bluuebunny/crossref_metadata_2025_split huggingface dataset:
Precursor Thymocyte Proliferation and Differentiation Are Controlled by Signals Unrelated to the Pre-TCR
Authors: Petrie Howard T. , Tourigny Michelle , Burtrum Douglas B. , Livak Ferenc 
Published: April 2014

Abstract
In-frame rearrangement of the TCR-β locus and expression of the pre-TCR are compulsory for the production of CD4+8+ thymocytes from CD4−8− precursors. Signals delivered via the pre-TCR are thought to induce the differentiation process as well as the extensive proliferation that accompanies this transition. However, it is equally possible that pre-TCR expression is required for the success of this transition, but does not play a direct role in the inductive process. In the present manuscript we examine this possibility using a variety of normal and genetically modified mouse models. Our evidence shows that differentiation and mitogenesis can both occur independently of pre-TCR expression. However, these processes are absolutely dependent on the presence of normal thymic architecture and cellular composition. These findings are consistent with a checkpoint role for the pre-TCR in regulating the divergence of survival and cell death fates at the CD4−8− to CD4+8+ transition. Further, our data suggest that precursor thymocyte differentiation is induced by other, probably ubiquitous, mechanisms that require the presence of normal thymic cellularity, composition, and architecture.
CD25 Is a Marker for CD4+ Thymocytes That Prevent Autoimmune Diabetes in Rats, But Peripheral T Cells with This Function Are Found in Both CD25+ and CD25− Subpopulations
Authors: Stephens Leigh Andrea , Mason Don 
Published: April 2014

Abstract
Previously we have shown that autoimmune diabetes, induced in rats by a protocol of adult thymectomy and split-dose gamma irradiation, can be prevented by the transfer of a subset of CD4+ T cells with a memory phenotype (CD45RC−), as well as by CD4+CD8− thymocytes, from syngeneic donors. Further studies now reveal that in the thymus the regulatory cells are observed in the CD25+ subset of CD4+CD8− cells, whereas transfer of the corresponding CD25− thymocyte subset leads to acceleration of disease onset in prediabetic recipients. However, in the periphery, not all regulatory T cells were found to be CD25+. In thoracic duct lymph, cells that could prevent diabetes were found in both CD25− and CD25+ subsets of CD4+CD45RC− cells. Further, CD25− regulatory T cells were also present within the CD4+CD45RC− cell subset from spleen and lymph nodes, but were effective in preventing diabetes only after the removal of CD25− recent thymic emigrants. Phenotypic analysis of human thymocytes showed the presence of CD25+ cells in the same proportions as in rat thymus. The possible developmental relationship between CD25+ and CD25− regulatory T cells is discussed.
Impaired Affinity Maturation inCr2−/− Mice Is Rescued by Adjuvants Without Improvement in Germinal Center Development
Authors: Wu Xiaobo , Jiang Ning , Fang Yi-Fu , Xu Chenguang , Mao Dailing , Singh Jasvinder , Fu Yang-Xin , Molina Hector 
Published: April 2014

AbstractCr2 −/− mice have an impairment in humoral immunity, as shown by the decrease in the Ab titers against T cell-dependent Ags and abnormalities in germinal center formation. Germinal centers are present, but they are decreased in size and number, indicating problems in their development. In this study, we investigated whether this abnormality in germinal center development is associated with problems in the establishment of optimal affinity maturation and the generation of memory B cells, processes closely related to the germinal center reaction. We immunized the Cr2−/− animals with different Ags with or without adjuvants. We showed that, when immunized without adjuvants, complement receptors are absolutely required for optimal affinity maturation. Although limited affinity maturation is elicited in the Cr2−/− Ab response, it is decreased as compared with normal animals. Memory B cell generation is also impaired. In the presence of adjuvants, germinal center development in the Cr2−/− mice is still abnormal, as demonstrated by their decreased size and number. Surprisingly, adjuvants establish optimal affinity maturation and partially restore the amount of Ab produced during the primary response and memory B cell generation. However, adjuvants cannot improve the ability of follicular dendritic cells to retain Ags in the form of immune complexes. These observations indicate that immunization with inflammatory Ags offset some of the immunological abnormalities found in the Cr2−/− mice and show that optimal affinity maturation in the Cr2−/− mice can be achieved in the absence of normal germinal centers.
The Brain Parenchyma Is Permissive for Full Antitumor CTL Effector Function, Even in the Absence of CD4 T Cells
Authors: Walker Paul R. , Calzascia Thomas , Schnuriger Valérie , Scamuffa Nathalie , Saas Philippe , de Tribolet Nicolas , Dietrich Pierre-Yves 
Published: April 2014

Abstract
Effective antitumor immune responses against cerebral malignancies have been demonstrated in several models, but precise cellular function of specific effector cells is poorly understood. We have explored this topic by analyzing the MHC class I-restricted T cell response elicited after implantation of HLA-CW3-transfected P815 mastocytoma cells (P815-CW3) in syngeneic mice. In this model, tumor-specific CTLs use a distinctive repertoire of TCRs that allows ex vivo assessment of the response by immunophenotyping and TCR spectratyping. Thus, for the first time in a brain tumor model, we are able to directly visualize ex vivo CTLs specific for a tumor-expressed Ag. Tumor-specific CTLs are detected in the CNS after intracerebral implantation of P815-CW3, together with other inflammatory cells. Moreover, despite observations in other models suggesting that CTLs infiltrating the brain may be functionally compromised and highly dependent upon CD4 T cells, in this syngeneic P815-CW3 model, intracerebral tumors were efficiently rejected, whether or not CD4 T cells were present. This observation correlated with potent ex vivo cytotoxicity of brain-infiltrating CTLs, specific for the immunodominant epitope CW3170–179 expressed on P815-CW3 tumor cells.
Production of Plasminogen Activator Inhibitor-1 by Human Mast Cells and Its Possible Role in Asthma
Authors: Cho Seong H. , Tam Sun W. , Demissie-Sanders Sossiena , Filler Scott A. , Oh Chad K. 
Published: April 2014

Abstract
The plasminogen activator inhibitor type 1 (PAI-1) has an essential role in tissue remodeling. The PAI-1 gene was induced by a combination of phorbol ester and calcium ionophore at the highest level among the inducible human mast cell genes that we have analyzed on a DNA microarray. PAI-1 was secreted by both a human mast cell line (HMC)-1 and primary cultured human mast cells upon stimulation, whereas PAI-1 was undetectable in either group of unstimulated cells. The secretion of PAI-1 was due to de novo synthesis of PAI-1 rather than secretion of preformed PAI-1. The functional significance of PAI-1 secretion was demonstrated by complete inhibition of tissue-type plasminogen activator activity with supernatants of stimulated HMC-1 cells. Furthermore, we were able to regulate PAI-1 gene expression in HMC-1 cells by known therapeutic agents. High-dose (1 μM) dexamethasone induced PAI-1 mRNA expression. Cyclosporin down-regulated the expression of the PAI-1 gene. Cycloheximide abrogated PAI-1 mRNA expression, suggesting that transcription of the PAI-1 gene requires de novo synthesis of early gene products, including transcription factors. Finally, we demonstrated PAI-1 in lung mast cells from a patient with asthmatic attack by double-immunofluorescence study. This is the first report demonstrating that activated human mast cells release a striking amount of functionally active PAI-1. These results suggest that PAI-1 could play an important role in airway remodeling of asthma, and inhibition of PAI-1 activity could represent a novel therapeutic approach in the management of airway remodeling.
IFN-γ Up-Regulates IL-18 Gene Expression Via IFN Consensus Sequence-Binding Protein and Activator Protein-1 Elements in Macrophages
Authors: Kim Yong-Man , Im Joo Young , Han Seung Hyun , Kang Hyung Sik , Choi Inpyo 
Published: April 2014

AbstractConstitutive IL-18 expression is detected from many different cells, including macrophages, keratinocytes, and osteoblasts. It has been known that IL-18 gene expression is regulated by two different promoters (p1 promoter and p2 promoter). When RAW 264.7 macrophages were treated with IFN-γ, IL-18 gene expression was increased in a dose- and time-dependent manner. IFN-γ activated the inducible promoter 1, but not the constitutive promoter 2. Mutagenesis studies indicated that an IFN consensus sequence-binding protein (ICSBP) binding site between −39 and −22 was critical for the IFN-γ inducibility. EMSA using an ICSBP oligonucleotide probe showed that IFN-γ treatment increased the formation of DNA-binding complex, which was supershifted with anti-IFN regulatory factor-1 Ab and anti-ICSBP Ab. Another element, an AP-1 site between −1120 and −1083, was important. EMSA using an AP-1-specific oligonucleotide demonstrated that IFN-γ or LPS treatment increased the AP-1-binding activity. The addition of anti-c-Jun Ab or anti-c-Fos Ab to IFN-γ- or LPS-treated nuclear extracts resulted in the reduction of AP-1 complex or the formation of a supershifted complex. Taken together, these results indicate that IFN-γ increased IL-18 gene expression via ICSBP and AP-1 elements.
The I-Ag7 MHC Class II Molecule Linked to Murine Diabetes Is a Promiscuous Peptide Binder
Authors: Stratmann Thomas , Apostolopoulos Vasso , Mallet-Designe Valérie , Corper Adam L. , Scott Christopher A. , Wilson Ian A. , Kang Angray S. , Teyton Luc 
Published: April 2014

AbstractSusceptibility to insulin-dependent diabetes mellitus is linked to MHC class II genes. The only MHC class II molecule expressed by nonobese diabetic (NOD) mice, I-Ag7, shares a common α-chain with I-Ad but has a peculiar β-chain. As with most β-chain alleles linked to diabetes susceptibility, I-Ag7 contains a nonaspartic residue at position β57. We have produced large amounts of empty I-Ag7 molecules using a fly expression system to characterize its biochemical properties and peptide binding by phage-displayed peptide libraries. The identification of a specific binding peptide derived from glutamic acid decarboxylase (GAD65) has allowed us to crystallize and obtain the three-dimensional structure of I-Ag7. Structural information was critical in evaluating the binding studies. I-Ag7, like I-Ad, appears to be very promiscuous in terms of peptide binding. Their binding motifs are degenerate and contain small and/or small hydrophobic residues at P4 and P6 of the peptide, a motif frequently found in most globular proteins. The degree of promiscuity is increased for I-Ag7 over I-Ad as a consequence of a larger P9 pocket that can specifically accommodate negatively charged residues, as well as possibly residues with bulky side chains. So, although I-Ad and I-Ag7 are structurally closely related, stable molecules and good peptide binders, they differ functionally in their ability to bind significantly different peptide repertoires that are heavily influenced by the presence or the absence of a negatively charged residue at position 57 of the β-chain. These characteristics link I-Ag7 with autoimmune diseases, such as insulin-dependent diabetes mellitus.
Serine 6 of Lck Tyrosine Kinase: A Critical Site for Lck Myristoylation, Membrane Localization, and Function in T Lymphocytes
Authors: Yasuda Koubun , Kosugi Atsushi , Hayashi Fumie , Saitoh Shin-ichiroh , Nagafuku Masakazu , Mori Yoshiko , Ogata Masato , Hamaoka Toshiyuki 
Published: April 2014

Abstract
Lck is a member of the Src family kinases expressed predominantly in T cells, and plays a pivotal role in TCR-mediated signal transduction. Myristoylation of glysine 2 in the N-terminal Src homology 4 (SH4) domain of Lck is essential for membrane localization and function. In this study, we examined a site within the SH4 domain of Lck regulating myristoylation, membrane localization, and function of Lck. A Lck mutant in which serine 6 (Ser6) was substituted by an alanine was almost completely cytosolic in COS-7 cells, and this change of localization was associated with a drastic inhibition of myristoylation in this mutant. To assess the role of Ser6 of Lck in T cell function, we established stable transfectants expressing various Lck mutants using Lck-negative JCaM1 cells. The Lck mutant of Ser6 to alanine, most of which did not target to the plasma membrane, was not able to reconstitute TCR-mediated signaling events in JCaM1 cells, as analyzed by tyrosine phosphorylation of intracellular proteins and CD69 expression. These results demonstrate that Ser6 is a critical factor for Lck myristoylation, membrane localization, and function in T cells, presumably because the residue is important for N-myristoyl transferase recognition.
An Intron Transcriptional Enhancer Element Regulates IL-4 Gene Locus Accessibility in Mast Cells
Authors: Hural John A. , Kwan Millie , Henkel Greg , Hock M. Benjamin , Brown Melissa A. 
Published: April 2014

AbstractThe cell type-specific expression of a gene is dependent on developmentally regulated modifications in chromatin structure that allow accessibility of basal and inducible transcription factors. In this study, we demonstrate that a cis-acting element in the second intron of the murine IL-4 gene has a dual function in regulating transcription in mast cells as well as chromatin accessibility of the IL-4 gene locus through its influence on the methylation state of the gene. Previous studies have shown that mast cell-restricted transcription factors GATA-1/2 and PU.1 associate with the intron element and regulate its activity. In this study, we use DNase I footprinting and mutational analyses to identify two additional sites that contribute to the element’s ability to enhance transcription. One of these sites associates preferentially with STAT5a and STAT5b. We also demonstrate that deletion of the element or mutation of the GATA binding site in the context of a stably integrated IL-4 genomic construct prevents maintenance of a demethylated locus in IL-4-producing mast cells. These data indicate that, analogous to Ig and TCR intron regulatory elements, the intron enhancer has an essential role in maintaining developmentally regulated demethylation at the IL-4 gene locus. In addition, they indicate that members of the GATA family of transcription factors likely play an important role in these processes.
The Primate Lentiviral Receptor Bonzo/STRL33 Is Coordinately Regulated with CCR5 and Its Expression Pattern Is Conserved Between Human and Mouse
Authors: Unutmaz Derya , Xiang Wenkai , Sunshine Mary Jean , Campbell Jim , Butcher Eugene , Littman Dan R. 
Published: April 2014

Abstract
Chemokines play necessary and important roles in regulating the trafficking of lymphocytes to intra- or interlymphoid tissues as well as to sites of inflammation. The complex migratory patterns of lymphoid lineage cells is governed by subset-specific expression of chemokine receptors and their access to specific ligands. Several chemokine receptors and chemokine receptor-like orphan receptors also serve, in conjunction with CD4, as coreceptors for infection by human and simian immunodeficiency viruses (HIV and SIV). Here we show that the expression pattern of Bonzo/STRL33, an orphan SIV/HIV coreceptor, is highly restricted to the memory subset of T cells and is up-regulated upon stimulation of these cells with IL-2 or IL-15. Both the pattern and the regulation of Bonzo expression closely paralleled that of CC family chemokine receptors CCR5 or CCR6 and inversely correlated with CXCR4 expression. However, in striking contrast to CCR5, Bonzo expression was not down-modulated by PMA or mitogen stimulation of T cells. Targeted replacement of the Bonzo gene with a gene encoding green fluorescent protein in mice revealed that the expression and cytokine regulation of mouse Bonzo are comparable to those of its human counterpart. The similar expression and regulation patterns of Bonzo and the HIV coreceptor CCR5 may have implications for understanding the role of HIV/SIV receptors in viral evolution and pathogenesis.
The Phagocytosis-Associated Respiratory Burst in Human Monocytes Is Associated with Increased Uptake of Glutathione
Authors: Seres Tamas , Knickelbein Roy G. , Warshaw Joseph B. , Johnston Richard B. 
Published: April 2014

Abstract
During the phagocytic respiratory burst, oxygen is converted to potent cytotoxic oxidants. Monocytes and macrophages are potentially long-lived, and we have hypothesized that protective mechanisms against oxidant stress are varied and fully expressed in these cells. We report here that the respiratory burst in monocytes is accompanied by an increase in the uptake of [35S]glutathione ([35S]GSH) after 20–30 min to levels up to 10-fold greater than those at baseline. By 30 min, 49% of the cell-associated radioactivity was in the cytosol, 41% was in membrane, and 10% was associated with the nuclear fraction. GSH uptake was inhibited by catalase, which removes hydrogen peroxide (H2O2), and micromolar H2O2 stimulated GSH uptake effectively in monocytes and also lymphocytes. Oxidation of GSH to glutathione disulfide with H2O2 and glutathione peroxidase prevented uptake. Acivicin, which inhibits GSH breakdown by γ-glutamyl transpeptidase (GGT), had no effect on the enhanced uptake seen during the respiratory burst. Uptake of cysteine or cystine, possible products of GGT activity, stayed the same or decreased during the respiratory burst. These results suggest that a GGT-independent mechanism is responsible for the enhanced GSH uptake seen during the respiratory burst. We describe here a sodium-independent, methionine-inhibitable transport system with a Km (8.5 μM) for GSH approximating the plasma GSH concentration. These results suggest that monocytes have a specific GSH transporter that is triggered by the release of H2O2 during the respiratory burst and that induces the uptake of GSH into the cell. Such a mechanism has the potential to protect the phagocyte against oxidant damage.
ICAM-1-Coupled Cytoskeletal Rearrangements and Transendothelial Lymphocyte Migration Involve Intracellular Calcium Signaling in Brain Endothelial Cell Lines
Authors: Etienne-Manneville Sandrine , Manneville Jean-Baptiste , Adamson Peter , Wilbourn Barry , Greenwood John , Couraud Pierre-Olivier 
Published: April 2014

AbstractEndothelium of the cerebral blood vessels, which constitutes the blood-brain barrier, controls adhesion and trafficking of leukocytes into the brain. Investigating signaling pathways triggered by the engagement of adhesion molecules expressed on brain endothelial cells using two rat brain endothelial cell lines (RBE4 and GP8), we report in this paper that ICAM-1 cross-linking induces a sustained tyrosine phosphorylation of the phosphatidylinositol-phospholipase C (PLC)γ1, with a concomitant increase in both inositol phosphate production and intracellular calcium concentration. Our results suggest that PLC are responsible, via a calcium- and protein kinase C (PKC)-dependent pathway, for p60Src activation and tyrosine phosphorylation of the p60Src substrate, cortactin. PKCs are also required for tyrosine phosphorylation of the cytoskeleton-associated proteins, focal adhesion kinase and paxillin, but not for ICAM-1-coupled p130Cas phosphorylation. PKC’s activation is also necessary for stress fiber formation induced by ICAM-1 cross-linking. Finally, cell pretreatment with intracellular calcium chelator or PKC inhibitors significantly diminishes transmonolayer migration of activated T lymphocytes, without affecting their adhesion to brain endothelial cells. In summary, our data demonstrate that ICAM-1 cross-linking induces calcium signaling which, via PKCs, mediates phosphorylation of actin-associated proteins and cytoskeletal rearrangement in brain endothelial cell lines. Our results also indicate that these calcium-mediated intracellular events are essential for lymphocyte migration through the blood-brain barrier.
Induction of Heme Oxygenase-1 Expression in Macrophages by Diesel Exhaust Particle Chemicals and Quinones via the Antioxidant-Responsive Element
Authors: Li Ning , Venkatesan M. Indira , Miguel Antonio , Kaplan Roman , Gujuluva Chandrasekhar , Alam Jawed , Nel Andre 
Published: April 2014

Abstract
Diesel exhaust particles (DEP) contain organic chemicals that contribute to the adverse health effects of inhaled particulate matter. Because DEP induce oxidative stress in the lung and in macrophages, effective antioxidant defenses are required. One type of defense is through the expression of the antioxidant enzyme, heme oxygenase I (HO-1). HO-1 as well as phase II detoxifying enzymes are induced via antioxidant response elements (ARE) in their promoters of that gene. We show that a crude DEP total extract, aromatic and polar DEP fractions, a benzo(a)pyrene quinone, and a phenolic antioxidant induce HO-1 expression in RAW264.7 cells in an ARE-dependent manner. N-acetyl cysteine and the flavonoid, luteolin, inhibited HO-1 protein expression. We also demonstrate that the same stimuli induce HO-1 mRNA expression in parallel with the activation of the SX2 enhancer of that gene. Mutation of the ARE core, but not the overlapping AP-1 binding sequence, disrupted SX2 activation. Finally, we show that biological agents, such as oxidized 1-palmitoyl-2-arachidonoyl-sn-glycero-3-phosphocholine, could also induce HO-1 expression via an ARE-dependent mechanism. Prior induction of HO-1 expression, using cobalt-protoporphyrin, protected RAW264.7 cells against DEP-induced toxicity. Taken together, these data show that HO-1 plays an important role in cytoprotection against redox-active DEP chemicals, including quinones.
A SAF Binding Site in the Promoter Region of Human γ-Fibrinogen Gene Functions as an IL-6 Response Element
Authors: Ray Alpana 
Published: April 2014

Abstract
Expression of fibrinogen is highly induced during inflammation, and such abnormal expression of this protein is considered as a major cardiovascular risk factor. IL-6 is one of the main mediators of abnormal expression of fibrinogen leading to the pathogenic conditions. Transient transfection and EMSA were performed to investigate the molecular mechanism of IL-6-induced γ-fibrinogen gene expression in hepatic cells. Using progressively deleted 5′ fragments of the γ-fibrinogen promoter coupled to chloramphenicol acetyltransferase gene, an IL-6 responsive element located between positions −273 and −259 was identified. Mutation of this element abrogates IL-6 responsiveness of the γ-fibrinogen promoter. Interaction of this promoter with a zinc finger transcription factor, serum amyloid A activating factor (SAF)-1, was demonstrated by EMSA. Furthermore, overexpression of wild-type SAF-1 in transfected liver cells can increase transcription of the γ-fibrinogen promoter. These data show that transcription factor SAF-1 is involved in the regulation of IL-6-mediated induction of the human γ-fibrinogen gene in liver cells.