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Khalifeh-Soltani A, Gupta D, Ha A, Podolsky MJ, Datta R, Atabai K. The Mfge8-α8β1-PTEN pathway regulates airway smooth muscle contraction in allergic inflammation. FASEB J 2018; 32:fj201800109R. [PMID: 29763381 DOI: 10.1096/fj.201800109r] [Citation(s) in RCA: 13] [Impact Index Per Article: 1.9] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 01/28/2023]
Abstract
Asthma affects ∼300 million people worldwide. Despite multiple treatment options, asthma treatment remains unsatisfactory in a subset of patients. Airway obstruction is a hallmark of allergic asthma and is largely due to airway smooth muscle hypercontractility induced by airway inflammation. Identification of molecular pathways that regulate airway smooth muscle hypercontractility is of considerable therapeutic interest. We previously identified roles for milk fat globule epidermal growth factor-like 8 (Mfge8) in opposing the effects of allergic inflammation on increasing airway smooth muscle contractile force. In this study, we delineate the signaling pathway by which Mfge8 mediates these effects. By using genetic and pharmacologic approaches, we show that the α8β1 integrin and the phosphatase and tensin homolog (PTEN) mediate the effects of Mfge8 on preventing IL-13-induced increases in airway contractility. Tracheal rings from mice with smooth muscle-specific deletion of α8β1 or PTEN have enhanced contraction in response to treatment with IL-13. Enhanced IL-13-induced tracheal ring contraction in Mfge8-/- mice was abolished by treatment with the PI3K inhibitor. Mechanistically, IL-13 induces ubiquitination and degradation of PTEN protein. Our findings identify a role for the Mfge8-α8β1-PTEN pathway in regulating the force of airway smooth muscle contraction in the setting of allergic inflammation.-Khalifeh-Soltani, A., Gupta, D., Ha, A., Podolsky, M. J., Datta, R., Atabai, K. The Mfge8-α8β1-PTEN pathway regulates airway smooth muscle contraction in allergic inflammation.
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Affiliation(s)
- Amin Khalifeh-Soltani
- Cardiovascular Research Institute, University of California, San Francisco, San Francisco, California, USA
| | - Deepti Gupta
- Cardiovascular Research Institute, University of California, San Francisco, San Francisco, California, USA
| | - Arnold Ha
- Cardiovascular Research Institute, University of California, San Francisco, San Francisco, California, USA
| | - Michael J Podolsky
- Cardiovascular Research Institute, University of California, San Francisco, San Francisco, California, USA
- Lung Biology Center, University of California, San Francisco, San Francisco, California, USA; and
- Department of Medicine, University of California, San Francisco, San Francisco, California, USA
| | - Ritwik Datta
- Cardiovascular Research Institute, University of California, San Francisco, San Francisco, California, USA
| | - Kamran Atabai
- Cardiovascular Research Institute, University of California, San Francisco, San Francisco, California, USA
- Lung Biology Center, University of California, San Francisco, San Francisco, California, USA; and
- Department of Medicine, University of California, San Francisco, San Francisco, California, USA
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Antileishmanial Activity and Inducible Nitric Oxide Synthase Activation by RuNO Complex. Mediators Inflamm 2016; 2016:2631625. [PMID: 27795620 PMCID: PMC5067336 DOI: 10.1155/2016/2631625] [Citation(s) in RCA: 10] [Impact Index Per Article: 1.1] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 02/23/2016] [Revised: 07/04/2016] [Accepted: 07/17/2016] [Indexed: 02/04/2023] Open
Abstract
Parasites of the genus Leishmania are capable of inhibiting effector functions of macrophages. These parasites have developed the adaptive ability to escape host defenses; for example, they inactivate the NF-κB complex and suppress iNOS expression in infected macrophages, which are responsible for the production of the major antileishmanial substance nitric oxide (NO), favoring then its replication and successful infection. Metal complexes with NO have been studied as potential compounds for the treatment of certain tropical diseases, such as ruthenium compounds, known to be exogenous NO donors. In the present work, the compound cis-[Ru(bpy)2SO3(NO)]PF6, or RuNO, showed leishmanicidal activity directly and indirectly on promastigote forms of Leishmania (Leishmania) amazonensis. In addition, treatment with RuNO increased NO production by reversing the depletion of NO caused by Leishmania. We also found increased expression of Akt, iNOS, and NF-κB in infected and treated macrophages. These results demonstrated that RuNO was able to kill the parasite by NO release and modulate the transcriptional capacity of the cell.
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Soufli I, Toumi R, Rafa H, Touil-Boukoffa C. Overview of cytokines and nitric oxide involvement in immuno-pathogenesis of inflammatory bowel diseases. World J Gastrointest Pharmacol Ther 2016; 7:353-360. [PMID: 27602236 PMCID: PMC4986402 DOI: 10.4292/wjgpt.v7.i3.353] [Citation(s) in RCA: 252] [Impact Index Per Article: 28.0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 03/29/2016] [Revised: 06/10/2016] [Accepted: 07/18/2016] [Indexed: 02/06/2023] Open
Abstract
Inflammatory bowel diseases (IBDs), including Crohn’s disease and ulcerative colitis are complex disorders with undetermined etiology. Several hypotheses suggest that IBDs result from an abnormal immune response against endogenous flora and luminal antigens in genetically susceptible individuals. The dysfunction of the mucosal immune response is implicated in the pathogenesis of IBD. The balance between pro-inflammatory cytokines [tumor necrosis factor (TNF)-α, interleukin (IL)-1β, IL-8, and IL-17A], anti-inflammatory cytokines (IL-4 and IL-13), and immunoregulatory cytokines (IL-10 and transforming growth factors β) is disturbed. Moreover, evidence from animal and clinical studies demonstrate a positive correlation between an increased concentration of nitric oxide (NO) and the severity of the disease. Interestingly, proinflammatory cytokines are involved in the up-regulation of inducible oxide synthase (iNOS) expression in IBD. However, anti-inflammatory and immunoregulatory cytokines are responsible for the negative regulation of iNOS. A positive correlation between NO production and increased pro-inflammatory cytokine levels (TNF-α, IL-6, IL-17, IL-12, and interferon-γ) were reported in patients with IBD. This review focuses on the role of cytokines in intestinal inflammation and their relationship with NO in IBD.
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Codogno P, Klionsky DJ. Opening new doors in autophagy research: Patrice Codogno. Autophagy 2016; 12:1063-8. [PMID: 27158743 DOI: 10.1080/15548627.2016.1170267] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 10/21/2022] Open
Abstract
Patrice Codogno ( Fig. 1 ), one of the associate editors of Autophagy since it was established, is well known in the autophagy field, and has played a particularly important role in France, serving as the first president of Club Francophone de l'AuTophaGie (CFATG). Patrice's research career spans from the predominantly biochemical analyses that were commonly used in the 1980s to the molecular studies that are the primary focus of many labs currently studying autophagy today. Anyone who has met Patrice knows that he is modest, which means his contributions to autophagy and to promoting the careers of scientists globally, are underappreciated. In addition, there is a fun-loving side to Patrice that is often hidden to the casual observer, and it is time to share some of his personality and thoughts with the rest of the autophagy community.
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Affiliation(s)
- Patrice Codogno
- a Institut Necker Enfants-Malades, INSERM U1151-CNRS UMR 8253 Université Paris Descartes-Sorbonne Paris Cité , Paris , France
| | - Daniel J Klionsky
- b Life Sciences Institute, University of Michigan , Ann Arbor , MI , USA
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Li J, Chanda D, Shiri-Sverdlov R, Neumann D. MSP: an emerging player in metabolic syndrome. Cytokine Growth Factor Rev 2014; 26:75-82. [PMID: 25466635 DOI: 10.1016/j.cytogfr.2014.10.007] [Citation(s) in RCA: 14] [Impact Index Per Article: 1.3] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 10/13/2014] [Accepted: 10/22/2014] [Indexed: 12/15/2022]
Abstract
MSP (Macrophage Stimulating Protein; also known as Hepatocyte Growth Factor-like protein (HGFL) and MST1) is a secreted protein and the ligand for transmembrane receptor tyrosine kinase Recepteur d'Origine Nantais (RON; also known as MST1R). Since its discovery, MSP has been demonstrated to play a key role in regulating inflammation in the peripheral tissues of multiple disease models. Recent evidences also point toward a beneficial role of MSP in the regulation of hepatic lipid and glucose metabolism, thereby implicating MSP as a crucial regulator in maintaining metabolic homeostasis while simultaneously suppressing inflammatory processes. In this review, we discuss the recent advances that demonstrate the significance of MSP in metabolic syndrome and build a strong case supporting its therapeutic potential.
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Affiliation(s)
- Jieyi Li
- Department of Molecular Genetics, NUTRIM School for Nutrition, Toxicology and Metabolism, Maastricht University, 6200 MD Maastricht, The Netherlands
| | - Dipanjan Chanda
- Department of Molecular Genetics, CARIM School for Cardiovascular Diseases, Maastricht University, 6200 MD Maastricht, The Netherlands.
| | - Ronit Shiri-Sverdlov
- Department of Molecular Genetics, NUTRIM School for Nutrition, Toxicology and Metabolism, Maastricht University, 6200 MD Maastricht, The Netherlands
| | - Dietbert Neumann
- Department of Molecular Genetics, CARIM School for Cardiovascular Diseases, Maastricht University, 6200 MD Maastricht, The Netherlands
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Resveratrol modulates cytokine-induced Jak/STAT activation more efficiently than 5-aminosalicylic acid: an in vitro approach. PLoS One 2014; 9:e109048. [PMID: 25271420 PMCID: PMC4182878 DOI: 10.1371/journal.pone.0109048] [Citation(s) in RCA: 41] [Impact Index Per Article: 3.7] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 06/03/2014] [Accepted: 09/03/2014] [Indexed: 12/21/2022] Open
Abstract
Background Many advances have been recently made focused on the valuable help of dietary polyphenols in chronic inflammatory diseases. On the other hand, current treatment options for intestinal bowel disease patients are unsatisfying and, for this reason, it is estimated that many patients use dietary supplements to achieve extra benefits. Aim The aim of this work was to analyze under a mechanistic perspective the anti-inflammatory potential of resveratrol, a natural polyphenolic compound, and to compare it with a pharmaceutical agent, 5-aminosalicylic acid, using the intestinal HT-29 cell line, as a cellular model. Methodology and Principal Findings In the present study, HT-29 colon epithelial cells were pre-treated with 25 µM resveratrol and/or 500 µM 5-aminosalicylic acid and then exposed to a combination of cytokines (IL-1α, TNF-α, IFN-γ) for a certain period of time. Our data showed that resveratrol, used in a concentration 20 times lower than 5-aminosalicylic acid, was able to significantly reduce NO and PGE2 production, iNOS and COX-2 expression and reactive oxidant species formation induced by the cytokine challenge. However, as already verified with 5-aminosalicylic acid, in spite of not exhibiting any effect on IkB-α degradation, resveratrol down-regulated JAK-STAT pathway, decreasing the levels of activated STAT1 in the nucleus. Additionally, resveratrol decreased the cytokine-stimulated activation of SAPK/JNK pathway but did not counteract the cytokine-triggered negative feedback mechanism of STAT1, through p38 MAPK. Conclusion/Significance Taken together, our results show that resveratrol may be considered a future nutraceutical approach, promoting remission periods, limiting the inflammatory process and preventing colorectal cancer, which is common in these patients.
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Jiang LQ, Franck N, Egan B, Sjögren RJO, Katayama M, Duque-Guimaraes D, Arner P, Zierath JR, Krook A. Autocrine role of interleukin-13 on skeletal muscle glucose metabolism in type 2 diabetic patients involves microRNA let-7. Am J Physiol Endocrinol Metab 2013; 305:E1359-66. [PMID: 24105413 DOI: 10.1152/ajpendo.00236.2013] [Citation(s) in RCA: 89] [Impact Index Per Article: 7.4] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Indexed: 12/28/2022]
Abstract
Low-grade inflammation associated with type 2 diabetes (T2DM) is postulated to exacerbate insulin resistance. We report that serum levels, as well as IL-13 secreted from cultured skeletal muscle, are reduced in T2DM vs. normal glucose-tolerant (NGT) subjects. IL-13 exposure increases skeletal muscle glucose uptake, oxidation, and glycogen synthesis via an Akt-dependent mechanism. Expression of microRNA let-7a and let-7d, which are direct translational repressors of the IL-13 gene, was increased in skeletal muscle from T2DM patients. Overexpression of let-7a and let-7d in cultured myotubes reduced IL-13 secretion. Furthermore, basal glycogen synthesis was reduced in cultured myotubes exposed to an IL-13-neutralizing antibody. Thus, IL-13 is synthesized and released by skeletal muscle through a mechanism involving let-7, and this effect is attenuated in skeletal muscle from insulin-resistant T2DM patients. In conclusion, IL-13 plays an autocrine role in skeletal muscle to increase glucose uptake and metabolism, suggesting a role in glucose homeostasis in metabolic disease.
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Affiliation(s)
- Lake Q Jiang
- Department of Physiology and Pharmacology, Section for Integrative Physiology, Karolinska Institutet, Stockholm, Sweden
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Gianchecchi E, Delfino DV, Fierabracci A. Recent insights on the putative role of autophagy in autoimmune diseases. Autoimmun Rev 2013; 13:231-41. [PMID: 24184881 DOI: 10.1016/j.autrev.2013.10.007] [Citation(s) in RCA: 33] [Impact Index Per Article: 2.8] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Journal Information] [Subscribe] [Scholar Register] [Received: 10/01/2013] [Accepted: 10/15/2013] [Indexed: 12/17/2022]
Abstract
The incidence of autoimmune pathologies is increasing worldwide. This has stimulated interest on their etiopathogenesis, caused by a complex interaction of genetic and environmental factors. With the advent of genome-wide linkage, candidate gene and genome wide association studies, risk polymorphisms in autophagy-related genes were discovered in several autoimmune conditions suggesting the possible contribution of autophagy to their etiopathogenesis. Autophagy represents the principal catabolic process mediated by lysosomes used by eukaryotic cells and is strictly regulated by proteins belonging to the Atg family. The function of autophagy has been well characterized in various tissues and systems, but its role in the regulation of innate and adaptive immune systems has been only recently discovered. It plays a fundamental role in the modulation of thymocyte selection and in the generation of T lymphocyte repertoire by participating in the intracellular antigen presentation on MHC class-II molecules by thymic epithelial cells. Furthermore, the generation of mice with knockout for specific autophagy-related genes induced several immunological alterations, including defects in B and T cell compartments and in T cell activation. In this review we report recent evidence on the role of autophagy in autoimmunity and discuss its relevance to the pathogenesis of these diseases. We finally highlight that future research may disclose potential new therapeutic targets for the treatment of this category of disorders by modulating the autophagic pathway.
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Affiliation(s)
- Elena Gianchecchi
- Autoimmunity Laboratory, Immunology Area, Bambino Gesù Children's Hospital IRCCS, Rome, Italy
| | - Domenico Vittorio Delfino
- Section of Pharmacology, Toxicology and Chemotherapy, Department of Clinical and Experimental Medicine, Perugia University, Perugia, Italy
| | - Alessandra Fierabracci
- Autoimmunity Laboratory, Immunology Area, Bambino Gesù Children's Hospital IRCCS, Rome, Italy.
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Cyanidin-3-glucoside suppresses cytokine-induced inflammatory response in human intestinal cells: comparison with 5-aminosalicylic acid. PLoS One 2013; 8:e73001. [PMID: 24039842 PMCID: PMC3765207 DOI: 10.1371/journal.pone.0073001] [Citation(s) in RCA: 90] [Impact Index Per Article: 7.5] [Reference Citation Analysis] [Abstract] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 03/26/2013] [Accepted: 07/16/2013] [Indexed: 12/26/2022] Open
Abstract
The potential use of polyphenols in the prevention and treatment of chronic inflammatory diseases has been extensively investigated although the mechanisms involved in cellular signaling need to be further elucidated. Cyanidin-3-glucoside is a typical anthocyanin of many pigmented fruits and vegetables widespread in the human diet. In the present study, the protection afforded by cyanidin-3-glucoside against cytokine-triggered inflammatory response was evaluated in the human intestinal HT-29 cell line, in comparison with 5-aminosalicylic acid, a well-established anti-inflammatory drug, used in inflammatory bowel disease. For this purpose, some key inflammatory mediators and inflammatory enzymes were examined. Our data showed that cyanidin-3-glucoside reduced cytokine-induced inflammation in intestinal cells, in terms of NO, PGE2 and IL-8 production and of iNOS and COX-2 expressions, at a much lower concentration than 5-aminosalicylic acid, suggesting a higher anti-inflammatory efficiency. Interestingly, cyanidin-3-glucoside and 5-aminosalicylic acid neither prevented IkB-α degradation nor the activation of NF-kB, but significantly reduced cytokine-induced levels of activated STAT1 accumulated in the cell nucleus. In addition, we established that phosphorylated p38 MAPK was not involved in the protective effect of cyanidin-3-glucoside or 5-aminosalicylic acid. Taking into account the high concentrations of dietary anthocyanins potentially reached in the gastrointestinal tract, cyanidin-3-glucoside may be envisaged as a promising nutraceutical giving complementary benefits in the context of inflammatory bowel disease.
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10
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Ratajczak-Wrona W, Jablonska E, Garley M, Jablonski J, Radziwon P, Iwaniuk A, Grubczak K. PI3K-Akt/PKB signaling pathway in neutrophils and mononuclear cells exposed to N-nitrosodimethylamine. J Immunotoxicol 2013; 11:231-7. [PMID: 23971717 DOI: 10.3109/1547691x.2013.826307] [Citation(s) in RCA: 9] [Impact Index Per Article: 0.8] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/26/2022] Open
Abstract
Neutrophils (PMN) play diverse regulatory and effector functions in the immune system through the release of reactive nitrogen species, including nitric oxide (NO). The enzyme responsible for NO synthesis in PMN is inducible nitric oxide synthase (iNOS) that is regulated by various signaling pathways, e.g. PI3K-Akt/PKB, and transcription factors. N-Nitrosodimethylamine (NDMA), a xenobiotic widespread in the human environment, affects immune cells. The study objective here was to examine the role of the PI3K-Akt/PKB pathway in induction of NO synthesis (with involvement of iNOS) in human PMN, as well as in autologous mononuclear cells (PBMC), exposed to NDMA. Isolated cells were incubated for 2 h with a sub-lethal dose of NDMA and then the expression of several select proteins in the cell cytoplasmic and nuclear fractions were determined by Western blot analyses. The results indicated that NDMA enhanced expression of iNOS, phospho-PI3K, and phospho-IκBα in the cytoplasmic fraction of the PMN and PBMC. The nuclear fraction of these cells also had a higher NF-κB expression. Moreover, in PMN, NDMA caused an increased expression of phospho-Akt (T308), phospho-Akt (S473), and phospho-IKKαβ in the cytoplasm, and c-Jun and FosB in the nuclear fraction. Blocking of PI3K caused a decrease in expression of all these proteins in NDMA-exposed PMN. However, inhibition of PI3K led to a drop in expression of iNOS, phospho-PI3K, and phospho-IκBα in the cytoplasm, and in NF-κB in the nuclear fraction, of PBMC. The results of these studies indicated to us that NDMA activates the PI3K-Akt/PKB pathway in human PMN and that this, in turn, contributes to the activation of transcription factors NF-κB, c-Jun, and FosB involved in NO production (through modulation of iNOS expression).
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Loss-of-function of inositol polyphosphate-4-phosphatase reversibly increases the severity of allergic airway inflammation. Nat Commun 2012; 3:877. [PMID: 22673904 DOI: 10.1038/ncomms1880] [Citation(s) in RCA: 50] [Impact Index Per Article: 3.8] [Reference Citation Analysis] [Abstract] [Journal Information] [Subscribe] [Scholar Register] [Received: 08/11/2011] [Accepted: 05/02/2012] [Indexed: 01/09/2023] Open
Abstract
Inositol polyphosphate phosphatases regulate the magnitude of phosphoinositide-3 kinase signalling output. Although inositol polyphosphate-4-phosphatase is known to regulate phosphoinositide-3 kinase signalling, little is known regarding its role in asthma pathogenesis. Here we show that modulation of inositol polyphosphate-4-phosphatase alters the severity of asthma. Allergic airway inflammation in mice led to calpain-mediated degradation of inositol polyphosphate-4-phosphatase. In allergic airway inflammation models, preventing inositol polyphosphate-4-phosphatase degradation by inhibiting calpain activity, or overexpression of inositol polyphosphate-4-phosphatase in mouse lungs, led to attenuation of the asthma phenotype. Conversely, knockdown of inositol polyphosphate-4-phosphatase severely aggravated the allergic airway inflammation and the asthma phenotype. Interestingly, inositol polyphosphate-4-phosphatase knockdown in lungs of naive mice led to spontaneous airway hyper-responsiveness, suggesting that inositol polyphosphate-4-phosphatase could be vital in maintaining the lung homeostasis. We suggest that inositol polyphosphate-4-phosphatase has an important role in modulating inflammatory response in asthma, and thus, uncover a new understanding of the complex interplay between inositol signalling and asthma, which could provide alternative strategies in asthma management.
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Saponaro C, Cianciulli A, Calvello R, Dragone T, Iacobazzi F, Panaro MA. The PI3K/Akt pathway is required for LPS activation of microglial cells. Immunopharmacol Immunotoxicol 2012; 34:858-65. [DOI: 10.3109/08923973.2012.665461] [Citation(s) in RCA: 73] [Impact Index Per Article: 5.6] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/21/2022]
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Tsai KD, Chang WW, Lin CC, Hsu SC, Lee YJ, Chen W, Shieh JC, Lin TH. Differential effects of LY294002 and wortmannin on inducible nitric oxide synthase expression in glomerular mesangial cells. Int Immunopharmacol 2012; 12:471-80. [PMID: 22240122 DOI: 10.1016/j.intimp.2011.12.017] [Citation(s) in RCA: 7] [Impact Index Per Article: 0.5] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 07/01/2011] [Revised: 11/26/2011] [Accepted: 12/22/2011] [Indexed: 01/07/2023]
Abstract
Nitric oxide (NO) that is produced by inducible nitric oxide synthase (iNOS) is associated with the pathophysiology of glomerulonephritis. Numerous studies have focused on the regulation of NO production by iNOS to reduce NO-mediated cytotoxicity. In the present study, we demonstrated the differential effects of two phosphatidylinositol 3-kinase (PI3K) inhibitors, LY294002 and wortmannin, on lipopolysaccharide- (LPS) and interferon (IFN)-γ-induced NO production in a glomerular mesangial cell line, MES-13 cells. At dosages without affecting cell viability of MES-13 cells, 5μM LY294002 showed a more-significant inhibitory effect on LPS/IFN-γ-induced NO production, and iNOS protein and gene expressions than did 1μM wortmannin. Akt phosphorylation in MES-13 cells declined upon the addition of wortmannin, but not upon treatment with LY294002. Suppression of PI3K expression by small interfering (si)RNA exhibited no effect on LPS/IFN-γ-stimulated NO production or iNOS protein expression in MES-13 cells. Neither LY294002 nor wortmannin reduced IFN-γ-induced STAT-1α phosphorylation. LY294002 exhibited a more-significant inhibitory effect on NF-κB luciferase activities than wortmannin in LPS/IFN-γ-stimulated MES-13 cells. Moreover, LY294002, but not wortmannin, accelerated iNOS protein degradation and reduced the iNOS dimer/monomer ratio in MES-13 cells. Although both LY294002 and wortmannin are known as PI3K inhibitors, their differential effects on iNOS expression in MES-13 cells indicate that the effects of LY294002 on inhibiting NF-κB activation and accelerating iNOS protein degradation are through a mechanism independent of PI3K.
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Affiliation(s)
- Kuen-Daw Tsai
- Department of Internal Medicine, China Medical University Beigang Hospital, 123 Sinde Road, Beigang Township, Yunlin County 65152, Taiwan, ROC
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Stavrum R, Stavrum AK, Valvatne H, Riley LW, Ulvestad E, Jonassen I, Assmus J, Doherty TM, Grewal HMS. Modulation of transcriptional and inflammatory responses in murine macrophages by the Mycobacterium tuberculosis mammalian cell entry (Mce) 1 complex. PLoS One 2011; 6:e26295. [PMID: 22039457 PMCID: PMC3200323 DOI: 10.1371/journal.pone.0026295] [Citation(s) in RCA: 14] [Impact Index Per Article: 1.0] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 06/07/2011] [Accepted: 09/23/2011] [Indexed: 11/22/2022] Open
Abstract
The outcome of many infections depends on the initial interactions between agent and host. Aiming at elucidating the effect of the M. tuberculosis Mce1 protein complex on host transcriptional and immunological responses to infection with M. tuberculosis, RNA from murine macrophages at 15, 30, 60 min, 4 and 10 hrs post-infection with M. tuberculosis H37Rv or Δ-mce1 H37Rv was analyzed by whole-genome microarrays and RT-QPCR. Immunological responses were measured using a 23-plex cytokine assay. Compared to uninfected controls, 524 versus 64 genes were up-regulated by 15 min post H37Rv- and Δ-mce1 H37Rv-infection, respectively. By 15 min post-H37Rv infection, a decline of 17 cytokines combined with up-regulation of Ccl24 (26.5-fold), Clec4a2 (23.2-fold) and Pparγ (10.5-fold) indicated an anti-inflammatory response initiated by IL-13. Down-regulation of Il13ra1 combined with up-regulation of Il12b (30.2-fold), suggested switch to a pro-inflammatory response by 4 hrs post H37Rv-infection. Whereas no significant change in cytokine concentration or transcription was observed during the first hour post Δ-mce1 H37Rv-infection, a significant decline of IL-1b, IL-9, IL-13, Eotaxin and GM-CSF combined with increased transcription of Il12b (25.1-fold) and Inb1 (17.9-fold) by 4 hrs, indicated a pro-inflammatory response. The balance between pro-and anti-inflammatory responses during the early stages of infection may have significant bearing on outcome.
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Affiliation(s)
- Ruth Stavrum
- Section of Microbiology and Immunology, the Gade Institute, University of Bergen, Bergen, Norway.
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Lin KW, Li J, Finn PW. Emerging pathways in asthma: innate and adaptive interactions. Biochim Biophys Acta Gen Subj 2011; 1810:1052-8. [PMID: 21596099 DOI: 10.1016/j.bbagen.2011.04.015] [Citation(s) in RCA: 12] [Impact Index Per Article: 0.9] [Reference Citation Analysis] [Abstract] [Journal Information] [Subscribe] [Scholar Register] [Received: 01/27/2011] [Revised: 03/17/2011] [Accepted: 04/28/2011] [Indexed: 12/19/2022]
Abstract
BACKGROUND Allergic asthma is a complex and chronic airway inflammatory disorder, and the prevalence of asthma has increased. Adaptive antigen-dependent immunity is a classical pathway of asthmatic pathology. Recent studies have focused on innate antigen-independent immunity in asthma. SCOPE OF REVIEW This review discusses updated research associating innate immunity with allergic asthma. We focus on innate molecules (Toll-like receptors and nucleotide-binding oligomerization domain-like receptors) and review studies regarding innate and adaptive interactions in allergic responses (surfactant protein D, lipopolysaccharide, and early life immune responses). We also highlight new emerging concepts in the field applicable to innate immunity and asthma. MAJOR CONCLUSIONS Innate immunity plays a key role in asthma. Understanding innate and adaptive interactions provide significant information in asthmatic research. Innate molecules not only contribute to classical pulmonary defense, but also modulate inflammatory responses. Emerging concepts in the analysis of the microbiome, microRNA and autophagy may provide new insights in searching therapeutic targets. GENERAL SIGNIFICANCE Finding specific mechanisms of innate and/or adaptive immunity in asthma are timely goals for further research. Integration of bioinformatics and systems biology tools, particularly in relation to microbiome analysis, may be helpful in providing an understanding to allergic immune responses. This article is part of a Special Issue entitled Biochemistry of Asthma.
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Affiliation(s)
- Ko-Wei Lin
- Division of Pulmonary and critical Care Medicine, Department of Medicine, School of Medicine, University of California, San Diego, La Jolla, CA 92093, USA
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Park HJ, Lee SJ, Kim SH, Han J, Bae J, Kim SJ, Park CG, Chun T. IL-10 inhibits the starvation induced autophagy in macrophages via class I phosphatidylinositol 3-kinase (PI3K) pathway. Mol Immunol 2010; 48:720-7. [PMID: 21095008 DOI: 10.1016/j.molimm.2010.10.020] [Citation(s) in RCA: 113] [Impact Index Per Article: 7.5] [Reference Citation Analysis] [Abstract] [Journal Information] [Subscribe] [Scholar Register] [Received: 08/07/2010] [Revised: 10/25/2010] [Accepted: 10/25/2010] [Indexed: 11/16/2022]
Abstract
Autophagy is an important process which maintains cellular homeostasis under stressful conditions such as starvation and pathogenic invasion. Previous observations have indicated that several cytokines are important regulators of the autophagic process. Among the various cytokines, IL-10 has a unique property which functions to suppress overall immunity. However, the functional role of IL-10 during the autophagic process has not been studied. In this study, we examined the effect of IL-10 during starvation induced autophagy of murine macrophages (J774). The results clearly indicated that IL-10 and IL-10 receptor signaling inhibits autophagy induction of murine macrophage. Further experiments revealed that IL-10 activates the class I phosphatidylinositol 3-kinase (PI3K) pathway, which results in the phosphorylation of p70S6K through the activation of Akt and a mammalian target of the rapamycin complex 1 (mTORC 1). These results will advance our understanding of the physiological function of IL-10 during the autophagic process of macrophage.
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Affiliation(s)
- Hun-Jung Park
- College of Life Sciences and Biotechnology, Korea University, Seoul 136-701, Republic of Korea
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17
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Brest P, Corcelle E, Cesaro A, Chargui A, Belaïd A, Klionsky D, Vouret-Craviari V, Hebuterne X, Hofman P, Mograbi B. Autophagy and Crohn's disease: at the crossroads of infection, inflammation, immunity, and cancer. Curr Mol Med 2010; 10:486-502. [PMID: 20540703 PMCID: PMC3655526 DOI: 10.2174/156652410791608252] [Citation(s) in RCA: 59] [Impact Index Per Article: 3.9] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 08/05/2009] [Accepted: 11/13/2009] [Indexed: 12/11/2022]
Abstract
Inflammatory bowel diseases (IBD) are common inflammatory disorders of the gastrointestinal tract that include ulcerative colitis (UC) and Crohn's disease (CD). The incidences of IBD are high in North America and Europe, affecting as many as one in 500 people. These diseases are associated with high morbidity and mortality. Colorectal cancer risk is also increased in IBD, correlating with inflammation severity and duration. IBD are now recognized as complex multigenetic disorders involving at least 32 different risk loci. In 2007, two different autophagy-related genes, ATG16L1 (autophagy-related gene 16-like 1) and IRGM (immunity-related GTPase M) were shown to be specifically involved in CD susceptibility by three independent genome-wide association studies. Soon afterwards, more than forty studies confirmed the involvement of ATG16L1 and IRGM variants in CD susceptibility and gave new information on the importance of macroautophagy (hereafter referred to as autophagy) in the control of infection, inflammation, immunity and cancer. In this review, we discuss how such findings have undoubtedly changed our understanding of CD pathogenesis. A unifying autophagy model then emerges that may help in understanding the development of CD from bacterial infection, to inflammation and finally cancer. The Pandora's box is now open, releasing a wave of hope for new therapeutic strategies in treating Crohn's disease.
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Affiliation(s)
- P. Brest
- Inserm ERI-21/EA 4319, Faculty of Medicine, University of Nice Sophia Antipolis, Nice, France
| | - E.A. Corcelle
- Apoptosis Department and Centre for Genotoxic Stress Research, Institute of Cancer Biology, Danish Cancer Society, Copenhagen, Denmark
| | - A. Cesaro
- Inserm ERI-21/EA 4319, Faculty of Medicine, University of Nice Sophia Antipolis, Nice, France
| | - A. Chargui
- Inserm ERI-21/EA 4319, Faculty of Medicine, University of Nice Sophia Antipolis, Nice, France
| | - A. Belaïd
- Inserm ERI-21/EA 4319, Faculty of Medicine, University of Nice Sophia Antipolis, Nice, France
| | - D.J. Klionsky
- University of Michigan, Life Sciences Institute, Ann Arbor, Michigan, USA
| | - V. Vouret-Craviari
- Inserm ERI-21/EA 4319, Faculty of Medicine, University of Nice Sophia Antipolis, Nice, France
| | - X. Hebuterne
- Inserm ERI-21/EA 4319, Faculty of Medicine, University of Nice Sophia Antipolis, Nice, France
- Centre Hospitalier Universitaire de Nice, Pôle Digestif, Hôpital L'Archet II, Nice, France
| | - P. Hofman
- Inserm ERI-21/EA 4319, Faculty of Medicine, University of Nice Sophia Antipolis, Nice, France
- Laboratory of Clinical and Experimental Pathology, Pasteur Hospital, Nice, France
| | - B. Mograbi
- Inserm ERI-21/EA 4319, Faculty of Medicine, University of Nice Sophia Antipolis, Nice, France
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18
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Xu Y, Eissa NT. Autophagy in innate and adaptive immunity. PROCEEDINGS OF THE AMERICAN THORACIC SOCIETY 2010; 7:22-8. [PMID: 20160145 PMCID: PMC3137146 DOI: 10.1513/pats.200909-103js] [Citation(s) in RCA: 53] [Impact Index Per Article: 3.5] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Subscribe] [Scholar Register] [Received: 09/22/2009] [Accepted: 10/12/2009] [Indexed: 12/16/2022]
Abstract
Autophagy (self-eating) is an evolutionary conserved simple process by which cells target their own cellular organelles and long-lived proteins for degradation. Recently, this simple ancient process has proved to be involved in many biological aspects, including host defense, cell survival and death, innate and adaptive immunity, and cancer. The implications of aberrant regulation of autophagy in human diseases are just beginning to unravel. This is a brief review of recent progress in the association of autophagy with innate and adaptive immunity relevant to lung biology and disease.
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Affiliation(s)
- Yi Xu
- Department of Medicine, Baylor College of Medicine, Houston, Texas
| | - N. Tony Eissa
- Department of Medicine, Baylor College of Medicine, Houston, Texas
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19
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Kimura A, Ishida Y, Wada T, Hisaoka T, Morikawa Y, Sugaya T, Mukaida N, Kondo T. The absence of interleukin-6 enhanced arsenite-induced renal injury by promoting autophagy of tubular epithelial cells with aberrant extracellular signal-regulated kinase activation. THE AMERICAN JOURNAL OF PATHOLOGY 2009; 176:40-50. [PMID: 20008137 DOI: 10.2353/ajpath.2010.090146] [Citation(s) in RCA: 35] [Impact Index Per Article: 2.2] [Reference Citation Analysis] [Abstract] [Track Full Text] [Subscribe] [Scholar Register] [Indexed: 12/28/2022]
Abstract
Sodium arsenite (NaAs)-induced autophagic cell death (ACD) of a mouse renal tubular epithelial cell line (mProx24), which expresses enhanced levels of interleukin-6 (IL-6), was reduced by the suppression of autophagy by 3-methyladenine or Atg7 knockdown. The inhibition of the IL-6/signal transducer and activator of transcription 3 (STAT3) signal pathway by anti-IL-6 antibody or a Jak2 inhibitor (AG490) exaggerated ACD of mProx24 cells after NaAs challenge, attenuating STAT3 activation and reciprocally enhancing extracellular signal-regulated kinase (ERK) phosphorylation. In contrast, an ERK inhibitor, PD98059, reduced NaAs-induced ACD in mProx24 cells. Subcutaneous injection of NaAs (12.5 mg/kg) into BALB/c (wild-type) mice enhanced intrarenal expression of IL-6, mainly produced by tubular cells, and caused severe renal injury characterized by hemorrhages, acute tubular necrosis, cast formation, and brush border disappearance, with increases in serum urea nitrogen (blood urea nitrogen) and creatinine levels. In addition, IL-6-deficient (IL-6(-/-)) mice exhibited exaggerated histopathological changes with higher blood urea nitrogen and creatinine levels. Moreover, in IL-6(-/-) mice treated with NaAs, ACD in renal tubular cells was significantly augmented, along with diminished STAT3 activation and reciprocal enhancement of ERK signaling, compared with wild-type mice. Finally, the administration of exogenous IL-6 into wild-type mice significantly reduced NaAs-induced ACD along with diminished ERK activation and eventually alleviated acute renal dysfunction. Thus, IL-6/STAT3 signal pathway could inhibit ERK activation, a crucial step for ACD, eventually attenuating NaAs-induced renal dysfunction.
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Affiliation(s)
- Akihiko Kimura
- Department of Forensic Medicine, Wakayama Medical University, Wakayama 641-8509, Japan
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20
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Delgado M, Singh S, De Haro S, Master S, Ponpuak M, Dinkins C, Ornatowski W, Vergne I, Deretic V. Autophagy and pattern recognition receptors in innate immunity. Immunol Rev 2009; 227:189-202. [PMID: 19120485 DOI: 10.1111/j.1600-065x.2008.00725.x] [Citation(s) in RCA: 143] [Impact Index Per Article: 8.9] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/21/2022]
Abstract
Autophagy is a physiologically and immunologically controlled intracellular homeostatic pathway that sequesters and degrades cytoplasmic targets including macromolecular aggregates, cellular organelles such as mitochondria, and whole microbes or their products. Recent advances show that autophagy plays a role in innate immunity in several ways: (i) direct elimination of intracellular microbes by digestion in autolysosomes, (ii) delivery of cytosolic microbial products to pattern recognition receptors (PRRs) in a process referred to as topological inversion, and (iii) as an anti-microbial effector of Toll-like receptors and other PRR signaling. Autophagy eliminates pathogens in vitro and in vivo but, when aberrant due to mutations, contributes to human inflammatory disorders such as Crohn's disease. In this review, we examine these relationships and propose that autophagy is one of the most ancient innate immune defenses that has possibly evolved at the time of alpha-protobacteria-pre-eukaryote relationships, leading up to modern eukaryotic cell-mitochondrial symbiosis, and that during the metazoan evolution, additional layers of immunological regulation have been superimposed and integrated with this primordial innate immunity mechanism.
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Affiliation(s)
- Monica Delgado
- Department of Molecular Genetics and Microbiology, University of New Mexico Health Sciences Center, Albuquerque, NM 87131, USA
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21
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Hsieh YC, Athar M, Chaudry IH. When apoptosis meets autophagy: deciding cell fate after trauma and sepsis. Trends Mol Med 2009; 15:129-38. [PMID: 19231289 DOI: 10.1016/j.molmed.2009.01.002] [Citation(s) in RCA: 94] [Impact Index Per Article: 5.9] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 11/24/2008] [Revised: 01/09/2009] [Accepted: 01/09/2009] [Indexed: 02/08/2023]
Abstract
Apoptotic cell death is considered to be an underlying mechanism in immunosuppression and multiple organ dysfunction after trauma-hemorrhage and sepsis. Although studied intensively over the last decade, the role of other cell death mechanisms under similar pathophysiological conditions has remained elusive. Recently, autophagy has emerged as an important mediator of programmed cell death pathways. Here, we review recent advances in our understanding of apoptosis and autophagy and the crosstalk between these processes. We explore the coexistence of these two processes and the effects of autophagy on apoptosis after trauma-hemorrhage and sepsis. The inter-relationship between autophagy and apoptosis might unveil novel therapeutic approaches for the detection and treatment of trauma-hemorrhage and sepsis.
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Affiliation(s)
- Ya-Ching Hsieh
- Department of Medical Research, E-Da Hospital, I-Shou University, Kaohsiung, Taiwan, ROC
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22
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Withaferin A inhibits iNOS expression and nitric oxide production by Akt inactivation and down-regulating LPS-induced activity of NF-kappaB in RAW 264.7 cells. Eur J Pharmacol 2008; 599:11-7. [PMID: 18838070 DOI: 10.1016/j.ejphar.2008.09.017] [Citation(s) in RCA: 75] [Impact Index Per Article: 4.4] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 04/07/2008] [Revised: 08/26/2008] [Accepted: 09/04/2008] [Indexed: 12/11/2022]
Abstract
Induction of inducible nitric oxide synthase (iNOS) expression and nitric oxide (NO) production is thought to have beneficial immunomodulatory effects in acute and chronic inflammatory disorders. In Raw 264.7 cells stimulated with lipopolysaccharide (LPS) to mimic inflammation, withaferin A inhibited LPS-induced expression of both iNOS protein and mRNA in a dose-dependent manner. To investigate the mechanism by which withaferin A inhibits iNOS gene expression, we examined activation of mitogen-activated protein kinases (MAPKs) and Akt in Raw 264.7 cells. We did not observe any significant changes in the phosphorylation of p38 MAPK in cells treated with LPS alone or LPS plus withaferin A. However, LPS-induced Akt phosphorylation was markedly inhibited by withaferin A, while the phosphorylation of p42/p44 extracellular signal-regulated kinases (ERKs) was slightly inhibited by withaferin A treatment. Withaferin A prevented IkappaB phosphorylation, blocking the subsequent nuclear translocation of nuclear factor-kappaB (NF-kappaB) and inhibiting its DNA binding activity. LPS-induced p65 phosphorylation, which is mediated by extracellular signal-regulated kinase (ERK) and Akt pathways, was attenuated by withaferin A treatment. Moreover, LPS-induced NO production and NF-kappaB activation were inhibited by SH-6, a specific inhibitor of Akt. Taken together, these results suggest that withaferin A inhibits inflammation through inhibition of NO production and iNOS expression, at least in part, by blocking Akt and subsequently down-regulating NF-kappaB activity.
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23
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Farghaly HSM, Blagbrough IS, Medina-Tato DA, Watson ML. Interleukin 13 increases contractility of murine tracheal smooth muscle by a phosphoinositide 3-kinase p110delta-dependent mechanism. Mol Pharmacol 2008; 73:1530-7. [PMID: 18276774 DOI: 10.1124/mol.108.045419] [Citation(s) in RCA: 29] [Impact Index Per Article: 1.7] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/11/2022] Open
Abstract
The Th2 cytokine interleukin (IL) 13 can elicit a number of responses consistent with a key role in the pathogenesis of asthma. We have used pharmacological and genetic approaches to demonstrate the role of signaling via the class I phosphoinositide 3-kinase p110delta isoform in IL-13-induced hyper-responsiveness of murine tracheal smooth muscle contractility in vitro. IL-13 treatment of tracheal tissue is associated with an early activation of phosphoinositide 3-kinase (PI3K), as assessed by phosphorylation of Akt. Tracheal smooth muscle contractility is enhanced by overnight incubation with IL-13, resulting in increased maximal contractions (E(max)) to carbachol (CCh) and KCl. Inhibition of PI3K by the non-isoform-selective inhibitors wortmannin or 2-(4-morpholinyl)-8-phenyl-4H-1-benzopyran-4-one (LY294002), or the selective inhibitor of the PI3K p110delta isoform 2-(6-aminopurin-9-ylmethyl)-5-methyl-3-O-tolyl-3H-quinazolin-4-one (IC87114), prevented IL-13-induced hyper-responsiveness. Consistent with a role for PI3K p110delta in IL-13-induced hyper-responsiveness, IL-13 was unable to induce hyper-responsiveness in tissues from mice expressing the catalytically inactive form of p110delta (p110delta(D910A)). These data indicate that IL-13 contributes to tracheal smooth muscle hyper-responsiveness via the PI3K p110delta isoform. In addition to previously reported effects on airway inflammation, inhibition of PI3K p110delta may be a useful target for the treatment of asthma by preventing IL-13-induced airway smooth muscle hyper-responsiveness.
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Affiliation(s)
- Hanan S M Farghaly
- Department of Pharmacy and Pharmacology, University of Bath, Bath BA2 7AY, UK
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24
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Abstract
Cells digest portions of their interiors in a process known as autophagy to recycle nutrients, remodel and dispose of unwanted cytoplasmic constituents. This ancient pathway, conserved from yeast to humans, is now emerging as a central player in the immunological control of bacterial, parasitic and viral infections. The process of autophagy may degrade intracellular pathogens, deliver endogenous antigens to MHC-class-II-loading compartments, direct viral nucleic acids to Toll-like receptors and regulate T-cell homeostasis. This Review describes the mechanisms of autophagy and highlights recent advances relevant to the role of autophagy in innate and adaptive immunity.
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Affiliation(s)
- Beth Levine
- Departments of Internal Medicine and Microbiology, University of Texas Southwestern Medical Center, Dallas, 75390 Texas USA
| | - Vojo Deretic
- Departments of Molecular Genetics and Microbiology, University of New Mexico School of Medicine, Albuquerque, 87131 New Mexico USA
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25
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Lleo A, Invernizzi P, Selmi C, Coppel RL, Alpini G, Podda M, Mackay IR, Gershwin ME. Autophagy: highlighting a novel player in the autoimmunity scenario. J Autoimmun 2007; 29:61-8. [PMID: 17693057 PMCID: PMC2063509 DOI: 10.1016/j.jaut.2007.06.003] [Citation(s) in RCA: 82] [Impact Index Per Article: 4.6] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 05/29/2007] [Revised: 06/19/2007] [Accepted: 06/20/2007] [Indexed: 11/26/2022]
Abstract
Autophagy is a physiological cellular mechanism that degrades and recycles proteins and other molecules to maintain an adequate amino acid level during nutritional starvation of the cell. Autophagy is involved in cellular homeostasis and differentiation, as well as in tissue remodeling, aging, cancer, and other diseases. Under particular environmental conditions, autophagy can also be a contributor to programmed cell death, or can act as a defense mechanism for the elimination of intracellular bacteria and viruses. According to recent experimental data, autophagy may be implicated in autoimmunity by promotion of major histocompatibility complex (MHC) class II presentation of cytosolic antigens and control of T lymphocyte homeostasis, and its induction by Th1 cytokines and perhaps by specific serum autoantibodies. We review herein the role of autophagy in immune function and its possible contribution to breakdown of tolerance and development of autoimmunity.
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Affiliation(s)
- Ana Lleo
- Division of Rheumatology, Allergy and Clinical Immunology, University of California at Davis, Davis, CA 95616, USA.
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26
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Shao L, Guo Z, Geller DA. Transcriptional suppression of cytokine-induced iNOS gene expression by IL-13 through IRF-1/ISRE signaling. Biochem Biophys Res Commun 2007; 362:582-6. [PMID: 17723228 PMCID: PMC2025693 DOI: 10.1016/j.bbrc.2007.07.203] [Citation(s) in RCA: 9] [Impact Index Per Article: 0.5] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 07/25/2007] [Accepted: 07/30/2007] [Indexed: 10/22/2022]
Abstract
IL-13 has been reported as one of the major down-regulators of iNOS expression in various tissues and cells. The molecular mechanism of iNOS suppression by IL-13 remains unclear, especially at the transcriptional stage. In this study, we found that IL-13 inhibited the expression of iNOS mRNA, protein, and NO product in a concentration-dependent manner for cytokine-stimulated rat hepatocytes. The most effective dose for IL-13 inhibitory effect is approximately 5 ng/ml. IL-13 also decreased the rat iNOS transcriptional activity by promoter analysis, but had no effect on iNOS mRNA stability. By using TranSignal Protein/DNA Combo Array, we identified cytokine-stimulated IRF-1/ISRE binding that was decreased by the addition of IL-13. Gel shift assay confirmed that IL-13 reduced the IRF-1/ISRE binding at nucleotides -913 to -923 of the rat iNOS promoter. Western blot revealed that IL-13 diminished the relative amount of IRF-1 protein translocated to the nucleus. Our data demonstrate that IL-13 down-regulates the cytokine-induced iNOS transcription by decreasing iNOS specific IRF-1/ISRE binding activity.
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Affiliation(s)
- Lifang Shao
- Department of Surgery, University of Pittsburgh, Pittsburgh, PA 15261, USA
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27
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Kolios G, Manousou P, Bourikas L, Notas G, Tsagarakis N, Mouzas I, Kouroumalis E. Ciprofloxacin inhibits cytokine-induced nitric oxide production in human colonic epithelium. Eur J Clin Invest 2006; 36:720-9. [PMID: 16968468 DOI: 10.1111/j.1365-2362.2006.01710.x] [Citation(s) in RCA: 21] [Impact Index Per Article: 1.1] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Indexed: 12/18/2022]
Abstract
BACKGROUND The fluoroquinolone ciprofloxacin is a broad-spectrum antibiotic that has been used in the treatment of inflammatory bowel diseases. There is evidence that quinolones have immunomodulating activities via the regulation of cytokine production. MATERIALS AND METHODS We investigated the effect of ciprofloxacin on the nitric oxide (NO) production by colonic epithelium. HT-29 cells and colonic biopsies from patients (n = 4) with ulcerative colitis (UC) and normal controls (n = 4) were cultured with various concentrations of ciprofloxacin (10-100 microg mL(-1)) in the presence and absence of pro-inflammatory cytokines. The production of NO was measured in culture supernatants with a spectrophotometric method and inducible nitric oxide synthase (iNOS) mRNA expression was examined by reverse transcription-polymerase chain reaction (RT-PCR). RESULTS Ciprofloxacin did not have any effect on the basal NO production by HT-29 cells. In contrast, ciprofloxacin significantly (P < 0.001) inhibited the pro-inflammatory cytokines (interleukin-1alpha + tumour necrosis factor-alpha + interferon-gamma)-induced NO production in HT-29, in a concentration-dependent manner, via the inhibition of the cytokine-induced iNOS mRNA expression. Wortmannin produced a concentration related reversal of the inhibitory effect of ciprofloxacin at both iNOS mRNA expression and NO production in HT-29 cells. A similar inhibitory effect of ciprofloxacin on the cytokine-induced NO production and iNOS mRNA expression was detected in vitro in cultures of normal colonic tissue. In addition, ciprofloxacin significantly inhibited the NO production and iNOS mRNA expression in cultures of colonic tissue from ulcerative colitis patients, in a concentration-dependent manner. CONCLUSIONS These data suggest that ciprofloxacin, in addition to its antimicrobial role, might have an immunoregulatory effect on intestinal inflammation, via the modulation of inflammatory mediators.
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Affiliation(s)
- G Kolios
- Department of Gastroenterology, Faculty of Medicine, University of Crete, Heraklion, Greece.
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28
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Deretic V. Autophagy as an immune defense mechanism. Curr Opin Immunol 2006; 18:375-82. [PMID: 16782319 DOI: 10.1016/j.coi.2006.05.019] [Citation(s) in RCA: 133] [Impact Index Per Article: 7.0] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Journal Information] [Subscribe] [Scholar Register] [Received: 03/14/2006] [Accepted: 05/05/2006] [Indexed: 12/19/2022]
Abstract
Autophagy is a homeostatic process whereby cytosol or intracellular organelles are sequestered by a double membrane structure termed autophagosome for subsequent delivery to lysosomes and degradation. Autophagy takes part in cell survival and death and has been implicated in development, aging, neurodegeneration and cancer. The newly discovered specialized role of autophagy in immune cells expands autophagic functions to defense against intracellular pathogens. Furthermore, autophagy is involved in acquired immunity, such as antigen processing for MHC II presentation, and is modulated by cytokines such as IFN-gamma. A further link has emerged between autophagy and defense against intracellular pathogens via the immunity-related GTPase lrgm1 (LRG-47), which has a protective role against Mycobacterium tuberculosis. We propose the term immunophagy for these defense processes.
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Affiliation(s)
- Vojo Deretic
- Department of Molecular Genetics and Microbiology, University of New Mexico School of Medicine, Albuquerque, NM 87131, USA.
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29
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Fichtner-Feigl S, Strober W, Kawakami K, Puri RK, Kitani A. IL-13 signaling through the IL-13alpha2 receptor is involved in induction of TGF-beta1 production and fibrosis. Nat Med 2005; 12:99-106. [PMID: 16327802 DOI: 10.1038/nm1332] [Citation(s) in RCA: 706] [Impact Index Per Article: 35.3] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 03/15/2005] [Accepted: 10/26/2005] [Indexed: 12/14/2022]
Abstract
Interleukin (IL)-13 is a major inducer of fibrosis in many chronic infectious and autoimmune diseases. In studies of the mechanisms underlying such induction, we found that IL-13 induces transforming growth factor (TGF)-beta(1) in macrophages through a two-stage process involving, first, the induction of a receptor formerly considered to function only as a decoy receptor, IL-13Ralpha(2). Such induction requires IL-13 (or IL-4) and tumor necrosis factor (TNF)-alpha. Second, it involves IL-13 signaling through IL-13Ralpha(2) to activate an AP-1 variant containing c-jun and Fra-2, which then activates the TGFB1 promoter. In vivo, we found that prevention of IL-13Ralpha(2) expression reduced production of TGF-beta(1) in oxazolone-induced colitis and that prevention of IL-13Ralpha(2) expression, Il13ra2 gene silencing or blockade of IL-13Ralpha(2) signaling led to marked downregulation of TGF-beta(1) production and collagen deposition in bleomycin-induced lung fibrosis. These data suggest that IL-13Ralpha(2) signaling during prolonged inflammation is an important therapeutic target for the prevention of TGF-beta(1)-mediated fibrosis.
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Affiliation(s)
- Stefan Fichtner-Feigl
- Mucosal Immunity Section, Laboratory of Host Defenses, National Institute of Allergy and Infectious Diseases, National Institutes of Health, Building 10-CRC 5W3864, 10 Center Drive, Bethesda, Maryland 20892, USA
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30
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Kolios G, Valatas V, Ward SG. Nitric oxide in inflammatory bowel disease: a universal messenger in an unsolved puzzle. Immunology 2005; 113:427-37. [PMID: 15554920 PMCID: PMC1782592 DOI: 10.1111/j.1365-2567.2004.01984.x] [Citation(s) in RCA: 383] [Impact Index Per Article: 19.2] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/15/2022] Open
Abstract
In recent years, nitric oxide (NO), a gas previously considered to be a potentially toxic chemical, has been established as a diffusible universal messenger that mediates cell-cell communication throughout the body. Constitutive and inducible NO production regulate numerous essential functions of the gastrointestinal mucosa, such as maintenance of adequate perfusion, regulation of microvascular and epithelial permeability, and regulation of the immune response. Up-regulation of the production of NO via expression of inducible nitric oxide synthase (iNOS) represents part of a prompt intestinal antibacterial response; however, NO has also been associated with the initiation and maintenance of inflammation in human inflammatory bowel disease (IBD). Recent studies on animal models of experimental IBD have shown that constitutive and inducible NO production seems to be beneficial during acute colitis, but sustained up-regulation of NO is detrimental. This fact is also supported by studies on mice genetically deficient in various NOS isoforms. However, the mechanism by which NO proceeds from being an indispensable homeostatic regulator to a harmful destructor remains unknown. Furthermore, extrapolation of data from animal colitis models to human IBD is questionable. The purpose of this review is to update our knowledge about the role of this universal mediator and the enzymes that generate it in the pathogenesis of IBD.
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Affiliation(s)
- George Kolios
- Department of Gastroenterology, Faculty of Medicine, University of Crete, Heraklion, Greece.
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31
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Valatas V, Kolios G, Manousou P, Xidakis C, Notas G, Ljumovic D, Kouroumalis EA. Secretion of inflammatory mediators by isolated rat Kupffer cells: the effect of octreotide. ACTA ACUST UNITED AC 2005; 120:215-25. [PMID: 15177940 DOI: 10.1016/j.regpep.2004.03.009] [Citation(s) in RCA: 36] [Impact Index Per Article: 1.8] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 09/23/2003] [Revised: 03/11/2004] [Accepted: 03/15/2004] [Indexed: 01/13/2023]
Abstract
AIMS We studied the production of inflammatory mediators by rat KC and the possible in vitro effect of the somatostatin analogue octreotide. METHODS Primary KC cultures were incubated with LPS added alone or with different concentrations of octreotide. The production of TNFalpha, IL-6, IL-10, IL-12 and IL-13 was assessed in culture supernatants by ELISA and that of nitric oxide (NO) by a modification of the Griess reaction. RESULTS Isolated KC produced a basal amount of TNFalpha, IL-6, IL-12, IL-13, and NO but not IL-10. LPS-stimulated KC secreted significantly increased amounts of TNFalpha (P < 0.001), IL-6 (P < 0.01), IL-10 (P < 0.001), IL-12 (P < 0.01), and NO (P < 0.001) whereas IL-13 production remained constant. Octreotide reduced IL-12 (P < 0.05) and increased IL-13 (P < 0.05) production by unstimulated KC. Furthermore, octreotide suppressed TNFalpha production (P < 0.05), without modifying TNFalpha mRNA expression and decreased iNOS expression and NO (P approximately 0.05) production by LPS-activated KC. These effects were reversed with Wortmannin pre-treatment suggesting that octreotide may act via interference with phosphatidylinositol 3-kinase pathways. CONCLUSIONS These data demonstrate that KC is a source of multiple inflammatory mediators, indicating a critical role in liver inflammatory disorders. Octreotide modulates inflammatory mediator production by isolated KC, suggesting that it might have immunoregulatory and anti-inflammatory effects in liver diseases.
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Affiliation(s)
- Vassilis Valatas
- Gastroenterology Department, Faculty of Medicine, University of Crete, P.O. Box 2208, Heraklion, GR-71409, Greece
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Wang IM, Lin H, Goldman SJ, Kobayashi M. STAT-1 is activated by IL-4 and IL-13 in multiple cell types. Mol Immunol 2004; 41:873-84. [PMID: 15261459 DOI: 10.1016/j.molimm.2004.04.027] [Citation(s) in RCA: 41] [Impact Index Per Article: 2.0] [Reference Citation Analysis] [Abstract] [MESH Headings] [Journal Information] [Subscribe] [Scholar Register] [Received: 11/27/2003] [Revised: 01/08/2004] [Accepted: 04/15/2004] [Indexed: 01/05/2023]
Abstract
While interleukin-4 (IL-4) and interleukin-13 (IL-13) can utilize a common receptor, composed of IL-4Ralpha and IL-13Ralpha1, IL-4 can also signal through a receptor with IL-4Ralpha and the common gamma chain (gammaC) as its subunits. IL-4 and IL-13 have been reported to elicit similar biological effects in a number of settings, including stimulating Ig isotype switching to IgE and inducing chemokines and cytokines in a variety of cell types whereas, depending on the receptor expression on responder cells, differential effects such as induction of type II helper T cell differentiation by IL-4 but not by IL-13 are also well documented. Recent data suggest distinct roles for these two cytokines in the 'in vivo' pathology of airway inflammatory diseases such as asthma. In this study, we examined the possibility of differential signaling by IL-4 and IL-13 on cells of the airway, by comparing expression of receptor chains and activation of different Signal Transducer and Activator of Transcription (STAT) family members. Five primary cultured cell lines representing four non-immune human lung tissue cell types (smooth muscle, epithelium, endothelium, and fibroblast) were utilized. While we readily detected expression of IL-4 Ralpha and IL-13Ralpha1 in all five cell lines, gammaC was not detectable in any of these cell lines. Consistent with previous reports, we detected STAT-6 activation in all five airway tissue cell lines examined in response to both cytokines. In addition, we also consistently detected STAT-1 activation in all of these cells. This observation was extended to include lymphoid as well as myeloid cells that express also gammaC chain. In conclusion, while the study found no differences in STAT activation in response to the two cytokines, the data show that in addition to STAT-6 activation, STAT-1 activation is also a part of the integral signaling pathways utilized by IL-4 and IL-13.
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Affiliation(s)
- I-Ming Wang
- Wyeth Research, Inflammation Biology, 200 Cambridge Park Drive, Cambridge, MA 02140, USA
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Jinnin M, Ihn H, Yamane K, Tamaki K. Interleukin-13 stimulates the transcription of the human alpha2(I) collagen gene in human dermal fibroblasts. J Biol Chem 2004; 279:41783-91. [PMID: 15271999 DOI: 10.1074/jbc.m406951200] [Citation(s) in RCA: 63] [Impact Index Per Article: 3.0] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 01/18/2023] Open
Abstract
Interleukin (IL)-13 is a novel lymphokine produced by activated Type 2 helper cells. In this study, we examined the target genes of IL-13 by the cDNA microarray analysis in human dermal fibroblasts. We focused on the human alpha2(I) collagen gene, which was one of the IL-13-induced genes by the microarray analysis. IL-13 induced type I collagen protein as well as mRNA in a dose-dependent manner. Actinomycin D, an RNA synthesis inhibitor, significantly blocked the IL-13-mediated up-regulation of alpha2(I) collagen mRNA expression, whereas cycloheximide, a protein synthesis inhibitor, did not block this up-regulation. In addition, IL-13 treatment induced the promoter activity of alpha2(I) collagen by nuclear run-on transcription assay and chloramphenicol acetyltransferase assay. IL-13-mediated transcriptional activation of alpha2(I) collagen gene or type I collagen protein up-regulation was inhibited by the treatment of fibroblasts with a selective phosphoinositide 3-kinase (PI3K) inhibitor, LY294002, or STAT6 antisense oligonucleotide, but not by PD98059, a specific inhibitor of MEK/ERK, or SB202190 or SB203580, specific inhibitors of p38 MAPK; IL-13 induced the phosphorylation of PI3K p85 regulatory subunit and STAT6. These results suggest that IL-13 may play a role in the regulation of extracellular matrix and indicate the possible therapeutic value of the blockade of IL-13 signaling pathways via PI3K and STAT6 in fibrosis.
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Affiliation(s)
- Masatoshi Jinnin
- Department of Dermatology, Faculty of Medicine, University of Tokyo, 7-3-1 Hongo, Bunkyo-ku, Tokyo 113-8655, Japan
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Hwang SY, Jung JS, Lim SJ, Kim JY, Kim TH, Cho KH, Han IO. LY294002 inhibits interferon-gamma-stimulated inducible nitric oxide synthase expression in BV2 microglial cells. Biochem Biophys Res Commun 2004; 318:691-7. [PMID: 15144893 DOI: 10.1016/j.bbrc.2004.04.082] [Citation(s) in RCA: 25] [Impact Index Per Article: 1.2] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 04/02/2004] [Indexed: 10/26/2022]
Abstract
The current study examined the potential involvement of phosphatidylinositol 3 phosphate kinase (PI3K) in interferon-gamma (IFN-gamma)-stimulated nitric oxide (NO) generation in BV2 murine microglial cells. We found that LY294002, a PI3K inhibitor, markedly reduced IFN-gamma-induced morphological changes, NO production, and cell death. The inhibitory effect of LY294002 on NO generation may be mediated through specific inhibition of signal transducer and activator-1 (STAT1) and NF-kappaB, which are activated by IFN-gamma. Induction of the mRNA for IFN-gamma-mediated interferon response factor (IRF-1) and inducible protein-10 (IP-10) was not significantly affected by LY294002, indicating that suppression of PI3K may not be sufficient for downregulation of these genes. Although it remains unclear how PI3K signaling is involved in IFN-gamma-mediated inflammatory reactions in the brain, our findings provide some insight into the inflammatory mechanisms of IFN-gamma in the brain and suggest that regulators of the PI3K pathway may act as anti-inflammatory agents in microglia.
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Affiliation(s)
- So-Young Hwang
- Research Institute, National Cancer Center, Goyang, Gyeonggi, Republic of Korea
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Kim WK, Hwang SY, Oh ES, Piao HZ, Kim KW, Han IO. TGF-β1 Represses Activation and Resultant Death of Microglia via Inhibition of Phosphatidylinositol 3-Kinase Activity. THE JOURNAL OF IMMUNOLOGY 2004; 172:7015-23. [PMID: 15153523 DOI: 10.4049/jimmunol.172.11.7015] [Citation(s) in RCA: 96] [Impact Index Per Article: 4.6] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Subscribe] [Scholar Register] [Indexed: 11/19/2022]
Abstract
Overactivation of microglial cells may cause severe brain tissue damage in various neurodegenerative diseases. Therefore, the overactivation of microglia should be repressed by any means. The present study investigated the potential mechanism and signaling pathway for the repressive effect of TGF-beta1, a major anti-inflammatory cytokine, on overactivation and resultant death of microglial cells. A bacterial endotoxin LPS stimulated expression of inducible NO synthase (iNOS) and caused death in cultured microglial cells. TGF-beta1 markedly blocked these LPS effects. However, the LPS-evoked death of microglial cells was not solely attributed to excess production of NO. Because phosphatidylinositol 3-kinase (PI3K) was previously shown to play a crucial role in iNOS expression and cell survival signals, we further studied whether PI3K signaling was associated with the suppressive effect of TGF-beta1. Like TGF-beta1, the PI3K inhibitor LY294002 blocked iNOS expression and death in cultured microglial cells. Both TGF-beta1 and LY294002 decreased the activation of caspases 3 and 11 and the mRNA expression of various kinds of inflammatory molecules caused by LPS. TGF-beta1 was further found to decrease LPS-induced activation of PI3K and Akt. TGF-beta1 and LY294002 suppressed LPS-induced p38 mitogen-activated kinase and c-Jun N-terminal kinase activity. In contrast, TGF-beta1 and LY294002 enhanced LPS-induced NF-kappaB activity. Our data indicate that TGF-beta1 protect normal or damaged brain tissue by repressing overactivation of microglial cells via inhibition of PI3K and its downstream signaling molecules.
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Affiliation(s)
- Won-Ki Kim
- Department of Pharmacology, College of Medicine, Division of Molecular Life Sciences and Center for Cell Signalling Research, Ewha Womans University, Seoul, Korea
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Wright KL, Weaver SA, Patel K, Coopman K, Feeney M, Kolios G, Robertson DAF, Ward SG. Differential regulation of prostaglandin E biosynthesis by interferon-gamma in colonic epithelial cells. Br J Pharmacol 2004; 141:1091-7. [PMID: 15023855 PMCID: PMC1574887 DOI: 10.1038/sj.bjp.0705719] [Citation(s) in RCA: 15] [Impact Index Per Article: 0.7] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 01/16/2023] Open
Abstract
1. Cyclooxygenase (COX)-2 expression and activity in response to pro-inflammatory cytokines TNF alpha and IFN gamma was evaluated in the colonic epithelial cell line HT29 and the airway epithelial cell line A549. 2. TNF alpha induced concentration- and time-dependent upregulation of COX-2 mRNA, protein and prostaglandin (PG)E(2) synthesis. 3. Co-stimulation of TNF alpha with IFN gamma resulted in reduced COX-2 mRNA and protein expression. 4. IFN gamma had no effect on the stability of TNF alpha-induced COX-2 mRNA. 5. TNF alpha-induced PGE(2) biosynthesis was significantly enhanced by the simultaneous addition of IFN gamma and was COX-2 dependent. 6. The combination of IFN gamma and TNF alpha induced the microsomal prostaglandin E synthase (mPGES), comensurate with the enhanced PGE(2) synthesis. 7. These results suggest that, in terms of PGE(2) biosynthesis, IFN gamma plays a negative regulatory role at the level of COX-2 expression and a positive regulatory role at the level of mPGES expression. This may have important implications for the clinical use of IFN gamma in inflammatory diseases.
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Affiliation(s)
- Karen L Wright
- Department of Pharmacy and Pharmacology, University of Bath, Claverton Down, Bath BA2 7AY
| | - Sean A Weaver
- Department of Pharmacy and Pharmacology, University of Bath, Claverton Down, Bath BA2 7AY
| | - Kajal Patel
- Department of Pharmacy and Pharmacology, University of Bath, Claverton Down, Bath BA2 7AY
| | - Karen Coopman
- Department of Pharmacy and Pharmacology, University of Bath, Claverton Down, Bath BA2 7AY
| | - Mark Feeney
- Department of Gastroenterology, Royal United Hospital, Bath BA1 3NG
| | - George Kolios
- Faculty of Medicine, University of Crete, P.O. Box 2208, Heraklion, Crete, GR-71003, Greece
| | | | - Stephen G Ward
- Department of Pharmacy and Pharmacology, University of Bath, Claverton Down, Bath BA2 7AY
- Author for correspondence:
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Monick MM, Robeff PK, Butler NS, Flaherty DM, Carter AB, Peterson MW, Hunninghake GW. Phosphatidylinositol 3-kinase activity negatively regulates stability of cyclooxygenase 2 mRNA. J Biol Chem 2002; 277:32992-3000. [PMID: 12072439 DOI: 10.1074/jbc.m203218200] [Citation(s) in RCA: 74] [Impact Index Per Article: 3.2] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/06/2022] Open
Abstract
Human alveolar macrophages have both lipopolysaccharide (LPS)-induced and constitutive phosphatidylinositol 3-kinase (PI3K) activity. We observed that blocking PI3K activity increased release of prostaglandin E2 after LPS exposure, and increasing PI3K activity (interleukin-13) decreased release of prostaglandin E2 after LPS exposure. This was not because of an effect of PI3K on phospholipase 2 activity. PI3K inhibition resulted in an increase in cyclooxygenase 2 (COX2) protein, mRNA, and mRNA stability. PI3K negatively regulated activation of the p38 pathway (p38, MKK3/6, and MAPKAP2), and an active p38 was necessary for COX2 production. The data suggest that PI3K inhibition of p38 modulates COX2 expression via destabilization of LPS-induced COX2 mRNA.
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Affiliation(s)
- Martha M Monick
- University of Iowa Roy J. and Lucille A. Carver College of Medicine and Veterans Administration Medical Center, Iowa City, Iowa 52242, USA.
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Yang MS, Park EJ, Sohn S, Kwon HJ, Shin WH, Pyo HK, Jin B, Choi KS, Jou I, Joe EH. Interleukin-13 and -4 induce death of activated microglia. Glia 2002; 38:273-80. [PMID: 12007140 DOI: 10.1002/glia.10057] [Citation(s) in RCA: 85] [Impact Index Per Article: 3.7] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/05/2022]
Abstract
When the brain suffers injury, microglia migrate to the damaged sites and become activated. These activated microglia are not detected several days later and the mechanisms underlying their disappearance are not well characterized. In this study, we demonstrate that interleukin (IL)-13, an anti-inflammatory cytokine, selectively induces cell death of activated microglia in vitro. Cell death was detected 4 days after the coaddition of IL-13 with any one of the microglial activators, lipopolysaccharide (LPS), ganglioside, or thrombin. This cell death occurred in a time-dependent manner. LPS, ganglioside, thrombin, or IL-13 alone did not induce cell death. Among anti-inflammatory cytokines, IL-4 mimicked the effect of IL-13, while TGF-beta did not. Cells treated with IL-13 plus LPS, or IL-13 plus ganglioside, showed the characteristics of apoptosis when analyzed by electron microscopy and terminal deoxynucleotidyl transferase-mediated dUTP nick end labeling staining. Electron micrographs also showed microglia engulfing neighboring dead cells. We propose that IL-13 and IL-4 induce death of activated microglia, and that this process is important for prevention of chronic inflammation that can cause tissue damage.
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Affiliation(s)
- Myung-Soon Yang
- Department of Pharmacology, Ajou University School of Medicine, Suwon, Korea
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Abstract
Carcinoma of the gastrointestinal tract is the most common internal malignancy affecting men and women in Western countries. Chronic intestinal inflammation, especially of the colon, is also a Western disease and correlates with a significantly increased risk of developing cancer. This has suggested that the immune processes involved in both conditions might share some common pathways. Indeed, there is increasing evidence that phosphatidylinositol 3-kinases (PI 3-kinases) are involved in both the pathogenesis of colorectal carcinoma and intestinal inflammation. Here, we discuss this rapidly progressing area of research, presenting evidence for a pivotal role of PI 3-kinase(s) in intestinal pathophysiology.
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Affiliation(s)
- S A Weaver
- Department of Pharmacy and Pharmacology, University of Bath, Claverton Down, Bath, UK BA2 7AY
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40
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Zeng C, Morrison AR. Disruption of the actin cytoskeleton regulates cytokine-induced iNOS expression. Am J Physiol Cell Physiol 2001; 281:C932-40. [PMID: 11502570 DOI: 10.1152/ajpcell.2001.281.3.c932] [Citation(s) in RCA: 31] [Impact Index Per Article: 1.3] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/22/2022]
Abstract
Interleukin-1beta (IL-1beta) induces the inducible nitric oxide synthase (iNOS), resulting in the release of nitric oxide (NO) from glomerular mesangial cells. In this study, we demonstrated that disruption of F-actin formation by sequestration of G-actin with the toxin latrunculin B (LatB) dramatically potentiated IL-1beta-induced iNOS protein expression in a dose-dependent manner. LatB by itself had little or no effect on iNOS expression. Staining of F-actin with nitrobenzoxadiazole (NBD)-phallacidin demonstrated that LatB significantly impaired F-actin stress fiber formation. Jasplakinolide (Jasp), which binds to and stabilizes F-actin, suppressed iNOS expression enhanced by LatB. These data strongly suggest that actin cytoskeletal dynamics regulates IL-1beta-induced iNOS expression. We demonstrated that LatB decreases serum response factor (SRF) activity as determined by reporter gene assays, whereas Jasp increases SRF activity. The negative correlation between SRF activity and iNOS expression suggests a negative regulatory role for SRF in iNOS expression. Overexpression of a dominant negative mutant of SRF increases the IL-1beta-induced iNOS expression, providing direct evidence that SRF inhibits iNOS expression.
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Affiliation(s)
- C Zeng
- Department of Medicine, Molecular Biology, and Pharmacology, Washington University School of Medicine, St. Louis, Missouri 63110, USA
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41
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Weaver SA, Russo MP, Wright KL, Kolios G, Jobin C, Robertson DA, Ward SG. Regulatory role of phosphatidylinositol 3-kinase on TNF-alpha-induced cyclooxygenase 2 expression in colonic epithelial cells. Gastroenterology 2001; 120:1117-27. [PMID: 11266376 DOI: 10.1053/gast.2001.23257] [Citation(s) in RCA: 54] [Impact Index Per Article: 2.3] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Indexed: 01/11/2023]
Abstract
BACKGROUND AND AIMS Cyclooxygenase (COX)-2 is up-regulated in most colonic cancers and in inflammatory bowel disease in which tumor necrosis factor (TNF)-alpha is believed to play a central role. There has been recent speculation on the activation of phosphatidylinositol 3-kinase (PI 3-kinase) by TNF-alpha and its role in the regulation of genes controlled by NF-kappaB. We investigated the regulatory role of PI 3-kinase on COX-2 expression in colonic epithelial cells. METHODS In HT-29 and Caco-2 colonic epithelial cells, COX-2 expression was induced by either TNF-alpha or interleukin (IL)-1alpha as observed by Northern and Western analyses. COX-2 activity was assessed by measuring prostaglandin E(2) (PGE2) production by enzyme-linked immunosorbent assay. NF-kappaB binding activity was assessed by electrophoretic mobility shift assay. PI 3-kinase activity was measured by quantifying the accumulation of PI 3-kinase-dependent D-3 lipid products by high-performance liquid chromatography. RESULTS The PI 3-kinase inhibitor wortmannin up-regulated induced COX-2 expression in a concentration-dependent manner in both HT-29 and Caco-2 cells. An alternative PI 3-kinase inhibitor, LY294002, caused up-regulation of induced COX-2 messenger RNA (mRNA) in HT-29 cells at concentrations of < or =1 micromol/L. IL-4 and IL-13, which are known to activate PI 3-kinase, down-regulated HT-29 COX-2 mRNA, protein, and PGE2 production. NF-kappaB binding activity was unaltered by PI 3-kinase inhibition in HT-29 cells, in which TNF-alpha was shown to activate PI 3-kinase directly. CONCLUSIONS COX-2 is negatively regulated by PI 3-kinase; we propose that the inhibitory effect of IL-4 and IL-13 is mediated via a PI 3-kinase-dependent pathway. This mechanism does not appear to involve NF-kappaB because PI 3-kinase inhibition did not alter NF-kappaB binding activity. TNF-alpha can activate PI 3-kinase directly in addition to inducing COX-2.
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Affiliation(s)
- S A Weaver
- Department of Pharmacy and Pharmacology, University of Bath, Claverton Down, Bath, BA2 7AY, England.
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Umeshita-Suyama R, Sugimoto R, Akaiwa M, Arima K, Yu B, Wada M, Kuwano M, Nakajima K, Hamasaki N, Izuhara K. Characterization of IL-4 and IL-13 signals dependent on the human IL-13 receptor alpha chain 1: redundancy of requirement of tyrosine residue for STAT3 activation. Int Immunol 2000; 12:1499-509. [PMID: 11058569 DOI: 10.1093/intimm/12.11.1499] [Citation(s) in RCA: 56] [Impact Index Per Article: 2.2] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/22/2022] Open
Abstract
IL-4 and IL-13 are pleiotropic cytokines whose biological activities overlap with each other. IL-13 receptor alpha chain 1 (IL-13R alpha 1) is necessary for binding to IL-13, and the heterodimer composed of IL-13R alpha 1 and IL-4R alpha chain transduces IL-13 and IL-4 signals; however, the functional mapping of the intracellular domain of IL-13R alpha 1 is not fully understood. In this study, we constructed wild and mutated types of human IL-13R alpha 1, and analyzed IL-4 and IL-13 signals using an IL-13R alpha 1-transfected human B cell line. Expression of IL-13R alpha 1 evoked STAT3 activation by IL-4 and IL-13, and in stimulated human B cells, on which IL-13R alpha 1 was highly expressed, IL-4 and IL-13 induced STAT3 activation. Replacement of the two tyrosine residues completely abolished STAT3 activation, although replacing either tyrosine residue alone retained it. Furthermore, we found that the Box1 region and the C-terminal tail of IL-13R alpha 1 were critical for binding to Tyk2, and activation of Jak1, Tyk2, the insulin receptor substrate-1 and STAT6 respectively. These results suggest that STAT3 activation is involved with IL-4 and IL-13 signals in human B cells along with the activation of STAT6, and that there is a unique sequence in IL-13R alpha 1 to activate STAT3.
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Affiliation(s)
- R Umeshita-Suyama
- Department of Clinical Chemistry and Laboratory Medicine, Graduate School of Medical Sciences, Kyushu University, 3-1-1 Maidashi, Higashi-ku, Fukuoka 812-8582, Japan
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Wright KL, Ward SG. Interactions between phosphatidylinositol 3-kinase and nitric oxide: explaining the paradox. MOLECULAR CELL BIOLOGY RESEARCH COMMUNICATIONS : MCBRC 2000; 4:137-43. [PMID: 11281727 DOI: 10.1006/mcbr.2001.0273] [Citation(s) in RCA: 13] [Impact Index Per Article: 0.5] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Subscribe] [Scholar Register] [Indexed: 01/08/2023]
Abstract
Nitric oxide (NO) and the many derivatives and reactive oxygen intermediates thereof are all molecules that are utilised by mammalian cells in the war against microbial pathogens and tumours. They are potentially toxic molecules and, with damage control being crucial, the production and metabolism of nitric oxide is a tightly regulated process. The duality of NO is well documented. On the one hand, beneficial effects include normal healing in the skin and intestinal mucosa, killing of certain bacteria, regulating T cell proliferation and differentiation (Th1 vs Th2), and regulating leukocyte recruitment, by affecting adhesion molecule expression. On the other hand, persistent high levels of NO can lead to the production of toxic metabolites (peroxynitrite and hydroxyls), which can have detrimental effects, such as increased microvascular and epithelial permeability, increased oxidative stress (which can damage DNA), and damage to iron-sulphur proteins in mitochondria. NO has been reported to modulate its own production and the mechanisms involved in this self-regulation are being hotly pursued. The purpose of this review is to update recent intriguing advances in our understanding of the interaction of the phosphatidylinositol (PI) 3-kinase-dependent signal transduction pathway in regulating the activity of the enzymes that generate NO, namely, the nitric oxide synthases.
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Affiliation(s)
- K L Wright
- Department of Pharmacy and Pharmacology, University of Bath, Claverton Down, Bath, BA2 7AY, United Kingdom
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Wang Q, Kim S, Wang X, Evers BM. Activation of NF-kappaB binding in HT-29 colon cancer cells by inhibition of phosphatidylinositol 3-kinase. Biochem Biophys Res Commun 2000; 273:853-8. [PMID: 10891336 DOI: 10.1006/bbrc.2000.3034] [Citation(s) in RCA: 16] [Impact Index Per Article: 0.6] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/22/2022]
Abstract
The ubiquitous transcription factor NF-kappaB, which is activated in cells by diverse stimuli including phosphatidylinositol 3-kinase (PI3-kinase), is a critical factor for cell survival and growth. Inhibition of PI3-kinase enhances enterocyte-like differentiation of the human colon cancer cell line HT-29. The purpose of our study was to determine whether PI3-kinase alters NF-kappaB in HT-29 cells. Wortmannin, a specific PI3-kinase inhibitor, stimulated NF-kappaB binding activity in HT-29 cells by 4 h after treatment. Activation of NF-kappaB occurred without degradation of IkappaBalpha, a protein that sequesters NF-kappaB in the cytosol. In addition to increasing NF-kappaB binding, either wortmannin or cotransfection with a dominant negative mutant of the p85 regulatory subunit of PI3-kinase (Deltap85) induced NF-kappaB transactivation. Taken together, these results suggest that inhibition of PI3-kinase in HT-29 cells results in induction of NF-kappaB binding activity and transactivation which is independent of IkappaBalpha degradation.
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Affiliation(s)
- Q Wang
- Department of Surgery, University of Texas Medical Branch, Galveston, Texas 77555, USA
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Castrillo A, Díaz-Guerra MJ, Hortelano S, Martín-Sanz P, Boscá L. Inhibition of IkappaB kinase and IkappaB phosphorylation by 15-deoxy-Delta(12,14)-prostaglandin J(2) in activated murine macrophages. Mol Cell Biol 2000; 20:1692-8. [PMID: 10669746 PMCID: PMC85352 DOI: 10.1128/mcb.20.5.1692-1698.2000] [Citation(s) in RCA: 222] [Impact Index Per Article: 8.9] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 02/07/2023] Open
Abstract
Activation of the macrophage cell line RAW 264.7 with lipopolysaccharide (LPS) and gamma interferon (IFN-gamma) induces the expression of gene products involved in host defense, among them type 2 nitric oxide synthase. Treatment of cells with 15-deoxy-Delta(12,14)-prostaglandin J(2) (15dPGJ(2)) inhibited the LPS- and IFN-gamma-dependent synthesis of NO, a process that was not antagonized by similar concentrations of prostaglandin J(2), prostaglandin E(2), or rosiglitazone, a peroxisomal proliferator-activated receptor gamma ligand. Incubation of activated macrophages with 15dPGJ(2) inhibited the degradation of IkappaBalpha and IkappaBbeta and increased their levels in the nuclei. NF-kappaB activity, as well as the transcription of NF-kappaB-dependent genes, such as those encoding type 2 nitric oxide synthase and cyclooxygenase 2, was impaired under these conditions. Analysis of the steps leading to IkappaB phosphorylation showed an inhibition of IkappaB kinase by 15dPGJ(2) in cells treated with LPS and IFN-gamma, resulting in an impaired phosphorylation of IkappaBalpha, at least in the serine 32 residue required for targeting and degradation of this protein. Incubation of partially purified activated IkappaB kinase with 2 microM 15dPGJ(2) reduced by 83% the phosphorylation in serine 32 of IkappaBalpha, suggesting that this prostaglandin exerts direct inhibitory effects on the activity of the IkappaB kinase complex. These results show rapid actions of 15dPGJ(2), independent of peroxisomal proliferator receptor gamma activation, in macrophages challenged with low doses of LPS and IFN-gamma.
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Affiliation(s)
- A Castrillo
- Instituto de Bioquímica (Centro Mixto CSIC-UCM), Facultad de Farmacia, Universidad Complutense, 28040 Madrid, Spain
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Petiot A, Ogier-Denis E, Blommaart EF, Meijer AJ, Codogno P. Distinct classes of phosphatidylinositol 3'-kinases are involved in signaling pathways that control macroautophagy in HT-29 cells. J Biol Chem 2000; 275:992-8. [PMID: 10625637 DOI: 10.1074/jbc.275.2.992] [Citation(s) in RCA: 942] [Impact Index Per Article: 37.7] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 01/11/2023] Open
Abstract
3-Methyladenine which stops macroautophagy at the sequestration step in mammalian cells also inhibits the phosphoinositide 3-kinase (PI3K) activity raising the possibility that PI3K signaling controls the macroautophagic pathway (Blommaart, E. F. C., Krause, U., Schellens, J. P. M., Vreeling-Sindelárová, H., and Meijer, A. J. (1997) Eur. J. Biochem. 243, 240-246). The aim of this study was to identify PI3Ks involved in the control of macroautophagic sequestration in human colon cancer HT-29 cells. An increase of class I PI3K products (phosphatidylinositol 3,4-bisphosphate and phosphatidylinositol 3,4,5-triphosphate) caused by either feeding cells with synthetic lipids (dipalmitoyl phosphatidylinositol 3, 4-bisphosphate and dipalmitoyl phosphatidylinositol 3,4, 5-triphosphate) or by stimulating the enzymatic activity by interleukin-13 reduced macroautophagy. In contrast, an increase in the class III PI3K product (phosphatidylinositol 3-phosphate), either by feeding cells with a synthetic lipid or by overexpressing the p150 adaptor, stimulates macroautophagy. Transfection of a specific class III PI3K antisense oligonucleotide greatly inhibited the rate of macroautophagy. In accordance with a role of class III PI3K, wortmannin (an inhibitor of PI3Ks) inhibits macroautophagic sequestration and protein degradation in the low nanomolar range (IC(50) 5-15 nM). Further in vitro enzymatic assay showed that 3-methyladenine inhibits the class III PI3K activity. Dipalmitoyl phosphatidylinositol 3-phosphate supplementation or p150 overexpression rescued the macroautophagic pathway in HT-29 cells overexpressing a GTPase-deficient mutant of the Galpha(i3) protein suggesting that both class III PI3K and trimeric G(i3) protein signaling are required in the control macroautophagy in HT-29 cells. In conclusion, our results demonstrate that distinct classes of PI3K control the macroautophagic pathway in opposite directions. The roles of PI3Ks in macroautophagy are discussed in the context of membrane recycling.
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Affiliation(s)
- A Petiot
- INSERM U504, Glycobiologie et Signalisation Cellulaire, 16 avenue Paul-Vaillant-Couturier, 94807 Villejuif, Cedex, France
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Edmunds C, Parry RV, Burgess SJ, Reaves B, Ward SG. CD28 stimulates tyrosine phosphorylation, cellular redistribution and catalytic activity of the inositol lipid 5-phosphatase SHIP. Eur J Immunol 1999; 29:3507-15. [PMID: 10556805 DOI: 10.1002/(sici)1521-4141(199911)29:11<3507::aid-immu3507>3.0.co;2-9] [Citation(s) in RCA: 28] [Impact Index Per Article: 1.1] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/08/2022]
Abstract
The D-3 phosphoinositide lipids phosphatidylinositol 3,4, 5-trisphophate [PtdIns(3,4,5)P(3)] and phosphatidylinositol 3, 4-bisphosphate [PtdIns(3,4)P(2)] represent upstream components of a major signaling pathway that is strongly activated by the T cell costimulatory molecule CD28. A major route for degradation of PtdIns(3,4,5)P(3) (and hence, regulation of PtdIns(3,4,5)P(3)-driven effector pathways), involves its conversion to PtdIns(3,4)P(2) by the 145-kDa SH2-containing inositol (poly)phosphate 5-phosphatase (SHIP). In this study, we demonstrate using the murine T cell hybridoma DC27.1, that SHIP is strongly tyrosine phosphorylated after ligation of CD28 by either mAb or the natural ligand B7.1. Ligation of CD3 also stimulates SHIP tyrosine phosphorylation and an additive effect on tyrosine phosphorylation of SHIP is observed when both CD3 and CD28 are ligated. The tyrosine phosphorylation of SHIP in response to CD28 ligation correlates with a marked redistribution of SHIP from the cytosol to the plasma membrane, as well as an increase in the in vitro 5-phosphatase activity associated with SHIP immunoprecipitates derived from CD28-stimulated cells. However, we have been unable to detect a direct association between CD28 and SHIP, so the mechanisms by which CD28 exerts the observed effects on SHIP remain unclear. This is the first demonstration that SHIP is a biochemical target for CD28 and suggests that SHIP may be involved in the regulation of T cell activation.
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Affiliation(s)
- C Edmunds
- Department of Pharmacy, Bath University, Bath, Great Britain
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Jung YD, Kim MS, Lee KS, Kang IC, Nah AS, Song DU, Yang SY, Kim JK, Ahn BW. 2-(4-morpholinyl)-8-phenyl-4H-1-benzopyran-4-one (LY294002) inhibits nitric oxide production in cultured murine astrocytes. Pharmacol Res 1999; 40:423-7. [PMID: 10527657 DOI: 10.1006/phrs.1999.0533] [Citation(s) in RCA: 11] [Impact Index Per Article: 0.4] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Indexed: 11/22/2022]
Abstract
The level of nitrite accumulation in the culture media of astrocytes activated with lipopolysaccharide (LPS) and interferon-gamma (IFN) was decreased by pretreatment of cells with LY294002, a quercetin derivative developed for phosphatidylinositol-3-kinase (PI3K) inhibitor, in a dose-dependent manner. The expression of iNOS mRNA in the astrocytes was inhibited by LY294002, as revealed by reverse transcriptional polymerase chain reaction and agarose gel electrophoresis. The catalytic activity of astrocytic iNOS was also inhibited by LY294002. On the other hand, wortmannin which was known to enhance endotoxin-induced NO production in macrophages by inhibiting PI3K did not cause any significant change in the NO production and iNOS mRNA expression of the astrocytes. These results suggest that LY294002 suppresses NO production in the astrocytes through not only the inhibition of iNOS mRNA expression but also the inhibition of the iNOS activity and that PI3K is not involved in the inhibitory actions of LY294002.pc 1999 Academic Press@p$hr
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Affiliation(s)
- Y D Jung
- Chonnam University Research Institute of Medical Sciences, Hakdong, Donggu, Kwangju, 501-190, Korea
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Orchansky PL, Kwan R, Lee F, Schrader JW. Characterization of the cytoplasmic domain of interleukin-13 receptor-alpha. J Biol Chem 1999; 274:20818-25. [PMID: 10409622 DOI: 10.1074/jbc.274.30.20818] [Citation(s) in RCA: 35] [Impact Index Per Article: 1.3] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 01/09/2023] Open
Abstract
Interleukin (IL)-13 and IL-4 are pleiotropic immunoregulatory cytokines that share many overlapping biological properties reflecting the fact that both can utilize a receptor complex composed of the IL-4 receptor-alpha (IL-4Ralpha) chain and the IL-13Ralpha chain. The cytoplasmic domain of the IL-13Ralpha is 60 amino acids long and is essential for IL-13-dependent growth. It contains a Pro-rich domain in the membrane-proximal region and two Tyr residues. Here we show that a truncated IL-13Ralpha, lacking the 38 carboxyl-terminal residues but retaining the Pro-rich region, can support IL-13-dependent proliferation, although with reduced efficiency. A Y402F mutant of the cytoplasmic domain of IL-13Ralpha supported normal IL-13-induced growth. However, tyrosine phosphorylation of signal transducer and activator of transcription 3 (STAT3), which we show is induced by IL-13 and IL-4 in cells that express the IL-13Ralpha, was significantly reduced. The cytoplasmic domain of IL-13Ralpha was constitutively associated with STAT3, Tyk2, and Janus kinase 1 (JAK1). IL-13-induced tyrosine phosphorylation of IL-13Ralpha in vivo could not be detected using anti-Tyr(P) antibodies. A glutathione S-transferase fusion protein of the cytoplasmic domain of IL-13Ralpha was phosphorylated on tyrosine in vitro by JAK1, JAK3, and Tyk2, although the tyrosine phosphorylation events mediated by Tyk2 and JAK3 were not detectable using anti-phosphotyrosine antibodies. These data, together with the demonstration that IL-13Ralpha associates constitutively with Tyk2 and that Tyr-402 is involved in IL-13-induced phosphorylation of STAT3, suggest that the latter is mediated by Tyk2. Tyrosine phosphorylation of STAT3, which was not necessary for IL-13-induced proliferation, may account for some of the effects of IL-4 and IL-13 on the function of their targets.
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Affiliation(s)
- P L Orchansky
- The Biomedical Research Centre, The University of British Columbia, Vancouver V6T 1Z3, British Columbia, Canada
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