bluuebunny/crossref_metadata_2025 huggingface dataset:
Mechanisms Underlying Resistance of Pancreatic Islets from ALR/Lt Mice to Cytokine-Induced Destruction
Authors: Mathews Clayton E. , Suarez-Pinzon Wilma L. , Baust Jeffrey J. , Strynadka Ken , Leiter Edward H. , Rabinovitch Alex
Published: April 2014

Nuclear and mitochondrial genomes combine in ALR/Lt mice to produce systemically elevated defenses against free radical damage, rendering these mice resistant to immune-mediated pancreatic islet destruction. We analyzed the mechanism whereby isolated islets from ALR mice resisted proinflammatory stress mediated by combined cytokines (IL-1β, TNF-α, and IFN-γ) in vitro. Such damage entails both superoxide and NO radical generation, as well as peroxynitrite, resulting from their combination. In contrast to islets from other mouse strains, ALR islets expressed constitutively higher glutathione reductase, glutathione peroxidase, and higher ratios of reduced to oxidized glutathione. Following incubation with combined cytokines, islets from control strains produced significantly higher levels of hydrogen peroxide and NO than islets from ALR mice. Nitrotyrosine was generated in NOD and C3H/HeJ islets but not by ALR islets. Western blot analysis showed that combined cytokines up-regulated the NF-κB inducible NO synthase in NOD-Rag and C3H/HeJ islets but not in ALR islets. This inability of cytokine-treated ALR islets to up-regulate inducible NO synthase and produce NO correlated both with reduced kinetics of IκB degradation and with markedly suppressed NF-κB p65 nuclear translocation. Hence, ALR/Lt islets resist cytokine-induced diabetogenic stress through enhanced dissipation and/or suppressed formation of reactive oxygen and nitrogen species, impaired IκB degradation, and blunted NF-κB activation. Nitrotyrosylation of β cell proteins may generate neoantigens; therefore, resistance of ALR islets to nitrotyrosine formation may, in part, explain why ALR mice are resistant to type 1 diabetes when reconstituted with a NOD immune system.
A Non-Glycosaminoglycan-Binding Variant of CC Chemokine Ligand 7 (Monocyte Chemoattractant Protein-3) Antagonizes Chemokine-Mediated Inflammation
Authors: Ali Simi , Robertson Helen , Wain Julie H. , Isaacs John D. , Malik Ghada , Kirby John A.
Published: April 2014

A non-glycosaminoglycan (GAG)-binding variant of the pleiotropic chemokine CCL7 was generated by mutating to alanine the basic (B) amino acids within an identified 44BXBXXB49 GAG-binding motif. Unlike wild-type (wt) CCL7, the mutant sequence had no affinity for heparin. However, the mutant retained a normal affinity for CCR1, CCR2b, and CCR3, and produced a normal calcium flux in mononuclear leukocytes. Both the wt and mutant proteins elicited an equal leukocyte chemotactic response within a solute diffusion gradient but, unlike the wt protein, the mutant failed to stimulate cell migration across a model endothelium. The number of leukocytes recruited to murine air pouches by the mutant sequence was lower than that recruited by wt CCL7. Furthermore, the presence of a mixture of a mutant and wt CCL7 within the air pouch elicited no significant cell accumulation. Cell recruitment also failed using a receptor-sharing mixture of mutant CCL7 and wt CCL5 or a nonreceptor sharing mixture of mutant CCL7 and wt CXCL12. The potential of the mutant sequence to modulate inflammation was confirmed by demonstration of its ability to inhibit the chemotactic response generated in vitro by synovial fluid from patients with active rheumatoid arthritis. A further series of experiments suggested that the non-GAG-binding mutant protein could potentially induce receptor desensitization before, and at a site remote from, any physiological recognition of GAG-bound chemokines. These data demonstrate that GAG binding is required for chemokine-driven inflammation in vivo and also suggest that a non-GAG-binding chemokine receptor agonist can inhibit the normal vectorial leukocyte migration mediated by chemokines.
Hom s 4, an IgE-Reactive Autoantigen Belonging to a New Subfamily of Calcium-Binding Proteins, Can Induce Th Cell Type 1-Mediated Autoreactivity
Authors: Aichberger Karl J. , Mittermann Irene , Reininger Renate , Seiberler Susanne , Swoboda Ines , Spitzauer Susanne , Kopp Tamara , Stingl Georg , Sperr Wolfgang R. , Valent Peter , Repa Andreas , Bohle Barbara , Kraft Dietrich , Valenta Rudolf
Published: April 2014

Skin inflammation in atopic dermatitis starts with Th2 and IgE-mediated responses against exogenous allergens and, for unknown reasons, resembles features of a Th1-driven reaction in the chronic stages. We report the characterization of a human protein, Hom s 4, recognized by IgE autoantibodies from atopic dermatitis patients. The complete Hom s 4 cDNA codes for a 54-kDa basic protein containing two typical calcium-binding domains separated by an unusually long α-helical domain. Therefore, Hom s 4 and homologous proteins found by sequence comparison in mice, fruit flies, and nematodes constitute a novel subfamily of calcium-binding proteins. Using Hom s 4-specific Abs, it is demonstrated that the protein is strongly expressed within epidermal keratinocytes and dermal endothelial cells. Purified Hom s 4 showed IgE cross-reactivity with exogenous calcium-binding allergens from plants and fish but, in contrast to the exogenous allergens, induced only weak histamine release from patient basophils. However, the analysis of Hom s 4-specific cytokine and humoral immune responses indicated that Hom s 4 strongly induces Th1 responses which are accompanied by the release of IFN-γ, a cytokine implicated in epithelial cell damage. Hom s 4-induced IFN-γ production was found in normal individuals, in patients with chronic inflammatory skin diseases and in Th2-prone atopic persons, suggesting that Hom s 4 represents a protein with an intrinsic property to induce Th1-mediated autoreactivity. It may thus contribute to chronic skin inflammation in atopic as well as in nonatopic persons.
Host-Residual Invariant NK T Cells Attenuate Graft-versus-Host Immunity
Authors: Haraguchi Kyoko , Takahashi Tsuyoshi , Matsumoto Akihiko , Asai Takashi , Kanda Yoshinobu , Kurokawa Mineo , Ogawa Seishi , Oda Hideaki , Taniguchi Masaru , Hirai Hisamaru , Chiba Shigeru
Published: April 2014

Invariant NK T (iNKT) cells have an invariant TCR-α chain and are activated in a CD1d-restricted manner. They are thought to regulate immune responses and play important roles in autoimmunity, allergy, infection, and tumor immunity. They also appear to influence immunity after hemopoietic stem cell transplantation. In this study, we examined the role of iNKT cells in graft-vs-host disease (GVHD) and graft rejection in a mouse model of MHC-mismatched bone marrow transplantation, using materials including α-galactosylceramide, NKT cells expanded in vitro, and Jα18 knockout mice that lack iNKT cells. We found that host-residual iNKT cells constitute effector cells which play a crucial role in reducing the severity of GVHD, and that this reduction is associated with a delayed increase in serum Th2 cytokine levels. Interestingly, we also found that host-residual iNKT cause a delay in engraftment and, under certain conditions, graft rejection. These results indicate that host-residual iNKT cells attenuate graft-vs-host immunity rather than host-vs-graft immunity.
DNA Vaccines: Progress and Challenges
Authors: Donnelly John J. , Wahren Britta , Liu Margaret A.
Published: April 2014

In the years following the publication of the initial in vivo demonstration of the ability of plasmid DNA to generate protective immune responses, DNA vaccines have entered into a variety of human clinical trials for vaccines against various infectious diseases and for therapies against cancer, and are in development for therapies against autoimmune diseases and allergy. They also have become a widely used laboratory tool for a variety of applications ranging from proteomics to understanding Ag presentation and cross-priming. Despite their rapid and widespread development and the commonplace usage of the term “DNA vaccines,” however, the disappointing potency of the DNA vaccines in humans underscores the challenges encountered in the efforts to translate efficacy in preclinical models into clinical realities. This review will provide a brief background of DNA vaccines including the insights gained about the varied immunological mechanisms that play a role in their ability to generate immune responses.
E2F1 and E2F2 Are Differentially Required for Homeostasis-Driven and Antigen-Induced T Cell Proliferation In Vivo
Authors: DeRyckere Deborah , DeGregori James
Published: April 2014

Homeostasis-driven T cell proliferation occurs in response to a lymphopenic environment and is mediated by TCR and IL-7 signaling. In this report, we demonstrate a defect in the proliferation of murine naive and memory T cells lacking both E2F1 and E2F2 in response to lymphopenic conditions, suggesting that E2F1 and E2F2 function redundantly downstream of TCR and/or IL-7 signaling during homeostasis-driven proliferation. In contrast, T cell proliferation in response to antigenic stimulation is either unaffected (in vivo) or potentiated (ex vivo) by loss of E2F1 and E2F2, indicating divergent requirements for these E2F factors in T cell proliferation mediated by distinct stimuli. E2F1/E2F2 double knockout (DKO) T cells enter S phase in response to homeostatic signaling, but fail to divide, suggesting that S phase progression is either incomplete or defective. In addition, E2F1/E2F2 DKO mice do not recover normal T cell numbers following exposure to a sublethal dose of radiation, indicating that this defect in homeostasis-driven proliferation is physiologically relevant. Consistent with their failure in cell cycle progression, the differentiation of DKO T cells into memory T cells in response to homeostatic signals is significantly reduced. These observations support the idea that proliferation is required for memory T cell formation and also have implications for the development of clinical strategies to minimize the occurrence of lymphopenia-induced autoimmunity.
Induction of a Regulatory Phenotype in Human CD4+ T Cells by Streptococcal M Protein
Authors: Price Jeffrey D. , Schaumburg Jessica , Sandin Charlotta , Atkinson John P. , Lindahl Gunnar , Kemper Claudia
Published: April 2014

Regulatory T cells (Tregs) participate in the control of the immune response. In the human system, an IL-10-secreting, T regulatory type 1 cell (Tr1)-like subset of Tregs can be induced by concurrent cross-linking of the TCR and CD46 on naive CD4+ T cells. Because many viral and bacterial pathogens, including the major human pathogen Streptococcus pyogenes, bind to CD46, we asked whether this bacterium can directly induce Tr1-like cells through the streptococcal ligand for CD46, the M protein. The M5 and M22 proteins were found to induce T cells to develop into the IL-10-producing Tr1-like phenotype. Moreover, whole M5-expressing bacteria, but not isogenic M-negative bacteria, led to proliferation and IL-10 secretion by T cells. The interaction between the M5 protein and T cells was dependent on CD46 and the conserved C repeat region of M5. Supernatants derived from T cells stimulated with M proteins or M protein-expressing bacteria suppressed bystander T cell proliferation through IL-10 secretion. In addition, activation of CD46 through streptococcal M protein induced the expression of granzyme B, providing a second means for these cells to regulate an immune response. These findings suggest that binding to CD46 and exploiting its signaling pathway may represent a strategy employed by a number of important human pathogens to induce directly an immunosuppressive/regulatory phenotype in T cells.
The Type 1 Cysteinyl Leukotriene Receptor Triggers Calcium Influx and Chemotaxis in Mouse αβ- and γδ Effector T Cells
Authors: Prinz Immo , Gregoire Claude , Mollenkopf Hans , Aguado Enrique , Wang Ying , Malissen Marie , Kaufmann Stefan H.E. , Malissen Bernard
Published: April 2014

Linker for activation of T cells (LAT) is essential for T cell activation. Mice with mutations of distinct LAT tyrosine residues (LatY136F and Lat3YF) develop lymphoproliferative disorders involving TCR αβ or γδ T cells that trigger symptoms resembling allergic inflammation. We analyzed whether these T cells share a pattern of gene expression that may account for their pathogenic properties. Both LatY136F αβ and Lat3YF γδ T cells expressed high levels of the type 1 cysteinyl leukotriene receptor (CysLT1). Upon binding to the 5(S)-hydroxy-6(R)-S-cysteinylglycyl-7,9-trans-11,14-cis-eicosatetraenoic acid (LTD4) cysteinyl leukotriene, CysLT1 induced Ca2+ flux and caused chemotaxis in both LatY136F αβ and Lat3YF γδ T cells. Wild-type in vitro-activated T cells, but not resting T cells, also migrated toward LTD4 however with a lower magnitude than T cells freshly isolated from LatY136F and Lat3YF mice. These results suggest that CysLT1 is likely involved in the recruitment of activated αβ and γδ T cells to inflamed tissues.
Systemic NKG2D Down-Regulation Impairs NK and CD8 T Cell Responses In Vivo
Authors: Wiemann Katrin , Mittrücker Hans-Willi , Feger Ute , Welte Stefan A. , Yokoyama Wayne M. , Spies Thomas , Rammensee Hans-Georg , Steinle Alexander
Published: April 2014

The immunoreceptor NKG2D stimulates activation of cytotoxic lymphocytes upon engagement with MHC class I-related NKG2D ligands of which at least some are expressed inducibly upon exposure to carcinogens, cell stress, or viruses. In this study, we investigated consequences of a persistent NKG2D ligand expression in vivo by using transgenic mice expressing MHC class I chain-related protein A (MICA) under control of the H2-Kb promoter. Although MICA functions as a potent activating ligand of mouse NKG2D, H2-Kb-MICA mice appear healthy without aberrations in lymphocyte subsets. However, NKG2D-mediated cytotoxicity of H2-Kb-MICA NK cells is severely impaired in vitro and in vivo. This deficiency concurs with a pronounced down-regulation of surface NKG2D that is also seen on activated CD8 T cells. As a consequence, H2-Kb-MICA mice fail to reject MICA-expressing tumors and to mount normal CD8 T cell responses upon Listeria infection emphasizing the importance of NKG2D in immunity against tumors and intracellular infectious agents.
Expansion and Hyperactivity of CD1d-Restricted NKT Cells during the Progression of Systemic Lupus Erythematosus in (New Zealand Black × New Zealand White)F1 Mice
Authors: Forestier Claire , Molano Alberto , Im Jin Seon , Dutronc Yves , Diamond Betty , Davidson Anne , Illarionov Petr A. , Besra Gurdyal S. , Porcelli Steven A.
Published: April 2014

CD1d-restricted NKT cells expressing invariant TCR α-chain rearrangements (iNKT cells) have been reported to be deficient in humans with a variety of autoimmune syndromes and in certain strains of autoimmune mice. In addition, injection of mice with α-galactosylceramide, a specific glycolipid agonist of iNKT cells, activates these T cells and ameliorates autoimmunity in several different disease models. Thus, deficiency and reduced function in iNKT cells are considered to be risk factors for the development of such diseases. In this study we report that the development of systemic lupus erythematosus in (New Zealand Black (NZB) × New Zealand White (NZW))F1 mice was paradoxically associated with an expansion and activation of iNKT cells. Although young (NZB × NZW)F1 mice had normal levels of iNKT cells, these expanded with age and became phenotypically and functionally hyperactive. Activation of iNKT cells in (NZB × NZW)F1 mice in vivo or in vitro with α-galactosylceramide indicated that the immunoregulatory role of iNKT cells varied over time, revealing a marked increase in their potential to contribute to production of IFN-γ with advancing age and disease progression. This evolution of iNKT cell function during the progression of autoimmunity may have important implications for the mechanism of disease in this model of systemic lupus erythematosus and for the development of therapies using iNKT cell agonists.
Analysis of the Underlying Cellular Mechanisms of Anti-CD154-Induced Graft Tolerance: The Interplay of Clonal Anergy and Immune Regulation
Authors: Quezada Sergio A. , Bennett Kathy , Blazar Bruce R. , Rudensky Alexander Y. , Sakaguchi Shimon , Noelle Randolph J.
Published: April 2014

Although it has been shown that CD4+CD25+ regulatory T cells (Treg) contribute to long-term graft acceptance, their impact on the effector compartment and the mechanism by which they exert suppression in vivo remain unresolved. Using a CD4+ TCR transgenic model for graft tolerance, we have unveiled the independent contributions of anergy and active suppression to the fate of immune and tolerant alloreactive T cells in vivo. First, it is shown that anti-CD154-induced tolerance resulted in the abortive expansion of the alloreactive, effector T cell pool. Second, commensurate with reduced expansion, there was a loss of cytokine production, activation marker expression, and absence of memory T cell markers. All these parameters defined the tolerant alloreactive T cells and correlated with the inability to mediate graft rejection. Third, the tolerant alloreactive T cell phenotype that is induced by CD154 was reversed by the in vivo depletion of Treg. Reversal of the tolerant phenotype was followed by rapid rejection of the allograft. Fourth, in addition to Treg depletion, costimulation of the tolerant alloreactive T cells or activation of the APC compartment also reverted alloreactive T cell tolerance and restored an activated phenotype. Finally, it is shown that the suppression is long-lived, and in the absence of anti-CD154 and donor-specific transfusion, these Treg can chronically suppress effector cell responses, allowing long-lived graft acceptance.
Cyclooxygenase 2 Inhibition Promotes IFN-γ-Dependent Enhancement of Antitumor Responses
Authors: Sharma Sherven , Zhu Li , Yang Seok Chul , Zhang Ling , Lin Jeff , Hillinger Sven , Gardner Brian , Reckamp Karen , Strieter Robert M. , Huang Min , Batra Raj K. , Dubinett Steven M.
Published: April 2014

In previous studies, we demonstrated an immune suppressive network in non-small cell lung cancer that is due to overexpression of tumor cyclooxygenase 2 (COX-2). In this study, we assessed the vaccination response to tumor challenge following either pharmacological or genetic inhibition of COX-2 in a murine lung cancer model. Treatment of naive mice with the COX-2 inhibitor, SC-58236, skewed splenocytes toward a type 1 cytokine response, inducing IFN-γ, IL-12, and IFN-γ-inducible protein 10, whereas the type 2 cytokines IL-4, IL-5, and IL-10 remained unaltered. Fifty percent of mice receiving SC-58236 and an irradiated tumor cell vaccine completely rejected tumors upon challenge. Those mice that did form tumors following challenge demonstrated a reduced tumor growth. In contrast, all mice either vaccinated with irradiated tumor cells alone or receiving SC-58236 alone showed progressive tumor growth. Studies performed in CD4 and CD8 knockout mice revealed a requirement for the CD4 T lymphocyte subset for the complete rejection of tumors. To determine the role of host COX-2 expression on the vaccination responses, studies were performed in COX-2 gene knockout mice. Compared with control littermates, COX-2−/− mice showed a significant tumor growth reduction, whereas heterozygous COX-2−/+ mice had an intermediate tumor growth reduction following vaccination. In vivo depletion of IFN-γ abrogated the COX-2 inhibitor-mediated enhancement of the vaccination effect. These findings provide a strong rationale for additional evaluation of the capacity of COX-2 inhibitors to enhance vaccination responses against cancer.
Dendritic Cell-Mediated Cross-Presentation of Antigens Derived from Colon Carcinoma Cells Exposed to a Highly Cytotoxic Multidrug Regimen with Gemcitabine, Oxaliplatin, 5-Fluorouracil, and Leucovorin, Elicits a Powerful Human Antigen-Specific CTL Response with Antitumor Activity in Vitro
Authors: Correale Pierpaolo , Cusi Maria Grazia , Del Vecchio Maria Teresa , Aquino Angelo , Prete Salvatore , Tsang Kwong Y. , Micheli Lucia , Nencini Cristina , La Placa Marco , Montagnani Francesco , Terrosi Chiara , Caraglia Michele , Formica Vincenzo , Giorgi Giorgio , Bonmassar Enzo , Francini Guido
Published: April 2014

Gemcitabine, oxaliplatin, leucovorin, and 5-fluorouracil (GOLF) is a novel multidrug regimen inducing high levels of necrosis and apoptosis in colon carcinoma cells. This regimen is also able to promote a process of Ag remodeling including up-regulation of immunotherapy targets like carcinoembryonic Ag (CEA), thymidylate synthase (TS). We have conducted a preclinical study aimed to investigate whether these drug-induced modifications would also enhance colon cancer cell immunogenicity. Several CTL lines were thus generated by in vitro stimulating human HLA-A(*)02.01+ PBMCs, from normal donors and colon cancer patients, with autologous dendritic cells cross-primed with cell lysates of colon cancer cells untreated, irradiated, or previously exposed to different drug treatments including the GOLF regimen. Class I HLA-restricted cytolytic activity of these CTL lines was tested against colon cancer cells and CEA and TS gene transfected target cells. These experiments revealed that CTLs sensitized with GOLF-treated cancer cells were much more effective than those sensitized with the untreated colon carcinoma cells or those exposed to the other treatments. CTL lines sensitized against the GOLF-treated colon cancer cells, also expressed a greater percentage of T-lymphocyte precursors able to recognize TS- and CEA-derived peptides. These results suggest that GOLF regimen is a powerful antitumor and immunomodulating regimen that can make the tumor cells a suitable means to induce an Ag-specific CTL response. These results suggest that a rationale combination of GOLF chemotherapy with cytokine-based immunotherapy could generate a chemotherapy-modulated Ag-specific T-lymphocyte response in cancer patients able to destroy the residual disease survived to the cytotoxic drugs.
TLR4 and Toll-IL-1 Receptor Domain-Containing Adapter-Inducing IFN-β, but Not MyD88, RegulateEscherichia coli-Induced Dendritic Cell Maturation and Apoptosis In Vivo
Authors: De Trez Carl , Pajak Bernard , Brait Maryse , Glaichenhaus Nicolas , Urbain Jacques , Moser Muriel , Lauvau Grégoire , Muraille Eric
Published: April 2014

Dendritic cells (DC) are short-lived, professional APCs that play a central role in the generation of adaptive immune responses. Induction of efficient immune responses is dependent on how long DCs survive in the host. Therefore, the regulation of DC apoptosis in vivo during infection remains an important question that requires further investigation. The impact of Escherichia coli bacteremia on DCs has never been analyzed. We show here that i.v. or i.p. administration of live or heat-killed E. coli in mice induces splenic DC migration, maturation, and apoptosis. We further characterize which TLR and Toll-IL-1R (TIR)-containing adaptor molecules regulate these processes in vivo. In this model, DC maturation is impaired in TLR2−/−, TLR4−/− and TIR domain-containing adapter-inducing IFN-β (TRIF)−/− mice. In contrast, DC apoptosis is reduced only in TLR4−/− and TRIF−/− mice. As expected, DC apoptosis induced by the TLR4 ligand LPS is also abolished in these mice. Injection of the TLR9 ligand CpG-oligodeoxynucleotide (synthetic bacterial DNA) induces DC migration and maturation, but only modest DC apoptosis when compared with LPS and E. coli. Together, these results suggest that E. coli bacteremia directly impacts on DC maturation and survival in vivo through a TLR4-TRIF-dependent signaling pathway.
T Cell Leukemia-1 Modulates TCR Signal Strength and IFN-γ Levels through Phosphatidylinositol 3-Kinase and Protein Kinase C Pathway Activation
Authors: Hoyer Katrina K. , Herling Marco , Bagrintseva Ksenia , Dawson David W. , French Samuel W. , Renard Mathilde , Weinger Jason G. , Jones Dan , Teitell Michael A.
Published: April 2014

A signaling role for T cell leukemia-1 (TCL1) during T cell development or in premalignant T cell expansions and mature T cell tumors is unknown. In this study, TCL1 is shown to regulate the growth and survival of peripheral T cells but not precursor thymocytes. Proliferation is increased by TCL1-induced lowering of the TCR threshold for CD4+ and CD8+ T cell activation through both PI3K-Akt and protein kinase C-MAPK-ERK signaling pathways. This effect is submaximal as CD28 costimulation coupled to TCL1 expression additively accelerates dose-dependent T cell growth. In addition to its role in T cell proliferation, TCL1 also increases IFN-γ levels from Th1-differentiated T cells, an effect that may provide a survival advantage during premalignant T cell expansions and in clonal T cell tumors. Combined, these data indicate a role for TCL1 control of growth and effector T cell functions, paralleling features provided by TCR-CD28 costimulation. These results also provide a more detailed mechanism for TCL1-augmented signaling and help explain the delayed occurrence of mature T cell expansions and leukemias despite tumorigenic TCL1 dysregulation that begins in early thymocytes.
Dendritic Cell Aggresome-Like-Induced Structure Formation and Delayed Antigen Presentation Coincide in Influenza Virus-Infected Dendritic Cells
Authors: Herter Sylvia , Osterloh Philipp , Hilf Norbert , Rechtsteiner Gerd , Höhfeld Jörg , Rammensee Hans-Georg , Schild Hansjörg
Published: April 2014

Influenza virus infection induces maturation of murine dendritic cells (DCs), which is most important for the initiation of an immune response. However, in contrast to EL-4 and MC57 cells, DCs present viral CTL epitopes with a delay of up to 10 h. This delay in Ag presentation coincides with the up-regulation of MHC class I molecules as well as costimulatory molecules on the cell surface and the accumulation of newly synthesized ubiquitinated proteins in large cytosolic structures, called DC aggresome-like-induced structures (DALIS). These structures were observed previously after LPS-induced maturation of DCs, and it was speculated that they play a role in the regulation of MHC class I Ag presentation. Our findings provide the first evidence for a connection between DC maturation, MHC class I-restricted Ag presentation, and DALIS formation, which is further supported by the observation that DALIS contain ubiquitinated influenza nucleoprotein.
IL-4 Inhibits Bone-Resorbing Activity of Mature Osteoclasts by Affecting NF-κB and Ca2+ Signaling
Authors: Mangashetti Latha S. , Khapli Shruti M. , Wani Mohan R.
Published: April 2014

IL-4 is an important immune cytokine that regulates bone homeostasis. We investigated the molecular mechanism of IL-4 action on bone-resorbing mature osteoclasts. Using a highly purified population of mature osteoclasts, we show that IL-4 dose-dependently inhibits receptor activator of NF-κB ligand (RANKL)-induced bone resorption by mature osteoclasts. We detected the existence of IL-4R mRNA in mature osteoclasts. IL-4 decreases TRAP expression without affecting multinuclearity of osteoclasts, and inhibits actin ring formation and migration of osteoclasts. Interestingly, IL-4 inhibition of bone resorption occurs through prevention of RANKL-induced nuclear translocation of p65 NF-κB subunit, and intracellular Ca2+ changes. Moreover, IL-4 rapidly decreases RANKL-stimulated ionized Ca2+ levels in the blood, and mature osteoclasts in IL-4 knockout mice are sensitive to RANKL action to induce bone resorption and hypercalcemia. Furthermore, IL-4 inhibits bone resorption and actin ring formation by human mature osteoclasts. Thus, we reveal that IL-4 acts directly on mature osteoclasts and inhibits bone resorption by inhibiting NF-κB and Ca2+ signaling.
Failure of CD25+ T Cells from Lupus-Prone Mice to Suppress Lupus Glomerulonephritis and Sialoadenitis
Authors: Bagavant Harini , Tung Kenneth S. K.
Published: April 2014

The development of organ-specific autoimmune diseases in mice thymectomized on day 3 of life (d3tx mice) can be prevented by transferring CD4+CD25+ T cells from syngeneic, normal adult mice. Using a d3tx model, we asked whether CD4+CD25+ T cell deficiency contributes to glomerulonephritis (GN) in lupus-prone mice. New Zealand Mixed 2328 (NZM2328) mice spontaneously develop autoantibodies to dsDNA and female-dominant, fatal GN. After d3tx, both male and female NZM2328 mice developed 1) accelerated dsDNA autoantibody response, 2) early onset and severe proliferative GN with massive mesangial immune complexes, and 3) autoimmune disease of the thyroid, lacrimal gland, and salivary gland. The d3tx male mice also developed autoimmune prostatitis. The transfer of CD25+ cells from 6-wk-old asymptomatic NZM2328 donors effectively suppressed dsDNA autoantibody and the development of autoimmune diseases, with the exception of proliferative lupus GN and sialoadenitis. This finding indicates that NZM2328 lupus mice have a selective deficiency in T cells that regulates the development of lupus GN and sialoadenitis. After d3tx, the proliferative GN of female mice progressed to fatal GN, but largely regressed in the male, thereby revealing a checkpoint in lupus GN progression that depends on gender.
Restraint of B Cell Activation by Foxj1-Mediated Antagonism of NF-κB and IL-6
Authors: Lin Ling , Brody Steven L. , Peng Stanford L.
Published: April 2014

The forkhead transcription factor Foxj1 inhibits spontaneous autoimmunity, in part by antagonizing NF-κB activation in T cells. We demonstrate here that Foxj1 also inhibits humoral immune responses intrinsically in B cells; Foxj1 deficiency in B cells results in spontaneous and accentuated germinal center formation, associated with the development of pathogenic autoantibodies and accentuated responses to immunizations—all reflecting excessive activity of NF-κB and its target gene IL-6, and correlating with a requirement for Foxj1 to regulate the inhibitory NF-κB component IκBβ. Thus, Foxj1 restrains B cell activation and the maturation of humoral responses, demonstrating a critical role for at least this forkhead transcription factor in the regulation of B lymphocyte homeostasis.
Crystal Structure of Mouse CD1d Bound to the Self Ligand Phosphatidylcholine: A Molecular Basis for NKT Cell Activation
Authors: Giabbai Barbara , Sidobre Stèphane , Crispin M. D. Max , Sanchez-Ruìz Yovan , Bachi Angela , Kronenberg Mitchell , Wilson Ian A. , Degano Massimo
Published: April 2014

NKT cells are immunoregulatory lymphocytes whose activation is triggered by the recognition of lipid Ags in the context of the CD1d molecules by the TCR. In this study we present the crystal structure to 2.8 Å of mouse CD1d bound to phosphatidylcholine. The interactions between the ligand acyl chains and the CD1d molecule define the structural and chemical requirements for the binding of lipid Ags to CD1d. The orientation of the polar headgroup toward the C terminus of the α1 helix provides a rationale for the structural basis for the observed Vα chain bias in invariant NKT cells. The contribution of the ligand to the protein surface suggests a likely mode of recognition of lipid Ags by the NKT cell TCR.
Effects of Bcl-2 Levels on Fas Signaling-Induced Caspase-3 Activation: Molecular Genetic Tests of Computational Model Predictions
Authors: Hua Fei , Cornejo Melanie G. , Cardone Michael H. , Stokes Cynthia L. , Lauffenburger Douglas A.
Published: April 2014

Fas-induced apoptosis is a critical process for normal immune system development and function. Although many molecular components in the Fas signaling pathway have been identified, a systematic understanding of how they work together to determine network dynamics and apoptosis itself has remained elusive. To address this, we generated a computational model for interpreting and predicting effects of pathway component properties. The model integrates current information concerning the signaling network downstream of Fas activation, through both type I and type II pathways, until activation of caspase-3. Unknown parameter values in the model were estimated using experimental data obtained from human Jurkat T cells. To elucidate critical signaling network properties, we examined the effects of altering the level of Bcl-2 on the kinetics of caspase-3 activation, using both overexpression and knockdown in the model and experimentally. Overexpression was used to distinguish among alternative hypotheses for inhibitory binding interactions of Bcl-2 with various components in the mitochondrial pathway. In comparing model simulations with experimental results, we find the best agreement when Bcl-2 blocks the release of cytochrome c by binding to both Bax and truncated Bid instead of Bax, truncated Bid, or Bid alone. Moreover, although Bcl-2 overexpression strongly reduces caspase-3 activation, Bcl-2 knockdown has a negligible effect, demonstrating a general model finding that varying the expression levels of signal molecules frequently has asymmetric effects on the outcome. Finally, we demonstrate that the relative dominance of type I vs type II pathways can be switched by varying particular signaling component levels without changing network structure.
Hierarchical Regulation of CTLA-4 Dimer-Based Lattice Formation and Its Biological Relevance for T Cell Inactivation
Authors: Darlington Peter J. , Kirchhof Mark G. , Criado Gabriel , Sondhi Jitin , Madrenas Joaquín
Published: April 2014

CTLA-4 is an activation-induced, homodimeric inhibitory receptor in T cells. Recent crystallographic reports have suggested that it may form lattice-like arrays on the cell surface upon binding B7.1/B7.2 (CD80, CD86) molecules. To test the biological relevance of these CTLA-4-B7 lattices, we introduced a C122A point mutation in human CTLA-4, because this residue was shown to be essential for dimerization in solution. Surprisingly, we found that up to 35% of C122A CTLA-4 dimerized in human T lymphocytes. Moreover, C122A CTLA-4 partitioned within lipid rafts, colocalized with the TCR in the immunological synapse, and inhibited T cell activation. C122-independent dimerization of CTLA-4 involved N-glycosylation, because further mutation of the N78 and N110 glycosylation sites abrogated dimerization. Despite being monomeric, the N78A/N110A/C122A triple mutant CTLA-4 localized in the immunological synapse and inhibited T cell activation. Such functionality correlated with B7-induced dimerization of these mutant molecules. Based on these data, we propose a model of hierarchical regulation of CTLA-4 oligomerization by which B7 binding ultimately determines the formation of dimer-dependent CTLA-4 lattices that may be necessary for triggering B7-dependent T cell inactivation.
Cutting Edge: Extracellular High Mobility Group Box-1 Protein Is a Proangiogenic Cytokine
Authors: Mitola Stefania , Belleri Mirella , Urbinati Chiara , Coltrini Daniela , Sparatore Bianca , Pedrazzi Marco , Melloni Edon , Presta Marco
Published: April 2014

The chromosomal high mobility group box-1 (HMGB1) protein acts as a proinflammatory cytokine when released in the extracellular environment by necrotic and inflammatory cells. In the present study, we show that HMGB1 exerts proangiogenic effects by inducing MAPK ERK1/2 activation, cell proliferation, and chemotaxis in endothelial cells of different origin. Accordingly, HMGB1 stimulates membrane ruffling and repair of a mechanically wounded endothelial cell monolayer and causes endothelial cell sprouting in a three-dimensional fibrin gel. In keeping with its in vitro properties, HMGB1 stimulates neovascularization when applied in vivo on the top of the chicken embryo chorioallantoic membrane whose blood vessels express the HMGB1 receptor for advanced glycation end products (RAGE). Accordingly, RAGE blockade by neutralizing Abs inhibits HMGB1-induced neovascularization in vivo and endothelial cell proliferation and membrane ruffling in vitro. Taken together, the data identify HMGB1/RAGE interaction as a potent proangiogenic stimulus.
Allograft Rejection Requires STAT5a/b-Regulated Antiapoptotic Activity in T Cells but Not B Cells
Authors: Zhang Ye , Kirken Robert A. , Furian Lucrezia , Janczewska Slawa , Qu Xiumei , Hancock Wayne W. , Wang Mouer , Tejpal Neelam , Kerman Ronald , Kahan Barry D. , Stepkowski Stanislaw M.
Published: April 2014

STATs play key roles in immune function. We examined the role of STAT5a/b in allograft rejection. STAT5a/b-deficient mice showed a 4-fold increased survival time of heart allografts (p < 0.01). Unlike wild type, purified STAT5a/b−/− T cells transferred to Rag1−/− recipients failed to mediate heart allograft rejection until supplemented with STAT5a/b−/− B cells. In vitro, STAT5a/b−/− T cells did not proliferate in response to Con A or alloantigens but entered apoptosis within 48 h (95%). Activated STAT5a/b−/− T cells showed increased expression of proapoptotic (caspases, DNA repair genes, TNF/TNFR-associated factor family genes) and decreased antiapoptotic mRNAs in microarrays, while Western blots confirmed reduced antiapoptotic Bcl-2 and elevated proapoptotic Bax protein expression. Interestingly, at 24 h postactivation, STAT5a/b+/+ and STAT5a/b−/− T cells produced similar levels of IL-2, IL-4, IL-10, and IFN-γ mRNA; ELISPOT assay showed an equivalent number of IL-4- and IFN-γ-producing T cells in both STAT5a/b+/+ and STAT5a/b−/− splenic populations. Sera from STAT5a/b+/+ and STAT5a/b−/− rejectors had donor-specific IgM, IgG1, IgG2a, and IgG2b Ab, while STAT5a/b deficiency had no impact on B cell survival or proliferation in response to LPS. Compared with allografts from STAT5a/b+/+ recipients, heart allografts from STAT5a/b−/− recipients had markedly reduced infiltration by CD4 and CD8 T cells but increased infiltration by B cells and dense endothelial deposition of C4d, a marker of humoral rejection. Thus, activated STAT5a/b−/− T cells produce cytokines prior to entering apoptosis, thereby promoting differentiation of B cells yielding donor-specific IgM and IgG Ab that mediate allograft rejection.
Helminth-Modified Pulmonary Immune Response Protects Mice from Allergen-Induced Airway Hyperresponsiveness
Authors: Mangan Niamh E. , van Rooijen Nico , McKenzie Andrew N. J. , Fallon Padraic G.
Published: April 2014

It has been shown that the presence of certain helminth infections in humans, including schistosomes, may reduce the propensity to develop allergies in infected populations. Using a mouse model of schistosome worm vs worm + egg infection, our objective was to dissect the mechanisms underlying the inverse relationship between helminth infections and allergies. We have demonstrated that conventional Schistosoma mansoni egg-laying male and female worm infection of mice exacerbates airway hyperresponsiveness. In contrast, mice infected with only schistosome male worms, precluding egg production, were protected from OVA-induced airway hyperresponsiveness. Worm-infected mice developed a novel modified type 2 cytokine response in the lungs, with elevated allergen-specific IL-4 and IL-13 but reduced IL-5, and increased IL-10. Although schistosome worm-only infection is a laboratory model, these data illustrate the complexity of schistosome modulation of host immunity by the worm vs egg stages of this helminth, with the potential of infections to aggravate or suppress allergic pulmonary inflammation. Thus, infection of mice with a human parasitic worm can result in reduced airway inflammation in response to a model allergen.
The TLR-7 Agonist, Imiquimod, Enhances Dendritic Cell Survival and Promotes Tumor Antigen-Specific T Cell Priming: Relation to Central Nervous System Antitumor Immunity
Authors: Prins Robert M. , Craft Noah , Bruhn Kevin W. , Khan-Farooqi Haumith , Koya Richard C. , Stripecke Renata , Miller Jeff F. , Liau Linda M.
Published: April 2014

Immunotherapy represents an appealing option to specifically target CNS tumors using the immune system. In this report, we tested whether adjunctive treatment with the TLR-7 agonist imiquimod could augment antitumor immune responsiveness in CNS tumor-bearing mice treated with human gp100 + tyrosine-related protein-2 melanoma-associated Ag peptide-pulsed dendritic cell (DC) vaccination. Treatment of mice with 5% imiquimod resulted in synergistic reduction in CNS tumor growth compared with melanoma-associated Ag-pulsed DC vaccination alone. Continuous imiquimod administration in CNS tumor-bearing mice, however, was associated with the appearance of robust innate immune cell infiltration and hemorrhage into the brain and the tumor. To understand the immunological mechanisms by which imiquimod augmented antitumor immunity, we tested whether imiquimod treatment enhanced DC function or the priming of tumor-specific CD8+ T cells in vivo. With bioluminescent, in vivo imaging, we determined that imiquimod dramatically enhanced both the persistence and trafficking of DCs into the draining lymph nodes after vaccination. We additionally demonstrated that imiquimod administration significantly increased the accumulation of tumor-specific CD8+ T cells in the spleen and draining lymph nodes after DC vaccination. The results suggest that imiquimod positively influences DC trafficking and the priming of tumor-specific CD8+ T cells. However, inflammatory responses induced in the brain by TLR signaling must also take into account the local microenvironment in the context of antitumor immunity to induce clinical benefit. Nevertheless, immunotherapeutic targeting of malignant CNS tumors may be enhanced by the administration of the innate immune response modifier imiquimod.
Reduced Ability of Neonatal and Early-Life Bone Marrow Stromal Cells to Support Plasmablast Survival
Authors: Pihlgren Maria , Friedli Mathieu , Tougne Chantal , Rochat Anne-Françoise , Lambert Paul-Henri , Siegrist Claire-Anne
Published: April 2014

In human infants (<1 year), circulating IgG Abs elicited in response to most T-dependent Ags rapidly decline and return to baseline within a few months after immunization for yet-unknown reasons. In mice immunized between 1 and 4 wk of age, a limited establishment of the bone marrow (BM) pool of long-lived plasma cells is observed. In this study, we show that tetanus toxoid (TT)-specific plasmablasts generated in the spleen are efficiently attracted in vitro and in vivo toward early-life BM stromal cells, which express adult levels of CXCL12. Similarly, adoptively transferred TT plasmablasts efficiently reach the BM compartment of 2-wk-old and adult mice. In contrast, TT plasmablasts fail to persist in the early-life BM compartment, as indicated by the persistence of a significantly lower number of TT plasmablasts in the early-life compartment than in the adult BM compartment 48 h after transfer. This limited persistence is associated with an increased rate of in vivo apoptosis of TT-specific plasmablasts that have reached the early-life BM and with a significantly lower survival rate of TT-specific plasmablasts cocultured on early-life BM stromal cells compared with adult BM stromal cells. Thus, early-life BM stromal cells fail to provide the molecular signals that support plasmablast survival and differentiation into surviving plasma cells.
The CD8+ T Cell Population Elicited by Recombinant Adenovirus Displays a Novel Partially Exhausted Phenotype Associated with Prolonged Antigen Presentation That Nonetheless Provides Long-Term Immunity
Authors: Yang Teng-Chih , Millar James , Groves Timothy , Grinshtein Natalie , Parsons Robin , Takenaka Shunsuke , Wan Yonghong , Bramson Jonathan L.
Published: April 2014

We have previously reported that the CD8+ T cell response elicited by recombinant adenovirus vaccination displayed a delayed contraction in the spleen. In our current study, we demonstrate that this unusual kinetic is a general phenomenon observed in multiple tissues. Phenotypic analysis of transgene-specific CD8+ T cells present 30 days postimmunization with recombinant adenovirus revealed a population with evidence of partial exhaustion, suggesting that the cells had been chronically exposed to Ag. Although Ag expression could no longer be detected 3 wk after immunization, examination of Ag presentation within the draining lymph nodes demonstrated that APCs were loaded with Ag peptide for at least 40 days postimmunization, suggesting that Ag remains available to the system for a prolonged period, although the exact source of this Ag remains to be determined. At 60 days postimmunization, the CD8+ T cell population continued to exhibit a phenotype consistent with partially exhausted effector memory cells. Nonetheless, these CD8+ T cells conferred sterilizing immunity against virus challenge 7–12 wk postimmunization, suggesting that robust protective immunity can be provided by CD8+ T cells with an exhausted phenotype. These data demonstrate that prolonged exposure to Ag may not necessarily impair protective immunity and prompt a re-evaluation of the impact of persistent exposure to Ag on T cell function.
Expression of CD161 (NKR-P1A) Defines Subsets of Human CD4 and CD8 T Cells with Different Functional Activities
Authors: Takahashi Tsuyoshi , Dejbakhsh-Jones Sussan , Strober Samuel
Published: April 2014

A subset of T cells in human peripheral blood expresses CD161 (NKR-P1A) receptors that are primarily associated with NK cells. In the current study we isolated blood T cell subsets according to the expression of CD161 and examined their contents of naive, central memory, and effector memory cells and their capacities for proliferation, cytokine secretion, and natural cytolysis. We found that CD4+CD161− and CD8+CD161− subsets contained predominantly naive T cells that secreted high levels of IL-2 after in vitro stimulation, and CD4+CD161int and CD8+CD161int subsets contained predominantly effector and central memory T cells that secreted high levels of IFN-γ and TNF-α. All of these subsets showed vigorous proliferation after stimulation in vitro, but none had NK lytic activity. Unexpectedly, the CD8+CD161+ cells contained an anergic CD8α+CD8βlow/−CD161high T cell subset that failed to proliferate, secrete cytokines, or mediate NK lytic activity.
IL-27 Limits IL-2 Production during Th1 Differentiation
Authors: Villarino Alejandro V. , Stumhofer Jason S. , Saris Christiaan J. M. , Kastelein Robert A. , de Sauvage Frederic J. , Hunter Christopher A.
Published: April 2014

Although the ability of IL-27 to promote T cell responses is well documented, the anti-inflammatory properties of this cytokine remain poorly understood. The current work demonstrates that during infection with Toxoplasma gondii, IL-27R-deficient mice generate aberrant IL-2 responses that are associated with the development of a lethal inflammatory disease. Because in vivo depletion of IL-2 prolongs the survival of infected IL-27R−/− mice, these data suggest that IL-27 curbs the development of immunopathology by limiting parasite-induced IL-2 production. Consistent with this hypothesis, IL-27R−/− CD4+ T cells produce more IL-2 than wild-type counterparts during in vitro differentiation, and when rIL-27 is introduced, it can suppress the expression of IL-2 mRNA and protein by the latter group. Additionally, these studies reveal that, like IL-27, IL-12 can inhibit IL-2 production, and although each employs distinct mechanisms, they can synergize to enhance the effect. In contrast, this property is not shared by closely related cytokines IL-6 and IL-23. Thus, while traditionally viewed as proinflammatory agents, the present findings establish that IL-27 and IL-12 cooperate to limit the availability of IL-2, a potent T cell growth and survival factor. Moreover, because the current studies demonstrate that both can induce expression of suppressor of cytokine signaling 3, a protein that tempers cytokine receptor signaling, they also suggest that IL-27 and IL-12 share additionally inhibitory properties.