bluuebunny/crossref_metadata_2025 huggingface dataset:
Requisite H2k Role in NK Cell-Mediated Resistance in Acute Murine Cytomegalovirus-Infected MA/My Mice
Authors: Dighe Abhijit , Rodriguez Marisela , Sabastian Pearl , Xie Xuefang , McVoy Michael , Brown Michael G.
Published: April 2014

Human CMV infections are a major health risk in patients with dysfunctional or compromised immunity, especially in patients with NK cell deficiencies, as these are frequently associated with high morbidity and mortality. In experimental murine CMV (MCMV) infections, Ly49H activation receptors on C57BL/6 (B6) NK cells engage m157 viral ligands on MCMV-infected cells and initiate dominant virus control. In this study, we report that MCMV resistance in MA/My relies on Ly49H-independent NK cell-mediated control of MCMV infection as NK cells in these mice do not bind anti-Ly49H mAb or soluble m157 viral ligands. We genetically compared MA/My resistance with MCMV susceptibility in genealogically and NK gene complex-Ly49 haplotype-related C57L mice. We found that MCMV resistance strongly associated with polymorphic H2k-linked genes, including MHC and non-MHC locations by analysis of backcross and intercross progeny. The H2b haplotype most frequently, but not absolutely, correlated with MCMV susceptibility, thus confirming a role for non-MHC genes in MCMV control. We also demonstrate a definite role for NK cells in H2k-type MCMV resistance because their removal from C57L.M-H2k mice before MCMV infection diminished immunity. NK gene complex-linked polymorphisms, however, did not significantly influence MCMV control. Taken together, effective NK cell-mediated MCMV control in this genetic system required polymorphic H2k genes without need of Ly49H-m157 interactions.
Antibodies and CD8+ T Cells Are Complementary and Essential for Natural Resistance to a Highly Lethal Cytopathic Virus
Authors: Fang Min , Sigal Luis J.
Published: April 2014

It is believed that CD8+ T lymphocytes or Abs can independently clear many primary viral infections, including those caused by Orthopoxviruses (OPV), a genus that includes the human pathogens variola and monkeypox and the vaccine species vaccinia virus. However, most experiments addressing the role of Abs and CD8+ T cells in protection have used viruses that are not specific for the host. In the present study, we used the mouse-specific OPV ectromelia virus and mice deficient in CD40, B cells, or CD8+ T cells and adoptive transfers of CD8+ T or B lymphocytes to show that the protection afforded by CD8+ T cells is incomplete. Despite sustained CD8+ T cell responses, in the absence of Ab responses ectromelia virus persists. This results in delayed disease and inexorably leads to death. Therefore, CD8+ T lymphocytes and Abs are not redundant but complementary and essential to survive infections with a highly pathogenic viruses in the natural host.
Increased Susceptibility to Apoptosis of CD56dimCD16+ NK Cells Induces the Enrichment of IFN-γ-Producing CD56bright Cells in Tuberculous Pleurisy
Authors: Schierloh Pablo , Yokobori Noemí , Alemán Mercedes , Musella Rosa M. , Beigier-Bompadre Macarena , Saab María A. , Alves Leandro , Abbate Eduardo , de la Barrera Silvia S. , Sasiain María C.
Published: April 2014

Tuberculous pleuritis is a good model for the study of specific cells at the site of active Mycobacterium tuberculosis (Mtb) infection. We investigated the frequency and phenotype of NK cells in paired samples of peripheral blood and pleural fluid (PF) from patients with tuberculosis (TB) or parapneumonic infection. We demonstrated for the first time a reduction of NK cells in PF from TB with an enrichment in the CD56brightCD16− subset. In agreement, in PF NK cells we observed an increased expression of CD94, NKG2A, CD62L, and CCR7 molecules and lower expression of Bcl-2 and perforin. The activation markers CD69 and HLA-DR were also increased. The enrichment in the CD56bright subset was due to an increased susceptibility to apoptosis of CD56+CD16+ NK cells mediated by heat-labile and stable soluble factors present in tuberculous effusions and not in PF from other etiologies. Furthermore, in TB patients, Mtb-induced IFN-γ production by PF NK cells was not dependent on the presence of CD3+, CD19+, and CD14+ cells, suggesting a direct interaction of CD56bright cells with Mtb and/or the involvement of other accessory cells present at the site of Mtb infection.
The Mechanism of Superantigen-Mediated Toxic Shock: Not a Simple Th1 Cytokine Storm
Authors: Faulkner Lee , Cooper Anneli , Fantino Cristina , Altmann Daniel M. , Sriskandan Shiranee
Published: April 2014

The profound clinical consequences of Gram-positive toxic shock are hypothesized to stem from excessive Th1 responses to superantigens. We used a new superantigen-sensitive transgenic model to explore the role of TCRαβ T cells in responses to staphylococcal enterotoxin B (SEB) in vitro and in two different in vivo models. The proliferative and cytokine responses of HLA-DR1 spleen cells were 100-fold more sensitive than controls and were entirely dependent on TCRαβ T cells. HLA-DR1 mice showed greater sensitivity in vivo to two doses of SEB with higher mortality and serum cytokines than controls. When d-galactosamine was used as a sensitizing agent with a single dose of SEB, HLA-DR1 mice died of toxic shock whereas controls did not. In this sensitized model of toxic shock there was a biphasic release of cytokines, including TNF-α, at 2 h and before death at 7 h. In both models, mortality and cytokine release at both time points were dependent on TCRαβ T cells. Anti-TNF-α pretreatment was protective against shock whereas anti-IFN γ pretreatment and delayed anti-TNF-α treatment were not. Importantly, anti-TNF-α pretreatment inhibited the early TNF-α response but did not inhibit the later TNF-α burst, to which mortality has previously been attributed. Splenic T cells were shown definitively to be the major source of TNF-α during the acute cytokine response. Our results demonstrate unequivocally that TCRαβ T cells are critical for lethality in toxic shock but it is the early TNF-α response and not the later cytokine surge that mediates lethal shock.
Beryllium Presentation to CD4+ T Cells Is Dependent on a Single Amino Acid Residue of the MHC Class II β-Chain
Authors: Bill Jerome R. , Mack Douglas G. , Falta Michael T. , Maier Lisa A. , Sullivan Andrew K. , Joslin Fenneke G. , Martin Allison K. , Freed Brian M. , Kotzin Brian L. , Fontenot Andrew P.
Published: April 2014

Chronic beryllium disease (CBD) is characterized by a CD4+ T cell alveolitis and granulomatous inflammation in the lung. Genetic susceptibility to this disease has been linked with HLA-DP alleles, particularly those possessing a glutamic acid at position 69 (Glu69) of the β-chain. However, 15% of CBD patients do not possess a Glu69-containing HLA-DP allele, suggesting that other MHC class II alleles may be involved in disease susceptibility. In CBD patients without a Glu69-containing HLA-DP allele, an increased frequency of HLA-DR13 alleles has been described, and these alleles possess a glutamic acid at position 71 of the β-chain (which corresponds to position 69 of HLA-DP). Thus, we hypothesized that beryllium presentation to CD4+ T cells was dependent on a glutamic acid residue at the identical position of both HLA-DP and -DR. The results show that HLA-DP Glu69- and HLA-DR Glu71-expressing molecules are capable of inducing beryllium-specific proliferation and IFN-γ expression by lung CD4+ T cells. Using fibroblasts expressing mutated HLA-DP2 and -DR13 molecules, beryllium recognition was dependent on the glutamic acid at position 69 of HLA-DP and 71 of HLA-DR, suggesting a critical role for this amino acid in beryllium presentation to Ag-specific CD4+ T cells. Thus, these results demonstrate that a single amino acid residue of the MHC class II β-chain dictates beryllium presentation and potentially, disease susceptibility.
Gliadin Fragments Induce Phenotypic and Functional Maturation of Human Dendritic Cells
Authors: Palová-Jelínková Lenka , Rožková Daniela , Pecharová Barbara , Bártová Jiřina , Šedivá Anna , Tlaskalová-Hogenová Helena , Spíšek Radek , Tučková Ludmila
Published: April 2014

Celiac disease is a chronic inflammatory disease developing in genetically predisposed individuals. Ingested gliadin, the triggering agent of the disease, can cross the epithelial barrier and elicit a harmful T cell-mediated immune response. Dendritic cells (DC) are supposed to play a pivotal role in shaping the immune response. The direction of the immune response toward immunity or tolerance depends on the stage of maturation and the functional properties of the DC. DC become fully functional APC upon maturation by various stimuli. We investigated the effect of a peptic digest of gliadin on the maturation of human monocyte-derived DC. Stimulation of cells with gliadin, in contrast with other tested food proteins, led to enhanced expression of maturation markers (CD80, CD83, CD86, and HLA-DR molecules) and increased secretion of chemokines and cytokines (mainly of IL-6, IL-8, IL-10, TNF-α, growth-related oncogene, MCP-1, MCP-2, macrophage-derived chemokine, and RANTES). Maturation was accompanied by a greater capacity to stimulate proliferation of allogeneic T cells and significantly reduced endocytic activity. Furthermore, gliadin-induced phosphorylation of members of three MAPK families (ERK1/2, JNK, and p38 MAPK) was demonstrated. The largest contribution of p38 MAPK was confirmed using its inhibitor SB203580, which markedly down-regulated the gliadin-triggered up-regulation of maturation markers and cytokine production. Gliadin treatment also resulted in increased NF-κB/DNA binding activity of p50 and p65 subunits. Taken together, gliadin peptides can contribute to overcoming the stage of unresponsiveness of immature DC by inducing phenotypic and functional DC maturation, resulting in more efficient processing and presentation of gliadin peptides to specific T lymphocytes.
Cutting Edge: CD8+CD122+ Regulatory T Cells Produce IL-10 to Suppress IFN-γ Production and Proliferation of CD8+ T Cells
Authors: Endharti Agustina Tri , Rifa’ Muhaimin , Shi Zhe , Fukuoka Yukari , Nakahara Yoshio , Kawamoto Yoshiyuki , Takeda Kozue , Isobe Ken-ichi , Suzuki Haruhiko
Published: April 2014

We recently identified CD8+CD122+ regulatory T cells that directly control CD8+ and CD4+ cells without intervention of APCs. In this study, we investigated the effector mechanism of CD8+CD122+ regulatory T cells by using an in vitro regulation system. The profile of cytokine expression revealed that IL-10 was predominantly produced by CD8+CD122+ cells, whereas other cytokines were similarly expressed in CD8+CD122+ cells and CD8+CD122− cells. Suppression of both proliferation and IFN-γ production by CD8+CD122− cells by CD8+CD122+ cells was blocked by adding anti-IL-10 Ab to the culture but not by adding anti-TGF-β Ab. When IL-10 was removed from the conditioned medium from CD8+CD122+ cells, the conditioned medium no longer showed regulatory activity. Finally, CD8+CD122+ cells from IL-10-deficient mice had no regulatory activity in vitro and reduced regulatory activity in vivo. Our results clearly indicate that IL-10 is produced by CD8+CD122+ cells and mediates the regulatory activity of these cells.
Cutting Edge: The Development of IL-4-Producing B Cells (B Effector 2 Cells) Is Controlled by IL-4, IL-4 Receptor α, and Th2 Cells
Authors: Harris David P. , Goodrich Stephen , Mohrs Katja , Mohrs Markus , Lund Frances E.
Published: April 2014

Although IL-4-producing B cells (B effector 2 cells) are found following infection and immunization, the signals regulating IL-4 production by Be2 cells are unknown. We show that culturing naive B cells with Th2 cells induces up-regulation of IL-4 in the B cells with a concomitant down-regulation of T-bet, IL-12Rβ2, and IFN-γ. Up-regulation of IL-4 in the Be2 cells is dependent on both T cells and IL-4 as IL-4Rα-deficient B cells primed with Th2 cells did not transcribe IL-4, and B cells primed in the presence of IL-4-deficient Th2 cells produced IFN-γ instead of IL-4. Likewise, the in vivo development of IL-4-expressing B cells in a nematode infection model was dependent on both T cells and IL-4Rα-mediated signals. Thus, the differentiation of naive B cells into IL-4-expressing Be2 cells is regulated by a combination of T cell-dependent signals and the cytokine environment and this process is critically dependent upon the IL-4/IL-4R signaling pathway.
CXCR5-Dependent Seeding of Follicular Niches by B and Th Cells Augments Antiviral B Cell Responses
Authors: Junt Tobias , Fink Katja , Förster Reinhold , Senn Beatrice , Lipp Martin , Muramatsu Masamichi , Zinkernagel Rolf M. , Ludewig Burkhard , Hengartner Hans
Published: April 2014

The chemokine receptor CXCR5 and its ligand CXCL13 define the structure of B cell follicles within secondary lymphoid organs. Here, we examined the impact of CXCR5 on antiviral B cell responses in vivo. CXCR5−/− mice showed a normal production of IgM and IgG acutely after infection with vesicular stomatitis virus (VSV) and developed VSV-specific germinal centers. However, impaired Ig class switch and Ab production were observed under conditions of limited availability of Ag (i.e., after immunization with nonreplicating viral particles or soluble Ag). Adoptive transfer of CXCR5-deficient, VSV-specific B and Th cells demonstrated that CXCR5 expression on both B and Th cells is required for an efficient Ig class switch. These experiments revealed that CXCR5 is critical for the coordinated interaction of antiviral T and B cells through its impact on initial B cell expansion and the recruitment of Ag-specific B and Th cells to germinal centers.
CD4+CD25+ T Cells Prevent the Development of Organ-Specific Autoimmune Disease by Inhibiting the Differentiation of Autoreactive Effector T Cells
Authors: DiPaolo Richard J. , Glass Deborah D. , Bijwaard Karen E. , Shevach Ethan M.
Published: April 2014

Thymic-derived, naturally occurring, CD4+CD25+ regulatory T cells (nTreg) are potent suppressors of immune responses. A detailed understanding of which components of the development and activation of pathogenic effector T cells are inhibited by nTreg during the course of T cell-mediated, organ-specific autoimmunity is as yet unknown. We have analyzed the effects of polyclonal nTreg on the development of autoimmune gastritis. The nTreg inhibited the development of disease, but failed to inhibit the migration of effector cells into the gastric lymph node or stomach. Notably, nTreg did not inhibit the expansion of autoreactive T cells in the gastric lymph node. The primary effect of nTreg appeared to be inhibition of differentiation of autoantigen-specific T cells to Th1 effector cells, as reflected by a decrease in Ag-stimulated IFN-γ production and a reduction in T-bet expression.
An Hour after Immunization Peritoneal B-1 Cells Are Activated to Migrate to Lymphoid Organs Where within 1 Day They Produce IgM Antibodies That Initiate Elicitation of Contact Sensitivity
Authors: Itakura Atsuko , Szczepanik Marian , Campos Regis A. , Paliwal Vipin , Majewska Monika , Matsuda Hiroshi , Takatsu Kiyoshi , Askenase Philip W.
Published: April 2014

Elicitation of contact sensitivity (CS), a classic example of T cell-mediated immunity, requires Ag-specific IgM Abs to trigger an initiation process. This early process leads to local recruitment of CS-effector T cells after secondary Ag challenge. These Abs are produced by the B-1 subset of B cells within 1 day after primary skin immunization. In this study we report the surprising observation that B-1 cells in the peritoneal cavity are activated as early as 1 h after naive mice are painted with a contact-sensitizing Ag on the skin of the trunk and feet to begin the initiation of CS. B-1 cells in the spleen and draining lymph nodes produce the initiating Abs by 1 day after immunization, when we found increased numbers of Ag-specific IgM Ab-producing cells in these tissues by ELISPOT assay. Importantly, we show that contact-activated peritoneal B-1 cells migrate to these lymphoid tissues and then differentiate into Ag-specific IgM Ab-producing cells, resulting in specific CS-initiating IgM Abs in the serum by 1 day. Furthermore, pertussis toxin, which is known to inhibit signaling via G protein-coupled chemokines, inhibited the migration of contact-activated peritoneal B-1 cells to the lymphoid tissues, probably due to BLR-1 (Burkitt lymphoma receptor-1). These findings indicate that within 1 h after contact skin immunization, B-1 cells in the peritoneal cavity are activated to migrate to the lymphoid tissues by chemokine-dependent mechanisms to produce serum Ag-specific IgM Abs within 1 day after immunization, leading to local recruitment of CS-effector T cells.
Interaction of Fas Ligand and Fas Expressed on Osteoclast Precursors Increases Osteoclastogenesis
Authors: Park Hyewon , Jung Youn-Kwan , Park Ok-Jin , Lee Yeon Ju , Choi Je-Yong , Choi Youngnim
Published: April 2014

We incidentally found that osteoclast precursors and mature osteoclasts express Fas ligand (FasL) as well as Fas, which was confirmed by flow cytometry, immunofluorescent staining, and RT-PCR. The aim of this study was to determine the role of FasL in differentiation and cell death of osteoclasts. To study the role of FasL in osteoclastogenesis, neutralizing anti-FasL mAb or rFasL was added during receptor activator of NF-κB ligand (RANKL)-induced osteoclastogenesis using bone marrow-derived macrophages. Neutralization of endogenous FasL by anti-FasL mAb decreased osteoclastogenesis, whereas rFasL enhanced osteoclast differentiation in a dose-dependent manner. In addition, rFasL up-regulated the secretion of osteoclastogenic cytokines, such as IL-1β and TNF-α, and the activation of NF-κB. Functional blocking of IL-1β and TNF-α using IL-1 receptor antagonist and soluble TNFR confirmed that those cytokines mediated the effect of FasL on osteoclastogenesis. The osteoclast precursors were relatively resistant to rFasL-induced apoptosis especially before RANKL treatment, resulting in minimal cell loss by rFasL treatment during osteoclastogenesis. Although rFasL increased the cell death of mature osteoclasts, growth factor withdrawal induced much more cell death. However, anti-FasL mAb did not affect the survival of mature osteoclasts, suggesting that the endogenous FasL does not have a role in the apoptosis of osteoclasts. Finally, in contrast to the effect on apoptosis, rFasL-assisted osteoclastogenesis was not mediated by caspases. In conclusion, FasL has a novel function in bone homeostasis by enhancing the differentiation of osteoclasts, which was not considered previously.
Identification of the Immunodominant HY H2-Dk Epitope and Evaluation of the Role of Direct and Indirect Antigen Presentation in HY Responses
Authors: Millrain Maggie , Scott Diane , Addey Caroline , Dewchand Hamlata , Ellis Pamela , Ehrmann Ingrid , Mitchell Michael , Burgoyne Paul , Simpson Elizabeth , Dyson Julian
Published: April 2014

Minor histocompatibility Ags derive from self-proteins and provoke allograft rejection and graft-vs-host disease in MHC-matched donor-recipient combinations. In this study, we define the HYDk epitope of the HY minor histocompatibility Ag as the 8mer peptide RRLRKTLL derived from the Smcy gene. Using HY tetramers, the response to this peptide was found to be immunodominant among the four characterized MHC class I-restricted HY epitopes (HYDkSmcy (defined here), HYKkSmcy, HYDbUty, and HYDbSmcy). Indirect presentation stimulated a robust primary HYDkSmcy response. Indirect presentation and priming of HY-specific CD8+ T cells is also operative in the presence of a full MHC mismatch. To determine whether the indirect route of Ag presentation is required for HY priming, female parent into F1 (H2bxk) female recipient bone marrow chimeras were immunized with male cells of the other parental haplotype, limiting presentation to the direct pathway. The dominant H2b HY response (HYDbUty) was dependent on indirect presentation. However, the dominant H2k HY response (HYDkSmcy) could be stimulated efficiently by the direct pathway. In contrast, secondary expansion of both HYDkSmcy and HYDbUty-specific CD8+ T cells was effective only when Ag was presented by the direct route. Transgenic overproduction of Smcy mRNA within the immunizing cells resulted in a corresponding increase in the HYDkSmcy, HYDbSmcy, and HYKkSmcy-specific CD8+ T cell responses when presented via the direct pathway but did not enhance indirect presentation demonstrating the independent regulation of MHC class I-peptide occupancy in the two Ag-processing pathways.
A Role for Double-Stranded RNA-Activated Protein Kinase PKR in Mycobacterium-Induced Cytokine Expression
Authors: Cheung Benny K. W. , Lee Davy C. W. , Li James C. B. , Lau Yu-Lung , Lau Allan S. Y.
Published: April 2014

Following infection of the host by Mycobacterium tuberculosis, induction of cytokines is a major defense mechanism to limit the pathogen invasion. Cytokines interact with each other to form an intertwined network of pathways. For example, IFN and TNF have been shown to interact through common pathways including IFN-inducible, dsRNA-activated serine/threonine protein kinase (PKR) induction. As a signal transducer, it has been conventionally known to regulate the induction of cytokine expression in response to virus infection through NF-κB. In light of the critical role of TNF in immunity and its cytotoxic effects mediated by PKR, we examined the role of the kinase in the regulation of immune response against M. tuberculosis using the interaction of bacillus Calmette-Guérin (BCG) and primary human blood monocytes as a model. Our results showed that BCG stimulates the induction of cytokine expression in human primary blood monocytes including TNF-α, IL-6, and IL-10. With the suppression of PKR by using PKR-mutant gene or 2-aminopurine as PKR inhibitor, we showed that the BCG-induced cytokine expression in human monocytes is regulated by the phosphorylation and activation of PKR. We also demonstrated that downstream of PKR induction is the activation of MAPK and translocation of NF-κB into the nucleus. NF-κB in turn mediates the transcription of specific cytokine genes. Taken together, PKR plays a critical role in the regulation of immune responses to mycobacterial infection and may serve as an important molecule in the innate antimycobacterial defense.
Primary Human T Lymphocytes Engineered with a Codon-Optimized IL-15 Gene Resist Cytokine Withdrawal-Induced Apoptosis and Persist Long-Term in the Absence of Exogenous Cytokine
Authors: Hsu Cary , Hughes Marybeth S. , Zheng Zhili , Bray Regina B. , Rosenberg Steven A. , Morgan Richard A.
Published: April 2014

IL-15 is a common γ-chain cytokine that has been shown to be more active than IL-2 in several murine cancer immunotherapy models. Although T lymphocytes do not produce IL-15, murine lymphocytes carrying an IL-15 transgene demonstrated superior antitumor activity in the immunotherapy of B16 melanoma. Thus, we sought to investigate the biological impact of constitutive IL-15 expression by human lymphocytes. In this report we describe the generation of a retroviral vector encoding a codon-optimized IL-15 gene. Alternate codon usage significantly enhanced the translational efficiency of this tightly regulated gene in retroviral vector-transduced cells. Activated human CD4+ and CD8+ human lymphocytes expressed IL-15Rα and produced high levels of cytokine upon retroviral transduction with the IL-15 vector. IL-15-transduced lymphocytes remained viable for up to 180 days in the absence of exogenous cytokine. IL-15 vector-transduced T cells showed continued proliferation after cytokine withdrawal and resistance to apoptosis while retaining specific Ag recognition. In the setting of adoptive cell transfer, IL-15-transduced lymphocytes may prolong lymphocyte survival in vivo and could potentially enhance antitumor activity.
Plasmacytoid Dendritic Cells of Different Origins Have Distinct Characteristics and Function: Studies of Lymphoid Progenitors versus Myeloid Progenitors
Authors: Yang Guo-Xiang , Lian Zhe-Xiong , Kikuchi Kentaro , Moritoki Yuki , Ansari Aftab A. , Liu Yong-Jun , Ikehara Susumu , Gershwin M. Eric
Published: April 2014

Plasmacytoid dendritic cells (pDCs) play a central role in host innate and adaptive immunity and are thought to be of lymphoid origin. However, in IL-7Rα−/− mice, which are deficient in T and B lymphocytes, pDCs are still found in lymphoid organs, which suggests that there is a lymphoid-independent pathway for the development of pDCs. Previous work has demonstrated that pDCs originate from both lymphoid and myeloid progenitors (MPs). However, it is not clear whether the function of pDCs is different relative to their origin. In an effort to compare the characteristics and functions between pDCs generated from different progenitors, we performed adoptive transfer studies using highly enriched populations of common lymphoid progenitors (CLPs) and MPs from the bone marrow of control mice and examined their potential and developmental kinetics for the generation of pDCs. Interestingly, although CLPs were polarized to generate pDCs, MPs were polarized to generate conventional dendritic cells and the kinetics of pDC generation from MPs was reached earlier than from CLPs. Furthermore, CLPs have the potential to generate more pDCs on a per cell basis. Moreover, MP-derived pDCs produce relatively higher levels of IFN-α than CLP-derived pDCs following CpG stimulation. These data indicate that MPs are multipotential and have the capacity to develop into not only myeloid cells, but also pDCs, which have distinct characteristics and function compared to that of lymphoid origin and, therefore, imply a more important role for MP-derived pDCs in conditions where the function of lymphoid progenitors is impaired or compromised.
Cooperation between 4-1BB and ICOS in the Immune Response to Influenza Virus Revealed by Studies of CD28/ICOS-Deficient Mice
Authors: Vidric Mariana , Suh Woong-Kyung , Dianzani Umberto , Mak Tak W. , Watts Tania H.
Published: April 2014

CD28, ICOS, and 4-1BB each play distinct roles in the CD8 T cell response to influenza virus. CD28−/− mice are severely impaired in primary CD8 T cell expansion and fail to mount a secondary response to influenza. Influenza-specific CD8 T cells expand normally in ICOS−/− mice, with only a small and transient defect late in the primary response and an unimpaired secondary response. Conversely, 4-1BB/4-1BBL interaction is dispensable for the primary CD8 T cell response to influenza, but maintains CD8 T cell survival and controls the size of the secondary response. Previous results showed that a single dose of agonistic anti-4-1BB Ab at priming allowed partial restoration of primary CD8 T cell expansion and full recovery of the secondary CD8 T cell responses to influenza in CD28−/− mice. In this study we show that anti-4-1BB fails to correct the CD8 T cell defect in CD28−/−ICOS−/− mice, suggesting that ICOS partially compensates for CD28 in this model. In support of this hypothesis, we found that anti-4-1BB enhances ICOS expression on both T cell subsets and that anti-4-1BB and anti-ICOS can synergistically activate CD4 and CD8 T cells. Furthermore, ICOS and 4-1BB can cooperate to directly stimulate isolated CD28−/− CD8 T cells. These results reveal a novel interaction between the ICOS and 4-1BB costimulatory pathways as well as unexpected redundancy between CD28 and ICOS in primary CD8 T cell expansion. These findings have implications for costimulation of human T cell responses in diseases such as AIDS or rheumatoid arthritis, in which CD28− T cells accumulate.
The Neuropeptide Vasoactive Intestinal Peptide Generates Tolerogenic Dendritic Cells
Authors: Delgado Mario , Gonzalez-Rey Elena , Ganea Doina
Published: April 2014

Tolerogenic dendritic cells (DCs) play an important role in maintaining peripheral tolerance through the induction/activation of regulatory T cells (Treg). Endogenous factors contribute to the functional development of tolerogenic DCs. In this report, we present evidence that two known immunosuppressive neuropeptides, the vasoactive intestinal peptide (VIP) and the pituitary adenylate cyclase-activating polypeptide (PACAP), contribute to the development of bone marrow-derived tolerogenic DCs in vitro and in vivo. The VIP/PACAP-generated DCs are CD11clowCD45RBhigh, do not up-regulate CD80, CD86, and CD40 following LPS stimulation, and secrete high amounts of IL-10. The induction of tolerogenic DCs is mediated through the VPAC1 receptor and protein kinase A, and correlates with the inhibition of IκB phosphorylation and of NF-κBp65 nuclear translocation. The VIP/PACAP-generated DCs induce functional Treg in vitro and in vivo. The VIP/DC-induced Treg resemble the previously described Tr1 in terms of phenotype and cytokine profile, suppress primarily Th1 responses including delayed-type hypersensitivity, and transfer suppression to naive hosts. The effect of VIP/PACAP on the DC-Treg axis represents an additional mechanism for their general anti-inflammatory role, particularly in anatomical sites which exhibit immune deviation or privilege.
Murine and Human IL-7 Activate STAT5 and Induce Proliferation of Normal Human Pro-B Cells
Authors: Johnson Sonja E. , Shah Nisha , Panoskaltsis-Mortari Angela , LeBien Tucker W.
Published: April 2014

The role of IL-7 in lymphoid development and T cell homeostasis has been extensively documented. However, the role of IL-7 in human B cell development remains unclear. We used a xenogeneic human cord blood stem cell/murine stromal cell culture to study the development of CD19+ B-lineage cells expressing the IL-7R. CD34+ cord blood stem cells were cultured on the MS-5 murine stromal cell line supplemented with human G-CSF and stem cell factor. Following an initial expansion of myeloid/monocytoid cells within the initial 2 wk, CD19+/pre-BCR− pro-B cells emerged, of which 25–50% expressed the IL-7R. FACS-purified CD19+/IL-7R+ cells were larger and, when replated on MS-5, underwent a dose-dependent proliferative response to exogenous human IL-7 (0.01–10.0 ng/ml). Furthermore, STAT5 phosphorylation was induced by the same concentrations of human IL-7. CD19+/IL-7R− cells were smaller and did not proliferate on MS-5 after stimulation with IL-7. In a search for cytokines that promote human B cell development in the cord blood stem cell/MS-5 culture, we made the unexpected finding that murine IL-7 plays a role. Murine IL-7 was detected in MS-5 supernatants by ELISA, recombinant murine IL-7 induced STAT5 phosphorylation in CD19+/IL-7R+ pro-B cells and human B-lineage acute lymphoblastic leukemias, and neutralizing anti-murine IL-7 inhibited development of CD19+ cells in the cord blood stem cell/MS-5 culture. Our results support a model wherein IL-7 transduces a replicative signal to normal human B-lineage cells that is complemented by additional stromal cell-derived signals essential for normal human B cell development.
Bone Marrow-Derived Dendritic Cells Reverse the Anergic State of CD4+CD25+ T Cells without Reversing Their Suppressive Function
Authors: Brinster Carine , Shevach Ethan M.
Published: April 2014

Dendritic cells (DC) are potent inducers of immunity to foreign Ags, but also contribute to self-tolerance by induction of regulatory T cells or deletion/anergy of self-reactive T cells. In this study, we have studied the capacity of DC to activate naturally occurring CD4+CD25+ regulatory T cells as well as the ability of CD4+CD25+ T cells to suppress the DC-mediated activation of CD4+CD25− T cells. Mature bone marrow-derived dendritic cells, but not splenic DC, were able to induce the proliferation of CD4+CD25+ T cells in the presence of a polyclonal stimulus and in the absence of exogenous IL-2. The DC-induced proliferative response of the CD4+CD25+ T cells was partially dependent on IL-2 produced by a small number of contaminating CD25+ effector cells. Because bone marrow-derived dendritic cells induce proliferation of both CD4+CD25+ and CD4+CD25− T cells in vitro, it was impossible to assay the suppressive function of the CD4+CD25+ T cells using [3H]TdR uptake or CFSE dilution. We therefore measured IL-2 production in cocultures of CD4+CD25+ and CD4+CD25− T cells using the IL-2 secretion assay. Surprisingly, CD4+CD25+ T cells markedly suppressed IL-2 secretion by the CD4+CD25− T cells without inhibiting their proliferation. Collectively, these results suggest that Ag presentation by DC can induce the expansion of CD4+CD25+ T cells while simultaneously activating their ability to suppress cytokine secretion by effector T cells.
The ICOS Molecule Plays a Crucial Role in the Development of Mucosal Tolerance
Authors: Miyamoto Katsuichi , Kingsley Cherry I. , Zhang Xingmin , Jabs Claudia , Izikson Leonid , Sobel Raymond A. , Weiner Howard L. , Kuchroo Vijay K. , Sharpe Arlene H.
Published: April 2014

The ICOS molecule stimulates production of the immunoregulatory cytokine IL-10, suggesting an important role for ICOS in controlling IL-10-producing regulatory T cells and peripheral T cell tolerance. In this study we investigate whether ICOS is required for development of oral, nasal, and high dose i.v. tolerance. Oral administration of encephalitogenic myelin oligodendrocyte glycoprotein (MOG) 35–55 peptide to ICOS-deficient (ICOS−/−) mice did not inhibit experimental autoimmune encephalomyelitis (EAE), T cell proliferation, or IFN-γ production, in striking contrast to wild-type mice. Similarly, intranasal administration of MOG35–55 before EAE induction suppressed EAE and T cell responses in wild-type, but not in ICOS−/−, mice. In contrast, ICOS−/− mice were as susceptible as wild-type mice to high dose tolerance. These results indicate that ICOS plays an essential and specific role in mucosal tolerance and that distinct costimulatory pathways differentially regulate different forms of peripheral tolerance. Surprisingly, CD4+ cells from MOG-fed wild-type and ICOS−/− mice could transfer suppression to wild-type recipients, indicating that functional regulatory CD4+ cells can develop in the absence of ICOS. However, CD4+ T cells from MOG-fed wild-type mice could not transfer suppression to ICOS−/− recipients, suggesting that ICOS may have a key role in controlling the effector functions of regulatory T cells. These results suggest that stimulating ICOS may provide an effective therapeutic approach for promoting mucosal tolerance.
Respiratory Tolerance Is Inhibited by the Administration of Corticosteroids
Authors: Stock Philippe , Akbari Omid , DeKruyff Rosemarie H. , Umetsu Dale T.
Published: April 2014

Corticosteroids constitute the most effective current anti-inflammatory therapy for acute and chronic forms of allergic diseases and asthma. Corticosteroids are highly effective in inhibiting the effector function of Th2 cells, eosinophils, and epithelial cells. However, treatment with corticosteroids may also limit beneficial T cell responses, including respiratory tolerance and the development of regulatory T cells (TReg), which actively suppress inflammation in allergic diseases. To examine this possibility, we investigated the effects of corticosteroid administration on the development of respiratory tolerance. Respiratory exposure to Ag-induced T cell tolerance and prevented the subsequent development of allergen-induced airway hyperreactivity. However, treatment with dexamethasone during the delivery of respiratory Ag prevented tolerance, such that allergen sensitization and severe airway hyperreactivity subsequently occurred. Treatment with dexamethasone during respiratory exposure to allergen eliminated the development of IL-10-secreting dendritic cells, which was required for the induction of IL-10-producing allergen-specific TReg cells. Therefore, because allergen-specific TReg cells normally develop to prevent allergic disease and asthma, our results suggest that treatment with corticosteroids, which limit the development of TReg cells and tolerance to allergens, could enhance subsequent Th2 responses and aggravate the long-term course of allergic diseases and asthma.
Multiple TLRs Are Expressed in Human Cholangiocytes and Mediate Host Epithelial Defense Responses to Cryptosporidium parvum via Activation of NF-κB
Authors: Chen Xian-Ming , O’Hara Steven P. , Nelson Jeremy B. , Splinter Patrick L. , Small Aaron J. , Tietz Pamela S. , Limper Andrew H. , LaRusso Nicholas F.
Published: April 2014

Infection of epithelial cells by Cryptosporidium parvum triggers a variety of host-cell innate and adaptive immune responses including release of cytokines/chemokines and up-regulation of antimicrobial peptides. The mechanisms that trigger these host-cell responses are unclear. Thus, we evaluated the role of TLRs in host-cell responses during C. parvum infection of cultured human biliary epithelia (i.e., cholangiocytes). We found that normal human cholangiocytes express all known TLRs. C. parvum infection of cultured cholangiocytes induces the selective recruitment of TLR2 and TLR4 to the infection sites. Activation of several downstream effectors of TLRs including IL-1R-associated kinase, p-38, and NF-κB was detected in infected cells. Transfection of cholangiocytes with dominant-negative mutants of TLR2 and TLR4, as well as the adaptor molecule myeloid differentiation protein 88 (MyD88), inhibited C. parvum-induced activation of IL-1R-associated kinase, p-38, and NF-κB. Short-interfering RNA to TLR2, TLR4, and MyD88 also blocked C. parvum-induced NF-κB activation. Moreover, C. parvum selectively up-regulated human β-defensin-2 in directly infected cells, and inhibition of TLR2 and TLR4 signals or NF-κB activation were each associated with a reduction of C. parvum-induced human β-defensin-2 expression. A significantly higher number of parasites were detected in cells transfected with a MyD88 dominant-negative mutant than in the control cells at 48–96 h after initial exposure to parasites, suggesting MyD88-deficient cells were more susceptible to infection. These findings demonstrate that cholangiocytes express a variety of TLRs, and suggest that TLR2 and TLR4 mediate cholangiocyte defense responses to C. parvum via activation of NF-κB.
Molecular and Cellular Basis of the Retrovirus Resistance in I/LnJ Mice
Authors: Case Laure K. , Purdy Alexandra , Golovkina Tatyana V.
Published: April 2014

Previously, we showed that IFN-γ elicited by mouse mammary tumor virus (MMTV) infection in I/LnJ mice stimulated production of virus-neutralizing Abs, mostly of the IgG2a isotype. These Abs coated virions secreted by infected I/LnJ cells, and thus completely prevented virus transmission to offspring. However, the mechanism of virus neutralization by isotype-specific Abs remained unknown. Ab coating is capable of blocking virus infection by interfering with receptor-virus binding, by virus opsonization, by complement activation, and via FcγR-mediated effector mechanisms. The aim of the studies described in this work was to uncover the cellular basis of anti-virus Ab production, to evaluate the importance of the IgG2a subclass of IgGs in virus neutralization, and to investigate which of the blocking mechanisms plays a role in virus neutralization. We showed that I/LnJ-derived bone marrow cells, specifically IFN-γ-producing CD4+ T cells, were key cells conferring resistance to MMTV infection in susceptible mice upon transfer. We also established that a unique bias in the subclass selection toward the IgG2a isotype in infected I/LnJ mice was not due to their potent neutralizing ability, as anti-virus Abs of other isotypes were also able to neutralize the virus, but were a product of virally induced IFN-γ. Finally, we demonstrated that F(ab′)2 of anti-MMTV IgGs neutralized the virus as efficiently as total IgGs, suggesting that Ab-mediated interference with viral entry is the sole factor inhibiting virus replication in I/LnJ mice. We propose and discuss possible mechanisms by which infected I/LnJ mice eradicate retrovirus.
Human CD4+ T Cells Induced by Synthetic Peptide Malaria Vaccine Are Comparable to Cells Elicited by AttenuatedPlasmodium falciparumSporozoites
Authors: Calvo-Calle Jaime Mauricio , Oliveira Giane A. , Nardin Elizabeth H.
Published: April 2014

Peptide vaccines containing minimal epitopes of protective Ags provide the advantages of low cost, safety, and stability while focusing host responses on relevant targets of protective immunity. However, the limited complexity of malaria peptide vaccines raises questions regarding their equivalence to immune responses elicited by the irradiated sporozoite vaccine, the “gold standard” for protective immunity. A panel of CD4+ T cell clones was derived from volunteers immunized with a peptide vaccine containing minimal T and B cell epitopes of the Plasmodium falciparum circumsporozoite protein to compare these with previously defined CD4+ T cell clones from volunteers immunized with irradiated P. falciparum sporozoites. As found following sporozoite immunization, the majority of clones from the peptide-immunized volunteers recognized the T* epitope, a predicted universal T cell epitope, in the context of multiple HLA DR and DQ molecules. Peptide-induced T cell clones were of the Th0 subset, secreting high levels of IFN-γ as well as variable levels of Th2-type cytokines (IL-4, IL-6). The T* epitope overlaps a polymorphic region of the circumsporozoite protein and strain cross-reactivity of the peptide-induced clones correlated with recognition of core epitopes overlapping the conserved regions of the T* epitope. Importantly, as found following sporozoite immunization, long-lived CD4+ memory cells specific for the T* epitope were detectable 10 mo after peptide immunization. These studies demonstrate that malaria peptides containing minimal epitopes can elicit human CD4+ T cells with fine specificity and potential effector function comparable to those elicited by attenuated P. falciparum sporozoites.
Impaired FcεRI-Dependent Gene Expression and Defective Eicosanoid and Cytokine Production as a Consequence of Fyn Deficiency in Mast Cells
Authors: Gomez Gregorio , Gonzalez-Espinosa Claudia , Odom Sandra , Baez Gabriela , Cid M. Eugenia , Ryan John J. , Rivera Juan
Published: April 2014

Fyn kinase is a key contributor in coupling FcεRI to mast cell degranulation. A limited macroarray analysis of FcεRI-induced gene expression suggested potential defects in lipid metabolism, eicosanoid and glutathione metabolism, and cytokine production. Biochemical analysis of these responses revealed that Fyn-deficient mast cells failed to secrete the inflammatory eicosanoid products leukotrienes B4 and C4, the cytokines IL-6 and TNF, and chemokines CCL2 (MCP-1) and CCL4 (MIP-1β). FcεRI-induced generation of arachidonic acid and normal induction of cytokine mRNA were defective. Defects in JNK and p38 MAPK activation were observed, whereas ERK1/2 and cytosolic phospholipase A2 (S505) phosphorylation was normal. Pharmacological studies revealed that JNK activity was associated with generation of arachidonic acid. FcεRI-mediated activation of IκB kinase β and IκBα phosphorylation and degradation was defective resulting in a marked decrease of the nuclear NF-κB DNA binding activity that drives IL-6 and TNF production in mast cells. However, not all cytokine were affected, as IL-13 production and secretion was enhanced. These studies reveal a major positive role for Fyn kinase in multiple mast cell inflammatory responses and demonstrate a selective negative regulatory role for certain cytokines.
Inhibitory Effects of Chloride on the Activation of Caspase-1, IL-1β Secretion, and Cytolysis by the P2X7 Receptor
Authors: Verhoef Philip A. , Kertesy Sylvia B. , Lundberg Kathleen , Kahlenberg J. Michelle , Dubyak George R.
Published: April 2014

The P2X7 receptor (P2X7R) is an ATP-gated cation channel that activates caspase-1 leading to the maturation and secretion of IL-1β. Because previous studies indicated that extracellular Cl− exerts a negative allosteric effect on ATP-gating of P2X7R channels, we tested whether Cl− attenuates the P2X7R→caspase-1→IL-1β signaling cascade in murine and human macrophages. In Bac1 murine macrophages, substitution of extracellular Cl− with gluconate produced a 10-fold increase in the rate and extent of ATP-induced IL-1β processing and secretion, while reducing the EC50 for ATP by 5-fold. Replacement of Cl− with gluconate also increased the potency of ATP as an inducer of mature IL-1β secretion in primary mouse bone marrow-derived macrophages and in THP-1 human monocytes/macrophages. Our observations were consistent with actions of Cl− at three levels: 1) a negative allosteric effect of Cl−, which limits the ability of ATP to gate the P2X7R-mediated cation fluxes that trigger caspase-1 activation; 2) an intracellular accumulation of Cl− via nonselective pores induced by P2X7R with consequential repression of caspase-1-mediated processing of IL-1β; and 3) a facilitative effect of Cl− substitution on the cytolytic release of unprocessed pro-IL-1β that occurs with sustained activation of P2X7R. This cytolysis was repressed by the cytoprotectant glycine, permitting dissociation of P2X7R-regulated secretion of mature IL-1β from the lytic release of pro-IL-1β. These results suggest that under physiological conditions P2X7R are maintained in a conformationally restrained state that limits channel gating and coupling of the receptor to signaling pathways that regulate caspase-1.
Protein Kinase Cθ Controls Th1 Cells in Experimental Autoimmune Encephalomyelitis
Authors: Salek-Ardakani Shahram , So Takanori , Halteman Beth S. , Altman Amnon , Croft Michael
Published: April 2014

Molecules that regulate encephalitogenic T cells are of interest for multiple sclerosis. In this study we show that protein kinase Cθ (PKCθ) is critical for the development of Ag-specific Th1 cells in experimental allergic encephalomyelitis (EAE), a model of multiple sclerosis. PKCθ-deficient mice immunized with myelin oligodendrocyte glycoprotein failed to develop cell infiltrates and Th1 cytokines in the CNS and were resistant to the development of clinical EAE. CD4 T cells became primed and accumulated in secondary lymphoid organs in the absence of PKCθ, but had severely diminished IFN-γ, TNF, and IL-17 production. Increasing Ag exposure and inflammatory conditions failed to induce EAE in PKCθ-deficient mice, showing a profound defect in the myelin oligodendrocyte glycoprotein-reactive T cell population. These data provide evidence of a pivotal role for PKCθ in the generation and effector function of autoimmune Th1 cells.
Resistance of Collagenase-2 (Matrix Metalloproteinase-8)-Deficient Mice to TNF-Induced Lethal Hepatitis
Authors: Van Lint Philippe , Wielockx Ben , Puimège Leen , Noël Agnès , López-Otin Carlos , Libert Claude
Published: April 2014

Acute fulminant liver failure is a serious worldwide health problem. Despite maximal supportive intensive care treatment, the disease offers a poor prognosis, with mortality rates of >80%. We have previously shown that a broad-spectrum inhibitor of matrix metalloproteinases (MMPs) confers complete protection in a mouse model of TNF-induced lethal hepatitis, thereby suggesting the possibility of protecting cancer patients against the deleterious side effects of TNF therapy. In our search for the individual matrix metalloproteinases involved, we found that the recently generated MMP-8-deficient mice are significantly protected against TNF-induced acute hepatitis. In contrast to their wild-type counterparts, MMP-8-null mice display very little hepatocyte necrosis and apoptosis, resulting in a much better survival outcome. We found that these animals clearly display impaired leukocyte influx into the liver and no release of the neutrophil-specific, LPS-induced CXC chemokine. Our findings provide evidence that MMP-8 plays an essential role in acute liver failure and might be a promising new target for the treatment for this illness.