bluuebunny/crossref_metadata_2025_split huggingface dataset:
Vaccination with Allergen-IL-18 Fusion DNA Protects Against, and Reverses Established, Airway Hyperreactivity in a Murine Asthma Model
Authors: Maecker Holden T. , Hansen Gesine , Walter David M. , DeKruyff Rosemarie H. , Levy Shoshana , Umetsu Dale T. 
Published: April 2014

AbstractVaccination with naked DNA encoding a specific allergen has been shown previously to prevent, but not reverse, the development of allergen-induced airway hyperresponsiveness (AHR). To enhance the effectiveness of DNA vaccine therapies and make possible the treatment of established AHR, we developed a DNA vaccination plasmid containing OVA cDNA fused to IL-18 cDNA. Vaccination of naive mice either with this fusion DNA construct or with an OVA cDNA-containing plasmid protected the mice from the subsequent induction of AHR. Protection from AHR correlated with increased IFN-γ production and reduced OVA-specific IgE production. The protection appeared to be mediated by IFN-γ and CD8+ cells because treatment of mice with neutralizing anti-IFN-γ mAb or with depleting anti-CD8 mAb abolished the protective effect. Moreover, vaccination of mice with preexisting AHR with the OVA-IL-18 fusion DNA, but not with the OVA cDNA plasmid, reversed established AHR, reduced allergen-specific IL-4, and increased allergen-specific IFN-γ production. Thus, combining IL-18 cDNA with OVA cDNA resulted in a vaccine construct that protected against the development of AHR, and that was unique among cDNA constructs in its capacity to reverse established AHR.
Quantifying the Frequency of Alloreactive T Cells In Vivo: New Answers to an Old Question
Authors: Suchin Elliot J. , Langmuir Peter B. , Palmer Ed , Sayegh Mohamed H. , Wells Andrew D. , Turka Laurence A. 
Published: April 2014

Abstract
Alloreactive T cell precursor frequency was measured in vivo using fluorescent dye labeling in combination with novel models based on lymphocyte activation and recovery. CFSE-labeled C57BL/6 (H-2b) spleen and lymph node cells were adoptively transferred to C57BL/6×DBA F1 (H-2b/d) recipients, a parent→F1 MHC mismatch in which only donor cells respond. Recipients were sacrificed at serial time points to assess engraftment efficiency, and the extent of donor cell activation and proliferation. These data were used to calculate alloreactive T cell frequencies that varied 30-fold (0.71 ± 0.31% to 21.05 ± 3.62%), depending upon whether it was assumed that all donor cells injected became established and were capable of responding, or that only those present at later time points (24–72 h) were available to respond. By measuring the number of cells established in the recipient 24 h after transfer, before proliferation, we calculated an in vivo alloreactive frequency of ∼7%. Using CD69 expression at 48 h to quantify activation, we found that 40–50% of the alloactivated CD4+ donor T cells do not divide. Studies of cotransferred congenic and allogeneic cells demonstrated that bystander proliferation does not occur. We conclude that accurate calculations of alloreactive precursor frequency must account for both proliferation and cell engraftment. When this is done, a high percentage of alloreactive T cells exists across an MHC mismatch, but not all alloreactive cells proliferate in vivo. Bystander proliferation is negligible, revealing exquisite specificity to the alloresponse. These data provide a novel approach to quantify alloreactive T cell responses during specific immunomodulatory strategies in vivo.
IL-7 and Not Stem Cell Factor Reverses Both the Increase in Apoptosis and the Decline in Thymopoiesis Seen in Aged Mice
Authors: Andrew Deborah , Aspinall Richard 
Published: April 2014

Abstract
Thymic atrophy is an age-associated decline in commitment to the T cell lineage considered to be associated with defective TCR β-chain rearrangement. Both IL-7 and stem cell factor (SCF) have dominant roles at this stage of triple negative (TN) thymocyte development. Because there is no age-associated decrease in the number of CD44+CD25−CD3−CD4−CD8− cells, this study investigated whether alterations in apoptosis within the TN pathway accounted for diminishing thymocyte numbers with age. Here we show significant age-associated increases in apoptotic TN thymocytes, specifically within CD44+CD25+ and CD44−CD25+ subpopulations, known to be the location of TCR β-chain rearrangement. IL-7 added to TN cultures established from old mice significantly both reduces apoptosis and increases the percentage of live cells within CD44+CD25+ and CD44−CD25+ subpopulations after 24 h, with prosurvival effects remaining after 5 days. SCF failed to demonstrate prosurvival effects in old or young cultures, and IL-7 and SCF together did not improve upon IL-7 alone. IL-7R expression did not decline with age, ruling out the possibility that the age-associated increase in apoptosis was attributed to reduced IL-7R expression. Compared with PBS, treatment of old mice with IL-7 produced significant increases in live TN cells. By comparison, treatment with SCF failed to increase live TN numbers, and IL-7 and SCF together failed to significantly improve thymopoiesis above that shown by IL-7 alone. Thus, treatment with IL-7 alone can reverse the age-associated defect in TN thymocyte development revealed by in vitro studies to be located at the stages of TCR β-chain rearrangement.
Autoimmune Intervention by CD154 Blockade Prevents T Cell Retention and Effector Function in the Target Organ
Authors: Howard Laurence M. , Miller Stephen D. 
Published: April 2014

AbstractThe CD40-CD154 interaction is an attractive target for therapeutic intervention in many autoimmune disorders, including multiple sclerosis. Previously, we showed that CD154 blockade both inhibited the onset of experimental autoimmune encephalomyelitis and blocked clinical disease progression (relapses) in mice with established disease. The mechanism of this protection is poorly understood. Because CD154 plays a role in Th1 development, its blockade has been thought to promote anti-inflammatory Th2 responses. However, these conclusions have primarily been based on extrapolated data from in vitro experiments, which may not accurately reflect the more complex events occurring in vivo. In this paper we determine how the immune response develops under the influence of therapeutic CD154 blockade in vivo. We demonstrate that anti-CD154 treatment does not alter the early expansion of Ag-specific T cells in secondary lymphoid organs or result in deviation to a Th2-dominant response. Interestingly, the late expansion and retention of Th1 cells in the lymph nodes were markedly reduced following immunization of Ab-treated mice, and this coincided with a recompartmentalization of these cells to the spleen. Most importantly, anti-CD154 treatment eliminated the retention/expansion of encephalitogenic Th1 cells, but not their entry into the CNS. These data indicate that a major mechanism by which CD154 blockade protects against autoimmune disease is by controlling the amplitude of acute phase Th1 responses in the draining lymph nodes and by preventing the sustained expansion of effector cells within the target organ.
Independent Regulation of Lymphocytic Choriomeningitis Virus-Specific T Cell Memory Pools: Relative Stability of CD4 Memory Under Conditions of CD8 Memory T Cell Loss
Authors: Varga Steven M. , Selin Liisa K. , Welsh Raymond M. 
Published: April 2014

AbstractInfection of mice with a series of heterologous viruses causes a reduction of memory CD8+ T cells specific to viruses from earlier infections, but the fate of the virus-specific memory CD4+ T cell pool following multiple virus infections has been unknown. We have previously reported that the virus-specific CD4+ Th precursor (Thp) frequency remains stable into long-term immunity following lymphocytic choriomeningitis virus (LCMV) infection. In this study, we questioned whether heterologous virus infections or injection with soluble protein CD4 Ags would impact this stable LCMV-specific CD4+ Thp memory pool. Limiting dilution analyses for IL-2-producing cells and intracellular cytokine staining for IFN-γ revealed that the LCMV-specific CD4+ Thp frequency remains relatively stable following multiple heterologous virus infections or protein Ag immunizations, even under conditions that dramatically reduce the LCMV-specific CD8+ CTL precursor frequency. These data indicate that the CD4+ and CD8+ memory T cell pools are regulated independently and that the loss in CD8+ T cell memory following heterologous virus infections is not a consequence of a parallel loss in the memory CD4+ T cell population.
Role of Phosphatidylinositol 3-Kinase in Anti-IgM- and Anti-IgD-Induced Apoptosis in B Cell Lymphomas
Authors: Carey Gregory B. , Scott David W. 
Published: April 2014

Abstract
Cross-linking of surface Ig receptors with anti IgM (anti-μ heavy chain, anti-μ), but not anti-IgD (anti-δ heavy chain, anti-δ), Abs leads to growth arrest and apoptosis in several extensively characterized B cell lymphomas. By poorly understood mechanisms, both Igs transiently stimulate c-Myc protein expression. However, ultimately, only anti-μ causes a severe loss in c-Myc and a large induction of p27Kip1 protein expression. Because phosphatidylinositol 3-kinase (PI3K) has been established as a major modulator of cellular growth and survival, we investigated its role in mediating anti-Ig-stimulated outcomes. Herein, we show that PI3K pathways regulate cell cycle progression and apoptosis in the ECH408 B cell lymphoma. Anti-μ and anti-δ driven c-Myc protein changes precisely follow their effects on the PI3K effector, p70S6K. Upstream of p70S6K, signaling through both Ig receptors depresses PI3K pathway phospholipids below control with time, which is followed by p27Kip1 induction. Conversely, anti-δ, but not anti-μ stimulated PI3K-dependent phospholipid return to control levels by 4–8 h. Abrogation of the PI3K pathway with specific inhibitors mimics anti-μ action, potentiates anti-μ-induced cell death and, importantly, converts anti-δ to a death signal. Transfection with active PI3K kinase construct induces anti-μ resistance, whereas transfection with dominant negative PI3K augments anti-μ sensitivity. Our results show that prolonged disengagement of PI3K or down-regulation of its products by anti-μ (and not anti-δ) determines B cell fate.
Prostaglandin E2 Up-Regulates Macrophage-Derived Chemokine Production but Suppresses IFN-Inducible Protein-10 Production by APC
Authors: Kuroda Etsushi , Sugiura Tsutomu , Okada Kazumasa , Zeki Kazuya , Yamashita Uki 
Published: April 2014

AbstractPGE2 has been known to suppress Th1 responses. We studied the role of PGE2 in two representative chemokines, macrophage-derived chemokine (MDC) and IFN-inducible protein-10, production by LPS- or CD40-stimulated spleen cells. The production of MDC, one of the ligands for CCR4 preferentially expressed on Th2, was enhanced in nonstimulated, LPS-, CD40-, or CD3-stimulated spleen cells by the pretreatment with PGE2, while the production of IFN-inducible protein-10, a representative ligand for CXC chemokine receptor 3 expressed on Th1, was suppressed. MDC production was also enhanced by IL-4, IL-5, and intracellular cAMP-elevating agents such as dibutyryl cAMP and 3-isobutyl-1-methylxanthine, and the effect of IL-4, IL-5, and PGE2 was additive. However, the pretreatment with IL-6, IL-10, or TGF-β, or the neutralization of IFN-γ or IL-12 had no effect on MDC production. B cells, macrophages, and dendritic cells were main producers of MDC, while T cells produced only a small amount of MDC. MDC production by B cells was equally stimulated by LPS and anti-CD40 Ab, while that by macrophages and dendritic cells was more markedly stimulated by anti-CD40 Ab, and PGE2 further enhanced MDC production by these stimulated cells. These results indicate that PGE2 regulates Th1/Th2-related chemokine production by B cells, macrophages, and dendritic cells, and that this is a new function of PGE2 for the regulation of Th2 immune responses at the induction and activation stages.
High-Avidity CTL Exploit Two Complementary Mechanisms to Provide Better Protection Against Viral Infection Than Low-Avidity CTL
Authors: Derby Michael A. , Alexander-Miller Martha A. , Tse Richard , Berzofsky Jay A. 
Published: April 2014

Abstract
Previously, we observed that high-avidity CTL are much more effective in vivo than low-avidity CTL in elimination of infected cells, but the mechanisms behind their superior activity remained unclear. In this study, we identify two complementary mechanisms: 1) high-avidity CTL lyse infected cells earlier in the course of a viral infection by recognizing lower Ag densities than those distinguished by low-avidity CTL and 2) they initiate lysis of target cells more rapidly at any given Ag density. Alternative mechanisms were excluded, including: 1) the possibility that low-avidity CTL might control virus given more time (virus levels remained as high at 6 days following transfer as at 3 days) and 2) that differences in efficacy might be correlated with homing ability. Furthermore, adoptive transfer of high- and low-avidity CTL into SCID mice demonstrated that transfer of a 10-fold greater amount of low-avidity CTL could only partially compensate for their decreased ability to eliminate infected cells. Thus, we conclude that high-avidity CTL exploit two complementary mechanisms that combine to prevent the spread of virus within the animal: earlier recognition of infected cells when little viral protein has been made and more rapid lysis of infected cells.
A New Regulatory Region of the IL-2 Locus That Confers Position-Independent Transgene Expression
Authors: Yui Mary A. , Hernández-Hoyos Gabriela , Rothenberg Ellen V. 
Published: April 2014

Abstract
Although the promoter/enhancer of the IL-2 gene mediates inducible reporter gene expression in vitro, it cannot drive consistent expression in transgenic mice. The location and existence of any regulatory elements that could open the IL-2 locus in vivo have remained unknown, preventing analysis of IL-2 regulation in developmental contexts. In this study, we report the identification of such a regulatory region, marked by novel DNase-hypersensitive sites upstream of the murine IL-2 promoter in unstimulated and stimulated T cells. Inclusion of most of these sites in an 8.4-kb IL-2 promoter green fluorescent protein transgene gives locus control region-like activity. Expression is efficient, tissue specific, and position independent. This transgene is expressed not only in peripheral T cells, but also in immature thymocytes and thymocytes undergoing positive selection, in agreement with endogenous IL-2 expression. In contrast, a 2-kb promoter green fluorescent protein transgene, lacking the new hypersensitive sites, is expressed in only a few founder lines, and expression is dysregulated in CD8+ cells. Thus, the 6.4 kb of additional upstream IL-2 sequence contains regulatory elements that provide integration site independence and differential regulation of transgene expression in CD8 vs CD4 cells.
The Integrity of the Ball-and-Socket Joint Between V and C Domains Is Essential for Complete Activity of a Humanized Antibody
Authors: Landolfi Nicholas F. , Thakur Archana B. , Fu Helen , Vásquez Max , Queen Cary , Tsurushita Naoya 
Published: April 2014

Abstract
AF2 is a high affinity murine Ab possessing potent neutralizing activity against human IFN-γ. In carrying out the modifications to humanize this Ab, we discovered that an initial version displayed affinity for IFN-γ that was slightly less than that of AF2, but exhibited IFN-γ-neutralizing activity that was severely diminished. Characterization via site-directed mutagenesis revealed that the majority of this loss in IFN-γ-neutralizing activity was due to altering the VH framework residue at position 11. VH position 11 is distal to the binding surface of the Ab; however, it, along with residues 110 and 112, have been identified as forming the socket of a molecular ball-and-socket joint between the V and C domains of the Ig Fab, which influences the elbow angle between these domains. To determine whether disrupting the structure of this joint was the basis for reduced IFN-γ-neutralizing capacity, we altered residue 148 of CH1, which with residue 149 comprises the corresponding ball portion of the joint. Changing this single CH1 domain residue diminished the ability of the Ab to neutralize IFN-γ to a level similar to that observed with the VH alteration. Thus, an intact ball-and-socket joint between the V and C domains in AF2 is required for potent neutralization of IFN-γ. These results suggest the importance of the elbow angle between Ig V and C domains in Ab activity, and support the hypothesis that this joint can be an important functional element of Ab structure.
The Interaction of FcαRI with IgA and Its Implications for Ligand Binding by Immunoreceptors of the Leukocyte Receptor Cluster
Authors: Wines Bruce D. , Sardjono Caroline T. , Trist Halina M. , Lay Chan-Sien , Hogarth P. Mark 
Published: April 2014

AbstractThis study defines the molecular basis of the FcαRI (CD89):IgA interaction, which is distinct from that of the other leukocyte Fc receptors and their Ig ligands. A comprehensive analysis using both cell-free (biosensor) and cell-based assays was used to define and characterize the IgA binding region of FcαRI. Biosensor analysis of mutant FcαRI proteins showed that residues Y35, Y81, and R82 were essential for IgA binding, and R52 also contributed. The role of the essential residues (Y35 and R82) was confirmed by analysis of mutant receptors expressed on the surface of mammalian cells. These receptors failed to bind IgA, but were detected by the mAb MY43, which blocks IgA binding to FcαRI, indicating that its epitope does not coincide with these IgA binding residues. A homology model of the ectodomains of FcαRI was generated based on the structures of killer Ig-like receptors, which share 30–34% identity with FcαRI. Key structural features of killer Ig-like receptors are appropriately reproduced in the model, including the structural conservation of the interdomain linker and hydrophobic core (residues V17, V97, and W183). In this FcαRI model the residues forming the IgA binding site identified by mutagenesis form a single face near the N-terminus of the receptor, distinct from other leukocyte Fc receptors where ligand binding is in the second domain. This taken together with major differences in kinetics and affinity for IgA:FcαRI interaction that were observed depending on whether FcαRI was immobilized or in solution suggest a mode of interaction unique among the leukocyte receptors.
NK1.1+ Cells and Murine Cytomegalovirus Infection: What Happens In Situ?
Authors: Andrews Daniel M. , Farrell Helen E. , Densley Eloise H. , Scalzo Anthony A. , Shellam Geoffrey R. , Degli-Esposti Mariapia A. 
Published: April 2014

Abstract
NK cells mediate early host defense against viral infection. In murine CMV (MCMV) infection NK cells play a critical role in controlling viral replication in target organs, such as spleen and liver. Until now it has not been possible to directly examine the role of NK cells in MCMV-induced inflammation in situ due to the inability to stain specifically for NK cells in infected tissues. In this study, we describe a method of in vivo fixation, resulting in the first identification of NK cells in situ using NK1.1 as the marker. Using this method, we characterize the NK1.1+ cellular component of the inflammatory response to wild-type MCMV in the spleen, liver, and lung of genetically susceptible and resistant mice following i.p. infection. This study provides the first in situ description of the cellular response mediated specifically by NK cells following MCMV infection.
The Application of a Plasmid DNA Encoding IFN-α1 Postinfection Enhances Cumulative Survival of Herpes Simplex Virus Type 2 Vaginally Infected Mice
Authors: Härle Peter , Noisakran Sansanee , Carr Daniel J. J. 
Published: April 2014

Abstract
Using a hormonally induced susceptibility mouse model to investigate vaginal HSV type 2 (HSV-2) infection, a study was undertaken to determine the efficacy of a plasmid DNA encoding IFN-α1 introduced into the vaginal lumen postinfection (PI). Mice infected with HSV-2 intravaginally and treated intravaginally 24 h later with 100 μg DNA encoding IFN-α1 showed enhanced survival (10/15) in comparison to mice treated with 100 μg plasmid DNA vector alone (3/10) or vehicle (4/27). In contrast, mice receiving recombinant IFN-αA (5–500 U/vagina) 24 h PI showed no significant survival in comparison to the vehicle (saline)-treated group. The protective effect was time dependent in that mice receiving the IFN-α1 transgene 48 h PI succumbed at a rate similar to the plasmid DNA vector-treated group. The increase in cumulative survival elicited by the transgene corresponded with a reduction in viral replication and Ag expressed in the vaginal epithelium early (i.e., 3 days PI) during acute infection and replicating virus recovered in the spinal cord day 7 PI. By day 7 PI, HSV-2 glycoprotein B transcript expression was no longer detectable in vaginal tissue from the IFN-α1 transgene-treated group (0/8) compared with levels expressed in plasmid vector-treated controls (4/6 mice surveyed were positive). Collectively, these results suggest the application of DNA encoding type I IFN is an effective and alternative approach to currently prescribed therapies in controlling vaginal HSV-2 infection by antagonizing viral replication.
Activated Antigen-Specific CD8+ T Cells Persist in the Lungs Following Recovery from Respiratory Virus Infections
Authors: Hogan Robert J. , Usherwood Edward J. , Zhong Weimin , Roberts Alan D. , Dutton Richard W. , Harmsen Allen G. , Woodland David L. 
Published: April 2014

Abstract
The poor correlation between cellular immunity to respiratory virus infections and the numbers of memory CD8+ T cells in the secondary lymphoid organs suggests that there may be additional reservoirs of T cell memory to this class of infection. Here we identify a substantial population of Ag-specific T cells in the lung that persist for several months after recovery from an influenza or Sendai virus infection. These cells are present in high numbers in both the airways and lung parenchyma and can be distinguished from memory cell populations in the spleen and peripheral lymph nodes in terms of the relative frequencies among CD8+ T cells, activation status, and kinetics of persistence. In addition, these cells are functional in terms of their ability to proliferate, express cytolytic activity, and secrete cytokines, although they do not express constitutive cytolytic activity. Adoptive transfer experiments demonstrated that the long-term establishment of activated T cells in the lung did not require infection in the lung by a pathogen carrying the inducing Ag. The kinetics of persistence of Ag-specific CD8+ T cells in the lung suggests that they play a key role in protective cellular immunity to respiratory virus infections.