bluuebunny/crossref_metadata_2025_split huggingface dataset:
Interleukin‐15‐induced maturation of human natural killer cells from early thymic precursors: selective expression of CD94/NKG2‐A as the only HLA class I‐specific inhibitory receptor
Authors: Mingari Maria Cristina , Vitale Chiara , Cantoni Claudia , Bellomo Rosa , Ponte Marco , Schiavetti Francesca , Bertone Stefania , Moretta Alessandro , Moretta Lorenzo 
Published: March 2007

AbstractImmature postnatal thymocytes were shown to contain precursors which, under suitable culture conditions, give rise to phenotypically and functionally mature natural killer (NK) cells. Here, we analyzed the effect of different cytokines for their ability to induce the expression of HLA class I‐specific inhibitory receptor(s) during the process of NK cell development from immature thymocytes. From thymocyte cell suspensions depleted of CD2+, CD3+, CD4+, CD8+, CD56+, and CD16+ cells, we further removed cells expressing HLA class I‐specific inhibitory receptors including CD94/NKG2‐A, p58.1, and p58.2 by immunomagnetic bead separation. The resulting cells did not contain any of the above NK receptors as determined by immunofluorescence and flow cytometric analysis, as well as by reverse transcriptase polymerase chain reaction (RT‐PCR) amplification using appropriate sets of primers. Although different cytokines have been used, including interleukin (IL)‐7, stem cell factor (SCF), IL‐2, and IL‐15, only IL‐2 or IL‐15 induced cell proliferation when used alone. Moreover, maturation towards CD3− CD56+ cells displaying cytolytic activity against the HLA class I− targets K562 or 221 was detectable in cultures containing IL‐15 used alone or in combination with IL‐7 or SCF. On the other hand, these CD3− CD56+ cell populations did not lyse HLA class I+ target cells, including autologous PHA blasts. Analysis of the expression of the various HLA class I‐specific inhibitory NK receptors revealed the presence of high proportions of CD94/NKG2‐A+ cells, while the NK receptors belonging to the Ig superfamily were undetectable both by immunofluorescence and by RT‐PCR analysis. The expression of CD94/NKG2‐A appeared to be responsible for the inability of cells to lyse HLA class I+ target cells. Thus, addition of anti‐CD94 monoclonal antibodies of IgM isotype resulted in lysis of autologous target cells. The use of 221 cells transfected with different HLA class I alleles as target cells confirmed the broad class I specificity of CD94/NKG2‐A receptor. Our experiments indicate that IL‐15 provides an appropriate stimulus to the expression of CD94/NKG2‐A, but not of other class I‐specific NK receptors in the process of maturation of NK cells from thymocyte precursors.
Thymocytes and RelB‐dependent medullary epithelial cells provide growth‐promoting and organization signals, respectively, to thymic medullary stromal cells
Authors: Naspetti Marianne , Aurrand‐Lions Michel , Dekoning Jenefer , Malissen Marie , Galland Franck , Lo David , Naquet Philippe 
Published: March 2007

AbstractThe thymic medulla is composed of distinct epithelial cell subsets, defined in this report by the reactivity of two novel antibodies, 95 and 29, raised against mouse thymic epithelial cell lines. These antibodies were used to probe the development of medulla in wild‐type or mutant thymuses. In CD3σ‐deficient mice where thymocyte maturation is arrested at the CD4− CD8− stage, few scattered 95+ and 29+ epithelial cells are found. When few mature thymocytes develop as in CD3‐ζ/η mice, expansion and organization of 95+ but not 29+ cells, becomes detectable. In RelB‐deficient mice, T cell maturation proceeds normally but negative selection is inefficient due to the lack of thymic medulla and dendritic cells. Strikingly, 29+ epithelial cells are absent and 95+ medullary epithelial cells are scattered throughout the thymus, intermingling with CDR1+ cortical epithelium. In chimeric mice lacking only dendritic cells, the corticomedullary junction persists and both 95+ and 29+ epithelial cells are localized in the medulla. These results suggest that two types of signals are required for development of thymic medulla. A growth signal depends upon the presence of maturing thymocytes, but organization of the thymic medulla requires the presence of activated 29+ medullary epithelial cells.
Lack of directed Vα14‐Jα281 rearrangements in NK1+ T cells
Authors: Shimamura Michio , Ohteki Toshiaki , Beutner Ulrich , Robson MacDonald H. 
Published: March 2007

AbstractNK1.1+ T cells are an unusual subset of TCRαβ cells distinguished by their highly restricted Vβ repertoire and predominant usage of an invariant Vα14‐Jα281 chain. To assess whether a directed rearrangement mechanism could be responsible for this invariant α chain, we have analyzed Vα14 rearrangements by polymerase chain reaction and Southern blot in a panel of cloned T‐T hybrids derived from thymic NK1.1+ T cells. As expected a high proportion (17/20) of the hybrids had rearranged Vα14 to Jα281. However, Vα14‐Jα281 rearrangements always occurred on only one chromosome and were accompanied by other Vα‐Ja rearrangements (not involving Vα14) on the homologous chromosome. These data argue that rigorous ligand selection rather than directed rearrangement is responsible for the high frequency of Vα14‐Jα281 rearrangements in NK1.1+ T cells.
On the mechanisms by which transforming growth factor‐β2 alters antigen‐presenting abilities of macrophages on T cell activation
Authors: Takeuchi Masaru , Kosiewicz Michele M. , Alard Pascale , Streilein J. Wayne 
Published: March 2007

AbstractPeritoneal exudate cells (PEC) incubated with antigen in the presence of transforming growth factor‐(TGF)‐β2 selectively suppress delayed hypersensitivity and IgG2a antibody production when injected intravenously into naive syngeneic recipients. In this study, we have examined in vitro the effects of TGF‐β2 on the antigen presenting abilities of PEC to activate DO11.10 T cells that express a transgenic T cell receptor that recognizes ovalbumin peptide fragment 323–339 in the context of I‐Ad. PEC were pretreated overnight with TGF‐β2, washed extensively, then co‐cultured with DO11.10 T cells in the presence of native OVA or P323–339. We found that TGF‐β2‐treated PEC induced the production of the T helper type 2 (Th2) cytokine, interleukin‐4 (IL‐4), but unlike untreated PEC, were unable to stimulate the Th1 cytokines, IL‐2 and interferon‐γ (IFN‐γ). Furthermore, TGF‐β2 was produced in an autocrine fashion by TGF‐β2‐treated PEC and was responsible for this shift to a Th2 response. This conclusion was supported by the following results. First, TGF‐β2‐treated PEC were found to express much more TGF‐β1 and TGF‐β2 mRNA than untreated PEC. Second, TGF‐β2‐treated PEC secreted large amounts of TGF‐β including its mature form. Third, addition of neutralizing anti‐TGF‐β2 antibodies, but not neutralizing anti‐TGF‐β1 antibodies, restored the ability of antigen‐pulsed, TGF‐β2‐pretreated PEC to stimulate DO11.10 T cells to secrete IL‐2 and IFN‐γ. These results indicate that antigen‐presenting cells that encounter antigen in a TGF‐β‐enriched environment (e.g., in the eye) shift responding naive T cells toward Th2 responses by producing TGF‐β during antigen presentation.
Analysis of HLA class Ib gene expression in male gametogenic cells
Authors: Fiszer Dorota , Ulbrecht Matthias , Fernandez Nelson , Johnson Judith P. , Weiss Elisabeth H. , Kurpisz Maciej 
Published: March 2007

AbstractWe have investigated mRNA expression for nonclassical MHC class I genes (HLA‐E,‐F,‐G) in human gametogenic cells. Testicular tissue was treated by collagenase and the resulting cell suspension was further purified by fractionation on Percoll gradients in a two‐step procedure. Three gametogenic cell fractions were analyzed: purified heterogenous suspension of gametogenic cells, fraction of round spermatids and fraction of elongated spermatids. Total RNA isolated from each cell population was subjected to both reverse transcriptase/polymerase chain reaction and Northern blot analysis using oligonucleotides specific for HLA‐E, ‐F and ‐G. Both method gave similar results. We have found a considerable level of HLA‐E mRNA, very low amounts of reamplified cDNA for HLA‐F and both a complete lack of mRNA and reamplified cDNA for the HLA‐G gene in the analyzed gametogenic cell fractions. Additionally, we have localized HLA‐E molecules on the cells of the adluminal compartment within seminiferous tubules using immunostaining with monoclonal antibodies specific for HLA‐E heavy chain followed by confocal microscopy analysis. The unique expression pattern of HLA class I antigens in the male gonad could play an important role in an efficient protection against an autoimmunological attack toward germ cells.
Crucial role of tumor necrosis factor (TNF) receptor 2 and membrane‐bound TNF in experimental cerebral malaria
Authors: Lucas Rudolf , Juillard Pierre , Decoster Els , Redard Mireille , Burger Danielle , Donati Yves , Giroud Christine , Monso‐Hinard Christine , De Kesel Toon , Buurman Wim A. , Moore Mark W. , Dayer Jean‐Michel , Fiers Walter , Bluethmann Horst , Grau Georges E. 
Published: March 2007

AbstractTumor necrosis factor (TNF) has been implicated in the pathogenesis of experimental cerebral malaria (CM), but the respective role of its two types of receptors has not been established. A significant increase in the expression of TNF‐receptor 2 (TNFR2, p75), but not of TNFR1 (p55), was found on brain microvessels at the time of CM in susceptible animals. Moreover, mice genetically deficient for TNFR2 (Tnfr2°) were significantly protected from experimental CM, in contrast to TNFR1‐deficient (Tnfr1°) mice, which were as susceptible as wild‐type mice. To identify the factors involved in the protection from CM conferred by the lack of TNFR2, we assessed in both knockout and control mice the serum concentrations of mediators that are critical for the development of CM, as well as the up‐regulation of intercellular adhesion molecule‐1 (ICAM‐1) in the brain microvessels. No significant difference in serum levels of TNF and interferon‐γ was found between infected wild‐type and Tnfr1° or Tnfr2° mice. Interestingly, the pronounced ICAM‐1 up‐regulation and leukocyte sequestration, typically occurring in brain microvessels of CM‐susceptible animals, was detected in infected control and Tnfr1° mice – both of which developed CM – whereas no such ICAM‐1 up‐regulation or leukocyte sequestration was observed in Tnfr2° mice, which were protected from CM. Making use of microvascular endothelium cells (MVEC) isolated from wild‐type, Tnfr1° or Tnfr2° mice, we show that soluble TNF requires the presence of both TNF receptors, whereas membrane‐bound TNF only needs TNFR2 for TNF‐mediated ICAM‐1 up‐regulation in brain MVEC. Thus, only in MVEC lacking TNFR2, neither membrane‐bound nor soluble TNF cause the up‐regulation of ICAM‐1 in vitro. In conclusion, these results indicate that the interaction between membrane TNF and TNFR2 is crucial in the development of this neurological syndrome.
T cell repertoire formation displays characteristics of qualitative models of thymic selection
Authors: Elliott James I. 
Published: March 2007

AbstractThe use of T cell receptor elements varies between mouse strains, reflecting a balance between positive and negative selection. The presence of H‐2E biases Vα and Vβ usage through major histocompatibility class II isotype preferences of V elements, and mammary tumor virus‐dependent, negative selection. Quantitative models of thymic selection predict that negative selection equates to ‘excess’ positive selection, whereas qualitative models suggest that positive and negative selection are opposing forces. This report attempts to distinguish between the models by assessing whether, at the level of the T cell repertoire, positive and negative selection have quantitative or qualitative characteristics. The data show that the effect of bearing Vα and Vβ regions which are both preferentially (or negatively) selected in the presence of H‐2E is additive or synergistic, whilst positive stimuli counteract negative ones. The data thus provide support for qualitative models of thymic selection.
PU.1 negatively regulates the CD11c integrin gene promoter through recognition of the major transcriptional start site
Authors: López‐Rodríguez Cristina , Corbí Angel L. 
Published: March 2007

AbstractCD11c integrin expression is restricted to myeloid cells and activated B lymphocytes, mainly through the collaborative action of Sp1 and members of the AP‐1 and C/EBP transcription factor families on the proximal region of the CD11c gene promoter. While analyzing the role of an initiator‐like sequence at the major transcriptional start site, an inverted consensus GGAA Ets binding site was identified as a negative regulatory element whose disruption increases the activity of the CD11c promoter. The GGAA element was specifically recognized by PU.1 in THP‐1 monocytic cells and by PU.1 and GABP‐related proteins in U937 promonocytic cells. Mutational analysis indicated that PU.1 recognition depends not only on the GGAA consensus element but also on flanking sequences. The functional relevance of PU.1 binding was assayed in transactivation experiments in HeLa cells, where PU.1 co‐expression led to a significant decrease in the activity of the CD11c promoter, demonstrating that PU.1 inhibits the activity of the CD11c promoter through a PU.1 binding site located at the major transcriptional start site (PU1‐5). The inhibitory action of PU.1 on CD11c is in contrast with its positive regulatory effect on the CD11b and CD18 integrin gene promoters, which might contribute to the differentially regulated expression of CD11b/CD18 and CD11c/CD18 during monocyte extravasation and terminal maturation. In addition, since PU.1 transcriptional activity correlates with macrophage proliferation, PU.1 might modulate CD11c gene transcription according to the proliferative state of the cell.
The leukocyte/neuron cell surface antigen OX2 binds to a ligand on macrophages
Authors: Preston Sandy , Wright Gavin J. , Starr Karen , Barclay A. Neil , Brown Marion H. 
Published: March 2007

AbstractThe OX2 membrane glycoprotein contains two immunoglobulin superfamily (IgSF) domains and seems likely to interact with other cell surface proteins. A soluble chimeric protein with the two IgSF domains of OX2 engineered onto domains 3 + 4 of rat CD4 antigen was expressed. To detect possible weak interactions, the chimeric protein was coupled to fluorescent covaspheresTM to provide a highly avid display of OX2. The OX2 covaspheres bound macrophages but not other cell types. The specificity of the interaction was demonstrated by blocking with Fab fragments of the OX2 monoclonal antibody (mAb). A new mAb, MRC OX88, was raised against macrophages which also blocked the interaction and presumably recognizes the ligand. The epitope for the MRC OX2 mAb and a site for ligand binding were mapped to domain 1 by site‐directed mutagenesis. The OX2 antigen is present on thymocytes, some lymphocytes, neurons and endothelial cells; thus, it has the potential to mediate interactions between these cell types and macrophages.
CD9, but not other tetraspans, associates with the β1 integrin precursor
Authors: Rubinstein Eric , Poindessous‐Jazat Virginie , Le Naour François , Billard Martine , Boucheix Claude 
Published: March 2007

AbstractThe molecules of the tetraspan superfamily have two unequal extracellular domains separated by four transmembrane (TM) domains. These molecules are associated on the cell surface with each other and with other partner molecules, in particular β1 integrins. We now show that CD9 associates with the precursor of the β1 integrin (preβ1). This association is detected as early as 15 min after metabolic labeling, and the use of Brefeldin A demonstrates that it does not require Golgi modifications of either CD9 or integrin β1. The specificity of this interaction is demonstrated by the fact that other tetraspans, CD63, CD81, and CD82, do not associate with preβ1, and CD9 does not associate with immature human histocompatibility leukocyte antigen class I. In order to localize the region of CD9 responsible for the association with the β1 integrin, we have generated two reciprocal chimeric CD9/CD82 molecules with the junction localized just after the third TM region. The large extracellular loop of CD9 or the fourth TM domain, or both, appear to be sufficient to mediate an association with the mature integrin with a high efficiency, compared to CD82. By contrast, association with preβ1 requires at least two regions of the molecule. Mutation of CD9 at the consensus site of the tetraspan superfamily, localized between the second and the third TM domain, did not impair the co‐precipitation of preβ1. Finally, because preβ1 associates with calnexin, we have investigated a possible association of CD9 with this chaperone molecule. CD9 associates with calnexin independently of its association with the β1 integrin, suggesting that calnexin could be involved in the processing of CD9.
Identification of a Stat‐6‐responsive element in the promoter of the human interleukin‐4 gene
Authors: Curiel Rafael E. , Lahesmaa Riitta , Subleski Jeff , Cippitelli Marco , Kirken Robert A. , Young Howard A. , Ghosh Paritosh 
Published: March 2007

AbstractInterleukin (IL)‐4 is an immunomodulatory cytokine produced by a number of cell types including T cells, basophils, and mast cells. This pleiotropic cytokine has a number of immunoregulatory functions; however, the molecular mechanisms controlling the transcription of this gene are not yet completely understood. Several studies have implicated a possible autoregulatory mechanism for its own expression. Here, we have identified a Stat‐6‐responsive element (Stat‐6RE) in the promoter of the human IL‐4 gene. Utilizing electrophoretic mobility shift analysis, we have demonstrated the presence of two specific IL‐4‐responsive DNA‐protein complexes in nuclear extracts of both human Th1 and Th2 clones. Phytohemagglutinin‐blasted peripheral blood T cells also generated an inducible complex in response to stimulation with IL‐4 and the IL‐4‐like cytokine IL‐13. Transient transfection of the murine pre‐B cell line BA/F3 stably transfected with the full‐length human IL‐4 receptor α chain demonstrated the ability of multicopy Stat‐6RE to initiate transcription from a heterologous promoter upon IL‐4 or IL‐13 stimulation. These results indicate a possible autocrine mechanism for the regulation of IL‐4 gene transcription through the Stat‐6RE as well as a possible mechanism for IL‐13 regulation of the human IL‐4 promoter.
Functional analysis of immunoreceptor tyrosinebased activation motif (ITAM)‐mediated signal transduction: the two YxxL segments within a single CD3ζITAM are functionally distinct
Authors: Sunder‐Plassmann Raute , Lialios Faye , Madsen Mari , Koyasu Shigeo , Reinherz Ellis L. 
Published: March 2007

AbstractFunctional analysis of the immunoreceptor tyrosine‐based activation motif (ITAM) derived from the membrane‐proximal ITAM of CD3ζ demonstrates that mutations at either the tyrosine or leucine residues in the N‐terminal YxxL segment of the ITAM abolish all signal transduction functions of this ITAM. In contrast, mutations at the tyrosine or leucine residues in the C‐terminal YxxL segment abrogate signals for interleukin (IL)‐2 production but do not prevent tyrosine phosphorylation of the N‐terminal tyrosine of the ITAM, lck association with the ITAM, activation of phospholipase C‐γl or calcium mobilization. Cross‐linking of chimeric receptors containing a C‐terminal YxxL leucine mutation induces tyrosine phosphorylation of ZAP70 but without stable binding to the phosphorylated ITAM. These results indicate that the two YxxL segments in an ITAM are functionally distinct and that both are essential for ZAP70 binding and IL‐2 production. Furthermore, tyrosine phosphorylation of ZAP70 per se is not sufficient to trigger the downstream events leading to IL‐2 production. Substitution of an alanine for the bulky side chain at the Y+1 position of the N‐terminal YxxL segment reduces the receptor cross‐linking requirement necessary to achieve cellular activation and the absolute dependence on lck in this process. Our results reveal that both the number of ITAM as well as the specific amino acid residues within a single ITAM determine the extent of chimeric receptor cross‐linking required to trigger tyrosine phosphorylation‐dependent signaling events.
Differentiation of CD4highCD8low coreceptor‐skewed thymocytes into mature CD8 single‐positive cells independent of MHC class I recognition
Authors: Barthlott Thomas , Kohler Hubertus , Pircher Hanspeter , Eichmann Klaus 
Published: March 2007

AbstractThymocytes with a CD4hiCD8lo coreceptor‐skewed (CRS) phenotype have been shown to contain precursors for CD8 single‐positive (SP) thymocytes, in addition to precursors for CD4 SP cells. The selection mechanisms that stimulate CD4hiCD8lo cells to revert to the CD8 lineage are not known. Mice transgenic (tg) for the major histocompatibility complex (MHC) class I‐restricted P14 T cell receptor (TCR), on the H‐2bm13 background, generate a large number of CD4hiCD8lo CRS thymocytes. We analyzed the developmental potential and the differentiation requirements of the CD4hiCD8lo population of these mice. Using reaggregate thymic organ cultures (RTOC), we observed that these cells efficiently and almost exclusively differentiate into CD8 SP cells. Differentiation occurred independent of whether or not the MHC haplotype of the thymic stroma corresponds to the MHC restriction of the tg TCR. Loss of CD4 was independent of thymic stroma, up‐regulation of CD8 to full levels was dependent on thymic stroma but independent of MHC haplotype. After trypsin treatment and overnight incubation, these CRS cells re‐expressed CD8 but failed to re‐express CD4, indicating that they are in the process of terminating CD4 synthesis. CD8 SP cells derived from the CRS cells proliferate in response to peptide‐pulsed antigen‐presenting cells. Our data suggest that CD4hiCD8lo CRS thymocytes bearing the P14 tg TCR have completed positive selection and differentiate autonomously into functionally competent CD8 SP cells.
B cell subpopulations separated by CD27 and crucial collaboration of CD27+ B cells and helper T cells in immunoglobulin production
Authors: Agematsu Kazunaga , Nagumo Haruo , Yang Fen‐Chun , Nakazawa Takayuki , Fukushima Keitaro , Ito Susumu , Sugita Kanji , Mori Tetsuo , Kobata Tetsuji , Morimoto Chikao , Komiyama Atsushi 
Published: March 2007

AbstractB cell immunoglobulin production is regulated by helper T cells through direct interaction and secreted cytokines. In the present study, we functionally analyzed CD27 in cord and peripheral blood B cells. Adult peripheral blood B cells were separated into CD27+ and CD27− cells, which differed in their morphology. Cord blood B cells did not express CD27, and CD27 expression on peripheral blood B cells increased with age. Only CD27+ B cells had the ability to produce immunoglobulin, which was increased by contact with a tumor necrosis factor‐related transmembrane ligand, CD70. Adult peripheral blood CD27+ B cells can be further subdivided into two discrete subtypes: IgD−CD27+ and IgD+ CD27+ B cells. IgD− CD27+ B cells produce IgG, IgM and IgA, whereas IgD+ CD27+ B cells predominantly produce IgM. The addition of activated CD4+ CD45RO T cells expressing CD70 caused down‐regulation of CD27 expression on activated B cells, and this down‐modulation was completely blocked by anti‐CD70 monoclonal antibody, indicating direct T‐B cell contact via CD27/CD70. The triggering via CD27 and CD40 additively increased the immunoglobulin production under Staphylococcus aureus Cowan strain plus interleukin‐2 stimulation. Taken together, our findings demonstrate that peripheral blood B cells are separated into subpopulations by CD27 and IgD expression and that CD27+ B cells produce large amounts of immunoglobulin by interaction with the CD70 molecule.