bluuebunny/crossref_metadata_2025 huggingface dataset:
Histamine Directly and Synergistically with Lipopolysaccharide Stimulates Cyclooxygenase-2 Expression and Prostaglandin I2 and E2 Production in Human Coronary Artery Endothelial Cells
Authors: Tan Xiaoyu , Essengue Suzanne , Talreja Jaya , Reese Jeff , Stechschulte Daniel J. , Dileepan Kottarappat N.
Published: April 2014

Although histamine plays an essential role in inflammation, its influence on cyclooxygenases (COX) and prostanoid homeostasis is not well understood. In this study, we investigated the effects of histamine on the expression of COX-1 and COX-2 and determined their contribution to the production of PGE2, prostacyclin (PGI2), and thromboxane A2 in human coronary artery endothelial cells (HCAEC). Incubation of HCAEC monolayers with histamine resulted in marked increases in the expression of COX-2 and production of PGI2 and PGE2 with no significant change in the expression of COX-1. Histamine-induced increases in PGI2 and PGE2 production were due to increased expression and function of COX-2 because gene silencing by small interfering RNA or inhibition of the catalytic activity by a COX-2 inhibitor blocked prostanoid production. The effects of histamine on COX-2 expression and prostanoid production were mediated through H1 receptors. In addition to the direct effect, histamine was found to amplify LPS-stimulated COX-2 expression and PGE2 and PGI2 production. In contrast, histamine did not stimulate thromboxane A2 production in resting or LPS-activated HCAEC. Histamine-induced increases in the production of PGE2 and PGI2 were associated with increased expression of mRNA encoding PGE2 and PGI2 synthases. The physiological role of histamine on the regulation of COX-2 expression in the vasculature is indicated by the findings that the expression of COX-2 mRNA, but not COX-1 mRNA, was markedly reduced in the aortic tissues of histidine decarboxylase null mice. Thus, histamine plays an important role in the regulation of COX-2 expression and prostanoid homeostasis in vascular endothelium.
FcγRIIa Genotype Predicts Progression of HIV Infection
Authors: Forthal Donald N. , Landucci Gary , Bream Jay , Jacobson Lisa P. , Phan Tran B. , Montoya Benjamin
Published: April 2014

Polymorphisms in FcγR genes are associated with susceptibility to or severity of a number of autoimmune and infectious diseases. We found that HIV-infected men in the Multicenter AIDS Cohort Study with the FcγRIIa RR genotype progressed to a CD4+ cell count of <200/mm3 at a faster rate than individuals with the RH or HH genotypes (relative hazard = 1.6; p = 0.0001). However, progression to AIDS (using the broad definition of either a CD4+ cell count <200/mm3 or development of an AIDS-defining illness) was less impacted by FcγRIIa genotype, largely because HH homozygotes had an increased risk of Pneumocystis jiroveci pneumonia as an AIDS-defining illness. We also showed that chronically infected subjects develop a substantial anti-gp120-specific IgG2 response. Moreover, HIV-1 immune complexes are more efficiently internalized by monocytes from HH subjects compared with RR subjects, likely because of the presence of IgG2 in the complexes. Finally, the FcγRIIIa F/V gene polymorphism was not associated with progression of HIV infection, but, as demonstrated previously, did predict the risk of Kaposi’s sarcoma. These results demonstrate the importance of FcγRs in AIDS pathogenesis and point toward a critical role for interactions between FcγRs and immune complexes in disease progression.
Cross-Reactive CD4+ T Cells against One Immunodominant Tumor-Derived Epitope in Melanoma Patients
Authors: Kudela Pavol , Janjic Bratislav , Fourcade Julien , Castelli Florence , Andrade Pedro , Kirkwood John M. , El-Hefnawy Talal , Amicosante Massimo , Maillere Bernard , Zarour Hassane M.
Published: April 2014

TCRs exhibit a high degree of specificity but may also recognize multiple and distinct peptide-MHC complexes, illustrating the so-called cross-reactivity of TCR-peptide-MHC recognition. In this study, we report the first evidence of CD4+ T cells recognizing the same tumor peptide-epitope from NY-ESO-1, in the context of multiple HLA-DR and HLA-DP molecules. These cross-reactive CD4+ T cells recognized not only autologous but also allogenic dendritic cells previously loaded with the relevant protein (i.e., the normally processed and presented epitope). Using clonotypic real-time RT-PCR, we have detected low frequencies of CD4+ T cells expressing one cross-reactive TCR from circulating CD4+ T cells of patients with stage IV melanoma either spontaneously or after immunization but not in normal donors. The maintenance of cross-reactive tumor Ag-specific CD4+ T cells in PBLs of cancer patients required the presence of tumor Ag/epitope in the context of the MHC molecule used to prime the Ag-specific CD4+ T cells. Our findings have significant implications for the optimization of TCR gene transfer immunotherapies widely applicable to cancer patients.
CD4+CD25int T Cells in Inflammatory Diseases Refractory to Treatment with Glucocorticoids
Authors: Lee Richard W. J. , Creed Thomas J. , Schewitz Lauren P. , Newcomb Paul V. , Nicholson Lindsay B. , Dick Andrew D. , Dayan Colin M.
Published: April 2014

Up to 30% of patients with autoimmune, allergic, and lymphoproliferative diseases are refractory to glucocorticoid therapy. The present study was undertaken to investigate whether such steroid resistance (SR) is limited to a subpopulation of CD4+ T cells and, as IL-2 is a putative driver of SR, whether T cell SR is associated with CD25 expression. We show that SR patients have a characteristic subgroup of activated CD4+ T cells that continue to proliferate despite exposure to high-dose Dexamethasone (Dex), demonstrate that CD4+CD25− cells are exquisitely sensitive to Dex whereas CD4+CD25int cells are highly SR, and further find that the combination of an anti-CD25 mAb with Dex enhances suppression of T cell proliferation compared with each agent alone. We therefore conclude that SR is not a general property of all lymphocytes but resides in T cell subpopulations, which are prevalent in SR patients and express intermediary levels of CD25. As a result, we propose a new paradigm for SR disease in which glucocorticoid therapy positively selects SR cells, generating a population of drug-resistant lymphocytes that perpetuate on-going inflammation.
Lack of Chromatin and Nuclear Fragmentation In Vivo Impairs the Production of Lupus Anti-Nuclear Antibodies
Authors: Frisoni Lorenza , McPhie Lenese , Kang Sun-Ah , Monestier Marc , Madaio Michael , Satoh Minoru , Caricchio Roberto
Published: April 2014

Nuclear autoantigens in systemic lupus erythematosus are thought to derive primarily from apoptotic cells, yet there is no direct evidence that interfering with apoptosis impairs the generation of lupus autoantibodies. Here we use a mouse model that lacks the endonuclease caspase-activated DNase (CAD), resulting in an absence of chromatin and nuclear fragmentation during apoptotic cell death. We show that in this mouse, production and release into circulation of chromatin is impaired after exposure to several apoptotic triggers, but that the absence of CAD does not interfere with upstream steps of apoptosis or immune system function. Finally we show that in CAD-mutant mice, impaired lupus autoimmunity is skewed toward known cytoplasmic components, and autoimmunity toward membrane autoantigens is preserved, while autoimmunity toward chromatin and other lupus nuclear targets is severely impaired or absent. We also show, as control, that the induction of experimental autoimmune encephalomyelitis is not affected by the absence of CAD. Thus, our work in vivo strongly suggests that apoptotic molecular steps during cell death generate nuclear autoantigens to sustain the specific autoimmune response in systemic lupus erythematosus.
Identification of an Antigenic and Immunogenic Motif Expressed by Two 7-Mer Rituximab-Specific Cyclic Peptide Mimotopes: Implication for Peptide-Based Active Immunotherapy
Authors: Perosa Federico , Favoino Elvira , Vicenti Chiara , Merchionne Francesca , Dammacco Franco
Published: April 2014

Two 7-mer cyclic peptides—Rp15-C and Rp13-C—which bear the antigenic motif recognized by the anti-CD20 mAb rituximab, but have different motif-surrounding amino acids, show a comparable avidity for rituximab and inhibit the binding of rituximab to raft-associated CD20 and rituximab-induced membrane ceramide on human lymphoid Daudi cells. Their immunogenic profiles differed: Abs recognizing CD20 were induced in two and five of five BALB/c mice immunized with Rp15-C and Rp13-C, respectively. Analysis of immunogenic motif, performed by panning a 7-mer phage-display peptide library with purified anti-peptide IgGs, showed that the motif defined by anti-Rp15-C mostly included amino acids surrounding the rituximab-specific antigenic motif , whereas that defined by anti-Rp13-C was . These data indicate that their motif-surrounding amino acids can markedly influence the specificity of Abs, even when elicited with a short 7-mer peptide. Because these anti-peptide Abs are of IgG isotype, their specificity is likely to reflect how peptides are processed at the T cell level and suggest that, within a short peptide, the motifs defined by T cells during the initial phase and upon their stimulation may be different. Our findings may account for the failure of most forms of peptide-based immunotherapy in cancer and autoimmune diseases in which anti-mimotope Abs are expected to play a relevant therapeutic effect. They also suggest strategies to implement the specificity of peptide-induced Abs against the target Ag.
CD28 Deficiency Exacerbates Joint Inflammation upon Borrelia burgdorferi Infection, Resulting in the Development of Chronic Lyme Arthritis
Authors: Iliopoulou Bettina P. , Alroy Joseph , Huber Brigitte T.
Published: April 2014

Lyme disease, caused by the tick-borne spirochete Borrelia burgdorferi (Bb), is a multisystem illness, affecting many organs, such as the heart, the nervous system, and the joints. Months after Bb infection, ∼60% of patients experience intermittent arthritic attacks, a condition that in some individuals progresses to chronic joint inflammation. Although mice develop acute arthritis in response to Bb infection, the joint inflammation clears after 2 wk, despite continuous infection, only very rarely presenting with chronic Lyme arthritis. Thus, the lack of an animal system has so far prevented the elucidation of this persistent inflammatory process that occurs in humans. In this study, we report that the majority of Bb-infected CD28−/− mice develop chronic Lyme arthritis. Consistent with observations in chronic Lyme arthritis patients, the infected mutant, but not wild-type mice present recurring monoarticular arthritis over an extended time period, as well as anti-outer surface protein A of Bb serum titers. Furthermore, we demonstrate that anti-outer surface protein A Abs develop in these mice only after establishment of chronic Lyme arthritis. Thus, the Bb-infected CD28−/− mice provide a murine model for studying chronic Lyme arthritis.
Analysis of Direct and Cross-Presentation of Antigens in TPPII Knockout Mice1
Authors: Firat Elke , Huai Jisen , Saveanu Loredana , Gaedicke Simone , Aichele Peter , Eichmann Klaus , van Endert Peter , Niedermann Gabriele
Published: April 2014

Tripeptidyl peptidase II (TPPII) is an oligopeptidase forming giant complexes in the cytosol that have high exo-, but also, endoproteolytic activity. Immunohistochemically, the complexes appear as distinct foci in the cytosol. In part controversial biochemical and functional studies have suggested that TPPII contributes, on the one hand, positively to Ag processing by generating epitope carboxyl termini or by trimming epitope precursors, and, on the other, negatively by destroying potentially antigenic peptides. To clarify which of these roles is predominant, we generated and analyzed TPPII-deficient mice. Cell surface levels of MHC class I peptide complexes tended to be increased on most cell types of these mice. Although presentation of three individual epitopes derived from lymphocytic choriomeningitis virus was not elevated on TPPII−/− cells, that of the immunodominant OVA epitope SIINFEKL was significantly enhanced. Consistent with this, degradation of a synthetic peptide corresponding to the OVA epitope and of another corresponding to a precursor thereof, both being proteasomally generated OVA fragments, was delayed in TPPII-deficient cytosolic extracts. In addition, dendritic cell cross-presentation of phagocytosed OVA and of OVA internalized as an immune complex was increased to about the same level as direct presentation of the Ag. The data suggest a moderate, predominantly destructive role of TPPII in class I Ag processing, in line with our finding that TPPII is not induced by IFN-γ, which up-regulates numerous, predominantly constructive components of the Ag processing and presentation machinery.
Atypical Memory Phenotype T Cells with Low Homeostatic Potential and Impaired TCR Signaling and Regulatory T Cell Function in Foxn1Δ/Δ Mutant Mice
Authors: Xiao Shiyun , Su Dong-ming , Manley Nancy R.
Published: April 2014

Foxn1 Δ / Δ mutants have a block in thymic epithelial cell differentiation at an intermediate progenitor stage, resulting in reduced thymocyte cellularity and blocks at the double-negative and double-positive stages. Whereas naive single-positive thymocytes were reduced >500-fold in the adult Foxn1Δ/Δ thymus, peripheral T cell numbers were reduced only 10-fold. The current data shows that Foxn1Δ/Δ peripheral T cells had increased expression of activation markers and the ability to produce IL-2 and IFN-γ. These cells acquired this profile immediately after leaving the thymus as early as the newborn stage and maintained high steady-state proliferation in vivo but decreased proliferation in response to TCR stimulation in vitro. Single-positive thymocytes and naive T cells also had constitutively low αβTCR and IL7R expression. These cells also displayed reduced ability to undergo homeostatic proliferation and increased rates of apoptosis. Although the frequency of Foxp3+CD4+CD25+ T cells was normal in Foxn1Δ/Δ mutant mice, these cells failed to have suppressor function, resulting in reduced regulatory T cell activity. Recent data from our laboratory suggest that T cells in the Foxn1Δ/Δ thymus develop from atypical progenitor cells via a noncanonical pathway. Our results suggest that the phenotype of peripheral T cells in Foxn1Δ/Δ mutant mice is the result of atypical progenitor cells developing in an abnormal thymic microenvironment with a deficient TCR and IL7 signaling system.
Critical Role for CXC Chemokine Ligand 16 (SR-PSOX) in Th1 Response Mediated by NKT Cells
Authors: Shimaoka Takeshi , Seino Ken-ichiro , Kume Noriaki , Minami Manabu , Nishime Chiyoko , Suematsu Makoto , Kita Toru , Taniguchi Masaru , Matsushima Kouji , Yonehara Shin
Published: April 2014

The transmembrane chemokine CXCL 16 (CXCL16), which is the same molecule as the scavenger receptor that binds phosphatidylserine and oxidized lipoprotein (SR-PSOX), has been shown to mediate chemotaxis and adhesion of CXC chemokine receptor 6-expressing cells such as NKT and activated Th1 cells. We generated SR-PSOX/CXCL16-deficient mice and examined the role of this chemokine in vivo. The mutant mice showed a reduced number of liver NKT cells, and decreased production of IFN-γ and IL-4 by administration of α-galactosylceramide (αGalCer). Of note, the αGalCer-induced production of IFN-γ was more severely impaired than the production of IL-4 in SR-PSOX-deficient mice. In this context, SR-PSOX-deficient mice showed impaired sensitivity to αGalCer-induced anti-tumor effect mediated by IFN-γ from NKT cells. NKT cells from wild-type mice showed impaired production of IFN-γ, but not IL-4, after their culture with αGalCer and APCs from mutant mice. Moreover, Propionibacterium acnes-induced in vivo Th1 responses were severely impaired in SR-PSOX-deficient as well as NKT KO mice. Taken together, SR-PSOX/CXCL16 plays an important role in not only the production of IFN-γ by NKT cells, but also promotion of Th1-inclined immune responses mediated by NKT cells.
Dendritic Cell Maturation Results in Pronounced Changes in Glycan Expression Affecting Recognition by Siglecs and Galectins
Authors: Bax Marieke , García-Vallejo Juan J. , Jang-Lee Jihye , North Simon J. , Gilmartin Tim J. , Hernández Gilberto , Crocker Paul R. , Leffler Hakon , Head Steven R. , Haslam Stuart M. , Dell Anne , van Kooyk Yvette
Published: April 2014

Dendritic cells (DC) are the most potent APC in the organism. Immature dendritic cells (iDC) reside in the tissue where they capture pathogens whereas mature dendritic cells (mDC) are able to activate T cells in the lymph node. This dramatic functional change is mediated by an important genetic reprogramming. Glycosylation is the most common form of posttranslational modification of proteins and has been implicated in multiple aspects of the immune response. To investigate the involvement of glycosylation in the changes that occur during DC maturation, we have studied the differences in the glycan profile of iDC and mDC as well as their glycosylation machinery. For information relating to glycan biosynthesis, gene expression profiles of human monocyte-derived iDC and mDC were compared using a gene microarray and quantitative real-time PCR. This gene expression profiling showed a profound maturation-induced up-regulation of the glycosyltransferases involved in the expression of LacNAc, core 1 and sialylated structures and a down-regulation of genes involved in the synthesis of core 2 O-glycans. Glycosylation changes during DC maturation were corroborated by mass spectrometric analysis of N- and O-glycans and by flow cytometry using plant lectins and glycan-specific Abs. Interestingly, the binding of the LacNAc-specific lectins galectin-3 and -8 increased during maturation and up-regulation of sialic acid expression by mDC correlated with an increased binding of siglec-1, -2, and -7.
Prolonged (E)-4-Hydroxy-3-Methyl-But-2-Enyl Pyrophosphate-Driven Antimicrobial and Cytotoxic Responses of Pulmonary and Systemic Vγ2Vδ2 T Cells in Macaques
Authors: Ali Zahida , Shao Lingyun , Halliday Lisa , Reichenberg Armin , Hintz Martin , Jomaa Hassan , Chen Zheng W.
Published: April 2014

Although phosphoantigen-specific Vγ2Vδ2 T cells appear to play a role in antimicrobial and anticancer immunity, mucosal immune responses and effector functions of these γδ T cells during infection or phospholigand treatment remain poorly characterized. In this study, we demonstrate that the microbial phosphoantigen (E)-4-hydroxy-3-methyl-but-2-enyl pyrophosphate (HMBPP) plus IL-2 treatment of macaques induced a prolonged major expansion of circulating Vγ2Vδ2 T cells that expressed CD8 and produced cytotoxic perforin during their peak expansion. Interestingly, HMBPP-activated Vγ2Vδ2 T cells underwent an extraordinary pulmonary accumulation, which lasted for 3–4 mo, although circulating Vγ2Vδ2 T cells had returned to baseline levels weeks prior. The Vγ2Vδ2 T cells that accumulated in the lung following HMBPP/IL-2 cotreatment displayed an effector memory phenotype, as follows: CCR5+CCR7−CD45RA−CD27+ and were able to re-recognize phosphoantigen and produce copious amounts of IFN-γ up to 15 wk after treatment. Furthermore, the capacity of massively expanded Vγ2Vδ2 T cells to produce cytokines in vivo coincided with an increase in numbers of CD4+ and CD8+ αβ T cells after HMBPP/IL-2 cotreatment as well as substantial perforin expression by CD3+Vγ2− T cells. Thus, the prolonged HMBPP-driven antimicrobial and cytotoxic responses of pulmonary and systemic Vγ2Vδ2 T cells may confer immunotherapeutics against infectious diseases and cancers.
Biochemical and Folding Defects in a RAG1 Variant Associated with Omenn Syndrome
Authors: Simkus Carrie , Anand Priyanka , Bhattacharyya Anamika , Jones Jessica M.
Published: April 2014

The RAG1 and RAG2 proteins are required to assemble mature Ag receptor genes in developing lymphocytes. Hypomorphic mutations in the gene encoding RAG1 are associated with Omenn syndrome, a primary immunodeficiency. We explored the biochemical defects resulting from a mutation identified in an Omenn syndrome patient which generates an amino acid substitution in the RAG1 RING finger/ubiquitin ligase domain (C325Y in murine RAG1) as well as an adjacent substitution (P326G). RAG1 C325Y demonstrated a 50-fold reduction in recombination activity in cultured pro-B cells despite the fact that its expression and localization to the nucleus were similar to the wild-type protein. The C325Y substitution severely abrogated ubiquitin ligase activity of the purified RAG1 RING finger domain, and the tertiary structure of the domain was altered. The P326G substitution also abrogated ubiquitin ligase activity but had a less severe effect on protein folding. RAG1 P326G also demonstrated a recombination impairment that was most pronounced when RAG1 levels were limiting. Thus, a correctly folded RAG1 RING finger domain is required for normal V(D)J recombination, and RAG1 ubiquitin ligase activity can contribute when the protein is present at relatively low levels.
Analysis of the 5q31–33 Locus Shows an Association between Single Nucleotide Polymorphism Variants in the IL-5 Gene and Symptomatic Infection with the Human Blood Fluke, Schistosoma japonicum
Authors: Ellis Magda K. , Zhao Zhen Zhen , Chen Hong-Gen , Montgomery Grant W. , Li Yue-Sheng , McManus Donald P.
Published: April 2014

Genetic studies of human susceptibility to Schistosoma (blood fluke) infections have previously identified a genetic locus determining infection intensity with the African species, Schistosoma mansoni, in the 5q31–33 region of the human genome that is known to contain the Th2 immune response cluster, including the genes encoding the IL-4, IL-5, and IL-13 cytokines. These cytokines are key players in inflammatory immune responses and have previously been implicated in human susceptibility to infection with the Asian species, S. japonicum. In a nested case control study, we genotyped 30 HapMap tagging single nucleotide polymorphisms (SNPs) across these three genes in 159 individuals identified as putatively susceptible to reinfection with S. japonicum and in 133 putatively resistant individuals. A third group comprising 113 individuals demonstrating symptomatic infection was also included. The results provided no significant association at a global level between reinfection predisposition and any of the individual SNPs or haplotype blocks. However, two tagging SNPs in IL-5 demonstrated globally significant association with susceptibility to symptomatic infection. They were in strong linkage disequilibrium with each other and were found to belong to the same haplotype block that also provided a significant association after permutation testing. This haplotype was located in the 3′-untranslated region of IL-5, suggesting that variants in this region of IL-5 may modulate the immune response in these individuals with symptomatic infection.
Mycobacterium tuberculosisAntigens Specifically Modulate CCR2 and MCP-1/CCL2 on Lymphoid Cells from Human Pulmonary Hilar Lymph Nodes
Authors: Arias Mauricio A. , Jaramillo Gabriela , López Yúrika P. , Mejía Natalia , Mejía Camila , Pantoja Adelis E. , Shattock Robin J. , García Luis F. , Griffin George E.
Published: April 2014

Macrophages and dendritic cells are involved in the immune response to Mycobacterium tuberculosis (Mtb). Such a response, although extensively studied using animal models and cells from human blood, has not been characterized in cells from pulmonary hilar lymph nodes (PHLN). We characterized populations of myeloid APC from PHLN and determined their expression of CCR2, CCR5, CCR7, CD40, CD54, CD80, and CD86 as well as the cytokine/chemokine microenvironment before and after purified protein derivative (PPD) and mannosilated lipoarabinomannan (ManLAM) stimulation. Results show that there are at least three APC populations in PHLN, defined as CD14highHLA-DRlow/−, CD14dimHLA-DRdim, and CD14−HLA-DRhigh/dendritic cells (DC), with the largest number represented by CD14dimHLA-DRdim cells (where dim indicates intermediate levels). CD14−HLA-DRhigh/DC expressed higher levels of costimulatory molecules and lower levels of CCR2 and CCR5, but all cell populations showed similar CCR7 levels. PPD and ManLAM specifically down-regulated CCR2 expression but not that of CCR5 and CCR7, and such down-regulation was observed on all APC populations. Mtb Ag did not affect the expression of costimulatory molecules. PPD but not ManLAM specifically induced MCP-1/CCL2 production, which was likely associated with the induction of IFN-γ because this cytokine was highly induced by PPD. We characterized, for the first time, different APC from human PHLN and show that Mtb Ag exert fine and specific regulation of molecules closely associated with the immune response to Mtb infection. Because knowledge of this response in secondary lymphoid tissues is still poorly understood in humans, such studies are necessary and important for a better understanding of lymphoid cell microenvironment and migrating capacities and their role in the immunopathogenesis of tuberculosis.
Blockade of Chemokine Receptor CXCR3 Inhibits T Cell Recruitment to Inflamed Joints and Decreases the Severity of Adjuvant Arthritis
Authors: Mohan Karkada , Issekutz Thomas B.
Published: April 2014

T lymphocytes expressing the chemokine receptors, CCR2, CCR5, CXCR3, and CXCR6 are increased in inflamed tissues in rheumatoid arthritis. The role of CXCR3 in autoimmune arthritis induced in Lewis rats was investigated. CXCR3+ T cells migrated 2- to 3-fold more than CXCR3− T cells to inflamed joints in arthritic animals. CXCR3-expressing in vivo Ag-activated T lymphoblasts and in vitro-activated lymph node cells from arthritic animals were strongly recruited to the arthritic joints, and treatment with anti-CXCR3 mAb significantly inhibited this T cell recruitment by 40–60%. Immune T cells from the spleen and lymph nodes of actively immunized arthritic donors adoptively transferred arthritis to naive rats. Treatment with anti-CXCR3 mAb delayed the onset of arthritis and significantly reduced the severity of joint inflammation with a >50% decrease in the clinical arthritis score. Blockade of CXCR3 also significantly reduced the weight loss in the arthritic animals and inhibited neutrophil accumulation in the joints by 50–60%. There was a marked reduction in the leukocyte infiltration of the synovium in the presence of CXCR3 blockade and a decrease in the loss of articular cartilage of the joints. In conclusion, CXCR3 on T cells has an essential role in T cell recruitment to inflamed joints and the development of joint inflammation in adjuvant arthritis.
Selective Proinflammatory Activation of Astrocytes by High-Mobility Group Box 1 Protein Signaling
Authors: Pedrazzi Marco , Patrone Mauro , Passalacqua Mario , Ranzato Elia , Colamassaro Diego , Sparatore Bianca , Pontremoli Sandro , Melloni Edon
Published: April 2014

Extracellular high-mobility group box 1 protein (HMGB1) triggers inflammatory events in the brain. We demonstrate that astrocytes, the main glial cells in the brain, acquire a specific reactive phenotype when exposed to HMGB1. This cell activation, which involves the receptor for advanced glycation end-products and the MAPK/ERK1/2 cascade, results in the transcriptional/translational induction of a restricted number of inflammatory mediators, including cyclooxygenase-2, matrix metalloproteinase-9, and several chemokines of the CC and CXC families. The mixture of factors released by HMGB1-reactive astrocytes displays a potent chemotactic activity on human monocytic cells. This study is the first to suggest that HMGB1/astrocyte interaction plays a specific functional role in the progression of inflammatory processes in the CNS by facilitating local leukocyte infiltration.
Inhibition of Neutrophil Apoptosis by ATP Is Mediated by the P2Y11 Receptor
Authors: Vaughan Kathryn R. , Stokes Leanne , Prince Lynne R. , Marriott Helen M. , Meis Sabine , Kassack Matthias U. , Bingle Colin D. , Sabroe Ian , Surprenant Annmarie , Whyte Moira K. B.
Published: April 2014

Neutrophils undergo rapid constitutive apoptosis that is delayed by a range of pathogen- and host-derived inflammatory mediators. We have investigated the ability of the nucleotide ATP, to which neutrophils are exposed both in the circulation and at sites of inflammation, to modulate the lifespan of human neutrophils. We found that physiologically relevant concentrations of ATP cause a concentration-dependent delay of neutrophil apoptosis (assessed by morphology, annexin V/To-Pro3 staining, and mitochondrial membrane permeabilization). We found that even brief exposure to ATP (10 min) was sufficient to cause a long-lasting delay of apoptosis and showed that the effects were not mediated by ATP breakdown to adenosine. The P2 receptor mediating the antiapoptotic actions of ATP was identified using a combination of more selective ATP analogs, receptor expression studies, and study of downstream signaling pathways. Neutrophils were shown to express the P2Y11 receptor and inhibition of P2Y11 signaling using the antagonist NF157 abrogated the ATP-mediated delay of neutrophil apoptosis, as did inhibition of type I cAMP-dependent protein kinases activated downstream of P2Y11, without effects on constitutive apoptosis. Specific targeting of P2Y11 could retain key immune functions of neutrophils but reduce the injurious effects of increased neutrophil longevity during inflammation.
Tumor-Associated E-Cadherin Mutations Affect Binding to the Killer Cell Lectin-Like Receptor G1 in Humans
Authors: Schwartzkopff Sabrina , Gründemann Carsten , Schweier Oliver , Rosshart Stephan , Karjalainen Klaus Erik , Becker Karl-Friedrich , Pircher Hanspeter
Published: April 2014

The killer cell lectin-like receptor G1 (KLRG1) is expressed by NK cells and memory T cells in man and mice. Cadherins were recently identified as ligands for mouse KLRG1 but ligands for human KLRG1 have not yet been defined. In this study, we first demonstrate that human E-cadherin is a ligand for human KLRG1. This finding is remarkable because human and mouse KLRG1 show only an intermediate degree of homology (57% aa identity). In addition, we show that E-cadherin, expressed on K562 target cells, inhibited polyclonal human NK cells. Inhibition of NK cell function was observed consistently in three independent functional assays but the extent of inhibition was modest and required high expression of E-cadherin on target cells. E-cadherin function is often inactivated during development of human carcinomas and splice-site mutations resulting in in-frame loss of exon 8 or 9 occur frequently in diffuse type gastric carcinomas. Our experiments further revealed that interaction of human KLRG1 to E-cadherin was susceptible to these tumor-associated mutations and that KLRG1+ NK cells were triggered more easily by K562 target cells carrying these mutations in comparison to target cells expressing wild-type E-cadherin. These results also indicate that the E-cadherin binding sites important for homophilic interaction are also involved in KLRG1 binding. Taken together, these data demonstrate that the main adhesion molecule of epithelial tissue, E-cadherin, is involved in regulation of NK cells in both humans and mice.
Membranal Cholesterol Is Not Required for L-Selectin Adhesiveness in Primary Lymphocytes but Controls a Chemokine-Induced Destabilization of L-Selectin Rolling Adhesions
Authors: Dwir Oren , Grabovsky Valentin , Pasvolsky Ronit , Manevich Eugenia , Shamri Revital , Gutwein Paul , Feigelson Sara W. , Altevogt Peter , Alon Ronen
Published: April 2014

Cholesterol-enriched lipid microdomains regulate L-selectin signaling, but the role of membrane cholesterol in L-selectin adhesion is unclear. Arrest chemokines are a subset of endothelial chemokines that rapidly activate leukocyte integrin adhesiveness under shear flow. In the absence of integrin ligands, these chemokines destabilize L-selectin-mediated leukocyte rolling. In the present study, we investigated how cholesterol extraction from the plasma membrane of peripheral blood T or B cells affects L-selectin adhesions and their destabilization by arrest chemokines. Unlike the Jurkat T cell line, whose L-selectin-mediated adhesion is cholesterol dependent, in primary human PBLs and in murine B cells and B cell lines, cholesterol depletion did not impair any intrinsic adhesiveness of L-selectin, consistent with low selectin partitioning into lipid rafts in these cells. However, cholesterol raft disruption impaired the ability of two arrest chemokines, CXCL12 and CXCL13, but not of a third arrest chemokine, CCL21, to destabilize L-selectin-mediated rolling of T lymphocytes. Actin capping by brief incubation with cytochalasin D impaired the ability of all three chemokines to destabilize L-selectin rolling. Blocking of the actin regulatory phosphatidylinositol lipid, phosphatidylinositol 4,5-bisphosphate, did not affect chemokine-mediated destabilization of L-selectin adhesions. Collectively, our results suggest that L-selectin adhesions are inhibited by actin-associated, cholesterol-stabilized assemblies of CXCL12- and CXCL13-binding receptors on both T and B lymphocytes. Thus, the regulation of L-selectin by cholesterol-enriched microdomains varies with the cell type as well as with the identity of the destabilizing chemokine.
Chronic Lymphocytic Choriomeningitis Virus Infection Actively Down-Regulates CD4+ T Cell Responses Directed against a Broad Range of Epitopes
Authors: Mothé Bianca R. , Stewart Barbara S. , Oseroff Carla , Bui Huynh-Hoa , Stogiera Stephanie , Garcia Zacarias , Dow Courtney , Rodriguez-Carreno Maria Pilar , Kotturi Maya , Pasquetto Valerie , Botten Jason , Crotty Shane , Janssen Edith , Buchmeier Michael J. , Sette Alessandro
Published: April 2014

Activation of CD4+ T cells helps establish and sustain CD8+ T cell responses and is required for the effective clearance of acute infection. CD4-deficient mice are unable to control persistent infection and CD4+ T cells are usually defective in chronic and persistent infections. We investigated the question of how persistent infection impacted pre-existing lymphocytic choriomeningitis virus (LCMV)-specific CD4+ T cell responses. We identified class II-restricted epitopes from the entire set of open reading frames from LCMV Armstrong in BALB/c mice (H-2d) acutely infected with LCMV Armstrong. Of nine epitopes identified, six were restricted by I-Ad, one by I-Ed and two were dually restricted by both I-Ad and I-Ed molecules. Additional experiments revealed that CD4+ T cell responses specific for these epitopes were not generated following infection with the immunosuppressive clone 13 strain of LCMV. Most importantly, in peptide-immunized mice, established CD4+ T cell responses to these LCMV CD4 epitopes as well as nonviral, OVA-specific responses were actively suppressed following infection with LCMV clone 13 and were undetectable within 12 days after infection, suggesting an active inhibition of established helper responses. To address this dysfunction, we performed transfer experiments using both the Smarta and OT-II systems. OT-II cells were not detected after clone 13 infection, indicating physical deletion, while Smarta cells proliferated but were unable to produce IFN-γ, suggesting impairment of the production of this cytokine. Thus, multiple mechanisms may be involved in the impairment of helper responses in the setting of early persistent infection.
CTCF-Independent, but Not CTCF-Dependent, Elements Significantly Contribute to TCR-α Locus Control Region Activity
Authors: Gomos-Klein Janette , Harrow Faith , Alarcón Jemma , Ortiz Benjamin D.
Published: April 2014

The mouse TCRα/TCRδ/Dad1 gene locus bears a locus control region (LCR) that drives high-level, position-independent, thymic transgene expression in chromatin. It achieves this through DNA sequences that enhance transcription and protect transgene expression from integration site-dependent position effects. The former activity maps to a classical enhancer region (Eα). In contrast, the elements supporting the latter capacity that suppresses position effects are incompletely understood. Such elements likely play important roles in their native locus and may resemble insulator/boundary sequences. Insulators support enhancer blocking and/or chromatin barrier activity. Most vertebrate enhancer-blocking insulators are dependent on the CTCF transcription factor and its cognate DNA binding site. However, studies have also revealed CTCF-independent enhancer blocking and barrier insulator activity in the vertebrate genome. The TCRα LCR contains a CTCF-dependent and multiple CTCF-independent enhancer-blocking regions whose roles in LCR activity are unknown. Using randomly integrated reporter transgenes in mice, we find that the CTCF region plays a very minor role in LCR function. In contrast, we report the in vivo function of two additional downstream elements located in the region of the LCR that supports CTCF-independent enhancer-blocking activity in cell culture. Internal deletion of either of these elements significantly impairs LCR activity. These results reveal that the position-effect suppression region of the TCRα LCR harbors an array of CTCF-independent, positive-acting gene regulatory elements, some of which share characteristics with barrier-type insulators. These elements may help manage the separate regulatory programs of the TCRα and Dad1 genes.
A Costimulation-Initiated Signaling Pathway Regulates NFATc1 Transcription in T Lymphocytes
Authors: Nurieva Roza I. , Chuvpilo Sergei , Wieder Eric D. , Elkon Keith B. , Locksley Richard , Serfling Edgar , Dong Chen
Published: April 2014

T cell activation and differentiation is accompanied and mediated by transcriptional reprogramming. The NFATc1 transcription factor is strongly induced upon T cell activation and controls numerous genes involved in the T cell effector function. However, its regulation by physiological stimuli in primary T cells has not been well understood. We previously found that ICOS synergizes with TCR and CD28 to greatly enhance NFATc1 expression in primary T cells. In this study, we have examined the signaling mechanisms whereby costimulation regulates NFATc1 expression. We found that CD28 and ICOS regulate sustained PI3K activity in primary T cells, which is required for NFATc1 up-regulation. CD28 and ICOS costimulation, possibly through Itk, a Tec kinase downstream of the PI3K, enhanced phosphorylation of phospholipase Cγ1 and increased and sustained Ca2+ flux in T cells. Costimulation of T cells potentiated transcription of the Nfatc1 gene P1 promoter in a PI3K-dependent manner. This work demonstrates an important role for costimulatory receptors in sustaining T cell activation programs leading to Nfatc1 gene transcription and has implications in our understanding of the immune response and tolerance.
Activation of TLR2 and TLR4 by Glycosylphosphatidylinositols Derived from Toxoplasma gondii
Authors: Debierre-Grockiego Françoise , Campos Marco A. , Azzouz Nahid , Schmidt Jörg , Bieker Ulrike , Resende Marianne Garcia , Mansur Daniel Santos , Weingart Ralf , Schmidt Richard R. , Golenbock Douglas T. , Gazzinelli Ricardo T. , Schwarz Ralph T.
Published: April 2014

GPIs isolated from Toxoplasma gondii, as well as a chemically synthesized GPI lacking the lipid moiety, activated a reporter gene in Chinese hamster ovary cells expressing TLR4, while the core glycan and lipid moieties cleaved from the GPIs activated both TLR4- and TLR2-expressing cells. MyD88, but not TLR2, TLR4, or CD14, is absolutely needed to trigger TNF-α production by macrophages exposed to T. gondii GPIs. Importantly, TNF-α response to GPIs was completely abrogated in macrophages from TLR2/4-double-deficient mice. MyD88−/− mice were more susceptible to death than wild-type (WT), TLR2−/−, TLR4−/−, TLR2/4−/−, and CD14−/− mice infected with the ME-49 strain of T. gondii. The cyst number was higher in the brain of TLR2/4−/−, but not TLR2−/−, TLR4−/−, and CD14−/−, mice, as compared with WT mice. Upon infection with the ME-49 strain of T. gondii, we observed no decrease of IL-12 and IFN-γ production in TLR2-, TLR4-, or CD14-deficient mice. Indeed, splenocytes from T. gondii-infected TLR2−/− and TLR2/4−/− mice produced more IFN-γ than cells from WT mice in response to in vitro stimulation with parasite extracts enriched in GPI-linked surface proteins. Together, our results suggest that both TLR2 and TLR4 receptors may participate in the host defense against T. gondii infection through their activation by the GPIs and could work together with other MyD88-dependent receptors, like other TLRs or even IL-18R or IL-1R, to obtain an effective host response against T. gondii infection.
The Involvement of CD44 and Its Novel Ligand Galectin-8 in Apoptotic Regulation of Autoimmune Inflammation
Authors: Eshkar Sebban Lora , Ronen Denise , Levartovsky David , Elkayam Ori , Caspi Dan , Aamar Suhail , Amital Howard , Rubinow Alan , Golan Ira , Naor David , Zick Yehiel , Golan Itshak
Published: April 2014

The synovial fluid (SF) cells of rheumatoid arthritis (RA) patients express a specific CD44 variant designated CD44vRA. Using a cellular model of this autoimmune disease, we show in this study that the mammalian lectin, galectin-8 (gal-8), is a novel high-affinity ligand of CD44vRA. By affinity chromatography, flow cytometry, and surface plasmon resonance, we demonstrate that gal-8 interacts with a high affinity (Kd, 6 × 10−9 M) with CD44vRA. We further demonstrate that SF cells from RA patients express and secrete gal-8, to a concentration of 25–65 nM, well within the concentration of gal-8 required to induce apoptosis of SF cells. We further show that not all gal-8 remains freely soluble in the SF and at least part forms triple complexes with CD44 and fibrinogen that can be detected, after fibrinogen immunoprecipitation, with Abs against fibrinogen, gal-8 and CD44. These triple complexes may therefore increase the inflammatory reaction by sequestering the soluble gal-8, thereby reducing its ability to induce apoptosis in the inflammatory cells. Our findings not only shed light on the receptor-ligand relationships between CD44 and gal-8, but also underline the biological significance of these interactions, which may affect the extent of the autoimmune inflammatory response in the SF of RA patients.
The Anti-inflammatory Effects of Heat Shock Protein 72 Involve Inhibition of High-Mobility-Group Box 1 Release and Proinflammatory Function in Macrophages
Authors: Tang Daolin , Kang Rui , Xiao Weimin , Wang Haichao , Calderwood Stuart K. , Xiao Xianzhong
Published: April 2014

High-mobility-group box 1 (HMGB1), a nuclear protein, has recently been identified as an important mediator of local and systemic inflammatory diseases when released into the extracellular milieu. Anti-inflammatory regulation by the stress response is an effective autoprotective mechanism when the host encounters harmful stimuli, but the mechanism of action remains incompletely delineated. In this study, we demonstrate that increases in levels of a major stress-inducible protein, heat shock protein 72 (Hsp72) by gene transfection attenuated LPS- or TNF-α-induced HMGB1 cytoplasmic translocation and release. The mechanisms involved inhibition of the chromosome region maintenance 1 (CRM1)-dependent nuclear export pathway. Overexpression of Hsp72 inhibited CRM1 translocation and interaction between HMGB1 and CRM1 in macrophages post-LPS and TNF-α treatment. In addition, overexpression of Hsp72 strongly inhibited HMGB1-induced cytokine (TNF-α, IL-1β) expression and release, which correlated closely with: 1) inhibition of the MAP kinases (p38, JNK, and ERK); and 2) inhibition of the NF-κB pathway. Taken together, these experiments suggest that the anti-inflammatory activity of Hsp72 is achieved by interfering with both the release and proinflammatory function of HMGB1. Our experimental data provide important insights into the anti-inflammatory mechanisms of heat shock protein protection.
Eotaxin-1/CC Chemokine Ligand 11: A Novel Eosinophil Survival Factor Secreted by Human Pulmonary Artery Endothelial Cells
Authors: Farahi Neda , Cowburn Andrew S. , Upton Paul D. , Deighton John , Sobolewski Anastasia , Gherardi Ermanno , Morrell Nicholas W. , Chilvers Edwin R.
Published: April 2014

Airway eosinophilia plays a major role in the pathogenesis of asthma with the inhibition of apoptosis by GM-CSF and IL-5 proposed as a mechanism underlying prolonged eosinophil survival. In vivo and ex vivo studies have indicated the capacity of interventions that drive human eosinophil apoptosis to promote the resolution of inflammation. Far less is known about the impact of transendothelial migration on eosinophil survival, in particular, the capacity of endothelial cell-derived factors to contribute toward the apoptosis-resistant phenotype characteristic of airway-resident eosinophils. We examined the effects of conditioned medium from human pulmonary artery endothelial cells (HPAEC-CM) on eosinophil apoptosis in vitro. HPAEC-CM inhibited eosinophil, but not neutrophil apoptosis. This effect was specific to HPAECs and comparable in efficacy to the survival effects of GM-CSF and IL-5. The HPAEC survival factor was shown, on the basis of GM-CSF, IL-5, and IL-3 detection assays, Ab neutralization, and sensitivity to PI3K inhibition, to be clearly discrete from these factors. Gel filtration of HPAEC-CM revealed a peak of eosinophil survival activity at 8–12 kDa, and PCR confirmed the presence of mRNA for CCL5, CCL11, CCL24, CCL26, and CCL27 in the HPAECs. The CCR3 antagonist GW782415 caused a major inhibition of the HPAEC-CM-induced survival effect, and Ab neutralization of individual CCR3 chemokines revealed CCL11 as the major survival factor present in the HPAEC-CM. Furthermore, chemokine Ab arrays demonstrated up-regulation of CCL11 in HPAEC-CM. These data demonstrate the capacity of HPAECs to generate CCR3 agonists and the ability of CCL11 to inhibit human eosinophil apoptosis.
Maternal Transmission of Resistance to Development of Allergic Airway Disease
Authors: Matson Adam P. , Zhu Li , Lingenheld Elizabeth G. , Schramm Craig M. , Clark Robert B. , Selander Dawn M. , Thrall Roger S. , Breen Elena , Puddington Lynn
Published: April 2014

Parental phenotype is known to influence the inheritance of atopic diseases, such as allergic asthma, with a maternal history being a more significant risk factor for progeny than paternal history. We hypothesized that recall Th1- or Th2-type immune responses during pregnancy would result in transfer of maternal factors that would differentially impact development of immune responsiveness in offspring. Following weaning, susceptibility and severity of allergic airway disease (a murine model of human asthma) was evaluated in progeny, disease being elicited by immunization with OVA-Al(OH)3 and challenge with aerosolized OVA. We found that progeny of mothers with Th1-biased immunity to OVA subjected to recall aerosol challenge during pregnancy had reduced levels of Ag-specific IgE and airway eosinophilia compared with progeny of mothers with Th2-biased immunity to OVA or naive mothers. Interestingly, progeny of mothers with Th1-type immunity to a heterologous albumin, BSA, were not protected from developing OVA-induced allergic airway disease. These findings demonstrated that maternal transfer of protection from development of allergic airway disease to offspring in this model of maternal Th1-type immunity was Ag specific.