alena; Janíčková, Helena; Zimčík, Pavel; Rudajev, Vladimír; Michal, Pavel; El-Fakahany, E. E.; Doležal, Vladimír

    2014-01-01

    Roč. 63, Suppl.1 (2014), S177-S189 ISSN 0862-8408 R&D Projects: GA ČR(CZ) GA305/09/0681; GA ČR(CZ) GAP304/12/0259; GA MŠk(CZ) 7E10060 Institutional support: RVO:67985823 Keywords : cholinergic transmission * muscarinic receptors * therapy * Alzheimer's disease, * schizophrenia Subject RIV: ED - Physiology Impact factor: 1.293, year: 2014

  4. Fas receptor-mediated apoptosis : a clinical application?

    NARCIS (Netherlands)

    Timmer, T; de Vries, EGE; de Jong, S

    Fas is a membrane protein belonging to the death receptor family. Cross-linking of Fas by its ligand, FasL, or agonistic anti-Fas antibodies, induces apoptosis of cells expressing Fas on the membrane by triggering a cascade of caspases. Since many different tumours express Fas on their membrane,

  5. Dynamics of Receptor-Mediated Nanoparticle Internalization into Endothelial Cells

    Science.gov (United States)

    Gonzalez-Rodriguez, David; Barakat, Abdul I.

    2015-01-01

    Nanoparticles offer a promising medical tool for targeted drug delivery, for example to treat inflamed endothelial cells during the development of atherosclerosis. To inform the design of such therapeutic strategies, we develop a computational model of nanoparticle internalization into endothelial cells, where internalization is driven by receptor-ligand binding and limited by the deformation of the cell membrane and cytoplasm. We specifically consider the case of nanoparticles targeted against ICAM-1 receptors, of relevance for treating atherosclerosis. The model computes the kinetics of the internalization process, the dynamics of binding, and the distribution of stresses exerted between the nanoparticle and the cell membrane. The model predicts the existence of an optimal nanoparticle size for fastest internalization, consistent with experimental observations, as well as the role of bond characteristics, local cell mechanical properties, and external forces in the nanoparticle internalization process. PMID:25901833

  6. Of pheromones and kairomones: what receptors mediate innate emotional responses?

    Science.gov (United States)

    Fortes-Marco, Lluis; Lanuza, Enrique; Martinez-Garcia, Fernando

    2013-09-01

    Some chemicals elicit innate emotionally laden behavioral responses. Pheromones mediate sexual attraction, parental care or agonistic confrontation, whereas predators' kairomones elicit defensive behaviors in their preys. This essay explores the hypothesis that the detection of these semiochemicals relies on highly specific olfactory and/or vomeronasal receptors. The V1R, V2R, and formyl-peptide vomeronasal receptors bind their ligands in highly specific and sensitive way, thus being good candidates for pheromone- or kairomone-detectors (e.g., secreted and excreted proteins, peptides and lipophilic volatiles). The olfactory epithelium also expresses specific receptors, for example trace amine-associated receptors (TAAR) and guanylyl cyclase receptors (GC-D and other types), some of which bind kairomones and putative pheromones. However, most of the olfactory neurons express canonical olfactory receptors (ORs) that bind many ligands with different affinity, being not suitable for mediating responses to pheromones and kairomones. In this respect, trimethylthiazoline (TMT) is considered a fox-derived kairomone for mice and rats, but it seems to be detected by canonical ORs. Therefore, we have reassessed the kairomonal nature of TMT by analyzing the behavioral responses of outbred (CD1) and inbred mice (C57BL/J6) to TMT. Our results confirm that both mouse strains avoid TMT, which increases immobility in C57BL/J6, but not CD1 mice. However, mice of both strains sniff at TMT throughout the test and show no trace of TMT-induced contextual conditioning (immobility or avoidance). This suggests that TMT is not a kairomone but, similar to a loud noise, in high concentrations it induces aversion and stress as unspecific responses to a strong olfactory stimulation. Copyright © 2013 Wiley Periodicals, Inc.

  7. Effect of ARA9 on dioxin receptor mediated transcription

    International Nuclear Information System (INIS)

    Lees, M.J.; Whitelaw, M.L.

    2002-01-01

    The dioxin (Aryl hydrocarbon) receptor (DR) is a unique bHLH transcription factor which is activated by binding of planar aromatic hydrocarbons typified by dioxin (TCDD). The active receptor is key to metabolism of aryl hydrocarbon xenobiotics by being a potent inducer of CYP1A1 gene activity. Chlorinated dioxins are inert to metabolism and initiate multifarious toxicities, including potent tumour promotion. These ill-effects are mediated by the activated DR and we are studying the mechanisms by which the ligand binding domain of the DR controls activity of the protein. The DR ligand binding domain resides within a PAS (Per/Arnt/Sim homology) region which is contiguous with the bHLH. The latent bHLH/PAS dioxin receptor (DR) is found in the cytoplasm of most mammalian cell types in a complex with heat shock protein 90, a novel immunophilin like protein termed ARA9/XAP2/AIP, and the co-chaperone p23. Here we use antisense ARA9 constructs to reveal that in the absence of ARA9, the DR is unable to form a transcriptionally active complex. Co-expression of antisense ARA9 with a form of the DR which is constitutively targeted to the nucleus leads to dramatically decreased levels of the nuclear DR protein, implying that ARA9 may function beyond its currently proposed role in cytoplasmic retention of the latent DR

  8. Ligand Receptor-Mediated Regulation of Growth in Plants.

    Science.gov (United States)

    Haruta, Miyoshi; Sussman, Michael R

    2017-01-01

    Growth and development of multicellular organisms are coordinately regulated by various signaling pathways involving the communication of inter- and intracellular components. To form the appropriate body patterns, cellular growth and development are modulated by either stimulating or inhibiting these pathways. Hormones and second messengers help to mediate the initiation and/or interaction of the various signaling pathways in all complex multicellular eukaryotes. In plants, hormones include small organic molecules, as well as larger peptides and small proteins, which, as in animals, act as ligands and interact with receptor proteins to trigger rapid biochemical changes and induce the intracellular transcriptional and long-term physiological responses. During the past two decades, the availability of genetic and genomic resources in the model plant species, Arabidopsis thaliana, has greatly helped in the discovery of plant hormone receptors and the components of signal transduction pathways and mechanisms used by these immobile but highly complex organisms. Recently, it has been shown that two of the most important plant hormones, auxin and abscisic acid (ABA), act through signaling pathways that have not yet been recognized in animals. For example, auxins stimulate cell elongation by bringing negatively acting transcriptional repressor proteins to the proteasome to be degraded, thus unleashing the gene expression program required for increasing cell size. The "dormancy" inducing hormone, ABA, binds to soluble receptor proteins and inhibits a specific class of protein phosphatases (PP2C), which activates phosphorylation signaling leading to transcriptional changes needed for the desiccation of the seeds prior to entering dormancy. While these two hormone receptors have no known animal counterparts, there are also many similarities between animal and plant signaling pathways. For example, in plants, the largest single gene family in the genome is the protein kinase family (approximately 5% of the protein coding genes), although the specific function for only a few dozen of these kinases is clearly established. Recent comparative genomics studies have revealed that parasitic nematodes and pathogenic microbes produce plant peptide hormone mimics that target specific plant plasma membrane receptor-like protein kinases, thus usurping endogenous signaling pathways for their own pathogenic purposes. With biochemical, genetic, and physiological analyses of the regulation of hormone receptor signal pathways, we are thus just now beginning to understand how plants optimize the development of their body shape and cope with constantly changing environmental conditions. © 2017 Elsevier Inc. All rights reserved.

  9. Environmental phthalate monoesters activate pregnane X receptor-mediated transcription

    International Nuclear Information System (INIS)

    Hurst, Christopher H.; Waxman, David J.

    2004-01-01

    Phthalate esters, widely used as plasticizers in the manufacture of products made of polyvinyl chloride, induce reproductive and developmental toxicities in rodents. The mechanism that underlies these effects of phthalate exposure, including the potential role of members of the nuclear receptor superfamily, is not known. The present study investigates the effects of phthalates on the pregnane X receptor (PXR), which mediates the induction of enzymes involved in steroid metabolism and xenobiotic detoxification. The ability of phthalate monoesters to activate PXR-mediated transcription was assayed in a HepG2 cell reporter assay following transfection with mouse PXR (mPXR), human PXR (hPXR), or the hPXR allelic variants V140M, D163G, and A370T. Mono-2-ethylhexyl phthalate (MEHP) increased the transcriptional activity of both mPXR and hPXR (5- and 15-fold, respectively) with EC 50 values of 7-8 μM. mPXR and hPXR were also activated by monobenzyl phthalate (MBzP, up to 5- to 6-fold) but were unresponsive to monomethyl phthalate and mono-n-butyl phthalate (M(n)BP) at the highest concentrations tested (300 μM). hPXR-V140M and hPXR-A370T exhibited patterns of phthalate responses similar to the wild-type receptor. By contrast, hPXR-D163G was unresponsive to all phthalate monoesters tested. Further studies revealed that hPXR-D163G did respond to rifampicin, but required approximately 40-fold higher concentrations than wild-type receptor, suggesting that the ligand-binding domain D163G variant has impaired ligand-binding activity. The responsiveness of PXR to activation by phthalate monoesters demonstrated here suggests that these ubiquitous environmental chemicals may, in part, exhibit their endocrine disruptor activities by altering PXR-regulated steroid hormone metabolism with potential adverse health effects in exposed individuals

  10. Investigating the Regulation of Estrogen Receptor-Mediated Transcription

    National Research Council Canada - National Science Library

    Thackray, Varykina

    2002-01-01

    ...-mediated regulation of specific target genes are still lacking. We have developed an estrogen responsive system in the fruit fly, Drosophila melanogaster in order to explore the functional interactions between ER and other cellular proteins...

  11. Investigating the Regulation of Estrogen Receptor-Mediated Transcription

    National Research Council Canada - National Science Library

    Thackray, Varykina

    2001-01-01

    ...-mediated regulation of specific target genes are still lacking. We have developed an estrogen responsive system in the fruit fly, Drosophila melanogaster in order to explore the functional interactions between ER and other cellular proteins...

  12. Androgen Receptor-Mediated Escape Mechanisms from Androgen Ablation Therapy

    Science.gov (United States)

    2005-10-01

    pathway independent of phosphatidylinositol 3’-kinase and Akt/protein kinase B. Cancer Res 59:1449-53. 158. Manin M, Baron S, Goossens K, et al. 2002...thyroid hormone-stimulated cell proliferation. Mol Cell Biol 21:6626-39 189. Richard -Parpaillon L, Heligon C, Chesnel F, Boujard D, Philpott A 2002 The IGF

  13. Erythropoietin's Beta Common Receptor Mediates Neuroprotection in Spinal Cord Neurons.

    Science.gov (United States)

    Foley, Lisa S; Fullerton, David A; Mares, Joshua; Sungelo, Mitchell; Weyant, Michael J; Cleveland, Joseph C; Reece, T Brett

    2017-12-01

    Paraplegia from spinal cord ischemia-reperfusion (SCIR) remains an elusive and devastating complication of complex aortic operations. Erythropoietin (EPO) attenuates this injury in models of SCIR. Upregulation of the EPO beta common receptor (βcR) is associated with reduced damage in models of neural injury. The purpose of this study was to examine whether EPO-mediated neuroprotection was dependent on βcR expression. We hypothesized that spinal cord neurons subjected to oxygen-glucose deprivation would mimic SCIR injury in aortic surgery and EPO treatment attenuates this injury in a βcR-dependent fashion. Lentiviral vectors with βcR knockdown sequences were tested on neuron cell cultures. The virus with greatest βcR knockdown was selected. Spinal cord neurons from perinatal wild-type mice were harvested and cultured to maturity. They were treated with knockdown or nonsense virus and transduced cells were selected. Three groups (βcR knockdown virus, nonsense control virus, no virus control; n = 8 each) were subjected to 1 hour of oxygen-glucose deprivation. Viability was assessed. βcR expression was quantified by immunoblot. EPO preserved neuronal viability after oxygen-glucose deprivation (0.82 ± 0.04 versus 0.61 ± 0.01; p neuron preservation was similar in the nonsense virus and control mice (0.82 ± 0.04 versus 0.80 ± 0.05; p = 0.77). EPO neuron preservation was lost in βcR knockdown mice compared with nonsense control mice (0.46 ± 0.03 versus 0.80 ± 0.05; p neuronal loss after oxygen-glucose deprivation in a βcR-dependent fashion. This receptor holds immense clinical promise as a target for pharmacotherapies treating spinal cord ischemic injury. Copyright © 2017 The Society of Thoracic Surgeons. Published by Elsevier Inc. All rights reserved.

  14. Molecular identification and biodegradation of 3-chloropropionic acid (3CP by filamentous fungi-Mucor and Trichoderma species isolated from UTM agricultural land

    Directory of Open Access Journals (Sweden)

    Parvizpour, S.

    2013-01-01

    Full Text Available Aims: This study was carried out to further characterize fungal species that could degrade 3-chloropropionic acid (3CPas sole source of carbon and energy. Methodology and Results: Both fungi were able to grow on 3CP after 10 days on solid minimal media. Based on sequencing of its segment of 18S rRNA these isolates were identified as Mucor sp. SP1 and Trichoderma sp. SP2. The isolated strains were not able to grow on media plates containing 10 mM of 2,2-dichloropropionate (2,2DCP as sole source of carbon. 3CP degradation was observed in liquid minimal medium containing 10 mM 3CP after 18 days cultureperiod. The chloride ion released was detected in both growth medium containing Mucor sp. SP1 and Trichoderma sp. SP2. At least 80% of 10 mM 3CP was utilized in the growth medium. Conclusion, significance and impact of study: Dehalogenase enzyme that can degrade α-chloro-substituted haloalkanoic acids for example 2,2DCP is well studied up to protein crystallization. Very few reports on the degradationof β-chloro-substituted haloalkanoic acids such as 3CP and none from fungi. This study is considered important because it can be compared to that of well-documented α-chloro-substituted haloalkanoic acids degradation. This is the first study to indicate fungal growth on 3CP as sole carbon and energy sources.

  15. Localization of tritiated 1-(2, 4-dichlorobenzyl)-1H-indazole-3-carboxylic acid ([3H] AF 1890) in rat testis using freeze-drying autoradiography

    International Nuclear Information System (INIS)

    Hilderbrandt-Stark, H.E.; Mills, J.W.; Fawcett, D.W.

    1982-01-01

    Tritiated 1-(2, 4-dichlorobenzyl)- 1 H-indazole-3-carboxylic acid ([ 3 H]AF 1890) was localized within rat testis using freeze-drying autoradiographic techniques. Ligated seminiferous tubule preparations were exposed in vitro to 80 microCi of [ 3 H]AF 1890 in the absence or presence of excess unlabeled AF 1890 for 30 min at 31 degrees C. Autoradiographs showed a generalized distribution of silver grains over the seminiferous epithelium, in the tubular lumen, and in the interstitial spaces. In addition, clusters of grains were concentrated over the seminiferous epithelium in regions of spermatid heads and tails and in the basal portion of the Sertoli cell cytoplasm, and over Leydig cells in the interstitial space. The generalized distribution was reduced and the grain clusters eliminated by incubation in an excess of unlabeled AF 1890. [ 3 H]inulin was used to assess the effectiveness of the tubular ligation because inulin does not normally penetrate the blood-testis barrier. This extracellular marker was not localized in the tubular lumen or in the seminiferous epithelium. Therefore, the labeling observed with [ 3 H]AF 1890 indicates that this compound crosses the blood-testis barrier

  16. Synthesis and physicochemical properties of the furan dicarboxylic acid, 3-carboxy-4-methyl-5-propyl-2-furanpropanoic acid, an inhibitor of plasma protein binding in uraemia.

    Science.gov (United States)

    Costigan, M G; Gilchrist, T L; Lindup, W E

    1996-06-01

    The furan dicarboxylic acid, 3-carboxy-4-methyl-5-propyl-2-furanpropanoic acid (5-propyl FPA) accumulates in the plasma of patients with chronic renal failure and is a major contributor to the drug binding defect of uraemic plasma. This acid has also been implicated in several other aspects of the uraemic syndrome: anaemia, irregularities of thyroid function, neurological symptoms and inhibition of active tubular secretion. The acid is not commercially available and its synthesis, starting with Meldrum's acid and methyl succinyl chloride, is described. The pKa values were measured by titration and values of 3.2 and 3.6 respectively were assigned to the carboxylic acid groups attached directly to the ring at position 3 and at position 2 (on the side-chain). The partition coefficient (log P) between hydrochloric acid and octanol was 1.2 and the distribution coefficient (log D; octanol-phosphate buffer pH 7.4) was -0.59. The pKa values and the degree of hydrophobic character of 5-propyl FPA are consistent with those of other protein-bound acids which undergo active tubular secretion by the kidney and this substance may serve as an endogenous marker for the effects of drugs and disease on this process.

  17. Protective effect of minocycline, a semi-synthetic second-generation tetracycline against 3-nitropropionic acid (3-NP)-induced neurotoxicity

    International Nuclear Information System (INIS)

    Ahuja, Manuj; Bishnoi, Mahendra; Chopra, Kanwaljit

    2008-01-01

    3-Nitropropionic acid (3-NP) is an irreversible inhibitor of the electron transport enzyme succinate dehydrogenase, a mitochondrial Complex II enzyme. Minocycline is a semi-synthetic second-generation tetracycline with neuroprotective activity and has the capability to effectively cross the blood-brain barrier. We investigated the effects of minocycline on behavioral, biochemical, inflammation related and neurochemical alterations induced by the sub-chronic administration of 3-nitropropionic acid to rats. Chronic pre-administration of minocycline (50 and 100 mg/kg) dose dependently prevented 3-NP-induced dysfunction behavioral (hypoactivity, memory retention, locomotor and rota-rod activity). In addition, 3-NP produced a marked increase in lipid peroxidation levels whereas decreased the activities of catalase and succinate dehydrogenase. In contrast, pretreatment of 3-NP injected rats with minocycline resulted in the attenuation of all these alterations. A marked increase in an inflammatory cytokine TNF-α by 3-NP was also decreased by minocycline treatment. Neurochemically, the administration of 3-NP significantly decreased the levels of catecholamines in the brain homogenates (dopamine, norepinephrine and serotonin) which were reversed by pretreatment of minocycline. The present finding explains the neuroprotective effect of minocycline against 3-NP toxicity by virtue of its antioxidant and anti-inflammatory activity

  18. Effects of self-assembly of 3-phosphonopropionic acid, 3-aminopropyltrimethoxysilane and dopamine on the corrosion behaviors and biocompatibility of a magnesium alloy

    International Nuclear Information System (INIS)

    Pan, Chang-Jiang; Hou, Yu; Wang, Ya-Nan; Gao, Fei; Liu, Tao; Hou, Yan-Hua; Zhu, Yu-Fu; Ye, Wei; Wang, Ling-Ren

    2016-01-01

    Magnesium based alloys are attracting tremendous interests as the novel biodegradable metallic biomaterials. However, the rapid in vivo degradation and the limited surface biocompatibility restrict their clinical applications. Surface modification represents one of the important approaches to control the corrosion rate of Mg based alloys and to enhance the biocompatibility. In the present study, in order to improve the corrosion resistance and surface biocompatibility, magnesium alloy (AZ31B) was modified by the alkali heating treatment followed by the self-assembly of 3-phosphonopropionic acid, 3-aminopropyltrimethoxysilane (APTMS) and dopamine, respectively. The results of attenuated total reflectance Fourier transform infrared spectroscopy (ATR-FTIR) and X-ray photoelectron spectra (XPS) indicated that the molecules were successfully immobilized on the magnesium alloy surface by the self-assembly. An excellent hydrophilic surface was obtained after the alkali heating treatment and the water contact angle increased to some degree after the self-assembly of dopamine, APTMS and 3-phosphonopropionic acid, however, the hydrophilicity of the modified samples was better than that of the pristine magnesium substrate. Due to the formation of the passivation layer after the alkali heating treatment, the corrosion resistance of the magnesium alloy was obviously improved. The corrosion rate further decreased to varying degrees after the self-assembly surface modification. The blood compatibility of the pristine magnesium was significantly improved after the surface modification. The hemolysis rate was reduced from 56% of the blank magnesium alloy to 18% of the alkali heating treated sample and the values were further reduced to about 10% of dopamine-modified sample and 7% of APTMS-modified sample. The hemolysis rate was below 5% for the 3-phosphonopropionic acid modified sample. As compared to the pristine magnesium alloy, fewer platelets were attached and activated on the

  19. Effects of self-assembly of 3-phosphonopropionic acid, 3-aminopropyltrimethoxysilane and dopamine on the corrosion behaviors and biocompatibility of a magnesium alloy

    Energy Technology Data Exchange (ETDEWEB)

    Pan, Chang-Jiang, E-mail: swjtupcj@163.com [Jiangsu Provincial Key Laboratory for Interventional Medical Devices, Huaiyin Institute of Technology, Huai' an 223003 (China); Hou, Yu; Wang, Ya-Nan [Jiangsu Provincial Key Laboratory for Interventional Medical Devices, Huaiyin Institute of Technology, Huai' an 223003 (China); Gao, Fei [Zhejiang Zylox Medical Devices Co., Ltd., Hangzhou 310000 (China); Liu, Tao; Hou, Yan-Hua; Zhu, Yu-Fu; Ye, Wei; Wang, Ling-Ren [Jiangsu Provincial Key Laboratory for Interventional Medical Devices, Huaiyin Institute of Technology, Huai' an 223003 (China)

    2016-10-01

    Magnesium based alloys are attracting tremendous interests as the novel biodegradable metallic biomaterials. However, the rapid in vivo degradation and the limited surface biocompatibility restrict their clinical applications. Surface modification represents one of the important approaches to control the corrosion rate of Mg based alloys and to enhance the biocompatibility. In the present study, in order to improve the corrosion resistance and surface biocompatibility, magnesium alloy (AZ31B) was modified by the alkali heating treatment followed by the self-assembly of 3-phosphonopropionic acid, 3-aminopropyltrimethoxysilane (APTMS) and dopamine, respectively. The results of attenuated total reflectance Fourier transform infrared spectroscopy (ATR-FTIR) and X-ray photoelectron spectra (XPS) indicated that the molecules were successfully immobilized on the magnesium alloy surface by the self-assembly. An excellent hydrophilic surface was obtained after the alkali heating treatment and the water contact angle increased to some degree after the self-assembly of dopamine, APTMS and 3-phosphonopropionic acid, however, the hydrophilicity of the modified samples was better than that of the pristine magnesium substrate. Due to the formation of the passivation layer after the alkali heating treatment, the corrosion resistance of the magnesium alloy was obviously improved. The corrosion rate further decreased to varying degrees after the self-assembly surface modification. The blood compatibility of the pristine magnesium was significantly improved after the surface modification. The hemolysis rate was reduced from 56% of the blank magnesium alloy to 18% of the alkali heating treated sample and the values were further reduced to about 10% of dopamine-modified sample and 7% of APTMS-modified sample. The hemolysis rate was below 5% for the 3-phosphonopropionic acid modified sample. As compared to the pristine magnesium alloy, fewer platelets were attached and activated on the

  20. Lysophosphatidic Acid Regulation and Roles in Human Prostate Cancer

    Science.gov (United States)

    2006-08-01

    IGF-II ( insulin -like growth factor-II), and hormones have been implicated in the growth and survival of prostate cancer cells [1]. A recent addition to...sphingomyelin. In the synthesis of sphingomyelin, a phosphocholine group is transferred from phosphatidylcholine to ceramide. Sphin- gomyelin synthesis...cytotoxicity of TNF-α [78]. A phosphatidylcholine -specific phospholipase C activity was also required for aSMase activa- tion [79,80]. It is assumed that

  1. Pro-lipogenic Action of Lysophosphatidic Acid in Ovarian Cancer

    Science.gov (United States)

    2014-04-01

    Function. IUBMB Life. 2008. 60: 347-354. 11. Tse C, Shoemaker AR, Adickes J, Anderson MG, Chen J, Jin S, Johnson EF, Marsh KC , Mitten MJ, Nimmer P...Ann. Oncol. 9, 437–442 14. Choi, J. W., Herr, D. R., Noguchi, K., Yung, Y. C., Lee, C. W., Mutoh, T., Lin, M. E., Teo , S. T., Park, K. E., Mosley, A

  2. Studies on radiolysis of amino acids, (3)

    International Nuclear Information System (INIS)

    Oku, Tadatake

    1978-01-01

    For the purpose of investigating the radiolysis of amino acids and the safeness to radiation, the radiolytic mechanism and radio-sensitivity of sulfur-containing amino acids in aqueous solution in the presence of air or in the atmosphere of nitrogen were studied. Aqueous solutions of L-methionine, cysteine (both 1mM) and L-cystine (0.3mM) were irradiated with γ-ray of 60 Co at the dose of 4.2 - 2,640 x 10 3 rad. The amino acids and the radiolytic products were determined with an amino acid analyzer. The volatile sulfur compounds formed from γ-irradiated methionine were estimated by a flame photometric detector-gas chromatograph. From the results obtained, G values of the radiolysis of sulfur-containing amino acids and the products were calculated, and the radiolytic mechanisms of methionine, cysteine and cystine were proposed. The radio-sensitivity of sulfur-containing amino acids was shown as follows: cysteine (C3-SH) > methionine (C5, -SCH 3 ) > cystine (C 6 , -S-S-). Off-flavor development from γ-irradiated methionine when oxidizing agent was added was less than that when reducing agent was added. (Kobatake, H.)

  3. Hindbrain GLP-1 receptor mediation of cisplatin-induced anorexia and nausea.

    Science.gov (United States)

    De Jonghe, Bart C; Holland, Ruby A; Olivos, Diana R; Rupprecht, Laura E; Kanoski, Scott E; Hayes, Matthew R

    2016-01-01

    While chemotherapy-induced nausea and vomiting are clinically controlled in the acute (anorexia, nausea, fatigue, and other illness-type behaviors during the delayed phase (>24 h) of chemotherapy are largely uncontrolled. As the hindbrain glucagon-like peptide-1 (GLP-1) system contributes to energy balance and mediates aversive and stressful stimuli, here we examine the hypothesis that hindbrain GLP-1 signaling mediates aspects of chemotherapy-induced nausea and reductions in feeding behavior in rats. Specifically, hindbrain GLP-1 receptor (GLP-1R) blockade, via 4th intracerebroventricular (ICV) exendin-(9-39) injections, attenuates the anorexia, body weight reduction, and pica (nausea-induced ingestion of kaolin clay) elicited by cisplatin chemotherapy during the delayed phase (48 h) of chemotherapy-induced nausea. Additionally, the present data provide evidence that the central GLP-1-producing preproglucagon neurons in the nucleus tractus solitarius (NTS) of the caudal brainstem are activated by cisplatin during the delayed phase of chemotherapy-induced nausea, as cisplatin led to a significant increase in c-Fos immunoreactivity in NTS GLP-1-immunoreactive neurons. These data support a growing body of literature suggesting that the central GLP-1 system may be a potential pharmaceutical target for adjunct anti-emetics used to treat the delayed-phase of nausea and emesis, anorexia, and body weight loss that accompany chemotherapy treatments. Copyright © 2015 Elsevier Inc. All rights reserved.

  4. Bovine lactoferrin counteracts Toll-like receptor mediated activation signals in antigen presenting cells.

    Directory of Open Access Journals (Sweden)

    Patrizia Puddu

    Full Text Available Lactoferrin (LF, a key element in mammalian immune system, plays pivotal roles in host defence against infection and excessive inflammation. Its protective effects range from direct antimicrobial activities against a large panel of microbes, including bacteria, viruses, fungi and parasites, to antinflammatory and anticancer activities. In this study, we show that monocyte-derived dendritic cells (MD-DCs generated in the presence of bovine LF (bLF fail to undergo activation by up-modulating CD83, co-stimulatory and major histocompatibility complex molecules, and cytokine/chemokine secretion. Moreover, these cells are weak activators of T cell proliferation and retain antigen uptake activity. Consistent with an impaired maturation, bLF-MD-DC primed T lymphocytes exhibit a functional unresponsiveness characterized by reduced expression of CD154 and impaired expression of IFN-γ and IL-2. The observed imunosuppressive effects correlate with an increased expression of molecules with negative regulatory functions (i.e. immunoglobulin-like transcript 3 and programmed death ligand 1, indoleamine 2,3-dioxygenase, and suppressor of cytokine signaling-3. Interestingly, bLF-MD-DCs produce IL-6 and exhibit constitutive signal transducer and activator of transcription 3 activation. Conversely, bLF exposure of already differentiated MD-DCs completely fails to induce IL-6, and partially inhibits Toll-like receptor (TLR agonist-induced activation. Cell-specific differences in bLF internalization likely account for the distinct response elicited by bLF in monocytes versus immature DCs, providing a mechanistic base for its multiple effects. These results indicate that bLF exerts a potent anti-inflammatory activity by skewing monocyte differentiation into DCs with impaired capacity to undergo activation and to promote Th1 responses. Overall, these bLF-mediated effects may represent a strategy to block excessive DC activation upon TLR-induced inflammation, adding further evidence for a critical role of bLF in directing host immune function.

  5. Effect of lipopolysaccharide and antidepressant drugs on glucocorticoid receptor-mediated gene transcription

    Czech Academy of Sciences Publication Activity Database

    Budziszewska, B.; Basta-Kaim, A.; Kubera, M.; Jaworska, L.; Leskiewicz, M.; Tetich, M.; Otczyk, M.; Zajícová, Alena; Holáň, Vladimír; Lasoń, W.

    2005-01-01

    Roč. 57, č. 4 (2005), s. 540-544 ISSN 1734-1140 Grant - others:State Committee for Scientific Research (KBN)(PL) 6P05A076 Institutional research plan: CEZ:AV0Z5052915 Keywords : glucocorticoid receptor * antidepressant drugs * interleukin-6 Subject RIV: EB - Genetics ; Molecular Biology

  6. Ultradian hormone stimulation induces glucocorticoid receptor-mediated pulses of gene transcription.

    Science.gov (United States)

    Stavreva, Diana A; Wiench, Malgorzata; John, Sam; Conway-Campbell, Becky L; McKenna, Mervyn A; Pooley, John R; Johnson, Thomas A; Voss, Ty C; Lightman, Stafford L; Hager, Gordon L

    2009-09-01

    Studies on glucocorticoid receptor (GR) action typically assess gene responses by long-term stimulation with synthetic hormones. As corticosteroids are released from adrenal glands in a circadian and high-frequency (ultradian) mode, such treatments may not provide an accurate assessment of physiological hormone action. Here we demonstrate that ultradian hormone stimulation induces cyclic GR-mediated transcriptional regulation, or gene pulsing, both in cultured cells and in animal models. Equilibrium receptor-occupancy of regulatory elements precisely tracks the ligand pulses. Nascent RNA transcripts from GR-regulated genes are released in distinct quanta, demonstrating a profound difference between the transcriptional programs induced by ultradian and constant stimulation. Gene pulsing is driven by rapid GR exchange with response elements and by GR recycling through the chaperone machinery, which promotes GR activation and reactivation in response to the ultradian hormone release, thus coupling promoter activity to the naturally occurring fluctuations in hormone levels. The GR signalling pathway has been optimized for a prompt and timely response to fluctuations in hormone levels, indicating that biologically accurate regulation of gene targets by GR requires an ultradian mode of hormone stimulation.

  7. VIP/PACAP receptor mediation of cutaneous active vasodilation during heat stress in humans.

    Science.gov (United States)

    Kellogg, Dean L; Zhao, Joan L; Wu, Yubo; Johnson, John M

    2010-07-01

    Vasoactive intestinal peptide (VIP) is implicated in cutaneous active vasodilation in humans. VIP and the closely related pituitary adenylate cyclase activating peptide (PACAP) act through several receptor types: VIP through VPAC1 and VPAC2 receptors and PACAP through VPAC1, VPAC2, and PAC1 receptors. We examined participation of VPAC2 and/or PAC1 receptors in cutaneous vasodilation during heat stress by testing the effects of their specific blockade with PACAP6-38. PACAP6-38 dissolved in Ringer's was administered by intradermal microdialysis at one forearm site while a control site received Ringer's solution. Skin blood flow was monitored by laser-Doppler flowmetry (LDF). Blood pressure was monitored noninvasively and cutaneous vascular conductance (CVC) calculated. A 5- to 10-min baseline period was followed by approximately 70 min of PACAP6-38 (100 microM) perfusion at one site in normothermia and a 3-min period of body cooling. Whole body heating was then performed to engage cutaneous active vasodilation and was maintained until CVC had plateaued at an elevated level at all sites for 5-10 min. Finally, 58 mM sodium nitroprusside was perfused through both microdialysis sites to effect maximal vasodilation. No CVC differences were found between control and PACAP6-38-treated sites during normothermia (19 +/- 3%max untreated vs. 20 +/- 3%max, PACAP6-38 treated; P > 0.05 between sites) or cold stress (11 +/- 2%max untreated vs. 10 +/- 2%max, PACAP6-38 treated, P > 0.05 between sites). PACAP6-38 attenuated the increase in CVC during whole body heating when compared with untreated sites (59 +/- 3%max untreated vs. 46 +/- 3%max, PACAP6-38 treated, P < 0.05). We conclude that VPAC2 and/or PAC1 receptor activation is involved in cutaneous active vasodilation in humans.

  8. β-Receptor-mediated increase in cerebral blood flow during hypoglycemia

    International Nuclear Information System (INIS)

    Hollinger, B.R.; Bryan, R.M.

    1987-01-01

    The authors tested the hypothesis that β-adrenergic receptor stimulation is involved with the increase in regional cerebral blood flow (rCBF) during hypoglycemia. Rats were surgically prepared with the use of halothane-nitrous oxide anesthesia. A plaster restraining cast was placed around the hindquarters, and anesthesia was discontinued. Hypoglycemia was produced by an intravenous injection of insulin; normoglycemic control rates were given saline. Propranolol was administered to some control and some hypoglycemic rats to block the β-adrenergic receptors. Regional CBF was measured using 4-[N-methyl- 14 C]iodoantipyrine. Regional CBF increased during hypoglycemia in rats that were not treated with propranolol. The increase varied from ∼60 to 200% depending on the brain region. During hypoglycemia, propranolol abolished the increase in rCBF in the hypothalamus, cerebellum, and pyramidal tract. In other regions the increase in rCBF was only 33-65% of the increase in hypoglycemic rats that were not treated with propranolol. They conclude that β-receptor stimulation plays a major role in the increase in rCBF during hypoglycemia

  9. Probenecid inhibits α-adrenergic receptor-mediated vasoconstriction in the human leg vasculature

    DEFF Research Database (Denmark)

    Nyberg, Michael Permin; Piil, Peter Bergmann; Kiehn, Oliver Thistrup

    2018-01-01

    to α1- and α2-adrenergic receptor stimulation in the human forearm and leg vasculature of young healthy male subjects (23±3 years). By use of immunolabeling and confocal microscopy, Panx1 channels were found to be expressed in vascular smooth muscle cells of arterioles in human leg skeletal muscle....... Probenecid treatment increased (Padrenergic receptor stimulation) by ≈15%, whereas the response to the α1-agonist phenylephrine was unchanged. Inhibition...

  10. β-Adrenergic receptor-mediated suppression of interleukin 2 receptors in human lymphocytes

    International Nuclear Information System (INIS)

    Feldman, R.D.; Hunninghake, G.W.; McArdle, W.L.

    1987-01-01

    Adrenergic receptor agonists are know to attenuate the proliferative response of human lymphocytes after activation; however, their mechanism of action is unknown. Since expression of interleukin 2 (IL-2) receptors is a prerequisite for proliferation, the effect of β-adrenergic receptor agonists on lymphocyte IL-2 receptors was studied on both mitogen-stimulated lymphocytes and IL-2-dependent T lymphocyte cell lines. In both cell types the β-adrenergic receptor agonist isoproterenol blocked the expression of IL-2 receptors, as determined with the IL-2 receptor anti-TAC antibody. To determine the effect of β-adrenergic agonists on expression of the high affinity IL-2 receptors, [ 125 I]IL-2 binding studies were performed at concentrations selective for high affinity sites. No significant effect of β-adrenergic agonists on high affinity IL-2 receptor sites could be detected. The data demonstrate that β-adrenergic receptor agonists down-regulate IL-2 receptors primarily affecting low affinity sites

  11. β2-adrenergic receptor-mediated negative regulation of group 2 innate lymphoid cell responses.

    Science.gov (United States)

    Moriyama, Saya; Brestoff, Jonathan R; Flamar, Anne-Laure; Moeller, Jesper B; Klose, Christoph S N; Rankin, Lucille C; Yudanin, Naomi A; Monticelli, Laurel A; Putzel, Gregory Garbès; Rodewald, Hans-Reimer; Artis, David

    2018-03-02

    The type 2 inflammatory response is induced by various environmental and infectious stimuli. Although recent studies identified group 2 innate lymphoid cells (ILC2s) as potent sources of type 2 cytokines, the molecular pathways controlling ILC2 responses are incompletely defined. Here we demonstrate that murine ILC2s express the β 2 -adrenergic receptor (β 2 AR) and colocalize with adrenergic neurons in the intestine. β 2 AR deficiency resulted in exaggerated ILC2 responses and type 2 inflammation in intestinal and lung tissues. Conversely, β 2 AR agonist treatment was associated with impaired ILC2 responses and reduced inflammation in vivo. Mechanistically, we demonstrate that the β 2 AR pathway is a cell-intrinsic negative regulator of ILC2 responses through inhibition of cell proliferation and effector function. Collectively, these data provide the first evidence of a neuronal-derived regulatory circuit that limits ILC2-dependent type 2 inflammation. Copyright © 2018 The Authors, some rights reserved; exclusive licensee American Association for the Advancement of Science. No claim to original U.S. Government Works.

  12. Serotonin(2) receptors mediate respiratory recovery after cervical spinal cord hemisection in adult rats.

    Science.gov (United States)

    Zhou, S Y; Basura, G J; Goshgarian, H G

    2001-12-01

    The aim of the present study was to specifically investigate the involvement of serotonin [5-hydroxytryptamine (5-HT(2))] receptors in 5-HT-mediated respiratory recovery after cervical hemisection. Experiments were conducted on C(2) spinal cord-hemisected, anesthetized (chloral hydrate, 400 mg/kg ip), vagotomized, pancuronium- paralyzed, and artificially ventilated female Sprague-Dawley rats in which CO(2) levels were monitored and maintained. Twenty-four hours after spinal hemisection, the ipsilateral phrenic nerve displayed no respiratory-related activity indicative of a functionally complete hemisection. Intravenous administration of the 5-HT(2A/2C)-receptor agonist (+/-)-2,5-dimethoxy-4-iodoamphetamine hydrochloride (DOI) induced respiratory-related activity in the phrenic nerve ipsilateral to hemisection under conditions in which CO(2) was maintained at constant levels and augmented the activity induced under conditions of hypercapnia. The effects of DOI were found to be dose dependent, and the recovery of activity could be maintained for up to 2 h after a single injection. DOI-induced recovery was attenuated by the 5-HT(2)-receptor antagonist ketanserin but not with the 5-HT(2C)-receptor antagonist RS-102221, suggesting that 5-HT(2A) and not necessarily 5-HT(2C) receptors may be involved in the induction of respiratory recovery after cervical spinal cord injury.

  13. Role of glucocorticoid receptor-mediated mechanisms in cocaine memory enhancement.

    Science.gov (United States)

    Stringfield, S J; Higginbotham, J A; Wang, R; Berger, A L; McLaughlin, R J; Fuchs, R A

    2017-09-01

    The basolateral amygdala (BLA) is a critical site for the reconsolidation of labile contextual cocaine memories following retrieval-induced reactivation/destabilization. Here, we examined whether glucocorticoid receptors (GR), which are abundant in the BLA, mediate this phenomenon. Rats were trained to lever press for cocaine reinforcement in a distinct environmental context, followed by extinction training in a different context. Rats were then briefly exposed to the cocaine-paired context (to elicit memory reactivation and reconsolidation) or their home cages (no reactivation control). Exposure to the cocaine-paired context elicited greater serum corticosterone concentrations than home cage stay. Interestingly, the GR antagonist, mifepristone (3-10 ng/hemisphere), administered into the BLA after memory reactivation produced a further, dose-dependent increase in serum corticosterone concentrations during the putative time of cocaine-memory reconsolidation but produced an inverted U-shaped dose-effect curve on subsequent cocaine-seeking behavior 72 h later. This effect was anatomically selective, dependent on memory reactivation (i.e., not observed after home cage exposure), and did not reflect protracted hyperactivity. However, the effect was also observed when mifepristone was administered after novelty stress that mimics drug context-induced hypothalamic-pituitary-adrenal (HPA) axis activation without explicit memory reactivation. Together, these findings suggest that, similar to explicit memory retrieval, a stressful event is sufficient to destabilize cocaine memories and permit their manipulation. Furthermore, BLA GR stimulation exerts inhibitory feedback upon HPA axis activation and thus suppresses cocaine-memory reconsolidation. Copyright © 2017 Elsevier Ltd. All rights reserved.

  14. High Throughput Synthesis and Screening for Agents Inhibiting Androgen Receptor Mediated Gene Transcription

    National Research Council Canada - National Science Library

    Boger, Dale L

    2005-01-01

    .... This entails the high throughput synthesis of DNA binding agents related to distamycin, their screening for binding to androgen response elements using a new high throughput DNA binding screen...

  15. High Throughout Synthesis and Screening for Agents Inhibiting Androgen Receptor Mediated Gene Transcription

    National Research Council Canada - National Science Library

    Boger, Dale

    2003-01-01

    .... This entails the high throughput synthesis of DNA binding agents related to distamycin, their screening for binding to androgen response elements using a new high throughput DNA binding screen...

  16. High Throughput Synthesis and Screening for Agents Inhibiting Androgen Receptor Mediated Gene Transcription

    National Research Council Canada - National Science Library

    Boger, Dale

    2004-01-01

    .... This entails the high throughput synthesis of DNA binding agents related to distamycin, their screening for binding to androgen response elements using a new high throughput DNA binding screen...

  17. Molecular and Biochemical Effects of a Kola Nut Extract on Androgen Receptor-Mediated Pathways

    Directory of Open Access Journals (Sweden)

    Rajasree Solipuram

    2009-01-01

    Full Text Available The low incidence of prostate cancer in Asians has been attributed to chemopreventative properties of certain chemicals found in their diet. This study characterized the androgenic and chemopreventative properties of the Jamaican bush tea “Bizzy,” using androgen receptor positive and negative cell lines. Exposure of prostate cells to Biz-2 resulted in a growth inhibition (GI50 of 15 ppm in LNCaP cells and 3.6 ppm in DU145 cells. Biz-2 elicited a 2-fold increase in the mRNA of the anti-apoptotic gene Bcl2, with a 10-fold increase in that of the proapoptotic gene Bax. We observed a 2.4- to 7.5-fold change in apoptotic cells in both cell lines. Biz-2 at 10 ppm elicited a time- and dose-dependent stimulation of both the protein and mRNA levels of several androgen-regulated genes. Biz-2 caused a 36% decrease in PSA secretion and a significant increase in PSA mRNA. The relative binding affinity (IC50 of Biz-2 for AR was 2- to 5-fold lower than that of the synthetic androgen R1881. Biz-2 was found to be a specific ligand for the AR in that the natural ligand, DHT, and the anti-androgen, flutamide, displaced Biz-2 bound to AR and inhibited Biz-2-induced transcription and PSA secretion. This study provided evidence that Biz-2 extract possesses the ability to modulate prostate cancer cell biology in an AR-dependent manner.

  18. Kappa Opioid Receptors Mediate where Fear Is Expressed Following Extinction Training

    Science.gov (United States)

    Cole, Sindy; Richardson, Rick; McNally, Gavan P.

    2011-01-01

    Six experiments used a within-subjects renewal design to examine the involvement of kappa opioid receptors (KORs) in regulating the expression and recovery of extinguished fear. Rats were trained to fear a tone conditioned stimulus (CS) via pairings with foot shock in a distinctive context (A). This was followed by extinction training of the CS in…

  19. The CRF Family of Neuropeptides and their Receptors - Mediators of the Central Stress Response

    Science.gov (United States)

    Dedic, Nina; Chen, Alon; Deussing, Jan M.

    2018-01-01

    Background: Dysregulated stress neurocircuits, caused by genetic and/or environmental changes, underlie the development of many neuropsychiatric disorders. Corticotropin-releasing factor (CRF) is the major physiological activator of the hypothalamic-pituitary-adrenal (HPA) axis and conse-quently a primary regulator of the mammalian stress response. Together with its three family members, urocortins (UCNs) 1, 2, and 3, CRF integrates the neuroendocrine, autonomic, metabolic and behavioral responses to stress by activating its cognate receptors CRFR1 and CRFR2. Objective: Here we review the past and current state of the CRF/CRFR field, ranging from pharmacologi-cal studies to genetic mouse models and virus-mediated manipulations. Results: Although it is well established that CRF/CRFR1 signaling mediates aversive responses, includ-ing anxiety and depression-like behaviors, a number of recent studies have challenged this viewpoint by revealing anxiolytic and appetitive properties of specific CRF/CRFR1 circuits. In contrast, the UCN/CRFR2 system is less well understood and may possibly also exert divergent functions on physiol-ogy and behavior depending on the brain region, underlying circuit, and/or experienced stress conditions. Conclusion: A plethora of available genetic tools, including conventional and conditional mouse mutants targeting CRF system components, has greatly advanced our understanding about the endogenous mecha-nisms underlying HPA system regulation and CRF/UCN-related neuronal circuits involved in stress-related behaviors. Yet, the detailed pathways and molecular mechanisms by which the CRF/UCN-system translates negative or positive stimuli into the final, integrated biological response are not completely un-derstood. The utilization of future complementary methodologies, such as cell-type specific Cre-driver lines, viral and optogenetic tools will help to further dissect the function of genetically defined CRF/UCN neurocircuits in the context of adaptive and maladaptive stress responses. PMID:28260504

  20. Astrocytic β2-adrenergic receptors mediate hippocampal long-term memory consolidation

    KAUST Repository

    Gao, Virginia; Suzuki, Akinobu; Magistretti, Pierre J.; Lengacher, Sylvain; Pollonini, Gabriella; Steinman, Michael Q.; Alberini, Cristina M.

    2016-01-01

    Emotionally relevant experiences form strong and long-lasting memories by critically engaging the stress hormone/neurotransmitter noradrenaline, which mediates and modulates the consolidation of these memories. Noradrenaline acts through adrenergic receptors (ARs), of which β2- Adrenergic receptors (βARs) are of particular importance. The differential anatomical and cellular distribution of βAR subtypes in the brain suggests that they play distinct roles in memory processing, although much about their specific contributions and mechanisms of action remains to be understood. Here we show that astrocytic rather than neuronal β2ARs in the hippocampus play a key role in the consolidation of a fear-based contextual memory. These hippocampal β2ARs, but not β1ARs, are coupled to the training-dependent release of lactate from astrocytes, which is necessary for long- Term memory formation and for underlying molecular changes. This key metabolic role of astrocytic β2ARs may represent a novel target mechanism for stress-related psychopathologies and neurodegeneration.

  1. Rab GTPases Regulate Endothelial Cell Protein C Receptor-Mediated Endocytosis and Trafficking of Factor VIIa

    Science.gov (United States)

    Nayak, Ramesh C.; Keshava, Shiva; Esmon, Charles T.; Pendurthi, Usha R.; Rao, L. Vijaya Mohan

    2013-01-01

    Recent studies have established that factor VIIa (FVIIa) binds to the endothelial cell protein C receptor (EPCR). FVIIa binding to EPCR may promote the endocytosis of this receptor/ligand complex. Rab GTPases are known to play a crucial role in the endocytic and exocytic pathways of receptors or receptor/ligand complexes. The present study was undertaken to investigate the role of Rab GTPases in the intracellular trafficking of EPCR and FVIIa. CHO-EPCR cells and human umbilical vein endothelial cells (HUVEC) were transduced with recombinant adenoviral vectors to express wild-type, constitutively active, or dominant negative mutant of various Rab GTPases. Cells were exposed to FVIIa conjugated with AF488 fluorescent probe (AF488-FVIIa), and intracellular trafficking of FVIIa, EPCR, and Rab proteins was evaluated by immunofluorescence confocal microscopy. In cells expressing wild-type or constitutively active Rab4A, internalized AF488-FVIIa accumulated in early/sorting endosomes and its entry into the recycling endosomal compartment (REC) was inhibited. Expression of constitutively active Rab5A induced large endosomal structures beneath the plasma membrane where EPCR and FVIIa accumulated. Dominant negative Rab5A inhibited the endocytosis of EPCR-FVIIa. Expression of constitutively active Rab11 resulted in retention of accumulated AF488-FVIIa in the REC, whereas expression of a dominant negative form of Rab11 led to accumulation of internalized FVIIa in the cytoplasm and prevented entry of internalized FVIIa into the REC. Expression of dominant negative Rab11 also inhibited the transport of FVIIa across the endothelium. Overall our data show that Rab GTPases regulate the internalization and intracellular trafficking of EPCR-FVIIa. PMID:23555015

  2. Probing multivalency in ligand–receptor-mediated adhesion of soft, biomimetic interfaces

    Directory of Open Access Journals (Sweden)

    Stephan Schmidt

    2015-05-01

    Full Text Available Many biological functions at cell level are mediated by the glycocalyx, a dense carbohydrate-presenting layer. In this layer specific interactions between carbohydrate ligands and protein receptors are formed to control cell–cell recognition, cell adhesion and related processes. The aim of this work is to shed light on the principles of complex formation between surface anchored carbohydrates and receptor surfaces by measuring the specific adhesion between surface bound mannose on a concanavalin A (ConA layer via poly(ethylene glycol-(PEG-based soft colloidal probes (SCPs. Special emphasis is on the dependence of multivalent presentation and density of carbohydrate units on specific adhesion. Consequently, we first present a synthetic strategy that allows for controlled density variation of functional groups on the PEG scaffold using unsaturated carboxylic acids (crotonic acid, acrylic acid, methacrylic acid as grafting units for mannose conjugation. We showed by a range of analytic techniques (ATR–FTIR, Raman microscopy, zeta potential and titration that this synthetic strategy allows for straightforward variation in grafting density and grafting length enabling the controlled presentation of mannose units on the PEG network. Finally we determined the specific adhesion of PEG-network-conjugated mannose units on ConA surfaces as a function of density and grafting type. Remarkably, the results indicated the absence of a molecular-level enhancement of mannose/ConA interaction due to chelate- or subsite-binding. The results seem to support the fact that weak carbohydrate interactions at mechanically flexible interfaces hardly undergo multivalent binding but are simply mediated by the high number of ligand–receptor interactions.

  3. Basolateral amygdala GABA-A receptors mediate stress-induced memory retrieval impairment in rats.

    Science.gov (United States)

    Sardari, Maryam; Rezayof, Ameneh; Khodagholi, Fariba; Zarrindast, Mohammad-Reza

    2014-04-01

    The present study was designed to investigate the involvement of GABA-A receptors of the basolateral amygdala (BLA) in the impairing effect of acute stress on memory retrieval. The BLAs of adult male Wistar rats were bilaterally cannulated and memory retrieval was measured in a step-through type passive avoidance apparatus. Acute stress was evoked by placing the animals on an elevated platform for 10, 20 and 30 min. The results indicated that exposure to 20 and 30 min stress, but not 10 min, before memory retrieval testing (pre-test exposure to stress) decreased the step-through latency, indicating stress-induced memory retrieval impairment. Intra-BLA microinjection of a GABA-A receptor agonist, muscimol (0.005-0.02 μg/rat), 5 min before exposure to an ineffective stress (10 min exposure to stress) induced memory retrieval impairment. It is important to note that pre-test intra-BLA microinjection of the same doses of muscimol had no effect on memory retrieval in the rats unexposed to 10 min stress. The blockade of GABA-A receptors of the BLA by injecting an antagonist, bicuculline (0.4-0.5 μg/rat), 5 min before 20 min exposure to stress, prevented stress-induced memory retrieval. Pre-test intra-BLA microinjection of the same doses of bicuculline (0.4-0.5 μg/rat) in rats unexposed to 20 min stress had no effect on memory retrieval. In addition, pre-treatment with bicuculline (0.1-0.4 μg/rat, intra-BLA) reversed muscimol (0.02 μg/rat, intra-BLA)-induced potentiation on the effect of stress in passive avoidance learning. It can be concluded that pre-test exposure to stress can induce memory retrieval impairment and the BLA GABA-A receptors may be involved in stress-induced memory retrieval impairment.

  4. Insulin receptors mediate growth effects in cultured fetal neurons. I. Rapid stimulation of protein synthesis

    International Nuclear Information System (INIS)

    Heidenreich, K.A.; Toledo, S.P.

    1989-01-01

    In this study we have examined the effects of insulin on protein synthesis in cultured fetal chick neurons. Protein synthesis was monitored by measuring the incorporation of [3H]leucine (3H-leu) into trichloroacetic acid (TCA)-precipitable protein. Upon addition of 3H-leu, there was a 5-min lag before radioactivity occurred in protein. During this period cell-associated radioactivity reached equilibrium and was totally recovered in the TCA-soluble fraction. After 5 min, the incorporation of 3H-leu into protein was linear for 2 h and was inhibited (98%) by the inclusion of 10 micrograms/ml cycloheximide. After 24 h of serum deprivation, insulin increased 3H-leu incorporation into protein by approximately 2-fold. The stimulation of protein synthesis by insulin was dose dependent (ED50 = 70 pM) and seen within 30 min. Proinsulin was approximately 10-fold less potent than insulin on a molar basis in stimulating neuronal protein synthesis. Insulin had no effect on the TCA-soluble fraction of 3H-leu at any time and did not influence the uptake of [3H]aminoisobutyric acid into neurons. The isotope ratio of 3H-leu/14C-leu in the leucyl tRNA pool was the same in control and insulin-treated neurons. Analysis of newly synthesized proteins by sodium dodecyl sulfate-polyacrylamide gel electrophoresis revealed that insulin uniformly increased the incorporation of 14C-leu into all of the resolved neuronal proteins. We conclude from these data that (1) insulin rapidly stimulates overall protein synthesis in fetal neurons independent of amino acid uptake and aminoacyl tRNA precursor pools; (2) stimulation of protein synthesis is mediated by the brain subtype of insulin receptor; and (3) insulin is potentially an important in vivo growth factor for fetal central nervous system neurons

  5. Pattern recognition receptor-mediated cytokine response in infants across 4 continents.

    Science.gov (United States)

    Smolen, Kinga K; Ruck, Candice E; Fortuno, Edgardo S; Ho, Kevin; Dimitriu, Pedro; Mohn, William W; Speert, David P; Cooper, Philip J; Esser, Monika; Goetghebuer, Tessa; Marchant, Arnaud; Kollmann, Tobias R

    2014-03-01

    Susceptibility to infection as well as response to vaccination varies among populations. To date, the underlying mechanisms responsible for these clinical observations have not been fully delineated. Because innate immunity instructs adaptive immunity, we hypothesized that differences between populations in innate immune responses may represent a mechanistic link to variation in susceptibility to infection or response to vaccination. Determine whether differences in innate immune responses exist among infants from different continents of the world. We determined the innate cytokine response following pattern recognition receptor (PRR) stimulation of whole blood from 2-year-old infants across 4 continents (Africa, North America, South America, and Europe). We found that despite the many possible genetic and environmental exposure differences in infants across 4 continents, innate cytokine responses were similar for infants from North America, South America, and Europe. However, cells from South African infants secreted significantly lower levels of cytokines than did cells from infants from the 3 other sites, and did so following stimulation of extracellular and endosomal but not cytosolic PRRs. Substantial differences in innate cytokine responses to PRR stimulation exist among different populations of infants that could not have been predicted. Delineating the underlying mechanism(s) for these differences will not only aid in improving vaccine-mediated protection but possibly also provide clues for the susceptibility to infection in different regions of the world. Copyright © 2013 The Authors. Published by Mosby, Inc. All rights reserved.

  6. Scavenger receptor-mediated endocytosis by sinusoidal cells in rat bone marrow

    International Nuclear Information System (INIS)

    Geoffroy, J.S.

    1987-01-01

    Endocytosis of serum albumin by sinusoidal endothelial cells in rat bone marrow was investigated initially at the ultrastructural level with subsequent biochemical investigation of the specificity mediating this event. Bovine serum albumin adsorbed to 20nm colloidal gold particles (AuBSA) was chosen as the electron microscopic probe. Morphological data strongly suggested that a receptor was involved in uptake of AuBSA. Confirmation of receptor involvement in the uptake of AuBSA by marrow sinusoidal endothelial cells was achieved utilizing an in situ isolated hind limb perfusion protocol in conjunction with unlabeled, radiolabeled, and radio-/colloidal gold labeled probes. The major findings of competition and saturation experiments were: (1) endocytosis of AuBSA was mediated by a receptor for modified/treated serum albumin; (2) endocytosis of formaldehyde-treated serum albumin was mediated by a binding site which may be the same or closely related to the site responsible for the uptake of AuBSA; and (3) endocytosis of native untreated albumin was not mediated by receptor and probably represents fluid-phase pinocitosis

  7. Presynaptic CRF1 Receptors Mediate the Ethanol Enhancement of GABAergic Transmission in the Mouse Central Amygdala

    Directory of Open Access Journals (Sweden)

    Zhiguo Nie

    2009-01-01

    Full Text Available Corticotropin-releasing factor (CRF is a 41-amino-acid neuropeptide involved in stress responses initiated from several brain areas, including the amygdala formation. Research shows a strong relationship between stress, brain CRF, and excessive alcohol consumption. Behavioral studies suggest that the central amygdala (CeA is significantly involved in alcohol reward and dependence. We recently reported that the ethanol augmentation of GABAergic synaptic transmission in rat CeA involves CRF1 receptors, because both CRF and ethanol significantly enhanced the amplitude of evoked GABAergic inhibitory postsynaptic currents (IPSCs in CeA neurons from wild-type (WT and CRF2 knockout (KO mice, but not in neurons of CRF1 KO mice. The present study extends these findings using selective CRF receptor ligands, gene KO models, and miniature IPSC (mIPSC analysis to assess further a presynaptic role for the CRF receptors in mediating ethanol effects in the CeA. In whole-cell patch recordings of pharmacologically isolated GABAAergic IPSCs from slices of mouse CeA, both CRF and ethanol augmented evoked IPSCs in a concentration-dependent manner, with low EC50s. A CRF1 (but not CRF2 KO construct and the CRF1-selective nonpeptide antagonist NIH-3 (LWH-63 blocked the augmenting effect of both CRF and ethanol on evoked IPSCs. Furthermore, the new selective CRF1 agonist stressin1, but not the CRF2 agonist urocortin 3, also increased evoked IPSC amplitudes. Both CRF and ethanol decreased paired-pulse facilitation (PPF of evoked IPSCs and significantly enhanced the frequency, but not the amplitude, of spontaneous miniature GABAergic mIPSCs in CeA neurons of WT mice, suggesting a presynaptic site of action. The PPF effect of ethanol was abolished in CeA neurons of CRF1 KO mice. The CRF1 antagonist NIH-3 blocked the CRF- and ethanol-induced enhancement of mIPSC frequency in CeA neurons. These data indicate that presynaptic CRF1 receptors play a critical role in permitting or mediating ethanol enhancement of GABAergic synaptic transmission in CeA, via increased vesicular GABA release, and thus may be a rational target for the treatment of alcohol abuse and alcoholism.

  8. Bradykinin B2 receptor-mediated phosphoinositide hydrolysis in bovine cultured tracheal smooth muscle cells.

    OpenAIRE

    Marsh, K. A.; Hill, S. J.

    1992-01-01

    1. Bovine tracheal smooth muscle cells were established in culture to study agonist-induced phosphoinositide (PI) hydrolysis in this tissue. 2. Bradykinin (0.1 nM-10 microM) evoked a concentration-dependent increase (log EC50 (M) = -9.4 +/- 0.2; n = 8) in the accumulation of total [3H]-inositol phosphates in cultured tracheal smooth muscle cells whereas the selective B1 receptor agonist des-Arg9-bradykinin (10 microM) was significantly less effective (16% of bradykinin maximal response; relat...

  9. Hippocampal GluA1-containing AMPA receptors mediate context-dependent sensitization to morphine.

    Science.gov (United States)

    Xia, Yan; Portugal, George S; Fakira, Amanda K; Melyan, Zara; Neve, Rachael; Lee, H Thomas; Russo, Scott J; Liu, Jie; Morón, Jose A

    2011-11-09

    Glutamatergic systems, including AMPA receptors (AMPARs), are involved in opiate-induced neuronal and behavioral plasticity, although the mechanisms underlying these effects are not fully understood. In the present study, we investigated the effects of repeated morphine administration on AMPAR expression, synaptic plasticity, and context-dependent behavioral sensitization to morphine. We found that morphine treatment produced changes of synaptic AMPAR expression in the hippocampus, a brain area that is critically involved in learning and memory. These changes could be observed 1 week after the treatment, but only when mice developed context-dependent behavioral sensitization to morphine in which morphine treatment was associated with drug administration environment. Context-dependent behavioral sensitization to morphine was also associated with increased basal synaptic transmission and disrupted hippocampal long-term potentiation (LTP), whereas these effects were less robust when morphine administration was not paired with the drug administration environment. Interestingly, some effects may be related to the prior history of morphine exposure in the drug-associated environment, since alterations of AMPAR expression, basal synaptic transmission, and LTP were observed in mice that received a saline challenge 1 week after discontinuation of morphine treatment. Furthermore, we demonstrated that phosphorylation of GluA1 AMPAR subunit plays a critical role in the acquisition and expression of context-dependent behavioral sensitization, as this behavior is blocked by a viral vector that disrupts GluA1 phosphorylation. These data provide evidence that glutamatergic signaling in the hippocampus plays an important role in context-dependent sensitization to morphine and supports further investigation of glutamate-based strategies for treating opiate addiction.

  10. Hippocampal GluA1-containing AMPA receptors mediate context-dependent sensitization to morphine

    OpenAIRE

    Xia, Yan; Portugal, George S.; Fakira, Amanda K.; Melyan, Zara; Neve, Rachael; Lee, H. Thomas; Russo, Scott J.; Liu, Jie; Morón, Jose A.

    2011-01-01

    Glutamatergic systems, including α-amino-3-hydroxy-5-methyl-4-isoxazole propionic acid receptors (AMPARs) are involved in opiate-induced neuronal and behavioral plasticity, although the mechanisms underlying these effects are not fully understood. In the present study, we investigated the effects of repeated morphine administration on AMPAR expression, synaptic plasticity, and context-dependent behavioral sensitization to morphine. We found that morphine treatment produced changes of synaptic...

  11. Alpha7 nicotinic receptor mediated protection against ethanol-induced cytotoxicity in PC12 cells.

    Science.gov (United States)

    Li, Y; King, M A; Grimes, J; Smith, N; de Fiebre, C M; Meyer, E M

    1999-01-16

    Ethanol caused a concentration-dependent loss of PC12 cells over a 24 h interval, accompanied by an increase in intracellular calcium. The specific alpha7 nicotinic receptor partial agonist DMXB attenuated both of these ethanol-induced actions at a concentration (3 microM) found previously to protect against apoptotic and necrotic cell loss. The alpha7 nicotinic receptor antagonist methylylaconitine blocked the neuroprotective action of DMXB when applied with but not 30 min after the agonist. These results indicate that activation of alpha7 nicotinic receptors may be therapeutically useful in preventing ethanol-neurotoxicity. Copyright 1999 Elsevier Science B.V.

  12. Alpha 7 nicotinic acetylcholine receptor-mediated protection against ethanol-induced neurotoxicity.

    Science.gov (United States)

    de Fiebre, NancyEllen C; de Fiebre, Christopher M

    2003-11-01

    The alpha(7)-selective nicotinic partial agonist 3-[2,4-dimethoxybenzylidene]anabaseine (DMXB) was examined for its ability to modulate ethanol-induced neurotoxicity in primary cultures of rat neurons. Primary cultures of hippocampal neurons were established from Long-Evans, embryonic day (E)-18 rat fetuses and maintained for 7 days. Ethanol (0-150 mM), DMXB (0-56 microM), or both were subsequently co-applied to cultures. Ethanol was added two additional times to the cultures to compensate for evaporation. After 5 days, neuronal viability was assessed with the MTT cell proliferation assay. Results demonstrated that ethanol reduces neuronal viability in a concentration-dependent fashion and that DMXB protects against this ethanol-induced neurotoxicity, also in a concentration-dependent fashion. These results support the suggestion that nicotinic partial agonists may be useful in treating binge drinking-induced neurotoxicity and may provide clues as to why heavy drinkers are usually smokers.

  13. Intrinsically disordered cytoplasmic domains of two cytokine receptors mediate conserved interactions with membranes

    DEFF Research Database (Denmark)

    Haxholm, Gitte Wolfsberg; Nikolajsen, Louise Fletcher; Olsen, Johan Gotthardt

    2015-01-01

    . This study presents the first comprehensive structural characterization of any cytokine receptor ICD and demonstrates that the human prolactin and growth hormone receptor ICDs are intrinsically disordered throughout their entire lengths. We show that they interact specifically with hallmark lipids...

  14. GABA(A) receptors mediate orexin-A induced stimulation of food intake.

    Science.gov (United States)

    Kokare, Dadasaheb M; Patole, Angad M; Carta, Anna; Chopde, Chandrabhan T; Subhedar, Nishikant K

    2006-01-01

    Although the role of orexins in sleep/wake cycle and feeding behavior is well established, underlying mechanisms have not been fully understood. An attempt has been made to investigate the role of GABA(A) receptors and their benzodiazepine site on the orexin-A induced response to feeding. Different groups of rats were food deprived overnight and next day injected intracerebroventricularly (icv) with vehicle (artificial CSF; 5 microl/rat) or orexin-A (20-50 nM/rat) and the animals were given free access to food. Cumulative food intake was measured during light phase of light/dark cycle at 1-, 2-, 4- and 6-h post-injection time points. Orexin-A (30-50 nM/rat, icv) stimulated food intake at all the time points (P GABA(A) receptor agonists muscimol (25 ng/rat, icv) and diazepam (0.5 mg/kg, ip) at subeffective doses significantly potentiated the hyperphagic effect of orexin-A (30 nM/rat, icv). However, the effect was negated by the GABA(A) receptor antagonist bicuculline (1 mg/kg, ip). Interestingly, benzodiazepine receptor antagonist flumazenil (5 ng/rat, icv), augmented the orexin-A (30 nM/rat, icv) induced hyperphagia; the effect may be attributed to the intrinsic activity of the agent. The results suggest that the hyperphagic effect of orexin-A, at least in part, is mediated by enhanced GABA(A) receptor activity.

  15. Acupuncture suppresses intravenous methamphetamine self-administration through GABA receptor's mediation.

    Science.gov (United States)

    Choi, Yi Jeong; Kim, Nam Jun; Zhao, Rong Jie; Kim, Da Hye; Yang, Chae Ha; Kim, Hee Young; Gwak, Young S; Jang, Eun Young; Kim, Jae Su; Lee, Yun Kyu; Lee, Hyun Jong; Lee, Sang Nam; Lim, Sung Chul; Lee, Bong Hyo

    2018-01-01

    Methamphetamine is one of the widely abused drugs. In spite of a number of studies, there is still little successful therapy to suppress the methamphetamine abuse. Acupuncture has shown to attenuate the reinforcing effects of psychostimulant. Based on, the present study investigated if acupuncture could suppress intravenous methamphetamine self-administration behavior. In addition, a possible neuronal mechanism was investigated. Male Sprague-Dawley rats weighing 270-300g were trained to intake food pellet. After catheter implantation, animal was trained to self-administer methamphetamine (0.05mg/kg) intravenously using fixed ratio 1 schedule in daily 2h session during 3 weeks. After training, rats who established baseline (infusion variation less than 20% of the mean for 3 consecutive days) received acupuncture treatment on the next day. Acupuncture was performed at each acupoint manually. In the second experiment, the selective antagonists of GABA A or GABA B receptor were given before acupuncture to investigate the possible neuronal involvement of GABA receptor pathway in the acupuncture effects. C-Fos expression was examined in the nucleus accumbens to support behavioral data. Acupuncture at HT7, but not at control acupoint LI5, reduced the self-administration behavior significantly. Also, the effects of acupuncture were blocked by the GABA receptor antagonists. C-Fos expression was shown to be parallel with the behavioral data. Results of this study have shown that acupuncture at HT7 suppressed methamphetamine self-administration through GABA receptor system, suggesting that acupuncture at HT7 can be a useful therapy for the treatment of methamphetamine abuse. Copyright © 2017 Elsevier B.V. All rights reserved.

  16. A putative octopamine/tyramine receptor mediating appetite in a hungry fly

    Science.gov (United States)

    Ishida, Yuko; Ozaki, Mamiko

    2011-07-01

    In the blowfly Phormia regina, experience of simultaneous feeding with d-limonene exposure inhibits proboscis extension reflex (PER) due to decreased tyramine (TA) titer in the brain. To elucidate the molecular mechanism of TA signaling pathway related to the associated feeding behavior, we cloned cDNA encoding the octopamine/TA receptor (PregOAR/TAR). The deduced protein is composed of 607 amino acid residues and has 7 predicted transmembrane domains. Based on homology and phylogenetic analyses, this protein belongs to the OAR/TAR family. The PregOAR/TAR was mainly expressed in head, with low levels of expression in other tissues at adult stages. Gene expression profile is in agreement with a plethora of functions ascribed to TA in various insect tissues. The immunolabeled cell bodies and processes were localized in the medial protocerebrum, outer layer of lobula, antennal lobe, and subesophageal ganglion. These results suggest that decrease of TA level in the brain likely affects neurons expressing PregOAR/TAR, causing mediation of the sensitivity in the sensillum and/or output of motor neurons for PER.

  17. 17-AAG sensitized malignant glioma cells to death-receptor mediated apoptosis.

    Science.gov (United States)

    Siegelin, Markus David; Habel, Antje; Gaiser, Timo

    2009-02-01

    17-AAG is a selective HSP90-inhibitor that exhibited therapeutic activity in cancer. In this study three glioblastoma cell lines (U87, LN229 and U251) were treated with 17-AAG, tumor necrosis factor-related apoptosis-inducing ligand (TRAIL) or the combination of both. Treatment with subtoxic doses of 17-AAG in combination with tumor necrosis factor-related apoptosis-inducing ligand (TRAIL) induces rapid apoptosis in TRAIL-resistant glioma cells, suggesting that this combined treatment may offer an attractive strategy for treating gliomas. 17-AAG treatment down-regulated survivin through proteasomal degradation. In addition, over-expression of survivin attenuated cytotoxicity induced by the combination of 17-AAG and TRAIL. In summary, survivin is a key regulator of TRAIL-17-AAG mediated cell death in malignant glioma.

  18. Serotonin₂A/C receptors mediate the aggressive phenotype of TLX gene knockout mice.

    Science.gov (United States)

    Juárez, Pablo; Valdovinos, Maria G; May, Michael E; Lloyd, Blair P; Couppis, Maria H; Kennedy, Craig H

    2013-11-01

    Deleting the tailless (TLX) gene in mice produces a highly aggressive phenotype yet to be characterized in terms of heterozygous animals or neurotransmitter mechanisms. We sought to establish pharmacological control over aggression and study the role of serotonin (5-HT)(2A/C) receptors in mediating changes in aggression. We analyzed aggression in mice heterozygous (+/-) or homozygous (-/-) for the TLX gene and wild-types (+/+) using a resident-intruder paradigm. No +/+ mice were aggressive, 36% of +/- TLX and 100% of -/- TLX mice showed aggression. Dose-effect functions were established for clozapine (0.1-1.5mg/kg, ip), ketanserin (0.3-1.25 mg/kg, ip), and (±)-1-(2,5-dimethoxy-4-iodophenyl)-2-aminopropane [(±)DOI] (0.5-2.0 mg/kg, ip). Injecting clozapine decreased the frequency and duration of attacks for +/- TLX and -/- TLX mice. Clozapine did not decrease grooming in either +/- TLX or -/- TLX mice but may have increased locomotion for -/- TLX mice. Injecting ketanserin, a 5-HT(2A/C) receptor antagonist, produced differential decreases in frequency and latency to aggression between genotypes and corresponding increases in locomotor behavior. Injecting (±)DOI, a 5-HT(2A/C) receptor agonist, increased the frequency and duration of attacks, decreased the latency to attacks, and decreased locomotion in +/- and -/- TLX mice. Results of the current study suggest aggression displayed by TLX null and heterozygous mice involves 5-HT(2A/C) receptors. Copyright © 2013 Elsevier B.V. All rights reserved.

  19. Gamma interferon augments Fc gamma receptor-mediated dengue virus infection of human monocytic cells.

    OpenAIRE

    Kontny, U; Kurane, I; Ennis, F A

    1988-01-01

    It has been reported that anti-dengue antibodies at subneutralizing concentrations augment dengue virus infection of monocytic cells. This is due to the increased uptake of dengue virus in the form of virus-antibody complexes by cells via Fc gamma receptors. We analyzed the effects of recombinant human gamma interferon (rIFN-gamma) on dengue virus infection of human monocytic cells. U937 cells, a human monocytic cell line, were infected with dengue virus in the form of virus-antibody complexe...

  20. Characterization of murine melanocortin receptors mediating adipocyte lipolysis and examination of signalling pathways involved

    DEFF Research Database (Denmark)

    Møller, Cathrine Laustrup; Raun, Kirsten; Jacobsen, Marianne Lambert

    2011-01-01

    hormone (a-MSH) generated from proopiomelanocortin (POMC), as well as synthetic MSH analogues to stimulate lipolysis in murine 3T3-L1 adipocytes it is shown that MC2R and MC5R are lipolytic mediators in differentiated 3T3-L1 adipocytes. Involvement of cAMP, phosphorylated extracellular signal...

  1. Ku proteins function as corepressors to regulate farnesoid X receptor-mediated gene expression

    International Nuclear Information System (INIS)

    Ohno, Masae; Kunimoto, Masaaki; Nishizuka, Makoto; Osada, Shigehiro; Imagawa, Masayoshi

    2009-01-01

    The farnesoid X receptor (FXR; NR1H4) is a member of the nuclear receptor superfamily and regulates the expression of genes involved in enterohepatic circulation and the metabolism of bile acids. Based on functional analyses, nuclear receptors are divided into regions A-F. To explore the cofactors interacting with FXR, we performed a pull-down assay using GST-fused to the N-terminal A/B region and the C region, which are required for the ligand-independent transactivation and DNA-binding, respectively, of FXR, and nuclear extracts from HeLa cells. We identified DNA-dependent protein kinase catalytic subunit (DNA-PKcs), Ku80, and Ku70 as FXR associated factors. These proteins are known to have an important role in DNA repair, recombination, and transcription. DNA-PKcs mainly interacted with the A/B region of FXR, whereas the Ku proteins interacted with the C region and with the D region (hinge region). Chromatin immunoprecipitation assays revealed that the Ku proteins associated with FXR on the bile salt export pump (BSEP) promoter. Furthermore, we demonstrated that ectopic expression of the Ku proteins decreased the promoter activity and expression of BSEP gene mediated by FXR. These results suggest that the Ku proteins function as corepressors for FXR.

  2. Voltage dependent anion channel-1 regulates death receptor mediated apoptosis by enabling cleavage of caspase-8

    International Nuclear Information System (INIS)

    Chacko, Alex D; Liberante, Fabio; Paul, Ian; Longley, Daniel B; Fennell, Dean A

    2010-01-01

    Activation of the extrinsic apoptosis pathway by tumour necrosis factor related apoptosis inducing ligand (TRAIL) is a novel therapeutic strategy for treating cancer that is currently under clinical evaluation. Identification of molecular biomarkers of resistance is likely to play an important role in predicting clinical anti tumour activity. The involvement of the mitochondrial type 1 voltage dependent anion channel (VDAC1) in regulating apoptosis has been highly debated. To date, a functional role in regulating the extrinsic apoptosis pathway has not been formally excluded. We carried out stable and transient RNAi knockdowns of VDAC1 in non-small cell lung cancer cells, and stimulated the extrinsic apoptotic pathway principally by incubating cells with the death ligand TRAIL. We used in-vitro apoptotic and cell viability assays, as well as western blot for markers of apoptosis, to demonstrate that TRAIL-induced toxicity is VDAC1 dependant. Confocal microscopy and mitochondrial fractionation were used to determine the importance of mitochondria for caspase-8 activation. Here we show that either stable or transient knockdown of VDAC1 is sufficient to antagonize TRAIL mediated apoptosis in non-small cell lung cancer (NSCLC) cells. Specifically, VDAC1 is required for processing of procaspase-8 to its fully active p18 form at the mitochondria. Loss of VDAC1 does not alter mitochondrial sensitivity to exogenous caspase-8-cleaved BID induced mitochondrial depolarization, even though VDAC1 expression is essential for TRAIL dependent activation of the intrinsic apoptosis pathway. Furthermore, expression of exogenous VDAC1 restores the apoptotic response to TRAIL in cells in which endogenous VDAC1 has been selectively silenced. Expression of VDAC1 is required for full processing and activation of caspase-8 and supports a role for mitochondria in regulating apoptosis signaling via the death receptor pathway

  3. Natural Modulators of Endosomal Toll-Like Receptor-Mediated Psoriatic Skin Inflammation

    Directory of Open Access Journals (Sweden)

    Chao-Yang Lai

    2017-01-01

    Full Text Available Psoriasis is a chronic inflammatory autoimmune disease that can be initiated by excessive activation of endosomal toll-like receptors (TLRs, particularly TLR7, TLR8, and TLR9. Therefore, inhibitors of endosomal TLR activation are being investigated for their ability to treat this disease. The currently approved biological drugs adalimumab, etanercept, infliximab, ustekinumab, ixekizumab, and secukizumab are antibodies against effector cytokines that participate in the initiation and development of psoriasis. Several immune modulatory oligonucleotides and small molecular weight compounds, including IMO-3100, IMO-8400, and CPG-52364, that block the interaction between endosomal TLRs and their ligands are under clinical investigation for their effectiveness in the treatment of psoriasis. In addition, several chemical compounds, including AS-2444697, PF-05387252, PF-05388169, PF-06650833, ML120B, and PHA-408, can inhibit TLR signaling. Although these compounds have demonstrated anti-inflammatory activity in animal models, their therapeutic potential for the treatment of psoriasis has not yet been tested. Recent studies demonstrated that natural compounds derived from plants, fungi, and bacteria, including mustard seed, Antrodia cinnamomea extract, curcumin, resveratrol, thiostrepton, azithromycin, and andrographolide, inhibited psoriasis-like inflammation induced by the TLR7 agonist imiquimod in animal models. These natural modulators employ different mechanisms to inhibit endosomal TLR activation and are administered via different routes. Therefore, they represent candidate psoriasis drugs and might lead to the development of new treatment options.

  4. Dopamine D3 receptors mediate the discriminative stimulus effects of quinpirole in free-feeding rats.

    Science.gov (United States)

    Baladi, Michelle G; Newman, Amy H; France, Charles P

    2010-01-01

    The discriminative stimulus effects of dopamine (DA) D3/D2 receptor agonists are thought to be mediated by D2 receptors. To maintain responding, access to food is often restricted, which can alter neurochemical and behavioral effects of drugs acting on DA systems. This study established stimulus control with quinpirole in free-feeding rats and tested the ability of agonists to mimic and antagonists to attenuate the effects of quinpirole. The same antagonists were studied for their ability to attenuate quinpirole-induced yawning and hypothermia. DA receptor agonists apomorphine and lisuride, but not amphetamine and morphine, occasioned responding on the quinpirole lever. The discriminative stimulus effects of quinpirole were attenuated by the D3 receptor-selective antagonist N-{4-[4-(2,3-dichlorophenyl)-piperazin-1-yl]-trans-but-2-enyl}-4-pyridine-2-yl-benzamide HCl (PG01037) and the nonselective D3/D2 receptor antagonist raclopride, but not by the D2 receptor-selective antagonist 3-[4-(4-chlorophenyl)-4-hydroxypiperidin-1-yl]methyl-1H-indole (L-741,626); the potencies of PG01037 and raclopride to antagonize this effect of quinpirole paralleled their potencies to antagonize the ascending limb of the quinpirole yawning dose-response curve (thought to be mediated by D3 receptors). L-741,626 selectively antagonized the descending limb of the quinpirole yawning dose-response curve, and both L-741,626 and raclopride, but not PG01037, antagonized the hypothermic effects of quinpirole (thought to be mediated by D2 receptors). Food restriction (10 g/day/7 days) significantly decreased quinpirole-induced yawning without affecting the quinpirole discrimination. Many discrimination studies on DA receptor agonists use food-restricted rats; together with those studies, the current experiment using free-feeding rats suggests that feeding conditions affecting the behavioral effects of direct-acting DA receptor agonists might also have an impact on the effects of indirect-acting agonists such as cocaine and amphetamine.

  5. Receptor-mediated internalization of [3H]-neurotensin in synaptosomal preparations from rat neostriatum.

    Science.gov (United States)

    Nguyen, Ha Minh Ky; Cahill, Catherine M; McPherson, Peter S; Beaudet, Alain

    2002-06-01

    Following its binding to somatodendritic receptors, the neuropeptide neurotensin (NT) internalizes via a clathrin-mediated process. In the present study, we investigated whether NT also internalizes presynaptically using synaptosomes from rat neostriatum, a region in which NT1 receptors are virtually all presynaptic. Binding of [(3)H]-NT to striatal synaptosomes in the presence of levocabastine to block NT2 receptors is specific, saturable, and has NT1 binding properties. A significant fraction of the bound radioactivity is resistant to hypertonic acid wash indicating that it is internalized. Internalization of [(3)H]-NT, like that of [(125)I]-transferrin, is blocked by sucrose and low temperature, consistent with endocytosis occurring via a clathrin-dependent pathway. However, contrary to what was reported at the somatodendritic level, neither [(3)H]-NT nor [(125)I]-transferrin internalization in synaptosomes is sensitive to the endocytosis inhibitor phenylarsine oxide. Moreover, treatment of synaptosomes with monensin, which prevents internalized receptors from recycling to the plasma membrane, reduces [(3)H]-NT binding and internalization, suggesting that presynaptic NT1 receptors, in contrast to somatodendritic ones, are recycled back to the plasma membrane. Taken together, these results suggest that NT internalizes in nerve terminals via an endocytic pathway that is related to, but is mechanistically distinct from that responsible for NT internalization in nerve cell bodies.

  6. The α7-nicotinic acetylcholine receptor mediates the sensitivity of gastric cancer cells to taxanes.

    Science.gov (United States)

    Tu, Chao-Chiang; Huang, Chien-Yu; Cheng, Wan-Li; Hung, Chin-Sheng; Uyanga, Batzorig; Wei, Po-Li; Chang, Yu-Jia

    2016-04-01

    Gastric cancer is difficult to cure because most patients are diagnosed at an advanced disease stage. Systemic chemotherapy remains an important therapy for gastric cancer, but both progression-free survival and disease-free survival associated with various combination regimens are limited because of refractoriness and chemoresistance. Accumulating evidence has revealed that the homomeric α7-nicotinic acetylcholine receptor (A7-nAChR) promotes human gastric cancer by driving cancer cell proliferation, migration, and metastasis. Therefore, A7-nAChR may serve as a potential therapeutic target for gastric cancer. However, the role of A7-nAChR in taxane therapy for gastric cancer was unclear. Cells were subjected to A7-nAChR knockdown (A7-nAChR KD) using short interfering RNA (siRNA). The anti-proliferative effects of taxane were assessed via 3-[4,5-dimethylthiazol-2-yl]-2,5-diphenyltetrazolium bromide (MTT), terminal deoxynucleotidyl transferase-mediated nick-end labeling (TUNEL), and cell cycle distribution assays. A7-nAChR-KD cells exhibited low resistance to docetaxel and paclitaxel treatment, as measured by the MTT assay. Following paclitaxel treatment, the proportion of apoptotic cells was higher among A7-nAChR-KD cells than among scrambled control cells, as measured by cell cycle distribution and TUNEL assays. Further molecular analyses showed a reduction in the pAKT levels and a dramatic increase in the Bad levels in paclitaxel-treated A7-nAChR-KD cells but not in scrambled control cells. Following paclitaxel treatment, the level of Bax was slightly increased in both cell populations, whereas Poly (ADP-ribose) polymerase (PARP) cleavage was increased only in A7-nAChR-KD cells. These findings indicate that A7-nAChR-KD cells are more sensitive to paclitaxel treatment. We conclude that A7-nAChR may be a key biomarker for assessing the chemosensitivity of gastric cancer cells to taxane.

  7. Toll-like receptor mediated activation is possibly involved in immunoregulating properties of cow's milk hydrolysates

    NARCIS (Netherlands)

    Kiewiet, M. B. Gea; Dekkers, Renske; Gros, Marjan; van Neerven, R. J. Joost; Groeneveld, Andre; de Vos, Paul; Faas, Marijke M.

    2017-01-01

    Immunomodulating proteins and peptides are formed during the hydrolysis of cow's milk proteins. These proteins are potential ingredients in functional foods used for the management of a range of immune related problems, both in infants and adults. However, the mechanism behind these effects is

  8. Glucocorticoid Receptor-Mediated Repression of Pro-Inflammatory Genes in Rheumatoid Arthritis

    Science.gov (United States)

    2015-10-01

    Braun TP, Zhu X, Szumowski M, Scott GD, Grossberg AJ, et al. 2011. Central nervous system inflam - mation induces muscle atrophy via activation of the...glucocorticoid receptor and nutritional sensor mTOR in skeletal muscle . Cell Metab. 13:170–82 133. Lutzner N, Kalbacher H, Krones-Herzig A, Rosl F. 2012...suggestions for reducing this burden to Department of Defense, Washington Headquarters Services, Directorate for Information Operations and Reports (0704

  9. Ursodeoxycholic Acid Induces Death Receptor-mediated Apoptosis in Prostate Cancer Cells

    Science.gov (United States)

    Lee, Won Sup; Jung, Ji Hyun; Panchanathan, Radha; Yun, Jeong Won; Kim, Dong Hoon; Kim, Hye Jung; Kim, Gon Sup; Ryu, Chung Ho; Shin, Sung Chul; Hong, Soon Chan; Choi, Yung Hyun; Jung, Jin-Myung

    2017-01-01

    Background Bile acids have anti-cancer properties in a certain types of cancers. We determined anticancer activity and its underlying molecular mechanism of ursodeoxycholic acid (UDCA) in human DU145 prostate cancer cells. Methods Cell viability was measured with an MTT assay. UDCA-induced apoptosis was determined with flow cytometric analysis. The expression levels of apoptosis-related signaling proteins were examined with Western blotting. Results UDCA treatment significantly inhibited cell growth of DU145 in a dose-dependent manner. It induced cellular shrinkage and cytoplasmic blebs and accumulated the cells with sub-G1 DNA contents. Moreover, UDCA activated caspase 8, suggesting that UDCA-induced apoptosis is associated with extrinsic pathway. Consistent to this finding, UDCA increased the expressions of tumor necrosis factor-related apoptosis-inducing ligand (TRAIL) receptor, death receptor 4 (DR4) and death receptor 5 (DR5), and TRAIL augmented the UDCA-induced cell death in DU145 cells. In addition, UDCA also increased the expressions of Bax and cytochrome c and decreased the expression of Bcl-xL in DU145 cells. This finding suggests that UDCA-induced apoptosis may be involved in intrinsic pathway. Conclusions UDCA induces apoptosis via extrinsic pathway as well as intrinsic pathway in DU145 prostate cancer cells. UDCA may be a promising anti-cancer agent against prostate cancer. PMID:28382282

  10. Adrenergic β2-receptors mediates visceral hypersensitivity induced by heterotypic intermittent stress in rats.

    Directory of Open Access Journals (Sweden)

    Chunhua Zhang

    Full Text Available Chronic visceral pain in patients with irritable bowel syndrome (IBS has been difficult to treat effectively partially because its pathophysiology is not fully understood. Recent studies show that norepinephrine (NE plays an important role in the development of visceral hypersensitivity. In this study, we designed to investigate the role of adrenergic signaling in visceral hypersensitivity induced by heterotypical intermittent stress (HIS. Abdominal withdrawal reflex scores (AWRs used as visceral sensitivity were determined by measuring the visceromoter responses to colorectal distension. Colon-specific dorsal root ganglia neurons (DRGs were labeled by injection of DiI into the colon wall and were acutely dissociated for whole-cell patch-clamp recordings. Blood plasma level of NE was measured using radioimmunoassay kits. The expression of β2-adrenoceptors was measured by western blotting. We showed that HIS-induced visceral hypersensitivity was attenuated by systemic administration of a β-adrenoceptor antagonist propranolol, in a dose-dependent manner, but not by a α-adrenoceptor antagonist phentolamine. Using specific β-adrenoceptor antagonists, HIS-induced visceral hypersensitivity was alleviated by β2 adrenoceptor antagonist but not by β1- or β3-adrenoceptor antagonist. Administration of a selective β2-adrenoceptor antagonist also normalized hyperexcitability of colon-innervating DRG neurons of HIS rats. Furthermore, administration of β-adrenoceptor antagonist suppressed sustained potassium current density (IK without any alteration of fast-inactivating potassium current density (IA. Conversely, administration of NE enhanced the neuronal excitability and produced visceral hypersensitivity in healthy control rats, and blocked by β2-adrenoceptor antagonists. In addition, HIS significantly enhanced the NE concentration in the blood plasma but did not change the expression of β2-adrenoceptor in DRGs and the muscularis externa of the colon. The present study might provide a potential molecular target for therapy of visceral hypersensitivity in patents with IBS.

  11. Cerebellar Kainate Receptor-Mediated Facilitation of Glutamate Release Requires Ca2+-Calmodulin and PKA

    Directory of Open Access Journals (Sweden)

    Rafael Falcón-Moya

    2018-06-01

    Full Text Available We elucidated the mechanisms underlying the kainate receptor (KAR-mediated facilitatory modulation of synaptic transmission in the cerebellum. In cerebellar slices, KA (3 μM increased the amplitude of evoked excitatory postsynaptic currents (eEPSCs at synapses between axon terminals of parallel fibers (PF and Purkinje neurons. KA-mediated facilitation was antagonized by NBQX under condition where AMPA receptors were previously antagonized. Inhibition of protein kinase A (PKA suppressed the effect of KA on glutamate release, which was also obviated by the prior stimulation of adenylyl cyclase (AC. KAR-mediated facilitation of synaptic transmission was prevented by blocking Ca2+ permeant KARs using philanthotoxin. Furthermore, depletion of intracellular Ca2+ stores by thapsigargin, or inhibition of Ca2+-induced Ca2+-release by ryanodine, abrogated the synaptic facilitation by KA. Thus, the KA-mediated modulation was conditional on extracellular Ca2+ entry through Ca2+-permeable KARs, as well as and mobilization of Ca2+ from intracellular stores. Finally, KAR-mediated facilitation was sensitive to calmodulin inhibitors, W-7 and calmidazolium, indicating that the increased cytosolic [Ca2+] sustaining KAR-mediated facilitation of synaptic transmission operates through a downstream Ca2+/calmodulin coupling. We conclude that, at cerebellar parallel fiber-Purkinje cell synapses, presynaptic KARs mediate glutamate release facilitation, and thereby enhance synaptic transmission through Ca2+-calmodulin dependent activation of adenylyl cyclase/cAMP/protein kinase A signaling.

  12. Identification of a novel modulator of thyroid hormone receptor-mediated action.

    Directory of Open Access Journals (Sweden)

    Bernhard G Baumgartner

    Full Text Available BACKGROUND: Diabetes is characterized by reduced thyroid function and altered myogenesis after muscle injury. Here we identify a novel component of thyroid hormone action that is repressed in diabetic rat muscle. METHODOLOGY/PRINCIPAL FINDINGS: We have identified a gene, named DOR, abundantly expressed in insulin-sensitive tissues such as skeletal muscle and heart, whose expression is highly repressed in muscle from obese diabetic rats. DOR expression is up-regulated during muscle differentiation and its loss-of-function has a negative impact on gene expression programmes linked to myogenesis or driven by thyroid hormones. In agreement with this, DOR enhances the transcriptional activity of the thyroid hormone receptor TR(alpha1. This function is driven by the N-terminal part of the protein. Moreover, DOR physically interacts with TR( alpha1 and to T(3-responsive promoters, as shown by ChIP assays. T(3 stimulation also promotes the mobilization of DOR from its localization in nuclear PML bodies, thereby indicating that its nuclear localization and cellular function may be related. CONCLUSIONS/SIGNIFICANCE: Our data indicate that DOR modulates thyroid hormone function and controls myogenesis. DOR expression is down-regulated in skeletal muscle in diabetes. This finding may be of relevance for the alterations in muscle function associated with this disease.

  13. Modulation of Toll-like receptor-mediated activation of Microglia

    NARCIS (Netherlands)

    Putten, C. M.-T. van der

    2015-01-01

    Microglia are the resident macrophages of the central nervous system (CNS). Like other tissue macrophages, microglia have many different functions under physiological as well as pathological conditions. Microglia can contribute to the initiation, progression and resolution of disease processes and

  14. Modeling receptor-mediated endocytosis of polymer-functionalized iron oxide nanoparticles by human macrophages

    Czech Academy of Sciences Publication Activity Database

    Lunov, O.; Zablotskyy, Vitaliy A.; Syrovets, T.; Röcker, C.; Tron, K.; Nienhaus, G.U.; Simmet, T.

    2011-01-01

    Roč. 32, č. 2 (2011), s. 547-555 ISSN 0142-9612 Institutional research plan: CEZ:AV0Z10100522 Keywords : macrophage * nanoparticle * bioabsorption * modeling * antisense * MRI (magnetic resonance imaging) Subject RIV: BM - Solid Matter Physics ; Magnetism Impact factor: 7.404, year: 2011 http://www.sciencedirect.com/science/article/pii/S014296121001149X

  15. Mechanisms of G Protein-Coupled Estrogen Receptor-Mediated Spinal Nociception

    DEFF Research Database (Denmark)

    Deliu, Elena; Brailoiu, G. Cristina; Arterburn, Jeffrey B.

    2012-01-01

    . Cytosolic calcium concentration elevates faster and with higher amplitude following G-1 intracellular microinjections compared to extracellular exposure, suggesting subcellular GPER functionality. Thus, GPER activation results in spinal nociception, and the downstream mechanisms involve cytosolic calcium......Human and animal studies suggest that estrogens are involved in the processing of nociceptive sensory information and analgesic responses in the central nervous system. Rapid pronociceptive estrogenic effects have been reported, some of which likely involve G protein-coupled estrogen receptor (GPER......) activation. Membrane depolarization and increases in cytosolic calcium and reactive oxygen species (ROS) levels are markers of neuronal activation, underlying pain sensitization in the spinal cord. Using behavioral, electrophysiological, and fluorescent imaging studies, we evaluated GPER involvement...

  16. An in vitro approach for prioritization and evaluation of chemical effects on glucocorticoid receptor mediated adipogenesis.

    Science.gov (United States)

    Hartman, Jessica K; Beames, Tyler; Parks, Bethany; Doheny, Daniel; Song, Gina; Efremenko, Alina; Yoon, Miyoung; Foley, Briana; Deisenroth, Chad; McMullen, Patrick D; Clewell, Rebecca A

    2018-05-18

    Rising obesity rates worldwide have socio-economic ramifications. While genetics, diet, and lack of exercise are major contributors to obesity, environmental factors may enhance susceptibility through disruption of hormone homeostasis and metabolic processes. The obesogen hypothesis contends that chemical exposure early in development may enhance adipocyte differentiation, thereby increasing the number of adipocytes and predisposing for obesity and metabolic disease. We previously developed a primary human adipose stem cell (hASC) assay to evaluate the effect of environmental chemicals on PPARG-dependent adipogenesis. Here, the assay was modified to determine the effects of chemicals on the glucocorticoid receptor (GR) pathway. In differentiation cocktail lacking the glucocorticoid agonist dexamethasone (DEX), hASCs do not differentiate into adipocytes. In the presence of GR agonists, adipocyte maturation was observed using phenotypic makers for lipid accumulation, adipokine secretion, and expression of key genes. To evaluate the role of environmental compounds on adipocyte differentiation, progenitor cells were treated with 19 prioritized compounds previously identified by ToxPi as having GR-dependent bioactivity, and multiplexed assays were used to confirm a GR-dependent mode of action. Five chemicals were found to be strong agonists. The assay was also modified to evaluate GR-antagonists, and 8/10 of the hypothesized antagonists inhibited adipogenesis. The in vitro bioactivity data was put into context with extrapolated human steady state concentrations (Css) and clinical exposure data (Cmax). These data support using a human adipose-derived stem cell differentiation assay to test the potential of chemicals to alter human GR-dependent adipogenesis. Copyright © 2017. Published by Elsevier Inc.

  17. Valerian inhibits rat hepatocarcinogenesis by activating GABA(A receptor-mediated signaling.

    Directory of Open Access Journals (Sweden)

    Anna Kakehashi

    Full Text Available Valerian is widely used as a traditional medicine to improve the quality of sleep due to interaction of several active components with the γ-aminobutyric acid (GABA A receptor (GABA(AR system. Recently, activation of GABA signaling in stem cells has been reported to suppress cell cycle progression in vivo. Furthermore, possible inhibitory effects of GABA(AR agonists on hepatocarcinogenesis have been reported. The present study was performed to investigate modulating effects of Valerian on hepatocarcinogenesis using a medium-term rat liver bioassay. Male F344 rats were treated with one of the most powerful Valerian species (Valeriana sitchensis at doses of 0, 50, 500 and 5000 ppm in their drinking water after initiation of hepatocarcinogenesis with diethylnitrosamine (DEN. Formation of glutathione S-transferase placental form positive (GST-P(+ foci was significantly inhibited by Valerian at all applied doses compared with DEN initiation control rats. Generation of 8-hydroxy-2'-deoxyguanosine in the rat liver was significantly suppressed by all doses of Valerian, likely due to suppression of Nrf2, CYP7A1 and induction of catalase expression. Cell proliferation was significantly inhibited, while apoptosis was induced in areas of GST-P(+ foci of Valerian groups associated with suppression of c-myc, Mafb, cyclin D1 and induction of p21(Waf1/Cip1, p53 and Bax mRNA expression. Interestingly, expression of the GABA(AR alpha 1 subunit was observed in GST-P(+ foci of DEN control rats, with significant elevation associated with Valerian treatment. These results indicate that Valerian exhibits inhibitory effects on rat hepatocarcinogenesis by inhibiting oxidative DNA damage, suppressing cell proliferation and inducing apoptosis in GST-P(+ foci by activating GABA(AR-mediated signaling.

  18. Chlorotoxin Fused to IgG-Fc Inhibits Glioblastoma Cell Motility via Receptor-Mediated Endocytosis

    Directory of Open Access Journals (Sweden)

    Tomonari Kasai

    2012-01-01

    Full Text Available Chlorotoxin is a 36-amino acid peptide derived from Leiurus quinquestriatus (scorpion venom, which has been shown to inhibit low-conductance chloride channels in colonic epithelial cells. Chlorotoxin also binds to matrix metalloproteinase-2 and other proteins on glioma cell surfaces. Glioma cells are considered to require the activation of matrix metalloproteinase-2 during invasion and migration. In this study, for targeting glioma, we designed two types of recombinant chlorotoxin fused to human IgG-Fcs with/without a hinge region. Chlorotoxin fused to IgG-Fcs was designed as a dimer of 60 kDa with a hinge region and a monomer of 30 kDa without a hinge region. The monomeric and dimeric forms of chlorotoxin inhibited cell proliferation at 300 nM and induced internalization in human glioma A172 cells. The monomer had a greater inhibitory effect than the dimer; therefore, monomeric chlorotoxin fused to IgG-Fc was multivalently displayed on the surface of bionanocapsules to develop a drug delivery system that targeted matrix metalloproteinase-2. The target-dependent internalization of bionanocapsules in A172 cells was observed when chlorotoxin was displayed on the bionanocapsules. This study indicates that chlorotoxin fused to IgG-Fcs could be useful for the active targeting of glioblastoma cells.

  19. Dopamine D(1) receptor-mediated control of striatal acetylcholine release by endogenous dopamine.

    Science.gov (United States)

    Acquas, E; Di Chiara, G

    1999-10-27

    The role of dopamine D(1) and D(2) receptors in the control of acetylcholine release in the dorsal striatum by endogenous dopamine was investigated by monitoring with microdialysis the effect of the separate or combined administration of the dopamine D(1) receptor antagonist, SCH 39166 ¿(-)-trans-6,7,7a,8,9, 13b-exahydro-3-chloro-2-hydroxy-N-methyl-5H-benzo-[d]-nap hto-[2, 1b]-azepine hydrochloride¿ (50 microg/kg subcutaneous (s.c.)), of the dopamine D(2)/D(3) receptor agonist, quinpirole (trans-(-)-4aR, 4a,5,6,7,8,8a,9-octahydro-5-propyl-1H-pyrazolo-(3,4-g)-quinoline hydrochloride) (5 and 10 microg/kg s.c.), and of the D(3) receptor selective agonist, PD 128,907 [S(+)-(4aR,10bR)-3,4,4a, 10b-tetrahydro-4-propyl-2H,5H-[1]benzopyrano-[4,3-b]-1,4-oxazin -9-ol hydrochloride] (50 microg/kg s.c.), on in vivo dopamine and acetylcholine release. Microdialysis was performed with a Ringer containing low concentrations (0.01 microM) of the acetylcholinesterase inhibitor, neostigmine. Quinpirole (10 microg/kg s.c.) decreased striatal dopamine and acetylcholine release. Administration of PD 128,907 (50 microg/kg) decreased dopamine but failed to affect acetylcholine release. SCH 39166 (50 microg/kg s.c.) stimulated dopamine release and reduced acetylcholine release. Pretreatment with quinpirole reduced (5 microg/kg s.c.) or completely prevented (10 microg/kg s.c.) the stimulation of dopamine release elicited by SCH 39166 (50 microg/kg s.c.); on the other hand, pretreatment with quinpirole (5 and 10 microg/kg) potentiated the reduction of striatal acetylcholine release induced by SCH 39166 (50 microg/kg s.c.). Similarly, pretreatment with PD 128,907 (50 microg/kg) which prevented the increase of dopamine release induced by SCH 39166 (50 microg/kg), potentiated the reduction of striatal acetylcholine transmission elicited by SCH 39166. Thus, pretreatment with low doses of quinpirole or PD 128,907 influences in opposite manner the effect of SCH 39166 on striatal dopamine and acetylcholine release, counteracting the increase of dopamine release and potentiating the decrease in acetylcholine release. These results provide further evidence for the existence of a tonic stimulatory input of endogenous dopamine on striatal acetylcholine transmission mediated by dopamine D(1) receptors.

  20. MicroRNA-219 modulates NMDA receptor-mediated neurobehavioral dysfunction

    DEFF Research Database (Denmark)

    Kocerha, Jannet; Faghihi, Mohammad Ali; Lopez-Toledano, Miguel A

    2009-01-01

    significantly modulated behavioral responses associated with disrupted NMDA receptor transmission. Furthermore, pretreatment with the antipsychotic drugs haloperidol and clozapine prevented dizocilpine-induced effects on miR-219. Taken together, these data support an integral role for miR-219 in the expression...

  1. Rab20 regulates phagosome maturation in RAW264 macrophages during Fc gamma receptor-mediated phagocytosis.

    Directory of Open Access Journals (Sweden)

    Youhei Egami

    Full Text Available Rab20, a member of the Rab GTPase family, is known to be involved in membrane trafficking, however its implication in FcγR-mediated phagocytosis is unclear. We examined the spatiotemporal localization of Rab20 during phagocytosis of IgG-opsonized erythrocytes (IgG-Es in RAW264 macrophages. By the live-cell imaging of fluorescent protein-fused Rab20, it was shown that Rab20 was transiently associated with the phagosomal membranes. During the early stage of phagosome formation, Rab20 was not localized on the membranes of phagocytic cups, but was gradually recruited to the newly formed phagosomes. Although Rab20 was colocalized with Rab5 to some extent, the association of Rab20 with the phagosomes persisted even after the loss of Rab5 from the phagosomal membranes. Then, Rab20 was colocalized with Rab7 and Lamp1, late endosomal/lysosomal markers, on the internalized phagosomes. Moreover, our analysis of Rab20 mutant expression revealed that the maturation of phagosomes was significantly delayed in cells expressing the GDP-bound mutant Rab20-T19N. These data suggest that Rab20 is an important component of phagosome and regulates the phagosome maturation during FcγR-mediated phagocytosis.

  2. Effects of Chronic Alcohol Exposure on Kainate Receptor-Mediated Neurotransmission in the Hippocampus

    Science.gov (United States)

    2004-09-01

    12917-12922, 1999. Frerking M, and Nicoll RA. Synaptic kainate receptors. Cur Opin Neurobio 10:342-351, 2000. Harvey J and Lacey MG. A postsynaptic...electrophysiological research. Ironically, this preparation was originally developed for use in biochemical studies characterizing energy metabolism in neuronal

  3. Activation of Brain Somatostatin Signaling Suppresses CRF Receptor-Mediated Stress Response

    Directory of Open Access Journals (Sweden)

    Andreas Stengel

    2017-04-01

    Full Text Available Corticotropin-releasing factor (CRF is the hallmark brain peptide triggering the response to stress and mediates—in addition to the stimulation of the hypothalamus-pituitary-adrenal (HPA axis—other hormonal, behavioral, autonomic and visceral components. Earlier reports indicate that somatostatin-28 injected intracerebroventricularly counteracts the acute stress-induced ACTH and catecholamine release. Mounting evidence now supports that activation of brain somatostatin signaling exerts a broader anti-stress effect by blunting the endocrine, autonomic, behavioral (with a focus on food intake and visceral gastrointestinal motor responses through the involvement of distinct somatostatin receptor subtypes.

  4. Activation of Brain Somatostatin Signaling Suppresses CRF Receptor-Mediated Stress Response.

    Science.gov (United States)

    Stengel, Andreas; Taché, Yvette F

    2017-01-01

    Corticotropin-releasing factor (CRF) is the hallmark brain peptide triggering the response to stress and mediates-in addition to the stimulation of the hypothalamus-pituitary-adrenal (HPA) axis-other hormonal, behavioral, autonomic and visceral components. Earlier reports indicate that somatostatin-28 injected intracerebroventricularly counteracts the acute stress-induced ACTH and catecholamine release. Mounting evidence now supports that activation of brain somatostatin signaling exerts a broader anti-stress effect by blunting the endocrine, autonomic, behavioral (with a focus on food intake) and visceral gastrointestinal motor responses through the involvement of distinct somatostatin receptor subtypes.

  5. Activation of Brain Somatostatin Signaling Suppresses CRF Receptor-Mediated Stress Response

    OpenAIRE

    Andreas Stengel; Yvette F. Taché; Yvette F. Taché

    2017-01-01

    Corticotropin-releasing factor (CRF) is the hallmark brain peptide triggering the response to stress and mediates—in addition to the stimulation of the hypothalamus-pituitary-adrenal (HPA) axis—other hormonal, behavioral, autonomic and visceral components. Earlier reports indicate that somatostatin-28 injected intracerebroventricularly counteracts the acute stress-induced ACTH and catecholamine release. Mounting evidence now supports that activation of brain somatostatin signaling exerts a br...

  6. Cobaltous chloride and hypoxia inhibit aryl hydrocarbon receptor-mediated responses in breast cancer cells

    International Nuclear Information System (INIS)

    Khan, Shaheen; Liu Shengxi; Stoner, Matthew; Safe, Stephen

    2007-01-01

    The aryl hydrocarbon receptor (AhR) is expressed in estrogen receptor (ER)-positive ZR-75 breast cancer cells. Treatment with 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) induces CYP1A1 protein and mRNA levels and also activates inhibitory AhR-ERα crosstalk associated with hormone-induced reporter gene expression. In ZR-75 cells grown under hypoxia, induction of these AhR-mediated responses by TCDD was significantly inhibited. This was not accompanied by decreased nuclear AhR levels or decreased interaction of the AhR complex with the CYP1A1 gene promoter as determined in a chromatin immunoprecipitation assay. Hypoxia-induced loss of Ah-responsiveness was not associated with induction of hypoxia-inducible factor-1α or other factors that sequester the AhR nuclear translocation (Arnt) protein, and overexpression of Arnt under hypoxia did not restore Ah-responsiveness. The p65 subunit of NFκB which inhibits AhR-mediated transactivation was not induced by hypoxia and was primarily cytosolic in ZR-75 cells grown under hypoxic and normoxic conditions. In ZR-75 cells maintained under hypoxic conditions for 24 h, BRCA1 (an enhancer of AhR-mediated transactivation in breast cancer cells) was significantly decreased and this contributed to loss of Ah-responsiveness. In cells grown under hypoxia for 6 h, BRCA1 was not decreased, but induction of CYP1A1 by TCDD was significantly decreased. Cotreatment of ZR-75 cells with TCDD plus the protein synthesis inhibitor cycloheximide for 6 h enhanced CYP1A1 expression in cells grown under hypoxia and normoxia. These results suggest that hypoxia rapidly induces protein(s) that inhibit Ah-responsiveness and these may be similar to constitutively expressed inhibitors of Ah-responsiveness (under normoxia) that are also inhibited by cycloheximide

  7. P2X7 receptor mediates activation of microglial cells in prostate of chemically irritated rats

    Directory of Open Access Journals (Sweden)

    Heng Zhang

    2013-04-01

    Full Text Available Purpose Evidence shows that adenosine triphosphate (ATP is involved in the transmission of multiple chronic pain via P2X7 receptor. This study was to investigate the P2X7 and microglial cells in the chronic prostatitis pain. Materials and Methods Rats were divided into control group and chronic prostatitis group (n = 24 per group. A chronic prostatitis animal model was established by injecting complete Freund's adjuvant (CFA to the prostate of rats, and the thermal withdrawal latency (TWL was detected on days 0, 4, 12 and 24 (n = 6 at each time point in each group. Animals were sacrificed and the pathological examination of the prostate, detection of mRNA expression of P2X7 and ionized calcium binding adaptor molecule 1 (IBA-1 and measurement of content of tumor necrosis factor-α (TNF-α and interleukin-1β (IL-1β in the dorsal horn of L5-S2 spinal cord were performed on days 0, 4, 12 and 24. In addition, the content of TNF-α and IL-1β in the dorsal horn of L5-S2 spinal cord was measured after intrathecal injection of inhibitors of microglial cells and/or P2X7 for 5 days. Results The chronic prostatitis was confirmed by pathological examination. The expression of P2X7 and IBA-1 and the content of TNF-α and IL-1β in rats with chronic prostatitis were significantly higher than those in the control group. On day 4, the expressions of pro-inflammatory cytokines became to increase, reaching a maximal level on day 12 and started to reduce on day 24, but remained higher than that in the control group. Following suppression of microglial cells and P2X7 receptor, the secretion of TNF-α and IL-1β was markedly reduced. Conclusion In chronic prostatitis pain, the microglial cells and P2X7 receptor are activated resulting in the increased expression of TNF-α and IL-1β in the L5-S2 spinal cord, which might attribute to the maintenance and intensification of pain in chronic prostatitis.

  8. The second extracellular loop of the adenosine A1 receptor mediates activity of allosteric enhancers.

    Science.gov (United States)

    Kennedy, Dylan P; McRobb, Fiona M; Leonhardt, Susan A; Purdy, Michael; Figler, Heidi; Marshall, Melissa A; Chordia, Mahendra; Figler, Robert; Linden, Joel; Abagyan, Ruben; Yeager, Mark

    2014-02-01

    Allosteric enhancers of the adenosine A1 receptor amplify signaling by orthosteric agonists. Allosteric enhancers are appealing drug candidates because their activity requires that the orthosteric site be occupied by an agonist, thereby conferring specificity to stressed or injured tissues that produce adenosine. To explore the mechanism of allosteric enhancer activity, we examined their action on several A1 receptor constructs, including (1) species variants, (2) species chimeras, (3) alanine scanning mutants, and (4) site-specific mutants. These findings were combined with homology modeling of the A1 receptor and in silico screening of an allosteric enhancer library. The binding modes of known docked allosteric enhancers correlated with the known structure-activity relationship, suggesting that these allosteric enhancers bind to a pocket formed by the second extracellular loop, flanked by residues S150 and M162. We propose a model in which this vestibule controls the entry and efflux of agonists from the orthosteric site and agonist binding elicits a conformational change that enables allosteric enhancer binding. This model provides a mechanism for the observations that allosteric enhancers slow the dissociation of orthosteric agonists but not antagonists.

  9. The Role of Estrogen Related Receptor in Modulating Estrogen Receptor Mediated Transcription in Breast Cancer Cells

    Science.gov (United States)

    2005-04-01

    tumors correlates with an unfavorable prognosis (Ariazi 2002; Lu 2001; Suzuki 2004; Vanacker 1999). The transcriptional activity of ERRa is not inhibited...SA. 101:6570-5. Needham, M ., S. Raines, J. McPheat, C. Stacey, J. Ellston, S. Hoare, and M . Parker. 2000. Differential interaction of steroid hormone...R. Graves, M . Wright, and B.M. Spiegelman. 1998. A cold- inducible coactivator of nuclear receptors linked to adaptive thermogenesis. Cell. 92:829- 39

  10. Muscarinic receptors mediate cold stress-induced detrusor overactivity in type 2 diabetes mellitus rats.

    Science.gov (United States)

    Imamura, Tetsuya; Ishizuka, Osamu; Ogawa, Teruyuki; Yamagishi, Takahiro; Yokoyama, Hitoshi; Minagawa, Tomonori; Nakazawa, Masaki; Gautam, Sudha Silwal; Nishizawa, Osamu

    2014-10-01

    This study determined if muscarinic receptors could mediate the cold stress-induced detrusor overactivity induced in type 2 diabetes mellitus rats. Ten-week-old female Goto-Kakizaki diabetic rats (n = 12) and Wister Kyoto non-diabetic rats (n = 12) were maintained on a high-fat diet for 4 weeks. Cystometric investigations of the unanesthetized rats were carried out at room temperature (27 ± 2°C) for 20 min. They were intravenously administered imidafenacin (0.3 mg/kg, n = 6) or vehicle (n = 6). After 5 min, the rats were transferred to a low temperature (4 ± 2°C) for 40 min where the cystometry was continued. The rats were then returned to room temperature for the final cystometric measurements. Afterwards, expressions of bladder muscarinic receptor M3 and M2 messenger ribonucleic acids and proteins were assessed by reverse transcription polymerase chain reaction and immunohistochemistry. In non-diabetic Wister Kyoto rats, imidafenacin did not reduce cold stress-induced detrusor overactivity. In diabetic Goto-Kakizaki rats, just after transfer to a low temperature, the cold stress-induced detrusor overactivity in imidafenacin-treated rats was reduced compared with vehicle-treated rats. Within the urinary bladders, the ratio of M3 to M2 receptor messenger ribonucleic acid in the diabetic Goto-Kakizaki rats was significantly higher than that of the non-diabetic Wister Kyoto rats. The proportion of muscarinic M3 receptor-positive area within the detrusor in diabetic Goto-Kakizaki rats was also significantly higher than that in non-diabetic Wister Kyoto rats. Imidafenacin partially inhibits cold stress-induced detrusor overactivity in diabetic Goto-Kakizaki rats. In this animal model, muscarinic M3 receptors partially mediate cold stress-induced detrusor overactivity. © 2014 The Japanese Urological Association.

  11. Testosterone increases renal anti-aging klotho gene expression via the androgen receptor-mediated pathway.

    Science.gov (United States)

    Hsu, Shih-Che; Huang, Shih-Ming; Lin, Shih-Hua; Ka, Shuk-Man; Chen, Ann; Shih, Meng-Fu; Hsu, Yu-Juei

    2014-12-01

    Gender is known to be associated with longevity and oestrogen administration induced longevity-associated gene expression is one of the potential mechanisms underlying the benefits of oestrogen on lifespan, whereas the role of testosterone in the regulation of longevity-associated gene expressions remains largely unclear. The klotho gene, predominantly expressed in the kidney, has recently been discovered to be an aging suppressor gene. In the present study, we investigated the regulatory effects of testosterone on renal klotho gene expression in vivo and in vitro. In testosterone-administered mouse kidney and NRK-52E cells, increased klotho expression was accompanied by the up-regulation of the nuclear androgen receptor (AR). Overexpression of AR enhanced the expression of klotho mRNA and protein. Conversely, testosterone-induced klotho expression was attenuated in the presence of flutamide, an AR antagonist. A reporter assay and a chromatin immunoprecipitation (ChIP) assay demonstrated that AR directly binds to the klotho promoter via androgen response elements (AREs) which reconfirmed its importance for AR binding via the element mutation. In summary, our study demonstrates that testosterone up-regulates anti-aging klotho together with AR expression in the kidney in vivo and in vitro by recruiting AR on to the AREs of the klotho promoter.

  12. Vitamin D receptor-mediated control of Soggy, Wise, and Hairless gene expression in keratinocytes.

    Science.gov (United States)

    Hsieh, Jui-Cheng; Estess, Rudolf C; Kaneko, Ichiro; Whitfield, G Kerr; Jurutka, Peter W; Haussler, Mark R

    2014-02-01

    The vitamin D receptor (VDR), but not its hormonal ligand, 1,25-dihydroxyvitamin D3 (1,25D), is required for the progression of the mammalian hair cycle. We studied three genes relevant to hair cycle signaling, DKKL1 (Soggy), SOSTDC1 (Wise), and HR (Hairless), to determine whether their expression is regulated by VDR and/or its 1,25D ligand. DKKL1 mRNA was repressed 49-72% by 1,25D in primary human and CCD-1106 KERTr keratinocytes; a functional vitamin D responsive element (VDRE) was identified at -9590 bp in murine Soggy. Similarly, SOSTDC1 mRNA was repressed 41-59% by 1,25D in KERTr and primary human keratinocytes; a functional VDRE was located at -6215 bp in human Wise. In contrast, HR mRNA was upregulated 1.56- to 2.77-fold by 1,25D in primary human and KERTr keratinocytes; a VDRE (TGGTGAgtgAGGACA) consisting of an imperfect direct repeat separated by three nucleotides (DR3) was identified at -7269 bp in the human Hairless gene that mediated dramatic induction, even in the absence of 1,25D ligand. In parallel, a DR4 thyroid hormone responsive element, TGGTGAggccAGGACA, was identified at +1304 bp in the human HR gene that conferred tri-iodothyronine (T3)-independent transcriptional activation. Because the thyroid hormone receptor controls HR expression in the CNS, whereas VDR functions in concert with the HR corepressor specifically in skin, a model is proposed wherein unliganded VDR upregulates the expression of HR, the gene product of which acts as a downstream comodulator to feedback-repress DKKL1 and SOSTDC1, resulting in integration of bone morphogenic protein and Wnt signaling to drive the mammalian hair cycle and/or influencing epidermal function.

  13. Collagen Type I as a Ligand for Receptor-Mediated Signaling

    Directory of Open Access Journals (Sweden)

    Iris Boraschi-Diaz

    2017-05-01

    Full Text Available Collagens form the fibrous component of the extracellular matrix in all multi-cellular animals. Collagen type I is the most abundant collagen present in skin, tendons, vasculature, as well as the organic portion of the calcified tissue of bone and teeth. This review focuses on numerous receptors for which collagen acts as a ligand, including integrins, discoidin domain receptors DDR1 and 2, OSCAR, GPVI, G6b-B, and LAIR-1 of the leukocyte receptor complex (LRC and mannose family receptor uPARAP/Endo180. We explore the process of collagen production and self-assembly, as well as its degradation by collagenases and gelatinases in order to predict potential temporal and spatial sites of action of different collagen receptors. While the interactions of the mature collagen matrix with integrins and DDR are well-appreciated, potential signals from immature matrix as well as collagen degradation products are possible but not yet described. The role of multiple collagen receptors in physiological processes and their contribution to pathophysiology of diseases affecting collagen homeostasis require further studies.

  14. Feedback regulation of mitochondria by caspase-9 in the B cell receptor-mediated apoptosis.

    Science.gov (United States)

    Eeva, J; Nuutinen, U; Ropponen, A; Mättö, M; Eray, M; Pellinen, R; Wahlfors, J; Pelkonen, J

    2009-12-01

    During the germinal centre reaction (GC), B cells with non-functional or self-reactive antigen receptors are negatively selected by apoptosis to generate B cell repertoire with appropriate antigen specificities. We studied the molecular mechanism of Fas/CD95- and B cell receptor (BCR)-induced apoptosis to shed light on the signalling events involved in the negative selection of GC B cells. As an experimental model, we used human follicular lymphoma (FL) cell line HF1A3, which originates from a GC B cell, and transfected HF1A3 cell lines overexpressing Bcl-x(L), c-FLIP(long) or dominant negative (DN) caspase-9. Fas-induced apoptosis was dependent on the caspase-8 activation, since the overexpression of c-FLIP(long), a natural inhibitor of caspase-8 activation, blocked apoptosis induced by Fas. In contrast, caspase-9 activation was not involved in Fas-induced apoptosis. BCR-induced apoptosis showed the typical characteristics of mitochondria-dependent (intrinsic) apoptosis. Firstly, the activation of caspase-9 was involved in BCR-induced DNA fragmentation, while caspase-8 showed only marginal role. Secondly, overexpression of Bcl-x(L) could block all apoptotic changes induced by BCR. As a novel finding, we demonstrate that caspase-9 can enhance the cytochrome-c release and collapse of mitochondrial membrane potential (DeltaPsi(m)) during BCR-induced apoptosis. The requirement of different signalling pathways in apoptosis induced by BCR and Fas may be relevant, since Fas- and BCR-induced apoptosis can thus be regulated independently, and targeted to different subsets of GC B cells.

  15. Targeting of liposomes to cells bearing nerve growth factor receptors mediated by biotinylated NGF

    International Nuclear Information System (INIS)

    Rosenberg, M.B.

    1986-01-01

    Previous studies of liposome targeting have concentrated on immunological systems, the use of ligand-receptor interactions has received little attention. The protein hormone beta-nerve growth factor (NGF) was modified by biotinylation via carboxyl group substitution (C-bio-NGF) under reaction conditions that yielded an average of 3 biotin additions per NGF subunit. NGF was also biotinylated through amino group substitution to produce derivatives with ratios of 1, 2 and 4 biotin moieties per NGF subunit (N-bio-NGF). These derivatives were compared with native NGF for their ability to compete with 125 I-NGF for binding to NGF receptors on rat pheochromocytoma (PC 12) cells at 4 0 C. C-bio-NGF was as effective as native NGF in binding to NGF receptors, while N-bio-NGF containing 1 biotin per NGF subunit was only 28% as active in binding as native NGF. C-bio-NGF, but not N-bio-NGF, mediated the specific binding of 125 I-streptavidin to PC12 cells. Biocytin-NGF, a derivative of C-bio-NGF with an extended spacer chain, was also synthesized and retained full biological and receptor binding activities. C-bio-NGF and biocytin-NGF were as effective as native NGF in a bioassay involving induction of neurite outgrowth from PC12 cells

  16. Ran GTPase promotes cancer progression via Met receptor-mediated downstream signaling

    Science.gov (United States)

    Yuen, Hiu-Fung; Chan, Ka-Kui; Platt-Higgins, Angela; Dakir, El-Habib; Matchett, Kyle B.; Haggag, Yusuf Ahmed; Jithesh, Puthen V.; Habib, Tanwir; Faheem, Ahmed; Dean, Fennell A.; Morgan, Richard; Rudland, Philip S.; El-Tanani, Mohamed

    2016-01-01

    It has been shown previously that cancer cells with an activated oncogenic pathway, including Met activation, require Ran for growth and survival. Here, we show that knockdown of Ran leads to a reduction of Met receptor expression in several breast and lung cancer cell lines. This, in turn suppressed HGF expression and the Met-mediated activation of the Akt pathway, as well as cell adhesion, migration, and invasion. In a cell line model where Met amplification has previously been shown to contribute to gefitinib resistance, Ran knockdown sensitized cells to gefitinib-mediated inhibition of Akt and ERK1/2 phosphorylation and consequently reduced cell proliferation. We further demonstrate that Met reduction-mediated by knockdown of Ran, occurs at the post-transcriptional level, probably via a matrix metalloproteinase. Moreover, the level of immunoreactive Ran and Met are positively associated in human breast cancer specimens, suggesting that a high level of Ran may be a pre-requisite for Met overexpression. Interestingly, a high level of immunoreactive Ran dictates the prognostic significance of Met, indicating that the co-overexpression of Met and Ran may be associated with cancer progression and could be used in combination as a prognostic indicator. PMID:27716616

  17. PTH/PTHrP Receptor Mediates Cachexia in Models of Kidney Failure and Cancer.

    Science.gov (United States)

    Kir, Serkan; Komaba, Hirotaka; Garcia, Ana P; Economopoulos, Konstantinos P; Liu, Wei; Lanske, Beate; Hodin, Richard A; Spiegelman, Bruce M

    2016-02-09

    Cachexia is a wasting syndrome associated with elevated basal energy expenditure and loss of adipose and muscle tissues. It accompanies many chronic diseases including renal failure and cancer and is an important risk factor for mortality. Our recent work demonstrated that tumor-derived PTHrP drives adipose tissue browning and cachexia. Here, we show that PTH is involved in stimulating a thermogenic gene program in 5/6 nephrectomized mice that suffer from cachexia. Fat-specific knockout of PTHR blocked adipose browning and wasting. Surprisingly, loss of PTHR in fat tissue also preserved muscle mass and improved muscle strength. Similarly, PTHR knockout mice were resistant to cachexia driven by tumors. Our results demonstrate that PTHrP and PTH mediate wasting through a common mechanism involving PTHR, and there exists an unexpected crosstalk mechanism between wasting of fat tissue and skeletal muscle. Targeting the PTH/PTHrP pathway may have therapeutic uses in humans with cachexia. Copyright © 2016 Elsevier Inc. All rights reserved.

  18. P2X7 receptor-mediated analgesia in cancer-induced bone pain

    DEFF Research Database (Denmark)

    Falk, Sarah; D. Schwab, Samantha; Frøsig-Jørgensen, Majbrit

    2015-01-01

    Pain is a common and debilitating complication for cancer patients significantly compromising their quality of life. Cancer-induced bone pain involves a complex interplay of molecular events, including mechanisms observed in inflammatory and neuropathic pain states, but also changes unique for ca...

  19. Coenzyme Q blocks biochemical but not receptor-mediated apoptosis by increasing mitochondrial antioxidant protection

    Czech Academy of Sciences Publication Activity Database

    Alleva, R.; Tomasetti, M.; Anděra, Ladislav; Gellert, N.; Borghi, B.; Weber, C.; Murphy, M. P.; Neužil, J.

    2001-01-01

    Roč. 503, č. 1 (2001), s. 46-50 ISSN 0014-5793 R&D Projects: GA ČR GA301/99/0350 Institutional research plan: CEZ:AV0Z5052915 Keywords : coenzyme Q * apoptosis Subject RIV: EB - Genetics ; Molecular Biology Impact factor: 3.644, year: 2001

  20. Evidence that 5-hydroxytryptamine/sub 3/ receptors mediate cytotoxic drug and radiation-evoked emesis

    Energy Technology Data Exchange (ETDEWEB)

    Miner, W.D.; Sanger, G.J.; Turner, D.H.

    1987-08-01

    The involvement of 5-hydroxytryptamine (5-HT) 5-HT/sub 3/ receptors in the mechanisms of severe emesis evoked by cytotoxic drugs or by total body irradiation have been studied in ferrets. Anti-emetic compounds tested were domperidone (a dopamine antagonist), metoclopramide (a gastric motility stimulant and dopamine antagonist at conventional doses, a 5-HT/sub 3/ receptor antagonist at higher doses) and BRL 24924 (a potent gastric motility stimulant and a 5-HT/sub 3/ receptor antagonist). Domperidone or metoclopramide prevented apomorphine-evoked emesis, whereas BRL 24924 did not. Similar doses of domperidone did not prevent emesis evoked by cis-platin or by total body irradiation, whereas metoclopramide or BRL 24924 greatly reduced or prevented these types of emesis. Metoclopramide and BRL 24924 also prevented emesis evoked by a combination of doxorubicin and cyclophosphamide. These results are discussed in terms of a fundamental role for 5-HT/sub 3/ receptors in the mechanisms mediating severely emetogenic cancer treatment therapies.

  1. Adaptive and innate immune reactions regulating mast cell activation: from receptor-mediated signaling to responses

    DEFF Research Database (Denmark)

    Tkaczyk, Christine; Jensen, Bettina M; Iwaki, Shoko

    2006-01-01

    differentially activate multiple signaling pathways within the mast cells required for the generation and/or release of inflammatory mediators. Thus, the composition of the suite of mediators released and the physiologic ramifications of these responses are dependent on the stimuli and the microenvironment...

  2. The effect of benfotiamine on mu-opioid receptor mediated antinociception in experimental diabetes.

    Science.gov (United States)

    Nacitarhan, C; Minareci, E; Sadan, G

    2014-03-01

    Diabetic neuropathy is a prevalent, disabling disorder. Currently, the only treatments available to patients with diabetic neuropathy are glucose control and pain management. B vitamin present neuroprotective effects, which are suggested to be related to their analgesic action in various models of neuropathic pain. According to our literature knowledge there is no report about antinociceptive effects of thiamine as benfotiamine and opioids together in diabetic mice. The purpose of this study was to determine the effects of benfotiamine on the antinociception produced by mu-opioid receptor agonist fentanyl in diabetic mice. The effects of benfotiamine on antinociception produced by fentanyl in diabetic mice were studied in 4 groups. Antinociceptive effect was determined with tail flick, hot plate and formalin test. Our results showed that, mu-opioid agonist fentanyl in benfotiamine applied diabetic group caused more potent antinociceptive effect than in diabetic group without benfotiamine treatment. In brief benfotiamine supplement in diet did not bring out antinociceptive effect itself, but during development of STZ diabetes, benfotiamine replacement increased the antinociceptive effect of fentanyl in mice tail-flick test. This effect is probably due to the replacement of benfotiamine efficiency occurring in diabetes mellitus. Finally, we suppose that oral benfotiamine replacement therapy may be useful to ameliorate analgesic effect of mu-opioid agonists on neuropathic pain in diabetic case. © J. A. Barth Verlag in Georg Thieme Verlag KG Stuttgart · New York.

  3. The TIM and TAM families of phosphatidylserine receptors mediate dengue virus entry.

    Science.gov (United States)

    Meertens, Laurent; Carnec, Xavier; Lecoin, Manuel Perera; Ramdasi, Rasika; Guivel-Benhassine, Florence; Lew, Erin; Lemke, Greg; Schwartz, Olivier; Amara, Ali

    2012-10-18

    Dengue viruses (DVs) are responsible for the most medically relevant arboviral diseases. However, the molecular interactions mediating DV entry are poorly understood. We determined that TIM and TAM proteins, two receptor families that mediate the phosphatidylserine (PtdSer)-dependent phagocytic removal of apoptotic cells, serve as DV entry factors. Cells poorly susceptible to DV are robustly infected after ectopic expression of TIM or TAM receptors. Conversely, DV infection of susceptible cells is inhibited by anti-TIM or anti-TAM antibodies or knockdown of TIM and TAM expression. TIM receptors facilitate DV entry by directly interacting with virion-associated PtdSer. TAM-mediated infection relies on indirect DV recognition, in which the TAM ligand Gas6 acts as a bridging molecule by binding to PtdSer within the virion. This dual mode of virus recognition by TIM and TAM receptors reveals how DVs usurp the apoptotic cell clearance pathway for infectious entry. Copyright © 2012 Elsevier Inc. All rights reserved.

  4. Astrocytic β2-adrenergic receptors mediate hippocampal long-term memory consolidation

    KAUST Repository

    Gao, Virginia

    2016-07-12

    Emotionally relevant experiences form strong and long-lasting memories by critically engaging the stress hormone/neurotransmitter noradrenaline, which mediates and modulates the consolidation of these memories. Noradrenaline acts through adrenergic receptors (ARs), of which β2- Adrenergic receptors (βARs) are of particular importance. The differential anatomical and cellular distribution of βAR subtypes in the brain suggests that they play distinct roles in memory processing, although much about their specific contributions and mechanisms of action remains to be understood. Here we show that astrocytic rather than neuronal β2ARs in the hippocampus play a key role in the consolidation of a fear-based contextual memory. These hippocampal β2ARs, but not β1ARs, are coupled to the training-dependent release of lactate from astrocytes, which is necessary for long- Term memory formation and for underlying molecular changes. This key metabolic role of astrocytic β2ARs may represent a novel target mechanism for stress-related psychopathologies and neurodegeneration.

  5. Interaction of medullary P2 and glutamate receptors mediates the vasodilation in the hindlimb of rat.

    Science.gov (United States)

    Korim, Willian Seiji; Ferreira-Neto, Marcos L; Pedrino, Gustavo R; Pilowsky, Paul M; Cravo, Sergio L

    2012-12-01

    In the nucleus tractus solitarii (NTS) of rats, blockade of extracellular ATP breakdown to adenosine reduces arterial blood pressure (AP) increases that follow stimulation of the hypothalamic defense area (HDA). The effects of ATP on NTS P2 receptors, during stimulation of the HDA, are still unclear. The aim of this study was to determine whether activation of P2 receptors in the NTS mediates cardiovascular responses to HDA stimulation. Further investigation was taken to establish if changes in hindlimb vascular conductance (HVC) elicited by electrical stimulation of the HDA, or activation of P2 receptors in the NTS, are relayed in the rostral ventrolateral medulla (RVLM); and if those responses depend on glutamate release by ATP acting on presynaptic terminals. In anesthetized and paralyzed rats, electrical stimulation of the HDA increased AP and HVC. Blockade of P2 or glutamate receptors in the NTS, with bilateral microinjections of suramin (10 mM) or kynurenate (50 mM) reduced only the evoked increase in HVC by 75 % or more. Similar results were obtained with the blockade combining both antagonists. Blockade of P2 and glutamate receptors in the RVLM also reduced the increases in HVC to stimulation of the HDA by up to 75 %. Bilateral microinjections of kynurenate in the RVLM abolished changes in AP and HVC to injections of the P2 receptor agonist α,β-methylene ATP (20 mM) into the NTS. The findings suggest that HDA-NTS-RVLM pathways in control of HVC are mediated by activation of P2 and glutamate receptors in the brainstem in alerting-defense reactions.

  6. DOTATOC: a powerful new tool for receptor-mediated radionuclide therapy

    International Nuclear Information System (INIS)

    Otte, A.; Jermann, E.; Behe, M.; Goetze, M.; Bucher, H.C.; Roser, H.W.; Heppeler, A.; Mueller-Brand, J.; Maecke, H.R.

    1997-01-01

    This study presents the first successful use of a peptidic vector, DOTATOC, labelled with the β-emitting radioisotope yttrium-90, for the treatment of a patient with somatostatin receptor-positive abdominal metastases of a neuroendocrine carcinoma of unknown localization. Tumour response and symptomatic relief were achieved. In addition, the new substance DOTATOC was labelled with the diagnostic chemical analogue indium-111 and studied in three patients with histopathologically verified neuroendocrine abdominal tumours for its diagnostic sensitivity and compared with the commercially available OctreoScan. In all patients the kidney-to-tumour uptake ratio (in counts per pixel) was on average 1.9-fold lower with 111 In-DOTATOC than with OctreoScan. DOTATOC could be a potential new diagnostic and therapeutic agent in the management of neuroendocrine tumours. (orig.). With 4 figs., 1 tab

  7. Effects of receptor-mediated endocytosis and tubular protein composition on volume retention in experimental glomerulonephritis

    DEFF Research Database (Denmark)

    Kastner, Christian; Pohl, Marcus; Sendeski, Mauricio

    2009-01-01

    Human glomerulonephritis (GN) is characterized by sustained proteinuria, sodium retention, hypertension, and edema formation. Increasing quantities of filtered protein enter the renal tubule, where they may alter epithelial transport functions. Exaggerated endocytosis and consequent protein...... overload may affect proximal tubules, but intrinsic malfunction of distal epithelia has also been reported. A straightforward assignment to a particular tubule segment causing salt retention in GN is still controversial. We hypothesized that 1) trafficking and surface expression of major transporters...

  8. Amygdala mu-opioid receptors mediate the motivating influence of cue-triggered reward expectations.

    Science.gov (United States)

    Lichtenberg, Nina T; Wassum, Kate M

    2017-02-01

    Environmental reward-predictive stimuli can retrieve from memory a specific reward expectation that allows them to motivate action and guide choice. This process requires the basolateral amygdala (BLA), but little is known about the signaling systems necessary within this structure. Here we examined the role of the neuromodulatory opioid receptor system in the BLA in such cue-directed action using the outcome-specific Pavlovian-to-instrumental transfer (PIT) test in rats. Inactivation of BLA mu-, but not delta-opioid receptors was found to dose-dependently attenuate the ability of a reward-predictive cue to selectively invigorate the performance of actions directed at the same unique predicted reward (i.e. to express outcome-specific PIT). BLA mu-opioid receptor inactivation did not affect the ability of a reward itself to similarly motivate action (outcome-specific reinstatement), suggesting a more selective role for the BLA mu-opioid receptor in the motivating influence of currently unobservable rewarding events. These data reveal a new role for BLA mu-opioid receptor activation in the cued recall of precise reward memories and the use of this information to motivate specific action plans. © 2016 Federation of European Neuroscience Societies and John Wiley & Sons Ltd.

  9. Thyrotropin modulates receptor-mediated processing of the atrial natriuretic peptide receptor in cultured thyroid cells

    International Nuclear Information System (INIS)

    Tseng, Y.L.; Burman, K.D.; Lahiri, S.; Abdelrahim, M.M.; D'Avis, J.C.; Wartofsky, L.

    1991-01-01

    In a prior study of atrial natriuretic peptide (ANP) binding to cultured thyroid cells, we reported that at 4 C, more than 95% of bound ANP is recovered on cell membranes, with negligible ANP internalization observed. Since ANP binding was inhibited by TSH, we have further studied TSH effects on postbinding ANP processing to determine whether this phenomenon reflects enhanced endocytosis of the ANP-receptor complex. An ANP chase study was initiated by binding [125I] ANP to thyroid cells at 4 C for 2 h, followed by incubation at 37 C. ANP processing was then traced by following 125I activity at various time intervals in three fractions: cell surface membranes, incubation medium, and inside the cells. Radioactivity released into medium represented processed ANP rather than ANP dissociated from surface membranes, since prebound [125I]ANP could not be competitively dissociated by a high concentration of ANP (1 mumol/L) at 37 C. Chase study results showed that prebound ANP quickly disappeared from cell membranes down to 34% by 30 min. Internalized ANP peaked at 10 min, with 21% of initial prebound ANP found inside the cells. At the same time, radioactivity recovered in incubation medium sharply increased between 10-30 min from 8% to 52%. Preincubation of cells with chloroquine (which blocks degradation of the ANP-receptor complex by inhibiting lysosomal hydrolase) caused a 146% increase in internalized [125I]ANP by 30 min (39% compared to 15% control), while medium radioactivity decreased from 52% to 16%, suggesting that processing of the receptor complex is mediated via lysosomal enzymes. In chase studies employing cells pretreated with chloroquine, TSH stimulated the internalization rate of ANP-receptor complex. By 30 min, TSH significantly reduced the membrane-bound ANP, and the decrease was inversely correlated to the increase in internalized radioactivity

  10. Direct and indirect endocrine disruption : aromatase and estrogen receptor-mediated processes in breast cancer development

    NARCIS (Netherlands)

    Heneweer, Marjoke

    2005-01-01

    Endocrine disrupting chemicals (EDCs) have been defined by the World Health Organization as: “exogenous substances or mixtures that alters function(s) of the endocrine system and causes adverse health effects in an intact organism, or its progeny, or (sub)populations”. Synthetic, as well as,

  11. Receptor-Mediated Entry of Pristine Octahedral DNA Nanocages in Mammalian Cells

    DEFF Research Database (Denmark)

    Vindigni, Giulia; Raniolo, Sofia; Ottaviani, Alessio

    2016-01-01

    , more recently, identified as a tumor marker. For this purpose a truncated octahedral DNA nanocage functionalized with a single biotin molecule, which allows DNA cage detection through the biotin–streptavidin assays, was constructed. The results indicate that DNA nanocages are stable in biological...

  12. Molecular and functional profiling of histamine receptor-mediated calcium ion signals in different cell lines.

    Science.gov (United States)

    Meisenberg, Annika; Kaschuba, Dagmar; Balfanz, Sabine; Jordan, Nadine; Baumann, Arnd

    2015-10-01

    Calcium ions (Ca(2+)) play a pivotal role in cellular physiology. Often Ca(2+)-dependent processes are studied in commonly available cell lines. To induce Ca(2+) signals on demand, cells may need to be equipped with additional proteins. A prominent group of membrane proteins evoking Ca(2+) signals are G-protein coupled receptors (GPCRs). These proteins register external signals such as photons, odorants, and neurotransmitters and convey ligand recognition into cellular responses, one of which is Ca(2+) signaling. To avoid receptor cross-talk or cross-activation with introduced proteins, the repertoire of cell-endogenous receptors must be known. Here we examined the presence of histamine receptors in six cell lines frequently used as hosts to study cellular signaling processes. In a concentration-dependent manner, histamine caused a rise in intracellular Ca(2+) in HeLa, HEK 293, and COS-1 cells. The concentration for half-maximal activation (EC50) was in the low micromolar range. In individual cells, transient Ca(2+) signals and Ca(2+) oscillations were uncovered. The results show that (i) HeLa, HEK 293, and COS-1 cells express sufficient amounts of endogenous receptors to study cellular Ca(2+) signaling processes directly and (ii) these cell lines are suitable for calibrating Ca(2+) biosensors in situ based on histamine receptor evoked responses. Copyright © 2015 Elsevier Inc. All rights reserved.

  13. Gamma-secretase activity of presenilin 1 regulates acetylcholine muscarinic receptor-mediated signal transduction

    DEFF Research Database (Denmark)

    Popescu, Bogdan O; Cedazo-Minguez, Angel; Benedikz, Eirikur

    2004-01-01

    , we studied the effect of two other FAD PS1 mutants (M146V and L250S) and two dominant negative PS1 mutants (D257A and D385N) on basal and carbachol-stimulated phosphoinositide (PI) hydrolysis and intracellular calcium concentrations ([Ca2+]i) in SH-SY5Y neuroblastoma cells. We found a significant...

  14. Dopamine D2 Receptor-Mediated Regulation of Pancreatic β Cell Mass

    Directory of Open Access Journals (Sweden)

    Daisuke Sakano

    2016-07-01

    Full Text Available Understanding the molecular mechanisms that regulate β cell mass and proliferation is important for the treatment of diabetes. Here, we identified domperidone (DPD, a dopamine D2 receptor (DRD2 antagonist that enhances β cell mass. Over time, islet β cell loss occurs in dissociation cultures, and this was inhibited by DPD. DPD increased proliferation and decreased apoptosis of β cells through increasing intracellular cAMP. DPD prevented β cell dedifferentiation, which together highly contributed to the increased β cell mass. DRD2 knockdown phenocopied the effects of domperidone and increased the number of β cells. Drd2 overexpression sensitized the dopamine responsiveness of β cells and increased apoptosis. Further analysis revealed that the adenosine agonist 5′-N-ethylcarboxamidoadenosine, a previously identified promoter of β cell proliferation, acted with DPD to increase the number of β cells. In humans, dopamine also modulates β cell mass through DRD2 and exerts an inhibitory effect on adenosine signaling.

  15. Regulation of granulocyte colony-stimulating factor receptor-mediated granulocytic differentiation by C-mannosylation.

    Science.gov (United States)

    Otani, Kei; Niwa, Yuki; Suzuki, Takehiro; Sato, Natsumi; Sasazawa, Yukiko; Dohmae, Naoshi; Simizu, Siro

    2018-04-06

    Granulocyte colony-stimulating factor (G-CSF) receptor (G-CSFR) is a type I cytokine receptor which is involved in hematopoietic cell maturation. G-CSFR has three putative C-mannosylation sites at W253, W318, and W446; however, it is not elucidated whether G-CSFR is C-mannosylated or not. In this study, we first demonstrated that G-CSFR was C-mannosylated at only W318. We also revealed that C-mannosylation of G-CSFR affects G-CSF-dependent downstream signaling through changing ligand binding capability but not cell surface localization. Moreover, C-mannosylation of G-CSFR was functional and regulated granulocytic differentiation in myeloid 32D cells. In conclusion, we found that G-CSFR is C-mannosylated at W318 and that this C-mannosylation has role(s) for myeloid cell differentiation through regulating downstream signaling. Copyright © 2018 Elsevier Inc. All rights reserved.

  16. Pharmacological and biochemical characterization of the D-1 dopamine receptor mediating acetylcholine release in rabbit retina

    International Nuclear Information System (INIS)

    Hensler, J.G.; Cotterell, D.J.; Dubocovich, M.L.

    1987-01-01

    Superfusion with dopamine (0.1 microM-10 mM) evokes calcium-dependent [ 3 H]acetylcholine release from rabbit retina labeled in vitro with [ 3 H]choline. This effect is antagonized by the D-1 dopamine receptor antagonist SCH 23390. Activation or blockade of D-2 dopamine, alpha-2 or beta receptors did not stimulate or attenuate the release of [ 3 H]acetylcholine from rabbit retina. Dopamine receptor agonists evoke the release of [ 3 H]acetylcholine with the following order of potency: apomorphine ≤ SKF(R)82526 3 H]acetylcholine: SCH 23390 (IC50 = 1 nM) 3 H]acetylcholine release is characteristic of the D-1 dopamine receptor. These potencies were correlated with the potencies of dopamine receptor agonists and antagonists at the D-1 dopamine receptor in rabbit retina as labeled by [ 3 H]SCH 23390, or as determined by adenylate cyclase activity. [ 3 H]SCH 23390 binding in rabbit retinal membranes was stable, saturable and reversible. Scatchard analysis of [ 3 H]SCH 23390 saturation data revealed a single high affinity binding site (Kd = 0.175 +/- 0.002 nM) with a maximum binding of 482 +/- 12 fmol/mg of protein. The potencies of dopamine receptor agonists to stimulate [ 3 H]acetylcholine release were correlated with their potencies to stimulate adenylate cyclase (r = 0.784, P less than .05, n = 7) and with their affinities at [ 3 H]SCH 23390 binding sites (r = 0.755, P < .05, n = 8)

  17. Receptor-mediated endocytosis of α-galactosidase A in human podocytes in Fabry disease.

    Directory of Open Access Journals (Sweden)

    Thaneas Prabakaran

    Full Text Available Injury to the glomerular podocyte is a key mechanism in human glomerular disease and podocyte repair is an important therapeutic target. In Fabry disease, podocyte injury is caused by the intracellular accumulation of globotriaosylceramide. This study identifies in the human podocyte three endocytic receptors, mannose 6-phosphate/insulin-like growth II receptor, megalin, and sortilin and demonstrates their drug delivery capabilities for enzyme replacement therapy. Sortilin, a novel α-galactosidase A binding protein, reveals a predominant intracellular expression but also surface expression in the podocyte. The present study provides the rationale for the renal effect of treatment with α-galactosidase A and identifies potential pathways for future non-carbohydrate based drug delivery to the kidney podocyte and other potential affected organs.

  18. Accumbens Shell AMPA Receptors Mediate Expression of Extinguished Reward Seeking through Interactions with Basolateral Amygdala

    Science.gov (United States)

    Millan, E. Zayra; McNally, Gavan P.

    2011-01-01

    Extinction is the reduction in drug seeking when the contingency between drug seeking behavior and the delivery of drug reward is broken. Here, we investigated a role for the nucleus accumbens shell (AcbSh). Rats were trained to respond for 4% (v/v) alcoholic beer in one context (Context A) followed by extinction in a second context (Context B).…

  19. Mannose Receptor Mediates the Immune Response to Ganoderma atrum Polysaccharides in Macrophages.

    Science.gov (United States)

    Li, Wen-Juan; Tang, Xiao-Fang; Shuai, Xiao-Xue; Jiang, Cheng-Jia; Liu, Xiang; Wang, Le-Feng; Yao, Yu-Fei; Nie, Shao-Ping; Xie, Ming-Yong

    2017-01-18

    The ability of mannose receptor (MR) to recognize the carbohydrate structures is well-established. Here, we reported that MR was crucial for the immune response to a Ganoderma atrum polysaccharide (PSG-1), as evidenced by elevation of MR in association with increase of phagocytosis and concentrations of interleukin-1β (IL-1β) and tumor necrosis factor-α (TNF-α) in normal macrophages. Elevation of MR triggered by PSG-1 also led to control lipopolysaccharide (LPS)-triggered inflammatory response via the increase of interleukin-10 (IL-10) and inhibition of phagocytosis and IL-1β. Anti-MR antibody partly attenuated PSG-1-mediated anti-inflammatory responses, while it could not affect TNF-α secretion, suggesting that another receptor was involved in PSG-1-triggered immunomodulatory effects. MR and toll-like receptor (TLR)4 coordinated the influences on the TLR4-mediated signaling cascade by the nuclear factor-κB (NF-κB) pathway in LPS-stimulated macrophages subjected to PSG-1. Collectively, immune response to PSG-1 required recognition by MR in macrophages. The NF-κB pathway served as a central role for the coordination of MR and TLR4 to elicit immune response to PSG-1.

  20. Ionotropic glutamate receptors mediate inducible defense in the water flea Daphnia pulex.

    Directory of Open Access Journals (Sweden)

    Hitoshi Miyakawa

    Full Text Available Phenotypic plasticity is the ability held in many organisms to produce different phenotypes with a given genome in response to environmental stimuli, such as temperature, nutrition and various biological interactions. It seems likely that environmental signals induce a variety of mechanistic responses that influence ontogenetic processes. Inducible defenses, in which prey animals alter their morphology, behavior and/or other traits to help protect against direct or latent predation threats, are among the most striking examples of phenotypic plasticity. The freshwater microcrustacean Daphnia pulex forms tooth-like defensive structures, "neckteeth," in response to chemical cues or signals, referred to as "kairomones," in this case released from phantom midge larvae, a predator of D. pulex. To identify factors involved in the reception and/or transmission of a kairomone, we used microarray analysis to identify genes up-regulated following a short period of exposure to the midge kairomone. In addition to identifying differentially expressed genes of unknown function, we also found significant up-regulation of genes encoding ionotropic glutamate receptors, which are known to be involved in neurotransmission in many animal species. Specific antagonists of these receptors strongly inhibit the formation of neckteeth in D. pulex, although agonists did not induce neckteeth by themselves, indicating that ionotropic glutamate receptors are necessary but not sufficient for early steps of neckteeth formation in D. pulex. Moreover, using co-exposure of D. pulex to antagonists and juvenile hormone (JH, which physiologically mediates neckteeth formation, we found evidence suggesting that the inhibitory effect of antagonists is not due to direct inhibition of JH synthesis/secretion. Our findings not only provide a candidate molecule required for the inducible defense response in D. pulex, but also will contribute to the understanding of complex mechanisms underlying the recognition of environmental changes, which form the basis of phenotypic plasticity.

  1. Evidence that 5-hydroxytryptamine3 receptors mediate cytotoxic drug and radiation-evoked emesis

    International Nuclear Information System (INIS)

    Miner, W.D.; Sanger, G.J.; Turner, D.H.

    1987-01-01

    The involvement of 5-hydroxytryptamine (5-HT) 5-HT 3 receptors in the mechanisms of severe emesis evoked by cytotoxic drugs or by total body irradiation have been studied in ferrets. Anti-emetic compounds tested were domperidone (a dopamine antagonist), metoclopramide (a gastric motility stimulant and dopamine antagonist at conventional doses, a 5-HT 3 receptor antagonist at higher doses) and BRL 24924 (a potent gastric motility stimulant and a 5-HT 3 receptor antagonist). Domperidone or metoclopramide prevented apomorphine-evoked emesis, whereas BRL 24924 did not. Similar doses of domperidone did not prevent emesis evoked by cis-platin or by total body irradiation, whereas metoclopramide or BRL 24924 greatly reduced or prevented these types of emesis. Metoclopramide and BRL 24924 also prevented emesis evoked by a combination of doxorubicin and cyclophosphamide. These results are discussed in terms of a fundamental role for 5-HT 3 receptors in the mechanisms mediating severely emetogenic cancer treatment therapies. (author)

  2. Antibody therapy of cancer : Fc receptor-mediated mechanisms of action

    NARCIS (Netherlands)

    Overdijk, M.B.

    2013-01-01

    Cancer, a class of malignant diseases characterized by unregulated cell growth, is still a leading cause of death worldwide. The high specificity of antibodies combined with the ability to engage multiple mechanisms of action (MoA) and minimal side-effects makes them attractive agents for targeted

  3. Cannabinoid Receptors Mediate Methamphetamine Induction of High Frequency Gamma Oscillations in the Nucleus Accumbens

    Science.gov (United States)

    Morra, Joshua T.; Glick, Stanley D.; Cheer, Joseph F.

    2012-01-01

    Patients suffering from amphetamine---induced psychosis display repetitive behaviors, partially alleviated by antipsychotics, which are reminiscent of rodent stereotypies. Due to recent evidence implicating endocannabinoid involvement in brain disorders, including psychosis, we studied the effects of endocannabinoid signaling on neuronal oscillations of rats exhibiting methamphetamine stereotypy. Neuronal network oscillations were recorded with multiple single electrode arrays aimed at the nucleus accumbens of freely moving rats. During the experiments, animals were dosed intravenously with the CB1 receptor antagonist rimonabant (0.3 mg/kg) or vehicle followed by an ascending dose regimen of methamphetamine (0.01, 0.1, 1, and 3 mg/kg; cumulative dosing). The effects of drug administration on stereotypy and local gamma oscillations were evaluated. Methamphetamine treatment significantly increased high frequency gamma oscillations (~ 80 Hz). Entrainment of a subpopulation of nucleus accumbens neurons to high frequency gamma was associated with stereotypy encoding in putative fast-spiking interneurons, but not in putative medium spiny neurons. The observed ability of methamphetamine to induce both stereotypy and high frequency gamma power was potently disrupted following CB1 receptor blockade. The present data suggest that CB1 receptor-dependent mechanisms are recruited by methamphetamine to modify striatal interneuron oscillations that accompany changes in psychomotor state, further supporting the link between endocannabinoids and schizophrenia spectrum disorders. PMID:22609048

  4. Optimization of stress response through the nuclear receptor-mediated cortisol signalling network

    NARCIS (Netherlands)

    Kolodkin, A.; Sahin, N.; Phillips, A.; Hood, S.R.; Bruggeman, F.J.; Westerhoff, H.V.; Plant, N.

    2013-01-01

    It is an accepted paradigm that extended stress predisposes an individual to pathophysiology. However, the biological adaptations to minimize this risk are poorly understood. Using a computational model based upon realistic kinetic parameters we are able to reproduce the interaction of the stress

  5. Optimization of stress response through the nuclear receptor-mediated cortisol signalling network.

    NARCIS (Netherlands)

    Kolodkin, A.; Sahin, N.; Phillips, A.; Hood, S.R.; Bruggeman, F.J.; Westerhoff, H.V.; Plant, N.

    2014-01-01

    It is an accepted paradigm that extended stress predisposes an individual to pathophysiology. However, the biological adaptations to minimize this risk are poorly understood. Using a computational model based upon realistic kinetic parameters we are able to reproduce the interaction of the stress

  6. Role of desensitization and subunit expression for kainate receptor-mediated neurotoxicity in murine neocortical cultures

    DEFF Research Database (Denmark)

    Jensen, J B; Schousboe, A; Pickering, D S

    1999-01-01

    ) toxicity mediated by alpha-amino-3-hydroxy-5-methyl-4-isoxazolepropionate (AMPA) receptors, and (3) toxicity that can be mediated by kainate receptors when desensitization of the receptors is blocked. The indirect action at NMDA receptors was discovered because (5R, 10S)-(+)-5-methyl-10,11-dihydro-5H...... nedioxy-5H-2,3-benzodiazepine (GYKI 53655), a selective AMPA receptor antagonist, abolished the remaining toxicity. These results indicated that kainate- and domoate-mediated toxicity involves both the NMDA and the AMPA receptors. Pretreatment of the cultures with concanavalin A to prevent desensitization...

  7. AMPA receptor mediated excitotoxicity in neocortical neurons is developmentally regulated and dependent upon receptor desensitization

    DEFF Research Database (Denmark)

    Jensen, J B; Schousboe, A; Pickering, D S

    1998-01-01

    with a fast and rapidly desensitizing response, this could explain the relatively low toxicity produced by 500 microM AMPA. This was investigated by blocking AMPA receptor desensitization with cyclothiazide. Using a lower concentration (25 microM) of AMPA, addition of 50 microM cyclothiazide increased...... the AMPA induced excitotoxicity in cultured cortical neurons at all DIV except for DIV 2. This combination of AMPA + cyclothiazide yielded 77% cell death for DIV 12 cultures. In contrast to the results observed with 500 microM AMPA, the neurotoxicity mediated directly by AMPA receptors when desensitization...

  8. AT(2) receptor-mediated vasodilation in the heart: effect of myocardial infarction

    NARCIS (Netherlands)

    M.P. Schuijt; M. Basdew; R. van Veghel; R.R.P. de Vries (René); P.R. Saxena (Pramod Ranjan); R.G. Schoemaker (Regien); A.H.J. Danser (Jan)

    2001-01-01

    textabstractTo investigate the functional consequences of postinfarct cardiac angiotensin (ANG) type 2 (AT(2)) receptor upregulation, rats underwent coronary artery ligation or sham operation and were infused with ANG II 3-4 wk later, when scar formation is complete. ANG II

  9. Signal transduction in cultered cardiomyocytes : alpha1-adrenergic and endothelin receptor mediated responses

    NARCIS (Netherlands)

    H.W. de Jonge (Jet)

    1996-01-01

    textabstractAlready in ancient times the Greek were aware of the heart in the human body and they gave it the name kardia, which is still in use in words as cardiac, myocardial, tachycardia and bradycardia. In those times the importance of the heart was appraised by Aristotle (384-322 B.C.), who

  10. Sigma-1 Receptor Mediates Acquisition of Alcohol Drinking and Seeking behavior in Alcohol-Preferring Rats

    Science.gov (United States)

    Blasio, Angelo; Valenza, Marta; Iyer, Malliga R.; Rice, Kenner C.; Steardo, Luca; Hayashi, T.; Cottone, Pietro; Sabino, Valentina

    2015-01-01

    Sigma-1 receptor (Sig-1R) has been proposed as a novel therapeutic target for drug and alcohol addiction. We have shown previously that Sig-1R agonists facilitate the reinforcing effects of ethanol and induce binge-like drinking, while Sig-1R antagonists block excessive drinking in both genetic and environmental models of alcoholism, without affecting intake in outbred non-dependent rats. Even though significant progress has been made in understanding the function of Sig-1Rs in alcohol reinforcement, its role in the early and late stage of alcohol addiction remains unclear. Administration of the selective Sig-1R antagonist BD-1063 dramatically reduced the acquisition of alcohol drinking behavior as well as the preference for alcohol in genetically selected TSRI Sardinian alcohol preferring (Scr:sP) rats; the treatment had no effect on total fluid intake, food intake or body weight gain, proving selectivity of action. Furthermore, BD-1063 dose-dependently decreased alcohol-seeking behavior in rats trained under a second-order schedule of reinforcement, in which responding is maintained by contingent presentation of a conditioned reinforcer. Finally, an innate elevation in Sig-1R protein levels was found in the nucleus accumbens of alcohol-preferring Scr:sP rats, compared to outbred Wistar rats, alteration which was normalized by chronic, voluntary alcohol drinking. Taken together these findings demonstrate that Sig-1R blockade reduces the propensity to both acquire alcohol drinking and to seek alcohol, and point to the nucleus accumbens as a potential key region for the effects observed. Our data suggest that Sig-1R antagonists may have therapeutic potential in multiple stages of alcohol addiction. PMID:25848705

  11. Pax6 represses androgen receptor-mediated transactivation by inhibiting recruitment of the coactivator SPBP.

    Directory of Open Access Journals (Sweden)

    Julianne Elvenes

    Full Text Available The androgen receptor (AR has a central role in development and maintenance of the male reproductive system and in the etiology of prostate cancer. The transcription factor Pax6 has recently been reported to act as a repressor of AR and to be hypermethylated in prostate cancer cells. SPBP is a transcriptional regulator that previously has been shown to enhance the activity of Pax6. In this study we have identified SPBP to act as a transcriptional coactivator of AR. We also show that Pax6 inhibits SPBP-mediated enhancement of AR activity on the AR target gene probasin promoter, a repression that was partly reversed by increased expression of SPBP. Enhanced expression of Pax6 reduced the amount of SPBP associated with the probasin promoter when assayed by ChIP in HeLa cells. We mapped the interaction between both AR and SPBP, and AR and Pax6 to the DNA-binding domains of the involved proteins. Further binding studies revealed that Pax6 and SPBP compete for binding to AR. These results suggest that Pax6 represses AR activity by displacing and/or inhibiting recruitment of coactivators to AR target promoters. Understanding the mechanism for inhibition of AR coactivators can give rise to molecular targeted drugs for treatment of prostate cancer.

  12. Impaired P2X1 Receptor-Mediated Adhesion in Eosinophils from Asthmatic Patients.

    Science.gov (United States)

    Wright, Adam; Mahaut-Smith, Martyn; Symon, Fiona; Sylvius, Nicolas; Ran, Shaun; Bafadhel, Mona; Muessel, Michelle; Bradding, Peter; Wardlaw, Andrew; Vial, Catherine

    2016-06-15

    Eosinophils play an important role in the pathogenesis of asthma and can be activated by extracellular nucleotides released following cell damage or inflammation. For example, increased ATP concentrations were reported in bronchoalveolar lavage fluids of asthmatic patients. Although eosinophils are known to express several subtypes of P2 receptors for extracellular nucleotides, their function and contribution to asthma remain unclear. In this article, we show that transcripts for P2X1, P2X4, and P2X5 receptors were expressed in healthy and asthmatic eosinophils. The P2X receptor agonist α,β-methylene ATP (α,β-meATP; 10 μM) evoked rapidly activating and desensitizing inward currents (peak 18 ± 3 pA/pF at -60 mV) in healthy eosinophils, typical of P2X1 homomeric receptors, which were abolished by the selective P2X1 antagonist NF449 (1 μM) (3 ± 2 pA/pF). α,β-meATP-evoked currents were smaller in eosinophils from asthmatic patients (8 ± 2 versus 27 ± 5 pA/pF for healthy) but were enhanced following treatment with a high concentration of the nucleotidase apyrase (17 ± 5 pA/pF for 10 IU/ml and 11 ± 3 pA/pF for 0.32 IU/ml), indicating that the channels are partially desensitized by extracellular nucleotides. α,β-meATP (10 μM) increased the expression of CD11b activated form in eosinophils from healthy, but not asthmatic, donors (143 ± 21% and 108 ± 11% of control response, respectively). Furthermore, α,β-meATP increased healthy (18 ± 2% compared with control 10 ± 1%) but not asthmatic (13 ± 1% versus 10 ± 0% for control) eosinophil adhesion. Healthy human eosinophils express functional P2X1 receptors whose activation leads to eosinophil αMβ2 integrin-dependent adhesion. P2X1 responses are constitutively reduced in asthmatic compared with healthy eosinophils, probably as the result of an increase in extracellular nucleotide concentration. Copyright © 2016 by The American Association of Immunologists, Inc.

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

    Science.gov (United States)

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

    2016-01-01

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

  14. Proteinase-activated receptors - mediators of early and delayed normal tissue radiation responses

    International Nuclear Information System (INIS)

    Hauer-Jensen, M.

    2003-01-01

    Proteinase-activated receptors (PARs) are G-protein coupled receptors that are activated by proteolytic exposure of a receptor-tethered ligand. The discovery of this receptor family represents one of the most intriguing recent developments in signal transduction. PARs are involved in the regulation of many normal and pathophysiological processes, notably inflammatory and fibroproliferative responses to injury. Preclinical studies performed in our laboratory suggest that proteinase-activated receptor-1 (PAR-1) plays a critical role in the mechanism of chronicity of radiation fibrosis, while proteinase-activated receptor-2 (PAR-2) may mediate important fibroproliferative responses in irradiated intestine. Specifically, activation of PAR-1 by thrombin, and PAR-2 by pancreatic trypsin and mast cell proteinases, appears to be involved in acute radiation-induced inflammation, as well as in subsequent extracellular matrix deposition, leading to the development of intestinal wall fibrosis and clinical complications. Pharmacological modulators of PAR-1 or PAR-2 expression or activation would be potentially useful as preventive or therapeutic agents in patients who receive radiation therapy, especially if blockade could be targeted to specific tissues or cellular compartments

  15. Xenobiotic Receptor-Mediated Regulation of Intestinal Barrier Function and Innate Immunity

    Directory of Open Access Journals (Sweden)

    Harmit S. Ranhotra

    2016-07-01

    Full Text Available The molecular basis for the regulation of the intestinal barrier is a very fertile research area. A growing body of knowledge supports the targeting of various components of intestinal barrier function as means to treat a variety of diseases, including the inflammatory bowel diseases. Herein, we will summarize the current state of knowledge of key xenobiotic receptor regulators of barrier function, highlighting recent advances, such that the field and its future are succinctly reviewed. We posit that these receptors confer an additional dimension of host-microbe interaction in the gut, by sensing and responding to metabolites released from the symbiotic microbiota, in innate immunity and also in host drug metabolism. The scientific evidence for involvement of the receptors and its molecular basis for the control of barrier function and innate immunity regulation would serve as a rationale towards development of non-toxic probes and ligands as drugs.

  16. Cholecystokinin-2 receptor mediated gene expression in neuronal PC12 cells

    DEFF Research Database (Denmark)

    Hansen, Thomas v O; Borup, Rehannah; Marstrand, Troels

    2007-01-01

    could be identified. Comparison with forskolin- and nerve growth factor (NGF)-treated PC12 cells showed that CCK induced a separate set of target genes. Taken together, we propose that neuronal CCK may have a role in the regulation of the circadian rhythm, the metabolism of cerebral cholesterol...... of neuronal CCK are incompletely understood. To identify genes regulated by neuronal CCK, we generated neuronal PC12 cells stably expressing the CCK-2 receptor (CCK-2R) and treated the cells with sulphated CCK-8 for 2-16 h, before the global expression profile was examined. The changes in gene expression...... peaked after 2 h, with 67 differentially expressed transcripts identified. A pathway analysis indicated that CCK was implicated in the regulation of the circadian clock system, the plasminogen system and cholesterol metabolism. But transcripts encoding proteins involved in dopamine signaling, ornithine...

  17. GABAA Receptor-Mediated Activity in a Model of Cortical Dysplasia

    Science.gov (United States)

    2012-06-29

    LiCl/pilocarpine- induced status epilepticus on brain mu and benzodiazepine receptor binding: regional and ontogenetic studies. Brain Res 1181:104-17...Z, Nadler V (2009) Enhanced tonic GABA current in normotopic and hilar ctopic dentate granule cells after pilocarpine-induced status epilepticus . J

  18. The Role of lysophosphatidic acide (LPA) in the insulin resistence of the pancreatic β-cells

    OpenAIRE

    Mourad Agha, Zein

    2016-01-01

    The pathogenesis of the type-2-diabetes mellitus underlying is characterized by a combination of peripheral insulin resistance, β-cell dysfunction and reduction in the β cell mass. The increasing of FFA level or their metabolites lead to inhibition of insulin signaling. Consequent, the ability of insulin is reduced and therefore lead to insulin resistance. LPA is a lipid mediator that is associated with a progression of T2D. It has been suggested that LPA and the development of obesity are st...

  19. The effects of S(-)-, R(+)-, and racemic bupivacaine on lysophosphatidate-induced priming of human neutrophils

    NARCIS (Netherlands)

    Hollmann, Markus W.; Kurz, Katrin; Herroeder, Susanne; Struemper, Danja; Hahnenkamp, Klaus; Berkelmans, Noud S.; den Bakker, Christel G.; Durieux, Marcel E.

    2003-01-01

    Local anesthetics modulate inflammatory responses and may therefore be potentially useful in mitigating perioperative inflammatory injury. The inflammatory modulating effects of S(-)-bupivacaine are not known. Therefore, we compared the effects of S(-)-bupivacaine, R(+)-bupivacaine, and racemic

  20. Lysophosphatidic acids are new substrates for the phosphatase domain of soluble epoxide hydrolase[S

    OpenAIRE

    Oguro, Ami; Imaoka, Susumu

    2012-01-01

    Soluble epoxide hydrolase (sEH) is a bifunctional enzyme that has a C-terminus epoxide hydrolase domain and an N-terminus phosphatase domain. The endogenous substrates of epoxide hydrolase are known to be epoxyeicosatrienoic acids, but the endogenous substrates of the phosphatase activity are not well understood. In this study, to explore the substrates of sEH, we investigated the inhibition of the phosphatase activity of sEH toward 4-methylumbelliferyl phosphate by using lecithin and its hyd...

  1. Identification of a dopamine receptor-mediated opiate reward memory switch in the basolateral amygdala-nucleus accumbens circuit.

    Science.gov (United States)

    Lintas, Alessandra; Chi, Ning; Lauzon, Nicole M; Bishop, Stephanie F; Gholizadeh, Shervin; Sun, Ninglei; Tan, Huibing; Laviolette, Steven R

    2011-08-03

    The basolateral amygdala (BLA), ventral tegmental area (VTA), and nucleus accumbens (NAc) play central roles in the processing of opiate-related associative reward learning and memory. The BLA receives innervation from dopaminergic fibers originating in the VTA, and both dopamine (DA) D1 and D2 receptors are expressed in this region. Using a combination of in vivo single-unit extracellular recording in the NAc combined with behavioral pharmacology studies, we have identified a double dissociation in the functional roles of DA D1 versus D2 receptor transmission in the BLA, which depends on opiate exposure state; thus, in previously opiate-naive rats, blockade of intra-BLA D1, but not D2, receptor transmission blocked the acquisition of associative opiate reward memory, measured in an unbiased conditioned place preference procedure. In direct contrast, in rats made opiate dependent and conditioned in a state of withdrawal, intra-BLA D2, but not D1, receptor blockade blocked opiate reward encoding. This functional switch was dependent on cAMP signaling as comodulation of intra-BLA cAMP levels reversed or replicated the functional effects of intra-BLA D1 or D2 transmission during opiate reward processing. Single-unit in vivo extracellular recordings performed in neurons of the NAc confirmed an opiate-state-dependent role for BLA D1/D2 transmission in NAc neuronal response patterns to morphine. Our results characterize and identify a novel opiate addiction switching mechanism directly in the BLA that can control the processing of opiate reward information as a direct function of opiate exposure state via D1 or D2 receptor signaling substrates.

  2. Solid lipid nanoparticles carrying chemotherapeutic drug across the blood-brain barrier through insulin receptor-mediated pathway.

    Science.gov (United States)

    Kuo, Yung-Chih; Shih-Huang, Chun-Yuan

    2013-09-01

    Carmustine (BCNU)-loaded solid lipid nanoparticles (SLNs) were grafted with 83-14 monoclonal antibody (MAb) (83-14 MAb/BCNU-SLNs) and applied to the brain-targeting delivery. Human brain-microvascular endothelial cells (HBMECs) incubated with 83-14 MAb/BCNU-SLNs were stained to demonstrate the interaction between the nanocarriers and expressed insulin receptors (IRs). The results revealed that the particle size of 83-14 MAb/BCNU-SLNs decreased with an increasing weight percentage of Dynasan 114 (DYN). Storage at 4 °C for 6 weeks slightly deformed the colloidal morphology. In addition, poloxamer 407 on 83-14 MAb/BCNU-SLNs induced cytotoxicity to RAW264.7 cells and inhibited phagocytosis by RAW264.7 cells. An increase in the weight percentage of DYN from 0% to 67% slightly reduced the viability of RAW264.7 cells and promoted phagocytosis. Moreover, the transport ability of 83-14 MAb/BCNU-SLNs across the blood-brain barrier (BBB) in vitro enhanced with an increasing weight percentage of Tween 80. 83-14 MAb on MAb/BCNU-SLNs stimulated endocytosis by HBMECs via IRs and enhanced the permeability of BCNU across the BBB. 83-14 MAb/BCNU-SLNs can be a promising antitumor drug delivery system for transporting BCNU to the brain.

  3. Turn a diarrhoea toxin into a receptor-mediated therapy for a plethora of CLDN-4-overexpressing cancers

    International Nuclear Information System (INIS)

    Yao, Qin; Cao, Siyu; Li, Chun; Mengesha, Asferd; Low, Pauline; Kong, Beihua; Dai, Shuzhen; Wei, Mingqian

    2010-01-01

    Research highlights: → CLDN-4 is the high-affinity receptor for Clostridium perfringens enterotoxin (CPE). → The targeted toxin C-CPE-ETA' utilises the C-terminal fragment of CPE for binding. → C-CPE-ETA' rapidly binds to and internalises into CLDN-4 positive cancer cells. → C-CPE-ETA' has anti-cancer ability in a range of CLDN-4 positive cancers. -- Abstract: Molecular targeted therapy (MTT) represents the new generation of anti-cancer arsenals. In this study, we report an alternative approach using a hybrid toxin that utilises the high-affinity of receptor-binding fragment of Clostridium perfringens enterotoxin (CPE). CPE naturally binds to CLDN-4 through the C-terminal 30 amino acid. However, recent studies have shown that CLDN-4 is also overexpressed on a range of cancer cells. We thus constructed a cDNA comprising C-CPE and a well characterised toxic domain of Pseudomonas aeruginosa exotoxin A (C-CPE-ETA'). The recombinant C-CPE-ETA' fusion protein was shown to retain the specificity of binding to CLDN-4 and initiating rapid penetration into cytosol in five different CLDN-4 positive cancer cells (Breast-MCF7, Skin-A431, Colon-SW480, Prostate-PC3 and DU145) but not to CLDN-4 negative cells (Hela, HUVEC). C-CPE-ETA' was strongly cytotoxic towards CLDN-4 positive cancer cell, as opposed to cells lacking CLDN-4 expression. Furthermore, we demonstrated that the recombinant fusion protein had significant anti-cancer ability in CLDN-4 positive cancer models in vivo. Subcutaneously implanted MCF7 and SW480 xenograft tumours were significantly decreased or abolished after three repeated injection of the hybrid toxin. Taken together, our results convincingly show that the hybrid toxin targets CLDN-4 positive cancer through receptor-binding, and causes significant tumour cell apoptosis, suggesting its potential as an alternative molecular targeted therapy against a plethora of CLDN-4 positive cancers.

  4. Cannabinoid receptor-mediated disruption of sensory gating and neural oscillations: A translational study in rats and humans.

    Science.gov (United States)

    Skosnik, Patrick D; Hajós, Mihály; Cortes-Briones, Jose A; Edwards, Chad R; Pittman, Brian P; Hoffmann, William E; Sewell, Andrew R; D'Souza, Deepak C; Ranganathan, Mohini

    2018-06-01

    Cannabis use has been associated with altered sensory gating and neural oscillations. However, it is unclear which constituent in cannabis is responsible for these effects, or whether these are cannabinoid receptor 1 (CB1R) mediated. Therefore, the present study in humans and rats examined whether cannabinoid administration would disrupt sensory gating and evoked oscillations utilizing electroencephalography (EEG) and local field potentials (LFPs), respectively. Human subjects (n = 15) completed four test days during which they received intravenous delta-9-tetrahydrocannabinol (Δ 9 -THC), cannabidiol (CBD), Δ 9 -THC + CBD, or placebo. Subjects engaged in a dual-click paradigm, and outcome measures included P50 gating ratio (S2/S1) and evoked power to S1 and S2. In order to examine CB1R specificity, rats (n = 6) were administered the CB1R agonist CP-55940, CP-55940+AM-251 (a CB1R antagonist), or vehicle using the same paradigm. LFPs were recorded from CA3 and entorhinal cortex. Both Δ 9 -THC (p < 0.007) and Δ 9 -THC + CBD (p < 0.004) disrupted P50 gating ratio compared to placebo, while CBD alone had no effect. Δ 9 -THC (p < 0.048) and Δ 9 -THC + CBD (p < 0.035) decreased S1 evoked theta power, and in the Δ 9 -THC condition, S1 theta negatively correlated with gating ratios (r = -0.629, p < 0.012 (p < 0.048 adjusted)). In rats, CP-55940 disrupted gating in both brain regions (p < 0.0001), and this was reversed by AM-251. Further, CP-55940 decreased evoked theta (p < 0.0077) and gamma (p < 0.011) power to S1, which was partially blocked by AM-251. These convergent human/animal data suggest that CB1R agonists disrupt sensory gating by altering neural oscillations in the theta-band. Moreover, this suggests that the endocannabinoid system mediates theta oscillations relevant to perception and cognition. Copyright © 2018 Elsevier Ltd. All rights reserved.

  5. LncRNA HOTAIR Enhances the Androgen-Receptor-Mediated Transcriptional Program and Drives Castration-Resistant Prostate Cancer

    Directory of Open Access Journals (Sweden)

    Ali Zhang

    2015-10-01

    Full Text Available Understanding the mechanisms of androgen receptor (AR activation in the milieu of low androgen is critical to effective treatment of castration-resistant prostate cancer (CRPC. Here, we report HOTAIR as an androgen-repressed lncRNA, and, as such, it is markedly upregulated following androgen deprivation therapies and in CRPC. We further demonstrate a distinct mode of lncRNA-mediated gene regulation, wherein HOTAIR binds to the AR protein to block its interaction with the E3 ubiquitin ligase MDM2, thereby preventing AR ubiquitination and protein degradation. Consequently, HOTAIR expression is sufficient to induce androgen-independent AR activation and drive the AR-mediated transcriptional program in the absence of androgen. Functionally, HOTAIR overexpression increases, whereas HOTAIR knockdown decreases, prostate cancer cell growth and invasion. Taken together, our results provide compelling evidence of lncRNAs as drivers of androgen-independent AR activity and CRPC progression, and they support the potential of lncRNAs as therapeutic targets.

  6. Identification of putative estrogen receptor-mediated endocrine disrupting chemicals using QSAR- and structure-based virtual screening approaches

    Energy Technology Data Exchange (ETDEWEB)

    Zhang, Liying; Sedykh, Alexander; Tripathi, Ashutosh [Eshelman School of Pharmacy, University of North Carolina, Chapel Hill, NC (United States); Zhu, Hao [The Rutgers Center for Computational and Integrative Biology, Rutgers University, Camden, NJ (United States); Department of Chemistry, Rutgers University, Camden, NJ (United States); Afantitis, Antreas; Mouchlis, Varnavas D.; Melagraki, Georgia [NovaMechanics Ltd., Nicosia (Cyprus); Rusyn, Ivan, E-mail: iir@unc.edu [Department of Environmental Sciences and Engineering, University of North Carolina, Chapel Hill, NC (United States); Tropsha, Alexander, E-mail: alex_tropsha@unc.edu [Eshelman School of Pharmacy, University of North Carolina, Chapel Hill, NC (United States)

    2013-10-01

    Identification of endocrine disrupting chemicals is one of the important goals of environmental chemical hazard screening. We report on the development of validated in silico predictors of chemicals likely to cause estrogen receptor (ER)-mediated endocrine disruption to facilitate their prioritization for future screening. A database of relative binding affinity of a large number of ERα and/or ERβ ligands was assembled (546 for ERα and 137 for ERβ). Both single-task learning (STL) and multi-task learning (MTL) continuous quantitative structure–activity relationship (QSAR) models were developed for predicting ligand binding affinity to ERα or ERβ. High predictive accuracy was achieved for ERα binding affinity (MTL R{sup 2} = 0.71, STL R{sup 2} = 0.73). For ERβ binding affinity, MTL models were significantly more predictive (R{sup 2} = 0.53, p < 0.05) than STL models. In addition, docking studies were performed on a set of ER agonists/antagonists (67 agonists and 39 antagonists for ERα, 48 agonists and 32 antagonists for ERβ, supplemented by putative decoys/non-binders) using the following ER structures (in complexes with respective ligands) retrieved from the Protein Data Bank: ERα agonist (PDB ID: 1L2I), ERα antagonist (PDB ID: 3DT3), ERβ agonist (PDB ID: 2NV7), and ERβ antagonist (PDB ID: 1L2J). We found that all four ER conformations discriminated their corresponding ligands from presumed non-binders. Finally, both QSAR models and ER structures were employed in parallel to virtually screen several large libraries of environmental chemicals to derive a ligand- and structure-based prioritized list of putative estrogenic compounds to be used for in vitro and in vivo experimental validation. - Highlights: • This is the largest curated dataset inclusive of ERα and β (the latter is unique). • New methodology that for the first time affords acceptable ERβ models. • A combination of QSAR and docking enables prediction of affinity and function. • The results have potential applications to green chemistry. • Models are publicly available for virtual screening via a web portal.

  7. Olfactory bulb glomerular NMDA receptors mediate olfactory nerve potentiation and odor preference learning in the neonate rat.

    Directory of Open Access Journals (Sweden)

    Rebecca Lethbridge

    Full Text Available Rat pup odor preference learning follows pairing of bulbar beta-adrenoceptor activation with olfactory input. We hypothesize that NMDA receptor (NMDAR-mediated olfactory input to mitral cells is enhanced during training, such that increased calcium facilitates and shapes the critical cAMP pattern. Here, we demonstrate, in vitro, that olfactory nerve stimulation, at sniffing frequencies, paired with beta-adrenoceptor activation, potentiates olfactory nerve-evoked mitral cell firing. This potentiation is blocked by a NMDAR antagonist and by increased inhibition. Glomerular disinhibition also induces NMDAR-sensitive potentiation. In vivo, in parallel, behavioral learning is prevented by glomerular infusion of an NMDAR antagonist or a GABA(A receptor agonist. A glomerular GABA(A receptor antagonist paired with odor can induce NMDAR-dependent learning. The NMDA GluN1 subunit is phosphorylated in odor-specific glomeruli within 5 min of training suggesting early activation, and enhanced calcium entry, during acquisition. The GluN1 subunit is down-regulated 3 h after learning; and at 24 h post-training the GluN2B subunit is down-regulated. These events may assist memory stability. Ex vivo experiments using bulbs from trained rat pups reveal an increase in the AMPA/NMDA EPSC ratio post-training, consistent with an increase in AMPA receptor insertion and/or the decrease in NMDAR subunits. These results support a model of a cAMP/NMDA interaction in generating rat pup odor preference learning.

  8. α7 Nicotinic receptor-mediated astrocytic gliotransmitter release: Aβ effects in a preclinical Alzheimer's mouse model.

    Directory of Open Access Journals (Sweden)

    Tiina Maria Pirttimaki

    Full Text Available It is now recognized that astrocytes participate in synaptic communication through intimate interactions with neurons. A principal mechanism is through the release of gliotransmitters (GTs such as ATP, D-serine and most notably, glutamate, in response to astrocytic calcium elevations. We and others have shown that amyloid-β (Aβ, the toxic trigger for Alzheimer's disease (AD, interacts with hippocampal α7 nicotinic acetylcholine receptors (nAChRs. Since α7nAChRs are highly permeable to calcium and are expressed on hippocampal astrocytes, we investigated whether Aβ could activate astrocytic α7nAChRs in hippocampal slices and induce GT glutamate release. We found that biologically-relevant concentrations of Aβ1-42 elicited α7nAChR-dependent calcium elevations in hippocampal CA1 astrocytes and induced NMDAR-mediated slow inward currents (SICs in CA1 neurons. In the Tg2576 AD mouse model for Aβ over-production and accumulation, we found that spontaneous astrocytic calcium elevations were of higher frequency compared to wildtype (WT. The frequency and kinetic parameters of AD mice SICs indicated enhanced gliotransmission, possibly due to increased endogenous Aβ observed in this model. Activation of α7nAChRs on WT astrocytes increased spontaneous inward currents on pyramidal neurons while α7nAChRs on astrocytes of AD mice were abrogated. These findings suggest that, at an age that far precedes the emergence of cognitive deficits and plaque deposition, this mouse model for AD-like amyloidosis exhibits augmented astrocytic activity and glutamate GT release suggesting possible repercussions for preclinical AD hippocampal neural networks that contribute to subsequent cognitive decline.

  9. Gamma-hydroxybutyric acid (GHB) and the mesoaccumbens reward circuit: evidence for GABA(B) receptor-mediated effects.

    Science.gov (United States)

    Pistis, M; Muntoni, A L; Pillolla, G; Perra, S; Cignarella, G; Melis, M; Gessa, G L

    2005-01-01

    Gamma-hydroxybutyric acid (GHB) is a short-chain fatty acid naturally occurring in the mammalian brain, which recently emerged as a major recreational drug of abuse. GHB has multiple neuronal mechanisms including activation of both the GABA(B) receptor, and a distinct GHB-specific receptor. This complex GHB-GABA(B) receptor interaction is probably responsible for the multifaceted pharmacological, behavioral and toxicological profile of GHB. Drugs of abuse exert remarkably similar effects upon reward-related circuits, in particular the mesolimbic dopaminergic system and the nucleus accumbens (NAc). We used single unit recordings in vivo from urethane-anesthetized rats to characterize the effects of GHB on evoked firing in NAc "shell" neurons and on spontaneous activity of antidromically identified dopamine (DA) cells located in the ventral tegmental area. GHB was studied in comparison with the GABA(B) receptor agonist baclofen and antagonist (2S)(+)-5,5-dimethyl-2-morpholineacetic acid (SCH50911). Additionally, we utilized a GHB analog, gamma-(p-methoxybenzil)-gamma-hydroxybutyric acid (NCS-435), devoid of GABA(B) binding properties, but with high affinity for specific GHB binding sites. In common with other drugs of abuse, GHB depressed firing in NAc neurons evoked by the stimulation of the basolateral amygdala. On DA neurons, GHB exerted heterogeneous effects, which were correlated to the baseline firing rate of the cells but led to a moderate stimulation of the DA system. All GHB actions were mediated by GABA(B) receptors, since they were blocked by SCH50911 and were not mimicked by NCS-435. Our study indicates that the electrophysiological profile of GHB is close to typical drugs of abuse: both inhibition of NAc neurons and moderate to strong stimulation of DA transmission are distinctive features of diverse classes of abused drugs. Moreover, it is concluded that addictive and rewarding properties of GHB do not necessarily involve a putative high affinity GHB receptor.

  10. Benzo[a]pyrene induces intercellular adhesion molecule-1 through a caveolae and aryl hydrocarbon receptor mediated pathway

    International Nuclear Information System (INIS)

    Oesterling, Elizabeth; Toborek, Michal; Hennig, Bernhard

    2008-01-01

    Toxicologic and epidemiologic studies have linked benzo[a]pyrene (B[a]P) exposure with cardiovascular diseases such as atherosclerosis. The mechanisms of action leading to these diseases have not been fully understood. One key step in the development of atherosclerosis is vascular endothelial dysfunction, which is characterized by increased adhesiveness. To determine if B[a]P could lead to increased endothelial adhesiveness, the effects of B[a]P on human endothelial cell intercellular adhesion molecule-1 (ICAM-1) expression was investigated. B[a]P was able to increase ICAM-1 protein only after pretreatment with the aryl hydrocarbon receptor (AhR) agonist β-naphthoflavone (β-NF). Knockdown of AhR by siRNA or treatment with AhR antagonist α-naphthoflavone (α-NF) eliminated the induction of ICAM-1 from B[a]P, confirming the necessity of AhR in this process. Likewise, B[a]P only increased monocyte adhesion to the vascular endothelium when cells were pretreated with β-NF. Experiments were done to define a signaling mechanism. B[a]P increased phosphorylation of MEK and p38-MAPK, and inhibitors to these proteins blunted the ICAM-1 induction. B[a]P was also able to increase AP-1 DNA binding and phosphorylation of cJun. Phosphorylation of cJun was disrupted by MEK and p38-MAPK inhibitors linking the signaling cascade. Finally, the importance of membrane microdomains, caveolae, was demonstrated by knockdown of the structural protein caveolin-1. Disruption of caveolae eliminated the B[a]P-induced ICAM-1 expression. These data suggest a possible pro-inflammatory mechanism of action of B[a]P involving caveolae, leading to increased vascular endothelial adhesiveness, and this inflammation may be a critical step in the development of B[a]P-induced atherosclerosis

  11. Identification of putative estrogen receptor-mediated endocrine disrupting chemicals using QSAR- and structure-based virtual screening approaches

    International Nuclear Information System (INIS)

    Zhang, Liying; Sedykh, Alexander; Tripathi, Ashutosh; Zhu, Hao; Afantitis, Antreas; Mouchlis, Varnavas D.; Melagraki, Georgia; Rusyn, Ivan; Tropsha, Alexander

    2013-01-01

    Identification of endocrine disrupting chemicals is one of the important goals of environmental chemical hazard screening. We report on the development of validated in silico predictors of chemicals likely to cause estrogen receptor (ER)-mediated endocrine disruption to facilitate their prioritization for future screening. A database of relative binding affinity of a large number of ERα and/or ERβ ligands was assembled (546 for ERα and 137 for ERβ). Both single-task learning (STL) and multi-task learning (MTL) continuous quantitative structure–activity relationship (QSAR) models were developed for predicting ligand binding affinity to ERα or ERβ. High predictive accuracy was achieved for ERα binding affinity (MTL R 2 = 0.71, STL R 2 = 0.73). For ERβ binding affinity, MTL models were significantly more predictive (R 2 = 0.53, p < 0.05) than STL models. In addition, docking studies were performed on a set of ER agonists/antagonists (67 agonists and 39 antagonists for ERα, 48 agonists and 32 antagonists for ERβ, supplemented by putative decoys/non-binders) using the following ER structures (in complexes with respective ligands) retrieved from the Protein Data Bank: ERα agonist (PDB ID: 1L2I), ERα antagonist (PDB ID: 3DT3), ERβ agonist (PDB ID: 2NV7), and ERβ antagonist (PDB ID: 1L2J). We found that all four ER conformations discriminated their corresponding ligands from presumed non-binders. Finally, both QSAR models and ER structures were employed in parallel to virtually screen several large libraries of environmental chemicals to derive a ligand- and structure-based prioritized list of putative estrogenic compounds to be used for in vitro and in vivo experimental validation. - Highlights: • This is the largest curated dataset inclusive of ERα and β (the latter is unique). • New methodology that for the first time affords acceptable ERβ models. • A combination of QSAR and docking enables prediction of affinity and function. • The results have potential applications to green chemistry. • Models are publicly available for virtual screening via a web portal

  12. P2X7 receptor-mediated calcium dynamics in HEK293 cells: experimental characterization and modelling approach

    International Nuclear Information System (INIS)

    Di Garbo, A; Alloisio, S; Nobile, M

    2012-01-01

    The P2X7 receptor (P2X7R) induces ionotropic Ca 2+  signalling in different cell types. It plays an important role in the immune response and in the nervous system. Here, the mechanisms underlying intracellular Ca 2+  variations evoked by 3′-O-(4-benzoyl)benzoyl-ATP (BzATP), a potent agonist of the P2X7R, in transfected HEK293 cells, are investigated both experimentally and theoretically. We propose a minimal model of P2X7R that is capable of reproducing, qualitatively and quantitatively, the experimental data. This approach was also adopted for the P2X7R variant, which lacks the entire C-terminus tail (trP2X7R). Then we introduce a biophysical model describing the Ca 2+  dynamics in HEK293. Our model gives an account of the ionotropic Ca 2+  influx evoked by BzATP on the basis of the kinetics model of P2X7R. To explain the complex Ca 2+  responses evoked by BzATP, the model predicted that an impairment in Ca 2+  extrusion flux through the plasma membrane is a key factor for Ca 2+ homeostasis in HEK293 cells. (paper)

  13. P2X7 receptor-mediated calcium dynamics in HEK293 cells: experimental characterization and modelling approach

    Science.gov (United States)

    Di Garbo, A.; Alloisio, S.; Nobile, M.

    2012-04-01

    The P2X7 receptor (P2X7R) induces ionotropic Ca2 + signalling in different cell types. It plays an important role in the immune response and in the nervous system. Here, the mechanisms underlying intracellular Ca2 + variations evoked by 3‧-O-(4-benzoyl)benzoyl-ATP (BzATP), a potent agonist of the P2X7R, in transfected HEK293 cells, are investigated both experimentally and theoretically. We propose a minimal model of P2X7R that is capable of reproducing, qualitatively and quantitatively, the experimental data. This approach was also adopted for the P2X7R variant, which lacks the entire C-terminus tail (trP2X7R). Then we introduce a biophysical model describing the Ca2 + dynamics in HEK293. Our model gives an account of the ionotropic Ca2 + influx evoked by BzATP on the basis of the kinetics model of P2X7R. To explain the complex Ca2 + responses evoked by BzATP, the model predicted that an impairment in Ca2 + extrusion flux through the plasma membrane is a key factor for Ca2 + homeostasis in HEK293 cells.

  14. p85α recruitment by the CD300f phosphatidylserine receptor mediates apoptotic cell clearance required for autoimmunity suppression

    Science.gov (United States)

    Tian, Linjie; Choi, Seung-Chul; Murakami, Yousuke; Allen, Joselyn; Morse, Herbert C., III; Qi, Chen-Feng; Krzewski, Konrad; Coligan, John E.

    2014-01-01

    Apoptotic cell (AC) clearance is essential for immune homeostasis. Here we show that mouse CD300f (CLM-1) recognizes outer membrane-exposed phosphatidylserine, and regulates the phagocytosis of ACs. CD300f accumulates in phagocytic cups at AC contact sites. Phosphorylation within CD300f cytoplasmic tail tyrosine-based motifs initiates signals that positively or negatively regulate AC phagocytosis. Y276 phosphorylation is necessary for enhanced CD300f-mediated phagocytosis through the recruitment of the p85α regulatory subunit of phosphatidylinositol-3-kinase (PI3K). CD300f-PI3K association leads to activation of downstream Rac/Cdc42 GTPase and mediates changes of F-actin that drive AC engulfment. Importantly, primary macrophages from CD300f-deficient mice have impaired phagocytosis of ACs. The biological consequence of CD300f deficiency is predisposition to autoimmune disease development, as FcγRIIB-deficient mice develop a systemic lupus erythematosus-like disease at a markedly accelerated rate if CD300f is absent. In this report we identify the mechanism and role of CD300f in AC phagocytosis and maintenance of immune homeostasis.

  15. Molecular Mechanisms Underlying β-Adrenergic Receptor-Mediated Cross-Talk between Sympathetic Neurons and Immune Cells

    Directory of Open Access Journals (Sweden)

    Dianne Lorton

    2015-03-01

    Full Text Available Cross-talk between the sympathetic nervous system (SNS and immune system is vital for health and well-being. Infection, tissue injury and inflammation raise firing rates of sympathetic nerves, increasing their release of norepinephrine (NE in lymphoid organs and tissues. NE stimulation of β2-adrenergic receptors (ARs in immune cells activates the cAMP-protein kinase A (PKA intracellular signaling pathway, a pathway that interfaces with other signaling pathways that regulate proliferation, differentiation, maturation and effector functions in immune cells. Immune–SNS cross-talk is required to maintain homeostasis under normal conditions, to develop an immune response of appropriate magnitude after injury or immune challenge, and subsequently restore homeostasis. Typically, β2-AR-induced cAMP is immunosuppressive. However, many studies report actions of β2-AR stimulation in immune cells that are inconsistent with typical cAMP–PKA signal transduction. Research during the last decade in non-immune organs, has unveiled novel alternative signaling mechanisms induced by β2-AR activation, such as a signaling switch from cAMP–PKA to mitogen-activated protein kinase (MAPK pathways. If alternative signaling occurs in immune cells, it may explain inconsistent findings of sympathetic regulation of immune function. Here, we review β2-AR signaling, assess the available evidence for alternative signaling in immune cells, and provide insight into the circumstances necessary for “signal switching” in immune cells.

  16. Androgen Receptor-Mediated Growth Suppression of HPr-1AR and PC3-Lenti-AR Prostate Epithelial Cells.

    Directory of Open Access Journals (Sweden)

    Young-Chae Kim

    Full Text Available The androgen receptor (AR mediates the developmental, physiologic, and pathologic effects of androgens including 5α-dihydrotestosterone (DHT. However, the mechanisms whereby AR regulates growth suppression and differentiation of luminal epithelial cells in the prostate gland and proliferation of malignant versions of these cells are not well understood, though they are central to prostate development, homeostasis, and neoplasia. Here, we identify androgen-responsive genes that restrain cell cycle progression and proliferation of human prostate epithelial cell lines (HPr-1AR and PC3-Lenti-AR, and we investigate the mechanisms through which AR regulates their expression. DHT inhibited proliferation of HPr-1AR and PC3-Lenti-AR, and cell cycle analysis revealed a prolonged G1 interval. In the cell cycle, the G1/S-phase transition is initiated by the activity of cyclin D and cyclin-dependent kinase (CDK complexes, which relieve growth suppression. In HPr-1AR, cyclin D1/2 and CDK4/6 mRNAs were androgen-repressed, whereas CDK inhibitor, CDKN1A, mRNA was androgen-induced. The regulation of these transcripts was AR-dependent, and involved multiple mechanisms. Similar AR-mediated down-regulation of CDK4/6 mRNAs and up-regulation of CDKN1A mRNA occurred in PC3-Lenti-AR. Further, CDK4/6 overexpression suppressed DHT-inhibited cell cycle progression and proliferation of HPr-1AR and PC3-Lenti-AR, whereas CDKN1A overexpression induced cell cycle arrest. We therefore propose that AR-mediated growth suppression of HPr-1AR involves cyclin D1 mRNA decay, transcriptional repression of cyclin D2 and CDK4/6, and transcriptional activation of CDKN1A, which serve to decrease CDK4/6 activity. AR-mediated inhibition of PC3-Lenti-AR proliferation occurs through a similar mechanism, albeit without down-regulation of cyclin D. Our findings provide insight into AR-mediated regulation of prostate epithelial cell proliferation.

  17. High-affinity α4β2 nicotinic receptors mediate the impairing effects of acute nicotine on contextual fear extinction.

    Science.gov (United States)

    Kutlu, Munir Gunes; Holliday, Erica; Gould, Thomas J

    2016-02-01

    Previously, studies from our lab have shown that while acute nicotine administered prior to training and testing enhances contextual fear conditioning, acute nicotine injections prior to extinction sessions impair extinction of contextual fear. Although there is also strong evidence showing that the acute nicotine's enhancing effects on contextual fear conditioning require high-affinity α4β2 nicotinic acetylcholine receptors (nAChRs), it is unknown which nAChR subtypes are involved in the acute nicotine-induced impairment of contextual fear extinction. In this study, we investigated the effects of acute nicotine administration on contextual fear extinction in knock-out (KO) mice lacking α4, β2 or α7 subtypes of nAChRs and their wild-type (WT) littermates. Both KO and WT mice were first trained and tested for contextual fear conditioning and received a daily contextual extinction session for 4 days. Subjects received intraperitoneal injections of nicotine (0.18 mg/kg) or saline 2-4 min prior to each extinction session. Our results showed that the mice that lack α4 and β2 subtypes of nAChRs showed normal contextual fear extinction but not the acute nicotine-induced impairment while the mice that lack the α7 subtype showed both normal contextual extinction and nicotine-induced impairment of contextual extinction. In addition, control experiments showed that acute nicotine-induced impairment of contextual fear extinction persisted when nicotine administration was ceased and repeated acute nicotine administrations alone did not induce freezing behavior in the absence of context-shock learning. These results clearly demonstrate that high-affinity α4β2 nAChRs are necessary for the effects of acute nicotine on contextual fear extinction. Copyright © 2015 Elsevier Inc. All rights reserved.

  18. Infralimbic dopamine D2 receptors mediate glucocorticoid-induced facilitation of auditory fear memory extinction in rats.

    Science.gov (United States)

    Dadkhah, Masoumeh; Abdullahi, Payman Raise; Rashidy-Pour, Ali; Sameni, Hamid Reza; Vafaei, Abbas Ali

    2018-03-01

    The infralimbic (IL) cortex of the medial prefrontal cortex plays an important role in the extinction of fear memory. Also, it has been showed that both brain glucocorticoid and dopamine receptors are involved in many processes such as fear extinction that drive learning and memory; however, the interaction of these receptors in the IL cortex remains unclear. We examined a putative interaction between the effects of glucocorticoid and dopamine receptors stimulation in the IL cortex on fear memory extinction in an auditory fear conditioning paradigm in male rats. Corticosterone (the endogenous glucocorticoid receptor ligand), or RU38486 (the synthetic glucocorticoid receptor antagonist) microinfusion into the IL cortex 10 min before test 1 attenuated auditory fear expression at tests 1-3, suggesting as an enhancement of fear extinction. The effect of corticosterone, but not RU38486 was counteracted by the dopamine D2 receptor antagonist sulpiride pre-treatment administered into the IL (at a dose that failed to alter freezing behavior on its own). In contrast, intra-IL infusion of the dopamine D1 receptor antagonist SCH23390 pre-treatment failed to alter freezing behavior. These findings provide evidence for the involvement of the IL cortex D2 receptors in CORT-induced facilitation of fear memory extinction. Copyright © 2018 Elsevier B.V. All rights reserved.

  19. CDK11{sup p58} represses vitamin D receptor-mediated transcriptional activation through promoting its ubiquitin-proteasome degradation

    Energy Technology Data Exchange (ETDEWEB)

    Chi, Yayun; Hong, Yi; Zong, Hongliang; Wang, Yanlin; Zou, Weiying; Yang, Junwu; Kong, Xiangfei; Yun, Xiaojing [Gene Research Center, Shanghai Medical College and Institutes of Biomedical, Shanghai 200032 (China); Gu, Jianxin, E-mail: jxgu@shmu.edu.cn [Gene Research Center, Shanghai Medical College and Institutes of Biomedical, Shanghai 200032 (China)

    2009-08-28

    Vitamin D receptor (VDR) is a member of the nuclear receptor superfamily and regulates transcription of target genes. In this study, we identified CDK11{sup p58} as a novel protein involved in the regulation of VDR. CDK11{sup p58}, a member of the large family of p34cdc2-related kinases, is associated with cell cycle progression, tumorigenesis, and apoptotic signaling. Our study demonstrated that CDK11{sup p58} interacted with VDR and repressed VDR-dependent transcriptional activation. Furthermore, overexpression of CDK11{sup p58} decreased the stability of VDR through promoting its ubiquitin-proteasome-mediated degradation. Taken together, these results suggest that CDK11{sup p58} is involved in the negative regulation of VDR.

  20. CDK11p58 represses vitamin D receptor-mediated transcriptional activation through promoting its ubiquitin-proteasome degradation

    International Nuclear Information System (INIS)

    Chi, Yayun; Hong, Yi; Zong, Hongliang; Wang, Yanlin; Zou, Weiying; Yang, Junwu; Kong, Xiangfei; Yun, Xiaojing; Gu, Jianxin

    2009-01-01

    Vitamin D receptor (VDR) is a member of the nuclear receptor superfamily and regulates transcription of target genes. In this study, we identified CDK11 p58 as a novel protein involved in the regulation of VDR. CDK11 p58 , a member of the large family of p34cdc2-related kinases, is associated with cell cycle progression, tumorigenesis, and apoptotic signaling. Our study demonstrated that CDK11 p58 interacted with VDR and repressed VDR-dependent transcriptional activation. Furthermore, overexpression of CDK11 p58 decreased the stability of VDR through promoting its ubiquitin-proteasome-mediated degradation. Taken together, these results suggest that CDK11 p58 is involved in the negative regulation of VDR.

  1. Structure of unliganded HSV gD reveals a mechanism for receptor-mediated activation of virus entry

    Energy Technology Data Exchange (ETDEWEB)

    Krummenacher, Claude; Supekar, Vinit M.; Whitbeck, J. Charles; Lazear, Eric; Connolly, Sarah A.; Eisenberg, Roselyn J.; Cohen, Gary H.; Wiley, Don C.; Carfi, Andrea (UPENN); (IRBM); (CHLMM)

    2010-07-19

    Herpes simplex virus (HSV) entry into cells requires binding of the envelope glycoprotein D (gD) to one of several cell surface receptors. The 50 C-terminal residues of the gD ectodomain are essential for virus entry, but not for receptor binding. We have determined the structure of an unliganded gD molecule that includes these C-terminal residues. The structure reveals that the C-terminus is anchored near the N-terminal region and masks receptor-binding sites. Locking the C-terminus in the position observed in the crystals by an intramolecular disulfide bond abolished receptor binding and virus entry, demonstrating that this region of gD moves upon receptor binding. Similarly, a point mutant that would destabilize the C-terminus structure was nonfunctional for entry, despite increased affinity for receptors. We propose that a controlled displacement of the gD C-terminus upon receptor binding is an essential feature of HSV entry, ensuring the timely activation of membrane fusion.

  2. 5-HT4 receptors mediating enhancement of contractility in canine stomach; an in vitro and in vivo study

    NARCIS (Netherlands)

    Prins, NH; van der Grijn, A; Lefebvre, RA; Akkermans, LMA; Schuurkes, JAJ

    1 We aimed to study 5-HT4 receptors in canine stomach contractility both in vivo and in vitro. 2 In anaesthetized Beagle dogs, the selective 5-HT4 receptor agonist prucalopride (i.v.) induced dose-dependent tonic stomach contractions under isobaric conditions, an effect that was antagonized by the

  3. Metabolic Adaptation of Human CD4+ and CD8+ T-Cells to T-Cell Receptor-Mediated Stimulation

    Directory of Open Access Journals (Sweden)

    Nicholas Jones

    2017-11-01

    Full Text Available Linking immunometabolic adaptation to T-cell function provides insight for the development of new therapeutic approaches in multiple disease settings. T-cell activation and downstream effector functions of CD4+ and CD8+ T-cells are controlled by the strength of interaction between the T-cell receptor (TCR and peptides presented by human leukocyte antigens (pHLA. The role of TCR–pHLA interactions in modulating T-cell metabolism is unknown. Here, for the first time, we explore the relative contributions of the main metabolic pathways to functional responses in human CD4+ and CD8+ T-cells. Increased expression of hexokinase II accompanied by higher basal glycolysis is demonstrated in CD4+ T-cells; cytokine production in CD8+ T-cells is more reliant on oxidative phosphorylation. Using antigen-specific CD4+ and CD8+ T-cell clones and altered peptide ligands, we demonstrate that binding affinity tunes the underlying metabolic shift. Overall, this study provides important new insight into how metabolic pathways are controlled during antigen-specific activation of human T-cells.

  4. Stromal progesterone receptors mediate induction of Indian Hedgehog (IHH) in uterine epithelium and its downstream targets in uterine stroma.

    Science.gov (United States)

    Simon, Liz; Spiewak, Kerry A; Ekman, Gail C; Kim, Jaeyeon; Lydon, John P; Bagchi, Milan K; Bagchi, Indrani C; DeMayo, Francesco J; Cooke, Paul S

    2009-08-01

    Uterine receptivity to embryo implantation depends on appropriate progesterone (P4) and estrogen stimulation. P4 rapidly stimulates production of the morphogen Indian hedgehog (IHH) in murine uterine epithelium as well as downstream molecules in the hedgehog pathway such as Patched homolog 1 (PTCH1) and nuclear receptor subfamily 2, group F, member 2 (NR2F2) in uterine stroma. Studies using IHH-null mice indicate that IHH is obligatory for the normal P4 response in the uterus. To determine whether IHH induction in uterine epithelium is mediated through P4 receptor (PR) in epithelium (E) and/or stroma (S), we produced tissue recombinants using uteri from neonatal PR knockout (ko) mice and wild-type (wt) mice containing PR in S and/or E or lacking PR altogether using a tissue recombinant methodology and assessed their response to P4. In tissue recombinants containing wt-S (wt-S + wt-E and wt-S + ko-E), P4 induced Ihh mRNA expression at 6 h that was 6-fold greater than in oil-treated controls (P Ihh mRNA expression was unaffected by P4 in ko-S + ko-E and ko-S + wt-E grafts despite epithelial PR expression in the latter. Nr2f2 and Ptch1 mRNA expression was similar in that it was stimulated by P4 only in recombinants containing stromal PR. These results indicate that stromal PR is both necessary and sufficient for P4 stimulation of epithelial IHH as well as downstream events such as PTCH1 and NR2F2 increases in stroma.

  5. Receptor-mediated radionuclide therapy with 90Y-DOTATOC in association with amino acid infusion: a phase I study

    International Nuclear Information System (INIS)

    Bodei, Lisa; Zoboli, Stefania; Grana, Chiara; Bartolomei, Mirco; Rocca, Paola; Caracciolo, Maurizio; Chinol, Marco; Paganelli, Giovanni; Cremonesi, Marta; Maecke, Helmut R.

    2003-01-01

    The aim of this study was to determine the maximum tolerated dose of 90 Y-DOTATOC per cycle administered in association with amino acid solution as kidney protection in patients with somatostatin receptor-positive tumours. Forty patients in eight groups received two cycles of 90 Y-DOTATOC, with activity increased by 0.37 GBq per group, starting at 2.96 and terminating at 5.55 GBq. All patients received lysine ± arginine infusion immediately before and after therapy. Forty-eight percent developed acute grade I-II gastrointestinal toxicity (nausea and vomiting) after amino acid infusion whereas no acute adverse reactions occurred after 90 Y-DOTATOC injection up to 5.55 GBq/cycle. Grade III haematological toxicity occurred in three of seven (43%) patients receiving 5.18 GBq, which was defined as the maximum tolerable activity per cycle. Objective therapeutic responses occurred. Five GBq per cycle is the recommended dosage of 90 Y-DOTATOC when amino acids are given to protect the kidneys. Although no patients developed acute kidney toxicity, delayed kidney toxicity remains a major concern, limiting the cumulative dose to 25 Gy. The way forward with this treatment would seem to be to identify more effective renal protective agents, in order to be able to increase the cumulative injectable activity and hence tumour dose. (orig.)

  6. Hypothalamic kappa opioid receptor mediates both diet- and MCH-induced liver damage through inflammation and ER stress

    NARCIS (Netherlands)

    Imbernon, Monica; Sanchez-Rebordelo, Estrella; Romero-Picó, Amparo; Kalló, Imre; Chee, Melissa J; Porteiro, Begoña; Al-Massadi, Omar; Contreras, Cristina; Fernø, Johan; Senra, Ana; Gallego, Rosalia; Folgueira, Cintia; Seoane, Luisa M; van Gestel, Margriet; Adan, Roger A; Liposits, Zsolt; Dieguez, Carlos; Lopez, Miguel; Nogueiras, Ruben

    2016-01-01

    The opioid system is widely known to modulate the brain reward system and thus affect human and animal behaviour, including feeding. We hypothesized that the hypothalamic opioid system might also control energy metabolism in peripheral tissues. Mice lacking the kappa opioid receptor (κOR) and

  7. Glutamate AMPA/kainate receptors, not GABA(A) receptors, mediate estradiol-induced sex differences in the hypothalamus.

    Science.gov (United States)

    Todd, Brigitte J; Schwarz, Jaclyn M; Mong, Jessica A; McCarthy, Margaret M

    2007-02-15

    Sex differences in brain morphology underlie physiological and behavioral differences between males and females. During the critical perinatal period for sexual differentiation in the rat, gonadal steroids act in a regionally specific manner to alter neuronal morphology. Using Golgi-Cox impregnation, we examined several parameters of neuronal morphology in postnatal day 2 (PN2) rats. We found that in the ventromedial nucleus of the hypothalamus (VMN) and in areas just dorsal and just lateral to the VMN that there was a sex difference in total dendritic spine number (males greater) that was abolished by treating female neonates with exogenous testosterone. Dendritic branching was similarly sexually differentiated and hormonally modulated in the VMN and dorsal to the VMN. We then used spinophilin, a protein that positively correlates with the amount of dendritic spines, to investigate the mechanisms underlying these sex differences. Estradiol, which mediates most aspects of masculinization and is the aromatized product of testosterone, increased spinophilin levels in female PN2 rats to that of males. Muscimol, an agonist at GABA(A) receptors, did not affect spinophilin protein levels in either male or female neonates. Kainic acid, an agonist at glutamatergic AMPA/kainate receptors, mimicked the effect of estradiol in females. Antagonizing AMPA/kainate receptors with NBQX prevented the estradiol-induced increase in spinophilin in females but did not affect spinophilin level in males. (c) 2007 Wiley Periodicals, Inc.

  8. Estradiol receptors mediate estradiol-induced inhibition of mitochondrial Ca^{2+} efflux in rat caudate nucleus and brain stem

    OpenAIRE

    PETROVIC, SNJEZANA; MILOSEVIC, MAJA; RISTIC-MEDIC, DANIJELA; VELICKOVIC, NATASA; DRAKULIC, DUNJA; GRKOVIC, IVANA; HORVAT, ANICA

    2015-01-01

    Our earlier studies found that in vitro estradiol modulates mitochondrial Ca2+ transport in discrete brain regions. The present study examined the role of estradiol receptors (ERs) in estradiol-induced inhibition of Ca^{2+} efflux from synaptosomal mitochondria isolated from rat caudate nuclei and brain stems. Radioactively labeled CaCl_2 (0.6?0.75 µCi ^45CaCl_{2}) was used for Ca^{2+} transport monitoring. The results revealed that in the presence of ER antagonist 7\\alpha,17ß-[9[(4,4,5,5,5-...

  9. Glucocorticoid receptor, but not mineralocorticoid receptor, mediates cortisol regulation of epidermal ionocyte development and ion transport in zebrafish (danio rerio.

    Directory of Open Access Journals (Sweden)

    Shelly Abad Cruz

    Full Text Available Cortisol is the major endogenous glucocorticoid (GC both in human and fish, mediated by corticosteroid receptors. Due to the absence of aldosterone production in teleost fish, cortisol is also traditionally accepted to function as mineralocorticoid (MC; but whether it acts through the glucocorticoid receptor (GR or the mineralocorticoid receptor (MR remains a subject of debate. Here, we used loss-of-function and rescue assays to determine whether cortisol affects zebrafish epidermal ionocyte development and function via the GR and/or the MR. GR knockdown morphants displayed a significant decrease in the major ionocytes, namely Na(+-K(+-ATPase-rich cells (NaRCs and H(+-ATPase-rich cells (HRCs, as well as other cells, including epidermal stem cells (ESCs, keratinocytes, and mucus cells; conversely, cell numbers were unaffected in MR knockdown morphants. In agreement, GR morphants, but not MR morphants, exhibited decreased NaRC-mediated Ca(2+ uptake and HRC-mediated H(+ secretion. Rescue via GR capped mRNA injection or exogenous cortisol incubation normalized the number of epidermal ionocytes in GR morphants. We also provide evidence for GR localization in epidermal cells. At the transcript level, GR mRNA is ubiquitously expressed in gill sections and present in both NaRCs and HRCs, supporting the knockdown and functional assay results in embryo. Altogether, we have provided solid molecular evidence that GR is indeed present on ionocytes, where it mediates the effects of cortisol on ionocyte development and function. Hence, cortisol-GR axis performs the roles of both GC and MC in zebrafish skin and gills.

  10. Enhanced effect of dimension of receptor-ligand complex and depletion effect on receptor-mediated endocytosis of nanoparticles

    Science.gov (United States)

    Liu, Ye; Gao, Qingqing; Liu, Yijun; Zhao, Chuang; Mao, Zongliang; Hu, Lin; Liu, Yanhui

    2017-12-01

    Not Available Project supported by the National Natural Science Foundation of China (Grant Nos. 11047022, 11204045, and 11464004), the Tracking Key Program of Social Development of Guizhou Province, China (Grant Nos. SY20123089 and SZ20113069), the General Financial Grant from the China Postdoctoral Science Foundation (Grant No. 2014M562341), the Research Foundation for Young University Teachers from Guizhou University (Grant No. 201311), and the College Innovation Talent Team of Guizhou Province, China (Grant No. (2014)32).

  11. Receptor-mediated radiotherapy with Y-DOTA-DPhe-Tyr-octreotide: the experience of the European Institute of Oncology Group.

    Science.gov (United States)

    Chinol, Marco; Bodei, Lisa; Cremonesi, Marta; Paganelli, Giovanni

    2002-04-01

    High concentrations of subtype 2 somatostatin tumor receptors (sst(2)) are expressed in numerous tumors, enabling primary and metastatic masses to be localized by scintigraphy after injecting (111)In-labeled somatostatin analogue octreotide. In addition to neuroendocrine tumors, somatostatin receptors have been identified on cancers of the central nervous system, breast, lung, and lymphatic tissue, and the use of radionuclide-labeled somatostatin analogues appeared promising for therapy as well as for diagnosis of such malignancies. The somatostatin analogue [DOTA-(D)Phe(1)-Tyr(3)] octreotide (DOTATOC) possesses favorable characteristics for its potential therapeutic use in that it shows high affinity for sst(2), moderately high affinity for sst(5), and intermediate affinity for sst(3), high hydrophilicity, stable and facile labeling with (111)In and (90)Y. We began to investigate the potential therapeutic applications of (90)Y DOTATOC in 1997 by performing a thorough dosimetric study in 18 patients who were administered (111)In DOTATOC to estimate the absorbed doses during(90)Y-DOTATOC therapy. Then, we moved on and treated an overall number of 256 patients, mostly recruited in 2 distinct protocols with and without the administration of kidney protecting agents, with (90)Y DOTATOC. No major acute reactions were observed up to the activity of 5.55 GBq per cycle. The MTD per cycle was defined as 5.18 GBq. Objective therapeutic responses were documented in more than 20% of patients in terms of partial and complete responses. The present article reports in details our clinical experience (still ongoing) and outcomes with the use of (90)Y DOTATOC. Copyright 2002, Elsevier Science (USA). All rights reserved.

  12. Pitavastatin attenuates the PDGF-induced LR11/uPA receptor-mediated migration of smooth muscle cells

    International Nuclear Information System (INIS)

    Jiang, Meizi; Bujo, Hideaki; Zhu, Yanjuan; Yamazaki, Hiroyuki; Hirayama, Satoshi; Kanaki, Tatsuro; Shibasaki, Manabu; Takahashi, Kazuo; Schneider, Wolfgang J.; Saito, Yasushi

    2006-01-01

    Statins, inhibitors of HMG-CoA reductase, elicit various actions on vascular cells including the modulation of proliferation and migration of smooth muscle cells (SMCs). Here, we have elucidated the mechanism by which statins, in particular pitavastatin, attenuate the migration activity of SMCs. The expression of LR11, a member of the LDL receptor family and an enhancer of cell surface localization of urokinase-type plasminogen activator receptor (uPAR), is increased in cultured SMCs by treatment with PDGF-BB. Pitavastatin attenuates the PDGF-BB -induced surface expression of LR11 and uPAR. The increased migration of SMCs observed both upon overexpression of LR11 and via stimulation of secretion of soluble LR11 is not reversed by pitavastatin. In vivo studies showed that the SMCs expressing LR11 in plaques are almost congruent with intimal cells expressing nonmuscle myosin heavy chain (SMemb). Pitavastatin reduced the expression of LR11 and SMemb, and the levels of LR11, uPAR, and SMemb in cultured intimal SMCs were reduced to those seen in medial SMCs. We propose that this statin reduces PDGF-induced migration through the attenuation of the LR11/uPAR system in SMCs. Modulation of the LR11/uPAR system with statins suggests a novel treatment strategy for atherogenesis based on suppression of intimal SMC migration

  13. Protein replacement by receptor-mediated endocytosis corrects the sensitivity of Fanconi anemia group C cells to mitomycin C

    NARCIS (Netherlands)

    Youssoufian, H; Kruyt, FAE; Li, XT

    1999-01-01

    Current methods for direct gene transfer into hematopoietic cells are inefficient. Here we show that functional complementation of Fanconi anemia (FA) group C cells by protein replacement can be as efficacious as by transfection with wild-type FAC cDNA, We expressed a chimeric protein (called

  14. UBXN1 Interferes with Rig-I-like Receptor-Mediated Antiviral Immune Response by Targeting MAVS

    Directory of Open Access Journals (Sweden)

    Penghua Wang

    2013-04-01

    Full Text Available RNA viruses are sensed by RIG-I-like receptors (RLRs, which signal through a mitochondria-associated adaptor molecule, MAVS, resulting in systemic antiviral immune responses. Although RLR signaling is essential for limiting RNA virus replication, it must be stringently controlled to prevent damage from inflammation. We demonstrate here that among all tested UBX-domain-containing protein family members, UBXN1 exhibits the strongest inhibitory effect on RNA-virus-induced type I interferon response. UBXN1 potently inhibits RLR- and MAVS-induced, but not TLR3-, TLR4-, or DNA-virus-induced innate immune responses. Depletion of UBXN1 enhances virus-induced innate immune responses, including those resulting from RNA viruses such as vesicular stomatitis, Sendai, West Nile, and dengue virus infection, repressing viral replication. Following viral infection, UBXN1 is induced, binds to MAVS, interferes with intracellular MAVS oligomerization, and disrupts the MAVS/TRAF3/TRAF6 signalosome. These findings underscore a critical role of UBXN1 in the modulation of a major antiviral signaling pathway.

  15. Cell-Type-Specific Regulation of the Retinoic Acid Receptor Mediated by the Orphan Nuclear Receptor TLX†

    Science.gov (United States)

    Kobayashi, Mime; Yu, Ruth T.; Yasuda, Kunio; Umesono, Kazuhiko

    2000-01-01

    Malformations in the eye can be caused by either an excess or deficiency of retinoids. An early target gene of the retinoid metabolite, retinoic acid (RA), is that encoding one of its own receptors, the retinoic acid receptor β (RARβ). To better understand the mechanisms underlying this autologous regulation, we characterized the chick RARβ2 promoter. The region surrounding the transcription start site of the avian RARβ2 promoter is over 90% conserved with the corresponding region in mammals and confers strong RA-dependent transactivation in primary cultured embryonic retina cells. This response is selective for RAR but not retinoid X receptor-specific agonists, demonstrating a principal role for RAR(s) in retina cells. Retina cells exhibit a far higher sensitivity to RA than do fibroblasts or osteoblasts, a property we found likely due to expression of the orphan nuclear receptor TLX. Ectopic expression of TLX in fibroblasts resulted in increased sensitivity to RA induction, an effect that is conserved between chick and mammals. We have identified a cis element, the silencing element relieved by TLX (SET), within the RARβ2 promoter region which confers TLX- and RA-dependent transactivation. These results indicate an important role for TLX in autologous regulation of the RARβ gene in the eye. PMID:11073974

  16. Cell-type-specific regulation of the retinoic acid receptor mediated by the orphan nuclear receptor TLX.

    Science.gov (United States)

    Kobayashi, M; Yu, R T; Yasuda, K; Umesono, K

    2000-12-01

    Malformations in the eye can be caused by either an excess or deficiency of retinoids. An early target gene of the retinoid metabolite, retinoic acid (RA), is that encoding one of its own receptors, the retinoic acid receptor beta (RARbeta). To better understand the mechanisms underlying this autologous regulation, we characterized the chick RARbeta2 promoter. The region surrounding the transcription start site of the avian RARbeta2 promoter is over 90% conserved with the corresponding region in mammals and confers strong RA-dependent transactivation in primary cultured embryonic retina cells. This response is selective for RAR but not retinoid X receptor-specific agonists, demonstrating a principal role for RAR(s) in retina cells. Retina cells exhibit a far higher sensitivity to RA than do fibroblasts or osteoblasts, a property we found likely due to expression of the orphan nuclear receptor TLX. Ectopic expression of TLX in fibroblasts resulted in increased sensitivity to RA induction, an effect that is conserved between chick and mammals. We have identified a cis element, the silencing element relieved by TLX (SET), within the RARbeta2 promoter region which confers TLX- and RA-dependent transactivation. These results indicate an important role for TLX in autologous regulation of the RARbeta gene in the eye.

  17. Deflation-activated receptors, not classical inflation-activated receptors, mediate the Hering-Breuer deflation reflex.

    Science.gov (United States)

    Yu, Jerry

    2016-11-01

    Many airway sensory units respond to both lung inflation and deflation. Whether those responses to opposite stimuli come from one sensor (one-sensor theory) or more than one sensor (multiple-sensor theory) is debatable. One-sensor theory is commonly presumed in the literature. This article proposes a multiple-sensor theory in which a sensory unit contains different sensors for sensing different forces. Two major types of mechanical sensors operate in the lung: inflation- and deflation-activated receptors (DARs). Inflation-activated sensors can be further divided into slowly adapting receptors (SARs) and rapidly adapting receptors (RARs). Many SAR and RAR units also respond to lung deflation because they contain DARs. Pure DARs, which respond to lung deflation only, are rare in large animals but are easily identified in small animals. Lung deflation-induced reflex effects previously attributed to RARs should be assigned to DARs (including pure DARs and DARs associated with SARs and RARs) if the multiple-sensor theory is accepted. Thus, based on the information, it is proposed that activation of DARs can attenuate lung deflation, shorten expiratory time, increase respiratory rate, evoke inspiration, and cause airway secretion and dyspnea.

  18. Ionotropic NMDA and P2X1/5 receptors mediate synaptically induced Ca2+ signalling in cortical astrocytes

    Czech Academy of Sciences Publication Activity Database

    Palygin, O.; Lalo, U.; Verkhratsky, Alexei; Pankratov, Y.

    2010-01-01

    Roč. 48, č. 4 (2010), s. 225-231 ISSN 0143-4160 R&D Projects: GA ČR GA305/08/1384 Institutional research plan: CEZ:AV0Z50390703 Keywords : Astroglia * Ca2+ signalling * NMDA receptors Subject RIV: FH - Neurology Impact factor: 3.553, year: 2010

  19. Investigation of the receptor-mediated endocytosis of transcobalamin/intrinsic factor-vitamin B12 complexes

    DEFF Research Database (Denmark)

    Beedholm, Rasmus; Grissom, Charles B.; Fedosov, Sergey N.

    receptor structure. This receptor is suggested to be regulated by the vitamin B12 level in the cells, which is interesting in relation to cancer growth. The cellular endocytosis of TC- B12 complex by this unknown receptor is being investigated, using confocal microscopy. Fluorescently labeled B12 molecules...... (Oregon green linked to B12) have been synthesized to determine the B12 uptake level in normal and various tumour-derived cells (e.g. Hela cells from cervix epithelioid carcinoma and BN- cells from rat yolk sac sarcoma). Costaining of the B12 binders has been performed using fluorescently labelled...

  20. Kinin B1 receptors mediate depression-like behavior response in stressed mice treated with systemic E. coli lipopolysaccharide

    Directory of Open Access Journals (Sweden)

    Campos Maria M

    2010-12-01

    Full Text Available Abstract Background Kinin B1 receptors are inducible molecules up-regulated after inflammatory stimuli. This study evaluated the relevance of kinin B1 receptors in a mouse depression behavior model. Methods Mice were exposed to a 5-min swimming session, and 30 min later they were injected with E. coli lipopolysaccharide (LPS. Depression-like behavior was assessed by determining immobility time in a tail suspension test. Different brain structures were collected for molecular and immunohistochemical studies. Anhedonia was assessed by means of a sucrose intake test. Results Our protocol elicited an increase in depression-like behavior in CF1 mice, as assessed by the tail-suspension test, at 24 h. This behavior was significantly reduced by treatment with the selective B1 receptor antagonists R-715 and SSR240612. Administration of SSR240612 also prevented an increase in number of activated microglial cells in mouse hippocampus, but did not affect a reduction in expression of mRNA for brain-derived neurotrophic factor. The increased immobility time following LPS treatment was preceded by an enhancement of hippocampal and cortical B1 receptor mRNA expression (which were maximal at 1 h, and a marked production of TNFα in serum, brain and cerebrospinal fluid (between 1 and 6 h. The depression-like behavior was virtually abolished in TNFα p55 receptor-knockout mice, and increased B1 receptor mRNA expression was completely absent in this mouse strain. Furthermore, treatment with SSR240612 was also effective in preventing anhedonia in LPS-treated mice, as assessed using a sucrose preference test. Conclusion Our data show, for the first time, involvement of kinin B1 receptors in depressive behavioral responses, in a process likely associated with microglial activation and TNFα production. Thus, selective and orally active B1 receptor antagonists might well represent promising pharmacological tools for depression therapy.

  1. Valsartan ameliorates ageing-induced aorta degeneration via angiotensin II type 1 receptor-mediated ERK activity

    Science.gov (United States)

    Shan, HaiYan; Zhang, Siyang; Li, Xuelian; yu, Kai; Zhao, Xin; Chen, Xinyue; Jin, Bo; Bai, XiaoJuan

    2014-01-01

    Angiotensin II (Ang II) plays important roles in ageing-related disorders through its type 1 receptor (AT1R). However, the role and underlying mechanisms of AT1R in ageing-related vascular degeneration are not well understood. In this study, 40 ageing rats were randomly divided into two groups: ageing group which received no treatment (ageing control), and valsartan group which took valsartan (selective AT1R blocker) daily for 6 months. 20 young rats were used as adult control. The aorta structure were analysed by histological staining and electron microscopy. Bcl-2/Bax expression in aorta was analysed by immunohistochemical staining, RT-PCR and Western blotting. The expressions of AT1R, AT2R and mitogen-activated protein kinases (MAPKs) were detected. Significant structural degeneration of aorta in the ageing rats was observed, and the degeneration was remarkably ameliorated by long-term administration of valsartan. With ageing, the expression of AT1R was elevated, the ratio of Bcl-2/Bax was decreased and meanwhile, an important subgroup of MAPKs, extracellular signal-regulated kinase (ERK) activity was elevated. However, these changes in ageing rats could be reversed to some extent by valsartan. In vitro experiments observed consistent results as in vivo study. Furthermore, ERK inhibitor could also acquire partial effects as valsartan without affecting AT1R expression. The results indicated that AT1R involved in the ageing-related degeneration of aorta and AT1R-mediated ERK activity was an important mechanism underlying the process. PMID:24548645

  2. The serotonin receptor mediates changes in autonomic neurotransmission and gastrointestinal transit induced by heat-killed Lactobacillus brevis SBC8803.

    Science.gov (United States)

    Horii, Y; Nakakita, Y; Misonou, Y; Nakamura, T; Nagai, K

    2015-01-01

    Lactobacilli exhibit several health benefits in mammals, including humans. Our previous reports established that heat-killed Lactobacillus brevis SBC8803 (SBC8803) increased both efferent gastric vagal nerve activity and afferent intestinal vagal nerve activity in rats. We speculated that this strain could be useful for the treatment of gastrointestinal (GI) disorders. In this study, we examined the effects of SBC8803 on peristalsis and the activity of the efferent celiac vagal nerve innervating the intestine in rats. First, we examined the effects of intraduodenal (ID) administration of SBC8803 on efferent celiac vagal nerve activity (efferent CVNA) in urethane-anesthetised rats using electrophysiological studies. The effects of intravenous injection of the serotonin 5-HT3 receptor antagonist granisetron on changes in efferent CVNA due to ID administration of SBC8803 were also investigated. Finally, the effects of oral gavage of SBC8803 on GI transit were analysed using the charcoal propulsion method in conscious rats treated with or without granisetron. ID administration of SBC8803 increased efferent CVNA. Pretreatment with granisetron eliminated SBC8803-dependent changes in efferent CVNA. Furthermore, oral gavage of SBC8803 significantly accelerated GI transit, while pretreatment with granisetron inhibited GI transit. Our findings suggested that SBC8803 increased efferent CVNA and GI transit of charcoal meal via 5-HT3 receptors. Moreover, SBC8803 enhanced the activity of efferent vagal nerve innervating the intestine and promoted peristalsis via 5-HT3 receptors.

  3. Selectivity and specificity of sphingosine-1-phosphate receptor ligands: caveats and critical thinking in characterizing receptor-mediated effects.

    Science.gov (United States)

    Salomone, Salvatore; Waeber, Christian

    2011-01-01

    Receptors for sphingosine-1-phosphate (S1P) have been identified only recently. Their medicinal chemistry is therefore still in its infancy, and few selective agonists or antagonists are available. Furthermore, the selectivity of S1P receptor agonists or antagonists is not well established. JTE-013 and BML-241 (also known as CAY10444), used extensively as specific S1P(2) and S1P(3) receptors antagonists respectively, are cases in point. When analyzing S1P-induced vasoconstriction in mouse basilar artery, we observed that JTE-013 inhibited not only the effect of S1P, but also the effect of U46619, endothelin-1 or high KCl; JTE-013 strongly inhibited responses to S1P in S1P(2) receptor knockout mice. Similarly, BML-241 has been shown to inhibit increases in intracellular Ca(2+) concentration via P(2) receptor or α(1A)-adrenoceptor stimulation and α(1A)-adrenoceptor-mediated contraction of rat mesenteric artery, while it did not affect S1P(3)-mediated decrease of forskolin-induced cyclic AMP accumulation. Another putative S1P(1/3) receptor antagonist, VPC23019, does not inhibit S1P(3)-mediated vasoconstriction. With these examples in mind, we discuss caveats about relying on available pharmacological tools to characterize receptor subtypes.

  4. Selectivity and specificity of sphingosine-1-phosphate receptor ligands: caveats and critical thinking in characterizing receptor-mediated effects

    Directory of Open Access Journals (Sweden)

    Christian eWaeber

    2011-02-01

    Full Text Available Receptors for sphingosine-1-phosphate (S1P have been identified only recently. Their medicinal chemistry is therefore still in its infancy, and few selective agonists or antagonists are available. Furthermore, the selectivity of S1P receptor agonists or antagonists is not well established. JTE-013 and BML-241 (also known as CAY10444, used extensively as specific S1P2 and S1P3 receptors antagonists respectively, are cases in point. When analyzing S1P-induced vasoconstriction in mouse basilar artery, we observed that JTE-013 inhibited not only the effect of S1P, but also the effect of U46619, endothelin-1 or high KCl; JTE-013 strongly inhibited responses to S1P in S1P2 receptor knockout mice. Similarly, BML-241 has been shown to inhibit increases in intracellular Ca2+ concentration via P2 receptor or α1A-adrenoceptor stimulation and α1A-adrenoceptor-mediated contraction of rat mesenteric artery, while it did not affect S1P3-mediated decrease of forskolin-induced cyclic AMP accumulation. Another putative S1P1/3 receptor antagonist, VPC23019, does not inhibit S1P3-mediated vasoconstriction. With these examples in mind, we discuss caveats about relying on available pharmacological tools to characterize receptor subtypes.

  5. Novel function of transcription factor Nrf2 as an inhibitor of RON tyrosine kinase receptor-mediated cancer cell invasion.

    Science.gov (United States)

    Thangasamy, Amalraj; Rogge, Jessica; Krishnegowda, Naveen K; Freeman, James W; Ammanamanchi, Sudhakar

    2011-09-16

    Recepteur d' origine nantais (RON), a tyrosine kinase receptor, is aberrantly expressed in human tumors and promotes cancer cell invasion. RON receptor activation is also associated with resistance to tamoxifen treatment in breast cancer cells. Nrf2 is a positive regulator of cytoprotective genes. Using chromatin immunoprecipitation (ChIP) and site-directed mutagenesis studies of the RON promoter, we identified Nrf2 as a negative regulator of RON gene expression. High Nrf2 and low RON expression was observed in normal mammary tissue whereas high RON and low or undetectable expression of Nrf2 was observed in breast tumors. The Nrf2 inducer sulforaphane (SFN) as well as ectopic Nrf2 expression or knock-down of the Nrf2 negative regulator keap1, which stabilizes Nrf2, inhibited RON expression and invasion of carcinoma cells. Consequently, our studies identified a novel functional role for Nrf2 as a "repressor" and RON kinase as a molecular target of SFN, which mediates the anti-tumor effects of SFN. These results are not limited to breast cancer cells since the Nrf2 inducer SFN stabilized Nrf2 and inhibited RON expression in carcinoma cells from various tumor types.

  6. NR4A nuclear receptors mediate carnitine palmitoyltransferase 1A gene expression by the rexinoid HX600

    Energy Technology Data Exchange (ETDEWEB)

    Ishizawa, Michiyasu [Division of Biochemistry, Department of Biomedical Sciences, Nihon University School of Medicine, 30-1 Oyaguchi-kamicho, Itabashi-ku, Tokyo 173-8610 (Japan); Kagechika, Hiroyuki [Graduate School of Biomedical Science, Institute of Biomaterials and Bioengineering, Tokyo Medical and Dental University, 2-3-10 Kanda-Surugadai, Chiyoda-ku, Tokyo 101-0062 (Japan); Makishima, Makoto, E-mail: makishima.makoto@nihon-u.ac.jp [Division of Biochemistry, Department of Biomedical Sciences, Nihon University School of Medicine, 30-1 Oyaguchi-kamicho, Itabashi-ku, Tokyo 173-8610 (Japan)

    2012-02-24

    Highlights: Black-Right-Pointing-Pointer The function of RXR heterodimers with NR4 receptors remains unknown. Black-Right-Pointing-Pointer The RXR ligand HX600 induces expression of carnitine palmitoyltransferase 1A (CPT1A). Black-Right-Pointing-Pointer HX600-induced CPT1A expression is mediated by the NR4 receptors, Nur77 and NURR1. Black-Right-Pointing-Pointer CPT1A induction by HX600 is not mediated by de novo protein synthesis. Black-Right-Pointing-Pointer CPT1A could be a target of the Nur77-RXR and NURR1-RXR heterodimers. -- Abstract: Retinoid X receptors (RXRs) are members of the nuclear receptor superfamily and can be activated by 9-cis retinoic acid (9CRA). RXRs form homodimers and heterodimers with other nuclear receptors such as the retinoic acid receptor and NR4 subfamily nuclear receptors, Nur77 and NURR1. Potential physiological roles of the Nur77-RXR and NURR1-RXR heterodimers have not been elucidated. In this study, we identified a gene regulated by these heterodimers utilizing HX600, a selective RXR agonist for Nur77-RXR and NURR1-RXR. While 9CRA induced many genes, including RAR-target genes, HX600 effectively induced only carnitine palmitoyltransferase 1A (CPT1A) in human teratocarcinoma NT2/D1 cells, which express RXR{alpha}, Nur77 and NURR1. HX600 also increased CPT1A expression in human embryonic kidney (HEK) 293 cells and hepatocyte-derived HepG2 cells. Although HX600 induced CPT1A less effectively than 9CRA, overexpression of Nur77 or NURR1 increased the HX600 response to levels similar to 9CRA in NT2/D1 and HEK293 cells. A dominant-negative form of Nur77 or NURR1 repressed the induction of CPT1A by HX600. A protein synthesis inhibitor did not alter HX600-dependent CPT1A induction. Thus, the rexinoid HX600 directly induces expression of CPT1A through a Nur77 or NURR1-mediated mechanism. CPT1A, a gene involved in fatty acid {beta}-oxidation, could be a target of RXR-NR4 receptor heterodimers.

  7. β2-Adrenergic Receptor-Mediated HIF-1α Upregulation Mediates Blood Brain Barrier Damage in Acute Cerebral Ischemia

    Directory of Open Access Journals (Sweden)

    Yanyun Sun

    2017-08-01

    Full Text Available Disruption of the blood brain barrier (BBB within the thrombolytic time window is an antecedent event to intracerebral hemorrhage in ischemic stroke. Our recent studies showed that 2-h cerebral ischemia induced BBB damage in non-infarcted area and secreted matrix metalloproteinase-2 (MMP-2 accounted for this disruption. However, the factors that affect MMP-2 secretion and regulate BBB damage remains unknown. Since hypoxia-inducible factor-1 alpha (HIF-1α was discovered as a mater regulator in hypoxia, we sought to investigate the roles of HIF-1α in BBB damage as well as the factors regulating HIF-1α expression in the ischemic brain. in vivo rat middle cerebral artery occlusion (MCAO and in vitro oxygen glucose deprivation (OGD models were used to mimic ischemia. Pretreatment with HIF-1α inhibitor YC-1 significantly inhibited 2-h MCAO-induced BBB damage, which was accompanied by suppressed occludin degradation and vascular endothelial growth factor (VEGF mRNA upregulation. Interestingly, β2-adrenergic receptor (β2-AR antagonist ICI 118551 attenuated ischemia-induced BBB damage by regulating HIF-1α expression. Double immunostaining showed that HIF-1α was upregulated in ischemic neurons but not in astrocytes andendothelial cells. Of note, HIF-1α inhibition with inhibitor YC-1 or siRNA significantly prevented OGD-induced VEGF upregulation as well as the secretion of VEGF and MMP-2 in neurons. More importantly, blocking β2-AR with ICI 118551 suppressedHIF-1α upregulation in ischemic neurons and attenuated occludin degradation induced by the conditioned media of OGD-treatedneurons. Taken together, blockade of β2-AR-mediated HIF-1α upregulation mediates BBB damage during acute cerebral ischemia. These findings provide new mechanistic understanding of early BBB damage in ischemic stroke and may help reduce thrombolysis-related hemorrhagic complications.

  8. H4 histamine receptors mediate cell cycle arrest in growth factor-induced murine and human hematopoietic progenitor cells.

    Directory of Open Access Journals (Sweden)

    Anne-France Petit-Bertron

    Full Text Available The most recently characterized H4 histamine receptor (H4R is expressed preferentially in the bone marrow, raising the question of its role during hematopoiesis. Here we show that both murine and human progenitor cell populations express this receptor subtype on transcriptional and protein levels and respond to its agonists by reduced growth factor-induced cell cycle progression that leads to decreased myeloid, erythroid and lymphoid colony formation. H4R activation prevents the induction of cell cycle genes through a cAMP/PKA-dependent pathway that is not associated with apoptosis. It is mediated specifically through H4R signaling since gene silencing or treatment with selective antagonists restores normal cell cycle progression. The arrest of growth factor-induced G1/S transition protects murine and human progenitor cells from the toxicity of the cell cycle-dependent anticancer drug Ara-C in vitro and reduces aplasia in a murine model of chemotherapy. This first evidence for functional H4R expression in hematopoietic progenitors opens new therapeutic perspectives for alleviating hematotoxic side effects of antineoplastic drugs.

  9. A cross-talk between TrkB and Ret tyrosine kinases receptors mediates neuroblastoma cells differentiation.

    Directory of Open Access Journals (Sweden)

    Carla Lucia Esposito

    Full Text Available Understanding the interplay between intracellular signals initiated by multiple receptor tyrosine kinases (RTKs to give the final cell phenotype is a major pharmacological challenge. Retinoic acid (RA-treatment of neuroblastoma (NB cells implicates activation of Ret and TrkB RTKs as critical step to induce cell differentiation. By studying the signaling interplay between TrkB and Ret as paradigmatic example, here we demonstrate the existence of a cross-talk mechanism between the two unrelated receptors that is needed to induce the cell differentiation. Indeed, we show that TrkB receptor promotes Ret phosphorylation by a mechanism that does not require GDNF. This reveals to be a key mechanism, since blocking either TrkB or Ret by small interfering RNA causes a failure in NB biochemical and morphological differentiation. Our results provide the first evidence that a functional transactivation between distinct tyrosine kinases receptors is required for an important physiological process.

  10. Quantitative analysis of receptor-mediated uptake and pro-apoptotic activity of mistletoe lectin-1 by high content imaging.

    Science.gov (United States)

    Beztsinna, N; de Matos, M B C; Walther, J; Heyder, C; Hildebrandt, E; Leneweit, G; Mastrobattista, E; Kok, R J

    2018-02-09

    Ribosome inactivating proteins (RIPs) are highly potent cytotoxins that have potential as anticancer therapeutics. Mistletoe lectin 1 (ML1) is a heterodimeric cytotoxic protein isolated from European Mistletoe and belongs to RIP class II. The aim of this project was to systematically study ML1 cell binding, endocytosis pathway(s), subcellular processing and apoptosis activation. For this purpose, state of the art cell imaging equipment and automated image analysis algorithms were used. ML1 displayed very fast binding to sugar residues on the membrane and energy-dependent uptake in CT26 cells. The co-staining with specific antibodies and uptake blocking experiments revealed involvement of both clathrin-dependent and -independent pathways in ML1 endocytosis. Co-localization studies demonstrated the toxin transport from early endocytic vesicles to Golgi network; a retrograde road to the endoplasmic reticulum. The pro-apoptotic and antiproliferative activity of ML1 were shown in time lapse movies and subsequently quantified. ML1 cytotoxicity was less affected in multidrug resistant tumor cell line 4T1 in contrast to commonly used chemotherapeutic drug (ML1 resistance index 6.9 vs 13.4 for doxorubicin; IC 50 : ML1 1.4 ng/ml vs doxorubicin 24000 ng/ml). This opens new opportunities for the use of ML1 as an alternative treatment in multidrug resistant cancers.

  11. Prolonged hypoxia modulates platelet activating factor receptor-mediated responses by fetal ovine pulmonary vascular smooth muscle cells.

    Science.gov (United States)

    Renteria, Lissette S; Raj, J Usha; Ibe, Basil O

    2010-12-01

    Hypoxia augments PAF receptor (PAFr) binding and PAFr protein expression in venous SMC (SMC-PV). We compared effect of acute and prolonged hypoxia (pO(2)<40 torr) on PAFr-mediated responses in arterial SMC (SMC-PA) and SMC-PV. Cells were studied for 30 min (acute) or for 48 h (prolonged) hypoxia and compared to normoxic (pO(2) ~100 torr) conditions. PAF binding was quantified in fmol/10(6) cells (mean ± SEM). PAF binding in normoxia were SMC-PA, 5.2 ± 0.2 and in SMC-PV, 19.3 ± 1.1; values in acute hypoxia were SMC-PA, 7.7 ± 0.4 and in SMC-PV, 27.8 ± 1.7. Prolonged hypoxia produced 6-fold increase in binding in SMC-PA, but only 2-fold increase in SMC-PV, but binding in SMC-PV was still higher. Acute hypoxia augmented inositol phosphate release by 50% and 40% in SMC-PA and SMC-PV, respectively. During normoxia, PAFr mRNA expression by both cell types was similar, but expression in hypoxia by SMC-PA was greater. In SMC-PA, hypoxia and PAF augmented intracellular calcium flux. Re-exposure of cells to 30 min normoxia after 48 h hypoxia decreased binding by 45-60%, suggesting immediate down-regulation of hypoxia-induced PAFr-mediated effects. We speculate that re-oxygenation immediately reverses hypoxia effect probably due to oxygen tension-dependent reversibility of PAFr activation and suggest that exposure of the neonate to prolonged state of hypoxia will vilify oxygen exchange capacity of the neonatal lungs. Copyright © 2010 Elsevier Inc. All rights reserved.

  12. Major Vault Protein Regulates Class A Scavenger Receptor-mediated Tumor Necrosis Factor-α Synthesis and Apoptosis in Macrophages*

    Science.gov (United States)

    Ben, Jingjing; Zhang, Yan; Zhou, Rongmei; Zhang, Haiyang; Zhu, Xudong; Li, Xiaoyu; Zhang, Hanwen; Li, Nan; Zhou, Xiaodan; Bai, Hui; Yang, Qing; Li, Donghai; Xu, Yong; Chen, Qi

    2013-01-01

    Atherosclerosis is considered a disease of chronic inflammation largely initiated and perpetuated by macrophage-dependent synthesis and release of pro-inflammatory mediators. Class A scavenger receptor (SR-A) expressed on macrophages plays a key role in this process. However, how SR-A-mediated pro-inflammatory response is modulated in macrophages remains ill defined. Here through immunoprecipitation coupled with mass spectrometry, we reported major vault protein (MVP) as a novel binding partner for SR-A. The interaction between SR-A and MVP was confirmed by immunofluorescence staining and chemical cross-linking assay. Treatment of macrophages with fucoidan, a SR-A ligand, led to a marked increase in TNF-α production, which was attenuated by MVP depletion. Further analysis revealed that SR-A stimulated TNF-α synthesis in macrophages via the caveolin- instead of clathrin-mediated endocytic pathway linked to p38 and JNK, but not ERK, signaling pathways. Importantly, fucoidan invoked an enrichment of MVP in lipid raft, a caveolin-reliant membrane structure, and enhanced the interaction among SR-A, caveolin, and MVP. Finally, we demonstrated that MVP elimination ameliorated SR-A-mediated apoptosis in macrophages. As such, MVP may fine-tune SR-A activity in macrophages which contributes to the development of atherosclerosis. PMID:23703615

  13. Scavenger receptor-mediated recognition of maleyl bovine plasma albumin and the demaleylated protein in human monocyte macrophages

    International Nuclear Information System (INIS)

    Haberland, M.E.; Fogelman, A.M.

    1985-01-01

    Maleyl bovine plasma albumin competed on an equimolar basis with malondialdehyde low density lipoprotein (LDL) in suppressing the lysosomal hydrolysis of 125 I-labeled malondialdehyde LDL mediated by the scavenger receptor of human monocyte macrophages. Maleyl bovine plasma albumin, in which 94% of the amino groups were modified, exhibited an anodic mobility in agarose electrophoresis 1.7 times that of the native protein. Incubation of maleyl bovine plasma albumin at pH 3.5 regenerated the free amino groups and restored the protein to the same electrophoretic mobility as native albumin. Although ligands recognized by the scavenger receptor typically are anionic, the authors propose that addition of new negative charge achieved by maleylation, rather than directly forming the receptor binding site(s), induces conformational changes in albumin as a prerequisite to expression of the recognition domain(s). They conclude that the primary sequence of albumin, rather than addition of new negative charge, provides the recognition determinant(s) essential for interaction of maleyl bovine plasma albumin with the scavenger receptor

  14. Dorsal hippocampal NMDA receptors mediate the interactive effects of arachidonylcyclopropylamide and MDMA/ecstasy on memory retrieval in rats.

    Science.gov (United States)

    Ghaderi, Marzieh; Rezayof, Ameneh; Vousooghi, Nasim; Zarrindast, Mohammad-Reza

    2016-04-03

    A combination of cannabis and ecstasy may change the cognitive functions more than either drug alone. The present study was designed to investigate the possible involvement of dorsal hippocampal NMDA receptors in the interactive effects of arachidonylcyclopropylamide (ACPA) and ecstasy/MDMA on memory retrieval. Adult male Wistar rats were cannulated into the CA1 regions of the dorsal hippocampus (intra-CA1) and memory retrieval was examined using the step-through type of passive avoidance task. Intra-CA1 microinjection of a selective CB1 receptor agonist, ACPA (0.5-4ng/rat) immediately before the testing phase (pre-test), but not after the training phase (post-training), impaired memory retrieval. In addition, pre-test intra-CA1 microinjection of MDMA (0.5-1μg/rat) dose-dependently decreased step-through latency, indicating an amnesic effect of the drug by itself. Interestingly, pre-test microinjection of a higher dose of MDMA into the CA1 regions significantly improved ACPA-induced memory impairment. Moreover, pre-test intra-CA1 microinjection of a selective NMDA receptor antagonist, D-AP5 (1 and 2μg/rat) inhibited the reversal effect of MDMA on the impairment of memory retrieval induced by ACPA. Pre-test intra-CA1 microinjection of the same doses of D-AP5 had no effect on memory retrieval alone. These findings suggest that ACPA or MDMA consumption can induce memory retrieval impairment, while their co-administration improves this amnesic effect through interacting with hippocampal glutamatergic-NMDA receptor mechanism. Thus, it seems that the tendency to abuse cannabis with ecstasy may be for avoiding cognitive dysfunction. Copyright © 2015. Published by Elsevier Inc.

  15. Hypothesis: The Psychedelic Ayahuasca Heals Traumatic Memories via a Sigma 1 Receptor-Mediated Epigenetic-Mnemonic Process

    Directory of Open Access Journals (Sweden)

    Antonio Inserra

    2018-04-01

    Full Text Available Ayahuasca ingestion modulates brain activity, neurotransmission, gene expression and epigenetic regulation. N,N-Dimethyltryptamine (DMT, one of the alkaloids in Ayahuasca activates sigma 1 receptor (SIGMAR1 and others. SIGMAR1 is a multi-faceted stress-responsive receptor which promotes cell survival, neuroprotection, neuroplasticity, and neuroimmunomodulation. Simultaneously, monoamine oxidase inhibitors (MAOIs also present in Ayahuasca prevent the degradation of DMT. One peculiarity of SIGMAR1 activation and MAOI activity is the reversal of mnemonic deficits in pre-clinical models. Since traumatic memories in post-traumatic stress disorder (PTSD are often characterised by “repression” and PTSD patients ingesting Ayahuasca report the retrieval of such memories, it cannot be excluded that DMT-mediated SIGMAR1 activation and the concomitant MAOIs effects during Ayahuasca ingestion might mediate such “anti-amnesic” process. Here I hypothesise that Ayahuasca, via hyperactivation of trauma and emotional memory-related centres, and via its concomitant SIGMAR1- and MAOIs- induced anti-amnesic effects, facilitates the retrieval of traumatic memories, in turn making them labile (destabilised. As Ayahuasca alkaloids enhance synaptic plasticity, increase neurogenesis and boost dopaminergic neurotransmission, and those processes are involved in memory reconsolidation and fear extinction, the fear response triggered by the memory can be reprogramed and/or extinguished. Subsequently, the memory is stored with this updated significance. To date, it is unclear if new memories replace, co-exist with or bypass old ones. Although the mechanisms involved in memory are still debated, they seem to require the involvement of cellular and molecular events, such as reorganisation of homo and heteroreceptor complexes at the synapse, synaptic plasticity, and epigenetic re-modulation of gene expression. Since SIGMAR1 mobilises synaptic receptor, boosts synaptic plasticity and modulates epigenetic processes, such effects might be involved in the reported healing of traumatic memories in PTSD patients. If this theory proves to be true, Ayahuasca could come to represent the only standing pharmacological treatment which targets traumatic memories in PTSD. Lastly, since SIGMAR1 activation triggers both epigenetic and immunomodulatory programmes, the mechanism here presented could help understanding and treating other conditions in which the cellular memory is dysregulated, such as cancer, diabetes, autoimmune and neurodegenerative pathologies and substance addiction.

  16. Fibroblast growth factor receptor mediates fibroblast-dependent growth in EMMPRIN-depleted head and neck cancer tumor cells.

    Science.gov (United States)

    Liu, Zhiyong; Hartman, Yolanda E; Warram, Jason M; Knowles, Joseph A; Sweeny, Larissa; Zhou, Tong; Rosenthal, Eben L

    2011-08-01

    Head and neck squamous cell carcinoma tumors (HNSCC) contain a dense fibrous stroma which is known to promote tumor growth, although the mechanism of stroma-mediated growth remains unclear. As dysplastic mucosal epithelium progresses to cancer, there is incremental overexpression of extracellular matrix metalloprotease inducer (EMMPRIN) which is associated with tumor growth and metastasis. Here, we present evidence that gain of EMMPRIN expression allows tumor growth to be less dependent on fibroblasts by modulating fibroblast growth factor receptor-2 (FGFR2) signaling. We show that silencing EMMPRIN in FaDu and SCC-5 HNSCC cell lines inhibits cell growth, but when EMMPRIN-silenced tumor cells were cocultured with fibroblasts or inoculated with fibroblasts into severe combined immunodeficient mice, the growth inhibition by silencing EMMPRIN was blunted by the presence of fibroblasts. Coculture experiments showed fibroblast-dependent tumor cell growth occurred via a paracrine signaling. Analysis of tumor gene expression revealed expression of FGFR2 was inversely related to EMMPRIN expression. To determine the role of FGFR2 signaling in EMMPRIN-silenced tumor cells, ligands and inhibitors of FGFR2 were assessed. Both FGF1 and FGF2 enhanced tumor growth in EMMPRIN-silenced cells compared with control vector-transfected cells, whereas inhibition of FGFR2 with blocking antibody or with a synthetic inhibitor (PD173074) inhibited tumor cell growth in fibroblast coculture, suggesting the importance of FGFR2 signaling in fibroblast-mediated tumor growth. Analysis of xenografted tumors revealed that EMMPRIN-silenced tumors had a larger stromal compartment compared with control. Taken together, these results suggest that EMMPRIN acquired during tumor progression promotes fibroblast-independent tumor growth.

  17. Fibroblast growth factor receptor mediates fibroblast-dependent growth in EMMPRIN depleted head and neck cancer tumor cells

    Science.gov (United States)

    Liu, Zhiyong; Hartman, Yolanda E.; Warram, Jason M.; Knowles, Joseph A.; Sweeny, Larrisa; Zhou, Tong; Rosenthal, Eben L.

    2011-01-01

    Head and neck squamous cell carcinoma tumors (HNSCC) contain a dense fibrous stroma which is known to promote tumor growth, although the mechanism of stroma mediated growth remains unclear. As dysplastic mucosal epithelium progresses to cancer there is incremental overexpression of extracellular matrix metalloprotease inducer (EMMPRIN) which is associated with tumor growth and metastasis. Here we present evidence that gain of EMMPRIN expression allows tumor growth to be less dependent on fibroblasts by modulating fibroblast growth factor receptor-2 (FGFR2) signaling. We show that silencing EMMPRIN in FaDu and SCC-5 HNSCC cell lines inhibits cell growth, but when EMMPRIN-silenced tumor cells were co-cultured with fibroblasts or inoculated with fibroblasts into SCID mice, the growth inhibition by silencing EMMPRIN was blunted by the presence of fibroblasts. Co-culture experiments demonstrated fibroblast-dependent tumor cell growth occurred via a paracrine signaling. Analysis of tumor gene expression revealed expression of FGFR2 was inversely related to EMMPRIN expression. To determine the role of FGFR2 signaling in EMMPRIN silenced tumor cells, ligands and inhibitors of FGFR2 were assessed. Both FGF1 and FGF2 enhanced tumor growth in EMMPRIN silenced cells compared to control vector transfected cells, while inhibition of FGFR2 with blocking antibody or with a synthetic inhibitor (PD173074) inhibited tumor cell growth in fibroblast co-culture, suggesting the importance of FGFR2 signaling in fibroblast mediated tumor growth. Analysis of xenografted tumors revealed EMMPRIN silenced tumors had a larger stromal compartment compared to control. Taken together, these results suggest that EMMPRIN acquired during tumor progression promotes fibroblast independent tumor growth. PMID:21665938

  18. Iptakalim inhibits nicotinic acetylcholine receptor-mediated currents in dopamine neurons acutely dissociated from rat substantia nigra pars compacta.

    Science.gov (United States)

    Hu, J; DeChon, J; Yan, K C; Liu, Q; Hu, G; Wu, J

    2006-07-31

    Iptakalim hydrochloride, a novel cardiovascular ATP-sensitive K(+) (K(ATP)) channel opener, has shown remarkable antihypertensive and neuroprotective effects in a variety of studies using in vivo and in vitro preparations. We recently found that iptakalim blocked human alpha4-containing nicotinic acetylcholine receptors (nAChRs) heterologously expressed in the human SH-EP1 cell line. In the present study, we examined the effects of iptakalim on several neurotransmitter-induced current responses in single DA neurons freshly dissociated from rat substantia nigra pars compacta (SNc), using perforated patch-clamp recordings combined with a U-tube rapid drug application. In identified DA neurons under voltage-clamp configuration, glutamate-, NMDA-, and GABA-induced currents were insensitive to co-application with iptakalim (100 microM), while whole-cell currents induced by ACh (1 mM+1 microM atropine) or an alpha4beta2 nicotinic acetylcholine receptors relatively selective agonist, RJR-2403 (300 microM), were eliminated by iptakalim. Iptakalim inhibited RJR-2403-induced current in a concentration-dependent manner, and reduced maximal RJR-2403-induced currents at the highest agonist concentration, suggesting a non-competitive block. In current-clamp mode, iptakalim failed to affect resting membrane potential and spontaneous action potential firing, but abolished RJR-2403-induced neuronal firing acceleration. Together, these results indicate that in dissociated SNc DA neurons, alpha4-containing nAChRs, rather than ionotropic glutamate receptors, GABA(A) receptors or perhaps K-ATP channels are the sensitive targets to mediate iptakalim's pharmacological roles.

  19. CB1 cannabinoid receptor-mediated anandamide signaling mechanisms of the inferior colliculus modulate the haloperidol-induced catalepsy.

    Science.gov (United States)

    Medeiros, P; de Freitas, R L; Silva, M O; Coimbra, N C; Melo-Thomas, L

    2016-11-19

    The inferior colliculus (IC), a midbrain structure that processes acoustic information of aversive nature, is distinguished from other auditory nuclei in the brainstem by its connections with structures of the motor system. Previous evidence relating the IC to motor behavior shows that glutamatergic and GABAergic mechanisms in the IC exert influence on systemic haloperidol-induced catalepsy. There is substantial evidence supporting a role played by the endocannabinoid system as a modulator of the glutamatergic neurotransmission, as well as the dopaminergic activity in the basal nuclei and therefore it may be considered as a potential pharmacological target for the treatment of movement disorders. The present study evaluated if the endocannabinoid system in the IC plays a role in the elaboration of systemic haloperidol-induced catalepsy. Male Wistar rats received intracollicular microinjection of either the endogenous cannabinoid anandamide (AEA) at different concentrations (5, 50 or 100pmol/0.2μl), the CB 1 cannabinoid receptor antagonist AM251 at 50, 100 or 200pmol/0.2μl or vehicle, followed by intraperitoneal (IP) administration of either haloperidol at 0.5 or 1mg/kg or physiological saline. Systemic injection of haloperidol at both doses (0.5 or 1mg/kg, IP) produced a cataleptic state, compared to vehicle/physiological saline-treated group, lasting 30 and 50min after systemic administration of the dopaminergic receptors non-selective antagonist. The midbrain microinjection of AEA at 50pmol/0.2μl increased the latency for stepping down from the horizontal bar after systemic administration of haloperidol. Moreover, the intracollicular administration of AEA at 50pmol/0.2μl was able to increase the duration of catalepsy as compared to AEA at 100pmol/0.2-μl-treated group. Intracollicular pretreatment with AM251 at the intermediate concentration (100pmol/0.2μl) was able to decrease the duration of catalepsy after systemic administration of haloperidol. However, neither the intracollicular microinjection of AM251 at the lowest (50pmol/0.2μl) nor at the highest (200pmol/0.2μl) concentration was able to block the systemic haloperidol-induced catalepsy. Furthermore, the intracollicular administration of AM251 at 100pmol/0.2μl was able to decrease the duration of catalepsy as compared to AM251 at 50pmol/0.2μl- and AM251 at 200pmol/0.2-μl-treated group. The latency for stepping down from the horizontal bar - induced by haloperidol administration - was decreased when microinjection of AEA at 50pmol/0.2μl was preceded with blockade of CB1 receptor with AM251 (100pmol/0.2μl). Our results strengthen the involvement of CB1-signaled endocannabinoid mechanisms of the IC in the neuromodulation of catalepsy induced by systemic administration of the dopaminergic receptors non-selective antagonist haloperidol. Copyright © 2016 IBRO. Published by Elsevier Ltd. All rights reserved.

  20. Toll-Like Receptor-Mediated Free Radical Generation in Clonorchis sinensis Excretory-Secretory Product-Treated Cholangiocarcinoma Cells.

    Science.gov (United States)

    Bahk, Young Yil; Pak, Jhang Ho

    2016-10-01

    Clonorchiasis, caused by direct contact with Clonorchis sinensis worms and their excretory-secretory products (ESPs), is associated with chronic inflammation, malignant changes in bile ducts, and even cholangiocarcinogenesis. Our previous report revealed that intracellular free radicals enzymatically generated by C. sinensis ESPs cause NF-κB-mediated inflammation in human cholangiocarcinoma cells (HuCCT1). Therefore, the present study was conducted to examine the role of upstream Toll-like receptors (TLRs) on the initial host innate immune responses to infection. We found that treatment of HuCCT1 cells with native ESPs induced changes in TLR mRNA levels in a time-dependent manner, concomitant with the generation of free radicals. ESP-mediated free radical generation was markedly attenuated by preincubation of the cells with TLR1-4-neutralizing antibodies, indicating that at least TLR1 through 4 participate in stimulation of the host innate immune responses. These findings indicate that free radicals triggered by ESPs are critically involved in TLR signal transduction. Continuous signaling by this pathway may function in initiating C. sinensis infection-associated inflammation cascades, a detrimental event leading to progression to more severe hepatobiliary diseases.

  1. Protein kinase Cδ signaling downstream of the EGF receptor mediates migration and invasiveness of prostate cancer cells

    International Nuclear Information System (INIS)

    Kharait, Sourabh; Dhir, Rajiv; Lauffenburger, Douglas; Wells, Alan

    2006-01-01

    Tumor progression to the invasive phenotype occurs secondary to upregulated signaling from growth factor receptors that drive key cellular responses like proliferation, migration, and invasion. We hypothesized that Protein kinase Cδ (PKCδ)-mediated transcellular contractility is required for migration and invasion of prostate tumor cells. Two invasive human prostate cancer cell lines, DU145 cells overexpressing wildtype human EGFR (DU145WT) and PC3 cells, were studied. PKCδ is overexpressed in these cells relative to normal prostate epithelial cells, and is activated downstream of EGFR leading to cell motility via modulation of myosin light chain activity. Abrogation of PKCδ using Rottlerin and specific siRNA significantly decreased migration and invasion of both cell lines in vitro. Both PKCδ and phosphorylated PKCδ protein levels were higher in human prostate cancer tissue relative to normal donor prostate as assessed by Western blotting and immunohistochemistry. Thus, we conclude that PKCδ inhibition can limit migration and invasion of prostate cancer cells

  2. Monocyte CD64 or CD89 targeting by surfactant protein D/anti-Fc receptor mediates bacterial uptake.

    NARCIS (Netherlands)

    Tacken, P.J.; Batenburg, J.J.

    2006-01-01

    We recently showed that a chimeric protein, consisting of a recombinant fragment of human surfactant protein D (rfSP-D) coupled to a Fab' fragment directed against the human Fcalpha receptor (CD89), effectively targets pathogens recognized by SP-D to human neutrophils. The present study evaluates

  3. Potentiation of oxycodone antinociception in mice by agmatine and BMS182874 via an imidazoline I2 receptor-mediated mechanism.

    Science.gov (United States)

    Bhalla, Shaifali; Ali, Izna; Lee, Hyaera; Andurkar, Shridhar V; Gulati, Anil

    2013-01-01

    The potentiation of oxycodone antinociception by BMS182874 (endothelin-A (ET(A)) receptor antagonist) and agmatine (imidazoline receptor/α(2)-adrenoceptor agonist) is well-documented. It is also known that imidazoline receptors but not α(2)-adrenoceptors are involved in potentiation of oxycodone antinociception by agmatine and BMS182874 in mice. However, the involvement of specific imidazoline receptor subtypes (I(1), I(2), or both) in this interaction is not clearly understood. The present study was conducted to determine the involvement of imidazoline I(1) and I(2) receptors in agmatine- and BMS182874-induced potentiation of oxycodone antinociception in mice. Antinociceptive (tail flick and hot-plate) latencies were determined in male Swiss Webster mice treated with oxycodone, agmatine, BMS182874, and combined administration of oxycodone with agmatine or BMS182874. Efaroxan (imidazoline I(1) receptor antagonist) and BU224 (imidazoline I(2) receptor antagonist) were used to determine the involvement of I(1) and I(2) imidazoline receptors, respectively. Oxycodone produced significant antinociceptive response in mice which was not affected by efaroxan but was blocked by BU224. Agmatine-induced potentiation of oxycodone antinociception was blocked by BU224 but not by efaroxan. Similarly, BMS182874-induced potentiation of oxycodone antinociception was blocked by BU224 but not by efaroxan. This is the first report demonstrating that BMS182874- or agmatine-induced enhancement of oxycodone antinociception is blocked by BU224 but not by efaroxan. We conclude that imidazoline I(2) receptors but not imidazoline I(1) receptors are involved in BMS182874- and agmatine-induced potentiation of oxycodone antinociception in mice. Copyright © 2012 Elsevier Inc. All rights reserved.

  4. Kappa-opioid receptors mediate the antidepressant-like activity of hesperidin in the mouse forced swimming test.

    Science.gov (United States)

    Filho, Carlos B; Del Fabbro, Lucian; de Gomes, Marcelo G; Goes, André T R; Souza, Leandro C; Boeira, Silvana P; Jesse, Cristiano R

    2013-01-05

    The opioid system has been implicated as a contributing factor for major depression and is thought to play a role in the mechanism of action of antidepressants. This study investigated the involvement of the opioid system in the antidepressant-like effect of hesperidin in the mouse forced swimming test. Our results demonstrate that hesperidin (0.1, 0.3 and 1 mg/kg; intraperitoneal) decreased the immobility time in the forced swimming test without affecting locomotor activity in the open field test. The antidepressant-like effect of hesperidin (0.3 mg/kg) in the forced swimming test was prevented by pretreating mice with naloxone (1 mg/kg, a nonselective opioid receptor antagonist) and 2-(3,4-dichlorophenyl)-Nmethyl-N-[(1S)-1-(3-isothiocyanatophenyl)-2-(1-pyrrolidinyl)ethyl] acetamide (DIPPA (1 mg/kg), a selective κ-opioid receptor antagonist), but not with naloxone methiodide (1 mg/kg, a peripherally acting opioid receptor antagonist), naltrindole (3 mg/kg, a selective δ-opioid receptor antagonist), clocinnamox (1 mg/kg, a selective μ-opioid receptor antagonist) or caffeine (3 mg/kg, a nonselective adenosine receptor antagonist). In addition, a sub-effective dose of hesperidin (0.01 mg/kg) produced a synergistic antidepressant-like effect in the forced swimming test when combined with a sub-effective dose of morphine (1 mg/kg). The antidepressant-like effect of hesperidin in the forced swimming test on mice was dependent on its interaction with the κ-opioid receptor, but not with the δ-opioid, μ-opioid or adenosinergic receptors. Taken together, these results suggest that hesperidin possesses antidepressant-like properties and may be of interest as a therapeutic agent for the treatment of depressive disorders. Published by Elsevier B.V.

  5. Striatal dopamine D2/3 receptor-mediated neurotransmission in major depression: Implications for anhedonia, anxiety and treatment response.

    Science.gov (United States)

    Peciña, Marta; Sikora, Magdalena; Avery, Erich T; Heffernan, Joseph; Peciña, Susana; Mickey, Brian J; Zubieta, Jon-Kar

    2017-10-01

    Dopamine (DA) neurotransmission within the brain's reward circuit has been implicated in the pathophysiology of depression and in both, cognitive and pharmacological mechanisms of treatment response. Still, a direct relationship between measures of DA neurotransmission and reward-related deficits in patients with depression has not been demonstrated. To gain insight into the symptom-specific alterations in the DA system in patients with depression, we used positron emission tomography (PET) and the D 2/3 receptor-selective radiotracer [ 11 C]raclopride in twenty-three non-smoking un-medicated Major Depressive Disorder (MDD) patients and sixteen healthy controls (HC). We investigated the relationship between D 2/3 receptor availability and baseline measures of depression severity, anxiety, anhedonia, and cognitive and pharmacological mechanisms of treatment response. We found that, compared to controls, patients with depression showed greater D 2/3 receptor availability in several striatal regions, including the bilateral ventral pallidum/nucleus accumbens (vPAL/NAc), and the right ventral caudate and putamen. In the depressed sample, D 2/3 receptor availability in the caudal portion of the ventral striatum (NAc/vPAL) correlated with higher anxiety symptoms, whereas D 2/3 receptor availability in the rostral area of the ventral striatum correlated negatively with the severity of motivational anhedonia. Finally, MDD non-remitters showed greater baseline anxiety, greater D 2/3 availability in the NAc/vPAL, and greater placebo-induced DA release in the bilateral NAc. Our results demonstrate abnormally high D 2/3 receptor availability in the ventral striatum of patients with MDD, which seem to be associated with comorbid anxiety symptoms and lack of response to antidepressants. Copyright © 2017 Elsevier B.V. and ECNP. All rights reserved.

  6. Central oxytocin receptors mediate mating-induced partner preferences and enhance correlated activation across forebrain nuclei in male prairie voles.

    Science.gov (United States)

    Johnson, Zachary V; Walum, Hasse; Jamal, Yaseen A; Xiao, Yao; Keebaugh, Alaine C; Inoue, Kiyoshi; Young, Larry J

    2016-03-01

    Oxytocin (OT) is a deeply conserved nonapeptide that acts both peripherally and centrally to modulate reproductive physiology and sociosexual behavior across divergent taxa, including humans. In vertebrates, the distribution of the oxytocin receptor (OTR) in the brain is variable within and across species, and OTR signaling is critical for a variety of species-typical social and reproductive behaviors, including affiliative and pair bonding behaviors in multiple socially monogamous lineages of fishes, birds, and mammals. Early work in prairie voles suggested that the endogenous OT system modulates mating-induced partner preference formation in females but not males; however, there is significant evidence that central OTRs may modulate pair bonding behavior in both sexes. In addition, it remains unclear how transient windows of central OTR signaling during sociosexual interaction modulate neural activity to produce enduring shifts in sociobehavioral phenotypes, including the formation of selective social bonds. Here we re-examine the role of the central OT system in partner preference formation in male prairie voles using a selective OTR antagonist delivered intracranially. We then use the same antagonist to examine how central OTRs modulate behavior and immediate early gene (Fos) expression, a metric of neuronal activation, in males during brief sociosexual interaction with a female. Our results suggest that, as in females, OTR signaling is critical for partner preference formation in males and enhances correlated activation across sensory and reward processing brain areas during sociosexual interaction. These results are consistent with the hypothesis that central OTR signaling facilitates social bond formation by coordinating activity across a pair bonding neural network. Copyright © 2015 Elsevier Inc. All rights reserved.

  7. Excitatory amino acid receptors mediate asymmetry and lateralization in the descending cardiovascular pathways from the dorsomedial hypothalamus.

    Directory of Open Access Journals (Sweden)

    Carlos Henrique Xavier

    Full Text Available The dorsomedial hypothalamus (DMH and lateral/dorsolateral periaqueductal gray (PAG are anatomically and functionally connected. Both the DMH and PAG depend on glutamatergic inputs for activation. We recently reported that removal of GABA-ergic tone in the unilateral DMH produces: asymmetry, that is, a right- (R- sided predominance in cardiac chronotropism, and lateralization, that is, a greater increase in ipsilateral renal sympathetic activity (RSNA. In the current study, we investigated whether excitatory amino acid (EAA receptors in the DMH-PAG pathway contribute to the functional interhemispheric difference. In urethane (1.2 to 1.4 g/kg, i.p. anesthetized rats, we observed that: (i nanoinjections of N-methyl D-aspartate (NMDA 100 pmol/100 nl into the unilateral DMH produced the same right-sided predominance in the control of cardiac chronotropy, (ii nanoinjections of NMDA into the ipsilateral DMH or PAG evoked lateralized RSNA responses, and (iii blockade of EAA receptors in the unilateral DMH attenuated the cardiovascular responses evoked by injection of NMDA into either the R- or left- (L- PAG. In awake rats, nanoinjection of kynurenic acid (1 nmol/100 nL into the L-DMH or R- or L-PAG attenuated the tachycardia evoked by air stress. However, the magnitude of stress-evoked tachycardia was smallest when the EAA receptors of the R-DMH were blocked. We conclude that EAA receptors contribute to the right-sided predominance in cardiac chronotropism. This interhemispheric difference that involves EAA receptors was observed in the DMH but not in the PAG.

  8. Cold Exposure Induces Proliferation of Mature Brown Adipocyte in a ß3-Adrenergic Receptor-Mediated Pathway.

    Science.gov (United States)

    Fukano, Keigo; Okamatsu-Ogura, Yuko; Tsubota, Ayumi; Nio-Kobayashi, Junko; Kimura, Kazuhiro

    2016-01-01

    Hyperplasia of brown adipose tissue (BAT) is a fundamental mechanism for adaptation to survive in the cold environment in rodents. To determine which cell types comprising BAT contribute to tissue hyperplasia, immunohistochemical analysis using a proliferative marker Ki67 was performed on the BAT from 6-week-old C57BL/6J mice housed at 23°C (control) or 10°C (cold) for 5 days. Interestingly, in the control group, the cell proliferative marker Ki67 was detected in the nuclei of uncoupling protein 1-positive mature brown adipocytes (7.2% ± 0.4% of brown adipocyte), as well as in the non-adipocyte stromal-vascular (SV) cells (19.6% ± 2.3% of SV cells), which include preadiopocytes. The percentage of Ki67-positive brown adipocytes increased to 25.6% ± 1.8% at Day 1 after cold exposure and was significantly higher than the non-cold acclimated control until Day 5 (21.8% ± 1.7%). On the other hand, the percentage of Ki67-positive SV cells gradually increased by a cold exposure and peaked to 42.1% ± 8.3% at Day 5. Injection of a ß3-adrenergic receptor (ß3-AR) agonist for continuous 5 days increased the number of Ki67-positive brown adipocytes even at Day 1 but not that of SV cells. In addition, the ß3-AR antagonist, but not ß1-AR antagonist, attenuated the cold exposure-induced increase in the number of Ki67-positive brown adipocytes. These results suggest that mature brown adipocytes proliferate immediately after cold exposure in a ß3-AR-mediated pathway. Thus, proliferation of mature brown adipocytes as well as preadipocytes in SV cells may contribute to cold exposure-induced BAT hyperplasia.

  9. Synthesis of 99mTc-oxybutynin for M3-receptor-mediated imaging of urinary bladder

    International Nuclear Information System (INIS)

    Moustapha, M.E.; Benha University, Benha; Motaleb, M.A.; Ibrahim, I.T.

    2011-01-01

    Radiolabeling of oxybutynin, a muscarinic acetylcholine (mACh) receptor antagonist agent with 99m Tc is of considerable interest for imaging of urinary bladder. This study is aimed to optimize radiolabeling yield of oxybutynin with 99m Tc using SnCl 2 x 2H 2 O as a reducing agent with respect to factors that affect the reaction conditions such as oxybutynin amount, stannous chloride amount, reaction time and pH of the reaction mixture. In vitro stability of the radiolabeled complex was checked and it was found to be stable for up to 8 h. 99m Tc-oxybutynin was injected via subcutaneous and intravenous administration routes into normal Sprague-Dawley rats. Biodistribution studies have revealed that 99m Tc-oxybutynin exhibits high affinity and specificity for the muscarinic M 3 subtype located on the smooth muscle of urinary bladder relative to the M 1 and M 2 subtypes of the G protein coupled receptor (GPCR) superfamily. In vivo uptake of subcutaneous 99m Tc-oxybutynin in urinary bladder was 19.6 ± 0.42% ID at 0.5 h, whereas in intravenous administration route the accumulation in the urinary bladder was found to be 9.4 ± 0.31% ID at 0.5 h post injection. Administration of cold oxybutynin effectively blocked urinary bladder uptake and further confirms the high specificity of this complex for the M 3 receptor. (author)

  10. Different forms of glycine- and GABAA-receptor mediated inhibitory synaptic transmission in mouse superficial and deep dorsal horn neurons

    Directory of Open Access Journals (Sweden)

    Brichta Alan M

    2009-11-01

    Full Text Available Abstract Background Neurons in superficial (SDH and deep (DDH laminae of the spinal cord dorsal horn receive sensory information from skin, muscle, joints and viscera. In both regions, glycine- (GlyR and GABAA-receptors (GABAARs contribute to fast synaptic inhibition. For rat, several types of GABAAR coexist in the two regions and each receptor type provides different contributions to inhibitory tone. Recent work in mouse has discovered an additional type of GlyR, (containing alpha 3 subunits in the SDH. The contribution of differing forms of the GlyR to sensory processing in SDH and DDH is not understood. Methods and Results Here we compare fast inhibitory synaptic transmission in mouse (P17-37 SDH and DDH using patch-clamp electrophysiology in transverse spinal cord slices (L3-L5 segments, 23°C. GlyR-mediated mIPSCs were detected in 74% (25/34 and 94% (25/27 of SDH and DDH neurons, respectively. In contrast, GABAAR-mediated mIPSCs were detected in virtually all neurons in both regions (93%, 14/15 and 100%, 18/18. Several Gly- and GABAAR properties also differed in SDH vs. DDH. GlyR-mediated mIPSC amplitude was smaller (37.1 ± 3.9 vs. 64.7 ± 5.0 pA; n = 25 each, decay time was slower (8.5 ± 0.8 vs. 5.5 ± 0.3 ms, and frequency was lower (0.15 ± 0.03 vs. 0.72 ± 0.13 Hz in SDH vs. DDH neurons. In contrast, GABAAR-mediated mIPSCs had similar amplitudes (25.6 ± 2.4, n = 14 vs. 25. ± 2.0 pA, n = 18 and frequencies (0.21 ± 0.08 vs. 0.18 ± 0.04 Hz in both regions; however, decay times were slower (23.0 ± 3.2 vs. 18.9 ± 1.8 ms in SDH neurons. Mean single channel conductance underlying mIPSCs was identical for GlyRs (54.3 ± 1.6 pS, n = 11 vs. 55.7 ± 1.8, n = 8 and GABAARs (22.7 ± 1.7 pS, n = 10 vs. 22.4 ± 2.0 pS, n = 11 in both regions. We also tested whether the synthetic endocanabinoid, methandamide (methAEA, had direct effects on Gly- and GABAARs in each spinal cord region. MethAEA (5 μM reduced GlyR-mediated mIPSC frequency in SDH and DDH, but did not affect other properties. Similar results were observed for GABAAR mediated mIPSCs, however, rise time was slowed by methAEA in SDH neurons. Conclusion Together these data show that Gly- and GABAARs with clearly differing physiological properties and cannabinoid-sensitivity contribute to fast synaptic inhibition in mouse SDH and DDH.

  11. A membrane glucocorticoid receptor mediates the rapid/non-genomic actions of glucocorticoids in mammalian skeletal muscle fibres.

    Science.gov (United States)

    Pérez, María Hernández-Alcalá; Cormack, Jonathan; Mallinson, David; Mutungi, Gabriel

    2013-10-15

    Glucocorticoids (GCs) are steroid hormones released from the adrenal gland in response to stress. They are also some of the most potent anti-inflammatory and immunosuppressive drugs currently in clinical use. They exert most of their physiological and pharmacological actions through the classical/genomic pathway. However, they also have rapid/non-genomic actions whose physiological and pharmacological functions are still poorly understood. Therefore, the primary aim of this study was to investigate the rapid/non-genomic effects of two widely prescribed glucocorticoids, beclomethasone dipropionate (BDP) and prednisolone acetate (PDNA), on force production in isolated, intact, mouse skeletal muscle fibre bundles. The results show that the effects of both GCs on maximum isometric force (Po) were fibre-type dependent. Thus, they increased Po in the slow-twitch fibre bundles without significantly affecting that of the fast-twitch fibre bundles. The increase in Po occurred within 10 min and was insensitive to the transcriptional inhibitor actinomycin D. Also, it was maximal at ∼250 nM and was blocked by the glucocorticoid receptor (GCR) inhibitor RU486 and a monoclonal anti-GCR, suggesting that it was mediated by a membrane (m) GCR. Both muscle fibre types expressed a cytosolic GCR. However, a mGCR was present only in the slow-twitch fibres. The receptor was more abundant in oxidative than in glycolytic fibres and was confined mainly to the periphery of the fibres where it co-localised with laminin. From these findings we conclude that the rapid/non-genomic actions of GCs are mediated by a mGCR and that they are physiologically/therapeutically beneficial, especially in slow-twitch muscle fibres.

  12. Protein expression from exogenous mRNA: uptake by receptor-mediated endocytosis and trafficking via the lysosomal pathway.

    NARCIS (Netherlands)

    Lorenz, C.; Fotin-Mleczek, M.; Roth, G.; Becker, C.; Dam, T.C.; Verdurmen, W.P.R.; Brock, R.E.; Probst, J.; Schlake, T.

    2011-01-01

    Insertional mutagenesis and the inherent risk of malignancy compromise the clinical use of DNA-based therapies. Being a transient copy of genetic material, mRNA is a safe alternative, overcoming this limitation. As a prerequisite for the development of efficient mRNA-based therapies, we investigated

  13. The brain cytoplasmic RNA BC1 regulates dopamine D-2 receptor-mediated transmission in the striatum

    OpenAIRE

    Centonze, Diego; Rossi, Silvia; Napoli, Ilaria; Mercaldo, Valentina; Lacoux, Caroline; Ferrari, Francesca; Ciotti, Maria Teresa; De Chiara, Valentina; Prosperetti, Chiara; Maccarrone, Mauro; Fezza, Filomena; Calabresi, Paolo; Bernardi, Giorgio; Bagni, Claudia

    2007-01-01

    Dopamine D-2 receptor (D2DR)-mediated transmission in the striatum is remarkably flexible, and changes in its efficacy have been heavily implicated in a variety of physiological and pathological conditions. Although receptor-associated proteins are clearly involved in specific forms of synaptic plasticity, the molecular mechanisms regulating the sensitivity of D-2 receptors in this brain area are essentially obscure. We have studied the physiological responses of the D2DR stimulations in mice...

  14. New generic approach to the treatment of organophosphate poisoning : Adenosine receptor mediated inhibition of ACh-release

    NARCIS (Netherlands)

    van Helden, HPM; Moor, E; Westerink, BHC; Bruijnzeel, PLB

    1998-01-01

    Current treatment of acute organophosphate (OP) poisoning includes a combined administration of a cholinesterase reactivator (oxime), a muscarinic receptor antagonist (atropine) and an anticonvulsant (diazepam). This treatment is not adequate since it does not prevent neuronal brain damage and

  15. The CRF1 and the CRF2 receptor mediate recognition memory deficits and vulnerability induced by opiate withdrawal.

    Science.gov (United States)

    Morisot, Nadège; Contarino, Angelo

    2016-06-01

    Opiate use disorders are associated with impaired cognitive function and altered stress-responsive systems. The corticotropin-releasing factor (CRF) system mediates stress responses via CRF1 and CRF2 receptors and may be implicated in substance use disorders. However, the specific role for each of the two known CRF receptor subtypes in cognitive impairment induced by opiate administration and withdrawal remains to be elucidated. In the present study, CRF1-/-, CRF2-/- and their respective wild-type mice are injected with escalating doses of morphine and cognitive function assessed by the novel object recognition (NOR) memory task throughout relatively long periods of opiate withdrawal. Early (2 days) phases of opiate withdrawal impair NOR memory in wild-type, CRF1-/- and CRF2-/- mice. However, the duration of opiate withdrawal-induced NOR memory deficits is prolonged in CRF1-/- but shortened in CRF2-/- mice, as compared to their respective wild-type mice, indicating opposite roles for the two CRF receptor subtypes. Nevertheless, following apparent recovery, exposure to an environmental stressor induces the reemergence of NOR memory deficits in long-term opiate-withdrawn wild-type but not CRF1-/- or CRF2-/- mice, indicating an essential role for both CRF receptor subtypes in stress vulnerability. These findings bring initial evidence of a complex physiopathological role for the CRF system in cognitive deficits and the long-lasting vulnerability induced by opiate drugs. Copyright © 2016 Elsevier Ltd. All rights reserved.

  16. Guanine nucleotide regulation of muscarinic receptor-mediated inositol phosphate formation in permeabilized 1321N1 cells

    International Nuclear Information System (INIS)

    Orellana, S.A.; Trilivas, I.; Brown, J.H.

    1986-01-01

    Carbachol and guanine nucleotides stimulate formation of the ( 3 H)inositol phosphates IP, IP2, and IP3 in saponin-permeabilized monolayers labelled with ( 3 H) inositol. Carbachol alone has little effect on formation of the ( 3 H) inositol phosphates (IPs), but GTPγS causes synergistic accumulation of ( 3 H)IPs to levels similar to those seen in intact cells. GTP, GppNHp, and GTPγS all support formation of the ( 3 H)IPs, with or without hormone, but GTPγS is the most effective. In the presence of GTPγS, the effect of carbachol is dose-dependent. Half-maximal and maximal accumulation of the ( 3 H)IPs occur at ∼ 5 μM and ∼ 100 μM carbachol, respectively; values close to those seen in intact cells. GTPγS alone stimulates formation of the ( 3 H)IPs after a brief lag time. The combination of GTPγS and carbachol both increases the rate of, and decreases the lag in, formation of the ( 3 H)IPs. LiCl increases ( 3 H)IP and IP2, but not IP3, accumulation; while 2,3-diphosphoglycerate substantially increases that of ( 3 H)IP3. GTPγS and carbachol cause formation of ( 3 H)IPs in the absence of Ca ++ , but formation induced by GTPγS with or without carbachol is Ca ++ -sensitive over a range of physiological concentrations. Although carbachol, Ca ++ , and GTPγS all have effects on formation of ( 3 H)IPs, GTPγS appears to be a primary and obligatory regulator of phosphoinositide hydrolysis in the permeabilized 1321N1 astrocytoma cell

  17. Region-specificity of GABAA receptor mediated effects on orientation and direction selectivity in cat visual cortical area 18.

    Science.gov (United States)

    Jirmann, Kay-Uwe; Pernberg, Joachim; Eysel, Ulf T

    2009-01-01

    The role of GABAergic inhibition in orientation and direction selectivity has been investigated with the GABA(A)-Blocker bicuculline in the cat visual cortex, and results indicated a region specific difference of functional contributions of GABAergic inhibition in areas 17 and 18. In area 17 inhibition appeared mainly involved in sculpturing orientation and direction tuning, while in area 18 inhibition seemed more closely associated with temporal receptive field properties. However, different types of stimuli were used to test areas 17 and 18 and further studies performed in area 17 suggested an important influence of the stimulus type (single light bars vs. moving gratings) on the evoked responses (transient vs. sustained) and inhibitory mechanisms (GABA(A) vs. GABA(B)) which in turn might be more decisive for the specific results than the cortical region. To insert the missing link in this chain of arguments it was necessary to study GABAergic inhibition in area 18 with moving light bars, which has not been done so far. Therefore, in the present study we investigated area 18 cells responding to oriented moving light bars with extracellular recordings and reversible microiontophoretic blockade of GABAergig inhibition with bicuculline methiodide. The majority of neurons was characterized by a pronounced orientation specificity and variable degrees of direction selectivity. GABA(A)ergic inhibition significantly influenced preferred orientation and preferred direction in area 18. During the action of bicuculline orientation tuning width increased and orientation and direction selectivity indices decreased. Our results obtained in area 18 with moving bar stimuli, although in the proportion of affected cells similar to those described in area 17, quantitatively matched the findings for direction and orientation specificity obtained with moving gratings in area 18. Accordingly, stimulus type is not decisive in area 18 and the GABA(A) dependent, inhibitory intracortical computations involved in orientation specificity are indeed region-specific and in comparison to area 17 less effective in area 18.

  18. Central oxytocin receptors mediate mating-induced partner preferences and enhance correlated activation across forebrain nuclei in male prairie voles

    Science.gov (United States)

    Johnson, Zachary V.; Walum, Hasse; Jamal, Yaseen A.; Xiao, Yao; Keebaugh, Alaine C.; Inoue, Kiyoshi; Young, Larry J.

    2016-01-01

    Oxytocin (OT) is a deeply conserved nonapeptide that acts both peripherally and centrally to modulate reproductive physiology and sociosexual behavior across divergent taxa, including humans. In vertebrates, the distribution of the oxytocin receptor (OTR) in the brain is variable within and across species, and OTR signaling is critical for a variety of species-typical social and reproductive behaviors, including affiliative and pair bonding behaviors in multiple socially monogamous lineages of fishes, birds, and mammals. Early work in prairie voles suggested that the endogenous OT system modulates mating-induced partner preference formation in females but not males; however, there is significant evidence that central OTRs may modulate pair bonding behavior in both sexes. In addition, it remains unclear how transient windows of central OTR signaling during sociosexual interaction modulate neural activity to produce enduring shifts in sociobehavioral phenotypes, including the formation of selective social bonds. Here we re-examine the role of the central OT system in partner preference formation in male prairie voles using a selective OTR antagonist delivered intracranially. We then use the same antagonist to examine how central OTRs modulate behavior and immediate early gene (Fos) expression, a metric of neuronal activation, in males during brief sociosexual interaction with a female. Our results suggest that, as in females, OTR signaling is critical for partner preference formation in males and enhances correlated activation across sensory and reward processing brain areas during sociosexual interaction. These results are consistent with the hypothesis that central OTR signaling facilitates social bond formation by coordinating activity across a pair bonding neural network. PMID:26643557

  19. PA1 Protein, a New Competitive Decelerator Acting at More than One Step to Impede Glucocorticoid Receptor-mediated Transactivation*

    Science.gov (United States)

    Zhang, Zhenhuan; Sun, Yunguang; Cho, Young-Wook; Chow, Carson C.; Simons, S. Stoney

    2013-01-01

    Numerous cofactors modulate the gene regulatory activity of glucocorticoid receptors (GRs) by affecting one or more of the following three major transcriptional properties: the maximal activity of agonists (Amax), the potency of agonists (EC50), and the partial agonist activity of antisteroids (PAA). Here, we report that the recently described nuclear protein, Pax2 transactivation domain interaction protein (PTIP)-associated protein 1 (PA1), is a new inhibitor of GR transactivation. PA1 suppresses Amax, increases the EC50, and reduces the PAA of an exogenous reporter gene in a manner that is independent of associated PTIP. PA1 is fully active with, and strongly binds to, the C-terminal half of GR. PA1 reverses the effects of the coactivator TIF2 on GR-mediated gene induction but is unable to augment the actions of the corepressor SMRT. Analysis of competition assays between PA1 and TIF2 with an exogenous reporter indicates that the kinetic definition of PA1 action is a competitive decelerator at two sites upstream from where TIF2 acts. With the endogenous genes IGFBP1 and IP6K3, PA1 also represses GR induction, increases the EC50, and decreases the PAA. ChIP and re-ChIP experiments indicate that PA1 accomplishes this inhibition of the two genes via different mechanisms as follows: PA1 appears to increase GR dissociation from and reduce GR transactivation at the IGFBP1 promoter regions but blocks GR binding to the IP6K3 promoter. We conclude that PA1 is a new competitive decelerator of GR transactivation and can act at more than one molecularly defined step in a manner that depends upon the specific gene. PMID:23161582

  20. PA1 protein, a new competitive decelerator acting at more than one step to impede glucocorticoid receptor-mediated transactivation.

    Science.gov (United States)

    Zhang, Zhenhuan; Sun, Yunguang; Cho, Young-Wook; Chow, Carson C; Simons, S Stoney

    2013-01-04

    Numerous cofactors modulate the gene regulatory activity of glucocorticoid receptors (GRs) by affecting one or more of the following three major transcriptional properties: the maximal activity of agonists (A(max)), the potency of agonists (EC(50)), and the partial agonist activity of antisteroids (PAA). Here, we report that the recently described nuclear protein, Pax2 transactivation domain interaction protein (PTIP)-associated protein 1 (PA1), is a new inhibitor of GR transactivation. PA1 suppresses A(max), increases the EC(50), and reduces the PAA of an exogenous reporter gene in a manner that is independent of associated PTIP. PA1 is fully active with, and strongly binds to, the C-terminal half of GR. PA1 reverses the effects of the coactivator TIF2 on GR-mediated gene induction but is unable to augment the actions of the corepressor SMRT. Analysis of competition assays between PA1 and TIF2 with an exogenous reporter indicates that the kinetic definition of PA1 action is a competitive decelerator at two sites upstream from where TIF2 acts. With the endogenous genes IGFBP1 and IP6K3, PA1 also represses GR induction, increases the EC(50), and decreases the PAA. ChIP and re-ChIP experiments indicate that PA1 accomplishes this inhibition of the two genes via different mechanisms as follows: PA1 appears to increase GR dissociation from and reduce GR transactivation at the IGFBP1 promoter regions but blocks GR binding to the IP6K3 promoter. We conclude that PA1 is a new competitive decelerator of GR transactivation and can act at more than one molecularly defined step in a manner that depends upon the specific gene.

  1. A role for Toll-like receptor mediated signals in neutrophils in the pathogenesis of the anti-phospholipid syndrome.

    Directory of Open Access Journals (Sweden)

    Gerd Gladigau

    Full Text Available The anti-phospholipid syndrome (APS is characterized by recurrent thrombosis and occurrence of anti-phospholipid antibodies (aPL. aPL are necessary, but not sufficient for the clinical manifestations of APS. Growing evidence suggests a role of innate immune cells, in particular polymorphonuclear neutrophils (PMN and Toll-like receptors (TLR to be additionally involved. aPL activate endothelial cells and monocytes through a TLR4-dependent signalling pathway. Whether this is also relevant for PMN in a similar way is currently not known. To address this issue, we used purified PMN from healthy donors and stimulated them in the presence or absence of human monoclonal aPL and the TLR4 agonist LPS monitoring neutrophil effector functions, namely the oxidative burst, phagocytosis, L-Selectin shedding and IL-8 production. aPL alone were only able to induce minor activation of PMN effector functions at high concentrations. However, in the additional presence of LPS the activation threshold was markedly lower indicating a synergistic activation pathway of aPL and TLR in PMN. In summary, our results indicate that PMN effector functions are directly activated by aPL and boosted by the additional presence of microbial products. This highlights a role for PMN as important innate immune effector cells that contribute to the pathophysiology of APS.

  2. New generic approach to the treatment of organophosphate poisoning: Adenosine receptor mediated inhibition of ACh-release

    NARCIS (Netherlands)

    Helden, H.P.M. van; Groen, B.; Moor, E.; Westerink, B.H.C.; Bruijnzeel, P.L.B.

    1998-01-01

    Current treatment of acute organophosphate (OP) poisoning includes a combined administration of a cholinesterase reactivator (oxime), a muscarinic receptor antagonist (atropine) and an anticonvulsant (diazepam). This treatment is not adequate since it does not prevent neuronal brain damage and

  3. The therapeutic CD38 monoclonal antibody daratumumab induces programmed cell death via fcg receptor-mediated cross-linking

    DEFF Research Database (Denmark)

    Overdijk, Marije B.; Jansen, J. H. Marco; Nederend, Maaike

    2016-01-01

    RIIb as well as activating FcgRs induce DARA cross-linking-mediated PCD. In conclusion, our in vitro and in vivo data show that FcgRmediated cross-linking of DARA induces PCD of CD38-expressing multiple myeloma tumor cells, which potentially contributes to the depth of response observed in DARA......Emerging evidence suggests that FcgR-mediated cross-linking of tumor-bound mAbs may induce signaling in tumor cells that contributes to their therapeutic activity. In this study, we show that daratumumab (DARA), a therapeutic human CD38 mAb with a broad-spectrum killing activity, is able to induce...... programmed cell death (PCD) of CD38+ multiple myeloma tumor cell lines when cross-linked in vitro by secondary Abs or via an FcgR. By comparing DARA efficacy in a syngeneic in vivo tumor model using FcRg-chain knockout or NOTAM mice carrying a signaling-inactive FcRg-chain, we found that the inhibitory Fcg...

  4. The phosphatidylinositol-3 kinase pathway is not essential for insulin-like growth factor I receptor-mediated clonogenic radioresistance

    International Nuclear Information System (INIS)

    Yu, Dong; Watanabe, Hiroshi; Shibuya, Hitoshi; Miura, Masahiko

    2002-01-01

    The insulin-like growth factor I receptor (IGF-IR) is known to induce clonogenic radioresistance in cells following ionizing irradiation. To explore the downstream signaling pathways, we focused on the phosphatidylinositol-3 kinase (PI3-K) pathway, which is thought to be the primary cell survival signal originating from the receptor. For this purpose, R- cells deficient in the endogenous IGF-IR were used as a recipient of the human IGF-IR with or without mutations at potential PI3-K activation sites: NPXY 950 and Y 1316 XXM. Mutats with double mutation at Y950/Y1316 exhibited not abrogated, but reduced activation of insulin receptor substance-1 (IRS-1), PI3-K, and Akt upon IGF-I stimulation. However, the mutants had the same clonogenic radioresistance as cells with wild type (WT) receptors. Neither wortmannin nor LY294002, specific inhibitors of PI3-K, affected the radioresistance of cells with WT receptors at concentrations specific for PI3-K. Collectively, these results indicate that the PI3-K pathway is not essential for IGF-IR-mediated clonogenic radioresistance. (author)

  5. The Effect of Thermal Fluctuation on the Receptor-Mediated Adhesion of a Cell Membrane to an Elastic Substrate

    Directory of Open Access Journals (Sweden)

    Bahador Marzban

    2017-04-01

    Full Text Available Mechanics of the bilayer membrane play an important role in many biological and bioengineering problems such as cell–substrate and cell–nanomaterial interactions. In this work, we study the effect of thermal fluctuation and the substrate elasticity on the cell membrane–substrate adhesion. We model the adhesion of a fluctuating membrane on an elastic substrate as a two-step reaction comprised of the out-of-plane membrane fluctuation and the receptor–ligand binding. The equilibrium closed bond ratio as a function of substrate rigidity was computed by developing a coupled Fourier space Brownian dynamics and Monte Carlo method. The simulation results show that there exists a crossover value of the substrate rigidity at which the closed bond ratio is maximal.

  6. Insulin-Independent GABAA Receptor-Mediated Response in the Barrel Cortex of Mice with Impaired Met Activity.

    Science.gov (United States)

    Lo, Fu-Sun; Erzurumlu, Reha S; Powell, Elizabeth M

    2016-03-30

    Autism spectrum disorder (ASD) is a neurodevelopmental disorder caused by genetic variants, susceptibility alleles, and environmental perturbations. The autism associated geneMETtyrosine kinase has been implicated in many behavioral domains and endophenotypes of autism, including abnormal neural signaling in human sensory cortex. We investigated somatosensory thalamocortical synaptic communication in mice deficient in Met activity in cortical excitatory neurons to gain insights into aberrant somatosensation characteristic of ASD. The ratio of excitation to inhibition is dramatically increased due to decreased postsynaptic GABAAreceptor-mediated inhibition in the trigeminal thalamocortical pathway of mice lacking active Met in the cerebral cortex. Furthermore, in contrast to wild-type mice, insulin failed to increase GABAAreceptor-mediated response in the barrel cortex of mice with compromised Met signaling. Thus, lacking insulin effects may be a risk factor in ASD pathogenesis. A proposed common cause of neurodevelopmental disorders is an imbalance in excitatory neural transmission, provided by the glutamatergic neurons, and the inhibitory signals from the GABAergic interneurons. Many genes associated with autism spectrum disorders impair synaptic transmission in the expected cell type. Previously, inactivation of the autism-associated Met tyrosine kinase receptor in GABAergic interneurons led to decreased inhibition. In thus report, decreased Met signaling in glutamatergic neurons had no effect on excitation, but decimated inhibition. Further experiments indicate that loss of Met activity downregulates GABAAreceptors on glutamatergic neurons in an insulin independent manner. These data provide a new mechanism for the loss of inhibition and subsequent abnormal excitation/inhibition balance and potential molecular candidates for treatment or prevention. Copyright © 2016 the authors 0270-6474/16/363691-07$15.00/0.

  7. AIB1 is required for the acquisition of epithelial growth factor receptor-mediated tamoxifen resistance in breast cancer cells

    International Nuclear Information System (INIS)

    Zhao Wenhui; Zhang Qingyuan; Kang Xinmei; Jin Shi; Lou Changjie

    2009-01-01

    Acquired resistance to tamoxifen has become a serious obstacle in breast cancer treatment. The underlying mechanism responsible for this condition has not been completely elucidated. In this study, a tamoxifen-resistant (Tam-R) MCF-7 breast cancer cell line was developed to mimic the occurrence of acquired tamoxifen resistance as seen in clinical practice. Increased expression levels of HER1, HER2 and the estrogen receptor (ER)-AIB1 complex were found in tamoxifen-resistant cells. EGF stimulation and gefitinib inhibition experiments further demonstrated that HER1/HER2 signaling and AIB1 were involved in the proliferation of cells that had acquired Tam resistance. However, when AIB1 was silenced with AIB1-siRNA in Tam-R cells, the cell growth stimulated by the HER1/HER2 signaling pathway was significantly reduced, and the cells were again found to be inhibited by tamoxifen. These results suggest that the AIB1 protein could be a limiting factor in the HER1/HER2-mediated hormone-independent growth of Tam-R cells. Thus, AIB1 may be a new therapeutic target, and the removal of AIB1 may decrease the crosstalk between ER and the HER1/HER2 pathway, resulting in the restoration of tamoxifen sensitivity in tamoxifen-resistant cells.

  8. Functional characterization of GABAA receptor-mediated modulation of cortical neuron network activity in microelectrode array recordings

    DEFF Research Database (Denmark)

    Bader, Benjamin M; Steder, Anne; Klein, Anders Bue

    2017-01-01

    The numerous γ-aminobutyric acid type A receptor (GABAAR) subtypes are differentially expressed and mediate distinct functions at neuronal level. In this study we have investigated GABAAR-mediated modulation of the spontaneous activity patterns of primary neuronal networks from murine frontal...... of the information extractable from the MEA recordings offers interesting insights into the contributions of various GABAAR subtypes/subgroups to cortical network activity and the putative functional interplay between these receptors in these neurons....... cortex by characterizing the effects induced by a wide selection of pharmacological tools at a plethora of activity parameters in microelectrode array (MEA) recordings. The basic characteristics of the primary cortical neurons used in the recordings were studied in some detail, and the expression levels...

  9. NMDA receptor-mediated long term modulation of electrically evoked field potentials in the rat medial vestibular nuclei.

    Science.gov (United States)

    Capocchi, G; Della Torre, G; Grassi, S; Pettorossi, V E; Zampolini, M

    1992-01-01

    The effect of high frequency stimulation (HFS) of the primary vestibular afferents on field potentials recorded in the ipsilateral Medial Vestibular Nuclei (MVN) was studied. Our results show that potentiation and depression can be induced in different portions of MVN, which are distinguishable by their anatomical organization. HFS induces potentiation of the monosynaptic component in the ventral portion of the MVN, whereas it provokes depression of the polysynaptic component in the dorsal portion of the same nucleus. The induction of both potentiation and depression was blocked under AP5 perfusion, thus demonstrating that NMDA receptor activation mediates both phenomena. Furthermore, the finding that the field potentials were not modified during perfusion with DL-AP5, as previously reported, supports the hypothesis that NMDA receptors are not involved in the normal synaptic transmission from the primary vestibular afferent fibres, but are only activated following hyperstimulation of this afferent system. Our results suggest that the mechanisms of long term modification of synaptic efficacy observed in MVN may underlie the plasticity phenomena occurring in vestibular nuclei.

  10. CHD8, A Novel Beta-Catenin Associated Chromatin Remodeling Enzyme, Regulates Androgen Receptor Mediated Gene Transcription

    National Research Council Canada - National Science Library

    Bochar, Daniel A

    2008-01-01

    .... To better understand the function of beta-catenin in AR mediated transcription, we have identified a novel chromatin remodeling enzyme, CHD8, that can associate with beta-catenin and functions in AR...

  11. Hypothesis: The Psychedelic Ayahuasca Heals Traumatic Memories via a Sigma 1 Receptor-Mediated Epigenetic-Mnemonic Process.

    Science.gov (United States)

    Inserra, Antonio

    2018-01-01

    Ayahuasca ingestion modulates brain activity, neurotransmission, gene expression and epigenetic regulation. N,N -Dimethyltryptamine (DMT, one of the alkaloids in Ayahuasca) activates sigma 1 receptor (SIGMAR1) and others. SIGMAR1 is a multi-faceted stress-responsive receptor which promotes cell survival, neuroprotection, neuroplasticity, and neuroimmunomodulation. Simultaneously, monoamine oxidase inhibitors (MAOIs) also present in Ayahuasca prevent the degradation of DMT. One peculiarity of SIGMAR1 activation and MAOI activity is the reversal of mnemonic deficits in pre-clinical models. Since traumatic memories in post-traumatic stress disorder (PTSD) are often characterised by "repression" and PTSD patients ingesting Ayahuasca report the retrieval of such memories, it cannot be excluded that DMT-mediated SIGMAR1 activation and the concomitant MAOIs effects during Ayahuasca ingestion might mediate such "anti-amnesic" process. Here I hypothesise that Ayahuasca, via hyperactivation of trauma and emotional memory-related centres, and via its concomitant SIGMAR1- and MAOIs- induced anti-amnesic effects, facilitates the retrieval of traumatic memories, in turn making them labile (destabilised). As Ayahuasca alkaloids enhance synaptic plasticity, increase neurogenesis and boost dopaminergic neurotransmission, and those processes are involved in memory reconsolidation and fear extinction, the fear response triggered by the memory can be reprogramed and/or extinguished. Subsequently, the memory is stored with this updated significance. To date, it is unclear if new memories replace, co-exist with or bypass old ones. Although the mechanisms involved in memory are still debated, they seem to require the involvement of cellular and molecular events, such as reorganisation of homo and heteroreceptor complexes at the synapse, synaptic plasticity, and epigenetic re-modulation of gene expression. Since SIGMAR1 mobilises synaptic receptor, boosts synaptic plasticity and modulates epigenetic processes, such effects might be involved in the reported healing of traumatic memories in PTSD patients. If this theory proves to be true, Ayahuasca could come to represent the only standing pharmacological treatment which targets traumatic memories in PTSD. Lastly, since SIGMAR1 activation triggers both epigenetic and immunomodulatory programmes, the mechanism here presented could help understanding and treating other conditions in which the cellular memory is dysregulated, such as cancer, diabetes, autoimmune and neurodegenerative pathologies and substance addiction.

  12. B Cell Receptor-Mediated Internalization of Salmonella: A Novel Pathway for Autonomous B Cell Activation and Antibody Production

    NARCIS (Netherlands)

    Souwer, Yuri; Griekspoor, Alexander; Jorritsma, Tineke; de Wit, Jelle; Janssen, Hans; Neefjes, Jacques; van Ham, S. Marieke

    2009-01-01

    The present paradigm is that primary B cells are nonphagocytosing cells. In this study, we demonstrate that human primary B cells are able to internalize bacteria when the bacteria are recognized by the BCR. BCR-mediated internalization of Salmonella typhimurium results in B cell differentiation and

  13. Cutaneous nociceptors lack sensitisation, but reveal μ-opioid receptor-mediated reduction in excitability to mechanical stimulation in neuropathy

    Directory of Open Access Journals (Sweden)

    Schmidt Yvonne

    2012-11-01

    Full Text Available Abstract Background Peripheral nerve injuries often trigger a hypersensitivity to tactile stimulation. Behavioural studies demonstrated efficient and side effect-free analgesia mediated by opioid receptors on peripheral sensory neurons. However, mechanistic approaches addressing such opioid properties in painful neuropathies are lacking. Here we investigated whether opioids can directly inhibit primary afferent neuron transmission of mechanical stimuli in neuropathy. We analysed the mechanical thresholds, the firing rates and response latencies of sensory fibres to mechanical stimulation of their cutaneous receptive fields. Results Two weeks following a chronic constriction injury of the saphenous nerve, mice developed a profound mechanical hypersensitivity in the paw innervated by the damaged nerve. Using an in vitro skin-nerve preparation we found no changes in the mechanical thresholds and latencies of sensory fibres from injured nerves. The firing rates to mechanical stimulation were unchanged or reduced following injury. Importantly, μ-opioid receptor agonist [D-Ala2,N-Me-Phe4,Gly5]-ol-enkephalin (DAMGO significantly elevated the mechanical thresholds of nociceptive Aδ and C fibres. Furthermore, DAMGO substantially diminished the mechanically evoked discharges of C nociceptors in injured nerves. These effects were blocked by DAMGO washout and pre-treatment with the selective μ-opioid receptor antagonist Cys2-Tyr3-Orn5-Pen7-amide. DAMGO did not alter the responses of sensory fibres in uninjured nerves. Conclusions Our findings suggest that behaviourally manifested neuropathy-induced mechanosensitivity does not require a sensitised state of cutaneous nociceptors in damaged nerves. Yet, nerve injury renders nociceptors sensitive to opioids. Prevention of action potential generation or propagation in nociceptors might represent a cellular mechanism underlying peripheral opioid-mediated alleviation of mechanical hypersensitivity in neuropathy.

  14. NOD1 cooperates with TLR2 to enhance T cell receptor-mediated activation in CD8 T cells.

    Directory of Open Access Journals (Sweden)

    Blandine C Mercier

    Full Text Available Pattern recognition receptors (PRR, like Toll-like receptors (TLR and NOD-like receptors (NLR, are involved in the detection of microbial infections and tissue damage by cells of the innate immune system. Recently, we and others have demonstrated that TLR2 can additionally function as a costimulatory receptor on CD8 T cells. Here, we establish that the intracytosolic receptor NOD1 is expressed and functional in CD8 T cells. We show that C12-iEDAP, a synthetic ligand for NOD1, has a direct impact on both murine and human CD8 T cells, increasing proliferation and effector functions of cells activated via their T cell receptor (TCR. This effect is dependent on the adaptor molecule RIP2 and is associated with an increased activation of the NF-κB, JNK and p38 signaling pathways. Furthermore, we demonstrate that NOD1 stimulation can cooperate with TLR2 engagement on CD8 T cells to enhance TCR-mediated activation. Altogether our results indicate that NOD1 might function as an alternative costimulatory receptor in CD8 T cells. Our study provides new insights into the function of NLR in T cells and extends to NOD1 the recent concept that PRR stimulation can directly control T cell functions.

  15. Entry of porcine reproductive and respiratory syndrome virus into porcine alveolar macrophages via receptor-mediated endocytosis.

    Science.gov (United States)

    Nauwynck, H J; Duan, X; Favoreel, H W; Van Oostveldt, P; Pensaert, M B

    1999-02-01

    Porcine alveolar macrophages (AMphi) are the dominant cell type that supports the replication of porcine reproductive and respiratory syndrome virus (PRRSV) in vivo and in vitro. In order to determine the characteristics of the virus-receptor interaction, the attachment of PRRSV to cells was examined by using biotinylated virus in a series of flow cytometric assays. PRRSV bound specifically to AMphi in a dose-dependent manner. Binding of PRRSV to AMphi increased gradually and reached a maximum within 60 min at 4 degrees C. By confocal microscopy, it was shown that different degrees of PRRSV binding exist and that entry is by endocytosis. Virus uptake in vesicles is a clathrin-dependent process, as it was blocked by the addition of cytochalasin D and co-localization of PRRSV and clathrin was found. Furthermore, by the use of two weak bases, NH4Cl and chloroquine, it was demonstrated that PRRSV uses a low pH-dependent entry pathway. In the presence of these reagents, input virions accumulated in large vacuoles, indicating that uncoating was prevented. These results indicate that PRRSV entry into AMphi involves attachment to a specific virus receptor(s) followed by a process of endocytosis, by which virions are taken into the cell within vesicles by a clathrin-dependent pathway. A subsequent drop in pH is required for proper virus replication.

  16. Adenosine A₁ and A₂A receptor-mediated modulation of acetylcholine release in the mice neuromuscular junction.

    Science.gov (United States)

    Garcia, Neus; Priego, Mercedes; Obis, Teresa; Santafe, Manel M; Tomàs, Marta; Besalduch, Nuria; Lanuza, M Angel; Tomàs, Josep

    2013-07-01

    Immunocytochemistry shows that purinergic receptors (P1Rs) type A1 and A2A (A1 R and A2 A R, respectively) are present in the nerve endings at the P6 and P30 Levator auris longus (LAL) mouse neuromuscular junctions (NMJs). As described elsewhere, 25 μm adenosine reduces (50%) acetylcholine release in high Mg(2+) or d-tubocurarine paralysed muscle. We hypothesize that in more preserved neurotransmission machinery conditions (blocking the voltage-dependent sodium channel of the muscle cells with μ-conotoxin GIIIB) the physiological role of the P1Rs in the NMJ must be better observed. We found that the presence of a non-selective P1R agonist (adenosine) or antagonist (8-SPT) or selective modulators of A1 R or A2 A R subtypes (CCPA and DPCPX, or CGS-21680 and SCH-58261, respectively) does not result in any changes in the evoked release. However, P1Rs seem to be involved in spontaneous release (miniature endplate potentials MEPPs) because MEPP frequency is increased by non-selective block but decreased by non-selective stimulation, with A1 Rs playing the main role. We assayed the role of P1Rs in presynaptic short-term plasticity during imposed synaptic activity (40 Hz for 2 min of supramaximal stimuli). Depression is reduced by micromolar adenosine but increased by blocking P1Rs with 8-SPT. Synaptic depression is not affected by the presence of selective A1 R and A2 A R modulators, which suggests that both receptors need to collaborate. Thus, A1 R and A2 A R might have no real effect on neuromuscular transmission in resting conditions. However, these receptors can conserve resources by limiting spontaneous quantal leak of acetylcholine and may protect synaptic function by reducing the magnitude of depression during repetitive activity. © 2013 Federation of European Neuroscience Societies and John Wiley & Sons Ltd.

  17. Cross-talk between an activator of nuclear receptors-mediated transcription and the D1 dopamine receptor signaling pathway.

    Science.gov (United States)

    Schmidt, Azriel; Vogel, Robert; Rutledge, Su Jane; Opas, Evan E; Rodan, Gideon A; Friedman, Eitan

    2005-03-01

    Nuclear receptors are transcription factors that usually interact, in a ligand-dependent manner, with specific DNA sequences located within promoters of target genes. The nuclear receptors can also be controlled in a ligand-independent manner via the action of membrane receptors and cellular signaling pathways. 5-Tetradecyloxy-2-furancarboxylic acid (TOFA) was shown to stimulate transcription from the MMTV promoter via chimeric receptors that consist of the DNA binding domain of GR and the ligand binding regions of the PPARbeta or LXRbeta nuclear receptors (GR/PPARbeta and GR/LXRbeta). TOFA and hydroxycholesterols also modulate transcription from NF-kappaB- and AP-1-controlled reporter genes and induce neurite differentiation in PC12 cells. In CV-1 cells that express D(1) dopamine receptors, D(1) dopamine receptor stimulation was found to inhibit TOFA-stimulated transcription from the MMTV promoter that is under the control of chimeric GR/PPARbeta and GR/LXRbeta receptors. Treatment with the D(1) dopamine receptor antagonist, SCH23390, prevented dopamine-mediated suppression of transcription, and by itself increased transcription controlled by GR/LXRbeta. Furthermore, combined treatment of CV-1 cells with TOFA and SCH23390 increased transcription controlled by the GR/LXRbeta chimeric receptor synergistically. The significance of this in vitro synergy was demonstrated in vivo, by the observation that SCH23390 (but not haloperidol)-mediated catalepsy in rats was potentiated by TOFA, thus showing that an agent that mimics the in vitro activities of compounds that activate members of the LXR and PPAR receptor families can influence D1 dopamine receptor elicited responses.

  18. Membrane receptor-mediated apoptosis and caspase activation in the differentiated EoL-1 eosinophilic cell line.

    Science.gov (United States)

    Al-Rabia, Mohammed W; Blaylock, Morgan G; Sexton, Darren W; Walsh, Garry M

    2004-06-01

    Caspases are key molecules in the control of apoptosis, but relatively little is known about their contribution to eosinophil apoptosis. We examined caspase-3, -8, and -9 activities in receptor ligation-dependent apoptosis induction in the differentiated human eosinophilic cell line EoL-1. Differentiated EoL-1 exhibited bi-lobed nuclei, eosinophil-associated membrane receptors, and basic granule proteins. Annexin-V fluorescein isothiocyanate binding to EoL-1 revealed significant (PEoL-1 but had no effect on constitutive (baseline) apoptosis at 16 and 20 h. Caspase activity was analyzed using the novel CaspaTag trade mark technique and flow cytometry. EoL-1 treated with pan-CD45, CD45RA, CD45RB, and CD95 mAb exhibited caspase-3 and -9 activation at 12 h post-treatment, which increased at 16 and 20 h. Activated caspase-8 was detected 12 and 16 h after ligation with CD45, CD45RA, CD45RB, and CD95 mAb followed by a trend toward basal levels at 20 h. CD69 ligation resulted in caspase-3 activation, a modest but significant activation of caspase-8, and a loss in mitochondrial transmembrane potential but had no significant effect on activation of caspase-9. Thus, the intrinsic and extrinsic caspase pathways are involved in controlling receptor ligation-mediated apoptosis induction in human eosinophils, findings that may aid the development of a more targeted, anti-inflammatory therapy for asthma.

  19. Role of Ca+2 and other second messengers in excitatory amino acid receptor mediated neurodegeneration: clinical perspectives

    DEFF Research Database (Denmark)

    Schousboe, A; Belhage, B; Frandsen, A

    1997-01-01

    Neurodegeneration associated with neurological disorders such as epilepsy, Huntington's Chorea, Alzheimer's disease, and olivoponto cerebellar atrophy or with energy failure such as ischemia, hypoxia, and hypoglycemia proceeds subsequent to overexposure of neurons to excitatory amino acids of which...... glutamate and aspartate may be quantitatively the most important. The toxic action of glutamate and aspartate is mediated through activation of glutamate receptors of the N-methyl-D-aspartate (NMDA) and non-NMDA subtypes. Antagonists for these receptors can act as neuroprotectants both in in vitro model...

  20. NK3 Receptors mediate an increase in firing rate of midbrain dopamine neurons of the rat and the guinea pig

    NARCIS (Netherlands)

    Werkman, T.R.; McCreary, A.C.; Kruse, C.G.; Wadman, W.J.

    2011-01-01

    This in vitro study investigates and compares the effects of NK3 receptor ligands on the firing rate of rat and guinea pig midbrain dopamine neurons. The findings are discussed in the light of choosing suitable animal models for investigating pharmacological properties of NK3 receptor antagonists,

  1. Corticotropin-releasing factor (CRF) and α 2 adrenergic receptors mediate heroin withdrawal-potentiated startle in rats.

    Science.gov (United States)

    Park, Paula E; Vendruscolo, Leandro F; Schlosburg, Joel E; Edwards, Scott; Schulteis, Gery; Koob, George F

    2013-09-01

    Anxiety is one of the early symptoms of opioid withdrawal and contributes to continued drug use and relapse. The acoustic startle response (ASR) is a component of anxiety that has been shown to increase during opioid withdrawal in both humans and animals. We investigated the role of corticotropin-releasing factor (CRF) and norepinephrine (NE), two key mediators of the brain stress system, on acute heroin withdrawal-potentiated ASR. Rats injected with heroin (2 mg/kg s.c.) displayed an increased ASR when tested 4 h after heroin treatment. A similar increase in ASR was found in rats 10-20 h into withdrawal from extended access (12 h) to i.v. heroin self-administration, a model that captures several aspects of heroin addiction in humans. Both the α 2 adrenergic receptor agonist clonidine (10 μg/kg s.c.) and CRF1 receptor antagonist N,N-bis(2-methoxyethyl)-3-(4-methoxy-2-methylphenyl)-2,5-dimethyl-pyrazolo[1,5-a] pyrimidin-7-amine (MPZP; 20 mg/kg s.c.) blocked heroin withdrawal-potentiated startle. To investigate the relationship between CRF1 and α 2 adrenergic receptors in the potentiation of the ASR, we tested the effect of MPZP on yohimbine (1.25 mg/kg s.c.)-potentiated startle and clonidine on CRF (2 μg i.c.v.)-potentiated startle. Clonidine blocked CRF-potentiated startle, whereas MPZP partially attenuated but did not reverse yohimbine-potentiated startle, suggesting that CRF may drive NE release to potentiate startle. These results suggest that CRF1 and α 2 receptors play an important role in the heightened anxiety-like behaviour observed during acute withdrawal from heroin, possibly via CRF inducing the release of NE in stress-related brain regions.

  2. Microarray evaluation of EP4 receptor-mediated prostaglandin E2 suppression of 3T3-L1 adipocyte differentiation

    International Nuclear Information System (INIS)

    Sugimoto, Yukihiko; Tsuboi, Hiroaki; Okuno, Yasushi; Tamba, Shigero; Tsuchiya, Soken; Tsujimoto, Gozo; Ichikawa, Atsushi

    2004-01-01

    Prostaglandin E 2 (PGE 2 ) has been shown to negatively regulate adipogenesis. To explore to what extent PGE 2 inhibits the differentiation of cells to adipocytes and to examine whether its effect could be due to EP4 receptor signaling, we used microarrays to analyze the gene expression profiles of 3T3-L1 cells exposed to a differentiation cocktail supplemented with PGE 2 , AE1-329 (an EP4 agonist), or vehicle. The differentiation-associated responses in genes such as adipocytokines and enzymes related to lipid metabolism were largely weakened upon PGE 2 treatment. In particular, the expression of peroxisome proliferator activated receptor-γ and CCAAT/enhancer binding protein-α, genes playing a central role in adipogenesis, was greatly suppressed. PGE 2 appears to be ineffective to a subclass of insulin target genes such as hexokinase 2 and phosphofructokinase. Similar responses were produced in the differentiation-associated genes upon AE1-329 treatment. These results suggest that PGE 2 inhibits a crucial step of the adipocyte differentiation process by acting on the EP4 receptor in 3T3-L1 cells

  3. Enhanced Y1-receptor-mediated vasoconstrictive action of neuropeptide Y (NPY) in superior mesenteric arteries in portal hypertension.

    Science.gov (United States)

    Wiest, Reiner; Jurzik, Lars; Moleda, Lukas; Froh, Matthias; Schnabl, Bernd; von Hörsten, Stephan; Schölmerich, Juergen; Straub, Rainer H

    2006-03-01

    Vascular hyporeactivity to catecholamines contributes to arterial vasodilation and hemodynamic dysregulation in portal hypertension. Neuropeptide Y (NPY) is a sympathetic neurotransmitter facilitating adrenergic vasoconstriction via Y1-receptors on the vascular smooth muscle. Therefore, we investigated its role for vascular reactivity in the superior mesenteric artery (SMA) of portal vein ligated (PVL) and sham operated rats. In vitro perfused SMA vascular beds of rats were tested for the cumulative dose-response to NPY dependent on the presence and level of alpha1-adrenergic vascular tone (methoxamine MT: 0.3-10 microM). Moreover, the effect of NPY (50 nM) on vascular responsiveness to alpha1-adrenergic stimulation (MT: 0.3-300 microM) was evaluated. Y1-receptor function was tested by Y1-selective inhibition using BIBP-3226 (1 microM). NPY dose-dependently and endothelium-independently enhanced MT-pre-constriction in SMA. This potentiation was increasingly effective with increasing adrenergic pre-stimulation and being more pronounced in PVL rats as compared to sham rats at high MT concentrations. NPY enhanced vascular contractility only in PVL rats correcting the adrenergic vascular hyporeactivity. Y1-receptor inhibition completely abolished NPY-evoked vasoconstrictive effects. NPY endothelium-independently potentiates adrenergic vasoconstriction via Y1-receptors being more pronounced in portal hypertension improving mesenteric vascular contractility and thereby correcting the splanchnic vascular hyporeactivity. This makes NPY a superior vasoconstrictor counterbalancing arterial vasodilation in portal hypertension.

  4. Sigma-1 receptor mediates cocaine-induced transcriptional regulation by recruiting chromatin-remodeling factors at the nuclear envelope.

    Science.gov (United States)

    Tsai, Shang-Yi A; Chuang, Jian-Ying; Tsai, Meng-Shan; Wang, Xiao-Fei; Xi, Zheng-Xiong; Hung, Jan-Jong; Chang, Wen-Chang; Bonci, Antonello; Su, Tsung-Ping

    2015-11-24

    The sigma-1 receptor (Sig-1R) chaperone at the endoplasmic reticulum (ER) plays important roles in cellular regulation. Here we found a new function of Sig-1R, in that it translocates from the ER to the nuclear envelope (NE) to recruit chromatin-remodeling molecules and regulate the gene transcription thereof. Sig-1Rs mainly reside at the ER-mitochondrion interface. However, on stimulation by agonists such as cocaine, Sig-1Rs translocate from ER to the NE, where Sig-1Rs bind NE protein emerin and recruit chromatin-remodeling molecules, including lamin A/C, barrier-to-autointegration factor (BAF), and histone deacetylase (HDAC), to form a complex with the gene repressor specific protein 3 (Sp3). Knockdown of Sig-1Rs attenuates the complex formation. Cocaine was found to suppress the gene expression of monoamine oxidase B (MAOB) in the brain of wild-type but not Sig-1R knockout mouse. A single dose of cocaine (20 mg/kg) in rats suppresses the level of MAOB at nuclear accumbens without affecting the level of dopamine transporter. Daily injections of cocaine in rats caused behavioral sensitization. Withdrawal from cocaine in cocaine-sensitized rats induced an apparent time-dependent rebound of the MAOB protein level to about 200% over control on day 14 after withdrawal. Treatment of cocaine-withdrawn rats with the MAOB inhibitor deprenyl completely alleviated the behavioral sensitization to cocaine. Our results demonstrate a role of Sig-1R in transcriptional regulation and suggest cocaine may work through this newly discovered genomic action to achieve its addictive action. Results also suggest the MAOB inhibitor deprenyl as a therapeutic agent to block certain actions of cocaine during withdrawal.

  5. Receptor-mediated targeting of 67Ga-Deferoxamine-Folate to folate-receptor-positive human kb tumor xenografts

    International Nuclear Information System (INIS)

    Mathias, Carla J.; Wang, Susan; Low, Philip S.; Waters, David J.; Green, Mark A.

    1999-01-01

    The radiochemical synthesis and stability of 67 Ga-deferoxamine-folate ([ 67 Ga]Ga-DF-Folate) were examined as a function of DF-Folate concentration. Optimal labeling occurred at DF-Folate concentrations ≥2.5 μg/mL. To define the possible biological significance of variations in product formulation, the biodistribution of [ 67 Ga]Ga-DF-Folate was examined as a function of administered deferoxamine-folate dose in an athymic mouse KB tumor model. The folate-receptor-positive KB tumors were found to concentrate the 67 Ga radiolabel in a dose-dependent fashion, consistent with saturable involvement of the folate receptor in mediating tumor accumulation of the radiopharmaceutical

  6. Degradation of Epidermal Growth Factor Receptor Mediates Dasatinib-Induced Apoptosis in Head and Neck Squamous Cell Carcinoma Cells

    Directory of Open Access Journals (Sweden)

    Yu-Chin Lin

    2012-06-01

    Full Text Available Epidermal growth factor receptor (EGFR is an important oncoprotein that promotes cell growth and proliferation. Dasatinib, a bcr-abl inhibitor, has been approved clinically for the treatment of chronic myeloid leukemia and demonstrated to be effective against solid tumors in vitro through Src inhibition. Here, we disclose that EGFR degradation mediated dasatinib-induced apoptosis in head and neck squamous cell carcinoma (HNSCC cells. HNSCC cells, including Ca9-22, FaDu, HSC3, SAS, SCC-25, and UMSCC1, were treated with dasatinib, and cell viability, apoptosis, and underlying signal transduction were evaluated. Dasatinib exhibited differential sensitivities against HNSCC cells. Growth inhibition and apoptosis were correlated with its inhibition on Akt, Erk, and Bcl-2, irrespective of Src inhibition. Accordingly, we found that down-regulation of EGFR was a determinant of dasatinib sensitivity. Lysosome inhibitor reversed dasatinib-induced EGFR down-regulation, and c-cbl activity was increased by dasatinib, indicating that dasatinib-induced EGFR down-regulation might be through c-cbl-mediated lysosome degradation. Increased EGFR activation by ligand administration rescued cells from dasatinib-induced apoptosis, whereas inhibition of EGFR enhanced its apoptotic effect. Estrogen receptor α (ERα was demonstrated to play a role in Bcl-2 expression, and dasatinib inhibited ERα at the pretranslational level. ERα was associated with EGFR in dasatinib-treated HNSCC cells. Furthermore, the xenograft model showed that dasatinib inhibited HSC3 tumor growth through in vivo down-regulation of EGFR and ERα. In conclusion, degradation of EGFR is a novel mechanism responsible for dasatinib-induced apoptosis in HNSCC cells.

  7. Adenosine A1 receptor-mediated inhibition of in vitro prolactin secretion from the rat anterior pituitary

    Directory of Open Access Journals (Sweden)

    D.L.W. Picanço-Diniz

    2006-11-01

    Full Text Available In previous studies, we demonstrated biphasic purinergic effects on prolactin (PRL secretion stimulated by an adenosine A2 agonist. In the present study, we investigated the role of the activation of adenosine A1 receptors by (R-N6-(2-phenylisopropyladenosine (R-PIA at the pituitary level in in vitro PRL secretion. Hemipituitaries (one per cuvette in five replicates from adult male rats were incubated. Administration of R-PIA (0.001, 0.01, 0.1, 1, and 10 µM induced a reduction of PRL secretion into the medium in a U-shaped dose-response curve. The maximal reduction was obtained with 0.1 µM R-PIA (mean ± SEM, 36.01 ± 5.53 ng/mg tissue weight (t.w. treatment compared to control (264.56 ± 15.46 ng/mg t.w.. R-PIA inhibition (0.01 µM = 141.97 ± 15.79 vs control = 244.77 ± 13.79 ng/mg t.w. of PRL release was blocked by 1 µM cyclopentyltheophylline, a specific A1 receptor antagonist (1 µM = 212.360 ± 26.560 ng/mg t.w., whereas cyclopentyltheophylline alone (0.01, 0.1, 1 µM had no effect. R-PIA (0.001, 0.01, 0.1, 1 µM produced inhibition of PRL secretion stimulated by both phospholipase C (0.5 IU/mL; 977.44 ± 76.17 ng/mg t.w. and dibutyryl cAMP (1 mM; 415.93 ± 37.66 ng/mg t.w. with nadir established at the dose of 0.1 µM (225.55 ± 71.42 and 201.9 ± 19.08 ng/mg t.w., respectively. Similarly, R-PIA (0.01 µM decreased (242.00 ± 24.00 ng/mg t.w. the PRL secretion stimulated by cholera toxin (0.5 mg/mL; 1050.00 ± 70.00 ng/mg t.w.. In contrast, R-PIA had no effect (468.00 ± 34.00 ng/mg t.w. on PRL secretion stimulation by pertussis toxin (0.5 mg/mL; 430.00 ± 26.00 ng/mg t.w.. These results suggest that inhibition of PRL secretion after A1 receptor activation by R-PIA is mediated by a Gi protein-dependent mechanism.

  8. The epidermal growth factor-like domains of the human EMR2 receptor mediate cell attachment through chondroitin sulfate glycosaminoglycans

    NARCIS (Netherlands)

    Stacey, Martin; Chang, Gin-Wen; Davies, John Q.; Kwakkenbos, Mark J.; Sanderson, Ralph D.; Hamann, Jörg; Gordon, Siamon; Lin, Hsi-Hsien

    2003-01-01

    Using multivalent protein probes, an evolutionarily conserved endogenous ligand for EMR2, a human myeloid cell-restricted EGF-TM7 receptor, was identified on the surface of a number of adherent cell lines. In addition, in situ staining of the ligand has revealed specific in vivo patterns consistent

  9. Multiple injections of electroporated autologous T cells expressing a chimeric antigen receptor mediate regression of human disseminated tumor.

    Science.gov (United States)

    Zhao, Yangbing; Moon, Edmund; Carpenito, Carmine; Paulos, Chrystal M; Liu, Xiaojun; Brennan, Andrea L; Chew, Anne; Carroll, Richard G; Scholler, John; Levine, Bruce L; Albelda, Steven M; June, Carl H

    2010-11-15

    Redirecting T lymphocyte antigen specificity by gene transfer can provide large numbers of tumor-reactive T lymphocytes for adoptive immunotherapy. However, safety concerns associated with viral vector production have limited clinical application of T cells expressing chimeric antigen receptors (CAR). T lymphocytes can be gene modified by RNA electroporation without integration-associated safety concerns. To establish a safe platform for adoptive immunotherapy, we first optimized the vector backbone for RNA in vitro transcription to achieve high-level transgene expression. CAR expression and function of RNA-electroporated T cells could be detected up to a week after electroporation. Multiple injections of RNA CAR-electroporated T cells mediated regression of large vascularized flank mesothelioma tumors in NOD/scid/γc(-/-) mice. Dramatic tumor reduction also occurred when the preexisting intraperitoneal human-derived tumors, which had been growing in vivo for >50 days, were treated by multiple injections of autologous human T cells electroporated with anti-mesothelin CAR mRNA. This is the first report using matched patient tumor and lymphocytes showing that autologous T cells from cancer patients can be engineered to provide an effective therapy for a disseminated tumor in a robust preclinical model. Multiple injections of RNA-engineered T cells are a novel approach for adoptive cell transfer, providing flexible platform for the treatment of cancer that may complement the use of retroviral and lentiviral engineered T cells. This approach may increase the therapeutic index of T cells engineered to express powerful activation domains without the associated safety concerns of integrating viral vectors. Copyright © 2010 AACR.

  10. Diosgenin promotes oligodendrocyte progenitor cell differentiation through estrogen receptor-mediated ERK1/2 activation to accelerate remyelination.

    Science.gov (United States)

    Xiao, Lin; Guo, Dazhi; Hu, Chun; Shen, Weiran; Shan, Lei; Li, Cui; Liu, Xiuyun; Yang, Wenjing; Zhang, Weidong; He, Cheng

    2012-07-01

    Differentiation of oligodendrocyte progenitor cells (OPCs) into mature oligodendrocytes is a prerequisite for remyelination after demyelination, and impairment of this process is suggested to be a major reason for remyelination failure. Diosgenin, a plant-derived steroid, has been implicated for therapeutic use in many diseases, but little is known about its effect on the central nervous system. In this study, using a purified rat OPC culture model, we show that diosgenin significantly and specifically promotes OPC differentiation without affecting the viability, proliferation, or migration of OPC. Interestingly, the effect of diosgenin can be blocked by estrogen receptor (ER) antagonist ICI 182780 but not by glucocorticoid and progesterone receptor antagonist RU38486, nor by mineralocorticoid receptor antagonist spirolactone. Moreover, it is revealed that both ER-alpha and ER-beta are expressed in OPC, and diosgenin can activate the extracellular signal-regulated kinase 1/2 (ERK1/2) in OPC via ER. The pro-differentiation effect of diosgenin can also be obstructed by the ERK inhibitor PD98059. Furthermore, in the cuprizone-induced demyelination model, it is demonstrated that diosgenin administration significantly accelerates/enhances remyelination as detected by Luxol fast blue stain, MBP immunohistochemistry and real time RT-PCR. Diosgenin also increases the number of mature oligodendrocytes in the corpus callosum while it does not affect the number of OPCs. Taking together, our results suggest that diosgenin promotes the differentiation of OPC into mature oligodendrocyte through an ER-mediated ERK1/2 activation pathway to accelerate remyelination, which implicates a novel therapeutic usage of this steroidal natural product in demyelinating diseases such as multiple sclerosis (MS). Copyright © 2012 Wiley Periodicals, Inc.

  11. Neurokinin 1 Receptor Mediates Membrane Blebbing and Sheer Stress-Induced Microparticle Formation in HEK293 Cells

    Science.gov (United States)

    Chen, Panpan; Douglas, Steven D.; Meshki, John; Tuluc, Florin

    2012-01-01

    Cell-derived microparticles participate in intercellular communication similar to the classical messenger systems of small and macro-molecules that bind to specialized membrane receptors. Microparticles have been implicated in the regulation of a variety of complex physiopathologic processes, such as thrombosis, the control of innate and adaptive immunity, and cancer. The neurokinin 1 receptor (NK1R) is a Gq-coupled receptor present on the membrane of a variety of tissues, including neurons in the central and peripheral nervous system, immune cells, endocrine and exocrine glands, and smooth muscle. The endogenous agonist of NK1R is the undecapeptide substance P (SP). We have previously described intracellular signaling mechanisms that regulate NK1R-mediated rapid cell shape changes in HEK293 cells and U373MG cells. In the present study, we show that the activation of NK1R in HEK293 cells, but not in U373MG cells, leads to formation of sheer-stress induced microparticles that stain positive with the membrane-selective fluorescent dye FM 2–10. SP-induced microparticle formation is independent of elevated intracellular calcium concentrations and activation of NK1R present on HEK293-derived microparticles triggers detectable calcium increase in SP-induced microparticles. The ROCK inhibitor Y27632 and the dynamin inhibitor dynasore inhibited membrane blebbing and microparticle formation in HEK293 cells, strongly suggesting that microparticle formation in this cell type is dependent on membrane blebbing. PMID:23024816

  12. Neurokinin 1 receptor mediates membrane blebbing and sheer stress-induced microparticle formation in HEK293 cells.

    Directory of Open Access Journals (Sweden)

    Panpan Chen

    Full Text Available Cell-derived microparticles participate in intercellular communication similar to the classical messenger systems of small and macro-molecules that bind to specialized membrane receptors. Microparticles have been implicated in the regulation of a variety of complex physiopathologic processes, such as thrombosis, the control of innate and adaptive immunity, and cancer. The neurokinin 1 receptor (NK1R is a Gq-coupled receptor present on the membrane of a variety of tissues, including neurons in the central and peripheral nervous system, immune cells, endocrine and exocrine glands, and smooth muscle. The endogenous agonist of NK1R is the undecapeptide substance P (SP. We have previously described intracellular signaling mechanisms that regulate NK1R-mediated rapid cell shape changes in HEK293 cells and U373MG cells. In the present study, we show that the activation of NK1R in HEK293 cells, but not in U373MG cells, leads to formation of sheer-stress induced microparticles that stain positive with the membrane-selective fluorescent dye FM 2-10. SP-induced microparticle formation is independent of elevated intracellular calcium concentrations and activation of NK1R present on HEK293-derived microparticles triggers detectable calcium increase in SP-induced microparticles. The ROCK inhibitor Y27632 and the dynamin inhibitor dynasore inhibited membrane blebbing and microparticle formation in HEK293 cells, strongly suggesting that microparticle formation in this cell type is dependent on membrane blebbing.

  13. Endothelin receptor mediated Ca(2+) signaling in coronary arteries after experimentally induced ischemia/reperfusion injury in rat

    DEFF Research Database (Denmark)

    Kristiansen, Sarah Brøgger; Haanes, Kristian A.; Sheykhzade, Majid

    2017-01-01

    a phenotypical shift, which includes increased evoked ETB induced contraction in the smooth muscle cell, and also a higher basal tone development which both are dependent on Ca(2+) influx through VGCCs. This is combined with alterations in the ETA calcium handling, which has a stronger dependence on Ca(2...

  14. Endothelial Mineralocorticoid Receptor Mediates Parenchymal Arteriole and Posterior Cerebral Artery Remodeling During Angiotensin II-Induced Hypertension.

    Science.gov (United States)

    Diaz-Otero, Janice M; Fisher, Courtney; Downs, Kelsey; Moss, M Elizabeth; Jaffe, Iris Z; Jackson, William F; Dorrance, Anne M

    2017-12-01

    The brain is highly susceptible to injury caused by hypertension because the increased blood pressure causes artery remodeling that can limit cerebral perfusion. Mineralocorticoid receptor (MR) antagonism prevents hypertensive cerebral artery remodeling, but the vascular cell types involved have not been defined. In the periphery, the endothelial MR mediates hypertension-induced vascular injury, but cerebral and peripheral arteries are anatomically distinct; thus, these findings cannot be extrapolated to the brain. The parenchymal arterioles determine cerebrovascular resistance. Determining the effects of hypertension and MR signaling on these arterioles could lead to a better understanding of cerebral small vessel disease. We hypothesized that endothelial MR signaling mediates inward cerebral artery remodeling and reduced cerebral perfusion during angiotensin II (AngII) hypertension. The biomechanics of the parenchymal arterioles and posterior cerebral arteries were studied in male C57Bl/6 and endothelial cell-specific MR knockout mice and their appropriate controls using pressure myography. AngII increased plasma aldosterone and decreased cerebral perfusion in C57Bl/6 and MR-intact littermates. Endothelial cell MR deletion improved cerebral perfusion in AngII-treated mice. AngII hypertension resulted in inward hypotrophic remodeling; this was prevented by MR antagonism and endothelial MR deletion. Our studies suggest that endothelial cell MR mediates hypertensive remodeling in the cerebral microcirculation and large pial arteries. AngII-induced inward remodeling of cerebral arteries and arterioles was associated with a reduction in cerebral perfusion that could worsen the outcome of stroke or contribute to vascular dementia. © 2017 American Heart Association, Inc.

  15. A novel ecdysone receptor mediates steroid-regulated developmental events during the mid-third instar of Drosophila.

    Directory of Open Access Journals (Sweden)

    Benjamin F B Costantino

    2008-06-01

    Full Text Available The larval salivary gland of Drosophila melanogaster synthesizes and secretes glue glycoproteins that cement developing animals to a solid surface during metamorphosis. The steroid hormone 20-hydroxyecdysone (20E is an essential signaling molecule that modulates most of the physiological functions of the larval gland. At the end of larval development, it is known that 20E--signaling through a nuclear receptor heterodimer consisting of EcR and USP--induces the early and late puffing cascade of the polytene chromosomes and causes the exocytosis of stored glue granules into the lumen of the gland. It has also been reported that an earlier pulse of hormone induces the temporally and spatially specific transcriptional activation of the glue genes; however, the receptor responsible for triggering this response has not been characterized. Here we show that the coordinated expression of the glue genes midway through the third instar is mediated by 20E acting to induce genes of the Broad Complex (BRC through a receptor that is not an EcR/USP heterodimer. This result is novel because it demonstrates for the first time that at least some 20E-mediated, mid-larval, developmental responses are controlled by an uncharacterized receptor that does not contain an RXR-like component.

  16. Histone H4 Lys 20 methyltransferase SET8 promotes androgen receptor-mediated transcription activation in prostate cancer

    Energy Technology Data Exchange (ETDEWEB)

    Yao, Lushuai [Laboratory of Genome Variations and Precision Bio-Medicine, Beijing Institute of Genomics, Chinese Academy of Sciences, Beijing 100101 (China); University of Chinese Academy of Sciences, Beijing 100049 (China); Li, Yanyan; Du, Fengxia [Laboratory of Genome Variations and Precision Bio-Medicine, Beijing Institute of Genomics, Chinese Academy of Sciences, Beijing 100101 (China); Han, Xiao [Laboratory of Genome Variations and Precision Bio-Medicine, Beijing Institute of Genomics, Chinese Academy of Sciences, Beijing 100101 (China); University of Chinese Academy of Sciences, Beijing 100049 (China); Li, Xiaohua [Laboratory of Genome Variations and Precision Bio-Medicine, Beijing Institute of Genomics, Chinese Academy of Sciences, Beijing 100101 (China); Niu, Yuanjie [Chawnshang Chang Sex Hormone Research Center, Tianjin Institute of Urology, Tianjin Medical University, Tianjin 300070 (China); Ren, Shancheng, E-mail: renshancheng@gmail.com [Department of Urology, Shanghai Changhai Hospital, Second Military Medical University, Shanghai 200433 (China); Sun, Yingli, E-mail: sunyl@big.ac.cn [Laboratory of Genome Variations and Precision Bio-Medicine, Beijing Institute of Genomics, Chinese Academy of Sciences, Beijing 100101 (China)

    2014-07-18

    Highlights: • Dihydrotestosterone stimulates H4K20me1 enrichment at the PSA promoter. • SET8 promotes AR-mediated transcription activation. • SET8 interacts with AR and promotes cell proliferation. - Abstract: Histone methylation status in different lysine residues has an important role in transcription regulation. The effect of H4K20 monomethylation (H4K20me1) on androgen receptor (AR)-mediated gene transcription remains unclear. Here we show that AR agonist stimulates the enrichment of H4K20me1 and SET8 at the promoter of AR target gene PSA in an AR dependent manner. Furthermore, SET8 is crucial for the transcription activation of PSA. Co-immunoprecipitation analyses demonstrate that SET8 interacts with AR. Therefore, we conclude that SET8 is involved in AR-mediated transcription activation, possibly through its interaction with AR and H4K20me1 modification.

  17. Timing is critical for effective glucocorticoid receptor mediated repression of the cAMP-induced CRH gene.

    Directory of Open Access Journals (Sweden)

    Siem van der Laan

    Full Text Available Glucocorticoid negative feedback of the hypothalamus-pituitary-adrenal axis is mediated in part by direct repression of gene transcription in glucocorticoid receptor (GR expressing cells. We have investigated the cross talk between the two main signaling pathways involved in activation and repression of corticotrophin releasing hormone (CRH mRNA expression: cyclic AMP (cAMP and GR. We report that in the At-T20 cell-line the glucocorticoid-mediated repression of the cAMP-induced human CRH proximal promoter activity depends on the relative timing of activation of both signaling pathways. Activation of the GR prior to or in conjunction with cAMP signaling results in an effective repression of the cAMP-induced transcription of the CRH gene. In contrast, activation of the GR 10 minutes after onset of cAMP treatment, results in a significant loss of GR-mediated repression. In addition, translocation of ligand-activated GR to the nucleus was found as early as 10 minutes after glucocorticoid treatment. Interestingly, while both signaling cascades counteract each other on the CRH proximal promoter, they synergize on a synthetic promoter containing 'positive' response elements. Since the order of activation of both signaling pathways may vary considerably in vivo, we conclude that a critical time-window exists for effective repression of the CRH gene by glucocorticoids.

  18. Lymphotropic Virions Affect Chemokine Receptor-Mediated Neural Signaling and Apoptosis: Implications for Human Immunodeficiency Virus Type 1-Associated Dementia

    Science.gov (United States)

    Zheng, Jialin; Ghorpade, Anuja; Niemann, Douglas; Cotter, Robin L.; Thylin, Michael R.; Epstein, Leon; Swartz, Jennifer M.; Shepard, Robin B.; Liu, Xiaojuan; Nukuna, Adeline; Gendelman, Howard E.

    1999-01-01

    Chemokine receptors pivotal for human immunodeficiency virus type 1 (HIV-1) infection in lymphocytes and macrophages (CCR3, CCR5, and CXCR4) are expressed on neural cells (microglia, astrocytes, and/or neurons). It is these cells which are damaged during progressive HIV-1 infection of the central nervous system. We theorize that viral coreceptors could effect neural cell damage during HIV-1-associated dementia (HAD) without simultaneously affecting viral replication. To these ends, we studied the ability of diverse viral strains to affect intracellular signaling and apoptosis of neurons, astrocytes, and monocyte-derived macrophages. Inhibition of cyclic AMP, activation of inositol 1,4,5-trisphosphate, and apoptosis were induced by diverse HIV-1 strains, principally in neurons. Virions from T-cell-tropic (T-tropic) strains (MN, IIIB, and Lai) produced the most significant alterations in signaling of neurons and astrocytes. The HIV-1 envelope glycoprotein, gp120, induced markedly less neural damage than purified virions. Macrophage-tropic (M-tropic) strains (ADA, JR-FL, Bal, MS-CSF, and DJV) produced the least neural damage, while 89.6, a dual-tropic HIV-1 strain, elicited intermediate neural cell damage. All T-tropic strain-mediated neuronal impairments were blocked by the CXCR4 antibody, 12G5. In contrast, the M-tropic strains were only partially blocked by 12G5. CXCR4-mediated neuronal apoptosis was confirmed in pure populations of rat cerebellar granule neurons and was blocked by HA1004, an inhibitor of calcium/calmodulin-dependent protein kinase II, protein kinase A, and protein kinase C. Taken together, these results suggest that progeny HIV-1 virions can influence neuronal signal transduction and apoptosis. This process occurs, in part, through CXCR4 and is independent of CD4 binding. T-tropic viruses that traffic in and out of the brain during progressive HIV-1 disease may play an important role in HAD neuropathogenesis. PMID:10482576

  19. The role of calcium in the receptor mediated control of potassium permeability in the rat lacrimal gland

    International Nuclear Information System (INIS)

    Parod, R.J.; Putney, J.W. Jr.

    1978-01-01

    In the presence of extracellular Ca, adrenaline stimulated by a large increase in the rate of K ( 86 Rb) release from rat lacrimal slices, followed by a lower, more sustained rate. In the absence of extracellular Ca, adrenaline elicited only a transient release of 86 Rb. The artificial introduction of Ca into the cytosol by the ionophore A-23187 could also initiate the release of 86 Rb. In a zero-Ca medium, if either adrenaline or carbachol produced a transient release of 86 Rb, the tissue could not respond to the other agonist with a transient release unless Ca was momentarily reintroduced to the medium. If Ca was present in a limiting concentration, the Ca-dependent rate of 86 Rb release elicited from a lacrimal slice exposed simultaneously to carbachol and adrenaline was not significantly different from the release seen with carbachol alone. It is concluded that the agonist-induced release of K from the lacrimal gland consists of both a Ca-independent phase which is initiated by the release of a limited pool of Ca, and a Ca-dependent phase which is mediated by the influx of extracellular Ca. It is also concluded that both α-adrenergic and muscarinic receptors occupation activate a common, post-receptor mechanism which may be responsible for both phases of K release. (author)

  20. Receptor-Mediated Melanoma Targeting with Radiolabeled α-Melanocyte-Stimulating Hormone: Relevance of the Net Charge of the Ligand

    Directory of Open Access Journals (Sweden)

    Alex N. Eberle

    2017-04-01

    Full Text Available A majority of melanotic and amelanotic melanomas overexpress melanocortin type 1 receptors (MC1Rs for α-melanocyte-stimulating hormone. Radiolabeled linear or cyclic analogs of α-MSH have a great potential as diagnostic or therapeutic tools for the management of malignant melanoma. Compounds such as [111In]DOTA-NAP-amide exhibit high affinity for the MC1R in vitro, good tumor uptake in vivo, but they may suffer from relatively high kidney uptake and retention in vivo. We have shown previously that the introduction of negative charges into radiolabeled DOTA-NAP-amide peptide analogs may enhance their excretion and reduce kidney retention. To address the question of where to place negative charges within the ligand, we have extended these studies by designing two novel peptides, Ac-Nle-Asp-His-d-Phe-Arg-Trp-Gly-Lys(DOTA-d-Asp-d-Asp-OH (DOTA-NAP-d-Asp-d-Asp with three negative charges at the C-terminal end (overall net charge of the molecule −2 and DOTA-Gly-Tyr(P-Nle-Asp-His-d-Phe-Arg-Trp-NH2 (DOTA-Phospho-MSH2-9 with two negative charges in the N-terminal region (net charge −1. The former peptide showed markedly reduced receptor affinity and biological activity by >10-fold compared to DOTA-NAP-amide as reference compound, and the latter peptide displayed similar bioactivity and receptor affinity as the reference compound. The uptake by melanoma tumor tissue of [111In]DOTA-Phospho-MSH2-9 was 7.33 ± 0.47 %ID/g 4 h after injection, i.e., almost equally high as with [111In]DOTA-NAP-amide. The kidney retention was 2.68 ± 0.18 %ID/g 4 h after injection and hence 44% lower than that of [111In]DOTA-NAP-amide. Over an observation period from 4 to 48 h, the tumor-to-kidney ratio of [111In]DOTA-Phospho-MSH2-9 was 35% more favorable than that of the reference compound. In a comparison of DOTA-NAP-d-Asp-d-Asp, DOTA-Phospho-MSH2-9 and DOTA-NAP-amide with five previously published analogs of DOTA-NAP-amide that altogether cover a range of peptides with an overall net charge between +2 and −2, we now demonstrate that a net charge of −1, with the extra negative charges preferably placed in the N-terminal region, has led to the lowest kidney uptake and retention. Charges of +2 or −2 markedly increased kidney uptake and retention. In conclusion, the novel DOTA-Phospho-MSH2-9 may represent a new lead compound for negatively charged linear MC1R ligands that can be further developed into a clinically relevant melanoma targeting radiopeptide.

  1. Genetically designed biomolecular capping system for mesoporous silica nanoparticles enables receptor-mediated cell uptake and controlled drug release

    Czech Academy of Sciences Publication Activity Database

    Datz, S.; Argyo, C.; Gattner, M.; Weiss, V.; Brunner, K.; Bretzler, J.; von Schirnding, C.; Torrano, A. A.; Spada, F.; Vrábel, Milan; Engelke, H.; Bräuchle, C.; Carell, T.; Bein, T.

    2016-01-01

    Roč. 8, č. 15 (2016), s. 8101-8110 ISSN 2040-3364 Institutional support: RVO:61388963 Keywords : responsive controlled release * Diels-Alder reactions * human carbonic anhydrase Subject RIV: CF - Physical ; Theoretical Chemistry Impact factor: 7.367, year: 2016 http://pubs.rsc.org/en/content/articlehtml/2016/nr/c5nr08163g

  2. Glycine and GABAA receptors mediate tonic and phasic inhibitory processes that contribute to prepulse inhibition in the goldfish startle network

    Directory of Open Access Journals (Sweden)

    Paul C.P. Curtin

    2015-03-01

    Full Text Available Prepulse inhibition (PPI is understood as an inhibitory process that attenuates sensory flow during early stages (20-1000ms of information processing. Here, we applied in vivo electrophysiology and pharmacology to determine if prepulse inhibition (PPI is mediated by glycine receptors (GlyRs and/or GABAA receptors (GABAARs in the goldfish auditory startle circuit. Specifically, we used selective antagonists to dissect the contributions of target receptors on sound-evoked postsynaptic potentials (PSPs recorded in the neurons that initiate startle, the Mauthner-cells (M-cell. We found that strychnine, a GlyR antagonist, disrupted a fast-activated (5 ms and rapidly (< 50ms decaying (feed-forward inhibitory process that disrupts PPI at 20 ms prepulse/pulse inter-stimulus intervals (ISI. Additionally we observed increases of the evoked postsynaptic potential (PSP peak amplitude (+87.43 ± 21.53%; N=9 and duration (+204 ± 48.91%, N=9. In contrast, treatment with bicuculline, a GABAAR antagonist, caused a general reduction in PPI across all tested ISIs (20-500 ms, essentially eliminating PPI at ISIs from 20-100 ms. Bicuculline also increased PSP peak amplitude (+133.8 ± 10.3%, N=5 and PSP duration (+284.95 ± 65.64%, N=5. Treatment with either antagonist also tonically increased post-synaptic excitability in the M-cells, reflected by an increase in the magnitude of antidromically-evoked action potentials (APs by 15.07 ± 3.21%, N=7 and 16.23 ± 7.08%, N=5 for strychnine and bicuculline, respectively. These results suggest that GABAARs and GlyRs are functionally segregated to short- and longer-lasting sound-evoked (phasic inhibitory processes that contribute to PPI, with the mediation of tonic inhibition by both receptor systems being critical for gain control within the M-cell startle circuit.

  3. P2X7 receptor-mediated PARP1 activity regulates astroglial death in the rat hippocampus following status epilepticus

    Directory of Open Access Journals (Sweden)

    Ji Yang eKim

    2015-09-01

    Full Text Available Poly(ADP-ribose polymerase-1 (PARP1 plays a regulatory role in apoptosis, necrosis, and other cellular processes after injury. Recently, we revealed that PARP1 regulates the differential neuronal/astroglial responses to pilocarpine-induced status epilepticus (SE in the distinct brain regions. In addition, P2X7 receptor (P2X7R, an ATP-gated ion channel, activation accelerates astroglial apoptosis, while it attenuates clasmatodendrosis (lysosome-derived autophagic astroglial death. Therefore, we investigated whether P2X7R regulates regional specific astroglial PARP1 expression/activation in response to SE. In the present study, P2X7R activation exacerbates SE-induced astroglial apoptosis, while P2X7R inhibition attenuates it accompanied by increasing PARP1 activity in the molecular layer of the dentate gyrus following SE. In the CA1 region, however, P2X7R inhibition deteriorates SE-induced clasmatodendrosis via PARP1 activation following SE. Taken together, our findings suggest that P2X7R function may affect SE-induced astroglial death by regulating PARP1 activation/expression in regional-specific manner. Therefore, the selective modulation of P2X7R-mediated PARP1 functions may be a considerable strategy for controls in various types of cell deaths.

  4. On the interaction of caffeine with nucleic acids. 3

    International Nuclear Information System (INIS)

    Fritzsche, H.; Petri, I.; Schuetz, H.; Weller, K.; Sedmera, P.; Lang, H.

    1980-01-01

    The authors have performed three experiments: 1) The self-association of both caffeine (Cf) and 5'-adenosine monophosphate (AMP) in aqueous solution has been re-investigated by 1 H NMR. 2) The interaction of Cf and AMP has been studied by 1 H NMR. 3) The interaction of Cf and poly(riboadenylate), (rA)sub(n), is indicated by a downfield shift of the H-8 line but an upfield shift of the H-2 line in the 1 H NMR spectra of (rA)sub(n). (Auth.)

  5. Lysophosphatidic acid inhibition of the accumulation of Pseudomonas aeruginosa PAO1 alginate, pyoverdin, elastase and LasA

    DEFF Research Database (Denmark)

    Laux, D.C.; Corson, J.M.; Givskov, Michael Christian

    2002-01-01

    . In the present study, a lysophospholipid, 1-paimitoyl-2-hydroxy-sn-glycero-3-phosphate [also called monopalmitoylphosphatidic acid (MPPA)], which accumulates in inflammatory exudates, was shown to inhibit the extracellular accumulation of P. aeruginosa PAO1 alginate, elastase, LasA protease and the siderophore...

  6. Alpha5 nicotinic acetylcholine receptor mediates nicotine-induced HIF-1α and VEGF expression in non-small cell lung cancer

    Energy Technology Data Exchange (ETDEWEB)

    Ma, Xiaoli; Jia, Yanfei; Zu, Shanshan [Central Laboratory, Jinan Central Hospital Affiliated to Shandong University, Jinan 250013 (China); Li, Ruisheng [Institute of Infectious Diseases, 302 Military Hospital, Beijing 100039 (China); Jia, Ying; Zhao, Yun; Xiao, Dongjie [Central Laboratory, Jinan Central Hospital Affiliated to Shandong University, Jinan 250013 (China); Dang, Ningning [Department of Dermatology, Jinan Central Hospital Affiliated to Shandong University, Jinan 250013 (China); Wang, Yunshan [Central Laboratory, Jinan Central Hospital Affiliated to Shandong University, Jinan 250013 (China)

    2014-07-15

    By binding to nicotinic acetylcholine receptors (nAChRs), nicotine induces the proliferation and apoptosis of non-small cell lung cancer (NSCLC). Previous studies have indicated that α5-nAChR is highly associated with lung cancer risk and nicotine dependence. However, the mechanisms through which α5-nAChRs may influence lung carcinogenesis are far from clear. In the present study, we investigated the roles of α5-nAChR in the nicotine-induced expression of hypoxia-inducible factor-1α (HIF-1α) and vascular endothelial growth factor (VEGF). Immunohistochemistry was used to detect the expression of α5-nAChR and HIF-1α in 60 specimens of lung cancer and para-carcinoma tissue. The correlations between the expression levels of α5-nAChR and HIF-1α and other clinicopathological data were analyzed. In a cell line that highly expressed α5-nAChR, the loss of α5-nAChR function by siRNA was used to study whether α5-nAChR is involved in the nicotine-induced expression of HIF-1α and VEGF through the activation of the ERK1/2 and PI3K/Akt signaling pathways. Cell growth was detected using the cell counting kit-8 (CCK-8). α5-nAChR (78.3%) and HIF-1α (88.3%) were both overexpressed in NSCLC, and their expression levels were found to be correlated with each other (P < 0.05). In the A549 cell line, α5-nAChR and HIF-1α were found to be expressed under normal conditions, and their expression levels were significantly increased in response to nicotine treatment. The silencing of α5-nAChR significantly inhibited the nicotine-induced cell proliferation compared with the control group and attenuated the nicotine-induced upregulation of HIF-1α and VEGF, and these effects required the cooperation of the ERK1/2 and PI3K/Akt signaling pathways. These results show that the α5-nAChR/HIF-1α/VEGF axis is involved in nicotine-induced tumor cell proliferation, which suggests that α5-nAChR may serve as a potential anticancer target in nicotine-associated lung cancer. - Highlights: • α5-nAChR and HIF-1α over-expressed in NSCLC and correlated with each other. • Nicotine induces cell proliferation and over-expressions of α5-nAChR, HIF-1α and VEGF in NSCLC. • Silence of α5-nAChR inhibited nicotine-induced A549 cell proliferation. • α5-nAChR involved in nicotine-induced HIF-1α and VEGF expression. • Down-regulation of α5-nAChR decreased the phosphorylation level of ERK1/2 and Akt.

  7. Elevated carbon dioxide blunts mammalian cAMP signaling dependent on inositol 1,4,5-triphosphate receptor-mediated Ca2+ release.

    Science.gov (United States)

    Cook, Zara C; Gray, Michael A; Cann, Martin J

    2012-07-27

    Elevated CO(2) is generally detrimental to animal cells, suggesting an interaction with core processes in cell biology. We demonstrate that elevated CO(2) blunts G protein-activated cAMP signaling. The effect of CO(2) is independent of changes in intracellular and extracellular pH, independent of the mechanism used to activate the cAMP signaling pathway, and is independent of cell context. A combination of pharmacological and genetic tools demonstrated that the effect of elevated CO(2) on cAMP levels required the activity of the IP(3) receptor. Consistent with these findings, CO(2) caused an increase in steady state cytoplasmic Ca(2+) concentrations not observed in the absence of the IP(3) receptor or under nonspecific acidotic conditions. We examined the well characterized cAMP-dependent inhibition of the isoform 3 Na(+)/H(+) antiporter (NHE3) to demonstrate a functional relevance for CO(2)-mediated reductions in cellular cAMP. Consistent with the cellular biochemistry, elevated CO(2) abrogated the inhibitory effect of cAMP on NHE3 function via an IP(3) receptor-dependent mechanism.

  8. Elevated Carbon Dioxide Blunts Mammalian cAMP Signaling Dependent on Inositol 1,4,5-Triphosphate Receptor-mediated Ca2+ Release*

    Science.gov (United States)

    Cook, Zara C.; Gray, Michael A.; Cann, Martin J.

    2012-01-01

    Elevated CO2 is generally detrimental to animal cells, suggesting an interaction with core processes in cell biology. We demonstrate that elevated CO2 blunts G protein-activated cAMP signaling. The effect of CO2 is independent of changes in intracellular and extracellular pH, independent of the mechanism used to activate the cAMP signaling pathway, and is independent of cell context. A combination of pharmacological and genetic tools demonstrated that the effect of elevated CO2 on cAMP levels required the activity of the IP3 receptor. Consistent with these findings, CO2 caused an increase in steady state cytoplasmic Ca2+ concentrations not observed in the absence of the IP3 receptor or under nonspecific acidotic conditions. We examined the well characterized cAMP-dependent inhibition of the isoform 3 Na+/H+ antiporter (NHE3) to demonstrate a functional relevance for CO2-mediated reductions in cellular cAMP. Consistent with the cellular biochemistry, elevated CO2 abrogated the inhibitory effect of cAMP on NHE3 function via an IP3 receptor-dependent mechanism. PMID:22654111

  9. Time-dependent effects of repeated THC treatment on dopamine D2/3 receptor-mediated signalling in midbrain and striatum.

    Science.gov (United States)

    Tournier, Benjamin B; Tsartsalis, Stergios; Dimiziani, Andrea; Millet, Philippe; Ginovart, Nathalie

    2016-09-15

    This study examined the time-course of alterations in levels and functional sensitivities of dopamine D2/3 receptors (D2/3R) during the course and up to 6 weeks following cessation of chronic treatment with Delta(9)-Tetrahydrocannabinol (THC) in rats. THC treatment led to an increase in D2/3R levels in striatum, as assessed using [(3)H]-(+)-PHNO, that was readily observable after one week of treatment, remained stably elevated during the subsequent 2 weeks of treatment, but fully reversed within 2 weeks of THC discontinuation. THC-induced D2/3R alterations were more pronounced and longer lasting in the dopamine cell body regions of the midbrain, wherein [(3)H]-(+)-PHNO binding was still elevated at 2 weeks but back to control values at 6 weeks after THC cessation. Parallel analyses of the psychomotor effects of pre- and post-synaptic doses of quinpirole also showed a pattern of D2/3R functional supersensitivity indicative of more rapid subsidence in striatum than in midbrain following drug cessation. These results indicate that chronic THC is associated with a biochemical and functional sensitization of D2/3R signaling, that these responses show a region-specific temporal pattern and are fully reversible following drug discontinuation. These results suggest that an increased post-synaptic D2/3R function and a decreased DA presynaptic signaling, mediated by increased D2/3R autoinhibition, may predominate during distinct phases of withdrawal and may contribute both to the mechanisms leading to relapse and to cannabinoid withdrawal symptoms. The different rates of normalization of D2/3R function in striatum and midbrain may be critical information for the development of new pharmacotherapies for cannabis dependence. Copyright © 2016 Elsevier B.V. All rights reserved.

  10. μ-opioid Receptor-Mediated Alterations of Allergen-Induced Immune Responses of Bronchial Lymph Node Cells in a Murine Model of Stress Asthma

    Directory of Open Access Journals (Sweden)

    Kaori Okuyama

    2012-01-01

    Conclusions: Restraint stress aggravated allergic airway inflammation in association with alterations in local immunity characterized by greater Th2-associated cytokine production and a reduced development of regulatory T cells, mediated by MORs.

  11. Mode of action and dose-response framework analysis for receptor-mediated toxicity : The aryl hydrocarbon receptor as a case study

    NARCIS (Netherlands)

    Budinsky, R. A.; Schrenk, D.; Simon, T.; Van Den Berg, M.; Reichard, J. F.; Silkworth, J. B.; Aylward, L. L.; Brix, A.; Gasiewicz, T.; Kaminski, N.; Perdew, G.; Starr, T. B.; Walker, N. J.; Rowlands, J. C.

    2014-01-01

    Dioxins and dioxin-like compounds are tumor promoters that cause liver cancer in rats and mice. The aryl hydrocarbon receptor (AHR) has been implicated as a key component in this tumor promotion response. Despite extensive knowledge of the toxicology of dioxins, no mode of action (MOA) hypothesis

  12. Role of G3BP1 in glucocorticoid receptor-mediated microRNA-15b and microRNA-23a biogenesis in endothelial cells

    KAUST Repository

    Kwok, Hoi-Hin

    2017-05-18

    MicroRNAs (miRNAs) are a family of non-coding RNAs that play crucial roles in regulating various normal cellular responses. Recent studies revealed that the canonical miRNA biogenesis pathway is subject to sophisticated regulation. Hormonal control of miRNA biogenesis by androgen and estrogen has been demonstrated, but the direct effects of the glucocorticoid receptor (GR) on miRNA biogenesis are unknown. This study revealed the role of GR in miRNA maturation. We showed that two GR agonists, dexamethasone and ginsenoside-Rg1 rapidly suppressed the expression of mature miR-15b, miR-23a, and miR-214 in human endothelial cells. RNA pulldown coupled with proteomic analysis identified GTPase-activating protein (SH3 domain) binding protein 1 (G3BP1) as one of the RNA-binding proteins mediating GR-regulated miRNA maturation. Activated GR induced phosphorylation of v-AKT Murine Thymoma Viral Oncogene Homologue (AKT) kinase, which in turn phosphorylated and promoted nuclear translocation of G3BP1. The nuclear G3BP1 bound to the G3BP1 consensus sequence located on primary miR-15b~16-2 and miR-23a~27a~24-2 to inhibit their maturation. The findings from this study have advanced our understanding of the non-genomic effects of GR in the vascular system.

  13. Paradoxical mineralocorticoid receptor-mediated effect in fear memory encoding and expression of rats submitted to an olfactory fear conditioning task

    NARCIS (Netherlands)

    Souza, R.R.; Dal Bó, S.; de Kloet, E.R.; Oitzl, M.S.; Carobrez, A.P.

    2014-01-01

    There is general agreement that the substantial modification in memory and motivational states exerted by corticosteroids after a traumatic experience is mediated in complementary manner by the mineralocorticoid (MR) and glucocorticoid (GR) receptors. Here we tested the hypothesis that

  14. Histone deacetylase inhibitors up-regulate LL-37 expression independent of toll-like receptor mediated signalling in airway epithelial cells.

    Science.gov (United States)

    Liu, Quan; Liu, Juan; Roschmann, Kristina Irene Lisolette; van Egmond, Danielle; Golebski, Korneliusz; Fokkens, Wytske Johanna; Wang, Dehui; van Drunen, Cornelis Maria

    2013-04-11

    HDAC inhibitors have been proposed as anticancer agents. However, their roles in innate genes expression remain not well known. Cathelicidin LL-37 is one of the few human bactericidal peptides, but the regulation of histone acetylation on LL-37 expression in airway epithelium remains largely unknown. Therefore, we investigated the effects of two non-selective HDACi, trichostatin A (TSA) and sodium butyrate (SB), on the expression of the cathelicidin LL-37 in human airway epithelial cells. LL37 in human NCI-H292 airway epithelial cells and the primary cultures of normal nasal epithelial cells(PNEC) in response to HDAC inhibitors with or without poly (I:C) stimulation was assessed using real-time PCR and western blot. In parallel, IL-6 expression was evaluated by ELISA. Our results showed that HDAC inhibitors up-regulated LL-37 gene expression independent of poly (I:C) stimulation in PNEC as well as in NCI-H292 cells. HDAC inhibitors increased LL37 protein expression in NCI-H292 cells but not in PNEC. In addition, HDAC inhibitors significantly inhibited poly (I:C)-induced IL-6 production in both of the epithelial cells. In conclusion, HDAC inhibitors directly up-regulated LL-37 gene expression in human airway epithelial cells.

  15. Histone deacetylase inhibitors up-regulate LL-37 expression independent of toll-like receptor mediated signalling in airway epithelial cells

    NARCIS (Netherlands)

    Liu, Quan; Liu, Juan; Roschmann, Kristina Irene Lisolette; van Egmond, Danielle; Golebski, Korneliusz; Fokkens, Wytske Johanna; Wang, Dehui; van Drunen, Cornelis Maria

    2013-01-01

    HDAC inhibitors have been proposed as anticancer agents. However, their roles in innate genes expression remain not well known. Cathelicidin LL-37 is one of the few human bactericidal peptides, but the regulation of histone acetylation on LL-37 expression in airway epithelium remains largely

  16. Histone deacetylase inhibitors up-regulate LL-37 expression independent of toll-like receptor mediated signalling in airway epithelial cells

    OpenAIRE

    Liu, Q.; Liu, J.; Roschmann, K.I.L.; Egmond, D. van; Golebski, K.; Fokkens, W.J.; Wang, D.; Drunen, C.M. van

    2013-01-01

    HDAC inhibitors have been proposed as anticancer agents. However, their roles in innate genes expression remain not well known. Cathelicidin LL-37 is one of the few human bactericidal peptides, but the regulation of histone acetylation on LL-37 expression in airway epithelium remains largely unknown. Therefore, we investigated the effects of two non-selective HDACi, trichostatin A (TSA) and sodium butyrate (SB), on the expression of the cathelicidin LL-37 in human airway epithelial cells. LL3...

  17. The ginsenoside PPD exerts anti-endometriosis effects by suppressing estrogen receptor-mediated inhibition of endometrial stromal cell autophagy and NK cell cytotoxicity.

    Science.gov (United States)

    Zhang, Bing; Zhou, Wen-Jie; Gu, Chun-Jie; Wu, Ke; Yang, Hui-Li; Mei, Jie; Yu, Jia-Jun; Hou, Xiao-Fan; Sun, Jian-Song; Xu, Feng-Yuan; Li, Da-Jin; Jin, Li-Ping; Li, Ming-Qing

    2018-05-14

    Endometriosis (EMS) is an estrogen-dependent gynecological disease with a low autophagy level of ectopic endometrial stromal cells (eESCs). Impaired NK cell cytotoxic activity is involved in the clearance obstruction of the ectopic endometrial tissue in the abdominopelvic cavity. Protopanaxadiol (PPD) and protopanaxatriol (PPT) are two metabolites of ginsenosides, which have profound biological functions, such as anti-cancer activities. However, the role and mechanism of ginsenosides and metabolites in endometriosis are completely unknown. Here, we found that the compounds PPD, PPT, ginsenoside-Rg3 (G-Rg3), ginsenoside-Rh2 (G-Rh2), and esculentoside A (EsA) led to significant decreases in the viability of eESCs, particularly PPD (IC50 = 30.64 µM). In vitro and in vivo experiments showed that PPD promoted the expression of progesterone receptor (PR) and downregulated the expression of estrogen receptor α (ERα) in eESCs. Treatment with PPD obviously induced the autophagy of eESCs and reversed the inhibitory effect of estrogen on eESC autophagy. In addition, eESCs pretreated with PPD enhanced the cytotoxic activity of NK cells in response to eESCs. PPD decreased the numbers and suppressed the growth of ectopic lesions in a mouse EMS model. These results suggest that PPD plays a role in anti-EMS activation, possibly by restricting estrogen-mediated autophagy regulation and enhancing the cytotoxicity of NK cells. This result provides a scientific basis for potential therapeutic strategies to treat EMS by PPD or further structural modification.

  18. Insulin receptors mediate growth effects in cultured fetal neurons. II. Activation of a protein kinase that phosphorylates ribosomal protein S6

    International Nuclear Information System (INIS)

    Heidenreich, K.A.; Toledo, S.P.

    1989-01-01

    As an initial attempt to identify early steps in insulin action that may be involved in the growth responses of neurons to insulin, we investigated whether insulin receptor activation increases the phosphorylation of ribosomal protein S6 in cultured fetal neurons and whether activation of a protein kinase is involved in this process. When neurons were incubated for 2 h with 32Pi, the addition of insulin (100 ng/ml) for the final 30 min increased the incorporation of 32Pi into a 32K microsomal protein. The incorporation of 32Pi into the majority of other neuronal proteins was unaltered by the 30-min exposure to insulin. Cytosolic extracts from insulin-treated neurons incubated in the presence of exogenous rat liver 40S ribosomes and [gamma-32P]ATP displayed a 3- to 8-fold increase in the phosphorylation of ribosomal protein S6 compared to extracts from untreated cells. Inclusion of cycloheximide during exposure of the neurons to insulin did not inhibit the increased cytosolic kinase activity. Activation of S6 kinase activity by insulin was dose dependent (seen at insulin concentration as low as 0.1 ng/ml) and reached a maximum after 20 min of incubation. Addition of phosphatidylserine, diolein, and Ca2+ to the in vitro kinase reaction had no effect on the phosphorylation of ribosomal protein S6. Likewise, treatment of neurons with (Bu)2cAMP did not alter the phosphorylation of ribosomal protein S6 by neuronal cytosolic extracts. We conclude that insulin activates a cytosolic protein kinase that phosphorylates ribosomal S6 in neurons and is distinct from protein kinase-C and cAMP-dependent protein kinase. Stimulation of this kinase may play a role in insulin signal transduction in neurons

  19. The GLP-1 Receptor Agonist Exendin-4 and Diazepam Differentially Regulate GABAA Receptor-Mediated Tonic Currents in Rat Hippocampal CA3 Pyramidal Neurons.

    Directory of Open Access Journals (Sweden)

    Sergiy V Korol

    Full Text Available Glucagon-like peptide-1 (GLP-1 is a metabolic hormone that is secreted in a glucose-dependent manner and enhances insulin secretion. GLP-1 receptors are also found in the brain where their signalling affects neuronal activity. We have previously shown that the GLP-1 receptor agonists, GLP-1 and exendin-4 enhanced GABA-activated synaptic and tonic currents in rat hippocampal CA3 pyramidal neurons. The hippocampus is the centre for memory and learning and is important for cognition. Here we examined if exendin-4 similarly enhanced the GABA-activated currents in the presence of the benzodiazepine diazepam. In whole-cell recordings in rat brain slices, diazepam (1 μM, an allosteric positive modulator of GABAA receptors, alone enhanced the spontaneous inhibitory postsynaptic current (sIPSC amplitude and frequency by a factor of 1.3 and 1.6, respectively, and doubled the tonic GABAA current normally recorded in the CA3 pyramidal cells. Importantly, in the presence of exendin-4 (10 nM plus diazepam (1 μM, only the tonic but not the sIPSC currents transiently increased as compared to currents recorded in the presence of diazepam alone. The results suggest that exendin-4 potentiates a subpopulation of extrasynaptic GABAA receptors in the CA3 pyramidal neurons.

  20. DMPD: Translational mini-review series on Toll-like receptors: networks regulated byToll-like receptors mediate innate and adaptive immunity. [Dynamic Macrophage Pathway CSML Database

    Lifescience Database Archive (English)

    Full Text Available 17223959 Translational mini-review series on Toll-like receptors: networks regulate...ol. 2007 Feb;147(2):199-207. (.png) (.svg) (.html) (.csml) Show Translational mini-review series on Toll-lik... immunity. PubmedID 17223959 Title Translational mini-review series on Toll-like receptors: networks regulat

  1. Pancreatic adenocarcinoma upregulated factor (PAUF) confers resistance to pancreatic cancer cells against oncolytic parvovirus H-1 infection through IFNA receptor-mediated signaling

    Energy Technology Data Exchange (ETDEWEB)

    Kaowinn, Sirichat; Cho, Il-Rae; Moon, Jeong; Jun, Seung Won; Kim, Chang Seok [BK21+, Department of Cogno-Mechatronics Engineering, Pusan National University, Busan 609-736 (Korea, Republic of); Kang, Ho Young [Department of Microbiology, Pusan National University, Busan 609-736 (Korea, Republic of); Kim, Manbok [Department of Medical Science, Dankook University College of Medicine, Cheonan 330-714 (Korea, Republic of); Koh, Sang Seok [Department of Biological Sciences, Dong-A University, Busan 604-714 (Korea, Republic of); Chung, Young-Hwa, E-mail: younghc@pusan.ac.kr [BK21+, Department of Cogno-Mechatronics Engineering, Pusan National University, Busan 609-736 (Korea, Republic of)

    2015-04-03

    Pancreatic adenocarcinoma upregulated factor (PAUF), a novel oncogene, plays a crucial role in the development of pancreatic cancer, including its metastasis and proliferation. Therefore, PAUF-expressing pancreatic cancer cells could be important targets for oncolytic virus-mediated treatment. Panc-1 cells expressing PAUF (Panc-PAUF) showed relative resistance to parvovirus H-1 infection compared with Panc-1 cells expressing an empty vector (Panc-Vec). Of interest, expression of type I IFN-α receptor (IFNAR) was higher in Panc-PAUF cells than in Panc-Vec cells. Increased expression of IFNAR in turn increased the activation of Stat1 and Tyk2 in Panc-PAUF cells compared with that in Panc-Vec cells. Suppression of Tyk2 and Stat1, which are important downstream molecules for IFN-α signaling, sensitized pancreatic cancer cells to parvovirus H-1-mediated apoptosis. Further, constitutive suppression of PAUF sensitized Bxpc3 pancreatic cancer cells to parvovirus H-1 infection. Taken together, these results suggested that PAUF conferred resistance to pancreatic cancer cells against oncolytic parvovirus H-1 infection through IFNAR-mediated signaling. - Highlights: • PAUF confers resistance against oncolytic parvovirus H-1 infection. • PAUF enhances the expression of IFNAR in Panc-1 cells. • Increased activation of Tyk2 or Stat1 by PAUF provides resistance to parvovirus H-1-mediated apoptosis. • Constitutive inhibition of PAUF enhances parvovirus H-1-mediated oncolysis of Bxpc3 pancreatic cancer cells.

  2. Pancreatic adenocarcinoma upregulated factor (PAUF) confers resistance to pancreatic cancer cells against oncolytic parvovirus H-1 infection through IFNA receptor-mediated signaling

    International Nuclear Information System (INIS)

    Kaowinn, Sirichat; Cho, Il-Rae; Moon, Jeong; Jun, Seung Won; Kim, Chang Seok; Kang, Ho Young; Kim, Manbok; Koh, Sang Seok; Chung, Young-Hwa

    2015-01-01

    Pancreatic adenocarcinoma upregulated factor (PAUF), a novel oncogene, plays a crucial role in the development of pancreatic cancer, including its metastasis and proliferation. Therefore, PAUF-expressing pancreatic cancer cells could be important targets for oncolytic virus-mediated treatment. Panc-1 cells expressing PAUF (Panc-PAUF) showed relative resistance to parvovirus H-1 infection compared with Panc-1 cells expressing an empty vector (Panc-Vec). Of interest, expression of type I IFN-α receptor (IFNAR) was higher in Panc-PAUF cells than in Panc-Vec cells. Increased expression of IFNAR in turn increased the activation of Stat1 and Tyk2 in Panc-PAUF cells compared with that in Panc-Vec cells. Suppression of Tyk2 and Stat1, which are important downstream molecules for IFN-α signaling, sensitized pancreatic cancer cells to parvovirus H-1-mediated apoptosis. Further, constitutive suppression of PAUF sensitized Bxpc3 pancreatic cancer cells to parvovirus H-1 infection. Taken together, these results suggested that PAUF conferred resistance to pancreatic cancer cells against oncolytic parvovirus H-1 infection through IFNAR-mediated signaling. - Highlights: • PAUF confers resistance against oncolytic parvovirus H-1 infection. • PAUF enhances the expression of IFNAR in Panc-1 cells. • Increased activation of Tyk2 or Stat1 by PAUF provides resistance to parvovirus H-1-mediated apoptosis. • Constitutive inhibition of PAUF enhances parvovirus H-1-mediated oncolysis of Bxpc3 pancreatic cancer cells

  3. Prenatal nicotine is associated with reduced AMPA and NMDA receptor-mediated rises in calcium within the laterodorsal tegmentum: a pontine nucleus involved in addiction processes

    DEFF Research Database (Denmark)

    Mc Nair, Laura Kristine Frendrup; Kohlmeier, Kristi Anne

    2015-01-01

    excitatory neurotransmitter within the laterodorsal tegmental nucleus (LDT), which is a brainstem region importantly involved in responding to motivational stimuli and critical in development of drug addiction-associated behaviours, however, it is unknown whether PNE alters glutamate signalling within...

  4. CaMKII inhibition with KN93 attenuates endothelin and serotonin receptor-mediated vasoconstriction and prevents subarachnoid hemorrhage-induced deficits in sensorimotor function

    DEFF Research Database (Denmark)

    Edvinsson, Lars; Povlsen, Gro Klitgaard; Ahnstedt, Hilda

    2014-01-01

    tested the hypothesis that inhibition of calcium calmodulin-dependent protein kinase II (CaMKII) may reduce cerebral vasoconstriction mediated by endothelin and serotonin receptors and improve neurological outcome after experimental SAH. METHODS: SAH was induced in adult rats by injection of 250 μ...

  5. Endocannabinoid CB1 Receptor Mediated Rises in Ca2+ and Depolarization-Induced Suppression of Inhibition within the Laterodorsal Tegmental Nucleus

    DEFF Research Database (Denmark)

    Soni, Neeraj; Kohlmeier, Kristi Anne

    2016-01-01

    -sensitive, inhibitory postsynaptic currents (IPSCs), which was an action sensitive to presence of a Ca(2+) chelator. Finally, actions of DHPG, a mGlu1R agonist, on IPSC activity were examined and found to result in an AM251- and BAPTA-sensitive inhibition of both the frequency and amplitude of sIPSCs. Taken...

  6. Assessment of the aryl hydrocarbon receptor-mediated activities of polycyclic aromatic hydrocarbons in a human cell-based reporter gene assay

    Czech Academy of Sciences Publication Activity Database

    Vondráček, Jan; Pěnčíková, K.; Neca, J.; Ciganek, M.; Grycová, A.; Dvořák, Z.; Machala, M.

    2017-01-01

    Roč. 220, JAN2017 (2017), s. 307-316 ISSN 0269-7491 R&D Projects: GA ČR GA13-07711S Institutional support: RVO:68081707 Keywords : rat primary hepatocytes * liver epithelial-cells * cancer-risk assessment Subject RIV: BO - Biophysics Impact factor: 5.099, year: 2016

  7. Fear potentiated startle increases phospholipase D (PLD) expression/activity and PLD-linked metabotropic glutamate receptor mediated post-tetanic potentiation in rat amygdala.

    Science.gov (United States)

    Krishnan, Balaji; Scott, Michael T; Pollandt, Sebastian; Schroeder, Bradley; Kurosky, Alexander; Shinnick-Gallagher, Patricia

    2016-02-01

    Long-term memory (LTM) of fear stores activity dependent modifications that include changes in amygdala signaling. Previously, we identified an enhanced probability of release of glutamate mediated signaling to be important in rat fear potentiated startle (FPS), a well-established translational behavioral measure of fear. Here, we investigated short- and long-term synaptic plasticity in FPS involving metabotropic glutamate receptors (mGluRs) and associated downstream proteomic changes in the thalamic-lateral amygdala pathway (Th-LA). Aldolase A, an inhibitor of phospholipase D (PLD), expression was reduced, concurrent with significantly elevated PLD protein expression. Blocking the PLD-mGluR signaling significantly reduced PLD activity. While transmitter release probability increased in FPS, PLD-mGluR agonist and antagonist actions were occluded. In the unpaired group (UNP), blocking the PLD-mGluR increased while activating the receptor decreased transmitter release probability, consistent with decreased synaptic potentials during tetanic stimulation. FPS Post-tetanic potentiation (PTP) immediately following long-term potentiation (LTP) induction was significantly increased. Blocking PLD-mGluR signaling prevented PTP and reduced cumulative PTP probability but not LTP maintenance in both groups. These effects are similar to those mediated through mGluR7, which is co-immunoprecipitated with PLD in FPS. Lastly, blocking mGluR-PLD in the rat amygdala was sufficient to prevent behavioral expression of fear memory. Thus, our study in the Th-LA pathway provides the first evidence for PLD as an important target of mGluR signaling in amygdala fear-associated memory. Importantly, the PLD-mGluR provides a novel therapeutic target for treating maladaptive fear memories in posttraumatic stress and anxiety disorders. Published by Elsevier Inc.

  8. The aryl hydrocarbon receptor-mediated and genotoxic effects of fractionated extract of standard reference diesel exhaust particle material in pulmonary, liver and prostate cells

    Czech Academy of Sciences Publication Activity Database

    Pálková, L.; Vondráček, Jan; Trilecová, L.; Ciganek, M.; Pěnčíková, K.; Neca, J.; Milcová, Alena; Topinka, Jan; Machala, M.

    2015-01-01

    Roč. 29, č. 3 (2015), s. 438-448 ISSN 0887-2333 R&D Projects: GA ČR(CZ) GBP503/12/G147 Institutional support: RVO:68081707 ; RVO:68378041 Keywords : Air pollution * PAHs * SRM 1650b Subject RIV: BO - Biophysics; DI - Air Pollution ; Quality (UEM-P) Impact factor: 3.338, year: 2015

  9. TRPV1 receptor-mediated expression of Toll-like receptors 2 and 4 following permanent middle cerebral artery occlusion in rats

    Directory of Open Access Journals (Sweden)

    Elham Hakimizadeh

    2017-08-01

    Full Text Available Objective(s: Stroke is known as a main cause of mortality and prolonged disability in adults. Both transient receptor potential V1 (TRPV1 channels and toll-like receptors (TLRs are involved in mediating the inflammatory responses. In the present study, the effects of TRPV1 receptor activation and blockade on stroke outcome and gene expression of TLR2 and TLR4 were assessed following permanent middle cerebral artery occlusion in rats Materials and Methods: Eighty male Wistar rats were divided into four groups as follows: sham, vehicle, AMG9810 (TRPV1 antagonist -treated and capsaicin (TRPV1 agonist -treated. For Stroke induction, the middle cerebral artery was permanently occluded and then behavioral functions were evaluated 1, 3 and 7 days after stroke. Results: TRPV1 antagonism significantly reduced the infarct volume compared to the stroke group. Also, neurological deficits were decreased by AMG9810 seven days after cerebral ischemia. In the ledged beam-walking test, the slip ratio was enhanced following ischemia. AMG9810 decreased this index in stroke animals. However, capsaicin improved the ratio 3 and 7 days after cerebral ischemia. Compared to the sham group, the mRNA expression of TLR2 and TLR4 was significantly increased in the stroke rats. AMG9810 Administration significantly reduced the mRNA expression of TLR2 and TLR4. However, capsaicin did not significantly affect the gene expression of TLR2 and TLR4. Conclusion: Our results demonstrated that TRPV1 antagonism by AMG9810 attenuates behavioral function and mRNA expression of TLR2 and TLR4. Thus, it might be useful to shed light on future therapeutic strategies for the treatment of ischemic stroke.

  10. Electrophysiological evidence of increased glycine receptor-mediated phasic and tonic inhibition by blockade of glycine transporters in spinal superficial dorsal horn neurons of adult mice

    Directory of Open Access Journals (Sweden)

    Misa Oyama

    2017-03-01

    Full Text Available To understand the synaptic and/or extrasynaptic mechanisms underlying pain relief by blockade of glycine transporter subtypes GlyT1 and GlyT2, whole-cell recordings were made from dorsal horn neurons in spinal slices from adult mice, and the effects of NFPS and ALX-1393, selective GlyT1 and GlyT2 inhibitors, respectively, on phasic evoked or miniature glycinergic inhibitory postsynaptic currents (eIPSCs or mIPSCs were examined. NFPS and ALX-1393 prolonged the decay phase of eIPSCs without affecting their amplitude. In the presence of tetrodotoxin to record mIPSCs, NFPS and ALX-1393 induced a tonic inward current that was reversed by strychnine. Although NFPS had no statistically significant influences on mIPSCs, ALX-1393 significantly increased their frequency. We then further explored the role of GlyTs in the maintenance of glycinergic IPSCs. To facilitate vesicular release of glycine, repetitive high-frequency stimulation (HFS was applied at 10 Hz for 3 min during continuous recordings of eIPSCs at 0.1 Hz. Prominent suppression of eIPSCs was evident after HFS in the presence of ALX-1393, but not NFPS. Thus, it appears that phasic and tonic inhibition may contribute to the analgesic effects of GlyT inhibitors. However, reduced glycinergic inhibition due to impaired vesicular refilling could hamper the analgesic efficacy of GlyT2 inhibitors.

  11. Epidermal growth factor receptor mediated proliferation depends on increased lipid droplet density regulated via a negative regulatory loop with FOXO3/Sirtuin6

    International Nuclear Information System (INIS)

    Penrose, Harrison; Heller, Sandra; Cable, Chloe; Makboul, Rania; Chadalawada, Gita; Chen, Ying; Crawford, Susan E.; Savkovic, Suzana D.

    2016-01-01

    The proliferation of colon cancer cells is mediated in part by epidermal growth factor receptor (EGFR) signaling and requires sustained levels of cellular energy to meet its high metabolic needs. Intracellular lipid droplets (LDs) are a source of energy used for various cellular functions and they are elevated in density in human cancer, yet their regulation and function are not well understood. Here, in human colon cancer cells, EGF stimulates increases in LD density, which depends on EGFR expression and activation as well as the individual cellular capacity for lipid synthesis. Increases in LDs are blockaded by inhibition of PI3K/mTOR and PGE2 synthesis, supporting their dependency on select upstream pathways. In colon cancer cells, silencing of the FOXO3 transcription factor leads to down regulation of SIRT6, a negative regulator of lipid synthesis, and consequent increases in the LD coat protein PLIN2, revealing that increases in LDs depend on loss of FOXO3/SIRT6. Moreover, EGF stimulates loss of FOXO3/SIRT6, which is blockaded by the inhibition of upstream pathways as well as lipid synthesis, revealing existence of a negative regulatory loop between LDs and FOXO3/SIRT6. Elevated LDs are utilized by EGF treatment and their depletion through the inhibition of lipid synthesis or silencing of PLIN2 significantly attenuates proliferation. This novel mechanism of proliferative EGFR signaling leading to elevated LD density in colon cancer cells could potentially be therapeutically targeted for the treatment of tumor progression. - Highlights: • In colon cancer cells, EGFR activation leads to increases in LD density. • EGFR signaling includes PI3K/mTOR and PGE2 leading to lipid synthesis. • Increases in LDs are controlled by a negative regulatory loop with FOXO3/SIRT6. • EGFR mediated colon cancer cell proliferation depends on increased LD density.

  12. TRPV1 receptor-mediated expression of Toll-like receptors 2 and 4 following permanent middle cerebral artery occlusion in rats

    Science.gov (United States)

    Hakimizadeh, Elham; Shamsizadeh, Ali; Roohbakhsh, Ali; Arababadi, Mohammad Kazemi; Hajizadeh, Mohammad Reza; Shariati, Mehdi; Fatemi, Iman; Moghadam-ahmadi, Amir; Bazmandegan, Gholamreza; Rezazadeh, Hossein; Allahtavakoli, Mohammad

    2017-01-01

    Objective(s): Stroke is known as a main cause of mortality and prolonged disability in adults. Both transient receptor potential V1 (TRPV1) channels and toll-like receptors (TLRs) are involved in mediating the inflammatory responses. In the present study, the effects of TRPV1 receptor activation and blockade on stroke outcome and gene expression of TLR2 and TLR4 were assessed following permanent middle cerebral artery occlusion in rats Materials and Methods: Eighty male Wistar rats were divided into four groups as follows: sham, vehicle, AMG9810 (TRPV1 antagonist) -treated and capsaicin (TRPV1 agonist) -treated. For Stroke induction, the middle cerebral artery was permanently occluded and then behavioral functions were evaluated 1, 3 and 7 days after stroke. Results: TRPV1 antagonism significantly reduced the infarct volume compared to the stroke group. Also, neurological deficits were decreased by AMG9810 seven days after cerebral ischemia. In the ledged beam-walking test, the slip ratio was enhanced following ischemia. AMG9810 decreased this index in stroke animals. However, capsaicin improved the ratio 3 and 7 days after cerebral ischemia. Compared to the sham group, the mRNA expression of TLR2 and TLR4 was significantly increased in the stroke rats. AMG9810 Administration significantly reduced the mRNA expression of TLR2 and TLR4. However, capsaicin did not significantly affect the gene expression of TLR2 and TLR4. Conclusion: Our results demonstrated that TRPV1 antagonism by AMG9810 attenuates behavioral function and mRNA expression of TLR2 and TLR4. Thus, it might be useful to shed light on future therapeutic strategies for the treatment of ischemic stroke. PMID:29085577

  13. Hypothalamic kappa opioid receptor mediates both diet-induced and melanin concentrating hormone-induced liver damage through inflammation and endoplasmic reticulum stress.

    Science.gov (United States)

    Imbernon, Monica; Sanchez-Rebordelo, Estrella; Romero-Picó, Amparo; Kalló, Imre; Chee, Melissa J; Porteiro, Begoña; Al-Massadi, Omar; Contreras, Cristina; Fernø, Johan; Senra, Ana; Gallego, Rosalia; Folgueira, Cintia; Seoane, Luisa M; van Gestel, Margriet; Adan, Roger A; Liposits, Zsolt; Dieguez, Carlos; López, Miguel; Nogueiras, Ruben

    2016-10-01

    The opioid system is widely known to modulate the brain reward system and thus affect the behavior of humans and other animals, including feeding. We hypothesized that the hypothalamic opioid system might also control energy metabolism in peripheral tissues. Mice lacking the kappa opioid receptor (κOR) and adenoviral vectors overexpressing or silencing κOR were stereotaxically delivered in the lateral hypothalamic area (LHA) of rats. Vagal denervation was performed to assess its effect on liver metabolism. Endoplasmic reticulum (ER) stress was inhibited by pharmacological (tauroursodeoxycholic acid) and genetic (overexpression of the chaperone glucose-regulated protein 78 kDa) approaches. The peripheral effects on lipid metabolism were assessed by histological techniques and western blot. We show that in the LHA κOR directly controls hepatic lipid metabolism through the parasympathetic nervous system, independent of changes in food intake and body weight. κOR colocalizes with melanin concentrating hormone receptor 1 (MCH-R1) in the LHA, and genetic disruption of κOR reduced melanin concentrating hormone-induced liver steatosis. The functional relevance of these findings was given by the fact that silencing of κOR in the LHA attenuated both methionine choline-deficient, diet-induced and choline-deficient, high-fat diet-induced ER stress, inflammation, steatohepatitis, and fibrosis, whereas overexpression of κOR in this area promoted liver steatosis. Overexpression of glucose-regulated protein 78 kDa in the liver abolished hypothalamic κOR-induced steatosis by reducing hepatic ER stress. This study reveals a novel hypothalamic-parasympathetic circuit modulating hepatic function through inflammation and ER stress independent of changes in food intake or body weight; these findings might have implications for the clinical use of opioid receptor antagonists. (Hepatology 2016;64:1086-1104). © 2016 The Authors. (Hepatology published by Wiley Periodicals, Inc., on behalf of the American Association for the Study of Liver Diseases.

  14. Human T-Cell Leukemia Virus Type 1 Tax-Deregulated Autophagy Pathway and c-FLIP Expression Contribute to Resistance against Death Receptor-Mediated Apoptosis

    Science.gov (United States)

    Wang, Weimin; Zhou, Jiansuo; Shi, Juan; Zhang, Yaxi; Liu, Shilian

    2014-01-01

    ABSTRACT The human T-cell leukemia virus type 1 (HTLV-1) Tax protein is considered to play a central role in the process that leads to adult T-cell leukemia/lymphoma (ATL) and HTLV-1-associated myelopathy/tropical spastic paraparesis (HAM/TSP). HTLV-1 Tax-expressing cells show resistance to apoptosis induced by Fas ligand (FasL) and tumor necrosis factor (TNF)-related apoptosis-inducing ligand (TRAIL). The regulation of Tax on the autophagy pathway in HeLa cells and peripheral T cells was recently reported, but the function and underlying molecular mechanism of the Tax-regulated autophagy are not yet well defined. Here, we report that HTLV-1 Tax deregulates the autophagy pathway, which plays a protective role during the death receptor (DR)-mediated apoptosis of human U251 astroglioma cells. The cellular FLICE-inhibitory protein (c-FLIP), which is upregulated by Tax, also contributes to the resistance against DR-mediated apoptosis. Both Tax-induced autophagy and Tax-induced c-FLIP expression require Tax-induced activation of IκB kinases (IKK). Furthermore, Tax-induced c-FLIP expression is regulated through the Tax-IKK-NF-κB signaling pathway, whereas Tax-triggered autophagy depends on the activation of IKK but not the activation of NF-κB. In addition, DR-mediated apoptosis is correlated with the degradation of Tax, which can be facilitated by the inhibitors of autophagy. IMPORTANCE Our study reveals that Tax-deregulated autophagy is a protective mechanism for DR-mediated apoptosis. The molecular mechanism of Tax-induced autophagy is also illuminated, which is different from Tax-increased c-FLIP. Tax can be degraded via manipulation of autophagy and TRAIL-induced apoptosis. These results outline a complex regulatory network between and among apoptosis, autophagy, and Tax and also present evidence that autophagy represents a new possible target for therapeutic intervention for the HTVL-1 related diseases. PMID:24352466

  15. Capsular Polysaccharide is a Main Component of Mycoplasma ovipneumoniae in the Pathogen-Induced Toll-Like Receptor-Mediated Inflammatory Responses in Sheep Airway Epithelial Cells.

    Science.gov (United States)

    Jiang, Zhongjia; Song, Fuyang; Li, Yanan; Xue, Di; Deng, Guangcun; Li, Min; Liu, Xiaoming; Wang, Yujiong

    2017-01-01

    Mycoplasma ovipneumoniae ( M. ovipneumoniae ) is characterized as an etiological agent of primary atypical pneumonia that specifically infects sheep and goat. In an attempt to better understand the pathogen-host interaction between the invading M. ovipneumoniae and airway epithelial cells, we investigated the host inflammatory responses against capsular polysaccharide (designated as CPS) of M. ovipneumoniae using sheep bronchial epithelial cells cultured in an air-liquid interface (ALI) model. Results showed that CPS derived from M. ovipneumoniae could activate toll-like receptor- (TLR-) mediated inflammatory responses, along with an elevated expression of nuclear factor kappa B (NF- κ B), activator protein-1 (AP-1), and interferon regulatory factor 3 (IRF3) as well as various inflammatory-associated mediators, representatively including proinflammatory cytokines, such as IL1 β , TNF α , and IL8, and anti-inflammatory cytokines such as IL10 and TGF β of TLR signaling cascade. Mechanistically, the CPS-induced inflammation was TLR initiated and was mediated by activations of both MyD88-dependent and MyD88-independent signaling pathways. Of importance, a blockage of CPS with specific antibody led a significant reduction of M. ovipneumoniae -induced inflammatory responses in sheep bronchial epithelial cells. These results suggested that CPS is a key virulent component of M. ovipneumoniae , which may play a crucial role in the inflammatory response induced by M. ovipneumoniae infections.

  16. Dimethyl sulfoxide potentiates death receptor-mediated apoptosis in the human myeloid leukemia U937 cell line through enhancement of mitochondrial membrane depolarization

    Czech Academy of Sciences Publication Activity Database

    Vondráček, Jan; Souček, Karel; Sheard, M. A.; Chramostová, Kateřina; Andrysík, Zdeněk; Hofmanová, Jiřina; Kozubík, Alois

    2006-01-01

    Roč. 30, č. 1 (2006), s. 81-89 ISSN 0145-2126 R&D Projects: GA ČR(CZ) GA524/03/0766 Institutional research plan: CEZ:AV0Z50040507 Keywords : human myeloid leukemia * DMSO * apoptosis Subject RIV: BO - Biophysics Impact factor: 2.483, year: 2006

  17. TDP-43 Loss-of-Function Causes Neuronal Loss Due to Defective Steroid Receptor-Mediated Gene Program Switching in Drosophila

    Directory of Open Access Journals (Sweden)

    Lies Vanden Broeck

    2013-01-01

    Full Text Available TDP-43 proteinopathy is strongly implicated in the pathogenesis of amyotrophic lateral sclerosis and related neurodegenerative disorders. Whether TDP-43 neurotoxicity is caused by a novel toxic gain-of-function mechanism of the aggregates or by a loss of its normal function is unknown. We increased and decreased expression of TDP-43 (dTDP-43 in Drosophila. Although upregulation of dTDP-43 induced neuronal ubiquitin and dTDP-43-positive inclusions, both up- and downregulated dTDP-43 resulted in selective apoptosis of bursicon neurons and highly similar transcriptome alterations at the pupal-adult transition. Gene network analysis and genetic validation showed that both up- and downregulated dTDP-43 directly and dramatically increased the expression of the neuronal microtubule-associated protein Map205, resulting in cytoplasmic accumulations of the ecdysteroid receptor (EcR and a failure to switch EcR-dependent gene programs from a pupal to adult pattern. We propose that dTDP-43 neurotoxicity is caused by a loss of its normal function.

  18. Low-dose dioxins alter gene expression related to cholesterol biosynthesis, lipogenesis, and glucose metabolism through the aryl hydrocarbon receptor-mediated pathway in mouse liver

    International Nuclear Information System (INIS)

    Sato, Shoko; Shirakawa, Hitoshi; Tomita, Shuhei; Ohsaki, Yusuke; Haketa, Keiichi; Tooi, Osamu; Santo, Noriaki; Tohkin, Masahiro; Furukawa, Yuji; Gonzalez, Frank J.; Komai, Michio

    2008-01-01

    2,3,7,8-Tetrachlorodibenzo-p-dioxin (TCDD) is a common environmental contaminant. TCDD binds and activates the transcription factor aryl hydrocarbon receptor (AHR), leading to adverse biological responses via the alteration of the expression of various AHR target genes. Although small amounts of TCDD are consumed via contaminated daily foodstuffs and environmental exposures, the effects of low-dose TCDD on gene expression in animal tissues have not been clarified, while a number of genes affected by high-dose TCDD were reported. In this study, we comprehensively analyzed gene expression profiles in livers of C57BL/6N mice that were orally administered relatively low doses of TCDD (5, 50, or 500 ng/kg body weight (bw) day -1 ) for 18 days. The hepatic TCDD concentrations, measured by gas chromatography-mass spectrometry, were 1.2, 17, and 1063 pg toxicity equivalent quantity (TEQ)/g, respectively. The mRNA level of the cytochrome P450 CYP1A1 was significantly increased by treatment with only TCDD 500 ng/kg bw day -1 . DNA microarray and quantitative RT-PCR analyses revealed changes in the expression of genes involved in the circadian rhythm, cholesterol biosynthesis, fatty acid synthesis, and glucose metabolism in the liver with at all doses of TCDD employed. However, repression of expression of genes involved in energy metabolism was not observed in the livers of Ahr-null mice that were administered the same dose of TCDD. These results indicate that changes in gene expression by TCDD are mediated by AHR and that exposure to low-dose TCDD could affect energy metabolism via alterations of gene expression

  19. α7 Nicotinic acetylcholine receptor-mediated anti-inflammatory effect in a chronic migraine rat model via the attenuation of glial cell activation

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    Liu Q

    2018-06-01

    Full Text Available Qing Liu,1 Chaoyang Liu,1 Li Jiang,1 Maolin Li,1 Ting Long,1 Wei He,1 Guangcheng Qin,2 Lixue Chen,2 Jiying Zhou1 1Department of Neurology, The First Affiliated Hospital, Chongqing Medical University, Chongqing, China; 2Laboratory Research Center, The First Affiliated Hospital, Chongqing Medical University, Chongqing, China Background: Evidence suggests that the activation of α7 nicotinic acetylcholine receptor (α7nAChR can greatly decrease the neuroinflammation response. Neuroinflammation plays a pivotal role in the pathogenesis of chronic migraine (CM. Clinical observations also show that nicotine gum induces analgesic effects in migraine patients. However, whether α7nAChR is involved in CM is unclear.Objective: To investigate the role of α7nAChR in CM and provide a new therapeutic target for CM.Materials and methods: Thirty-six male Sprague–Dawley rats were distributed randomly into control, CM, PNU-282987, and α-bungarotoxin groups (n=9 rats in each group. The CM model was established by the recurrent daily administration of inflammatory soup on the dura over the course of 1 week. The hind paw threshold and facial allodynia were assessed by the von Frey test. The expression levels of α7nAChR, tumor necrosis factor-alpha, and interleukin-1 beta were analyzed by Western blot and real-time fluorescence quantitative polymerase chain reaction. The location of α7nAChR in the hippocampus was quantified by immunofluorescence, as well as the microglial and astrocyte alterations. Changes in the calcitonin gene-related peptide and the phosphorylated JNK protein among different groups were measured by Western blot.Results: We found that the expression of α7nAChR was reduced after repeated inflammatory soup administration. The increased expression of tumor necrosis factor-alpha, interleukin-1 beta, and calcitonin gene-related peptide in CM group were significantly decreased by PNU-282987 and aggravated by α-bungarotoxin. Moreover, PNU-282987 decreased the numbers of astrocytes and microglia compared with the numbers in the CM group in both hippocampal CA1 and CA3 regions. In contrast, α-bungarotoxin activated the astrocytes and microglia, but the differences with respect to the CM group were not significant. Activated c-Jun N-terminal kinase signaling was observed in CM rats and was also blocked by PNU-282987.Conclusion: The activation of α7nAChR increased the mechanical threshold and alleviated pain in the CM rat model. α7nAChR activation also decreased the upregulation of astrocytes and microglia through the p-c-Jun N-terminal kinase–mitogen-activated protein kinase signaling pathway. Keywords: chronic migraine, α7nAChR, analgesia, glial activation, neuroinflammation, nociception 

  20. Epidermal growth factor receptor mediated proliferation depends on increased lipid droplet density regulated via a negative regulatory loop with FOXO3/Sirtuin6

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    Penrose, Harrison; Heller, Sandra; Cable, Chloe [Department of Pathology and Laboratory Medicine, Tulane University School of Medicine, 1430 Tulane Ave SL-79, New Orleans, LA 70112 (United States); Makboul, Rania [Department of Pathology and Laboratory Medicine, Tulane University School of Medicine, 1430 Tulane Ave SL-79, New Orleans, LA 70112 (United States); Pathology Department, Assiut University, Assiut (Egypt); Chadalawada, Gita; Chen, Ying [Department of Pathology and Laboratory Medicine, Tulane University School of Medicine, 1430 Tulane Ave SL-79, New Orleans, LA 70112 (United States); Crawford, Susan E. [Department of Pathology, Saint Louis University School of Medicine, 1402 South Grand Blvd, Saint Louis, MO 63104 (United States); Savkovic, Suzana D., E-mail: ssavkovi@tulane.edu [Department of Pathology and Laboratory Medicine, Tulane University School of Medicine, 1430 Tulane Ave SL-79, New Orleans, LA 70112 (United States)

    2016-01-15

    The proliferation of colon cancer cells is mediated in part by epidermal growth factor receptor (EGFR) signaling and requires sustained levels of cellular energy to meet its high metabolic needs. Intracellular lipid droplets (LDs) are a source of energy used for various cellular functions and they are elevated in density in human cancer, yet their regulation and function are not well understood. Here, in human colon cancer cells, EGF stimulates increases in LD density, which depends on EGFR expression and activation as well as the individual cellular capacity for lipid synthesis. Increases in LDs are blockaded by inhibition of PI3K/mTOR and PGE2 synthesis, supporting their dependency on select upstream pathways. In colon cancer cells, silencing of the FOXO3 transcription factor leads to down regulation of SIRT6, a negative regulator of lipid synthesis, and consequent increases in the LD coat protein PLIN2, revealing that increases in LDs depend on loss of FOXO3/SIRT6. Moreover, EGF stimulates loss of FOXO3/SIRT6, which is blockaded by the inhibition of upstream pathways as well as lipid synthesis, revealing existence of a negative regulatory loop between LDs and FOXO3/SIRT6. Elevated LDs are utilized by EGF treatment and their depletion through the inhibition of lipid synthesis or silencing of PLIN2 significantly attenuates proliferation. This novel mechanism of proliferative EGFR signaling leading to elevated LD density in colon cancer cells could potentially be therapeutically targeted for the treatment of tumor progression. - Highlights: • In colon cancer cells, EGFR activation leads to increases in LD density. • EGFR signaling includes PI3K/mTOR and PGE2 leading to lipid synthesis. • Increases in LDs are controlled by a negative regulatory loop with FOXO3/SIRT6. • EGFR mediated colon cancer cell proliferation depends on increased LD density.