bluuebunny/crossref_metadata_2025_split huggingface dataset:
Distal Enhancer Elements Transcribe Intergenic RNA in the IL-10 Family Gene Cluster
Authors: Jones Elizabeth A. , Flavell Richard A. 
Published: April 2014

Abstract
The IL-10 gene and homologs IL-19, IL-20, and IL-24 are expressed within a highly conserved 145-kb cytokine gene cluster. Like the Th2 IL-4 cytokine gene cluster, it is feasible that there is coordinate regulation of these cytokines by distal regulatory elements spanning the locus. We initiated a search to characterize regulatory elements within the IL-10 family locus and present data herein on a conserved 40-kb region between the IL-19 and IL-10 genes. We map the location of 17 DNase I-hypersensitive sites in different murine T cell populations and identify three enhancer elements, which function in T cells in vitro. Two of these enhancer elements, located 9 kb upstream and 6.45 kb downstream of IL-10, display cell-specific function in the Th1-Th2 cell clones AE7 and D10 and also exhibit basic promoter activity. The downstream element, IL-10CNS+6.45, binds AP-1 in the absence of NFAT and expresses intergenic RNA in a Th2-specific manner, further validating its role as a Th2-specific enhancer/promoter element. We show that the five most highly conserved noncoding sequences in the 40-kb region transcribe intergenic RNA; four of these regions possess promoter activity in vitro that could account for the expression of these transcripts. Hence, we speculate that these novel regulatory elements in the IL-10 family gene locus function via an intermediate regulatory RNA.
Human Cytomegalovirus Encodes an MHC Class I-Like Molecule (UL142) That Functions to Inhibit NK Cell Lysis
Authors: Wills Mark R. , Ashiru Omodele , Reeves Matthew B. , Okecha Georgina , Trowsdale John , Tomasec Peter , Wilkinson Gavin W. G. , Sinclair John , Sissons J. G. Patrick 
Published: April 2014

Abstract
Clinical and low passage strains of human CMV (HCMV) encode an additional MHC class I-related molecule UL142, in addition to the previously described UL18. The UL142 open reading frame is encoded within the ULb′ region which is missing from a number of common high passage laboratory strains. Cells expressing UL142 following transfection, and fibroblasts infected with a recombinant adenovirus-expressing UL142, were used to screen both polyclonal NK cells and NK cell clones, in a completely autologous system. Analysis of 100 NK cell clones derived from five donors, revealed 23 clones that were inhibited by fibroblasts expressing UL142 alone. Small-interfering RNA-mediated knockdown of UL142 mRNA expression in HCMV-infected cells resulted in increased sensitivity to lysis. From these data we conclude that UL142 is a novel HCMV-encoded MHC class I-related molecule which inhibits NK cell killing in a clonally dependent manner.
Heterogeneous Human NK Cell Responses to Plasmodium falciparum-Infected Erythrocytes
Authors: Korbel Daniel S. , Newman Kirsty C. , Almeida Catarina R. , Davis Daniel M. , Riley Eleanor M. 
Published: April 2014

Abstract
Human NK cells can respond rapidly to Plasmodium falciparum-infected RBC (iRBC) to produce IFN-γ. In this study, we have examined the heterogeneity of this response among malaria-naive blood donors. Cells from all donors become partially activated (up-regulating CD69, perforin, and granzyme) upon exposure to iRBC but cells from only a subset of donors become fully activated (additionally up-regulating CD25, IFN-γ, and surface expression of lysosomal-associated membrane protein 1 (LAMP-1)). Although both CD56dim and CD56bright NK cell populations can express IFN-γ in response to iRBC, CD25 and LAMP-1 are up-regulated only by CD56dim NK cells and CD69 is up-regulated to a greater extent in this subset; by contrast, perforin and granzyme A are preferentially up-regulated by CD56bright NK cells. NK cells expressing IFN-γ in response to iRBC always coexpress CD69 and CD25 but rarely LAMP-1, suggesting that individual NK cells respond to iRBC either by IFN-γ production or cytotoxicity. Furthermore, physical contact with iRBC can, in a proportion of donors, lead to NK cell cytoskeletal reorganization suggestive of functional interactions between the cells. These observations imply that individuals may vary in their ability to mount an innate immune response to malaria infection with obvious implications for disease resistance or susceptibility.
Coordinate Expression of CC Chemokine Ligand 5, Granulysin, and Perforin in CD8+ T Cells Provides a Host Defense Mechanism againstMycobacterium tuberculosis
Authors: Stegelmann Frank , Bastian Max , Swoboda Kay , Bhat Rauf , Kiessler Viviane , Krensky Alan M. , Roellinghoff Martin , Modlin Robert L. , Stenger Steffen 
Published: April 2014

AbstractThe ability of CD8+ T cells to kill intracellular pathogens depends upon their capacity to attract infected cells as well as their secretion of cytolytic and antimicrobial effector molecules. We examined the Ag-induced expression of three immune effector molecules contained within cytoplasmic granules of human CD8+ T cells: the chemokine CCL5, the cytolytic molecule perforin, and the antimicrobial protein granulysin. Macrophages infected with virulent Mycobacterium tuberculosis triggered the expression of CCL5 in CD8+ T cells only in donors with previous exposure to the tuberculosis bacteria, not in naive donors. Functionally, CCL5 efficiently attracted M. tuberculosis-infected macrophages, but failed to exert direct antibacterial activity. Infected macrophages also triggered the expression of granulysin in CD8+ T cells, and granulysin was found to be highly active against drug-susceptible and drug-resistant M. tuberculosis clinical isolates. The vast majority of CCL5-positive cells coexpressed granulysin and perforin. Taken together, this report provides evidence that a subset of CD8+ T cells coordinately expresses CCL5, perforin and granulysin, thereby providing a host mechanism to attract M. tuberculosis-infected macrophages and kill the intracellular pathogen.
Toll-IL-1 Receptor Domain-Containing Adaptor Protein Is Critical for Early Lung Immune Responses againstEscherichia coliLipopolysaccharide and ViableEscherichia coli
Authors: Jeyaseelan Samithamby , Manzer Rizwan , Young Scott K. , Yamamoto Masahiro , Akira Shizuo , Mason Robert J. , Worthen G. Scott 
Published: April 2014

AbstractPulmonary bacterial diseases are a leading cause of mortality in the U.S. Innate immune response is vital for bacterial clearance from the lung, and TLRs play a critical role in this process. Toll-IL-1R domain-containing adaptor protein (TIRAP) is a key molecule in the TLR4 and 2 signaling. Despite its potential importance, the role of TIRAP-mediated signaling in lung responses has not been examined. Our goals were to determine the role of TIRAP-dependent signaling in the induction of lung innate immune responses against Escherichia coli LPS and viable E. coli, and in lung defense against E. coli in mice. LPS-induced neutrophil sequestration; NF-κB translocation; keratinocyte cell-derived chemokine, MIP-2, TNF-α, and IL-6 expression; histopathology; and VCAM-1 and ICAM-1 expression were abolished in the lungs of TIRAP−/− mice. A cell-permeable TIRAP blocking peptide attenuated LPS-induced lung responses. Furthermore, immune responses in the lungs of TIRAP−/− mice were attenuated against E. coli compared with TIRAP+/+ mice. TIRAP−/− mice also had early mortality, higher bacterial burden in the lungs, and more bacterial dissemination following E. coli inoculation. Moreover, we used human alveolar macrophages to examine the role of TIRAP signaling in the human system. The TIRAP blocking peptide abolished LPS-induced TNF-α, IL-6, and IL-8 expression in alveolar macrophages, whereas it attenuated E. coli-induced expression of these cytokines and chemokines. Taken together, this is the first study illustrating the crucial role of TIRAP in the generation of an effective early immune response against E. coli LPS and viable E. coli, and in lung defense against a bacterial pathogen.
A Model Vaccine Exploiting Fungal Mannosylation to Increase Antigen Immunogenicity
Authors: Lam Jennifer S. , Mansour Michael K. , Specht Charles A. , Levitz Stuart M. 
Published: April 2014

AbstractAg mannosylation represents a promising strategy to augment vaccine immunogenicity by targeting Ag to mannose receptors (MRs) on dendritic cells. Because fungi naturally mannosylate proteins, we hypothesized that Ags engineered in fungi would have an enhanced capacity to stimulate T cell responses. Using the model Ag OVA, we generated proteins that differentially expressed N- and O-linked mannosylation in the yeast Pichia pastoris and compared them to their unglycosylated counterparts produced in Escherichia coli. We found that yeast-derived OVA proteins containing N-linkages, extensive O-linkages, or both were more potent than the unmannosylated Ags at inducing OVA-specific CD4+ T cell proliferation. This elevated response to fungal Ags was inhibited by mannan, suggesting involvement of MRs. However, the macrophage MR (CD206) was not essential, because macrophage MR-deficient dendritic cells were fully competent in presenting yeast-derived OVA Ags. Thus, the use of fungal glycosylation to provide N-linked and/or extensive O-linked mannosylation increased the capacity of the model Ag OVA to stimulate Ag-specific T cell responses in an MR-dependent manner. These data have implications for vaccine design by providing proof of principle that yeast-derived mannosylation can enhance immunogenicity.
The Impairment of CD8 Responses Limits the Selection of Escape Mutations in Acute Hepatitis C Virus Infection
Authors: Urbani Simona , Amadei Barbara , Cariani Elisabetta , Fisicaro Paola , Orlandini Alessandra , Missale Gabriele , Ferrari Carlo 
Published: April 2014

Abstract
Evasion from protective CD8 responses by mutations within immunodominant epitopes represents a potential strategy of HCV persistence. To investigate the pathogenetic relevance of this mechanism, a careful search for immunodominant CD8 epitopes was conducted in six patients with chronic evolution of HCV infection by analyzing their global CD8 response with a panel of overlapping synthetic peptides covering the overall HCV sequence and by studying the CD8 frequency by tetramer staining. Immunodominant responses were followed longitudinally from the time of acute onset in relation to the evolution of the epitopic sequences. Although intensity of CD8 responses and frequency of HCV-specific CD8 cells declined over time in all patients, mutations emerged in only three of the six acute patients studied. Variant sequences were less efficiently recognized by CD8 cells than parental epitopes and were poorly efficient in inducing a CD8 response in vitro. CD8 epitopes undergoing mutations were targeted by high avidity CD8 cells more efficient in effector function. Our data support the view that immunodominant CD8 responses are affected by inhibitory mechanisms operating early after infection and that the emergence of escape mutations represents an additional mechanism of virus evasion from those CD8 responses that are functionally preserved.
Lung NF-κB Activation and Neutrophil Recruitment Require IL-1 and TNF Receptor Signaling during Pneumococcal Pneumonia
Authors: Jones Matthew R. , Simms Benjamin T. , Lupa Michal M. , Kogan Mariya S. , Mizgerd Joseph P. 
Published: April 2014

AbstractPulmonary inflammation is an essential component of the host defense against Streptococcus pneumoniae infection of the lungs. The early response cytokines, TNF-α and IL-1, are rapidly induced upon microbial exposure. Mice deficient in all TNF- and IL-1-dependent signaling receptors were used to determine the roles of these cytokines during pneumococcal pneumonia. The deficiency of signaling receptors for TNF and IL-1 decreased bacterial clearance. Neutrophil recruitment to alveolar air spaces was impaired by receptor deficiency, as was pulmonary expression of the neutrophil chemokines KC and MIP-2. Because NF-κB mediates the expression of both chemokines, we assessed NF-κB activation in the lungs. During pneumococcal pneumonia, NF-κB proteins translocate to the nucleus and activate gene expression; these functions were largely abrogated by the deficiency of receptors for TNF-α and IL-1. Thus, the combined deficiency of TNF and IL-1 signaling reduces innate immune responses to S. pneumoniae in the lungs, probably due to essential roles for these receptors in activating NF-κB.
Diversity in the Acute CD8 T Cell Response to Vaccinia Virus in Humans
Authors: Jing Lichen , Chong Tiana M. , McClurkan Christopher L. , Huang Jay , Story Brian T. , Koelle David M. 
Published: April 2014

AbstractOrthopoxviruses have complex proteomes. Infection provokes a brisk CD8 response, which is required in some systems for recovery from primary infection. Little is known concerning the Ags and epitopes recognized by CD8 T cells. We examined the fine specificity of cloned and bulk human vaccinia-specific CD8 CTL by expressing polypeptide fragments from a library of vaccinia genomic DNA. This epitope discovery method emphasizes virus-specific biological activity, as the responder cells are all reactive with whole vaccinia virus. Sixteen novel epitopes, restricted by several HLA A and B alleles, were defined to the nomamer peptide level in diverse vaccinia open reading frames. An additional seven epitope were mapped to short regions of vaccinia proteins. Targets of the CD8 response included proteins assigned to structural, enzymatic, transcription factor, and immune evasion functions, and included members of all viral kinetic classes. Most epitopes were conserved in other orthopoxviruses. Responses to at least 18 epitopes were detected within a single blood sample, revealing a surprising degree of diversity. These epitopes will be useful in natural history studies of CD8 responses to vaccinia, a nonpersisting virus with long-term memory, and in the design and evaluation of attenuated and replication-incompetent vaccinia strains being tested for variola and monkeypox prevention and for the delivery of heterologous Ags.
Cationic Polypeptides Are Required for Anti-HIV-1 Activity of Human Vaginal Fluid
Authors: Venkataraman Nitya , Cole Amy L. , Svoboda Pavel , Pohl Jan , Cole Alexander M. 
Published: April 2014

Abstract
Mucosal surfaces of the vagina are the portals for heterosexual transmission of HIV-1 and therefore play a fundamental role in the pathogenesis of primary infection. In the search for direct biological evidence for the role of human vaginal fluid in innate host defense, we characterized the anti-HIV-1 function of cationic polypeptides within minimally manipulated vaginal fluid. In the current study we revealed that vaginal fluid confers intrinsic anti-HIV-1 properties against both X4 and R5 strains of HIV-1 and could protect against HIV-1 infection and reduce proviral genome integration in organotypic cultures of human cervicovaginal tissue. The majority of this activity was contained in the cationic polypeptide fraction, and the depletion of cationic polypeptides using a selective cation exchange resin ablated most of the intrinsic activity against HIV-1. By adding the cationic polypeptide fraction to depleted vaginal fluid, we were able to restore activity against HIV-1. Using a proteomic approach, we identified 18 cationic polypeptides within vaginal fluid, nearly all of which are either known antimicrobials or have other purported roles in host defense. Interestingly, physiologic concentrations of 13 of the cationic polypeptides were not active alone against HIV-1, yet in concert they partially restored the anti-HIV-1 activity of cation-depleted vaginal fluid. These results suggest that synergism between cationic polypeptides is complex, and full anti-HIV-1 activity probably involves the aggregate of the cationic peptides and proteins in vaginal fluid.
Human NK Cells Inhibit Cytomegalovirus Replication through a Noncytolytic Mechanism Involving Lymphotoxin-Dependent Induction of IFN-β
Authors: Iversen Ann-Charlotte , Norris Paula S. , Ware Carl F. , Benedict Chris A. 
Published: April 2014

AbstractNK cells play a key role in host defense against the β-herpesvirus CMV through perforin-dependent cytolysis. In this study, we show that human NK cells can also control human CMV (HCMV) infection by a noncytolytic mechanism involving induction of IFN-β in the virus-infected cell. Both IL-2-activated primary NK cells and an IL-2-dependent NK cell line (NK-92) exhibited potent, noncytolytic anti-HCMV activity at very low E:T cell ratios (&lt;0.1:1). Activated NK cells expressed lymphotoxin (LT)αβ on their cell surface, and secreted LTα and TNF, all of which contributed to the NF-κB-dependent release of IFN-β from infected fibroblasts. IFN-β produced by fibroblasts and NK cell-produced IFN-γ combined to inhibit HCMV replication after immediate early gene expression. These results highlight an efficient mechanism used by NK cells to activate IFN-β expression in the infected target cell that contributes to the arrest of virion production and virus spread without cellular elimination.
Preferential Targeting of CD4-CCR5 Complexes with Bifunctional Inhibitors: A Novel Approach to Block HIV-1 Infection
Authors: Mack Matthias , Pfirstinger Jochen , Haas Jürgen , Nelson Peter J. , Kufer Peter , Riethmüller Gert , Schlöndorff Detlef 
Published: April 2014

Abstract
Two receptors, CD4 and one of several chemokine receptors, are required for cellular HIV-1 infection, with CCR5 being the main coreceptor for macrophage-tropic strains. We have designed bifunctional fusion proteins, consisting of RANTES/CCL5 and a single-chain Fv Ab fragment against CD4 to simultaneously block CD4 and CCR5. The fusion proteins bind to both receptors, compete with RANTES/CCL5 binding, and induce down-modulation of CCR5 ∼10 times more efficiently on CD4+ compared with CD8+ T cells. Moreover, after short incubation and subsequent washout, a significant down-modulation of CCR5 was only seen with the fusion proteins and only on CD4+ cells, but not with unmodified RANTES or on CD4− cells, indicating a preferential targeting of CCR5 on CD4+ T cells. The fusion proteins block M-tropic HIV infection more efficiently than RANTES/CCL5 and CD4 Abs alone or in combination. To our knowledge this is the first report of simultaneous blockade of an HIV-1 receptor and coreceptor with bifunctional inhibitors.
Viral Inhibition of IL-1- and Neutrophil Elastase-Induced Inflammatory Responses in Bronchial Epithelial Cells
Authors: Carroll Tomás P. , Greene Catherine M. , Taggart Clifford C. , Bowie Andrew G. , O’Neill Shane J. , McElvaney Noel G. 
Published: April 2014

AbstractPreviously, we elucidated the intracellular mechanisms by which neutrophil elastase (NE) up-regulates inflammatory gene expression in bronchial epithelial cells. In this study, we examine the effects of both IL-1 and NE on inflammatory gene expression in 16HBE14o− bronchial epithelial cells and investigate approaches to abrogate these inflammatory responses. IL-1 induced IL-8 protein production in time- and dose-dependent fashions, an important observation given that IL-8 is a potent neutrophil chemoattractant and a key inflammatory mediator. IL-1 and NE were shown to activate the p38 MAPK pathway in 16HBE14o− cells. Western blot analysis demonstrated IL-1R-associated kinase 1 (IRAK-1) degradation in response to stimulation with both IL-1 and NE. In addition, the expression of dominant negative IRAK-1 (IRAK-1Δ), IRAK-2Δ, or IRAK-4Δ inhibited IL-1- and NE-induced NF-κB-linked reporter gene expression. Dominant negative versions of the intracellular adaptor proteins MyD88 (MyD88Δ) and MyD88 adaptor-like (Mal P/H) abrogated NE-induced NF-κB reporter gene expression. In contrast, only MyD88Δ was found to inhibit IL-1-induced NF-κB reporter activity. We also investigated the vaccinia virus proteins, A46R and A52R, which have been shown to antagonize IL-1 signaling. Transfection with A46R or A52R cDNA inhibited IL-1- and NE-induced NF-κB and IL-8R gene expression and IL-8 protein production in primary and transformed bronchial epithelial cells. Furthermore, cytokine array studies demonstrated that IL-1 and NE can up-regulate the expression of IL-6, oncostatin M, epithelial cell-derived neutrophil activating peptide-78, growth-related oncogene family members, vascular endothelial growth factor, and GM-CSF, with induction of these proteins inhibited by the viral proteins. These findings identify vaccinia virus proteins as possible therapeutic agents for the manifestations of several inflammatory lung diseases.
Effector Function of Diabetogenic CD4 Th1 T Cell Clones: A Central Role for TNF-α
Authors: Cantor Joseph , Haskins Kathryn 
Published: April 2014

AbstractEffector function of T cells in autoimmune diabetes has been widely studied with mixed populations of lymphoid T cells stimulated ex vivo, but this approach does not permit evaluation of the contribution by a single T cell clone in the inflammatory site during pathogenesis. We have investigated cytokine production both in vitro and in vivo in a panel of diabetogenic CD4 Th1 T cell clones derived from the NOD mouse. SuperArray analysis showed a common pattern of mRNA expression for inflammatory cytokines and receptors in vitro after TCR stimulation. Ex vivo intracellular cytokine staining demonstrated that two important inflammatory cytokines, IFN-γ and TNF-α, were being made by these T cells recovered from the pancreas 6 days following adoptive transfer. TNF-α produced in the pancreas by pathogenic T cell clones and recruited macrophages was not the membrane-bound form. Secreted TNF-α can lead to production of multiple inflammatory chemokines, as were observed in the pathogenic clones by intracellular cytokine staining. Our results not only define the nature of an inflammatory cytokine response critical to development of diabetes, but also suggest its role in the regulation of other events during pathogenesis induced by CD4 T cells. Similar analyses in other models demonstrated that disease induced by CD4 T cell clones closely resembles spontaneous autoimmune diabetes in which both CD4 and CD8 T cells are required. Thus, cloned T cells in effect amplify effector function of T cells which otherwise may be difficult to detect without ex vivo stimulation.
In Vivo Correction of Complement Regulatory Protein Deficiency with an Inhibitor Targeting the Red Blood Cell Membrane
Authors: Spitzer Dirk , Unsinger Jacqueline , Mao Dailing , Wu Xiaobo , Molina Hector , Atkinson John P. 
Published: April 2014

AbstractBecause of the complement system’s involvement in many human diseases and potential complications associated with its systemic blockade, site-specific regulation of this effector system is an attractive concept. We report on further developments of such an approach using a single-chain Ab fragment as a vehicle to deliver complement regulatory proteins to a defined cell type. In a model system in which RBCs deficient in complement receptor 1-related gene/protein y (Crry) are rapidly cleared after injection into wild-type animals by a complement-dependent mechanism, we selectively reconstituted these cells with N- and C-terminally targeted recombinant forms of Crry. Transfusion of Crry-coated knockout RBCs into C57BL/6 mice extended their in vivo half-life from &lt;5 min to ∼2 days. Maintenance of protective levels of Crry (by a combined treatment of donor and recipient RBCs) led to nearly normal RBC survival. Uniform in vitro and in vivo coating of the RBCs and the more efficient complement inhibitory capacity of C-terminally tagged Crry were other interesting features of this experimental system. These results suggest the possibility of using the single-chain Ab fragment-mediated targeting concept of complement regulatory proteins to restrict complement inhibition to the site of its excessive activation.