bluuebunny/crossref_metadata_2025_split huggingface dataset:
Identification of a CD28 Response Element in the CD40 Ligand Promoter
Authors: Parra Eduardo , Mustelin Tomas , Dohlsten Mikael , Mercola Dan 
Published: April 2014

Abstract
Ligation of the T cell coreceptor CD28 or CD2 by its cognate ligands B7-1 or LFA-3, respectively, greatly aids the Ag-induced up-regulation of several genes, including IL-2 and CD40 ligand (CD40L). Using luciferase reporter constructs under the control of the 1.2 kb of 5′ noncoding region of the human CD40L gene, we have found that stimulation through CD28 was required for a strong transcriptional activity of the CD40L promoter in response to TCR ligation, while the activity induced by CD2 was slightly lower than CD28. Deletion analysis demonstrated that the transcriptional elements mediating this effect were located within a 300-bp region upstream of the start site. Further dissection of this region and gel shift analyses demonstrated the presence of a CD28 response element in a region located between nucleotides −170 to −164 relative to the start site. Transcriptional studies with a CD40L enhancer-promoter carrying a mutation in this putative CD28 response element revealed that the activity was reduced by 80 and 70% after B7-1 and LFA-3 costimulation, respectively. The transcription factor complex bound to this site contained at least JunD, c-Fos, p50, p65, and c-Rel, but not c-Jun. Mutations introduced into the CD28RE also blocked the binding of this complex. These observations identify an important role for the CD28 signaling pathway in the regulation of CD40L promoter transcriptional activity.
Cloning of Human Preprotachykinin-I Promoter and the Role of Cyclic Adenosine 5′-Monophosphate Response Elements in Its Expression by IL-1 and Stem Cell Factor
Authors: Qian Jing , Yehia Ghassan , Molina Carlos A. , Fernandes Annemarie , Donnelly Robert J. , Anjaria Devashish J. , Gascon Pedro , Rameshwar Pranela 
Published: April 2014

Abstract
Preprotachykinin-I gene (PPT-I) encodes several peptides with organ-specific functions that link the neuroendocrine-immune-hemopoietic axis. We cloned upstream of the initiation site of human PPT-I promoter and identified consensus sequences for two cAMP response elements (CRE). PPT-I is induced by cytokines including those that signal through the cAMP pathway. Therefore, we studied the role of the two CRE in IL-1α and stem cell factor (SCF) stimulation of bone marrow stroma because both cytokines induce endogenous PPT-I in these cells and activate the cAMP pathway. Furthermore, bone marrow stroma expresses the transcription factors regulated by the cAMP pathways such as the repressor (ICERIIγ) and activator (CREMτ). Mutagenesis of the two CRE and/or cotransfection with vectors that express ICERIIγ or CREMτ indicated that the two CRE have major roles in PPT-I expression. The two CRE are also required for optimal promoter activity by SCF and IL-1α. A particular cytokine could concomitantly induce PPT-I and the high affinity G protein-coupled receptor for PPT-I peptides, NK-1R. We showed that SCF, a representative cytokine, induced PPT-I and NK-1R leading to autocrine and/or paracrine cell activation. Because NK-1R activates cAMP through the G protein, the results suggest that the presence of CRE sequences within PPT-I promoter could be important in the regulation of PPT-I expression by cytokines, irrespective of their ability to signal through cAMP. As PPT-I is implicated in hemopoietic regulation, immune responses, breast cancer, and other neural functions, these studies add to the basic biology of these processes and could provide targets for drug development.
The A′ and F′ Pockets of Human CD1b Are Both Required for Optimal Presentation of Lipid Antigens to T Cells
Authors: Niazi Kayvan R. , Chiu Melvin W. , Mendoza Richard M. , Degano Massimo , Khurana Sumit , Moody D. Branch , Melián Agustín , Wilson Ian A. , Kronenberg Mitchell , Porcelli Steven A. , Modlin Robert L. 
Published: April 2014

Abstract
CD1 proteins are unique in their ability to present lipid Ags to T cells. Human CD1b shares significant amino acid homology with mouse CD1d1, which contains an unusual putative Ag-binding groove formed by two large hydrophobic pockets, A′ and F′. We investigated the function of the amino acid residues that line the A′ and F′ pockets of CD1b by engineering 36 alanine-substitution mutants and analyzing their ability to present mycobacterial glycolipid Ags. Two lipid Ags presented by CD1b were studied, a naturally occurring glucose monomycolate (GMM) isolated from mycobacteria, which contains two long alkyl chains (C54-C62 and C22-C24) and synthetic GMM (sGMM), which includes two short alkyl chains (C18 and C14). We identified eight residues in both the A′ and F′ pockets that were involved in the presentation of both GMM and sGMM to T cells. Interestingly, four additional residues located in the distal portion of the A′ pocket were required for the optimal presentation of GMM, but not sGMM. Conversely, nine residues located between the center of the groove and the F′ pocket were necessary for the optimal presentation of sGMM, but not GMM. These data indicate that both the A′ and F′ pockets of human CD1b are required for the presentation of lipid Ags to T cells.
Engineered Antibodies with Increased Activity to Recruit Complement
Authors: Idusogie Esohe E. , Wong Pin Yee , Presta Leonard G. , Gazzano-Santoro Helene , Totpal Klara , Ultsch Mark , Mulkerrin Michael G. 
Published: April 2014

Abstract
This manuscript describes two sites in a human IgG1 that, when mutated individually or in combination, result in a dramatic increase in C1q binding and complement-dependent cytotoxicity activity. These two residues, K326 and E333, are located at the extreme ends of the C1q binding epicenter in the CH2 domain of a human IgG. A mutation to tryptophan at K326 debilitates Ab-dependent cell-mediated cytotoxicity activity. In addition, substitutions of the residues E333 with serine and of K326 with tryptophan in a human IgG2 confer biological activity in the complement-dependent cytotoxicity assay in which the wild-type IgG2 is inactive. This study reveals that the residues K326 and E333 play a significant role in the control of the biological activity of an IgG molecule and can rescue the activity of an inactive IgG isotype.
A Redundant Role of the CD3γ-Immunoreceptor Tyrosine-Based Activation Motif in Mature T Cell Function
Authors: Haks Mariëlle C. , Cordaro Tanina A. , van den Brakel Jeroen H. N. , Haanen John B. A. G. , de Vries Evert F. R. , Borst Jannie , Krimpenfort Paul , Kruisbeek Ada M. 
Published: April 2014

AbstractAt least four different CD3 polypeptide chains are contained within the mature TCR complex, each encompassing one (CD3γ, CD3δ, and CD3ε) or three (CD3ζ) immunoreceptor tyrosine-based activation motifs (ITAMs) within their cytoplasmic domains. Why so many ITAMs are required is unresolved: it has been speculated that the different ITAMs function in signal specification, but they may also serve in signal amplification. Because the CD3ζ chains do not contribute unique signaling functions to the TCR, and because the ITAMs of the CD3-γδε module alone can endow the TCR with normal signaling capacity, it thus becomes important to examine how the CD3γ-, δ-, and ε-ITAMs regulate TCR signaling. We here report on the role of the CD3γ chain and the CD3γ-ITAM in peripheral T cell activation and differentiation to effector function. All T cell responses were reduced or abrogated in T cells derived from CD3γ null-mutant mice, probably because of decreased expression levels of the mature TCR complex lacking CD3γ. Consistent with this explanation, T cell responses proceed undisturbed in the absence of a functional CD3γ-ITAM. Loss of integrity of the CD3γ-ITAM only slightly impaired the regulation of expression of activation markers, suggesting a quantitative contribution of the CD3γ-ITAM in this process. Nevertheless, the induction of an in vivo T cell response in influenza A virus-infected CD3γ-ITAM-deficient mice proceeds normally. Therefore, if ITAMs can function in signal specification, it is likely that either the CD3δ and/or the CD3ε chains endow the TCR with qualitatively unique signaling functions.
A Particular TCR β Variable Region Used by T Cells Infiltrating Kidney Transplants
Authors: Baron Christophe , Sachs David H. , LeGuern Christian 
Published: April 2014

Abstract
Immune tolerance to MHC class II identical renal grafts is achievable in miniature swine following a short immunosuppressive treatment. Like in clinical transplants, swine-accepted allografts are primarily infiltrated by CD8+ T cells, which are noncytotoxic to the renal tissue. However, the actual specificity and function of these intragraft-infiltrating lymphocytes remain poorly understood. To develop the molecular tools to study TCR-associated functions of graft-infiltrating cells in a preclinical transplantation model, we have determined the nucleotide sequence of 19 pig Vβ, 12 Jβ, and two Dβ. Sequence comparisons identified 17 different Vβ families and two Jβ clusters homologous to the human Jβ1 and Jβ2. The fact that the pig Jβ1 segments were always found joined to the Dβ1-like sequence in numerous rearranged TCR β cDNA suggests the existence of two d-J clusters in swine. These results also imply that the polymorphism of the porcine TCR β segments is similar to that found in human. Finally, we report the discovery of a new and functional Vβ subfamily named Vβ100, which exhibited similarity to the murine Vβ2 sequence but had no described Vβ homolog in humans. Pilot spectratyping studies on Vβ usage revealed a clonal dominance of Vβ100+ T cell subsets among infiltrating cells in two accepted grafts.
Formation of High-Affinity C5 Convertases of the Alternative Pathway of Complement
Authors: Rawal Nenoo , Pangburn Michael K. 
Published: April 2014

Abstract
Cleavage of C5 by C5 convertase is the last enzymatic step in the complement activation cascade leading to the formation of the cytolytic proteolytically activated form of C5 (C5b)-9 complex. In the present study, we examined the effect of the density of C3b (the proteolytically activated form of C3) on the function of the noncatalytic subunit of natural surface-bound forms of the enzyme. A comparison of the kinetic parameters of C5 convertases assembled on three surfaces (zymosan, rabbit erythrocytes, and sheep erythrocytes) were similar and revealed that the average Km decreased ∼28-fold (5.2–0.18 μM) when the density of C3b was increased from ∼18,000 to 400,000 C3b/cell. Very-high-affinity C5 convertases were generated when preformed C3 convertases were allowed to self amplify by giving them excess C3. These convertases exhibited Km from 0.016 to 0.074 μM, well below the normal plasma concentration of C5 in blood (0.37 μM). The results suggest that in serum convertases formed with monomeric C3b will be relatively inefficient in capturing C5 but will continue to cleave C3 opsonizing the cell surface for phagocytosis, whereas convertases formed with C3b-C3b complexes in areas of high C3b density will primarily cleave C5. The catalytic rate of these convertases approaches maximum velocity, thereby switching the enzyme from cleavage of C3 to cleavage of C5, and production of the cytolytic C5b-9 complex.
Elastase Controls the Binding of the Vitamin D-Binding Protein (Gc-Globulin) to Neutrophils: A Potential Role in the Regulation of C5a Co-Chemotactic Activity
Authors: DiMartino Stephen J. , Shah Anisha B. , Trujillo Glenda , Kew Richard R. 
Published: April 2014

AbstractThe vitamin D-binding protein (DBP) binds to the plasma membranes of numerous cell types and mediates a diverse array of cellular functions. DBP bound to the surface of leukocytes serves as a co-chemotactic factor for C5a, significantly enhancing the chemotactic activity of pM concentrations of C5a. This study investigated the regulation of DBP binding to neutrophils as a possible key step in the process of chemotaxis enhancement to C5a. Using radioiodinated DBP as a probe, neutrophils released 70% of previously bound DBP into the extracellular media during a 60-min incubation at 37°C. This was suppressed by serine protease inhibitors (PMSF, Pefabloc SC), but not by metallo- or thiol-protease inhibitors. DBP shed from neutrophils had no detectable alteration in its m.w., suggesting that a serine protease probably cleaves the DBP binding site, releasing DBP in an unaltered form. Cells treated with PMSF accumulate DBP vs time with over 90% of the protein localized to the plasma membrane. Purified neutrophil plasma membranes were used to screen a panel of protease inhibitors for their ability to suppress shedding of the DBP binding site. Only inhibitors to neutrophil elastase prevented the loss of membrane DBP-binding capacity. Moreover, treatment of intact neutrophils with elastase inhibitors prevented the generation of C5a co-chemotactic activity from DBP. These results indicate that steady state binding of DBP is essential for co-chemotactic activity, and further suggest that neutrophil elastase may play a critical role in the C5a co-chemotactic mechanism.
Expression of the Th1 Chemokine IFN-γ-Inducible Protein 10 in the Airway Alters Mucosal Allergic Sensitization in Mice
Authors: Wiley Ryan E. , Palmer Kay , Gajewska Beata U. , Stämpfli Martin R. , Alvarez David , Coyle Anthony J. , Gutierrez-Ramos José-Carlos , Jordana Manel 
Published: April 2014

AbstractAlthough the preliminary characterization of chemokines and their receptors has been prolific, comparatively little is known about the role of chemokines in the evolution of immune responses. We speculate that the preferential recruitment of a particular immune cell population has implications for the short- and long-term features of an adaptive response. To test this hypothesis, we employed adenovirus-mediated gene transfer to express the Th1-affiliated, CXC chemokine IFN-γ-inducible protein (IP) 10 in the airways of mice undergoing a mucosal sensitization regimen known to result in a Th2-polarized allergic response. This resulted in a ∼60–75% inhibition of eosinophils in the bronchoalveolar lavage (BAL); these inflammatory changes were accompanied by enhanced IFN-γ, ablated IL-4, and, peculiarly, unaltered IL-5 and eotaxin levels in the BAL. The effect of IP-10 expression was shown to be dependent on IFN-γ, as there was no statistically significant reduction in BAL eosinophilia in IFN-γ knockout mice subjected to the IP-10 intervention. Flow cytometric analysis of mononuclear cells in the lung revealed a ∼60% reduction in the fraction of CD4+ cells expressing T1/ST2, a putative Th2 marker, and a parallel increase in the proportion expressing intracellular IFN-γ following IP-10 treatment. The effect of IP-10 expression at the time of initial Ag encounter is persistent, as mice rechallenged with OVA following the resolution of acute inflammation exhibited reduced eosinophilia and IL-4 in the BAL. Collectively, these data illustrate that local expression of the chemokine IP-10 can introduce Th1 phenomena to a Th2-predisposed context and subvert the development of a Th2 response.
Mast Cell Tissue Inhibitor of Metalloproteinase-1 Is Cleaved and Inactivated Extracellularly by α-Chymase
Authors: Frank Brendon T. , Rossall J. Caleb , Caughey George H. , Fang Kenneth C. 
Published: April 2014

Abstract
We previously reported that mast cell α-chymase cleaves and activates progelatinase B (progel B). Outside of cells, progel B is complexed with tissue inhibitor of metalloproteinase (TIMP)-1, which hinders zymogen activation and inhibits activity of mature forms. The current work demonstrates that dog BR mastocytoma cells, HMC-1 cells, and murine bone marrow-derived mast cells secrete TIMP-1 whose electrophoretic profile in supernatants suggests degranulation-dependent proteolysis. α-Chymase cleaves uncomplexed TIMP-1, reducing its ability to inhibit gel B, whereas tryptase has no effect. Sequencing of TIMP-1’s α-chymase-mediated cleavage products reveals hydrolysis at Phe12-Cys13 and Phe23-Val24 in loop 1 and Phe101-Val102 and Trp105-Asn106 in loop 3 of the NH2-terminal domain. TIMP-1 in a ternary complex with progel B and neutrophil gelatinase-associated lipocalin is also susceptible to α-chymase cleavage, yielding products like those resulting from processing of free TIMP-1. Thus, α-chymase cleaves free and gel B-bound TIMP-1. Incubation of the progel B-TIMP-1-neutrophil gelatinase-associated lipocalin complex with α-chymase increases gel B activity 2- to 5-fold, suggesting that α-chymase activates progel B whether it exists as free monomer or as a complex with TIMP-1. Furthermore, inhibition of α-chymase blocks degranulation-induced TIMP-1 processing (absent in α-chymase-deficient HMC-1 cells). Purified α-chymase processes TIMP-1 in BR supernatants, generating products like those induced by degranulation. In summary, these results suggest that controlled exocytosis of mast cell α-chymase activates progel B even in the presence of TIMP-1. This is the first identification of a protease that overcomes inhibition by bound TIMP-1 to activate progel B without involvement of other proteases.
Effect of Thymectomy on Human Peripheral Blood T Cell Pools in Myasthenia Gravis
Authors: Sempowski Gregory D. , Thomasch James R. , Gooding Maria E. , Hale Laura P. , Edwards Lloyd J. , Ciafaloni Emma , Sanders Donald B. , Massey Janice M. , Douek Daniel C. , Koup Richard A. , Haynes Barton F. 
Published: April 2014

Abstract
The human thymus is required for establishment of the T cell pool in fetal life, but postnatal thymectomy does not lead to immunodeficiency in humans. Because thymectomy in humans is performed for treatment of myasthenia gravis (MG), we have studied patients with MG for effects of thymectomy on peripheral blood (PB) naive (CD45RA+, CD62L+) and memory (CD45RO+) T cells. We have also determined the effect of thymectomy on levels of PB cells containing signal joint TCR δ excision circles (TRECs), a molecular marker of thymus emigrants that have divided few times after leaving the thymus. In 17 nonthymectomized and 26 thymectomized MG patients studied at varying times after thymectomy (1 day to 41 years), we found no significant mean difference in PB T cell TREC levels between ages 40 and 80 years. However, both thymectomized and nonthymectomized MG patients had lower PB T cell TREC levels than did age-matched normal subjects (p &lt; 0.0001 for both). These data demonstrated that MG itself or treatment for MG decreased thymopoiesis independent of thymectomy. Next, to control for disease activity and treatment, we prospectively studied 10 MG patients before and from 27 to 517 days after thymectomy. We found that thymectomy decreased CD4 or CD8 T cell TREC concentrations most when thymopoiesis was active before thymectomy (six of six patients), but had little effect in patients when thymopoiesis was minimal (four of four patients). In contrast, there was no significant effect of thymectomy on absolute numbers of naive PB T cells. Thus, in MG, removal of a thymus with active thymopoiesis resulted in a significant fall in PB TREC+ T cells postthymectomy.
Human B Cell Growth and Differentiation in the Spleen of Immunodeficient Mice
Authors: Depraetere Stany , Verhoye Lieven , Leclercq Georges , Leroux-Roels Geert 
Published: April 2014

AbstractHuman mAbs (HumAbs) have therapeutic potential against infectious diseases and cancer. Heretofore, their production has been hampered by ethical constraints preventing the isolation of Ag-specific activated B cells by in vivo immunization. Alternatively, severe combined immune deficient (SCID) mice, transplanted i.p. with human (Hu)-PBLs, allow the in vivo stimulation of human Ab responses without the usual constraints. Unfortunately, human B cells only represent a minor fraction of the surviving graft, they are scattered all over the animal body, and thus are hard to isolate for subsequent immortalization procedures. To prevent this dispersion and to provide the human B cells with a niche for expansion and maturation, SCID mice were engrafted with Hu-PBL directly into the spleen. Simultaneously endogenous murine NK cell activity was depleted by treatment with an anti-mouse IL-2 receptor β-chain Ab. During engraftment, human B lymphocytes became activated, divided intensely, and differentiated into plasmacytoid cells. In vivo exposure to a recall Ag after cell transfer induced expansion of Ag-specific B cell clones. One week after inoculation, human B cells were abundant in the spleen and could easily be recovered for fusion with a heteromyeloma line. This resulted in the formation of stable hybridoma cell lines that secreted Ag-specific HumAbs. Thus transplantation of human lymphoid cells in the spleens of immune deficient mice represents a model for the study of human T cell-dependent B cell activation and proves to be an excellent tool for the successful production of HumAbs.
Thioredoxin Acts as a B Cell Growth Factor in Channel Catfish
Authors: Khayat Morad , Stuge Tor B. , Wilson Melanie , Bengtén Eva , Miller Norman W. , Clem L. William 
Published: April 2014

AbstractTo identify differentially expressed genes from channel catfish macrophages, a cDNA library from LPS-stimulated catfish macrophages was screened by subtractive hybridization. This screening yielded a 552-bp cDNA coding for catfish thioredoxin (CF-TRX). The deduced amino acid sequence revealed that CF-TRX contains 107 amino acids and is 59% homologous to human adult T cell leukemia-derived factor/TRX, originally described as an IL-2Rα-inducing factor. Northern blot analyses showed that CF-TRX is expressed in catfish T and macrophage cell lines, but weakly in B cell lines. Similar results were also observed in Western blot analyses using a mAb specific for recombinant CF-TRX (rTRX). The use of rTRX in functional studies demonstrated that rTRX induces in vitro proliferative responses of catfish PBL that were synergistically enhanced by the addition of culture supernatants from catfish T cell lines. In addition, cell separation studies and flow cytometric analyses revealed that the cells proliferating in rTRX-stimulated cultures were mostly B cells. These results suggest that CF-TRX may have an important role in the activation and proliferation of channel catfish B cells.
Induction of Polyclonal Prostate Cancer-Specific CTL Using Dendritic Cells Transfected with Amplified Tumor RNA
Authors: Heiser Axel , Maurice Margaret A. , Yancey Donna R. , Wu Ning Z. , Dahm Philipp , Pruitt Scott K. , Boczkowski David , Nair Smita K. , Ballo Michael S. , Gilboa Eli , Vieweg Johannes 
Published: April 2014

Abstract
Polyvalent cancer vaccines targeting the entire antigenic spectrum on tumor cells may represent a superior therapeutic strategy for cancer patients than vaccines solely directed against single Ags. In this study, we show that autologous dendritic cells (DC) transfected with RNA amplified from microdissected tumor cells are capable of stimulating CTL against a broad set of unidentified and critical prostate-specific Ags. Although the polyclonal CTL responses generated with amplified tumor RNA-transfected DC encompassed as a subcomponent a response against prostate-specific Ag (PSA) as well as against telomerase reverse transcriptase, the tumor-specific CTL were consistently more effective than PSA or telomerase reverse transcriptase CTL to lyse tumor targets, suggesting the superiority of the polyclonal response. Although tumor RNA-transfected DC stimulated CTL, which recognized not only tumor but also self-Ags expressed by benign prostate tissue, these cross-reactive CTL were exclusively specific for the PSA, indicating an immunodominant role of PSA in the prostate cancer-specific immune response. Our data suggest that tumor RNA-transfected DC may represent a broadly applicable, potentially clinically effective vaccine strategy for prostate cancer patients, which is not limited by tumor tissue availability for Ag preparation and may minimize the risk of clonal tumor escape.