bluuebunny/crossref_metadata_2025 huggingface dataset:
CD11b+Ly-6Chi Suppressive Monocytes in Experimental Autoimmune Encephalomyelitis
Authors: Zhu Bing , Bando Yoshio , Xiao Sheng , Yang Kaiyong , Anderson Ana C. , Kuchroo Vijay K. , Khoury Samia J.
Published: April 2014

Innate immune cells may regulate adaptive immunity by balancing different lineages of T cells and providing negative costimulation. In addition, CD11b+Gr-1+ myeloid-derived suppressor cells have been described in tumor, parasite infection, and severe trauma models. In this study, we observe that splenic CD11b+ cells markedly increase after experimental autoimmune encephalomyelitis (EAE) immunization, and they suppress T cell proliferation in vitro. Although >80% of CD11b+ cells express varying levels of Gr-1, only a small population of CD11b+Ly-6Chigh inflammatory monocytes (IMC) can efficiently suppress T cell proliferation and induce T cell apoptosis through the production of NO. IFN-γ produced by activated T cells is essential to induce IMC suppressive function. EAE immunization increases the frequencies of IMC in the bone marrow, spleen, and blood, but not in the lymph nodes. At the peak of EAE, IMC represent ∼30% of inflammatory cells in the CNS. IMC express F4/80 and CD93 but not CD31, suggesting that they are immature monocytes. Furthermore, IMC have the plasticity to up-regulate NO synthase 2 or arginase 1 expression upon different cytokine treatments. These findings indicate that CD11b+Ly-6Chigh IMC induced during EAE priming are powerful suppressors of activated T cells. Further understanding of suppressive monocytes in autoimmune disease models may have important clinical implications for human autoimmune diseases.
Protein Kinase C Regulates Expression and Function of Inhibitory Killer Cell Ig-Like Receptors in NK Cells
Authors: Alvarez-Arias Diana A. , Campbell Kerry S.
Published: April 2014

The inhibitory killer cell Ig-like receptors (KIR) negatively regulate NK cell cytotoxicity by activating the Src homology 2 domain-containing protein tyrosine phosphatases 1 and 2 following ligation with MHC class I molecules expressed on normal cells. This requires tyrosine phosphorylation of KIR on ITIMs in the cytoplasmic domain. Surprisingly, we have found that KIR3DL1 is strongly and constitutively phosphorylated on serine and weakly on threonine residues. In this study, we have mapped constitutive phosphorylation sites for casein kinases, protein kinase C, and an unidentified kinase on the KIR cytoplasmic domain. Three of these phosphorylation sites are highly conserved in human inhibitory KIR. Functional studies of the wild-type receptor and serine/threonine mutants indicated that phosphorylation of Ser394 by protein kinase C slightly suppresses KIR3DL1 inhibitory function, and reduces receptor internalization and turnover. Our results provide evidence that serine/threonine phosphorylation is an important regulatory mechanism of KIR function.
A Fas-Associated Death Domain Protein/Caspase-8-Signaling Axis Promotes S-Phase Entry and Maintains S6 Kinase Activity in T Cells Responding to IL-2
Authors: Arechiga Adrian F. , Bell Bryan D. , Leverrier Sabrina , Weist Brian M. , Porter Melissa , Wu Zhengqi , Kanno Yuka , Ramos Stephanie J. , Ong S. Tiong , Siegel Richard , Walsh Craig M.
Published: April 2014

Fas-associated death domain protein (FADD) constitutes an essential component of TNFR-induced apoptotic signaling. Paradoxically, FADD has also been shown to be crucial for lymphocyte development and activation. In this study, we report that FADD is necessary for long-term maintenance of S6 kinase (S6K) activity. S6 phosphorylation at serines 240 and 244 was only observed after long-term stimulation of wild-type cells, roughly corresponding to the time before S-phase entry, and was poorly induced in T cells expressing a dominantly interfering form of FADD (FADDdd), viral FLIP, or possessing a deficiency in caspase-8. Defects in S6K1 phosphorylation were also observed. However, defective S6K1 phosphorylation was not a consequence of a wholesale defect in mammalian target of rapamycin function, because 4E-BP1 phosphorylation following T cell activation was unaffected by FADDdd expression. Although cyclin D3 up-regulation and retinoblastoma hypophosphorylation occurred normally in FADDdd T cells, cyclin E expression and cyclin-dependent kinase 2 activation were markedly impaired in FADDdd T cells. These results demonstrate that a FADD/caspase-8-signaling axis promotes T cell cycle progression and sustained S6K activity.
STAT5 Is Essential for Akt/p70S6 Kinase Activity during IL-2-Induced Lymphocyte Proliferation
Authors: Lockyer Heather M. , Tran Eric , Nelson Brad H.
Published: April 2014

IL-2R activates two distinct signaling pathways mediated by the adaptor protein Shc and the transcription factor STAT5. Prior mutagenesis studies of the IL-2R have indicated that the Shc and STAT5 pathways are redundant in the ability to induce lymphocyte proliferation. Yet paradoxically, T cells from STAT5-deficient mice fail to proliferate in response to IL-2, suggesting that the Shc pathway is unable to promote mitogenesis in the genetic absence of STAT5. Here we show in the murine lymphocyte cell line Ba/F3 that low levels of STAT5 activity are essential for Shc signaling. In the absence of STAT5 activity, Shc was unable to sustain activation of the Akt/p70S6 kinase pathway or promote lymphocyte proliferation and viability. Restoring STAT5 activity via a heterologous receptor rescued Shc-induced Akt/p70S6 kinase activity and cell proliferation with kinetics consistent with a transcriptional mechanism. Thus, STAT5 appears to regulate the expression of one or more unidentified components of the Akt pathway. Our results not only explain the severe proliferative defect in STAT5-deficient T cells but also provide mechanistic insight into the oncogenic properties of STAT5 in various leukemias and lymphomas.
Analysis of Epitope-Specific Immune Responses Induced by Vaccination with Structurally Folded and Unfolded Recombinant Bet v 1 Allergen Derivatives in Man
Authors: Pree Ines , Reisinger Jürgen , Focke Margit , Vrtala Susanne , Pauli Gabrielle , van Hage Marianne , Cromwell Oliver , Gadermaier Elisabeth , Egger Cornelia , Reider Norbert , Horak Friedrich , Valenta Rudolf , Niederberger Verena
Published: April 2014

Previously, we have constructed recombinant derivatives of the major birch pollen allergen, Bet v 1, with a more than 100-fold reduced ability to induce IgE-mediated allergic reactions. These derivatives differed from each other because the two recombinant Bet v 1 fragments represented unfolded molecules whereas the recombinant trimer resembled most of the structural fold of the Bet v 1 allergen. In this study, we analyzed the Ab (IgE, IgG subclass, IgA, IgM) response to Bet v 1, recombinant and synthetic Bet v 1-derived peptides in birch pollen allergic patients who had been vaccinated with the derivatives or adjuvant alone. Furthermore, we studied the induction of IgE-mediated skin responses in these patients using Bet v 1 and Bet v 1 fragments. Both types of vaccines induced a comparable IgG1 and IgG4 response against new sequential epitopes which overlap with the conformational IgE epitopes of Bet v 1. This response was 4- to 5-fold higher than that induced by immunotherapy with birch pollen extract. Trimer more than fragments induced also IgE responses against new epitopes and a transient increase in skin sensitivity to the fragments at the beginning of therapy. However, skin reactions to Bet v 1 tended to decrease one year after treatment in both actively treated groups. We demonstrate that vaccination with folded and unfolded recombinant allergen derivatives induces IgG Abs against new epitopes. These data may be important for the development of therapeutic as well as prophylactic vaccines based on recombinant allergens.
A Population of CD19highCD45R−/lowCD21low B Lymphocytes Poised for Spontaneous Secretion of IgG and IgA Antibodies
Authors: de Andrés Belén , Cortegano Isabel , Serrano Natalia , del Rio Borja , Martín Paloma , Gonzalo Pilar , Marcos Miguel A. R. , Gaspar María Luisa
Published: April 2014

Ab responses to selected Ags are produced by discrete B cell populations whose presence and functional relevance vary along the ontogeny. The earliest B lineage-restricted precursors in gestational day 11 mouse embryos display the CD19+CD45R/B220− phenotype. Phenotypically identical cells persist throughout gestation and in postnatal life, in parallel to the later-arising, CD19+CD45R+ B cells. Very early after birth, the CD19+CD45R− B cell subset included high frequencies of spontaneously Ig-secreting cells. In the adult spleen, a small subset of CD19highCD45R−/lowIgM+/−IgD−CD21/Cr2−/low cells, which was detected in perifollicular areas, displayed genetic and phenotypical traits of highly differentiated B cells, and was enriched in IgG- and IgA-secreting plasma cells. In vitro differentiation and in vivo adoptive transfer experiments of multipotent hemopoietic progenitors revealed that these CD19highCD45R−/low B cells were preferentially regenerated by embryo-, but not by adult bone marrow-, derived progenitors, except when the latter were inoculated into newborn mice. Both the early ontogenical emergence and the natural production of serum Igs, are shared features of this CD19highCD45R−/low B cell population with innate-like B lymphocytes such as B1 and marginal zone B cells, and suggest that the new population might be related to that category.
The Acquired Immune Response to the Mucosal Adjuvant LTK63 Imprints the Mouse Lung with a Protective Signature
Authors: Tritto Elaine , Muzzi Alessandro , Pesce Isabella , Monaci Elisabetta , Nuti Sandra , Galli Grazia , Wack Andreas , Rappuoli Rino , Hussell Tracy , De Gregorio Ennio
Published: April 2014

LTK63, a nontoxic mutant of Escherichia coli heat labile enterotoxin (LT), is a potent and safe mucosal adjuvant that has also been shown to confer generic protection to several respiratory pathogens. To understand the mechanisms of action underlying the LTK63 protective effect, we analyzed the molecular and cellular events triggered by its administration in vivo. We show here that LTK63 intrapulmonary administration induced in the mouse lung a specific gene expression signature characterized by the up-regulation of cell cycle genes, several host defense genes, chemokines, chemokine receptors, and immune cell-associated genes. Such a transcriptional profile reflected the activation of alveolar macrophages and the recruitment to the lung of T and B cells and innate immune cells such as granulocytes, NK, and dendritic cells. All of these events were T cell dependent and specific for LTK63 because they were absent in SCID and nude mice. Additionally, we showed that LTK63 induces a potent adaptive immune response against itself directed to the lung. We propose that acquired response to LTK63 is the driving force for the local recruitment of both adaptive and innate immune cells. Our data suggest that LTK63 acts as an airway infection mimic that establishes a generic protective environment limiting respiratory infection by innate immune mechanisms and by improving adaptive responses to invading pathogens.
Bone Marrow of Persistently Hepatitis C Virus-Infected Individuals Accumulates Memory CD8+ T Cells Specific for Current and Historical Viral Antigens: A Study in Patients with Benign Hematological Disorders
Authors: Racanelli Vito , Frassanito Maria Antonia , Leone Patrizia , Brunetti Claudia , Ruggieri Simona , Dammacco Franco
Published: April 2014

The role of virus-specific T cells in hepatitis C virus (HCV) pathogenesis is not clear. Existing knowledge on the frequency, phenotype, and behavior of these cells comes from analyses of blood and liver, but other lymphoid compartments that may be important sites for functionally mature T cells have not yet been analyzed. We studied HCV-specific T cells from bone marrow, in comparison to those from peripheral blood and liver biopsy tissue, from 20 persistently HCV-infected patients with benign hematological disorders. Bone marrow contained a sizeable pool of CD8+ T cells specific for epitopes from structural and nonstructural HCV proteins. These cells displayed the same effector memory phenotype as liver-derived equivalents and the same proliferative potential as blood-derived equivalents but had greater antiviral effector functions such as Ag-specific cytotoxicity and IFN-γ production. These features were not shared by influenza virus-specific CD8+ T cells in the same bone marrow samples. Despite their highly differentiated phenotype and activated status, some bone marrow-resident HCV-specific CD8+ T cells were not directed against the infecting virus but, instead, against historical HCV Ags (i.e., viral species of a previous infection or minor viral species of the current infection). These findings provide a snapshot view of the distribution, differentiation, and functioning of virus-specific memory T cells in patients with persistent HCV infection.
The Proinflammatory Cytokine Macrophage Migration Inhibitory Factor Regulates Glucose Metabolism during Systemic Inflammation
Authors: Atsumi Toshiya , Cho You-Ree , Leng Lin , McDonald Courtney , Yu Tim , Danton Cheryl , Hong Eun-Gyoung , Mitchell Robert A. , Metz Christine , Niwa Hirokatsu , Takeuchi Jun , Onodera Shin , Umino Tomomi , Yoshioka Narihito , Koike Takao , Kim Jason K. , Bucala Richard
Published: April 2014

Inflammation provokes significant abnormalities in host metabolism that result from the systemic release of cytokines. An early response of the host is hyperglycemia and resistance to the action of insulin, which progresses over time to increased glucose uptake in peripheral tissue. Although the cytokine TNF-α has been shown to exert certain catabolic effects, recent studies suggest that the metabolic actions of TNF-α occur by the downstream regulation of additional mediators, such as macrophage migration inhibitory factor (MIF). We investigated the glycemic responses of endotoxemic mice genetically deficient in MIF (MIF−/−). In contrast to wild-type mice, MIF−/− mice exhibit normal blood glucose and lactate responses following the administration of endotoxin, or TNF-α. MIF−/− mice also show markedly increased glucose uptake into white adipose tissue in vivo in the endotoxemic state. Treatment of adipocytes with MIF, or anti-MIF mAb, modulates insulin-mediated glucose transport and insulin receptor signal transduction; these effects include the phosphorylation of insulin receptor substrate-1, its association with the p85 regulatory subunit of PI3K, and the downstream phosphorylation of Akt. Genetic MIF deficiency also promotes adipogenesis, which is in accord with a downstream role for MIF in the action of TNF-α. These studies support an important role for MIF in host glucose metabolism during sepsis.
CD8 T Cells Inhibit Respiratory Syncytial Virus (RSV) Vaccine-Enhanced Disease
Authors: Olson Matthew R. , Varga Steven M.
Published: April 2014

Vaccination of children with a formalin-inactivated (FI) respiratory syncytial virus (RSV) vaccine led to exacerbated disease including pulmonary eosinophilia following a natural RSV infection. Immunization of BALB/c mice with FI-RSV or a recombinant vaccinia virus (vv) expressing the RSV attachment (G) protein (vvG) results in a pulmonary Th2 response and eosinophilia after RSV challenge that closely mimics the RSV vaccine-enhanced disease observed in humans. The underlying causes of RSV vaccine-enhanced disease remain poorly understood. We demonstrate here that RSV M2-specific CD8 T cells reduce the Th2-mediated pathology induced by vvG-immunization and RSV challenge in an IFN-γ-independent manner. We also demonstrate that FI-RSV immunization does not induce a measurable RSV-specific CD8 T cell response and that priming FI-RSV-immunized mice for a potent memory RSV-specific CD8 T cell response abrogates pulmonary eosinophilia after subsequent RSV challenge. Our results suggest that the failure of the FI-RSV vaccine to induce a CD8 T cell response may have contributed to the development of pulmonary eosinophilia and augmented disease that occurred in vaccinated individuals.
Activation of Innate Immune Defense Mechanisms by Signaling through RIG-I/IPS-1 in Intestinal Epithelial Cells
Authors: Hirata Yoshihiro , Broquet Alexis H. , Menchén Luis , Kagnoff Martin F.
Published: April 2014

Intestinal epithelial cells (IECs) are a first line of defense against microbial pathogens that enter the host through the intestinal tract. Moreover, viral pathogens that infect the host via the intestinal epithelium are an important cause of morbidity and mortality. However, the mechanisms by which viral pathogens activate antiviral defense mechanisms in IECs are largely unknown. The synthetic dsRNA analog polyinosinic-polycytidylic acid and infection with live virus were used to probe the molecules that are activated and the mechanisms of signaling in virus-infected human IECs. Polyinosinic-polycytidylic acid activated IFN regulatory factor 3 dimerization and phosphorylation, increased activity of the IFN-stimulated response element, induced a significant increase in IFN-β mRNA transcripts and IFN-β secretion, and up-regulated the expression of IFN-regulated genes in IECs. Those responses were dependent upon activation of the dsRNA binding protein retinoic acid inducible gene I (RIG-I) and the RIG-I interacting protein IFN promoter stimulator-1, but not on dsRNA-activated protein kinase or TLR3, which also were expressed by IECs. Virus replication and virus-induced cell death increased in IECs in which RIG-I was silenced, consistent with the importance of the RIG-I signaling pathway in IEC antiviral innate immune defense mechanisms.
Helicobacter pylori Vacuolating Cytotoxin Inhibits Activation-Induced Proliferation of Human T and B Lymphocyte Subsets
Authors: Torres Victor J. , VanCompernolle Scott E. , Sundrud Mark S. , Unutmaz Derya , Cover Timothy L.
Published: April 2014

Helicobacter pylori are Gram-negative bacteria that persistently colonize the human gastric mucosa despite the recruitment of immune cells. The H. pylori vacuolating cytotoxin (VacA) recently has been shown to inhibit stimulation-induced proliferation of primary human CD4+ T cells. In this study, we investigated effects of VacA on the proliferation of various other types of primary human immune cells. Intoxication of PBMC with VacA inhibited the stimulation-induced proliferation of CD4+ T cells, CD8+ T cells, and B cells. VacA also inhibited the proliferation of purified primary human CD4+ T cells that were stimulated by dendritic cells. VacA inhibited both T cell-induced and PMA/anti-IgM-induced proliferation of purified B cells. Intoxication with VacA did not alter the magnitude of calcium flux that occurred upon stimulation of CD4+ T cells or B cells, indicating that VacA does not alter early signaling events required for activation and proliferation. VacA reduced the mitochondrial membrane potential of CD4+ T cells, but did not reduce the mitochondrial membrane potential of B cells. We propose that the immunomodulatory actions of VacA on T and B lymphocytes, the major effectors of the adaptive immune response, may contribute to the ability of H. pylori to establish a persistent infection in the human gastric mucosa.
Specific Leukotriene Receptors Couple to Distinct G Proteins to Effect Stimulation of Alveolar Macrophage Host Defense Functions
Authors: Peres Camila M. , Aronoff David M. , Serezani Carlos H. , Flamand Nicolas , Faccioli Lucia H. , Peters-Golden Marc
Published: April 2014

Leukotrienes (LTs) are lipid mediators implicated in asthma and other inflammatory diseases. LTB4 and LTD4 also participate in antimicrobial defense by stimulating phagocyte functions via ligation of B leukotriene type 1 (BLT1) receptor and cysteinyl LT type 1 (cysLT1) receptor, respectively. Although both Gαi and Gαq proteins have been shown to be coupled to both BLT1 and cysLT1 receptors in transfected cell systems, there is little known about specific G protein subunit coupling to LT receptors, or to other G protein-coupled receptors, in primary cells. In this study we sought to define the role of specific G proteins in pulmonary alveolar macrophage (AM) innate immune responses to LTB4 and LTD4. LTB4 but not LTD4 reduced cAMP levels in rat AM by a pertussis toxin (PTX)-sensitive mechanism. Enhancement of FcγR-mediated phagocytosis and bacterial killing by LTB4 was also PTX-sensitive, whereas that induced by LTD4 was not. LTD4 and LTB4 induced Ca2+ and intracellular inositol monophosphate accumulation, respectively, highlighting the role of Gαq protein in mediating PTX-insensitive LTD4 enhancement of phagocytosis and microbicidal activity. Studies with liposome-delivered G protein blocking Abs indicated a dependency on specific Gαq/11 and Gαi3 subunits, but not Gαi2 or Gβγ, in LTB4-enhanced phagocytosis. The selective importance of Gαq/11 protein was also demonstrated in LTD4-enhanced phagocytosis. The present investigation identifies differences in specific G protein subunit coupling to LT receptors in antimicrobial responses and highlights the importance of defining the specific G proteins coupled to heptahelical receptors in primary cells, rather than simply using heterologous expression systems.
Identification of the IL-17 Receptor Related Molecule IL-17RC as the Receptor for IL-17F
Authors: Kuestner Rolf E. , Taft David W. , Haran Aaron , Brandt Cameron S. , Brender Ty , Lum Karen , Harder Brandon , Okada Shannon , Ostrander Craig D. , Kreindler James L. , Aujla Shean J. , Reardon Brian , Moore Margaret , Shea Pamela , Schreckhise Randall , Bukowski Thomas R. , Presnell Scott , Guerra-Lewis Patricia , Parrish-Novak Julia , Ellsworth Jeff L. , Jaspers Stephen , Lewis Katherine E. , Appleby Mark , Kolls Jay K. , Rixon Mark , West James W. , Gao Zeren , Levin Steven D.
Published: April 2014

The proinflammatory cytokines IL-17A and IL-17F have a high degree of sequence similarity and share many biological properties. Both have been implicated as factors contributing to the progression of inflammatory and autoimmune diseases. Moreover, reagents that neutralize IL-17A significantly ameliorate disease severity in several mouse models of human disease. IL-17A mediates its effects through interaction with its cognate receptor, the IL-17 receptor (IL-17RA). We report here that the IL-17RA-related molecule, IL-17RC is the receptor for IL-17F. Notably, both IL-17A and IL-17F bind to IL-17RC with high affinity, leading us to suggest that a soluble form of this molecule may serve as an effective therapeutic antagonist of IL-17A and IL-17F. We generated a soluble form of IL-17RC and demonstrate that it effectively blocks binding of both IL-17A and IL-17F, and that it inhibits signaling in response to these cytokines. Collectively, our work indicates that IL-17RC functions as a receptor for both IL-17A and IL-17F and that a soluble version of this protein should be an effective antagonist of IL-17A and IL-17F mediated inflammatory diseases.
The Chemokine CCL6 Promotes Innate Immunity via Immune Cell Activation and Recruitment
Authors: Coelho Ana L. , Schaller Matthew A. , Benjamim Claudia F. , Orlofsky Amos Z. , Hogaboam Cory M. , Kunkel Steven L.
Published: April 2014

Septic syndrome is a consequence of innate immune failure. Recent studies showed that the CC chemokine CCL6 enhanced antimicrobial immunity during experimental sepsis through an unknown mechanism. The present study demonstrates that transgenic CCL6 expression abolishes mortality in a septic peritonitis model via the modulation of resident peritoneal cell activation and, more importantly, through the recruitment of IFN-producing NK cells and killer dendritic cells into the peritoneum. Thus, CCL6 attenuates the immune failure during sepsis, in part, through a protective type 1-cytokine mediated mechanism.
Proteinase-Activated Receptor-2 Exerts Protective and Pathogenic Cell Type-Specific Effects in Alzheimer’s Disease
Authors: Afkhami-Goli Amir , Noorbakhsh Farshid , Keller Avril J. , Vergnolle Nathalie , Westaway David , Jhamandas Jack H. , Andrade-Gordon Patricia , Hollenberg Morley D. , Arab Hosseinali , Dyck Richard H. , Power Christopher
Published: April 2014

The proteinase-activated receptors (PARs) are a novel family of G protein-coupled receptors, and their effects in neurodegenerative diseases remain uncertain. Alzheimer’s disease (AD) is a neurodegenerative disorder defined by misfolded protein accumulation with concurrent neuroinflammation and neuronal death. We report suppression of proteinase-activated receptor-2 (PAR2) expression in neurons of brains from AD patients, whereas PAR2 expression was increased in proximate glial cells, together with up-regulation of proinflammatory cytokines and chemokines and reduced IL-4 expression (p < 0.05). Glial PAR2 activation increased expression of formyl peptide receptor-2 (p < 0.01), a cognate receptor for a fibrillar 42-aa form of β-amyloid (Aβ1–42), enhanced microglia-mediated proinflammatory responses, and suppressed astrocytic IL-4 expression, resulting in neuronal death (p < 0.05). Conversely, neuronal PAR2 activation protected human neurons against the toxic effects of Aβ1–42 (p < 0.05), a key component of AD neuropathogenesis. Amyloid precursor protein-transgenic mice, displayed glial fibrillary acidic protein and IL-4 induction (p < 0.05) in the absence of proinflammatory gene up-regulation and neuronal injury, whereas PAR2 was up-regulated at this early stage of disease progression. PAR2-deficient mice, after hippocampal Aβ1–42 implantation, exhibited enhanced IL-4 induction and less neuroinflammation (p < 0.05), together with improved neurobehavioral outcomes (p < 0.05). Thus, PAR2 exerted protective properties in neurons, but its activation in glia was pathogenic with secretion of neurotoxic factors and suppression of astrocytic anti-inflammatory mechanisms contributing to Aβ1–42-mediated neurodegeneration.
Hepatocyte Growth Factor Significantly Suppresses Collagen-Induced Arthritis in Mice
Authors: Okunishi Katsuhide , Dohi Makoto , Fujio Keishi , Nakagome Kazuyuki , Tabata Yasuhiko , Okasora Takahiro , Seki Makoto , Shibuya Mihoko , Imamura Mitsuru , Harada Hiroaki , Tanaka Ryoichi , Yamamoto Kazuhiko
Published: April 2014

Hepatocyte growth factor (HGF) plays an important role in angiogenesis, cell proliferation, antifibrosis, and antiapoptosis. Moreover, recent studies have highlighted the immunosuppressive effect of HGF in animal models of allogenic heart transplantation and autoimmune myocarditis and in studies in vitro as well. We also reported that HGF significantly suppresses dendritic cell function, thus down-regulating Ag-induced Th1-type and Th2-type immune responses in allergic airway inflammation. However, the immunosuppressive effect of HGF in many other situations has not been fully clarified. In the present study, using a mouse model of collagen-induced arthritis (CIA) and experiments in vitro, we examined the effect of HGF on autoimmune arthritis and then elucidated the mechanisms of action of HGF. To achieve sufficient delivery of HGF, we used biodegradable gelatin hydrogels as a carrier. HGF suppressed Ag-induced T cell priming by regulating the functions of dendritic cells in the Ag-sensitization phase with down-regulation of IL-10. In contrast, under continuous Ag stimulation HGF induced IL-10-producing immunocytes both in vivo and in vitro. Moreover, HGF potently inhibited the development of CIA with enhancing the Th2-type immune response. We also confirmed that HGF significantly suppressed the production of IL-17 by immunocytes. These results indicate that HGF suppresses the development of CIA through different ways at different phases. They also suggest that HGF could be an attractive tool for treating patients with rheumatoid arthritis.
The Heat Shock Protein HSP70 Promotes Mouse NK Cell Activity against Tumors That Express Inducible NKG2D Ligands
Authors: Elsner Leslie , Muppala Vijayakumar , Gehrmann Mathias , Lozano Jingky , Malzahn Dörthe , Bickeböller Heike , Brunner Edgar , Zientkowska Marta , Herrmann Thomas , Walter Lutz , Alves Frauke , Multhoff Gabriele , Dressel Ralf
Published: April 2014

The stress-inducible heat shock protein (HSP) 70 is known to function as an endogenous danger signal that can increase the immunogenicity of tumors and induce CTL responses. We show in this study that HSP70 also activates mouse NK cells that recognize stress-inducible NKG2D ligands on tumor cells. Tumor size and the rate of metastases derived from HSP70-overexpressing human melanoma cells were found to be reduced in T and B cell-deficient SCID mice, but not in SCID/beige mice that lack additionally functional NK cells. In the SCID mice with HSP70-overexpressing tumors, NK cells were activated so that they killed ex vivo tumor cells that expressed NKG2D ligands. In the tumors, the MHC class I chain-related (MIC) A and B molecules were found to be expressed. Interestingly, a counter selection was observed against the expression of MICA/B in HSP70-overexpressing tumors compared with control tumors in SCID, but not in SCID/beige mice, suggesting a functional relevance of MICA/B expression. The melanoma cells were found to release exosomes. HSP70-positive exosomes from the HSP70-overexpressing cells, in contrast to HSP70-negative exosomes from the control cells, were able to activate mouse NK cells in vitro to kill YAC-1 cells, which express NKG2D ligands constitutively, or the human melanoma cells, in which MICA/B expression was induced. Thus, HSP70 and inducible NKG2D ligands synergistically promote the activation of mouse NK cells resulting in a reduced tumor growth and suppression of metastatic disease.
Intravenous Immunoglobulin Therapy Affects T Regulatory Cells by Increasing Their Suppressive Function
Authors: Kessel Aharon , Ammuri Hana , Peri Regina , Pavlotzky Elsa R. , Blank Miri , Shoenfeld Yehuda , Toubi Elias
Published: April 2014

Intravenous Ig therapy (IVIg) is reported to be a useful regimen in treating autoimmune diseases. In this study, we asked whether IVIg (in vitro) could increase the expression of TGF-β, IL-10, and the transcription factor FoxP3 in T regulatory (Treg) cells, and the idea that IVIg could enhance suppressive properties of these cells. CD4+ T cells from 12 healthy individuals were cultured in the presence or absence of IVIg vs human control IgG during 16, 24, and 36 h. Using FACS analysis and gating on CD4+CD25high Treg cells, we assessed the expression of intracellular TGF-β, IL-10, and FoxP3. In addition, the production of TNF-α by stimulated CD4+ T cells alone or in culture with CD25+ by itself or together with IVIg was also assessed. The presence of IVIg with Treg cells in culture significantly increased the intracellular expression of TGF-β (17.7 ± 8.5% vs 29.8 ± 13%; p = 0.02), IL-10 (20.7 ± 4.7% vs 34.2 ± 5.2%; p = 0.008) and FoxP3 (20.8 ± 5.2% vs 33.7 ± 5.9%; p = 0.0006) when compared with cells cultured alone or with control human IgG. The suppressive effect of CD4+CD25+ T cells presented as the decrease of TNF-α production by stimulated CD4+CD25− (effector T cells) was further increased by adding IVIg to cell culture. We hereby demonstrate an additional mechanism by which IVIg could maintain self-tolerance and decrease immune-mediated inflammation.
Exacerbation of Collagen-Induced Arthritis by Oligoclonal, IL-17-Producing γδ T Cells
Authors: Roark Christina L. , French Jena D. , Taylor Molly A. , Bendele Alison M. , Born Willi K. , O’Brien Rebecca L.
Published: April 2014

Murine γδ T cell subsets, defined by their Vγ chain usage, have been shown in various disease models to have distinct functional roles. In this study, we examined the responses of the two main peripheral γδ T cell subsets, Vγ1+ and Vγ4+ cells, during collagen-induced arthritis (CIA), a mouse model that shares many hallmarks with human rheumatoid arthritis. We found that whereas both subsets increased in number, only the Vγ4+ cells became activated. Surprisingly, these Vγ4+ cells appeared to be Ag selected, based on preferential Vγ4/Vδ4 pairing and very limited TCR junctions. Furthermore, in both the draining lymph node and the joints, the vast majority of the Vγ4/Vδ4+ cells produced IL-17, a cytokine that appears to be key in the development of CIA. In fact, the number of IL-17-producing Vγ4+ γδ T cells in the draining lymph nodes was found to be equivalent to the number of CD4+αβ+ Th-17 cells. When mice were depleted of Vγ4+ cells, clinical disease scores were significantly reduced and the incidence of disease was lowered. A decrease in total IgG and IgG2a anti-collagen Abs was also seen. These results suggest that Vγ4/Vδ4+ γδ T cells exacerbate CIA through their production of IL-17.
IL-10- and TGF-β-Mediated Susceptibility in Kala-azar and Post-kala-azar Dermal Leishmaniasis: The Significance of Amphotericin B in the Control of Leishmania donovani Infection in India
Authors: Saha Samiran , Mondal Smriti , Ravindran Rajesh , Bhowmick Swati , Modak Dolanchampa , Mallick Sudeshna , Rahman Mehboobar , Kar Sourjya , Goswami Ramaprasad , Guha Subhasis Kamal , Pramanik Netai , Saha Bibhuti , Ali Nahid
Published: April 2014

Visceral leishmaniasis (VL) or kala-azar is known to be associated with a mixed Th1-Th2 response, and effective host defense requires the induction of IFN-γ and IL-12. We address the role of the differential decline of IL-10 and TGF-β in response to sodium antimony gluconate (SAG) and amphotericin B (AmB), the therapeutic success of SAG and AmB in Indian VL, and the significance of IL-10 and TGF-β in the development and progression of post-kazla-azar dermal leishmaniasis (PKDL). In the active disease, PBMC from VL patients showed suppressed Ag-specific lymphoproliferation, IFN-γ and IL-12 production, and elevation of IL-10 and TGF-β. Cure corresponded with an elevation in IFN-γ and IL-12 production and down-regulation of IL-10 and TGF-β. Both CD4+ and CD8+ T cells were involved in IFN-γ and IL-10 production. Interestingly, the retention and maintenance of residual IL-10 and TGF-β in some SAG-treated individuals and the elevation of IL-10 and TGF-β in PKDL, a sequel to kala-azar, probably reflects the role of these cytokines in reactivation of the disease in the form of PKDL. Contrastingly, AmB treatment of VL resulted in negligible TGF-β levels and absolute elimination of IL-10, reflecting the better therapeutic activity of AmB and its probable role in the recent decline in PKDL occurrences in India. Moreover, elucidation of immune responses in Indian PKDL patients revealed a spectral pattern of disease progression where disease severity could be correlated inversely with lymphoproliferation and directly with TGF-β, IL-10, and Ab production. In addition, the enhancement of CD4+CD25+ T cells in active VL, their decline at cure, and reactivation in PKDL suggest their probable immunosuppressive role in these disease forms.
Perspectives on Mucosal Vaccines: Is Mucosal Tolerance a Barrier?
Authors: Mestecky Jiri , Russell Michael W. , Elson Charles O.
Published: April 2014

Mucosal administration of Ags induces specific Abs in external secretions and systemic unresponsiveness termed oral or mucosal tolerance. The dominant response depends on the species studied, the nature, dose, frequency, route of Ag application, and the use of adjuvants. The temporal sequence of Ag exposure determines the quality of the ensuing immune response; although initial mucosal Ag exposure results in systemic T cell hyporesponsiveness, pre-existing systemic responses are refractory to the tolerizing effects of mucosal Ag encounter. Mucosal and systemic humoral responses may be induced concomitantly with diminished systemic T cell responses, thereby permitting Ab-mediated containment of mucosal Ags without stimulation of the systemic immune compartment. B cell Ig isotype switching and differentiation toward IgA production share common regulatory mechanisms with the suppression of T cells. Optimization of mucosal vaccination strategies has the potential for enhancing protective immune responses and suppressing systemic responses to autoantigens desirable for the treatment of autoimmune diseases.
Cutting Edge: TCR Revision Affects Predominantly Foxp3− Cells and Skews Them toward the Th17 Lineage
Authors: Zehn Dietmar , Bevan Michael J. , Fink Pamela J.
Published: April 2014

CD4+ T cells respond to peripheral endogenous superantigen stimulation by undergoing deletion or TCR revision. The latter involves RAG re-expression, TCR gene rearrangement, and expression of a novel TCR. TCR-revised T cells are functional and express a diverse TCR repertoire. Because TCR revision harbors the potential to create self-reactivity, it is important to explore whether T cells known to be self-reactive (regulatory T cells) or those involved in autoimmunity (Th17 cells) arise from TCR revision. Interestingly, we observed that Foxp3+ cells are excluded from revising their TCR and that only a small fraction of postrevision cells expresses Foxp3. In contrast, Th17 cells are 20 times more frequent among revised than among C57BL/6 CD4+ T cells, indicating that postrevision cells are biased toward the Th17 lineage. The link between Th17 differentiation and TCR revision might be highly relevant to the role of Th17 cells in promoting autoimmunity.
Targeting the Gut Vascular Endothelium Induces Gut Effector CD8 T Cell Responses Via Cross-Presentation by Dendritic Cells
Authors: Bourges Dorothee , Zhan Yifan , Brady Jamie L. , Braley Hal , Caminschi Irina , Prato Sandro , Villadangos José A. , Lew Andrew M.
Published: April 2014

Systemic delivery of Ag usually induces poor mucosal immunity. To improve the CD8 T cell response at mucosal sites, we targeted the Ag to MAdCAM-1, a mucosal addressin cell adhesion molecule expressed mainly by high endothelial venules (HEV) in mesenteric lymph nodes (MLN) and Peyer’s patches of gut-associated lymphoid tissue. When chemical conjugates of anti-MAdCAM-1 Ab and model Ag OVA were injected i.v., a greatly enhanced proliferative response of Ag-specific OT-I CD8 T cells was detected in MLN. This was preceded by prolonged accumulation, up to 2 wk, of the anti-MAdCAM OVA conjugate on HEV of Peyer’s patches and MLN. In contrast, nontargeted OVA conjugate was very inefficient in inducing OT-I CD8 T cell proliferation in MLN and required at least 20-fold more Ag to induce a comparable response. In addition, MAdCAM targeting elicits an endogenous OVA-specific CD8 T cell response, evident by IFN-γ production and target killing. Induced response offers protection against an OVA-expressing B cell lymphoma. We propose that the augmentation of gut CD8 T cell responses by MAdCAM targeting is due to both accumulation of Ag in the HEV and conversion of a soluble Ag to a cell-associated one, allowing cross-presentation by DCs.
A Distinct Role of Neutrophil Lactoferrin in RelA/p65 Phosphorylation on Ser536 by Recruiting TNF Receptor-Associated Factors to IκB Kinase Signaling Complex
Authors: Oh Sang-Muk , Lee Shin-Hee , Lee Bum-Jin , Pyo Chul-Woong , Yoo Na-Kyung , Lee Soo Young , Kim Jiyoung , Choi Sang-Yun
Published: April 2014

The activation of NF-κB by neutrophil lactoferrin (Lf) is regulated via the IκB kinase (IKK) signaling cascade, resulting in the sequential phosphorylation and degradation of IκB. In this study, we observed that Lf protein augmented p65 phosphorylation at the Ser536, but not the Ser276 residue, and stimulated the translocation of p65 into the nucleus. Lf was also shown to enhance the association between p65 and CREB-binding protein/p300 in vivo. To elucidate the mechanism by which Lf triggers these signaling pathways, we attempted to delineate the roles of the upstream components of the IKK complex, using their dominant-negative mutants and IKKα−/− and IKKβ−/− mouse embryonic cells. We demonstrated that both IKKα and IKKβ as well as NF-κB-inducing kinase are indispensable for Lf-induced p65 phosphorylation. However, MAPK kinase kinase 1 is not essentially required for this activation. We also observed that Lf-induced p65 phosphorylation was either partially or completely abrogated as the result of treatment with the mutant forms of TNFR-associated factor (TRAF) 2, TRAF5, or TRAF6. Moreover, we demonstrated that Lf directly interacted with TRAF5. Expression of the dominant-negative mutant of TRAF5 or its small interfering RNA almost completely abrogated the Lf-induced p65 phosphorylation. These results suggest that signaling pathways, including TRAFs/NF-κB-inducing kinase/IKKs, may be involved in the regulation of Lf-induced p65 activation, thereby resulting in the activation of members of the NF-κB family.
Lymphotoxin Pathway and Aire Influences on Thymic Medullary Epithelial Cells Are Unconnected
Authors: Venanzi Emily S. , Gray Daniel H. D. , Benoist Christophe , Mathis Diane
Published: April 2014

The lymphotoxin pathway is critical for the development and maintenance of peripheral lymphoid organs. Mice with deficiencies in members of this pathway lack lymph nodes and Peyer’s patches and have abnormal spleen architecture. These animals also develop autoantibodies to and lymphocytic infiltrates of multiple organs, provoking speculation that the lymphotoxin pathway may play a role in central tolerance induction. Indeed, a series of reports has claimed that lymphotoxin signals control the expression of Aire, a transcriptional regulator that is expressed in medullary epithelial cells of the thymus, mediates ectopic transcription of genes encoding a variety of peripheral tissue Ags, and promotes clonal deletion of self-reactive thymocytes. However, one report argued that lymphotoxin signals regulate the composition and organization of the thymus, particularly of the medullary epithelial compartment. Herein, we resolve this controversy in favor of the latter view. The expression and function of Aire were unaffected in medullary epithelial cells of mice lacking either lymphotoxin β receptor or the lymphotoxin α-chain, and there was minimal overlap between the sets of genes controlled by Aire and lymphotoxin. Instead, both knockout lines showed abnormal medullary epithelial cell organization, and the line lacking the β receptor had significantly fewer medullary epithelial cells. In short, the lymphotoxin pathway drives the developmental rather than selectional properties of thymic stromal cells.
Sialyl-Lewisx on P-Selectin Glycoprotein Ligand-1 Is Regulated during Differentiation and Maturation of Dendritic Cells: A Mechanism Involving the Glycosyltransferases C2GnT1 and ST3Gal I
Authors: Julien Sylvain , Grimshaw Matthew J. , Sutton-Smith Mark , Coleman Julia , Morris Howard R. , Dell Anne , Taylor-Papadimitriou Joyce , Burchell Joy M.
Published: April 2014

To fulfil their function as APCs, dendritic cells (DC) and their precursors need to travel from blood to the peripheral tissues and, upon activation, migrate from tissues to draining lymph nodes. Because O-glycans play a role in T cell trafficking, we investigated the O-glycosylation profile of human monocyte-derived DC. Sialyl-Lewisx (sLex), a glycan involved in extravasation via selectin binding, was found to be expressed exclusively on P-selectin glycoprotein ligand-1 in monocytes and immature DC. However, sLex was lost from mature DC even though these cells retained expression of P-selectin glycoprotein ligand-1. Maturation of DC led to a rapid change in the expression of glycosyltransferases involved in O-linked glycosylation. A down-regulation of C2GnT1 mRNA and enzymatic activity was observed with a concurrent up-regulation of ST3Gal I and ST6GalNAc II mRNA resulting in a loss of the core 2 structures required for sLex expression as a P-selectin ligand. Interestingly, the early regulation of these glycosyltransferases was mediated by PGE2, which is known to be required for human DC migration. The pattern of O-glycosylation seen in mature cells was very similar to that expressed by naive T cells, which home to lymph nodes. Our data show that the regulation of O-glycosylation controls sLex expression, and also suggest that O-glycans may have a function in DC migration.
Molecular Architecture of the TAP-Associated MHC Class I Peptide-Loading Complex
Authors: Rufer Elke , Leonhardt Ralf M. , Knittler Michael R.
Published: April 2014

Tapasin organizes the peptide-loading complex (PLC) by recruiting peptide-receptive MHC class I (MHC-I) and accessory chaperones to the N-terminal regions of the TAP subunits TAP1 and TAP2. Despite numerous studies have shown that the formation of the PLC is essential to facilitate proper MHC-I loading, the molecular architecture of this complex is still highly controversial. We studied the stoichiometry of the PLC by blue native-PAGE in combination with Ab-shift assays and found that TAP/tapasin complexes exist at steady state as a mixture of two distinct oligomers of 350 and 450 kDa. Only the higher m.w. complex contains MHC-I and disulfide-linked tapasin/ER60 conjugates. Moreover, we show for the first time to our knowledge that the fully assembled PLC comprises two tapasin, two ER60, but only one complex of MHC-I and calreticulin. Based hereon we postulate that the TAP subunits alternate in the recruitment and loading of a single MHC-I.
Increasing the Survival of Dendritic Cells In Vivo Does Not Replace the Requirement for CD4+ T Cell Help during Primary CD8+ T Cell Responses
Authors: Matthews Kate E. , Qin Jim S. , Yang Jianping , Hermans Ian F. , Palmowski Michael J. , Cerundolo Vincenzo , Ronchese Franca
Published: April 2014

The survival of dendritic cells (DC) in vivo determines the duration of Ag presentation and is critical in determining the strength and magnitude of the resulting T cell response. We used a mouse model to show that Ag-loaded C57BL/6 DC (MHC class II+/+ (MHC II+/+)) that reach the lymph node survived longer than Ag-loaded MHC II−/− DC, with the numbers of C57BL/6 DC being ∼2.5-fold the number of the MHC II−/− DC by day 4 and ∼5-fold by day 7. The differential survival of DC in vivo was not affected by low doses of LPS, but in vitro pretreatment with CD40L or with high doses of LPS increased the numbers of MHC II−/− DC to levels approaching those of C57BL/6 DC. Regardless of their numbers and relative survival in lymph nodes, MHC II−/− DC were profoundly defective in their ability to induce CTL responses against the gp33 peptide epitope, and were unable to induce expansion and optimal cytotoxic activity of CD8+ T cells specific for the male Ag UTY. We conclude that CD4+ T cell help for CD8+ responses involves mechanisms other than the increased survival of Ag-presenting DC in the lymph node.
Allergic Airways Disease Develops after an Increase in Allergen Capture and Processing in the Airway Mucosa
Authors: von Garnier Christophe , Wikstrom Matthew E. , Zosky Graeme , Turner Debra J. , Sly Peter D. , Smith Miranda , Thomas Jennifer A. , Judd Samantha R. , Strickland Deborah H. , Holt Patrick G. , Stumbles Philip A.
Published: April 2014

Airway mucosal dendritic cells (AMDC) and other airway APCs continuously sample inhaled Ags and regulate the nature of any resulting T cell-mediated immune response. Although immunity develops to harmful pathogens, tolerance arises to nonpathogenic Ags in healthy individuals. This homeostasis is thought to be disrupted in allergic respiratory disorders such as allergic asthma, such that a potentially damaging Th2-biased, CD4+ T cell-mediated inflammatory response develops against intrinsically nonpathogenic allergens. Using a mouse model of experimental allergic airways disease (EAAD), we have investigated the functional changes occurring in AMDC and other airway APC populations during disease onset. Onset of EAAD was characterized by early and transient activation of airway CD4+ T cells coinciding with up-regulation of CD40 expression exclusively on CD11b− AMDC. Concurrent enhanced allergen uptake and processing occurred within all airway APC populations, including B cells, macrophages, and both CD11b+ and CD11b− AMDC subsets. Immune serum transfer into naive animals recapitulated the enhanced allergen uptake observed in airway APC populations and mediated activation of naive allergen-specific, airway CD4+ T cells following inhaled allergen challenge. These data suggest that the onset of EAAD is initiated by enhanced allergen capture and processing by a number of airway APC populations and that allergen-specific Igs play a role in the conversion of normally quiescent AMDC subsets into those capable of inducing airway CD4+ T cell activation.