bluuebunny/crossref_metadata_2025 huggingface dataset:
HIV-1 Tat Mediates Degradation of RON Receptor Tyrosine Kinase, a Regulator of Inflammation
Authors: Kalantari Parisa , Harandi Omid F. , Hankey Pamela A. , Henderson Andrew J.
Published: April 2014

HIV encodes several proteins, including Tat, that have been demonstrated to modulate the expression of receptors critical for innate immunity, including MHC class I, mannose receptor, and β2-microglobulin. We demonstrate that Tat targets the receptor tyrosine kinase recepteur d’origine nantais (RON), which negatively regulates inflammation and HIV transcription, for proteosome degradation. Tat decreases cell surface RON expression in HIV-infected monocytic cells, and Tat-mediated degradation of RON protein is blocked by inhibitors of proteosome activity. Tat specifically induced down-regulation of RON and not other cell surface receptors, such as the transferrin receptor, the receptor tyrosine kinase TrkA, or monocytic markers CD14 and ICAM-1. The Tat trans activation domain is required for RON degradation, and this down-regulation is dependent on the integrity of the kinase domain of RON receptor. We propose that Tat mediates degradation of RON through a ubiquitin-proteosome pathway, and suggest that by targeting signals that modulate inflammation, Tat creates a microenvironment that is optimal for HIV replication and progression of AIDS-associated diseases.
Evidence for Genes in Addition to Tlr7 in the Yaa Translocation Linked with Acceleration of Systemic Lupus Erythematosus
Authors: Santiago-Raber Marie-Laure , Kikuchi Shuichi , Borel Paula , Uematsu Satoshi , Akira Shizuo , Kotzin Brian L. , Izui Shozo
Published: April 2014

The accelerated development of systemic lupus erythematosus (SLE) in male BXSB mice is associated with the genetic abnormality in its Y chromosome, designated Yaa (Y-linked autoimmune acceleration). Recently, the Yaa mutation was identified to be a translocation from the telomeric end of the X chromosome (containing the gene encoding TLR7) onto the Y chromosome. In the present study, we determined whether the Tlr7 gene duplication is indeed responsible for the Yaa-mediated acceleration of SLE. Analysis of C57BL/6 mice congenic for the Nba2 (NZB autoimmunity 2) locus (B6.Nba2) bearing the Yaa mutation revealed that introduction of the Tlr7 null mutation on the X chromosome significantly reduced serum levels of IgG autoantibodies against DNA and ribonucleoproteins, as well as the incidence of lupus nephritis. However, the protection was not complete, because these mice still developed high titers of anti-chromatin autoantibodies and retroviral gp70-anti-gp70 immune complexes, and severe lupus nephritis, which was not the case in male B6.Nba2 mice lacking the Yaa mutation. Moreover, we found that the Tlr7 gene duplication contributed to the development of monocytosis, but not to the reduction of marginal zone B cells, which both are cellular abnormalities causally linked to the Yaa mutation. Our results indicate that the Yaa-mediated acceleration of SLE as well as various Yaa-linked cellular traits cannot be explained by the Tlr7 gene duplication alone, and suggest additional contributions from other duplicated genes in the translocated X chromosome.
Genetic and Molecular Basis of Quantitative Trait Loci of Arthritis in Rat: Genes and Polymorphisms
Authors: Xiong Qing , Jiao Yan , Hasty Karen A. , Stuart John M. , Postlethwaite Arnold , Kang Andrew H. , Gu Weikuan
Published: April 2014

Rheumatoid arthritis (RA) is an autoimmune disease, the pathogenesis of which is affected by multiple genetic and environmental factors. To understand the genetic and molecular basis of RA, a large number of quantitative trait loci (QTL) that regulate experimental autoimmune arthritis have been identified using various rat models for RA. However, identifying the particular responsible genes within these QTL remains a major challenge. Using currently available genome data and gene annotation information, we systematically examined RA-associated genes and polymorphisms within and outside QTL over the whole rat genome. By the whole genome analysis of genes and polymorphisms, we found that there are significantly more RA-associated genes in QTL regions as contrasted with non-QTL regions. Further experimental studies are necessary to determine whether these known RA-associated genes or polymorphisms are genetic components causing the QTL effect.
Cutting Edge: Chromatin Remodeling as a Molecular Basis for the Enhanced Functionality of Memory CD8 T Cells
Authors: Northrop John K. , Wells Andrew D. , Shen Hao
Published: April 2014

Memory CD8 T cells, unlike their naive precursors, are capable of rapidly producing high levels of cytokines, killing target cells, and proliferating into numerous secondary effectors immediately upon Ag encounter. This ready-to-respond state contributes to their superior ability to confer protective immunity, yet the underlying molecular basis remains unknown. In this study, we show that memory CD8 T cells have increased histone acetylation compared with naive CD8 T cells; however, those activated without CD4 T cell help (“unhelped”) remain hypoacetylated and fail to develop into functional, protective memory. Treatment with a histone deacetylase inhibitor during activation results in increased histone acetylation in unhelped CD8 T cells and restores their ability to differentiate into functional memory cells capable of immediate cytokine production and providing protective immunity. These results demonstrate that CD4 T help-dependent chromatin remodeling provides a molecular basis for the enhanced responsiveness of memory CD8 T cells.
Impaired SLAM-SLAM Homotypic Interaction between Invariant NKT Cells and Dendritic Cells Affects Differentiation of IL-4/IL-10-Secreting NKT2 Cells in Nonobese Diabetic Mice
Authors: Baev Denis V. , Caielli Simone , Ronchi Francesca , Coccia Margherita , Facciotti Federica , Nichols Kim E. , Falcone Marika
Published: April 2014

The regulatory function of invariant NKT (iNKT) cells for tolerance induction and prevention of autoimmunity is linked to a specific cytokine profile that comprises the secretion of type 2 cytokines like IL-4 and IL-10 (NKT2 cytokine profile). The mechanism responsible for iNKT cell differentiation toward a type 2 phenotype is unknown. Herein we show that costimulatory signals provided by the surface receptor signaling lymphocytic activation molecule (SLAM) on myeloid dendritic cells (mDC) to iNKT cells is crucial for NKT2 orientation. Additionally, we demonstrate that the impaired acquisition of an NKT2 cytokine phenotype in nonobese diabetic (NOD) mice that spontaneously develop autoimmune diabetes is due to defective SLAM-induced signals generated by NOD mDC. Mature mDC of C57BL/6 mice express SLAM and induce C57BL/6 or NOD iNKT cells to acquire a predominant NKT2 cytokine phenotype in response to antigenic stimulation with the iNKT cell-specific Ag, the α-galactosylceramide. In contrast, mature NOD mDC express significantly lower levels of SLAM and are unable to promote GATA-3 (the SLAM-induced intracellular signal) up-regulation and IL-4/IL-10 production in iNKT cells from NOD or C57BL/6 mice. NOD mice carry a genetic defect of the Slamf1 gene that is associated with reduced SLAM expression on double-positive thymocytes and altered iNKT cell development in the thymus. Our data suggest that the genetic Slamf1 defect in NOD mice also affects SLAM expression on other immune cells such as the mDC, thus critically impairing the peripheral differentiation of iNKT cells toward a regulatory NKT2 type.
The Complement Inhibitor Low Molecular Weight Dextran Sulfate Prevents TLR4-Induced Phenotypic and Functional Maturation of Human Dendritic Cells
Authors: Spirig Rolf , van Kooten Cees , Obregon Carolina , Nicod Laurent , Daha Mohamed , Rieben Robert
Published: April 2014

Low molecular weight dextran sulfate (DXS) has been reported to inhibit the classical, alternative pathway as well as the mannan-binding lectin pathway of the complement system. Furthermore, it acts as an endothelial cell protectant inhibiting complement-mediated endothelial cell damage. Endothelial cells are covered with a layer of heparan sulfate (HS), which is rapidly released under conditions of inflammation and tissue injury. Soluble HS induces maturation of dendritic cells (DC) via TLR4. In this study, we show the inhibitory effect of DXS on human DC maturation. DXS significantly prevents phenotypic maturation of monocyte-derived DC and peripheral myeloid DC by inhibiting the up-regulation of CD40, CD80, CD83, CD86, ICAM-1, and HLA-DR and down-regulates DC-SIGN in response to HS or exogenous TLR ligands. DXS also inhibits the functional maturation of DC as demonstrated by reduced T cell proliferation, and strongly impairs secretion of the proinflammatory mediators IL-1β, IL-6, IL-12p70, and TNF-α. Exposure to DXS leads to a reduced production of the complement component C1q and a decreased phagocytic activity, whereas C3 secretion is increased. Moreover, DXS was found to inhibit phosphorylation of IκB-α and activation of NF-κB. These findings suggest that DXS prevents TLR-induced maturation of human DC and may therefore be a useful reagent to impede the link between innate and adaptive immunity.
Oral or Nasal Antigen Induces Regulatory T Cells That Suppress Arthritis and Proliferation of Arthritogenic T Cells in Joint Draining Lymph Nodes
Authors: Broere Femke , Wieten Lotte , Klein Koerkamp Elles I. , van Roon Joel A. G. , Guichelaar Teun , Lafeber Floris P. J. G. , van Eden Willem
Published: April 2014

The propagation of mucosal tolerance as a therapeutic approach in autoimmune diseases remains a difficult goal to achieve, and therefore further mechanistic studies are necessary to develop potential clinical protocols to induce mucosal regulatory T cells (Tr cells). In this study we addressed whether oral or nasal proteoglycan induced functional Tr cells in the cartilage proteoglycan-induced chronic arthritis model. Both nasal and oral application of human proteoglycan before induction of disease suppressed arthritis severity and incidence. Tolerized mice showed enhanced numbers of IL-10 producing CD4+ cells in the paw-draining lymph nodes. Furthermore, CD4+ spleen cells displayed enhanced expression of molecules associated with Tr cells, such as IL-10, Foxp3, and TGF-β. Transfer of CD4+ spleen cells from mucosally tolerized donors into proteoglycan-immunized mice abolished arthritis and reduced humoral responses, indicative of Tr cells with the capacity to inhibit already induced immune responses. Tr cells were activated upon transfer, because enhanced proliferation was observed in the joint draining lymph nodes compared with activated T cells from nontolerized donors. Upon cotransfer with naive proteoglycan-specific T cells, mucosally induced Tr cells inhibited proliferation of these arthritogenic T cells in vivo. Herein we show that both oral and nasal Ag application induced Tr cells, which had a direct inhibitory effect on already established pathogenic B and T cell responses.
Histone Acetylation and Flagellin Are Essential for Legionella pneumophila-Induced Cytokine Expression
Authors: Schmeck Bernd , Lorenz Janina , N′Guessan Philippe Dje , Opitz Bastian , van Laak Vincent , Zahlten Janine , Slevogt Hortense , Witzenrath Martin , Flieger Antje , Suttorp Norbert , Hippenstiel Stefan
Published: April 2014

Legionella pneumophila causes severe pneumonia. Acetylation of histones is thought to be an important regulator of gene transcription, but its impact on L. pneumophila-induced expression of proinflammatory cytokines is unknown. L. pneumophila strain 130b induced the expression of the important chemoattractant IL-8 and genome-wide histone modifications in human lung epithelial A549 cells. We analyzed the IL-8-promoter and found that histone H4 was acetylated and H3 was phosphorylated at Ser10 and acetylated at Lys14, followed by transcription factor NF-κB. Recruitment of RNA polymerase II to the IL-8 promoter corresponded with increases in gene transcription. Histone modification and IL-8 release were dependent on p38 kinase and NF-κB pathways. Legionella-induced IL-8 expression was decreased by histone acetylase (HAT) inhibitor anacardic acid and enhanced by histone deacetylase (HDAC) inhibitor trichostatin A. After Legionella infection, HATs p300 and CREB-binding protein were time-dependently recruited to the IL-8 promoter, whereas HDAC1 and HDAC5 first decreased and later reappeared at the promoter. Legionella specifically induced expression of HDAC5 but not of other HDACs in lung epithelial cells, but knockdown of HDAC1 or 5 did not alter IL-8 release. Furthermore, Legionella-induced cytokine release, promoter-specific histone modifications, and RNA polymerase II recruitment were reduced in infection with flagellin-deletion mutants. Legionella-induced histone modification as well as HAT-/HDAC-dependent IL-8 release could also be shown in primary lung epithelial cells. In summary, histone acetylation seems to be important for the regulation of proinflammatory gene expression in L. pneumophila infected lung epithelial cells. These pathways may contribute to the host response in Legionnaires’ disease.
Transfusion of IgG-Opsonized Foreign Red Blood Cells Mediates Reduction of Antigen-Specific B Cell Priming in a Murine Model
Authors: Brinc Davor , Le-Tien Hoang , Crow Andrew R. , Siragam Vinayakumar , Freedman John , Lazarus Alan H.
Published: April 2014

Hemolytic disease of the fetus and newborn can be effectively prevented by administration of anti-D to the mother. The administered IgG results in the attenuation of RBC-specific Ab production, a process termed Ab-mediated immune suppression (AMIS). Because in animal models of AMIS no major effect on T cell priming occurs, we hypothesized that the effect of the IgG on the immune system under AMIS conditions may involve a deficiency in B cell priming. We therefore challenged mice with either untreated RBCs or IgG-opsonized RBCs (AMIS) and assessed B cell priming. B cells from mice transfused with untreated RBCs, but not from mice treated under AMIS conditions, were primed as assessed by their ability to function as Ag-specific APCs to appropriate T cells. To our knowledge, this is the first report demonstrating that AMIS inhibits the appearance of Ag-primed RBC-specific B cells.
IL-13 Production by Regulatory T Cells Protects against Experimental Autoimmune Encephalomyelitis Independently of Autoantigen
Authors: Ochoa-Repáraz Javier , Rynda Agnieszka , Ascón Miguel A. , Yang Xinghong , Kochetkova Irina , Riccardi Carol , Callis Gayle , Trunkle Theresa , Pascual David W.
Published: April 2014

Treatment with an anti-inflammatory Salmonella vaccine expressing enterotoxigenic Escherichia coli colonization factor Ag 1 (CFA/I) proved effective in stimulating protective, potent CD25+CD4+ regulatory T (Treg) cells in susceptible mice challenged with experimental autoimmune encephalomyelitis (EAE). Because the Salmonella vector was considerably less protective, we questioned whether altering fimbrial subunit expression to resemble conventional Salmonella expression may impact Treg cell potency. The Salmonella-CFA/I vaccine was modified to limit fimbrial subunit expression to the intracellular compartment (Salmonella-CFA/IIC). SJL mice were challenged with proteolipid protein peptide 139–151 to induce EAE and orally treated with one of three Salmonella vaccines 6 days postchallenge. Treatment with Salmonella-CFA/IIC greatly reduced clinical disease, similarly as Salmonella-CFA/I, by subduing IL-17 and IL-21; however, mechanisms of protection differed as evident by increased IL-13 and IFN-γ but diminished TGF-β production by Treg cells from Salmonella-CFA/IIC-treated mice. Adoptive transfer of Treg cells from both CFA/I-expressing constructs was equivalent in protecting against EAE, showing minimal disease. Although not as potent in its protection, CD25−CD4+ T cells from Salmonella-CFA/IIC showed minimal Th2 cells, but vaccination did prime these Th2 cells rendering partial protection against EAE challenge. In vivo IL-13 but not IFN-γ neutralization compromised protection conferred by adoptive transfer with Salmonella-CFA/IIC-induced Treg cells. Thus, the Salmonella-CFA/IIC vaccine elicits Treg cells with attributes from both the Salmonella vector and Salmonella-CFA/I vaccines. Importantly, these Treg cells can be induced to high potency by simply vaccinating against irrelevant Ags, offering a novel approach to treat autoimmune diseases independently of the autoantigen.
Medroxyprogesterone Acetate Inhibits CD8+ T Cell Viral-Specific Effector Function and Induces Herpes Simplex Virus Type 1 Reactivation
Authors: Cherpes Thomas L. , Busch James L. , Sheridan Brian S. , Harvey Stephen A. K. , Hendricks Robert L.
Published: April 2014

Clinical research suggests hormonal contraceptive use is associated with increased frequencies of HSV reactivation and shedding. We examined the effects of medroxyprogesterone acetate (MPA), the compound most commonly used for injectable hormonal contraception, on HSV type 1 (HSV-1) reactivation and CD8+ T cell function in murine trigeminal ganglia (TG). In ex vivo TG cultures, MPA dramatically inhibited canonical CD8+ T cell effector functions, including IFN-γ production and lytic granule release, and increased HSV-1 reactivation from latency. In vivo, MPA treatment of latently infected ovariectomized mice inhibited IFN-γ production and lytic granule release by TG resident CD8+ T cells stimulated directly ex vivo. RNA specific for the essential immediate early viral gene ICP4 as well as viral genome DNA copy number were increased in mice that received MPA during latency, suggesting that treatment increased in vivo reactivation. The increase in HSV-1 copy number appeared to be the result of a two-tine effect, as MPA induced higher reactivation frequencies from latently infected explanted TG neurons in the presence or absence of CD45+ cells. Our data suggest hormonal contraceptives that contain MPA may promote increased frequency of HSV reactivation from latency through the combinatory effects of inhibiting protective CD8+ T cell responses and by a leukocyte-independent effect on infected neurons.
Increased Frequency of EBV-Specific Effector Memory CD8+ T Cells Correlates with Higher Viral Load in Rheumatoid Arthritis
Authors: Lünemann Jan D. , Frey Oliver , Eidner Thorsten , Baier Michael , Roberts Susanne , Sashihara Junji , Volkmer Rudolf , Cohen Jeffrey I. , Hein Gert , Kamradt Thomas , Münz Christian
Published: April 2014

EBV is a candidate trigger of rheumatoid arthritis (RA). We determined both EBV-specific T cell and B cell responses and cell-associated EBV DNA copies in patients with RA and demographically matched healthy virus carriers. Patients with RA showed increased and broadened IgG responses to lytic and latent EBV-encoded Ags and 7-fold higher levels of EBV copy numbers in circulating blood cells. Additionally, patients with RA exhibited substantial expansions of CD8+ T cells specific for pooled EBV Ags expressed during both B cell transformation and productive viral replication and the frequency of CD8+ T cells specific for these Ags correlated with cellular EBV copy numbers. In contrast, CD4+ T cell responses to EBV and T cell responses to human CMV Ags were unchanged, altogether arguing against a defective control of latent EBV infection in RA. Our data show that the regulation of EBV infection is perturbed in RA and suggest that increased EBV-specific effector T cell and Ab responses are driven by an elevated EBV load in RA.
Auditing Protein Therapeutics Management by Professional APCs: Toward Prevention of Immune Responses against Therapeutic Proteins
Authors: Dasgupta Suryasarathi , Bayry Jagadeesh , André Sebastien , Dimitrov Jordan D. , Kaveri Srinivas V. , Lacroix-Desmazes Sebastien
Published: April 2014

Alloimmunization is a crippling concern in the management of patients undergoing administration of protein therapeutics as evidenced in replacement therapy and other treatment procedures. Several issues in the genesis and modulation of such deleterious immune responses have been studied. While authors have focused on the downstream events of the specific immune response and suggested modification of protein therapeutics to eliminate epitopes that interact with B cell receptors, T cell receptors, or MHCII molecules, the mechanisms underlying Ag interaction with APCs, a step upstream of immune effectors, have been grossly neglected. We wish to emphasize that the recent knowledge in understanding the capacities of an APC to handle an Ag and the importance of the surrounding microenvironment in this process are crucial for designing novel protein therapeutics with reduced immunogenicity.
Cutting Edge: Multiple Sclerosis-Like Lesions Induced by Effector CD8 T Cells Recognizing a Sequestered Antigen on Oligodendrocytes
Authors: Saxena Amit , Bauer Jan , Scheikl Tanja , Zappulla Jacques , Audebert Marc , Desbois Sabine , Waisman Ari , Lassmann Hans , Liblau Roland S. , Mars Lennart T.
Published: April 2014

CD8 T cells are emerging as important players in multiple sclerosis (MS) pathogenesis, although their direct contribution to tissue damage is still debated. To assess whether autoreactive CD8 T cells can contribute to the pronounced loss of oligodendrocytes observed in MS plaques, we generated mice in which the model Ag influenza hemagglutinin is selectively expressed in oligodendrocytes. Transfer of preactivated hemagglutinin-specific CD8 T cells led to inflammatory lesions in the optic nerve, spinal cord, and brain. These lesions, associating CD8 T cell infiltration with focal loss of oligodendrocytes, demyelination, and microglia activation, were very reminiscent of active MS lesions. Thus, our study demonstrates the potential of CD8 T cells to induce oligodendrocyte lysis in vivo as a likely consequence of direct Ag-recognition. These results provide new insights with regard to CNS tissue damage mediated by CD8 T cells and for understanding the role of CD8 T cells in MS.
Cutting Edge: Priming of NK Cells by IL-18
Authors: Chaix Julie , Tessmer Marlowe S. , Hoebe Kasper , Fuséri Nicolas , Ryffel Bernhard , Dalod Marc , Alexopoulou Lena , Beutler Bruce , Brossay Laurent , Vivier Eric , Walzer Thierry
Published: April 2014

Recent evidence suggests that NK cells require priming to display full effector activity. In this study, we demonstrate that IL-18 contributed to this phenomenon. IL-18 signaling-deficient NK cells were found to be unable to secrete IFN-γ in response to ex vivo stimulation with IL-12. This was not due to a costimulatory role of IL-18, because blocking IL-18 signaling during the ex vivo stimulation with IL-12 did not alter IFN-γ production by wild-type NK cells. Rather, we demonstrate that IL-18 primes NK cells in vivo to produce IFN-γ upon subsequent stimulation with IL-12. Importantly, IL-12-induced IFN-γ transcription by NK cells was comparable in IL-18 signaling-deficient and -sufficient NK cells. This suggests that priming by IL-18 leads to an improved translation of IFN-γ mRNA. These results reveal a novel type of cooperation between IL-12 and IL-18 that requires the sequential action of these cytokines.
Type I IFN-Induced, NKT Cell-Mediated Negative Control of CD8 T Cell Priming by Dendritic Cells
Authors: Bochtler Petra , Kröger Andrea , Schirmbeck Reinhold , Reimann Jörg
Published: April 2014

We investigated the negative effect of type I IFN (IFN-I) on the priming of specific CD8 T cell immunity. Priming of murine CD8 T cells is down-modulated if Ag is codelivered with IFN-I-inducing polyinosinic:polycytidylic acid (pI/C) that induces (NK cell- and T/B cell-independent) acute changes in the composition and surface phenotype of dendritic cells (DC). In wild-type but not IFN-I receptor-deficient mice, pI/C reduces the plasmacytoid DC but expands the CD8+ conventional DC (cDC) population and up-regulates surface expression of activation-associated (CD69, BST2), MHC (class I/II), costimulator (CD40, CD80/CD86), and coinhibitor (PD-L1/L2) molecules by cDC. Naive T cells are efficiently primed in vitro by IFN-I-stimulated CD8 cDC (the key APC involved in CD8 T cell priming) although these DC produced less IL-12 p40 and IL-6. pI/C (IFN-I)-mediated down modulation of CD8 T cell priming in vivo was not observed in NKT cell-deficient CD1d−/− mice. CD8 cDC from pI/C-treated mice inefficiently stimulated IFN-γ, IL-4, and IL-2 responses of NKT cells. In vitro, CD8 cDC that had activated NKT cells in the presence of IFN-I primed CD8 T cells that produced less IFN-γ but more IL-10. The described immunosuppressive effect of IFN-I thus involves an NKT cell-mediated change in the phenotype of CD8 cDC that favors priming of IL-10-producing CD8 T cells. In the presence of IFN-I, NKT cells hence impair the competence of CD8 cDC to prime proinflammatory CD8 T cell responses.
The Transcription Factor Fli-1 Modulates Marginal Zone and Follicular B Cell Development in Mice
Authors: Zhang Xian K. , Moussa Omar , LaRue Amanda , Bradshaw Sarah , Molano Ivan , Spyropoulos Demetri D. , Gilkeson Gary S. , Watson Dennis K.
Published: April 2014

Fli-1 belongs to the Ets transcription factor family and is expressed primarily in hematopoietic cells, including most cells active in immunity. To assess the role of Fli-1 in lymphocyte development in vivo, we generated mice that express a truncated Fli-1 protein, lacking the C-terminal transcriptional activation domain (Fli-1ΔCTA). Fli-1ΔCTA/Fli-1ΔCTA mice had significantly fewer splenic follicular B cells, and an increased number of transitional and marginal zone B cells, compared with wild-type controls. Bone marrow reconstitution studies demonstrated that this phenotype is the result of lymphocyte intrinsic effects. Expression of Igα and other genes implicated in B cell development, including Pax-5, E2A, and Egr-1, are reduced, while Id1 and Id2 are increased in Fli-1ΔCTA/Fli-1ΔCTA mice. Proliferation of B cells from Fli-1ΔCTA/Fli-1ΔCTA mice was diminished, although intracellular Ca2+ flux in B cells from Fli-1ΔCTA/Fli-1ΔCTA mice was similar to that of wild-type controls after anti-IgM stimulation. Immune responses and in vitro class switch recombination were also altered in Fli-1ΔCTA/Fli-1ΔCTA mice. Thus, Fli-1 modulates B cell development both centrally and peripherally, resulting in a significant impact on the in vivo immune response.
Allele-Selective Effect of PA28 in MHC Class I Antigen Processing
Authors: Yamano Taketoshi , Sugahara Hidetoshi , Mizukami Shusaku , Murata Shigeo , Chiba Tomoki , Tanaka Keiji , Yui Katsuyuki , Udono Heiichiro
Published: April 2014

PA28 is an IFN-γ-inducible proteasome activator and its genetic ablation causes complete loss of processing of certain Ags, but not all of them. The reason why this occurs and how PA28 influences the formation of peptide repertoires for MHC class I molecules remains unknown. In this study, we show the allele-specific role of PA28 in Ag processing. Retrovirus-transduced overexpression of PA28α decreased expression of Kd (Dd) while it increased Kb and Ld on the cell surface. By contrast, overexpression of PA28αΔC5, a mutant carrying a deletion of its five C-terminal residues and capable of attenuating the activity of endogenous PA28, produced the opposite effect on expression of those MHC class I molecules. Moreover, knockdown of both PA28α and β by small-interfering RNA profoundly augmented expression of Kd and Dd, but not of Ld, on the cell surface. Finally, we found that PA28-associated proteasome preferentially digested within epitopic sequences of Kd, although correct C-terminal flankings were removed, which in turn hampered production of Kd ligands. Our results indicate that whereas PA28 negatively influences processing of Kd (Dd) ligands, thereby, down-regulating Ag presentation by those MHC class I molecules, it also efficiently produces Kb (Ld) epitopes, leading to up-regulation of the MHC molecules.
HIF-1α Is Up-Regulated in Activated Mast Cells by a Process That Involves Calcineurin and NFAT
Authors: Walczak-Drzewiecka Aurelia , Ratajewski Marcin , Wagner Waldemar , Dastych Jaroslaw
Published: April 2014

Mast cells play important roles in many pathological conditions where local hypoxia is observed, including asthma, rheumatic diseases, and certain types of cancer. Here, we investigated how expression of the hypoxia-inducible factor 1, α subunit gene (HIF1A), is regulated in mast cells. The product of HIF1A is hypoxia-inducible factor 1α (HIF-1α), is a major nuclear transcription factor modulating gene expression in response to hypoxic conditions. We observed that under hypoxic conditions, exposure of mast cells to ionomycin and substance P resulted in significant up-regulation of HIF1A expression as compared with resting mast cells incubated under identical conditions. The ionomycin-mediated increase in HIF-1α protein levels was sensitive to the transcription inhibitor actinomycin D and to inhibitors of calcineurin, cyclosporin A (CsA), and FK506. The increased HIF-1α protein level was paralleled by a severalfold increase in HIF-1α mRNA that could be also inhibited with actinomycin D and CsA. The HIF1A promoter activity was significantly increased in ionomycin-activated mast cells, and the promoter activity could be inhibited by CsA and FK506. Furthermore, in situ mutagenesis experiments showed that the ionomycin-mediated HIF1A promoter activity depends on a conservative NFAT-binding site. Thus, accumulation of HIF-1α in activated mast cells requires up-regulation of HIF1A gene transcription and depends on the calcineurin-NFAT signaling pathway.
Acquisition of Suppressive Function by Activated Human CD4+CD25− T Cells Is Associated with the Expression of CTLA-4 Not FoxP3
Authors: Zheng Yong , Manzotti Claire N. , Burke Fiona , Dussably Laure , Qureshi Omar , Walker Lucy S. K. , Sansom David M.
Published: April 2014

The role of CTLA-4 in regulatory T cell (Treg) function is not well understood. We have examined the role of CTLA-4 and its relationship with the transcription factor FoxP3 using a model of Treg induction in human peripheral blood. Activation of human CD4+CD25− T cells resulted in the appearance of a de novo population of FoxP3-expressing cells within 48 h. These cells expressed high levels of CTLA-4 and cell sorting on expression of CTLA-4 strongly enriched for FoxP3+-expressing cells with suppressive function. Culture in IL-2 alone also generated cells with suppressive capacity that also correlated with the appearance of CTLA-4. To directly test the role of CTLA-4, we transfected resting human T cells with CTLA-4 and found that this method conferred suppression, similar to that of natural Tregs, even though these cells did not express FoxP3. Furthermore, transfection of FoxP3 did not induce CTLA-4 and these cells were not suppressive. By separating the expression of CTLA-4 and FoxP3, our data show that FoxP3 expression alone is insufficient to up-regulate CTLA-4; however, activation of CD4+CD25− T cells can induce both FoxP3 and CTLA-4 in a subpopulation of T cells that are capable of suppression. These data suggest that the acquisition of suppressive behavior by activated CD4+CD25− T cells requires the expression of CTLA-4, a feature that appears to be facilitated by, but is not dependent on, expression of FoxP3.
IFN-γ Induces the Erosion of Preexisting CD8 T Cell Memory during Infection with a Heterologous Intracellular Bacterium
Authors: Dudani Renu , Murali-Krishna Kaja , Krishnan Lakshmi , Sad Subash
Published: April 2014

Memory T cells are critical for the control of intracellular pathogens and require few signals for maintenance; however, erosion of established preexisting memory CD8+ T cells has been shown to occur during infection with heterologous viral infections. We evaluated whether this also occurs during infection with various intracellular bacteria and what mechanisms may be involved. We demonstrate that erosion of established memory is also induced during infection of mice with various intracellular bacteria, such as Listeria monocytogenes, Salmonella typhimurium, and Mycobacterium bovis (bacillus Calmette-Guérin). The extent of erosion of established CD8+ T cell memory was dependent on the virulence of the heterologous pathogen, not persistence. Furthermore, when antibiotics were used to comprehensively eliminate the heterologous pathogen, the numbers of memory CD8+ T cells were not restored, indicating that erosion of preexisting memory CD8+ T cells was irreversible. Irrespective of the initial numbers of memory CD8+ T cells, challenge with the heterologous pathogen resulted in a similar extent of erosion of memory CD8+ T cells, suggesting that cellular competition was not responsible for erosion. After challenge with the heterologous pathogen, effector memory CD8+ T cells were rapidly eliminated. More importantly, erosion of preexisting memory CD8+ T cells was abrogated in the absence of IFN-γ. These studies help reveal the paradoxical role of IFN-γ. Although IFN-γ promotes the control of intracellular bacterial replication during primary infection, this comes at the expense of erosion of preexisting memory CD8+ T cells in the wake of infection with heterologous pathogens.
The Agonists of TLR4 and 9 Are Sufficient to Activate Memory B Cells to Differentiate into Plasma Cells In Vitro but Not In Vivo
Authors: Richard Katharina , Pierce Susan K. , Song Wenxia
Published: April 2014

Memory B cells can persist for a lifetime and be reactivated to yield high affinity, isotype switched plasma cells. The generation of memory B cells by Ag immunization requires adjuvants that generally contain TLR agonists. However, requirements for memory B cell activation and the role of TLRs in this activation are not well understood. In this study, we analyzed the response of memory B cells from immunized mice to TLR9 and 4 agonists CpG oligodeoxynucleotides (ODN) and LPS. Mouse memory B cells express both TLR9 and 4, and respond to both CpG ODN and LPS in vitro by differentiating into high affinity IgG secreting plasma cells. In contrast, neither CpG ODN nor LPS alone is sufficient to activate memory B cells in vivo. Ag is required for the clonal expansion of Ag-specific memory B cells, the differentiation of memory B cells to high affinity IgG secreting plasma cells, and the recall of high affinity Ab responses. The Ag-specific B cells that have not yet undergone isotype switching showed a relatively higher expression of TLR4 than memory B cells, which was reflected in a heightened response to LPS, but in both cases yielded mostly low affinity IgM secreting plasma cells. Thus, although memory B cells are sensitive to TLR agonists in vitro, TLR agonists alone appear to have little affect on B cell memory in vivo.
Influence of a Non-NK Complex Region of Chromosome 6 on CD4+ Invariant NK T Cell Homeostasis
Authors: Vallois David , Gagnerault Marie-Claude , Avner Philip , Rogner Ute C. , Boitard Christian , Benlagha Kamel , Herbelin André , Lepault Françoise
Published: April 2014

The number and function of immunoregulatory invariant NKT (iNKT) cells are genetically controlled. A defect of iNKT cell ontogeny and function has been implicated as one causal factor of NOD mouse susceptibility to type 1 diabetes. Other factors of diabetes susceptibility, such as a decrease of regulatory T cell function or an increase in TLR1 expression, are corrected in diabetes-resistant Idd6 NOD.C3H 6.VIII congenic mice. Thus, we surmised that the iNKT cell defects found in NOD mice may also be rescued in congenic mice. Unexpectedly, we found, in both the thymus and the periphery, a 50% reduction in iNKT cell number in NOD.C3H 6.VIII mice as compared with NOD mice. This reduction only affected CD4+ iNKT cells, and left the double negative iNKT cells unchanged. In parallel, the production of IL-4 and IFN-γ following α-GalCer stimulation was proportionally reduced. Using three subcongenic strains, we have narrowed down the region controlling iNKT development within Idd6 (5.8 Mb) to Idd6.2 region (2.5 Mb). Idd6 region had no effect on NK cell number and in vivo cytotoxic activity. These results indicate that the role of iNKT cells in diabetes development is equivocal and more complex than initially considered. In addition, they bring strong evidence that the regulation of CD4+ iNKT cell production is independent from that of DN iNKT cells, and involves genes of the Idd6 locus.
IL-21-Induced Isotype Switching to IgG and IgA by Human Naive B Cells Is Differentially Regulated by IL-4
Authors: Avery Danielle T. , Bryant Vanessa L. , Ma Cindy S. , de Waal Malefyt Rene , Tangye Stuart G.
Published: April 2014

Naive B cells can alter the effector function of their Ig molecule by isotype switching, thereby allowing them to secrete not only IgM, but also the switched isotypes IgG, IgA, and IgE. Different isotypes are elicited in response to specific pathogens. Similarly, dysregulated production of switched isotypes underlies the development of various diseases, such as autoimmunity and immunodeficiency. Thus, it is important to characterize mediators controlling isotype switching, as well as their contribution to the overall B cell response. Isotype switching in human naive B cells can be induced by CD40L together with IL-4, IL-10, IL-13, and/or TGF-β. Recently, IL-21 was identified as a switch factor for IgG1 and IgG3. However, the effect of IL-21 on switching to IgA, as well as the interplay between IL-21 and other switch factors, remains unknown. We found that IL-4 and IL-21 individually induced CD40L-stimulated human naive B cells to undergo switching to IgG, with IL-4 predominantly inducing IgG1+ cells and IL-21 inducing IgG3. Culture of naive B cells with CD40L and IL-21, but not IL-4, also yielded IgA+ cells. Combining IL-4 and IL-21 had divergent effects on isotype switching. Specifically, while IL-4 and IL-21 synergistically increased the generation of IgG1+ cells from CD40L-stimulated B cells, IL-4 concomitantly abolished IL-21-induced switching to IgA. Our findings demonstrate the dynamic interplay between IL-4 and IL-21 in regulating the production of IgG subclasses and IgA, and suggest temporal roles for these cytokines in humoral immune responses to specific pathogens.
The E3 Ubiquitin Ligase Ro52 Negatively Regulates IFN-β Production Post-Pathogen Recognition by Polyubiquitin-Mediated Degradation of IRF3
Authors: Higgs Rowan , Gabhann Joan Ní , Larbi Nadia Ben , Breen Eamon P. , Fitzgerald Katherine A. , Jefferies Caroline A.
Published: April 2014

Induction of type I IFNs is a fundamental cellular response to both viral and bacterial infection. The role of the transcription factor IRF3 is well established in driving this process. However, equally as important are cellular mechanisms for turning off type I IFN production to limit this response. In this respect, IRF3 has previously been shown to be targeted for ubiquitin-mediated degradation postviral detection to turn off the IFN-β response. In this study, we provide evidence that the E3 ligase Ro52 (TRIM21) targets IRF3 for degradation post-pathogen recognition receptor activation. We demonstrate that Ro52 interacts with IRF3 via its C-terminal SPRY domain, resulting in the polyubiquitination and proteasomal degradation of the transcription factor. Ro52-mediated IRF3 degradation significantly inhibits IFN-β promoter activity, an effect that is reversed in the presence of the proteasomal inhibitor MG132. Specific targeting of Ro52 using short hairpin RNA rescues IRF3 degradation following polyI:C-stimulation of HEK293T cells, with a subsequent increase in IFN-β production. Additionally, shRNA targeting of murine Ro52 enhances the production of the IRF3-dependent chemokine RANTES following Sendai virus infection of murine fibroblasts. Collectively, this demonstrates a novel role for Ro52 in turning off and thus limiting IRF3-dependent type I IFN production by targeting the transcription factor for polyubiquitination and subsequent proteasomal degradation.
CTLA4 Expression Is an Indicator and Regulator of Steady-State CD4+FoxP3+ T Cell Homeostasis
Authors: Tang Anita L. , Teijaro John R. , Njau Modesta N. , Chandran Smita S. , Azimzadeh Agnes , Nadler Steven G. , Rothstein David M. , Farber Donna L.
Published: April 2014

The presence of FoxP3+ regulatory T cells (Tregs) is necessary for control of deleterious immune responses in the steady state; however, mechanisms for maintaining the frequency and quality of endogenous Tregs are not well defined. In this study, we used in vivo modulators of the CD28 and CTLA4 pathways administered to intact mice to reveal mechanisms controlling the homeostasis and phenotype of endogenous Tregs. We demonstrate that expression of the negative costimulatory regulator CTLA4 on FoxP3+ Tregs in vivo is a direct consequence of their rapid, perpetual homeostasis. Up-regulation of CTLA4 expression occurs only on FoxP3+ Tregs undergoing extensive proliferation and can be abrogated by inhibiting the CD28 pathway, coinciding with a reduction in FoxP3+ Treg proliferation and frequency. We further demonstrate that CTLA4 negatively regulates steady-state Treg homeostasis, given that inhibiting CTLA4 signaling with an anti-CTLA4 blocking Ab greatly enhances Treg proliferation and overall Treg frequency. Our findings provide new insight into the origin and role of CTLA4 expression on natural FoxP3+ Tregs and reveal opposing effects of costimulation modulators on the steady-state level and quality of Tregs, with implications regarding their effects on endogenous Tregs in patients receiving immunotherapy.
Endometrial NK Cells Are Special Immature Cells That Await Pregnancy
Authors: Manaster Irit , Mizrahi Saar , Goldman-Wohl Debra , Sela Hen Y. , Stern-Ginossar Noam , Lankry Dikla , Gruda Raizy , Hurwitz Arye , Bdolah Yuval , Haimov-Kochman Ronit , Yagel Simcha , Mandelboim Ofer
Published: April 2014

NK cells populate the human endometrium before pregnancy. Unlike decidual NK cells that populate the decidua during pregnancy, the NK cells present in the human endometrium, before pregnancy, have not been fully characterized. In this study, we provide a detailed analysis of the origin, phenotype, and function of endometrial NK cells (eNK). We show that eNK cells have a unique receptor repertoire. In particular, they are negative for NKp30 and chemokine receptor expression, which distinguishes them from any other NK subset described so far. We further show that eNK cells lack NK-specific functional phenotype and activity such as cytokine secretion and cytotoxicity, before IL-15 stimulation. Following such stimulation, endometrial NK cells acquire phenotype and function that are similar to those of decidual NK cells. We therefore suggest that eNK cells are inactive cells (before IL-15 activation and in relation to the known NK activity) that are present in the endometrium before conception, waiting for pregnancy.
Th1, Th2, and Th17 Effector T Cell-Induced Autoimmune Gastritis Differs in Pathological Pattern and in Susceptibility to Suppression by Regulatory T Cells
Authors: Stummvoll Georg H. , DiPaolo Richard J. , Huter Eva N. , Davidson Todd S. , Glass Deborah , Ward Jerrold M. , Shevach Ethan M.
Published: April 2014

Th cells can be subdivided into IFN-γ-secreting Th1, IL-4/IL-5-secreting Th2, and IL-17-secreting Th17 cells. We have evaluated the capacity of fully differentiated Th1, Th2, and Th17 cells derived from a mouse bearing a transgenic TCR specific for the gastric parietal cell antigen, H+K+-ATPase, to induce autoimmune gastritis after transfer to immunodeficient recipients. We have also determined the susceptibility of the disease induced by each of the effector T cell types to suppression by polyclonal regulatory T cells (Treg) in vivo. Each type of effector cell induced autoimmune gastritis with distinct histological patterns. Th17 cells induced the most destructive disease with cellular infiltrates composed primarily of eosinophils accompanied by high levels of serum IgE. Polyclonal Treg could suppress the capacity of Th1 cells, could moderately suppress Th2 cells, but could suppress Th17-induced disease only at early time points. The major effect of the Treg was to inhibit the expansion of the effector T cells. However, effector cells isolated from protected animals were not anergic and were fully competent to proliferate and produce effector cytokines ex vivo. The strong inhibitory effect of polyclonal Treg on the capacity of some types of differentiated effector cells to induce disease provides an experimental basis for the clinical use of polyclonal Treg in the treatment of autoimmune disease in humans.
IL-2 Receptor β-Chain Signaling Controls Immunosuppressive CD4+ T Cells in the Draining Lymph Nodes and Lung during Allergic Airway Inflammation In Vivo
Authors: Doganci Aysefa , Karwot Roman , Maxeiner Joachim H. , Scholtes Petra , Schmitt Edgar , Neurath Markus F. , Lehr Hans Anton , Ho I-Cheng , Finotto Susetta
Published: April 2014

IL-2 influences both survival and differentiation of CD4+ T effector and regulatory T cells. We studied the effect of i.n. administration of Abs against the α- and the β-chains of the IL-2R in a murine model of allergic asthma. Blockade of the β- but not the α-chain of the IL-2R after allergen challenge led to a significant reduction of airway hyperresponsiveness. Although both treatments led to reduction of lung inflammation, IL-2 signaling, STAT-5 phosphorylation, and Th2-type cytokine production (IL-4 and IL-5) by lung T cells, IL-13 production and CD4+ T cell survival were solely inhibited by the blockade of the IL-2R β-chain. Moreover, local blockade of the common IL-2R/IL-15R β-chain reduced NK cell number and IL-2 production by lung CD4+CD25+ and CD4+CD25− T cells while inducing IL-10- and TGF-β-producing CD4+ T cells in the lung. This cytokine milieu was associated with reduced CD4+ T cell proliferation in the draining lymph nodes. Thus, local blockade of the β-chain of the IL-2R restored an immunosuppressive cytokine milieu in the lung that ameliorated both inflammation and airway hyperresponsiveness in experimental allergic asthma. These findings provide novel insights into the functional role of IL-2 signaling in experimental asthma and suggest that blockade of the IL-2R β-chain might be useful for therapy of allergic asthma in humans.
Lack of Plasma Protein Hemopexin Dampens Mercury-Induced Autoimmune Response in Mice
Authors: Fagoonee Sharmila , Caorsi Cristiana , Giovarelli Mirella , Stoltenberg Meredin , Silengo Lorenzo , Altruda Fiorella , Camussi Giovanni , Tolosano Emanuela , Bussolati Benedetta
Published: April 2014

Several factors affect the autoimmune response, including iron-dependent modulation of T cells. Hemopexin is the plasma protein with the highest binding affinity to heme. It mediates heme-iron recovery in the liver, thus controlling heme-iron availability in peripheral cells. The aim of the present study was to investigate the role of hemopexin in the progress of an autoimmune response. To this end, we chose a mouse model of mercury-induced autoimmunity and evaluated the susceptibility of hemopexin-null mice to mercury treatment compared with wild-type controls. In this study we show that lack of hemopexin dampens mercury-induced autoimmune responses in mice. Hemopexin-null mice produced fewer antinuclear autoantibodies and had reduced deposits of immune complexes in the kidney after mercuric chloride treatment compared with wild-type mice. These features were associated with a reduction in activated T cells and lower absolute B cell number in spleen and impaired IgG1 and IgG2a production. In contrast, in hemopexin-null mice the response to OVA/CFA immunization was maintained. In addition, hemopexin-null mice had reduced transferrin receptor 1 expression in T cells, possibly due to the increase in heme-derived iron. Interestingly, CD4+T cells isolated from mercury-treated hemopexin-null mice show reduced IFN-γ-dependent STAT1 phosphorylation compared with that of wild-type mice. Our data suggest that hemopexin, by controlling heme-iron availability in lymphocytes, modulates responsiveness to IFN-γ and, hence, autoimmune responses.