bluuebunny/crossref_metadata_2025 huggingface dataset:
Adiponectin Enhances IL-6 Production in Human Synovial Fibroblast via an AdipoR1 Receptor, AMPK, p38, and NF-κB Pathway
Authors: Tang Chih-Hsin , Chiu Yung-Cheng , Tan Tzu-Wei , Yang Rong-Sen , Fu Wen-Mei
Published: April 2014

Articular adipose tissue is a ubiquitous component of human joints, and adiponectin is a protein hormone secreted predominantly by differentiated adipocytes and involved in energy homeostasis. We investigated the signaling pathway involved in IL-6 production caused by adiponectin in both rheumatoid arthritis synovial fibroblasts and osteoarthritis synovial fibroblasts. Rheumatoid arthritis synovial fibroblasts and osteoarthritis synovial fibroblasts expressed the AdipoR1 and AdipoR2 isoforms of the adiponectin receptor. Adiponectin caused concentration- and time-dependent increases in IL-6 production. Adiponectin-mediated IL-6 production was attenuated by AdipoR1 and 5′-AMP-activated protein kinase (AMPK)α1 small interference RNA. Pretreatment with AMPK inhibitor (araA and compound C), p38 inhibitor (SB203580), NF-κB inhibitor, IκB protease inhibitor, and NF-κB inhibitor peptide also inhibited the potentiating action of adiponectin. Adiponectin increased the kinase activity and phosphorylation of AMPK and p38. Stimulation of synovial fibroblasts with adiponectin activated IκB kinase α/β (IKK α/β), IκBα phosphorylation, IκBα degradation, p65 phosphorylation at Ser (276), p65 and p50 translocation from the cytosol to the nucleus, and κB-luciferase activity. Adiponectin-mediated an increase of IKK α/β activity, κB-luciferase activity, and p65 and p50 binding to the NF-κB element and was inhibited by compound C, SB203580 and AdipoR1 small interference RNA. Our results suggest that adiponectin increased IL-6 production in synovial fibroblasts via the AdipoR1 receptor/AMPK/p38/IKKαβ and NF-κB signaling pathway.
Mature B Cells Preferentially Lose Tolerance in the Chronic Graft-versus-Host Disease Model of Systemic Lupus Erythematosus
Authors: Choudhury Arpita , Cohen Philip L. , Eisenberg Robert A.
Published: April 2014

Chronic graft-vs-host (cGVH) disease is a well-characterized systemic lupus erythematosus (SLE) model. Induction of cGVH in anti-DNA H chain knockin (3H9KI) transgenic mice results in specific activation of anti-dsDNA B cells. In this study, we show that B cells from 3H9KI mice were activated by cGVH even when adoptively transferred into irradiated JHT−/− recipients that lack endogenous B cells. This process of activation was reflected by high autoantibody titers and changes in phenotypic markers. We have used this system to characterize the particular B cell subsets that were responsible for secreting autoantibodies during cGVH response. We isolated splenic B cell subsets based on their expression of specific cell surface markers and used them in our adoptive transfer studies. We found that mature B cells were the most vulnerable to the allostimulus and were the major source of autoantibodies compared with immature B cells. The greater susceptibility of mature B cells to become activated and thereby lose tolerance was unanticipated and has implications for maintenance of peripheral tolerance and for the development of autoimmunity. Furthermore, of the mature B cells, marginal zone B cells were particularly responsible for mounting the initial response to the cGVH stimulus. This observation underscores the critical role of marginal zone B cells in activation and production of autoantibodies.
How Regulatory CD25+CD4+ T Cells Impinge on Tumor Immunobiology: The Differential Response of Tumors to Therapies
Authors: Leon Kalet , Garcia Karina , Carneiro Jorge , Lage Agustin
Published: April 2014

Aiming to get a better insight on the impact of regulatory CD25+CD4+ T cells in tumor-immunobiology, a simple mathematical model was previously formulated and studied. This model predicts the existence of two alternative modes of uncontrolled tumor growth, which differ on their coupling with the immune system, providing a plausible explanation to the observation that the development of some tumors expand regulatory T cells whereas others do not. We report now the study of how these two tumor classes respond to different therapies, namely vaccination, immune suppression, surgery, and their different combinations. We show 1) how the timing and the dose applied in each particular treatment determine whether the tumor will be rejected, with or without concomitant autoimmunity, or whether it will continue progressing with slower or faster pace; 2) that both regulatory T cell-dependent and independent tumors are equally sensitive to vaccination, although the former are more sensitive to T cell depletion treatments and are unresponsive to partial surgery alone; 3) that surgery, suppression, and vaccination treatments, can synergistically improve their individual effects, when properly combined. Particularly, we predict rational combinations helping to overcome the limitation of these individual treatments on the late stage of tumor development.
Extent of Stimulation Controls the Formation of Memory CD8 T Cells
Authors: Quigley Michael , Huang Xiaopei , Yang Yiping
Published: April 2014

Only a small fraction of effector CD8 T cells survives to become long-lived memory cells, whereas the majority of them die after an acute infection. What controls the formation of memory CD8 T cells remains mostly unknown. In this study, we showed CD8 T cells primed earlier during vaccinia viral infection received stronger stimulation, divided more extensively, and survived better than those primed later, leading to generation of a larger memory pool. Despite differentiation into effectors, the late-primed CD8 T cells lacked full cell division, displayed increased apoptosis, and failed to develop into memory cells, suggesting that the extent of stimulation influences the survival of effector CD8 T cells. We further demonstrated that the extent of stimulation, which included both the duration and the levels of antigenic stimulation/costimulation, during priming determined the formation of memory CD8 T cells via controlling the extent of Akt activation, and functional suppression of Akt led to defective CD8 memory formation in vivo. Collectively, our data suggest that the extent of stimulation controls CD8 memory formation via activation of Akt and may provide important insights into the design of effective vaccines.
Autoimmunity Stimulated by Adoptively Transferred Dendritic Cells Is Initiated by Both αβ and γδ T Cells but Does Not Require MyD88 Signaling
Authors: Martin David A. , Zhang Kang , Kenkel Justin , Hughes Grant , Clark Edward , Davidson Anne , Elkon Keith B.
Published: April 2014

Vaccination of nonautoimmune prone mice with syngeneic dendritic cells (DC) readily induces anti-DNA autoantibodies but does not trigger systemic disease. We observed that anti-DNA autoantibody generation absolutely required αβ T cells and that γδ T cells also contributed to the response, but that regulatory T cells restrained autoantibody production. Although both NZB/W F1 mice and DC vaccinated C57/BL6 mice produced autoantibodies against dsDNA, vaccinated mice had higher levels of Abs against H1 histone and lower levels of antinucleosome Abs than NZB/W F1 mice. Despite a 100-fold increase in IL-12 and Th1 skewing to a foreign Ag, OVA, synergistic TLR activation of DC in vitro failed to augment anti-DNA Abs or promote class switching beyond that induced by LPS alone. TLR stimulation was not absolutely required for the initial loss of B cell tolerance because anti-DNA levels were similar when wild-type (WT) or MyD88-deficient DC were used for vaccination or WT and MyD88-deficient recipients were vaccinated with WT DC. In contrast, systemic administration of LPS, augmented anti-DNA Ab levels and promoted class switching, and this response was dependent on donor DC signaling via MyD88. LPS also augmented responses in the MyD88-deficient recipients, suggesting that LPS likely exerts its effects on both transferred DC and host B cells in vivo. These results indicate that both the αβ and γδ subsets are necessary for promoting autoantibody production by DC vaccination, and that although TLR/MyD88 signaling is not absolutely required for initiation, this pathway does promote augmentation, and Th1-mediated skewing, of anti-DNA autoantibodies.
A Functionally Coupled μ3-Like Opiate Receptor/Nitric Oxide Regulatory Pathway in Human Multi-Lineage Progenitor Cells
Authors: Cadet Patrick , Mantione Kirk J. , Zhu Wei , Kream Richard M. , Sheehan Melinda , Stefano George B.
Published: April 2014

Ongoing studies from our group support the existence and biological importance of a distinct cellular signaling pathway involving endogenously synthesized, chemically authentic, l-morphine, its cognate μ3 opiate receptor subtype, and constitutive NO synthase. Based on prior studies indicating evolutionary conservation and adaptation of morphinergic/NO-coupled signaling to mediate autocrine/paracrine control of cellular functions, our goal was to determine whether a functionally competent μ3 opiate receptor/NO-coupled regulatory pathway exists in human multilineage progenitor cells (MLPC) prepared from umbilical cord blood. Real-time PCR analysis indicated significant expression of μ3 opiate receptor-encoding RNA by undifferentiated human MLPC, in the absence of traditional μ1 opioid receptor-encoding RNA expression. Unpredictably, confirmatory RT-PCR analyses indicated cellular expression of a splice variant of the previously characterized μ3 opiate receptor-encoding mRNA. Pharmacological analyses provided critical validating evidence of functional μ3-like opiate receptor/NO-coupled signaling within primary cultures of undifferentiated human MLPC via morphine-evoke real-time release of NO. Control analyses indicated that morphine-stimulated NO release was markedly inhibited by prior treatment with the opiate antagonist l-naloxone or the constitutive NO synthase inhibitor N(G)-nitro-l-arginine methyl ester and unresponsive to stimulation by the opioid peptide methionine enkephalin. Complementary microarray analysis demonstrated that traditional μ1, δ, and κ opioid receptor gene expression is not detected in both undifferentiated and differentiated MLPC. Chemical differentiation of MLPC into neuronal progenitor cells effected significant phenotypic expression of a variety of neurally-associated genes. Our data provide compelling evidence in support of both the evolutionary primacy and primordial regulatory role of μ3-like opiate receptor/NO signaling in embryogenesis.
B Cell Activation State-Governed Formation of Germinal Centers following Viral Infection
Authors: Fink Katja , Manjarrez-Orduño Nataly , Schildknecht Anita , Weber Jacqueline , Senn Beatrice M. , Zinkernagel Rolf M. , Hengartner Hans
Published: April 2014

Germinal centers are structures that promote humoral memory cell formation and affinity maturation, but the triggers for their development are not entirely clear. Activated extrafollicular B cells can form IgM-producing plasmablasts or enter a germinal center reaction and differentiate into memory or plasma cells, mostly of the IgG isotype. Vesicular stomatitis virus (VSV) induces both types of response, allowing events that promote each of these pathways to be studied. In this work, extrafollicular vs germinal center responses were examined at a cellular level, analyzing VSV-specific B cells in infected mice. We show that VSV-specific germinal centers are transiently formed when insufficient proportions of specific T cell help is available and that strong B cell activation in cells expressing high levels of the VSV-specific BCR promoted their differentiation into early blasts, whereas moderate stimulation of B cells or interaction with Th cells restricted extrafollicular responses and promoted germinal center formation.
Regulatory Role of Lymphoid Chemokine CCL19 and CCL21 in the Control of Allergic Rhinitis
Authors: Takamura Kaoru , Fukuyama Satoshi , Nagatake Takahiro , Kim Dong-Young , Kawamura Aya , Kawauchi Hideyuki , Kiyono Hiroshi
Published: April 2014

The lymphoid chemokines CCL19 and CCL21 are known to be crucial both for lymphoid cell trafficking and for the structural organization of lymphoid tissues such as nasopharynx-associated lymphoid tissue (NALT). However, their role in allergic responses remains unclear, and so our current study aims to shed light on the role of CCL19/CCL21 in the development of allergic rhinitis. After nasal challenge with OVA, OVA-sensitized plt (paucity of lymph node T cells) mice, which are deficient in CCL19/CCL21, showed more severe allergic symptoms than did identically treated wild-type mice. OVA-specific IgE production, eosinophil infiltration, and Th2 responses were enhanced in the upper airway of plt mice. Moreover, in plt mice, the number of CD4+CD25+ regulatory T cells declined in the secondary lymphoid tissues, whereas the number of Th2-inducer-type CD8α−CD11b+ myeloid dendritic cells (m-DCs) increased in cervical lymph nodes and NALT. Nasal administration of the plasmid-encoding DNA of CCL19 resulted in the reduction of m-DCs in the secondary lymphoid tissues and the suppression of allergic responses in plt mice. These results suggest that CCL19/CCL21 act as regulatory chemokines for the control of airway allergic disease and so may offer a new strategy for the control of allergic disease.
CD86 Has Sustained Costimulatory Effects on CD8 T Cells
Authors: Thomas Ian J. , Petrich de Marquesini Liliana G. , Ravanan Rommel , Smith Richard M. , Guerder Sylvie , Flavell Richard A. , Wraith David C. , Wen Li , Wong F. Susan
Published: April 2014

CD80 and CD86 both costimulate T cell activation. Their individual effects in vivo are difficult to study as they are coordinately up-regulated on APCs. We have studied mice expressing rat insulin promoter (RIP)-CD80 and RIP-CD86 on the NOD and NOD.scid genetic background to generate in vivo models, using diabetes as a readout for cytotoxic T cell activation. Accelerated spontaneous diabetes onset was observed in NOD-RIP-CD80 mice and the transfer of diabetes from 6-wk-old NOD mice to NOD.scid-RIP-CD80 mice was greater compared with NOD-RIP-CD86 and NOD.scid-RIP-CD86 mice, respectively. However, the secondary in vivo response was maintained if T cells were activated through CD86 costimulation compared with CD80. This was demonstrated by greater ability to cause recurrent diabetes in NOD-RIP-CD86 diabetic mice transplanted with 6-wk-old NOD islets and adoptively transferred diabetes from diabetic NOD-RIP-CD86 mice to NOD.scid mice. In vitro, CD80 costimulation enhanced cytotoxicity, proliferation, and cytokine secretion in activated CD8 T cells compared with CD86 costimulation. We demonstrated increased CTLA-4 and programmed death-1 inhibitory molecule expression following costimulation by both CD80 and CD86 (CD80 > CD86). Furthermore, T cells stimulated by CD80 were more susceptible to inhibition by CD4+CD25+ T cells. Overall, while CD86 does not stimulate an initial response as strongly as CD80, there is greater sustained activity that is seen even in the absence of continued costimulation. These functions have implications for the engineered use of costimulatory molecules in altering immune responses in a therapeutic setting.
Anti-Inflammatory Properties and Regulatory Mechanism of a Novel Derivative of Artemisinin in Experimental Autoimmune Encephalomyelitis
Authors: Wang Zhaojun , Qiu Ju , Guo Taylor B. , Liu Ailian , Wang Ying , Li Yin , Zhang Jingwu Z.
Published: April 2014

Ethyl 2-[4-(12-β-artemisininoxy)]phenoxylpropionate (SM933) is a novel derivative of artemisinin, an herbal compound approved for the treatment of malaria. In this study, we show that SM933 has unique anti-inflammatory properties through regulation of signaling pathways, leading to amelioration of experimental autoimmune encephalomyelitis. The anti-inflammatory properties of SM933 were characterized by inhibition of encephalitogenic T cell responses that were altered to exhibit a Th2 immune deviation and reduced activity and concentration of NO and inducible NO synthase. The observed effect of SM933 was mediated through regulatory mechanisms involving the NFκB and the Rig-G/JAB1 signaling pathways. SM933 was found to inhibit the activity of NFκB by up-regulating IκB, which accounted for various down-stream anti-inflammatory actions. Furthermore, it up-regulated Rig-G through the action of IFN-α and prevented JAB1, a master cell cycle regulator, from entering the nucleus to promote p27 degradation, resulting in down-regulation of CDK2 and cyclin A and cell cycle progression. Regulation of the Rig-G/JAB1 pathway by SM933 led to altered cell cycle activity of encephalitogenic T cells as a result of its selective effect on activated, but not resting, T cells. The study indicates that SM933 is a novel anti-inflammatory agent acting through defined signaling mechanisms and provides regulatory mechanisms required for effective drug targeting in treatment of autoimmune disease and inflammation.
Jagged1 and Delta1 Differentially Regulate the Outcome of Experimental Autoimmune Encephalomyelitis
Authors: Elyaman Wassim , Bradshaw Elizabeth M. , Wang Yue , Oukka Mohamed , Kivisäkk Pia , Chiba Shigeru , Yagita Hideo , Khoury Samia J.
Published: April 2014

Notch signaling plays an important role during T cell development in the thymus and in T cell activation but the role of Notch in autoimmunity is not clear. We investigated the role of Jagged1 and Delta1 in experimental autoimmune encephalomyelitis. During experimental autoimmune encephalomyelitis, Delta1 expression is up-regulated on dendritic cells and B cells after priming while Jagged1 is up-regulated only on dendritic cells. Administration of anti-Jagged1 Ab exacerbated clinical disease while that of anti-Delta1 Ab reduced the severity of the clinical disease. In contrast, administration of Jagged1-Fc protected from disease, that of Delta1-Fc exacerbated disease. Treatment with Jagged1-Fc was associated with increased IL-10-producing Ag-specific cells in the CNS, while anti-Jagged1 decreased the frequency of IL-10-producing cells. Treatment with Delta1-Fc increased Th1 cells in the CNS, while anti-Delta-1 decreased the frequency of Th1 cells. Manipulation of Delta1 or Jagged1 had no effect on the frequency of Th17 cells or FoxP3+ cells. Moreover, Jagged1 may play a role in CNS homeostasis because murine astrocytes specifically express Jagged1 that is up-regulated by TGF-β, whereas IFN-γ, TNF-α, and IL-17 decrease Jagged1 expression. Our study provides novel data about differential roles of Notch ligands in regulating inflammation in the periphery as well as in the CNS.
Functional yet Balanced Reactivity to Candida albicans Requires TRIF, MyD88, and IDO-Dependent Inhibition of Rorc
Authors: De Luca Antonella , Montagnoli Claudia , Zelante Teresa , Bonifazi Pierluigi , Bozza Silvia , Moretti Silvia , D’Angelo Carmen , Vacca Carmine , Boon Louis , Bistoni Francesco , Puccetti Paolo , Fallarino Francesca , Romani Luigina
Published: April 2014

The ability of regulatory T (Treg) cells to inhibit aspects of innate and adaptive immunity is central to their protective function in fungal infections. In murine candidiasis, CD4+CD25+ Treg cells prevent excessive inflammation but enable fungal persistence in the gastrointestinal tract, which underlies the onset of durable antifungal protection. In this study, we show that fungal growth, inflammatory immunity, and tolerance to the fungus were all controlled by the coordinate activation of naturally occurring Treg cells, which limited early inflammation at the sites of infection, and pathogen-induced Treg cells (that regulated the expression of adaptive Th immunity in secondary lymphoid organs). Naturally occurring Treg cells required the TRIF pathway for migration to inflamed sites, where the MyD88 pathway would then restrain their suppressive function. Subsequent inflammatory Th1-type immunity was modulated by induced Treg cells, which required the TRIF pathway as well, and acted through activation of IDO in dendritic cells and Th17 cell antagonism. In vitro, using naive CD4+ cells from TRIF-deficient mice, tryptophan metabolites were capable of inducing the Foxp3-encoding gene transcriptionally and suppressing the gene encoding RORγt, Th17 lineage specification factor. This is the first study to show that the same tryptophan catabolites can foster dendritic cell-supported generation of Foxp3+ cells and mediate, at the same time, inhibition of RORγt-expressing T cells.
Purinergic P2Y2 Receptors Induce Increased MCP-1/CCL2 Synthesis and Release from Rat Alveolar and Peritoneal Macrophages
Authors: Stokes Leanne , Surprenant Annmarie
Published: April 2014

Macrophages play a key role in inflammation by synthesis and release of proinflammatory cytokines and chemokines. Extracellular nucleotides released at sites of tissue damage may be an early danger signal for immune cells, and ATP-gated P2X7 receptors are well known to mediate the rapid release of proinflammatory IL-18 and IL-1β. However, there is little direct evidence for the involvement of other purine receptor subtypes in the release of other cytokines or chemokines. We initially used protein arrays to address whether extracellular ATP can release cytokines and/or chemokines from rat NR8383 alveolar macrophage, which lack the P2X7 receptor. ATPγS increased the release of the proinflammatory chemokine, MCP-1 (MCP-1/CCL2). Pharmacological profiling identified the receptor responsible as the P2Y2 receptor. Brief activation (10 min) of P2Y2 receptors increased MCP-1 mRNA levels within 30 min and increased its release at 60 min. Similar results were obtained from rat peritoneal macrophages. We investigated likely downstream signaling cascades that may be involved, specifically the canonical Gq-mediated phospholipase C (PLC) and subsequent MAP kinase pathways, and Gi/Go-mediated signaling. We could find no evidence for these pathways being involved in the P2Y2R-induced increase in mRNA levels although inhibition of PLC blocked the UTP-induced increased release of MCP-1. Thus, the PLC-activated pathway can account for the increased release of MCP-1, but a novel signaling pathway may be involved in the increase in MCP-1 mRNA by activation of P2Y2 receptors in alveolar and peritoneal macrophage.
TLR4/MD-2 Monoclonal Antibody Therapy Affords Protection in Experimental Models of Septic Shock
Authors: Daubeuf Bruno , Mathison John , Spiller Stephan , Hugues Stephanie , Herren Suzanne , Ferlin Walter , Kosco-Vilbois Marie , Wagner Hermann , Kirschning Carsten J. , Ulevitch Richard , Elson Greg
Published: April 2014

Overactivation of the immune system upon acute bacterial infection leads to septic shock. Specific bacterial products potently stimulate immune cells via toll-like receptors (TLRs). Gram-negative bacteria induce a predominantly TLR4-driven signal through LPS release. To neutralize LPS signaling in experimental models of sepsis, we generated mAbs toward the TLR4/myeloid differentiation protein-2 (MD-2) complex. The binding properties of an array of selected rat mAbs differed in respect to their specificity for TLR4/MD-2 complex. The specificity of one such mAb, 5E3, to murine TLR4 was confirmed by its recognition of an epitope within the second quarter of the ectodomain. 5E3 inhibited LPS-dependent cell activation in vitro and prevented proinflammatory cytokine production in vivo following LPS challenge in a dose-dependent manner. Furthermore, 5E3 protected mice from lethal shock-like syndrome when applied using both preventative and therapeutic protocols. Most notably, in the colon ascendens stent peritonitis model of polymicrobial abdominal sepsis, administration of a single dose of 5E3 (50 μg) protected mice against mortality. These results demonstrate that neutralizing TLR4/MD-2 is highly efficacious in protecting against bacterial infection-induced toxemia and offers TLR4/MD-2 mAb treatment as a potential therapy for numerous clinical indications.
Characterization of Antigen-Specific Immune Responses Induced by Canarypox Virus Vaccines
Authors: Bos Rinke , van Duikeren Suzanne , van Hall Thorbald , Lauwen Marjolein M. , Parrington Mark , Berinstein Neil L. , McNeil Bryan , Melief Cornelis J. M. , Verbeek J. Sjef , van der Burg Sjoerd H. , Offringa Rienk
Published: April 2014

Avipoxvirus-based vectors, such as recombinant canarypox virus ALVAC, are studied extensively as delivery vehicles for vaccines against cancer and infectious diseases. Effective use of such vaccines is expected to benefit from proper understanding of the interaction between these viral vectors and the host immune system. We performed preclinical vaccination experiments in a murine tumor model to analyze the immunogenic properties of an ALVAC-based vaccine against carcinoembryonic Ag (ALVAC-CEA), a tumor-associated autoantigen commonly overexpressed in colorectal cancers. The protective CEA-specific immunity induced by this vaccine consisted of CD4+ T cell responses with a mixed Th1/Th2 cytokine profile that were accompanied by potent humoral responses, but not by CEA-specific CD8+ CTL immunity. In contrast, protective immunity induced by a CEA-specific DNA vaccine (DNA-CEA) consisted of Th1 and CTL responses. Modification of the ALVAC-CEA vaccine through coinjection of DNA-CEA, admixture with CpG oligodeoxynucleotides, or supplementation with additional transgenes encoding a triad of costimulatory molecules (TRICOM) did not result in induction of CEA-specific CTL responses. Even though these results suggested that ALVAC does not elicit Ag-specific CTLs, immunization with ALVAC vaccines against other Ags efficiently induced CTL responses. Our data show that the capacity of ALVAC vaccines to elicit CTL immunity against transgene-encoded Ags critically depends on the presence of highly immunogenic CTL epitopes in these Ags. This consideration needs to be taken into account with respect to the design and evaluation of vaccination strategies that use ALVAC-based vaccine.
Macrophages from 11β-Hydroxysteroid Dehydrogenase Type 1-Deficient Mice Exhibit an Increased Sensitivity to Lipopolysaccharide Stimulation Due to TGF-β-Mediated Up-Regulation of SHIP1 Expression
Authors: Zhang Tian Y. , Daynes Raymond A.
Published: April 2014

11β-Hydroxysteroid dehydrogenase type 1 (11βHSD1) performs end-organ metabolism of glucocorticoids (GCs) by catalyzing the conversion of C11-keto-GCs to C11-hydroxy-GCs, thereby generating activating ligands for the GC receptor. In this study, we report that 11βHSD1−/− mice are more susceptible to endotoxemia, evidenced by increased weight loss and serum TNF-α, IL-6, and IL-12p40 levels following LPS challenge in vivo. Peritoneal and splenic macrophage (splnMφ) from these genetically altered mice overproduce inflammatory cytokines following LPS stimulation in vitro. Inflammatory cytokine overexpression by 11βHSD1−/− splnMφ results from an increased activation of NF-κB- and MAPK-signaling cascades and an attenuated PI3K-dependent Akt activation. The expression of SHIP1 is augmented in 11βHSD1−/− Mφ and contributes to inflammatory cytokine production because overexpression of SHIP1 in primary bone marrow Mφ (BMMφ) leads to a similar type of hyperresponsiveness to subsequent LPS stimulation. 11βHSD1+/+ and 11βHSD1−/− BMMφ responded to LPS similarly. However, 11βHSD1−/− BMMφ derived in the presence of elevated GC levels up-regulated SHIP1 expression and increased their capacity to produce inflammatory cytokines following their activation with LPS. These observations suggest the hyperresponsiveness of 11βHSD1−/− splnMφ results from myeloid cell differentiation in the presence of moderately elevated GC levels found within 11βHSD1−/− mice. GC-conditioning of BMMφ enhanced SHIP1 expression via up-regulation of bioactive TGF-β. Consistently, TGF-β protein expression was increased in unstimulated CD11b− cells residing in the BM and spleen of 11βHSD1−/− mice. Our results suggest that modest elevations in plasma GC levels can modify the LPS responsiveness of Mφ by augmenting SHIP1 expression through a TGF-β-dependent mechanism.
Cutting Edge: Nonglycosidic CD1d Lipid Ligands Activate Human and Murine Invariant NKT Cells
Authors: Silk Jonathan D. , Salio Mariolina , Reddy B. Gopal , Shepherd Dawn , Gileadi Uzi , Brown James , Masri S. Hajar , Polzella Paolo , Ritter Gerd , Besra Gurdyal S. , Jones E. Yvonne , Schmidt Richard R. , Cerundolo Vincenzo
Published: April 2014

Invariant NKT cells (iNKT cells) recognize CD1d/glycolipid complexes. We demonstrate that the nonglycosidic compound threitolceramide efficiently activates iNKT cells, resulting in dendritic cell (DC) maturation and the priming of Ag-specific T and B cells. Threitolceramide-pulsed DCs are more resistant to iNKT cell-dependent lysis than α-galactosylceramide-pulsed DCs due to the weaker affinity of the human iNKT TCR for CD1d/ threitolceramide than CD1d/α-galactosylceramide complexes. iNKT cells stimulated with threitolceramide also recover more quickly from activation-induced anergy. Kinetic and functional experiments showed that shortening or lengthening the threitol moiety by one hydroxymethylene group modulates ligand recognition, as human and murine iNKT cells recognize glycerolceramide and arabinitolceramide differentially. Our data broaden the range of potential iNKT cell agonists. The ability of these compounds to assist the priming of Ag-specific immune responses while minimizing iNKT cell-dependent DC lysis makes them attractive adjuvants for vaccination strategies.
Cutting Edge: Central Nervous System Plasmacytoid Dendritic Cells Regulate the Severity of Relapsing Experimental Autoimmune Encephalomyelitis
Authors: Bailey-Bucktrout Samantha L. , Caulkins Sarah C. , Goings Gwendolyn , Fischer Jens A. A. , Dzionek Andrzej , Miller Stephen D.
Published: April 2014

Plasmacytoid dendritic cells (pDCs) have both stimulatory and regulatory effects on T cells. pDCs are a major CNS-infiltrating dendritic cell population during experimental autoimmune encephalomyelitis but, unlike myeloid dendritic cells, have a minor role in T cell activation and epitope spreading. We show that depletion of pDCs during either the acute or relapse phases of experimental autoimmune encephalomyelitis resulted in exacerbation of disease severity. pDC depletion significantly enhanced CNS but not peripheral CD4+ T cell activation, as well as IL-17 and IFN-γ production. Moreover, CNS pDCs suppressed CNS myeloid dendritic cell-driven production of IL-17, IFN-γ, and IL-10 in an IDO-independent manner. The data demonstrate that pDCs play a critical regulatory role in negatively regulating pathogenic CNS CD4+ T cell responses, highlighting a new role for pDCs in inflammatory autoimmune disease.
Cutting Edge: Murine Cytomegalovirus Induces a Polyfunctional CD4 T Cell Response
Authors: Arens Ramon , Wang Peng , Sidney John , Loewendorf Andrea , Sette Alessandro , Schoenberger Stephen P. , Peters Bjoern , Benedict Chris A.
Published: April 2014

CD4 T lymphocytes regulate the adaptive immune response to most viruses, both by providing help to CD8 T cells and B cells as well as through direct antiviral activity. Currently, no mouse cytomegalovirus (MCMV)-specific CD4 T cell responses are known. In this study, we identify and characterize 15 I-Ab-restricted CD4 T cell responses specific for MCMV epitopes. CD4 T cells accumulate to high levels in the spleen and lungs during acute infection and produce multiple cytokines (IFN-γ, TNF, IL-2, IL-10, and IL-17). Interestingly, IL-17 and IFN-γ production within epitope-specific cells was found to be mutually exclusive. CD4 T cells recognizing a peptide derived from m09 were only detectable at later times of infection and displayed a unique cytokine production profile. In total, this study reveals that the MCMV-specific CD4 T cell response is complex and functionally diverse, highlighting its important role in controlling this persistent pathogen.
Imprinting of CCR9 on CD4 T Cells Requires IL-4 Signaling on Mesenteric Lymph Node Dendritic Cells
Authors: Elgueta Raul , Sepulveda Fernando E. , Vilches Felipe , Vargas Leonardo , Mora J. Rodrigo , Bono Maria Rosa , Rosemblatt Mario
Published: April 2014

It has recently been shown that IL-4 can educate dendritic cells (DC) to differentially affect T cell effector activity. In this study, we show that IL-4 can also act upon DC to instruct naive T cells to express the gut-associated homing receptor CCR9. Thus, effector T cells generated after coculture with mesenteric lymph node (MLN)-DC show a higher expression of CCR9 when activated in the presence of IL-4. In contrast, IL-4 had no effect on CCR9 expression when naive T cells were polyclonally activated in the absence of MLN-DC, suggesting that the effect of IL-4 on CCR9 expression passed through DC. Indeed, T cells activated by MLN-DC from IL-4Rα−/− mice showed a much lower CCR9 expression and a greatly reduced migration to the small intestine than T cells activated by wild-type MLN-DC even in the presence of IL-4. Consistent with the finding that the vitamin A metabolite retinoic acid (RA) induces gut-homing molecules on T cells, we further demonstrate that IL-4 up-regulated retinaldehyde dehydrogenase 2 mRNA on MLN-DC, a critical enzyme involved in the synthesis of RA. Moreover, LE135, a RA receptor antagonist, blocked the increased expression of CCR9 driven by IL-4-treated MLN-DC. Thus, besides the direct effect of RA on T cell gut tropism, our results show that the induction of a gut-homing phenotype on CD4+ T cells is also influenced by the effect of IL-4 on gut-associated DC.
Actin-Binding Protein 1 Regulates B Cell Receptor-Mediated Antigen Processing and Presentation in Response to B Cell Receptor Activation
Authors: Onabajo Olusegun O. , Seeley Margaret K. , Kale Amruta , Qualmann Britta , Kessels Michael , Han Jin , Tan Tse-Hua , Song Wenxia
Published: April 2014

The BCR serves as both signal transducer and Ag transporter. Binding of Ags to the BCR induces signaling cascades and Ag processing and presentation, two essential cellular events for B cell activation. BCR-initiated signaling increases BCR-mediated Ag-processing efficiency by increasing the rate and specificity of Ag transport. Previous studies showed a critical role for the actin cytoskeleton in these two processes. In this study, we found that actin-binding protein 1 (Abp1/HIP-55/SH3P7) functioned as an actin-binding adaptor protein, coupling BCR signaling and Ag-processing pathways with the actin cytoskeleton. Gene knockout of Abp1 and overexpression of the Src homology 3 domain of Abp1 inhibited BCR-mediated Ag internalization, consequently reducing the rate of Ag transport to processing compartments and the efficiency of BCR-mediated Ag processing and presentation. BCR activation induced tyrosine phosphorylation of Abp1 and translocation of both Abp1 and dynamin 2 from the cytoplasm to plasma membrane, where they colocalized with the BCR and cortical F-actin. Mutations of the two tyrosine phosphorylation sites of Abp1 and depolymerization of the actin cytoskeleton interfered with BCR-induced Abp1 recruitment to the plasma membrane. The inhibitory effect of a dynamin proline-rich domain deletion mutant on the recruitment of Abp1 to the plasma membrane, coimmunoprecipitation of dynamin with Abp1, and coprecipitation of Abp1 with GST fusion of the dyanmin proline-rich domain demonstrate the interaction of Abp1 with dynamin 2. These results demonstrate that the BCR regulates the function of Abp1 by inducing Abp1 phosphorylation and actin cytoskeleton rearrangement, and that Abp1 facilitates BCR-mediated Ag processing by simultaneously interacting with dynamin and the actin cytoskeleton.
Dynamics of Proximal Signaling Events after TCR/CD8-Mediated Induction of Proliferation or Apoptosis in Mature CD8+ T Cells
Authors: Wang Xiaoqian , Simeoni Luca , Lindquist Jonathan A. , Saez-Rodriguez Julio , Ambach Andreas , Gilles Ernst D. , Kliche Stefanie , Schraven Burkhart
Published: April 2014

Engagement of the TCR can induce different functional outcomes such as activation, proliferation, survival, or apoptosis. How the TCR-mediated signaling cascades generating these distinct cellular responses are organized on the molecular level is so far not completely understood. To obtain insight into this question, we analyzed TCR/CD8-mediated signaling events in mature OT-I TCR transgenic T cells under conditions of stimulation that lead to either proliferation or apoptosis. These experiments revealed major differences in the phosphorylation dynamics of LAT, ZAP70, protein kinase B, phospholipase C-γ1, protein kinase D1, and ERK1/2. Moreover, input signals leading to apoptosis induced a strong, but transient activation of ERK1/2 mainly at sites of TCR-engagement. In contrast, stimuli promoting survival/proliferation generated a low and sustained activation of ERK1/2, which colocalizes with Ras in recycling endosomal vesicles. The transient activation of ERK1/2 under pro-apoptotic conditions of stimulation is at least partially due to the rapid polyubiquitination and subsequent degradation of ZAP70, whereas the sustained activation of ERK1/2 under survival promoting conditions is paralleled by the induction/phosphorylation of anti-apoptotic molecules such as protein kinase B and Bcl-xL. Collectively, our data provide signaling signatures that are associated with proliferation or apoptosis of T cells.
Decreased Pathology and Prolonged Survival of Human DC-SIGN Transgenic Mice during Mycobacterial Infection
Authors: Schaefer Martin , Reiling Norbert , Fessler Cornelia , Stephani Johannes , Taniuchi Ichiro , Hatam Farahnaz , Yildirim Ali Oender , Fehrenbach Heinz , Walter Kerstin , Ruland Juergen , Wagner Hermann , Ehlers Stefan , Sparwasser Tim
Published: April 2014

Dendritic cell (DC)-specific intercellular adhesion molecule-3 grabbing nonintegrin (DC-SIGN: CD209) is a C-type lectin that binds ICAM-2,3 and various pathogens such as HIV, helicobacter, and mycobacteria. It has been suggested that Mycobacterium tuberculosis, the causative agent of pulmonary tuberculosis, interacts with DC-SIGN to evade the immune system. To directly analyze the role of human DC-SIGN during mycobacterial infection, we generated conventional transgenic (tg) mice (termed “hSIGN”) using CD209 cDNA under the control of the murine CD11c promoter. Upon mycobacterial infection, DCs from hSIGN mice produced significantly less IL-12p40 and no significant differences were be observed in the secretion levels of IL-10 relative to control DCs. After high dose aerosol infection with the strain M. tuberculosis H37Rv, hSIGN mice showed massive accumulation of DC-SIGN+ cells in infected lungs, reduced tissue damage and prolonged survival. Based on our in vivo data, we propose that instead of favoring the immune evasion of mycobacteria, human DC-SIGN may have evolved as a pathogen receptor promoting protection by limiting tuberculosis-induced pathology.
Coinfection with Nonlethal Murine Malaria Parasites Suppresses Pathogenesis Caused by Plasmodium berghei NK65
Authors: Niikura Mamoru , Kamiya Shigeru , Kita Kiyoshi , Kobayashi Fumie
Published: April 2014

Mixed infection with different Plasmodium species is often observed in endemic areas, and the infection with benign malaria parasites such as Plasmodium vivax or P. malariae has been considered to reduce the risk of developing severe pathogenesis caused by P. falciparum. However, it is still unknown how disease severity is reduced in hosts during coinfection. In the present study, we investigated the influence of coinfection with nonlethal parasites, P. berghei XAT (Pb XAT) or P. yoelii 17X (Py 17X), on the outcome of P. berghei NK65 (Pb NK65) lethal infection, which caused high levels of parasitemia and severe pathogenesis in mice. We found that the simultaneous infection with nonlethal Pb XAT or Py 17X suppressed high levels of parasitemia, liver injury, and body weight loss caused by Pb NK65 infection, induced high levels of reticulocytemia, and subsequently prolonged survival of mice. In coinfected mice, the immune response, including the expansion of B220intCD11c+ cells and CD4+ T cells and expression of IL-10 mRNA, was comparable to that in nonlethal infection. Moreover, the suppression of liver injury and body weight loss by coinfection was reduced in IL-10−/− mice, suggesting that IL-10 plays a role for a reduction of severity by coinfection with nonlethal malaria parasites.
Macrophage Proinflammatory Response to Francisella tularensis Live Vaccine Strain Requires Coordination of Multiple Signaling Pathways
Authors: Cole Leah E. , Santiago Araceli , Barry Eileen , Kang Tae Jin , Shirey Kari Ann , Roberts Zachary J. , Elkins Karen L. , Cross Alan S. , Vogel Stefanie N.
Published: April 2014

The macrophage proinflammatory response to Francisella tularensis (Ft) live vaccine strain (LVS) was shown previously to be TLR2 dependent. The observation that intracellular Ft LVS colocalizes with TLR2 and MyD88 inside macrophages suggested that Ft LVS might signal from within the phagosome. Macrophages infected with LVSΔiglC, a Ft LVS mutant that fails to escape from the phagosome, displayed greatly increased expression of a subset of TLR2-dependent, proinflammatory genes (e.g., Tnf) but decreased expression of others (e.g., Ifnb1). This latter subset was similarly mitigated in IFN-β−/− macrophages indicating that while Ft LVS-induced TLR2 signaling is necessary, cytosolic sensing of Ft to induce IFN-β is required for full induction of the macrophage proinflammatory response. Although LVSΔiglC greatly increased IL-1β mRNA in wild-type macrophages, protein secretion was not observed. IL-1β secretion was also diminished in Ft LVS-infected IFN-β−/− macrophages. rIFN-β failed to restore IL-1β secretion in LVSΔiglC-infected macrophages, suggesting that signals in addition to IFN-β are required for assembly of the inflammasome and activation of caspase-1. IFN-β plays a central role in controlling the macrophage bacterial burden: bacterial recovery was greater in IFN-β−/− than in wild-type macrophages and treatment of Ft LVS-infected macrophages with rIFN-β or 5,6-dimethylxanthenone-4-acetic acid, a potent IFN-β inducer, greatly decreased the intracellular Ft LVS burden. In toto, these observations support the hypothesis that the host inflammatory response to Ft LVS is complex and requires engagement of multiple signaling pathways downstream of TLR2 including production of IFN-β via an unknown cytosolic sensor and activation of the inflammasome.
Dual Role of NOX2 in Respiratory Syncytial Virus- and Sendai Virus-Induced Activation of NF-κB in Airway Epithelial Cells
Authors: Fink Karin , Duval Annick , Martel Alexis , Soucy-Faulkner Anton , Grandvaux Nathalie
Published: April 2014

Human respiratory syncytial virus (RSV), a member of the Paramyxoviridae family, is the most important viral agent of pediatric respiratory tract disease worldwide. Human airway epithelial cells (AEC) are the primary targets of RSV. AEC are responsible for the secretion of a wide spectrum of cytokines and chemokines that are important mediators of the exacerbated airway inflammation triggered by the host in response to RSV infection. NF-κB is a key transcription factor responsible for the regulation of cytokine and chemokine gene expression and thus represents a potential therapeutic target. In the present study, we sought to delineate the role of RSV-induced reactive oxygen species in the regulation of the signaling pathways leading to NF-κB activation. First, we demonstrate that besides the well-characterized IκBα-dependent pathway, phosphorylation of p65 at Ser536 is an essential event regulating NF-κB activation in response to RSV in A549. Using antioxidant and RNA-interference strategies, we show that a NADPH oxidase 2 (NOX2)-containing NADPH oxidase is an essential regulator of RSV-induced NF-κB activation. Molecular analyses revealed that NOX2 acts upstream of both the phosphorylation of IκBα at Ser32 and of p65 at Ser536 in A549 and normal human bronchial epithelial cells. Similar results were obtained in the context of infection by Sendai virus, thus demonstrating that the newly identified NOX2-dependent NF-κB activation pathway is not restricted to RSV among the Paramyxoviridae. These results illustrate a previously unrecognized dual role of NOX2 in the regulation of NF-κB in response to RSV and Sendai virus in human AEC.
Toll-Like Receptor-4 Modulates Survival by Induction of Left Ventricular Remodeling after Myocardial Infarction in Mice
Authors: Riad Alexander , Jäger Sebastian , Sobirey Meike , Escher Felicitas , Yaulema-Riss Aymaru , Westermann Dirk , Karatas Aysun , Heimesaat Markus M. , Bereswill Stefan , Dragun Duska , Pauschinger Matthias , Schultheiss Heinz P. , Tschöpe Carsten
Published: April 2014

Left ventricular (LV) remodeling is known to contribute to morbidity and mortality after myocardial infarction (MI). Because LV remodeling is strongly associated with an inflammatory response, we investigated whether or not TLR-4 influences LV remodeling and survival in a mice model of MI. Six days after MI induction, TLR4 knockout (KO)-MI mice showed improved LV function 32 and reduced LV remodeling as indexed by reduced levels of atrial natriuretic factor and total collagen as well as by a reduced heart weight to body weight ratio when compared with WT-MI mice. This was associated with a reduction of protein levels of the intracellular TLR4 adapter protein MyD88 and enhanced protein expression of the anti-hypertrophic JNK in KO-MI mice when compared with wild-type (WT)-MI mice. In contrast, protein activation of the pro-hypertrophic kinases protein kinase Cδ and p42/44 were not regulated in KO-MI mice when compared with WT-MI mice. Improved LV function, reduced cardiac remodeling, and suppressed intracellular TLR4 signaling in KO-MI mice were associated with significantly improved survival compared with WT-MI mice (62 vs 23%; p < 0.0001). TLR4 deficiency led to improved survival after MI mediated by attenuated left ventricular remodeling.
Properdin Deficiency in Murine Models of Nonseptic Shock
Authors: Ivanovska Nina D. , Dimitrova Petya A. , Luckett Jeni C. , El-Rachkidy Lonnen Rana , Schwaeble Wilhelm J. , Stover Cordula M.
Published: April 2014

Hereditary properdin deficiency is linked to susceptibility to meningococcal disease (Neisseria meningitidis serotypes Y and W-135) with high mortality. Its relative contribution toward the outcome of nonseptic shock has not been investigated. Using properdin-deficient C57BL/6 mice and their littermates, this study examines their survival of zymosan-induced and LPS-induced shock. Properdin-deficient mice were more resistant to zymosan shock compared with wild-type mice, which showed greater impairment of end-organ function 24 h after zymosan injection, higher TNF-α production by alveolar and peritoneal macrophages, higher TNF-α, and, inversely, lower IL-10 levels in peritoneal lavage and circulation and higher plasma C5a levels. Properdin-deficient mice showed significantly higher mortality in LPS shock, elevated TNF-α, and, inversely, reduced IL-10 production by peritoneal macrophages as well as lower plasma C5a levels compared with wild-type littermates. NO production by peritoneal macrophages and plasma α1-antitrypsin levels at 24 h after the injection of LPS or zymosan were decreased in properdin-deficient mice in both models, and fewer histopathologic changes in liver were observed in properdin-deficient animals. This study provides evidence that properdin deficiency attenuates zymosan-induced shock and exacerbates LPS-induced shock.