bluuebunny/crossref_metadata_2025 huggingface dataset:
Galectin-3 Deficiency Reduces the Severity of Experimental Autoimmune Encephalomyelitis
Authors: Jiang Hui-Rong , Al Rasebi Zakeya , Mensah-Brown Eric , Shahin Allen , Xu Damo , Goodyear Carl S. , Fukada Sandra Y. , Liu Fu-Tong , Liew Foo Y. , Lukic Miodrag L.
Published: April 2014

Galectin-3 (Gal-3) is a member of the β-galactoside-binding lectin family and plays an important role in inflammation. However, the precise role of Gal-3 in autoimmune diseases remains obscure. We have investigated the functional role of Gal-3 in experimental autoimmune encephalomyelitis (EAE) following immunization with myelin oligodendrocyte glycoprotein (MOG)35–55 peptide. Gal-3 deficient (Gal-3−/−) mice developed significantly milder EAE and markedly reduced leukocyte infiltration in the CNS compared with similarly treated wild-type (WT) mice. Gal-3−/− mice also contained fewer monocytes and macrophages but more apoptotic cells in the CNS than did WT mice. Following Ag stimulation in vitro, lymph node cells from the immunized Gal-3−/− mice produced less IL-17 and IFN-γ than did those of the WT mice. In contrast, Gal-3−/− mice produced more serum IL-10, IL-5, and IL-13 and contained higher frequency of Foxp3+ regulatory T cells in the CNS than did the WT mice. Furthermore, bone marrow-derived dendritic cells from Gal-3−/− mice produced more IL-10 in response to LPS or bacterial lipoprotein than did WT marrow-derived dendritic cells. Moreover, Gal-3−/− dendritic cells induced Ag-specific T cells to produce more IL-10, IL-5, and IL-12, but less IL-17, than did WT dendritic cells. Taken together, our data demonstrate that Gal-3 plays an important disease-exacerbating role in EAE through its multifunctional roles in preventing cell apoptosis and increasing IL-17 and IFN-γ synthesis, but decreasing IL-10 production.
Acquisition of Anergy to Proinflammatory Cytokines in Nonimmune Cells through Endoplasmic Reticulum Stress Response: A Mechanism for Subsidence of Inflammation
Authors: Hayakawa Kunihiro , Hiramatsu Nobuhiko , Okamura Maro , Yamazaki Hiroaki , Nakajima Shotaro , Yao Jian , Paton Adrienne W. , Paton James C. , Kitamura Masanori
Published: April 2014

Acute endoplasmic reticulum (ER) stress causes induction of inflammatory molecules via activation of NF-κB. However, we found that, under ER stress conditions, renal mesangial cells acquire anergy to proinflammatory stimuli. Priming of the cells with ER stress inducers (tunicamycin, thapsigargin, A23187, and AB5 subtilase cytotoxin) caused blunted induction of MCP-1 in response to TNF-α, IL-1β, macrophage-derived factors, or bystander macrophages. The magnitude of suppression was closely correlated with the level of GRP78, an endogenous indicator of ER stress. The suppression of MCP-1 under ER stress conditions was reversible and observed in general regardless of cell types or triggers of ER stress. The decrease in the level of MCP-1 mRNA was ascribed to transcriptional suppression via unexpected inhibition of NF-κB, but not to accelerated mRNA degradation. Subsequent experiments revealed that TNFR-associated factor 2, an essential component for TNF-α signaling, was down-regulated by ER stress. We also found that, under ER stress conditions, expression of NF-κB suppressor A20 was induced. Overexpression of A20 resulted in suppression of cytokine-triggered NF-κB activation and knockdown of A20 by RNA interference significantly attenuated induction of anergy by ER stress. In contrast, other ER stress-inducible/-related molecules that may suppress NF-κB (e.g., GRP78, NO, reactive oxygen species, and IκB) were not involved in the inhibitory effects of ER stress. These results elucidated ER stress-dependent mechanisms by which nonimmune cells acquire anergy to inflammatory stimuli under pathological situations. This self-defense machinery may play a role in halting progression of acute inflammation and in its spontaneous subsidence.
Potent Induction of IFN-α and Chemokines by Autoantibodies in the Cerebrospinal Fluid of Patients with Neuropsychiatric Lupus
Authors: Santer Deanna M. , Yoshio Taku , Minota Seiji , Möller Thomas , Elkon Keith B.
Published: April 2014

Neuropsychiatric disease in systemic lupus erythematosus (NPSLE) is a poorly understood, but potentially fatal, disease manifestation. A pathogenetic role for autoantibodies is suspected, but the mechanism is unclear. Since immune complexes in SLE can stimulate IFN-α and there is strong evidence in humans and in mice that IFN-α can cause neuropsychiatric manifestations, we asked whether NPSLE patient serum and/or cerebrospinal fluid (CSF) contain abnormally high IFN-α-inducing activity. In a bioassay containing plasmacytoid dendritic cells and a source of Ag, NPSLE CSF induced significantly higher IFN-α compared with CSF from patients with multiple sclerosis or other autoimmune disease controls. When normalized for IgG concentration, NPSLE CSF was 800-fold more potent at inducing IFN-α compared with paired serum due to inhibitors present in serum. Analysis of Ig-deficient patient serum, depletion of IgG from normal serum, as well as addition of purified IgG to NPSLE CSF and serum in the bioassays revealed that one inhibitor was contained within the IgG fraction itself. In addition to IFN-α, immune complexes formed by CSF autoantibodies produced significantly increased levels of IFN-γ-inducible protein 10 (IP-10/CXCL), IL-8, and MCP-1, all of which have been reported to be elevated in CSF from NPSLE patients. Taken together, these findings are consistent with a two-step model of NPSLE whereby CSF autoantibodies bind to Ags released by neurocytotoxic Abs or other brain cell injury, and the resulting immune complexes stimulate IFN-α and proinflammatory cytokines and chemokines.
Mucosally Delivered Salmonella Typhi Expressing the Yersinia pestis F1 Antigen Elicits Mucosal and Systemic Immunity Early in Life and Primes the Neonatal Immune System for a Vigorous Anamnestic Response to Parenteral F1 Boost
Authors: Ramirez Karina , Capozzo Alejandra V. E. , Lloyd Scott A. , Sztein Marcelo B. , Nataro James P. , Pasetti Marcela F.
Published: April 2014

Neonates respond poorly to conventional vaccines. This has been attributed, in part, to the immaturity of neonatal dendritic cells that lack full capacity for Ag presentation and T cell stimulation. We engineered an attenuated Salmonella Typhi strain to express and export the F1 Ag of Y. pestis (S. Typhi(F1)) and investigated its immunogenicity early in life using a heterologous prime-boost regimen. Newborn mice primed intranasally with a single dose of S. Typhi(F1) elicited mucosal Ab- and IFN-γ-secreting cells 1 wk after immunization. They also developed a potent and fast anamnestic response to a subsequent parenteral boost with F1-alum, which surpassed those of mice primed and boosted with S. Typhi(F1) or F1-alum. Neonatal priming with S. Typhi(F1), as opposed to priming with F1-alum, resulted in a more balanced IgG2a/IgG1 profile, enhanced avidity maturation and stimulation of B memory cells, and strong Th1-type cell-mediated immunity. S. Typhi(F1) enhanced the activation and maturation of neonatal CD11c+ dendritic cells, shown by increased expression of CD80, CD86, CD40, and MHC-II cell surface markers and production of proinflammatory cytokines IL-12, TNF-α, IL-6, and MCP-1. S. Typhi(F1)-stimulated neonatal DC had improved capacity for Ag presentation and T cell stimulation in vitro and induced F1-specific CD4+ and CD8+ T cell responses when adoptively transferred to newborn mice. Mucosal immunization with S. Typhi expressing a foreign Ag effectively primes the neonatal immune system for potent, fast, and broader responses to a parenteral Ag boost. Such a strategy can prevent infectious diseases, including those considered biowarfare threats, early in life.
Cutting Edge: Ikaros Is a Regulator of Th2 Cell Differentiation
Authors: Quirion Mary R. , Gregory Gregory D. , Umetsu Sarah E. , Winandy Susan , Brown Melissa A.
Published: April 2014

Ikaros, a hematopoietic transcription factor, has well defined effects on early lymphocyte development in the bone marrow and thymus. In this study we demonstrate that Ikaros is a positive regulator of Th2 cytokine gene expression in peripheral T cells. CD4+ T cells from naive Ikarosnull mice cultured under Th2-skewing conditions express the Th1 cytokine IFN-γ and have reduced IL-4, IL-5, and IL-13 expression. Ikaros directly associates with several Th2 locus regulatory regions in naive CD4+ T cells. The decreased ability to express Th2 cytokines in Ikarosnull T cells corresponds with histone 3 hypoacetylation across the Th2 cytokine locus as well as decreased GATA3 and cMaf and increased T-bet and STAT1 expression. These data support a model whereby Ikaros directly activates Th2 gene expression by promoting local chromatin accessibility during CD4+ T cell differentiation and also acts indirectly to regulate expression of Th2- and Th1-specific transcription factors.
Cutting Edge: Developmental Up-Regulation of IFN-γ-Inducible Lysosomal Thiol Reductase Expression Leads to Reduced T Cell Sensitivity and Less Severe Autoimmunity
Authors: Maric Maja , Barjaktarevic Igor , Bogunovic Branka , Stojakovic Milica , Maric Christine , Vukmanovic Stanislav
Published: April 2014

Reactivity to self-peptide/MHC complexes is required for selection of the TCR repertoire in the thymus but can also promote autoimmunity. Reduced TCR sensitivity of mature T cells is thought to help control the autoreactivity in peripheral T cells. The molecular basis for reduced sensitivity of peripheral T cells is not known. We found that peripheral T cells, but not immature thymocytes, lacking IFN-γ-inducible lysosomal thiol reductase (GILT) display increased sensitivity to TCR ligation. GILT−/− peripheral T cells express reduced levels of mitochondrial superoxide dismutase 2 and consequently display higher levels of reactive oxygen radicals and ERK1/2 phosphorylation following activation. The increased sensitivity of GILT-deficient T cells results in a more severe hyperglycemia associated with streptozotocin-induced diabetes. GILT expression levels progressively increase in T cells with maturation. These data suggest that regulation of GILT expression may be a mechanism of T cell differentiation-associated changes in sensitivity to TCR engagement.
Pre-TCR-Induced β-Catenin Facilitates Traversal through β-Selection
Authors: Xu Mai , Sharma Archna , Wiest David L. , Sen Jyoti Misra
Published: April 2014

Pre-TCR induced signals regulate development of the αβ TCR lineage cells at the β-selection checkpoint. We have previously shown that conditional deletion of β-catenin, a central mediator of Wnt-β-catenin-T cell factor signaling pathway, impairs traversal through the β-selection checkpoint. We now provide a molecular basis for the impairment. We demonstrate that pre-TCR signals specifically stabilize β-catenin in CD4−CD8− double negative thymocytes during β-selection. Pre-TCR induced Erk activity was required to stabilize β-catenin. Enforced expression of stabilized β-catenin was sufficient to mediate aspects of β-selection including sustained expression of early growth response (Egr) genes. Consistently, deletion of β-catenin reduced induction of Egr gene expression by the pre-TCR signal and blocked efficient β-selection. Thus, we demonstrate that pre-TCR induced β-catenin sustains expression of Egr genes that facilitate traversal through the β-selection checkpoint.
Sustained Expression of Pre-TCR Induced β-Catenin in Post-β-Selection Thymocytes Blocks T Cell Development
Authors: Xu Mai , Sharma Archna , Hossain M. Zulfiquer , Wiest David L. , Sen Jyoti Misra
Published: April 2014

Pre-TCR and IL-7R signals regulate β-selection of thymocytes and then must be down-regulated for further development. However, the molecular events that control down-regulation remain unknown. We and others have previously shown that β-catenin in cooperation with TCF regulates β-selection. In this paper, we demonstrate that β-catenin expression is stringently regulated by intrathymic signals, it is expressed at the highest levels in the pre-TCR signaled thymocytes, and is down-regulated in post-β-selection thymocytes. Pre-TCR-induced β-catenin regulates initial stages of pre-TCR signaling including expression of early growth response (Egr) genes but must be down-regulated to express RORγt, which is essential for maturation to the CD4+CD8+ double positive (DP) stage. Sustained expression of β-catenin results in the generation of IL-7R-, Egr-, and TGFβ-expressing pre-DP thymocytes that are blocked in development. These data are consistent with a model in which post-β-selection, pre-TCR-induced β-catenin expression must return to background levels for efficient transition to the DP stage.
Cross-Linking of CD80 on CD4+ T Cells Activates a Calcium-Dependent Signaling Pathway
Authors: Podojil Joseph R. , Miller Stephen D.
Published: April 2014

CD80 expressed on the surface of APCs provides a positive costimulatory signal to naive CD4+ T cells via CD28 during activation. However, CD80 is also expressed on the surface of activated CD4+ T cells, and cross-linking CD80 on the surface of CD4+ T cells activated in the presence of Th1-promoting cytokines induces a direct up-regulation of T-bet, IFN-γ, and Bcl(XL) expression in primary CD4+ T cells. The present data show that naive CD4+ T cells activated in Th1-promoting conditions in the presence of anti-CD80 mAb increase the level of IFN-γ produced by increasing the rate of IFN-γ mRNA transcription, which is supported by an increase in the level of T-bet phosphorylation and T-bet binding to the third intronic enhancer in the IFN-γ locus. Furthermore, anti-CD80 mAb-induced increase in IFN-γ expression and T-bet phosphorylation is dependent upon the activation of a Ca2+-dependent pathway as shown by anti-CD80 mAb-induced intracellular Ca2+ flux following CD80 cross-linking. These findings indicate a novel regulatory role for CD80-mediated intracellular signals in CD4+ T cells and have important implications for disease therapies using anti-costimulatory mAbs as use of an intact CD80 mAb may lead to CD80 cross-linking on activated T cells and enhanced proinflammatory cytokine production.
Priming of CD8+ and CD4+ T Cells in Experimental Leishmaniasis Is Initiated by Different Dendritic Cell Subtypes
Authors: Brewig Nancy , Kissenpfennig Adrien , Malissen Bernard , Veit Alexandra , Bickert Thomas , Fleischer Bernhard , Mostböck Sven , Ritter Uwe
Published: April 2014

The biological role of Langerin+ dendritic cells (DCs) such as Langerhans cells and a subset of dermal DCs (dDCs) in adaptive immunity against cutaneous pathogens remains enigmatic. Thus, we analyzed the impact of Langerin+ DCs in adaptive T cell-mediated immunity toward Leishmania major parasites in a Lang-DTR mouse model that allows conditional diphtheria toxin (DT)-induced ablation of Langerin+ DCs in vivo. For the first time, infection experiments with DT-treated Lang-DTR mice revealed that proliferation of L. major-specific CD8+ T cells is significantly reduced during the early phase of the immune response following depletion of Langerin+ DCs. Consequently, the total number of activated CD8+ T cells within the draining lymph node and at the site of infection is diminished. Furthermore, we show that the impaired CD8+ T cell response is due to the absence of Langerin+ dDCs and not Langerhans cells. Nevertheless, the CD4+ T cell response is not altered and the infection is cleared as effectively in DT-treated Lang-DTR mice as in control mice. This clearly demonstrates that Langerin+ DCs are, in general, dispensable for an efficient adaptive immune response against L. major parasites. Thus, we propose a novel concept that, in the experimental model of leishmaniasis, priming of CD4+ T cells is mediated by Langerin− dDCs, whereas Langerin+ dDCs are involved in early priming of CD8+ T cells.
Clonal Expansions and Loss of Receptor Diversity in the Naive CD8 T Cell Repertoire of Aged Mice
Authors: Ahmed Mushtaq , Lanzer Kathleen G. , Yager Eric J. , Adams Pamela S. , Johnson Lawrence L. , Blackman Marcia A.
Published: April 2014

There are well-characterized age-related changes in the peripheral repertoire of CD8 T cells characterized by reductions in the ratio of naive:memory T cells and the development of large clonal expansions in the memory pool. In addition, the TCR repertoire of naive T cells is reduced with aging. Because a diverse repertoire of naive T cells is essential for a vigorous response to new infections and vaccinations, there is much interest in understanding the mechanisms responsible for declining repertoire diversity. It has been proposed that one reason for declining repertoire diversity in the naive T cell pool is an increasing dependence on homeostatic proliferation in the absence of new thymic emigrants for maintenance of the naive peripheral pool. In this study, we have analyzed the naive CD8 T cell repertoire in young and aged mice by DNA spectratype and sequence analysis. Our data show that naive T cells from aged mice have perturbed spectratype profiles compared with the normally Gaussian spectratype profiles characteristic of naive CD8 T cells from young mice. In addition, DNA sequence analysis formally demonstrated a loss of diversity associated with skewed spectratype profiles. Unexpectedly, we found multiple repeats of the same sequence in naive T cells from aged but not young mice, consistent with clonal expansions previously described only in the memory T cell pool. Clonal expansions among naive T cells suggests dysregulation in the normal homeostatic proliferative mechanisms that operate in young mice to maintain diversity in the naive T cell repertoire.
Tuberculosis-Induced Variant IL-4 mRNA Encodes a Cytokine Functioning As Growth Factor for (E)-4-Hydroxy-3-Methyl-But-2-Enyl Pyrophosphate-Specific Vγ2Vδ2 T Cells
Authors: Yuan ZhuQing , Wang Richard , Lee Yuyang , Chen Crystal Y. , Yu XingBing , Wu ZhongDao , Huang Dan , Shen Ling , Chen Zheng W.
Published: April 2014

The possibility that mycobacterial infections induce variant cytokine mRNA encoding a functionally distinct protein for immune regulation has not been addressed. In this study, we reported that Mycobacterium tuberculosis and bacillus Calmette-Guérin infections of macaques induced expression of variant IL-4 (VIL-4) mRNA encoding a protein comprised of N-terminal 97 aa identical with IL-4, and unique C-terminal 96 aa including a signaling-related proline-rich motif. While VIL-4 could be stably produced as intact protein, the purified VIL-4 induced apparent expansion of phosphoantigen (E)-4-hydroxy-3-methyl-but-2-enyl pyrophosphate (HMBPP)-specific Vγ2Vδ2 T cells in dose- and time-dependent manners. The unique C-terminal 96 aa bearing the proline-rich motif (PPPCPP) of VIL-4 appeared to confer the ability to expand Vγ2Vδ2 T cells, since simultaneously produced IL-4 had only a subtle effect on these γδ T cells. Moreover, VIL-4 seemed to use IL-4R α for signaling and activation, as the VIL-4-induced expansion of Vγ2Vδ2 T cells was blocked by anti-IL-4R α mAb but not anti-IL-4 mAb. Surprisingly, VIL-4-expanded Vγ2Vδ2 T cells after HMBPP stimulation appeared to be heterologous effector cells capable of producing IL-4, IFN-γ, and TNF-α. Thus, mycobacterial infections of macaques induced variant mRNA encoding VIL-4 that functions as growth factor promoting expansion of HMBPP-specific Vγ2Vδ2 T effector cells.
Plasmacytoid Dendritic Cells Are Dispensable during Primary Influenza Virus Infection
Authors: Wolf Amaya I. , Buehler Darya , Hensley Scott E. , Cavanagh Lois L. , Wherry E. John , Kastner Philippe , Chan Susan , Weninger Wolfgang
Published: April 2014

Plasmacytoid dendritic cells (pDC) are thought to be pivotal in the first line of defense against viral infections. Although previous studies have suggested that pDC regulate the immune response against respiratory syncytial virus, their role in pulmonary infection with influenza virus has remained unclear. Using mice with GFP-tagged pDC, we observed a marked increase in pDC numbers in the lung airways 3 days after intranasal infection with influenza virus A/PR/8/34. To further investigate their potential involvement in the disease, we made use of pDC-deficient IkarosL/L mice. In the absence of pDC, the recruitment of T cells to the bronchoalveolar space was delayed, which could be reversed by the adoptive transfer of pDC before infection. Surprisingly, however, when compared with wild-type animals, IkarosL/L mice revealed a similar course of disease, as determined by weight loss, viral titers, levels of neutralizing Ab, and lung pathology. Moreover, the activation and differentiation of influenza-specific CD8+ effector T cells was unaltered in the absence of pDC, as was the generation of CD8+ memory T cells. Taken together, our study suggests that pDC regulate the accumulation of T cells in the bronchoalveolar space during early influenza virus infection, but are dispensable for the control of this disease.
Modified Adenine (9-Benzyl-2-Butoxy-8-Hydroxyadenine) Redirects Th2-Mediated Murine Lung Inflammation by Triggering TLR7
Authors: Vultaggio Alessandra , Nencini Francesca , Fitch Paul M. , Filì Lucia , Maggi Laura , Fanti Paola , deVries Annick , Beccastrini Enrico , Palandri Francesca , Manuelli Cinzia , Bani Daniele , Giudizi Maria Grazia , Guarna Antonio , Annunziato Francesco , Romagnani Sergio , Maggi Enrico , Howie Sarah E. M. , Parronchi Paola
Published: April 2014

Substitute adenine (SA)-2, a synthetic heterocycle chemically related to adenine with substitutions in positions 9-, 2-, and 8- (i.e., 9-benzyl-2-butoxy-8-hydroxyadenine), induces in vitro immunodeviation of Th2 cells to a Th0/Th1 phenotype. In this article, we evaluate the in vivo ability of SA-2 to affect Th2-mediated lung inflammation and its safety. TLR triggering and NF-κB activation by SA-2 were analyzed on TLR-transfected HEK293 cells and on purified bone marrow dendritic cells. The in vivo effect of SA-2 on experimental airway inflammation was evaluated in both prepriming and prechallenge protocols by analyzing lung inflammation, including tissue eosinophilia and goblet cell hyperplasia, bronchoalveolar lavage fluid cell types, and the functional profile of Ag-specific T cells from draining lymph nodes and spleens. SA-2 induced mRNA expression and production of proinflammatory (IL-6, IL-12, and IL-27) and regulatory (IL-10) cytokines and chemokines (CXCL10) in dendritic cells but down-regulated TGF-β. Prepriming administration of SA-2 inhibited OVA-specific Abs and Th2-driven lung inflammation, including tissue eosinophilia and goblet cells, with a prevalent Foxp3-independent regulatory mechanism. Prechallenge treatment with SA-2 reduced the lung inflammation through the induction of a prevalent Th1-related mechanism. In this model the activity of SA-2 was route-independent, but adjuvant- and Ag dose-dependent. SA-2-treated mice did not develop any increase of serum antinuclear autoantibodies. In conclusion, critical substitutions in the adenine backbone creates a novel synthetic TLR7 ligand that shows the ability to ameliorate Th2-mediated airway inflammation by a complex mechanism, involving Th1 redirection and cytokine-mediated regulation, which prevents autoreactivity.
Differential Capability of Human Cutaneous Dendritic Cell Subsets to Initiate Th17 Responses
Authors: Mathers Alicia R. , Janelsins Brian M. , Rubin Joseph P. , Tkacheva Olga A. , Shufesky William J. , Watkins Simon C. , Morelli Adrian E. , Larregina Adriana T.
Published: April 2014

Human skin-migratory dendritic cells (DCs) have the ability to prime and bias Th1 and Th2 CD4+ T lymphocytes. However, whether human cutaneous DCs are capable of initiating proinflammatory Th17 responses remains undetermined. We report that skin-migratory DCs stimulate allogeneic naive CD4+ T cells that differentiate simultaneously into two distinct effector Th17 and Th1 populations capable of homing to the skin, where they induce severe cutaneous damage. Skin-migratory Langerhans cells (smiLCs) were the main cutaneous DC subset capable of inducing Th17 responses dependent on the combined effects of IL-15 and stabilized IL-6, which resulted in IL-6 trans-signaling of naive CD4+ T cells. Different from smiLCs, purified skin-migratory dermal DCs did not synthesize IL-15 and were unable to bias Th17 responses. Nevertheless, these dermal DCs were capable of differentiating Th17 cells in mixed leukocyte cultures supplemented with IL-15 and stabilized IL-6. Overall, our data demonstrate that human epidermal smiLCs induce Th17 responses by mechanisms different from those previously described and highlight the need to target clinical treatments based on these variations.
Cutting Edge: An IL-17F-CreEYFP Reporter Mouse Allows Fate Mapping of Th17 Cells
Authors: Croxford Andrew L. , Kurschus Florian C. , Waisman Ari
Published: April 2014

The need for reporter lines able to faithfully track Th17 cells in vivo has become an issue of exceptional importance. To address this, we generated a mouse strain in which Cre recombinase is expressed from the IL-17F promoter. Crossing the IL-17F-Cre allele to a conditional enhanced yellow fluorescent protein (EYFP) reporter mouse yielded the IL-17F-CreEYFP strain, in which IL-17F expression is twinned with EYFP in live IL-17F-expressing cells. Although we demonstrate that IL-17F expression is restricted to CD4+ T cells during experimental autoimmune encephalomyelitis, IL-17F-CreEYFP CD8 T cells robustly expressed IL-17F in response to TGF-β, IL-6, and IL-23. Fate mapping of IL-17F-expressing reporter T cells revealed a significant down-regulation of Th17 cytokines after homeostatic expansion in RAG1-deficient animals. Despite this loss of effector phenotype, committed Th17 cells were resistant to Foxp3 expression in vitro or in vivo. Thus, the IL-17F-Cre strain furthers our understanding of Th17 biology.
Cutting Edge: Permissive MHC Class II Allele Changes the Pattern of Antitumor Immune Response Resulting in Failure of Tumor Rejection
Authors: Klyushnenkova Elena N. , Kouiavskaia Diana V. , Berard Carla A. , Alexander Richard B.
Published: April 2014

We studied the growth of transgenic adenocarcinoma of mouse prostate (TRAMP)-C1 tumor cells expressing human prostate-specific Ag (PSA) in HLA-DRB1*1501 (DR2b) transgenic mice. TRAMP-PSA tumors were frequently rejected by HLA-DR2b− mice but had increased incidence in HLA-DR2b+ littermates. The levels of PSA-specific CD8 T cell responses were significantly higher in the HLA-DR2b− mice that rejected TRAMP-PSA tumors compared with HLA-DR2b+ tumor-bearing littermates. In contrast, Ab responses to PSA were strong in HLA-DR2b+ mice bearing TRAMP-PSA tumors and were virtually undetectable in HLA-DR2b− littermates. The analysis of CD4 T cell responses to PSA revealed the presence of several CD4 T cell epitopes in HLA-DR2b+ mice but failed to identify strong I-Ab-restricted epitopes in HLA-DR2b− mice. Our data demonstrate that the expression of a permissive HLA class II allele can change the pattern of the immune response to a tumor Ag, resulting in the failure of tumor rejection.
ISCOMATRIX Adjuvant Induces Efficient Cross-Presentation of Tumor Antigen by Dendritic Cells via Rapid Cytosolic Antigen Delivery and Processing via Tripeptidyl Peptidase II
Authors: Schnurr Max , Orban Martin , Robson Neil C. , Shin Amanda , Braley Hal , Airey Denise , Cebon Jonathan , Maraskovsky Eugene , Endres Stefan
Published: April 2014

Cancer vaccines aim to induce antitumor CTL responses, which require cross-presentation of tumor Ag to CTLs by dendritic cells (DCs). Adjuvants that facilitate cross-presentation of vaccine Ag are therefore key for inducing antitumor immunity. We previously reported that human DCs could not efficiently cross-present the full-length cancer/testis Ag NY-ESO-1 to CTL unless formulated as either an immune complex (NY-ESO-1/IC) or with ISCOMATRIX adjuvant. We now demonstrate that NY-ESO-1/ICs induce cross-presentation of HLA-A2- and HLA-Cw3-restricted epitopes via a proteasome-dependent pathway. In contrast, cross-presentation of NY-ESO-1/ISCOMATRIX vaccine was proteasome independent and required the cytosolic protease tripeptidyl peptidase II. Trafficking studies revealed that uptake of ICs and ISCOMATRIX vaccine by DCs occurred via endocytosis with delivery to lysosomes. Interestingly, ICs were retained in lysosomes, whereas ISCOMATRIX adjuvant induced rapid Ag translocation into the cytosol. Ag translocation was dependent on endosomal acidification and IL-4-driven differentiation of monocytes into DCs. This study demonstrates that Ag formulation determines Ag processing and supports a role for tripeptidyl peptidase II in cross-presentation of CTL epitopes restricted to diverse HLA alleles.
A New Triggering Receptor Expressed on Myeloid Cells (Trem) Family Member, Trem-Like 4, Binds to Dead Cells and Is a DNAX Activation Protein 12-Linked Marker for Subsets of Mouse Macrophages and Dendritic Cells
Authors: Hemmi Hiroaki , Idoyaga Juliana , Suda Koji , Suda Nao , Kennedy Kathleen , Noda Masaki , Aderem Alan , Steinman Ralph M.
Published: April 2014

Dendritic cells (DCs) are professional APCs that can control immune responses against self and altered self, typically foreign, determinants. DCs can be divided into several subsets, including CD8α+ and CD8α− DCs. These subsets possess specific functions. For example, mouse splenic CD8α+, but not CD8α− DCs selectively take up dying cells and cross-present cell-associated Ags to naive T cells. In this study, we identified genes that were more expressed in CD8α+ than CD8α− DCs by microarray analysis. Only one of these genes, when the extracellular domains were linked to human IgG Fc domain, could bind to late apoptotic or necrotic cells. This gene was a new member of the triggering receptor expressed on myeloid cells (Trem) family, Trem-like 4 (Treml4). Treml4 mRNA and protein, the latter detected with a new mAb, were predominantly expressed in spleen. Treml4, like other Trem family members, could associate with the adaptor molecule DNAX activation protein 12 kDa, but neither DNAX activation protein 10 kDa nor FcRγ. Consistent with the microarray data, we confirmed that Treml4 protein was more expressed on CD8α+ than CD8α− DCs, and we also found that Treml4 was expressed at high levels on splenic macrophages in spleen, particularly red pulp and marginal metallophilic macrophages. In addition, Treml4 expression on DCs was not changed after maturation induced by TLR ligands. Thus, Treml4 is a new Trem family molecule that is abundantly expressed on CD8α+ DCs and subsets of splenic resident macrophages, and can recognize dead cells by different types of phagocytes in spleen.
CXC Chemokine Ligand 12 Promotes CCR7-Dependent Naive T Cell Trafficking to Lymph Nodes and Peyer’s Patches
Authors: Bai Zhongbin , Hayasaka Haruko , Kobayashi Masayoshi , Li Wenzhe , Guo Zijin , Jang Myoung Ho , Kondo Akihiro , Choi Byung-il , Iwakura Yoichiro , Miyasaka Masayuki
Published: April 2014

A number of chemokines, including CCL21, CCL19, CXCL12, and CXCL13, are coexpressed on the lumen or basal lamina of high endothelial venules (HEVs) in lymph nodes (LNs) and Peyer’s patches (PPs), consistent with the idea that they might cooperate to regulate lymphocyte trafficking into these lymphoid tissues. In this study we report that CXCL12, acting through its receptor, CXCR4, cooperates with CCR7 ligands to promote T cell trafficking across HEVs. CXCL12 enhanced the CCR7-induced chemotaxis of wild-type but not CXCR4-deficient T cells in vitro at suboptimal concentrations of a CCR7 ligand, but without affecting the expression level or ligand-binding ability of CCR7. Real-time chemotaxis analysis showed that CXCL12 substantially shortened the lag time before cell migration began in vitro, but not the migration speed of T cells responding to suboptimal CCR7 ligand concentrations. In addition, CXCL12 augmented the CCR7 ligand-driven ERK phosphorylation and actin polymerization in T cells under the same conditions. In adoptive transfer experiments, CXCL12 promoted naive T cell trafficking to LNs and PPs in wild-type but not CCR7 ligand-deficient plt/plt recipient mice; this increased T cell trafficking was associated with enhanced binding of the T cells to HEVs and their subsequent migration into the LN parenchyma. Thus, CXCL12 synergizes with CCR7 ligands to promote T cell migration by sensitizing T cells through CXCR4, thus enabling them to respond to lower concentrations of CCR7 ligands. Such concerted action of chemokines provides an additional, previously unknown mechanism for efficient lymphocyte trafficking across HEVs into LNs and PPs.
MicroRNA-513 Regulates B7-H1 Translation and Is Involved in IFN-γ-Induced B7-H1 Expression in Cholangiocytes
Authors: Gong Ai-Yu , Zhou Rui , Hu Guoku , Li Xiaoqing , Splinter Patrick L. , O'Hara Steven P. , LaRusso Nicholas F. , Soukup Garrett A. , Dong Haidong , Chen Xian-Ming
Published: April 2014

Biliary epithelial cells (cholangiocytes) respond to proinflammatory cytokines such as IFN-γ and actively participate in the regulation of biliary inflammatory response in the liver. B7-H1 (also known as CD274 or PD-L1) is a member of the B7 costimulatory molecules and plays a critical immunoregulatory role in cell-mediated immune responses. In this study, we show that resting human cholangiocytes in culture express B7-H1 mRNA, but not B7-H1 protein. IFN-γ induces B7-H1 protein expression and alters the microRNA (miRNA) expression profile in cholangiocytes. Of those IFN-γ-down-regulated miRNAs, we identified microRNA-513 (miR-513) with complementarity to the 3′-untranslated region of B7-H1 mRNA. Targeting of the B7-H1 3′-untranslated region by miR-513 results in translational repression. Transfection of cholangiocytes with an antisense oligonucleotide to miR-513 induces B7-H1 protein expression. Additionally, transfection of miR-513 precursor decreases IFN-γ-induced B7-H1 protein expression and consequently influences B7-H1-associated apoptotic cell death in cocultured Jurkat cells. Thus, miR-513 regulates B7-H1 translation and is involved in IFN-γ-induced B7-H1 expression in human cholangiocytes, suggesting a role for miRNA-mediated gene silencing in the regulation of cholangiocyte response to IFN-γ.
Protein Tyrosine Phosphatases Regulate Asthma Development in a Murine Asthma Model
Authors: Pouliot Philippe , Camateros Pierre , Radzioch Danuta , Lambrecht Bart N. , Olivier Martin
Published: April 2014

Allergic asthma is a chronic inflammatory disease characterized by Th2-type inflammation. Although the cellular interactions are now well studied, the intracellular signaling involved in asthma development is still a developing field. Protein tyrosine kinases are one focus of such research and their inhibition shows improvement of asthmatic features. Interestingly, very little attention was given to protein tyrosine phosphatases (PTPs), the counterparts to protein tyrosine kinases, in the development of asthma. Previous studies from our laboratory showed that pharmacological inhibition of PTPs induced a transient Th1 response in the spleen. Therefore, we hypothesized that modulation of PTPs could influence asthma development. To assess PTP functions, we used the PTP inhibitor bis-peroxovanadium bpV(phen) in a murine model of asthma during either allergen sensitization or challenge. Inhibition of PTPs during allergen sensitization resulted in the reduction of key features of allergic asthma: serum IgE levels, lung tissue inflammation, eosinophilia, and airway hyperresponsiveness. Of utmost interest, PTP inhibition at allergen challenge resulted in a very similar improvement of asthmatic features. Of further importance, we observed that bpV(phen) treatment modulated cytokine expression in the spleen and, more specifically, favored Th1 cytokines while inhibiting Th2 cytokines. Collectively, we show for the first time that intact activity of PTPs is required for a complete induction of asthma in a mouse model. This clearly suggests that PTPs have a pivotal regulatory role in the development of asthmatic diseases, which opens the possibility of new therapeutic avenues.
Affinity-Based Selection of Regulatory T Cells Occurs Independent of Agonist-Mediated Induction of Foxp3 Expression
Authors: Relland Lance M. , Mishra Manoj K. , Haribhai Dipica , Edwards Brandon , Ziegelbauer Jennifer , Williams Calvin B.
Published: April 2014

Natural regulatory T (nTreg) cells recognize self-peptides with high affinity, yet the understanding of how affinity influences their selection in the thymus is incomplete. We use altered peptide ligands in transgenic mice and in organ culture to create thymic environments spanning a broad range of ligand affinity. We demonstrate that the nTreg TCR repertoire is shaped by affinity-based selection, similar to conventional T cells. The effect of each ligand on the two populations is distinct, consistent with early nTreg cell lineage specification. Foxp3 expression is an independent process that does not rely on “high affinity” binding per se, but requires a high-potency agonistic interaction for its induction. The timing of ligand exposure, TGFβ signaling, and the organization of the thymic architecture are also important. The development of nTreg cells is therefore a multistep process in which ligand affinity, potency, and timing of presentation all play a role in determining cell fate.
Placental Cytokine Expression Covaries with Maternal Asthma Severity and Fetal Sex
Authors: Scott Naomi M. , Hodyl Nicolette A. , Murphy Vanessa E. , Osei-Kumah Annette , Wyper Hayley , Hodgson Deborah M. , Smith Roger , Clifton Vicki L.
Published: April 2014

In the presence of maternal asthma, we have previously reported reduced placental blood flow, decreased cortisol metabolism, and reductions in fetal growth in response to maternal asthma and asthma exacerbations. We have proposed that these changes in placental function and fetal development may be related to activation of proinflammatory pathways in the placenta in response to maternal asthma. In the present study, we examined the influence of maternal asthma severity, inhaled glucocorticoid treatment, maternal cigarette use, placental macrophage numbers, and fetal sex on placental cytokine mRNA expression from a prospective cohort study of pregnant women with and without asthma. Placental expression of TNF-α, IL-1β, IL-6, IL-8, and IL-5 mRNA were all increased significantly in placentae of female fetuses whose mothers had mild asthma, but no changes were observed in placentae of male fetuses. The proinflammatory cytokines TNF-α, IL-1β, and IL-6 were negatively correlated with female cord blood cortisol, but there were no such correlations in placentae from males. Multivariate analysis indicated the strongest predictor of both cytokine mRNA expression in the placenta and birth weight was fetal cortisol but only in females. Placental cytokine mRNA levels were not significantly altered by inhaled glucocorticoid use, placental macrophage numbers, cigarette use, moderate-severe asthma, or male sex. These data suggest that placental basal cytokine mRNA expression is sex specifically regulated in pregnancies complicated by asthma, and interestingly these changes are more prevalent in mild rather than severe asthma.
Endogenously Produced IL-4 Nonredundantly Stimulates CD8+ T Cell Proliferation
Authors: Morris Suzanne C. , Heidorn Stephanie M. , Herbert De'Broski R. , Perkins Charles , Hildeman David A. , Khodoun Marat V. , Finkelman Fred D.
Published: April 2014

T cell proliferation and survival are regulated by the cytokine receptor common γ-chain-associated cytokines IL-2, IL-7, and IL-15, while IL-4, another γ-chain-associated cytokine, is thought to primarily affect T cell quality rather than quantity. In contrast, our experiments reveal that endogenously produced IL-4 is a direct, nonredundant, and potent stimulator of CD8+ T cell proliferation in Ag- and pathogen-induced CD8+ T cell responses. These stimulatory effects of IL-4 are observed in both BALB/c and C57BL/6 mice and activate both naive and memory/activated phenotype CD8+ T cells, although the former are stimulated less than are the latter. IL-4 effects are IL-7- and IL-15-independent, but MHC class I-dependent stimulation appears to be required for the mitogenic effect of IL-4 on naive phenotype CD8+ T cells. Thus, endogenously produced IL-4 is an important regulator of quantitative as well as qualitative aspects of T cell immunity.
B and T Lymphocyte Attenuator Regulates B Cell Receptor Signaling by Targeting Syk and BLNK
Authors: Vendel Andrew C. , Calemine-Fenaux Jill , Izrael-Tomasevic Anita , Chauhan Vandana , Arnott David , Eaton Dan L.
Published: April 2014

B and T lymphocyte attenuator (BTLA) functions as a negative regulator of T cell activation and proliferation. Although the role of BTLA in regulating T cell responses has been characterized, a thorough investigation into the precise molecular mechanisms involved in BTLA-mediated lymphocyte attenuation and, more specifically, its role in regulating B cell activation has not been presented. In this study, we have begun to elucidate the biochemical mechanisms by which BTLA functions to inhibit B cell activation. We describe the cell surface expression of BTLA on various human B cell subsets and confirm its ability to attenuate B cell proliferation upon associating with its known ligand, herpesvirus entry mediator (HVEM). BTLA associates with the BCR and, upon binding to HVEM, recruits the tyrosine phosphatase Src homology 2 domain-containing phosphatase 1 and reduces activation of signaling molecules downstream of the BCR. This is exemplified by a quantifiable decrease in tyrosine phosphorylation of the protein tyrosine kinase Syk, as measured by absolute quantification mass spectrometry. Furthermore, effector molecules downstream of BCR signaling, including the B cell linker protein, phospholipase Cγ2, and NF-κB, display decreased activation and nuclear translocation, respectively, after BTLA activation by HVEM. These results begin to provide insight into the mechanism by which BTLA negatively regulates B cell activation and indicates that BTLA is an inhibitory coreceptor of the BCR signaling pathway and attenuates B cell activation by targeting the downstream signaling molecules Syk and B cell linker protein.
Complement Inhibitor C4b-Binding Protein Interacts Directly with Small Glycoproteins of the Extracellular Matrix
Authors: Happonen Kaisa E. , Sjöberg Andreas P. , Mörgelin Matthias , Heinegård Dick , Blom Anna M.
Published: April 2014

Components derived from cartilage have been suggested to maintain the inflammation in joints in arthritis. Small leucine-rich repeat proteins (SLRPs) are structural components of cartilage important in organizing the meshwork of extracellular matrix components. It has recently been shown that the SLRP fibromodulin interacts with complement initiator C1q, leading to complement activation. The complement response is limited since fibromodulin also interacts with the complement inhibitor factor H. We have now found that osteoadherin, chondroadherin, fibromodulin, and proline arginine-rich end leucine-rich repeat protein bind to the complement inhibitor C4b-binding protein (C4BP). Using direct binding assays with C4BP fragments and C4BP mutants lacking individual domains in combination with electron microscopy, we have demonstrated that mainly the central core of C4BP mediated binding to SLRPs. Binding of SLRPs to C4BP did not affect its ability to inhibit complement. Osteoadherin, fibromodulin, and chondroadherin, which bind C1q and activate complement, were found to cause significantly higher C9 deposition in C4BP-depleted serum compared with Igs, indicating that the level of complement activation initiated by SLRPs is regulated by simultaneous binding to C4BP. A similar dual binding of C1q and complement inhibitors was observed previously for other endogenous ligands (amyloid, prions, C-reactive protein, and apoptotic cells) but not for exogenous activators (bacteria-bound Igs). These interactions can be significant during inflammatory joint diseases, such as rheumatoid arthritis, where cartilage is degraded, and cartilage components are released into synovial fluid, where they can interact with factors of the complement system.
A Comparative Study of HLA Binding Affinity and Ligand Diversity: Implications for Generating Immunodominant CD8+ T Cell Responses
Authors: Rao Xiangyu , Fontaine Costa Ana Isabel C. A. , van Baarle Debbie , Keşmir Can
Published: April 2014

Conventional CD8+ T cell responses against intracellular infectious agents are initiated upon recognition of pathogen-derived peptides presented at the cell surface of infected cells in the context of MHC class I molecules. Among the major MHC class I loci, HLA-B is the swiftest evolving and the most polymorphic locus. Additionally, responses restricted by HLA-B molecules tend to be dominant, and most associations with susceptibility or protection against infectious diseases have been assigned to HLA-B alleles. To assess whether the differences in responses mediated via two major HLA class I loci, HLA-B and HLA-A, may already begin at the Ag presentation level, we have analyzed the diversity and binding affinity of their peptide repertoire by making use of curated pathogen-derived epitope data retrieved from the Immune Epitope Database and Analysis Resource, as well as in silico predicted epitopes. In contrast to our expectations, HLA-B alleles were found to have a less diverse peptide repertoire, which points toward a more restricted binding motif, and the respective average peptide binding affinity was shown to be lower than that of HLA-A-restricted epitopes. This unexpected observation gives rise to new hypotheses concerning the mechanisms underlying immunodominance of CD8+ T cell responses.
Soluble IL-2RA Levels in Multiple Sclerosis Subjects and the Effect of Soluble IL-2RA on Immune Responses
Authors: Maier Lisa M. , Anderson David E. , Severson Christopher A. , Baecher-Allan Clare , Healy Brian , Liu David V. , Wittrup K. Dane , De Jager Philip L. , Hafler David A.
Published: April 2014

Multiple sclerosis (MS) is an organ-specific autoimmune disorder that is in part genetically determined. The gene encoding the α-chain of the IL-2 receptor, IL2RA, harbors alleles associated with risk to MS and other autoimmune diseases. In addition, IL2RA genetic variants correlate with the levels of a soluble form of the IL-2 receptor in subjects with type 1 diabetes and multiple sclerosis. Here, we show that the IL2RA genotypes differentially affects soluble IL-2RA (sIL-2RA) levels in MS cases vs healthy controls; the two variants associated with MS (rs12722489 and rs2104286) account for 15 and 18% of the total variance in log10-transformed sIL-2RA concentration in control subjects but less so in subjects with MS (2 and 5%), suggesting that perturbations associated with disease or treatment may influence sIL-2RA levels in subjects with MS. Whereas analyses demonstrate that sIL-2RA serum concentrations are a remarkably stable phenotype in both healthy controls and untreated MS subjects, a difference is observed between benign and malignant MS. These data indicate that, in addition to specific allelic variants at IL2RA, immunological perturbations associated with aggressive forms of the disease can influence sIL-2RA levels in serum of MS subjects. We also demonstrate, functionally, that sIL-2RA can inhibit IL-2 signaling, yet enhance T cell proliferation and expansion. In summary, we propose that before disease onset, strong genetic factors associated with disease risk dictate sIL-2RA levels that may be further modulated with onset of chronic systemic inflammation associated with MS.