bluuebunny/crossref_metadata_2025 huggingface dataset:
Allergen-Specific Th1 Cells Counteract Efferent Th2 Cell-Dependent Bronchial Hyperresponsiveness and Eosinophilic Inflammation Partly Via IFN-γ
Authors: Huang Tung-Jung , MacAry Paul A. , Eynott Paul , Moussavi Artin , Daniel Kenn C. , Askenase Philip W. , Kemeny David M. , Chung K. Fan
Published: April 2014

Th2 T cell immune-driven inflammation plays an important role in allergic asthma. We studied the effect of counterbalancing Th1 T cells in an asthma model in Brown Norway rats that favors Th2 responses. Rats received i.v. transfers of syngeneic allergen-specific Th1 or Th2 cells, 24 h before aerosol exposure to allergen, and were studied 18–24 h later. Adoptive transfer of OVA-specific Th2 cells, but not Th1 cells, and OVA, but not BSA exposure, induced bronchial hyperresponsiveness (BHR) to acetylcholine and eosinophilia in a cell number-dependent manner. Importantly, cotransfer of OVA-specific Th1 cells dose-dependently reversed BHR and bronchoalveolar lavage (BAL) eosinophilia, but not mucosal eosinophilia. OVA-specific Th1 cells transferred alone induced mucosal eosinophilia, but neither BHR nor BAL eosinophilia. Th1 suppression of BHR and BAL eosinophilia was allergen specific, since cotransfer of BSA-specific Th1 cells with the OVA-specific Th2 cells was not inhibitory when OVA aerosol alone was used, but was suppressive with OVA and BSA challenge. Furthermore, recipients of Th1 cells alone had increased gene expression for IFN-γ in the lungs, while those receiving Th2 cells alone showed increased IL-4 mRNA. Importantly, induction of these Th2 cytokines was inhibited in recipients of combined Th1 and Th2 cells. Anti-IFN-γ treatment attenuated the down-regulatory effect of Th1 cells. Allergen-specific Th1 cells down-regulate efferent Th2 cytokine-dependent BHR and BAL eosinophilia in an asthma model via mechanisms that depend on IFN-γ. Therapy designed to control the efferent phase of established asthma by augmenting down-regulatory Th1 counterbalancing mechanisms should be effective.
Ocular Immune Privilege Promoted by the Presentation of Peptide on Tolerogenic B Cells in the Spleen. II. Evidence for Presentation by Qa-1
Authors: D’Orazio Thomas J. , Mayhew Elizabeth , Niederkorn Jerry Y.
Published: April 2014

Ocular immune privilege is the result of several unique features of the eye, including the systemic down-regulation of Th1 immune responses to Ags encountered in the anterior chamber of the eye—a phenomenon termed anterior chamber-associated immune deviation (ACAID). The induction of ACAID requires the participation of three cell populations: the ocular ACAID APC, the splenic B cell, and the splenic T cell. Because B cells have been implicated in tolerogenic Ag presentation in other systems, we hypothesized that B cells were responsible for the induction of regulatory T cells in ACAID. The central hypothesis for this study is that APC from the eye migrate to the spleen where they release antigenic peptides (OVA) that are captured and presented to T cells by splenic B cells. A combination of in vitro and in vivo studies demonstrated that splenic B cells, incubated with ACAID APC in vitro, were capable of inducing ACAID when transferred to naive mice. The induction of ACAID required the normal expression of β2-microglobulin on both the B cell and ACAID APC, but not on the T suppressor cells. Moreover, the induction of ACAID regulatory cells required histocompatibility between the B cells and regulatory T cells at the TL/Qa region. The results indicate that: 1) B cells are necessary for the induction of ACAID; 2) ACAID B cells do not directly suppress the expression of delayed-type hypersensitivity; and 3) the induction of Ag-specific regulatory T cells by ACAID B cells requires histocompatibility at the TL/Qa region.
Control of T Cell Development In Vivo by Subdomains Within the IL-7 Receptor α-Chain Cytoplasmic Tail
Authors: Porter Brian O. , Scibelli Paul , Malek Thomas R.
Published: April 2014

IL-7/IL-7R signaling functions in both growth and differentiation during T cell development. In this study, we examined the extent these activities were controlled by signaling associated with distinct IL-7Rα cytoplasmic domains by transgenic expression of wild-type or cytoplasmic deletion mutants of IL-7Rα in the thymi of IL-7Rα−/− mice. We show an essential requirement for the tyrosine-containing carboxyl-terminal T domain in restoring thymic cellularity, pro-/pre-T cell progression, and survival. In contrast, the functional differentiation of TCR αβ cells and the development of TCR γδ cells are partially independent of the T domain. Thus, separate cytoplasmic domains of the IL-7Rα chain differentially control distinct functions during T cell development, whereas normal IL-7R-dependent thymic development requires the integrated activity of all these domains.
Enhanced Secretion of IFN-γ by Activated Th1 Cells Occurs Via Reverse Signaling Through TNF-Related Activation-Induced Cytokine
Authors: Chen Nien-Jung , Huang Mei-Wen , Hsieh Shie-Liang
Published: April 2014

Growing evidence has demonstrated that members of TNF superfamily transduce signals after engagement with their receptors. TNF-related activation-induced cytokine (TRANCE), a member of TNF superfamily, is preferentially expressed on the surface of activated CD4+ Th1 cells. The soluble receptor activator of NF-κB (RANK).Fc fusion protein suppresses IFN-γ secretion by activated Th1 cells, but does not affect IL-4 secretion by Th2 cells. The suppressive effect on IFN-γ secretion is observed when Th1 cells are activated by APCs, but not by immobilized anti-TCRβ mAb. In contrast, immobilized RANK.Fc fusion protein augments IFN-γ secretion by Th1 cells, indicating the occurrence of reverse signaling through TRANCE during T cell/APC interaction. The enhanced secretion of IFN-γ mediated via TRANCE correlates with the activation of p38 mitogen-activated protein kinase and is blocked by SB203580, a p38 mitogen-activated protein kinase-specific inhibitor. Thus, in addition to its role in activating dendritic cells by binding to the receptor RANK, TRANCE itself can signal the augmentation of IFN-γ secretion via a p38-dependent pathway, and this provides yet another example of reverse signaling by a member of TNF superfamily.
Multiple Signals Required for Cyclic AMP-Responsive Element Binding Protein (CREB) Binding Protein Interaction Induced by CD3/CD28 Costimulation
Authors: Yu Cheng-Tai , Shih Hsiu-ming , Lai Ming-Zong
Published: April 2014

The optimal activation of cAMP-responsive element binding protein (CREB), similar to the full activation of T lymphocytes, requires the stimulation of both CD3 and CD28. Using a reporter system to detect interaction of CREB and CREB-binding protein (CBP), in this study we found that CREB binds to CBP only by engagement of both CD3 and CD28. CD3/CD28-promoted CREB-CBP interaction was dependent on p38 mitogen-activated protein kinase (MAPK) and calcium/calmodulin-dependent protein kinase (CaMK) IV in addition to the previously identified extracellular signal-regulated kinase pathway. Extracellular signal-regulated kinase, CaMKIV, and p38 MAPK were also the kinases involved in CREB Ser133 phosphorylation induced by CD3/CD28. A reconstitution experiment illustrated that optimum CREB-CBP interaction and CREB trans-activation were attained when these three kinase pathways were simultaneously activated in T cells. Our results demonstrate that coordinated activation of different kinases leads to full activation of CREB. Notably, CD28 ligation activated p38 MAPK and CaMKIV, the kinases stimulated by CD3 engagement, suggesting that CD28 acts by increasing the activation extent of p38 MAPK and CaMKIV. These results support the model of a minimum activation threshold for CREB-CBP interaction that can be reached only when both CD3 and CD28 are stimulated.
MIP-2 Recruits NKT Cells to the Spleen During Tolerance Induction
Authors: Faunce Douglas E. , Sonoda Koh-Hei , Stein-Streilein Joan
Published: April 2014

Peripheral tolerance occurs after intraocular administration of Ag and is dependent on an increase in splenic NKT cells. New data here show that macrophage inflammatory protein-2 (MIP-2) is selectively up-regulated in tolerance-conferring APCs and serves to recruit NKT cells to the splenic marginal zone, where they form clusters with APCs and T cells. In the absence of the high-affinity receptor for MIP-2 (as in CXCR2-deficient mice) or in the presence of a blocking Ab to MIP-2, peripheral tolerance is prevented, and Ag-specific T regulatory cells are not generated. Understanding the regulation of lymphocyte traffic during tolerance induction may lead to novel therapies for autoimmunity, graft acceptance, and tumor rejection.
Fascin Is Involved in the Antigen Presentation Activity of Mature Dendritic Cells
Authors: Al-Alwan Monther M. , Rowden Geoffrey , Lee Timothy D. G. , West Kenneth A.
Published: April 2014

Maturation of dendritic cells (DC) is critical to their development into potent APCs. Upon maturation, DC up-regulate the expression of MHC class II as well as costimulatory and adhesion molecules, all of which are important in Ag presentation. In addition, they undergo structural changes characterized by the expression of numerous long dendrites. Fascin is an actin-bundling protein that has been reported to be important for the development of dendrites. In this study, we evaluated fascin expression and function during DC maturation into potent APC. In vitro, treatment of bone marrow-derived DC (BM-DC) with GM-CSF resulted in increased levels of fascin expression. This increase correlated directly with an increase in MHC class II and B7-2 expression. Fascin expression was decreased by the addition of TGF-β and increased by the addition TNF-α to the culture. These cytokines suppress or enhance DC maturation, respectively. Increased levels of fascin expression were found to correlate with increased APC activity in a one-way MLR. Specific inhibition of fascin expression, using antisense oligonucleotides, markedly reduced this APC allostimulatory activity. These data demonstrate that fascin expression correlates with DC maturation into APC, and it plays a significant role in the ability of DC to function as APC. This observation is the first evidence linking fascin-mediated dendrite formation with the APC activity of DC.
T Lymphocytes Promote the Development of Bone Marrow-Derived APC in the Central Nervous System
Authors: Subramanian Sandhya , Bourdette Dennis N. , Corless Christopher , Vandenbark Arthur A. , Offner Halina , Jones Richard E.
Published: April 2014

Certain cells within the CNS, microglial cells and perivascular macrophages, develop from hemopoietic myelomonocytic lineage progenitors in the bone marrow (BM). Such BM-derived cells function as CNS APC during the development of T cell-mediated paralytic inflammation in diseases such as experimental autoimmune encephalomyelitis and multiple sclerosis. We used a novel, interspecies, rat-into-mouse T cell and/or BM cell-transfer method to examine the development and function of BM-derived APC in the CNS. Activated rat T cells, specific for either myelin or nonmyelin Ag, entered the SCID mouse CNS within 3–5 days of cell transfer and caused an accelerated recruitment of BM-derived APC into the CNS. Rat APC in the mouse CNS developed from transferred rat BM within an 8-day period and were entirely sufficient for induction of CNS inflammation and paralysis mediated by myelin-specific rat T cells. The results demonstrate that T cells modulate the development of BM-derived CNS APC in an Ag-independent fashion. This previously unrecognized regulatory pathway, governing the presence of functional APC in the CNS, may be relevant to pathogenesis in experimental autoimmune encephalomyelitis, multiple sclerosis, and/or other CNS diseases involving myelomonocytic lineage cells.
RAG1 and RAG2 Expression by B Cell Subsets from Human Tonsil and Peripheral Blood
Authors: Girschick Hermann J. , Grammer Amrie C. , Nanki Toshihiro , Mayo Marlyn , Lipsky Peter E.
Published: April 2014

It has been suggested that B cells acquire the capacity for secondary V(D)J recombination during germinal center (GC) reactions. The nature of these B cells remains controversial. Subsets of tonsil and blood B cells and also individual B cells were examined for the expression of recombination-activating gene (RAG) mRNA. Semiquantitative analysis indicated that RAG1 mRNA was present in all tonsil B cell subsets, with the largest amount found in naive B cells. RAG2 mRNA was only found in tonsil naive B cells, centrocytes, and to a lesser extent in centroblasts. Neither RAG1 nor RAG2 mRNA was routinely found in normal peripheral blood B cells. In individual tonsil B cells, RAG1 and RAG2 mRNAs were found in 18% of naive B cells, 22% of GC founder cells, 0% of centroblasts, 13% of centrocytes, and 9% of memory B cells. Individual naive tonsil B cells containing both RAG1 and RAG2 mRNA were activated (CD69+). In normal peripheral blood ∼5% of B cells expressed both RAG1 and RAG2. These cells were uniformly postswitch memory B cells as documented by the coexpression of IgG mRNA. These results indicate that coordinate RAG expression is not found in normal peripheral naive B cells but is up-regulated in naive B cells which are activated in the tonsil. With the exception of centroblasts, RAG1 and RAG2 expression can be found in all components of the GC, including postswitch memory B cells, some of which may circulate in the blood of normal subjects.
A Novel Chicken Membrane-Associated Complement Regulatory Protein: Molecular Cloning and Functional Characterization
Authors: Inoue Naokazu , Fukui Aya , Nomura Midori , Matsumoto Misako , Toyoshima Kumao , Seya Tsukasa
Published: April 2014

A cDNA encoding a membrane-associated complement (C) regulatory protein was identified here for the first time in an oviparous vertebrate, chicken. This protein, named Cremp, possessed five short consensus repeats (SCRs) and one SCR-like domain followed by a transmembrane domain and a cytoplasmic tail. SCR1/SCR2 of Cremp were 43.6% identical with SCR2/SCR3 of human decay-accelerating factor (CD55), and SCR3/SCR4 were 45.3% identical with those of human membrane cofactor protein (CD46). Cremp is likely to be an ancestral hybrid protein of human decay-accelerating factor and membrane cofactor protein rather than a homolog of rodent C receptor 1-related protein y, which structurally resembles human CR1 (CD35). Chinese hamster ovary cells transfected with Cremp were efficiently protected from chicken C but not from human or rabbit C in both classical and alternative pathways. Thus, chicken Cremp is a membrane C regulator for cell protection against homologous C. Cremp mRNA was seen as a doublet comprised of a faint band of 2.2 kb and a thick band of 3.0 kb on RNA blotting analysis. An Ab against chicken Cremp recognized a single band of 46.8 kDa on immunoblotting. mRNA and protein of Cremp were ubiquitously expressed in all chicken organs tested. Minute amounts of dimer were present in some tissues. Surface expression of Cremp was confirmed by flow cytometry and immunofluorescence analysis. These results suggested that even in nonmammals a C regulatory membrane protein with ubiquitous tissue distribution should be a prerequisite for protection of host cells from homologous C attack.
Classically Restricted Human CD8+ T Lymphocytes Derived fromMycobacterium tuberculosis-Infected Cells: Definition of Antigenic Specificity
Authors: Lewinsohn David M. , Zhu Liqing , Madison Valerie J. , Dillon Davin C. , Fling Steven P. , Reed Steven G. , Grabstein Kenneth H. , Alderson Mark R.
Published: April 2014

Previous studies in murine and human models have suggested an important role for HLA Ia-restricted CD8+ T cells in host defense to Mycobacterium tuberculosis (Mtb). Therefore, understanding the Ags presented via HLA-Ia will be important in understanding the host response to Mtb and in rational vaccine design. We have used monocyte-derived dendritic cells in a limiting dilution analysis to generate Mtb-specific CD8+ T cells. Two HLA-Ia-restricted CD8+ T cell clones derived by this method were selected for detailed analysis. One was HLA-B44 restricted, and the other was HLA-B14 restricted. Both were found to react with Mtb-infected, but not bacillus Calmette-Guérin-infected, targets. For both these clones, the Ag was identified as culture filtrate protein 10 (CFP10)/Mtb11, a 10.8-kDa protein not expressed by bacillus Calmette-Guérin. Both clones were inhibited by the anti-class I Ab and anti-HLA-B,C Abs. Using a panel of CFP10/Mtb11-derived 15-aa peptides overlapping by 11 aa, the region containing the epitopes for both clones has been defined. Minimal 10-aa epitopes were defined for both clones. CD8+ effector cells specific for these two epitopes are present at high frequency in the circulating pool. Moreover, the CD8+ T cell response to CFP10/Mtb11 can be largely accounted for by the two epitopes defined herein, suggesting that this is the immunodominant response for this purified protein derivative-positive donor. This study represents the first time CD8+ T cells generated against Mtb-infected APC have been used to elucidate an Mtb-specific CD8+ T cell Ag.
Lipoprotein Access to MHC Class I Presentation During Infection of Murine Macrophages with Live Mycobacteria
Authors: Neyrolles Olivier , Gould Keith , Gares Marie-Pierre , Brett Sara , Janssen Riny , O’Gaora Peadar , Herrmann Jean-Louis , Prévost Marie-Christine , Perret Emmanuelle , Thole Jelle E. R. , Young Douglas
Published: April 2014

Following uptake by macrophages, live mycobacteria initially reside within an immature phagosome that resists acidification and retains access to recycling endosomes. Glycolipids are exported from the mycobacterial phagosome and become available for immune recognition by CD1-restricted T cells. The aim of this study was to explore the possibility that lipoproteins might similarly escape from the phagosome and act as immune targets in cells infected with live mycobacteria. We have focused on a 19-kDa lipoprotein from Mycobacterium tuberculosis that was previously shown to be recognized by CD8+ T cells. The 19-kDa Ag was found to traffic separately from live mycobacteria within infected macrophages by a pathway that was dependent on acylation of the protein. When expressed as a recombinant protein in rapid-growing mycobacteria, the 19-kDa Ag was able to deliver peptides for recognition by MHC class I-restricted T cells by a TAP-independent mechanism. Entry into the class I pathway was rapid, dependent on acylation, and could be blocked by killing the mycobacteria by heating before infection. Although the pattern of 19-kDa trafficking was similar with different mycobacterial species, preliminary experiments suggest that class I presentation is more efficient during infection with rapid-growing mycobacteria than with the slow-growing bacillus Calmette-Guérin vaccine strain.
Biased Vβ Usage in Immature Thymocytes Is Independent of DJβ Proximity and pTα Pairing
Authors: Wilson Anne , Maréchal Céline , MacDonald H. Robson
Published: April 2014

During thymus development, the TCR β locus rearranges before the TCR α locus. Pairing of productively rearranged TCR β-chains with an invariant pTα chain leads to the formation of a pre-TCR and subsequent expansion of immature pre-T cells. Essentially nothing is known about the TCR Vβ repertoire in pre-T cells before or after the expression of a pre-TCR. Using intracellular staining, we show here that the TCR Vβ repertoire is significantly biased at the earliest developmental stage in which VDJβ rearrangement has occurred. Moreover (and in contrast to the VH repertoire in immature B cells), Vβ repertoire biases in immature T cells do not reflect proximity of Vβ gene segments to the DJβ cluster, nor do they depend upon preferential Vβ pairing with the pTα chain. We conclude that V gene repertoires in developing T and B cells are controlled by partially distinct mechanisms.
Activation of Extracellular Signal-Regulated Kinases, NF-κB, and Cyclic Adenosine 5′-Monophosphate Response Element-Binding Protein in Lung Neutrophils Occurs by Differing Mechanisms After Hemorrhage or Endotoxemia
Authors: Abraham Edward , Arcaroli John , Shenkar Robert
Published: April 2014

Acute lung injury is frequently associated with sepsis or blood loss and is characterized by a proinflammatory response and infiltration of activated neutrophils into the lungs. Hemorrhage or endotoxemia result in activation of cAMP response element-binding protein (CREB) and NF-κB in lung neutrophils as well as increased expression of proinflammatory cytokines, such as TNF-α and macrophage-inflammatory peptide-2, by these cells. Activation of the extracellular regulated kinase (ERK) pathway occurs in stress responses and is involved in CREB activation. In the present experiments, hemorrhage or endotoxemia produced increased activation of mitogen-activated protein kinase kinase (MEK)1/2 and ERK2 (p42), but not of ERK1 (p44), in lung neutrophils. ERK1, ERK2, and MEK1/2 were not activated in peripheral blood neutrophils after hemorrhage or endotoxemia. Inhibition of xanthine oxidase led to further increase in the activation of MEK1/2 and ERK2 in lung neutrophils after hemorrhage, but not after endotoxemia. α-Adrenergic blockade before hemorrhage resulted in increased activation in lung neutrophils of MEK1/2, ERK1, ERK2, and CREB, but decreased activation of NF-κB. In contrast, α-adrenergic blockade before endotoxemia was associated with decreased activation of MEK1/2, ERK2, and CREB, but increased activation of NF-κB. β-Adrenergic blockade before hemorrhage did not alter MEK1/2 or ERK1 activation in lung neutrophils, but decreased activation of ERK2 and CREB, while increasing activation of NF-κB. β-Adrenergic inhibition before endotoxemia did not affect activation of MEK1/2, ERK1, ERK2, CREB, or NF-κB. These data indicate that the pathways leading to lung neutrophil activation after hemorrhage are different from those induced by endotoxemia.
Annexin V Binds to Positively Selected B Cells
Authors: Dillon Stacey R. , Constantinescu Andrei , Schlissel Mark S.
Published: April 2014

Recombinant annexin V (rAnV) has been used in flow cytometry to identify cells undergoing apoptosis, based on its ability to bind to phosphatidylserine, a negatively charged lipid normally restricted to the cytoplasmic face of the plasma membrane but externalized early during apoptosis. When we stained murine bone marrow (BM) cells with fluorescently labeled rAnV, we found that a surprisingly large fraction of BM B cells bearing selectable transgenic Ag receptors bind significant amounts of rAnV, but that these cells are not apoptotic. Here, we show that binding of rAnV to developing B cells in normal mice correlates with B cell receptor-dependent selection events at several stages of development within both B-1 and B-2 cell subsets. In fact, nearly all B-1 B cells and splenic marginal zone B cells bind rAnV, suggesting that the externalization of phosphatidylserine occurs once mature B cells are selected through BCR-mediated signaling. However, this plasma membrane alteration is apparently not shared by all lymphocytes, because we did not find a parallel population of rAnV-binding viable T cells in vivo in normal or TCR transgenic mice. We also show that BM stromal cell lines can influence the extent of rAnV binding by viable BM B cells during coculture in vitro. We suggest that rAnV detects a potentially important membrane alteration that occurs as B cells develop in the BM and are readied for export to the peripheral lymphoid organs and again among mature B cells recruited to the marginal zone or the B-1 compartment.
p38 Mitogen-Activated Protein Kinase Inhibition Increases Cytokine Release by Macrophages In Vitro and During Infection In Vivo
Authors: van den Blink Bernt , Juffermans Nicole P. , ten Hove Tessa , Schultz Mark J. , van Deventer Sander J. H. , van der Poll Tom , Peppelenbosch Maikel P.
Published: April 2014

p38 mitogen-activated protein kinase (MAPK) has been suggested as a mediator of cytokine release and is currently being targeted for anti-inflammatory therapy. However, experimental data are contradictory and lack sufficient affirmation in vivo. We tested the effect of p38 MAPK inhibition in several cell types and in different murine models of infectious disease. We observed that most cell types react to p38 MAPK inhibition with diminished cytokine release, but that this treatment induced increased cytokine release in macrophages. Furthermore, we observed increased cytokine production in mouse models of pneumococcal pneumonia and tuberculosis accompanied by severely reduced bacterial clearance. This apparent inefficacy of p38 MAPK inhibition in reducing cytokine release in infectious disease, as well as its immune-compromising action, suggest that targeting p38 MAPK may not be a suitable anti-cytokine strategy in patients with such disease or at risk for infection.
Homing of In Vitro-Generated Donor Antigen-Reactive CD4+ T Lymphocytes to Renal Allografts Is α4β1 But Not αLβ2 Integrin Dependent
Authors: Hammer Markus H. , Zhai Yuan , Katori Masamichi , Ritter Thomas , Volk Hans-Dieter , Coito Ana J. , Kupiec-Weglinski Jerzy W.
Published: April 2014

The extravasation and sequestration of Ag-reactive T lymphocytes into vascularized organ allografts depend on a cascade of complex interactions among circulating lymphocytes, endothelial cells, and extracellular matrix proteins. Ag-activated donor-specific CD4 T cells are major initiators and effectors in the allograft rejection response. Interfering with the intragraft homing of activated CD4 T cells may represent a novel therapeutic approach in transplant recipients. We have developed a FACS-based short-term homing assay that allows tracing in vitro-generated Ag-reactive CD4 T cells after adoptive transfer in test rat recipients. Allospecific cell lines were preincubated with anti-α4β1 or anti-αLβ2 mAb, because of enhanced expression of both integrin receptors after alloactivation. The pretreated LewisBN lymphocytes were carboxyfluorescein diacetate succinimidyl ester labeled and adoptively transferred into Lewis rat recipients of Brown Norway kidney allografts. The injection of equal numbers of PKH-26-labeled untreated cells allowed quantitative comparison of both populations in the same animal. Ex vivo treatment with anti-α4β1 mAb diminished intragraft infiltration of adoptively transferred T cells by 85% in a donor-specific fashion. In contrast, treatment with anti-αLβ2 mAb did not affect intragraft cell sequestration. Hence, blocking α4β1 integrin interactions represents a novel strategy in preventing local intragraft recruitment of Ag-reactive CD4 T cells in transplant recipients.
Cutting Edge: A Role for B Lymphocyte Stimulator in Systemic Lupus Erythematosus
Authors: Zhang Jun , Roschke Viktor , Baker Kevin P. , Wang Zheng , Alarcón Graciela S. , Fessler Barri J. , Bastian Holly , Kimberly Robert P. , Zhou Tong
Published: April 2014

Increased levels of B lymphocyte stimulator (BLyS) are associated with systemic autoimmunity in animal models of spontaneous autoimmune disease, and transgenic animals expressing BLyS develop typical autoimmune disease. Here, we demonstrate significant elevations of BLyS in the patients with systemic lupus erythematosus (SLE). The BLyS isolated from the sera of SLE patients had the same m.w. as the natural soluble form and was able to stimulate B cell activation in vitro. Increased BLyS in SLE patients was partially associated with higher levels of anti-dsDNA Ab of the IgG, IgM, and IgA classes, but not associated with the disease activity. Our results suggest that BLyS may be a useful marker for early activation of an autoimmune diathesis and likely plays a critical role in triggering activation of self-Ag-driven autoimmune B cells in human SLE. BLyS may provide an effective therapeutic target in systemic autoimmunity.
Genetically Modified Bone Marrow-Derived Vehicle Cells Site Specifically Deliver an Anti-Inflammatory Cytokine to Inflamed Interstitium of Obstructive Nephropathy
Authors: Yamagishi Hiroko , Yokoo Takashi , Imasawa Toshiyuki , Mitarai Tetsuya , Kawamura Tetsuya , Utsunomiya Yasunori
Published: April 2014

In this study, we used genetically modified bone marrow-derived CD11b+CD18+ vehicle cells to deliver IL-1 receptor antagonist (IL-1ra) for treatment of inflamed renal interstitium in an animal model of unilateral ureteral obstruction (UUO). Vehicle cells that expressed the ICAM-1 ligands, CD11b and CD18, were obtained from bone marrow cells of DBA/2j mice and adenovirally transduced with the IL-1ra gene or glucocerebrosidase (GC) gene ex vivo. In kidneys treated to develop UUO, levels of ICAM-1, IL-1β, and IL-1R expression increased within 3 days compared with contralateral untreated kidneys in the same mice. Similarly, the macrophage infiltration in the cortical interstitium increased after 3 days in UUO kidneys, but not untreated kidneys. After UUO developed, DBA/2j mice were injected i.v. with either IL-1ra+ vehicle cells (IL-1ra-treated mice) or GC+ vehicle cells (GC-treated mice) at 24 h after UUO. Six days after the injection of these vehicle cells, marked increase of CD11b+ IL-1ra+ vehicle cells was observed in the ICAM-1-positive interstitium of UUO kidneys from IL-1ra-treated mice. In contrast, no CD11b+ IL-1ra+ cells appeared in ICAM-1-negative contralateral kidneys from these mice. Furthermore, the infiltration of macrophages (p < 0.001), expression of ICAM-1 (p < 0.005), and presence of α-smooth muscle actin (p = 0.005) in the interstitium of UUO kidneys were significantly decreased in IL-1ra-treated mice compared with GC-treated mice. These findings suggest that IL-1 may contribute to the development of renal interstitial injury and that our method can deliver a functioning gene encoding an antiinflammatory cytokine gene specifically at that site by interacting with local adhesion molecules.
Lymphoid Chemokine B Cell-Attracting Chemokine-1 (CXCL13) Is Expressed in Germinal Center of Ectopic Lymphoid Follicles Within the Synovium of Chronic Arthritis Patients
Authors: Shi Kenrin , Hayashida Kenji , Kaneko Motoharu , Hashimoto Jun , Tomita Tetsuya , Lipsky Peter E. , Yoshikawa Hideki , Ochi Takahiro
Published: April 2014

A unique feature in inflammatory tissue of rheumatoid arthritis (RA) is the formation of ectopic lymphoid aggregates with germinal center (GC)-like structures that can be considered to contribute to the pathogenesis of RA, because local production of the autoantibody, rheumatoid factor, is thought to be a causative factor in tissue damage. However, the factors governing the formation of GC in RA are presently unknown. To begin to address this, the expression of B cell attracting chemokine (BCA-1) (CXCL13), a potent chemoattractant of B cells, was examined in the synovium of patients with RA or with osteoarthritis (OA). Expression of BCA-1 mRNA was detected in all RA samples, but in only one of five OA samples. Lymphoid follicles were observed in four of seven RA samples and in two of eight OA samples, and in most of them BCA-1 protein was detected in GC. BCA-1 was not detected in tissues lacking lymphoid follicles. Notably, BCA-1 was detected predominantly in follicular dendritic cells in GC. CD20-positive B cells were aggregated in regions of BCA-1 expression, but not T cells or macrophages. These data suggest that BCA-1 produced by follicular dendritic cells may attract B cells and contribute to the formation of GC-like structures in chronic arthritis.
Distinct T Cell Developmental Consequences in Humans and Mice Expressing Identical Mutations in the DLAARN Motif of ZAP-70
Authors: Elder Melissa E. , Skoda-Smith Suzanne , Kadlecek Theresa A. , Wang Fengling , Wu Jun , Weiss Arthur
Published: April 2014

The protein tyrosine kinase, ZAP-70, is pivotally involved in transduction of Ag-binding signals from the TCR required for T cell activation and development. Defects in ZAP-70 result in SCID in humans and mice. We describe an infant with SCID due to a novel ZAP-70 mutation, comparable with that which arose spontaneously in an inbred mouse colony. The patient inherited a homozygous missense mutation within the highly conserved DLAARN motif in the ZAP-70 kinase domain. Although the mutation only modestly affected protein stability, catalytic function was absent. Despite identical changes in the amino acid sequence of ZAP-70, the peripheral T cell phenotypes of our patient and affected mice are distinct. ZAP-70 deficiency in this patient, as in other humans, is characterized by abundant nonfunctional CD4+ T cells and absent CD8+ T cells. In contrast, ZAP-70-deficient mice lack both major T cell subsets. Although levels of the ZAP-70-related protein tyrosine kinase, Syk, may be sufficiently increased in human thymocytes to rescue CD4 development, survival of ZAP-70-deficient T cells in the periphery does not appear to be dependent on persistent up-regulation of Syk expression.
Costimulation-Dependent Modulation of Experimental Autoimmune Encephalomyelitis by Ligand Stimulation of Vα14 NK T Cells
Authors: Pál Endre , Tabira Takeshi , Kawano Tetsu , Taniguchi Masaru , Miyake Sachiko , Yamamura Takashi
Published: April 2014

Experimental autoimmune encephalomyelitis (EAE) is a Th1 cell-mediated autoimmune disease that can be protected against by stimulating regulatory cells. Here we examined whether EAE can be purposefully modulated by stimulating Vα14 NK T cells with the CD1d-restricted ligand α-galactosylceramide (α-GC). EAE induced in wild-type C57BL/6 (B6) mice was not appreciably altered by injection of α-GC. However, EAE induced in IL-4 knockout mice and IFN-γ knockout mice was enhanced or suppressed by α-GC, respectively. This indicates that the IL-4 and IFN-γ triggered by α-GC may play an inhibitory or enhancing role in the regulation of EAE. We next studied whether NK T cells of wild-type mice may switch their Th0-like phenotype toward Th1 or Th2. Notably, in the presence of blocking B7.2 (CD86) mAb, α-GC stimulation could bias the cytokine profile of NK T cells toward Th2, whereas presentation of α-GC by CD40-activated APC induced a Th1 shift of NK T cells. Furthermore, transfer of the α-GC-pulsed APC preparations suppressed or enhanced EAE according to their ability to polarize NK T cells toward Th2 or Th1 in vitro. These results have important implications for understanding the role of NK T cells in autoimmunity and for designing a therapeutic strategy targeting NK T cells.
Increased Production of Immature Myeloid Cells in Cancer Patients: A Mechanism of Immunosuppression in Cancer
Authors: Almand Bond , Clark Joseph I. , Nikitina Ekaterina , van Beynen James , English Nicholas R. , Knight Stella C. , Carbone David P. , Gabrilovich Dmitry I.
Published: April 2014

Defective dendritic cell (DC) function caused by abnormal differentiation of these cells is an important mechanism of tumor escape from immune system control. Previously, we have demonstrated that the number and function of DC were dramatically reduced in cancer patients. This effect was closely associated with accumulation of immature cells (ImC) in peripheral blood. In this study, we investigated the nature and functional role of those ImC. Using flow cytometry, electron microscopy, colony formation assays, and cell differentiation in the presence of different cell growth factors, we have determined that the population of ImC is composed of a small percentage (<2%) of hemopoietic progenitor cells, with all other cells being represented by MHC class I-positive myeloid cells. About one-third of ImC were immature macrophages and DC, and the remaining cells were immature myeloid cells at earlier stages of differentiation. These cells were differentiated into mature DC in the presence of 1 μM all-trans-retinoic acid. Removal of ImC from DC fractions completely restored the ability of the DC to stimulate allogeneic T cells. In two different experimental systems ImC inhibited Ag-specific T cell responses. Thus, immature myeloid cells generated in large numbers in cancer patients are able to directly inhibit Ag-specific T cell responses. This may represent a new mechanism of immune suppression in cancer and may suggest a new approach to cancer treatment.
Gangliosides GD1b, GT1b, and GQ1b Enhance IL-2 and IFN-γ Production and Suppress IL-4 and IL-5 Production in Phytohemagglutinin-Stimulated Human T Cells
Authors: Kanda Naoko , Watanabe Shinichi
Published: April 2014

Gangliosides are sialic acid-containing glycolipids. We studied the in vitro effects of gangliosides on Th1 and Th2 cytokine production in PHA-stimulated human T cells. Gangliosides GD1b, GT1b, and GQ1b (each 100 nM) enhanced PHA-induced IL-2 secretion of peripheral blood T cells ∼4-fold and enhanced that of IFN-γ 3- to 4-fold compared with controls. These gangliosides decreased PHA-induced IL-4 secretion by 50–53% and that of IL-5 by 53–63% compared with controls, respectively. The other gangliosides did not alter the secretion of Th1 or Th2 cytokines. RT-PCR showed that GD1b, GT1b, and GQ1b enhanced PHA-induced IL-2 and IFN-γ transcription and suppressed that of IL-4 and IL-5. Transient transfection assays of Jurkat T cells showed that GD1b, GT1b, and GQ1b enhanced PHA-induced IL-2 and IFN-γ promoter activities but suppressed those of IL-4 and IL-5. The cAMP analogue dibutyryl cAMP and the cAMP-elevating agents forskolin and 3-isobutyl-1-methylxanthine each reversed GD1b-, GT1b-, and GQ1b-induced stimulation of IL-2 and IFN-γ production and inhibition of IL-4 and IL-5 production at the levels of proteins, transcription, and promoter activities. GD1b, GT1b, and GQ1b suppressed PHA-induced increase in cAMP level in T cells. These gangliosides suppressed PHA-stimulated adenylate cyclase activity in T cells. These results suggest that GD1b, GT1b, and GQ1b may enhance Th1 cytokine production while suppressing Th2 production by inhibiting adenylate cyclase activity.
Anti-CD40 Antibody Induces Antitumor and Antimetastatic Effects: The Role of NK Cells
Authors: Turner Joel G. , Rakhmilevich Alexander L. , Burdelya Lyudmila , Neal Zane , Imboden Michael , Sondel Paul M. , Yu Hua
Published: April 2014

We assessed the effect of the stimulatory anti-CD40 Ab on NK cell activation in vivo and the therapeutic potential of activated NK cells in tumor-bearing mice. Single-dose i.p. injection of the anti-CD40 Ab resulted in production of IL-12 and IFN-γ in vivo, followed by a dramatic increase in NK cell cytolytic activity in PBLs. NK cell activation by anti-CD40 Ab was also observed in CD40 ligand knockout mice. Because NK cells express CD40 ligand but not CD40, our results suggest that NK activation is mediated by increased cytokine production upon CD40 ligation of APCs. Treatment of tumor-bearing mice with anti-CD40 Ab resulted in substantial antitumor and antimetastatic effects in three tumor models. Depletion of NK cells with anti-asialo GM1 Ab reduced or abrogated the observed antitumor effects in all the tested models. These results indicate that a stimulatory CD40 Ab indirectly activates NK cells, which can produce significant antitumor and antimetastatic effects.
CD44-Deficient Mice Exhibit Enhanced Hepatitis After Concanavalin A Injection: Evidence for Involvement of CD44 in Activation-Induced Cell Death
Authors: Chen Dawei , McKallip Robert J. , Zeytun Ahmet , Do Yoonkyung , Lombard Catherine , Robertson John L. , Mak Tak W. , Nagarkatti Prakash S. , Nagarkatti Mitzi
Published: April 2014

Administration of Con A induces severe injury to hepatocytes in mice and is considered to be a model for human hepatitis. In the current study, we investigated the role of CD44 in Con A-induced hepatitis. Intravenous administration of Con A (20 mg/kg) caused 100% mortality in C57BL/6 CD44-knockout (KO) mice, although it was not lethal in C57BL/6 CD44 wild-type (WT) mice. Administration of lower doses of Con A (12 mg/kg body weight) into CD44 WT mice induced hepatitis as evident from increased plasma aspartate aminotransferase levels accompanied by active infiltration of mononuclear cells and neutrophils, and significant induction of apoptosis in the liver. Interestingly, CD44 KO mice injected with similar doses of Con A exhibited more severe acute suppurative hepatitis. Transfer of spleen cells from Con A-injected CD44 KO mice into CD44 WT mice induced higher levels of hepatitis when compared with transfer of similar cells from CD44 WT mice into CD44 WT mice. The increased hepatitis seen in CD44 KO mice was accompanied by increased production of cytokines such as TNF-α, IL-2 and IFN-γ, but not Fas or Fas ligand. The increased susceptibility of CD44 KO mice to hepatitis correlated with the observation that T cells from CD44 KO mice were more resistant to activation-induced cell death when compared with the CD44 WT mice. Together, these data demonstrate that activated T cells use CD44 to undergo apoptosis, and dysregulation in this pathway could lead to increased pathogenesis in a number of diseases, including hepatitis.
Distinct Mechanisms Contribute to Generate and Change the CD4:CD8 Cell Ratio During Thymus Development: A Role for the Notch Ligand, Jagged1
Authors: Jiménez Eva , Vicente Angeles , Sacedón Rosa , Muñoz Juan J. , Weinmaster Gerry , Zapata Agustín G. , Varas Alberto
Published: April 2014

In adult life, the high CD4:CD8 cell ratio observed in peripheral lymphoid organs originates in the thymus. Our results show that the low peripheral CD4:CD8 cell ratio seen during fetal life also has an intrathymic origin. This distinct production of CD4+CD8− and CD4−CD8+ thymocytes is regulated by the developmental age of the thymic stroma. The differential expression of Notch receptors and their ligands, especially Jagged1, throughout thymus development plays a key role in the generation of the different CD4:CD8 cell ratios. We also show that the intrathymic CD4:CD8 cell ratio sharply changes from fetal to adult values around birth. Differences in the proliferation and emigration rates of the mature thymocyte subsets contribute to this change.
Definition of a Novel Cellular Constituent of the Bone Marrow That Regulates the Response of Immature B Cells to B Cell Antigen Receptor Engagement
Authors: Sandel Peter C. , Gendelman Mariya , Kelsoe Garnett , Monroe John G.
Published: April 2014

Previously we defined a Thy1dull bone marrow-derived cell population that regulated fate decisions by immature B cells after Ag receptor signaling. The microenvironmental signals provided by this cell population were shown to redirect the B cell Ag receptor -induced apoptotic response of immature B cells toward continued recombination-activating gene (RAG) expression and secondary light chain recombination (receptor editing). Neither the identity of the cell responsible for this activity nor its role in immature B cell development in vivo were addressed by these previous studies. Here we show that this protective microenvironmental niche is defined by the presence of a novel Thy1dull, DX5pos cell that can be found in close association with immature B cells in vivo. Depletion of this cell eliminates the anti-apoptotic effect of bone marrow in vitro and leads to a significant decrease in the number and frequency of bone marrow immature B cells in vivo. We propose that, just as the bone marrow environment is essential for the survival and progression of pro-B and pre-B cells through their respective developmental checkpoints, this cellular niche regulates the progression of immature stage B cells through negative selection.
Protein Kinase C-θ Mediates a Selective T Cell Survival Signal Via Phosphorylation of BAD
Authors: Villalba Martin , Bushway Paul , Altman Amnon
Published: April 2014

Protein kinase C (PKC)-activating phorbol esters protect T cells from Fas-induced apoptosis. However, the mechanism of this protective effect and the identity of the relevant PKC isoform(s) are poorly understood. Here, we show that PKCθ plays a selective and important role in this protection. Fas triggering led to a selective caspase-3-dependent cleavage of the enzyme and proteasome-mediated degradation and inactivation of its catalytic fragment. These events preceded the onset of apoptosis. Pharmacological inhibition of PKCθ promoted Fas-mediated apoptosis in three different types of T cells. Conversely, constitutively active PKCθ (and, to a lesser degree, PKCε) selectively protected T cells from Fas-induced apoptosis. We provide evidence that the distant Bcl-2 family member, BAD, is a PKCθ substrate, is phosphorylated by TCR stimulation, and can mediate at least in part the anti-apoptotic effect of PKCθ.
Generation of Macrophages from Early T Progenitors In Vitro
Authors: Lee Chong-Kil , Kim Jeong Ki , Kim Youngsoo , Lee Myung-Koo , Kim Kyungjae , Kang Jong-Koo , Hofmeister Robert , Durum Scott K. , Han Seong Sun
Published: April 2014

Early T progenitors in the thymus have been reported to have the capacity to develop into B cells, thymic dendritic cells, and NK cells. Here we describe conditions that induce early T progenitors to develop into macrophages. Initially, we observed that early T progenitors could be induced to develop into macrophages by cytokines produced from a thymic stromal cell line, TFGD, and later we found that the cytokine mixture of M-CSF plus IL-6 plus IL-7 also induced macrophage differentiation from pro-T cells. M-CSF by itself was unable to induce macrophage differentiation from early T progenitors. To correlate this observation with the developmental potential of early T progenitors, mouse embryonic thymocytes were sorted into four populations, pro-T1 to pro-T4, based on the expression of CD44 and CD25, and then cultured with TFGD culture supernatant. We found that pro-T1 and pro-T2 cells, but not pro-T3 and pro-T4 cells, generate macrophages. Limiting dilution analysis of the differentiation capability of sorted pro-T2 cells also confirmed that pro-T2 cells could generate macrophages. These results suggest that T cells and thymic macrophages could originate from a common intrathymic precursor.
Early Intestinal Th1 Inflammation and Mucosal T Cell Recruitment During Acute Graft-Versus-Host Reaction
Authors: Snider Denis , Liang Hong
Published: April 2014

Little is understood about the earliest cytokine responses and the role(s) of donor CD4 T cells in the intestine during the induced graft-vs-host reaction (GVHR). We investigated the activation and mucosal homing phenotype of the donor CD4 cells and the kinetics of cytokine responses within the intestine and associated lymphoid tissues during early GVHR. Significant frequencies of donor CD4 cells accumulated within recipient Peyer’s patches (PP), mesenteric lymph nodes (MLN), lamina propria (LP), and spleen (SP), during the first 9 days of GVHR. Many donor CD4 cells in SP, MLN, and LP expressed CD44 and also expressed de novo the mucosal homing integrin α4β7 (LPAM-1). A large IFN-γ response occurred by day 3 in cells from PP and MLN, but much later (day 9) in SP and LP cells. IL-10 production by SP and MLN cells was elevated initially but declined substantially by day 9. IL-4 production by SP, MLN, and PP cells was low on day 3 and showed gradual decline in LP by day 9. IL-5 production by LP cells gradually increased in direct contrast to IL-5 production by MLN cells. The MLN CD4 cells showed the most dynamic changes, with high numbers of activated/effector donor CD4 cells and altered cytokine production consistent with a developing Th1 response. The IFN-γ responses in PP and MLN preceded that of the SP, suggesting an intestinal origin for some Th1 effector cells in GVHR. Donor CD4 T cells apparently acquire the ability to home to the LP during early GVHR.