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Joseph Alroy - One of the best experts on this subject based on the ideXlab platform.

  • Expression of CD3 Antigens in renal tubule epithelium and renal oncocytomas
    Pathology research and practice, 2005
    Co-Authors: Joseph Alroy, Barbara F. Banner, Angelo A. Ucci, Gisso Azabdoaftari, John C. Cheville
    Abstract:

    CD3 Antigen, formerly thought to be specific for T lymphocytes, has been identified in Purkinje cells of the cerebellum and gastric parietal cells in several species, including humans. The antibodies commonly used to recognize CD3 are directed against the e-subunit of the T cell receptor. This subunit has a role in signal transduction in T lymphocytes and possibly other types of cells. We immunostained sections for CD3 from normal kidneys of several species, including humans, and from different primary human renal cortical neoplasms to determine if CD3 Antigen is expressed in normal and in neoplastic tubular epithelium. CD3 expression was strong in normal proximal and distal tubular epithelium in most species and in renal oncocytomas, weak in chromophobe carcinoma, and negative in clear cell carcinomas, in papillary renal cell carcinoma, and in a transitional cell carcinoma. These findings suggest that this marker may be useful in the diagnostic workup and classification of renal cortical neoplasms.

  • Expression of CD3 ɛ Subunit in Gastric Parietal Cells: A Possible Role in Signal Transduction?
    Pathology research and practice, 2003
    Co-Authors: Barbara F. Banner, Zachary Spicer, Joseph Alroy
    Abstract:

    CD3 Antigen, formerly thought to be specific for T lymphocytes, has been found on gastric parietal cells in animals and humans. The common anti-CD3 antibodies recognize the epsilon subunit, which has a role in signal transduction. The aim of this study was to immunostain stomach specimens from humans and different animal species for CD3 Antigen to determine if CD3 Antigen is conserved across species and if CD3 Antigen expression is altered in humans by use of certain drugs or the presence of gastritis. Gastric biopsies from 50 humans and necropsy sections from 13 different animals were immunostained using commercial anti CD3 epsilon antibodies on an automated immunostainer. Sections of stomach from four mice lacking the gastric H+,K(+)-ATPase alpha-subunit and four control mice were similarly immunostained. CD3 epsilon Antigen expression in cytoplasm and cell membranes of gastric parietal cells was graded subjectively based on the number of positive cells. CD3 epsilon Antigen was found on gastric parietal cells in all but one species studied, with varying expression in membranes and cytoplasm. There was a trend toward a decreased frequency of CD3+ cells in biopsies from patients on drugs (n = 23) compared to those on no drugs (n = 27). This trend was most marked in patients on H2 receptor antagonists. There was no correlation between CD3 expression and inflammation or Helicobacter pylori (H. pylori) infection. There was loss of CD3 expression in parietal cells in mice lacking the alpha-subunit of H+,K(+)-ATPase. These findings support previous observations that CD3 Antigen is present in gastric parietal cells, and suggest that it may function in signal transduction during acid secretion.

Denis R. Alexander - One of the best experts on this subject based on the ideXlab platform.

  • CD3 Antigen mediated calcium signals and protein kinase c activation are higher in cd45r0 than in cd45ra human t lymphocyte subsets
    European Journal of Immunology, 1993
    Co-Authors: Anne T Robinson, Nigel G A Miller, Denis R. Alexander
    Abstract:

    T lymphocytes may be separated into subsets according to their expression of CD45 isoforms. The CD45R0+ T cell subset has been reported to proliferate in response to recall Antigen and to mitogenic mAb to a much greater extent than the CD45RA+ subset. This difference could be due to more efficient coupling of the T cell Antigen receptor complex to mitogenic signaling pathways. To investigate this possibility, CD3 Antigen-induced calcium signals, diacylglycerol (DAG) production and protein kinase C (PKC) activation levels were compared in CD45RA+ and CD45R0+ human T lymphocyte subsets derived from peripheral blood. The mean CD3-induced rise in intracellular calcium was 80% greater in CD45R0+ than in CD45RA+ cells. Basal DAG levels in CD45R0+ cells were found to be, on average, 60% higher than in CD45RA+ cells (p = 0.002), but the CD3-induced production of DAG over background was not different in the two subsets (p = 0.4). Basal PKC activity, and CD3-induced PKC activation levels over background, were found to be 50% and 140% higher, respectively, in CD45R0+ cells than in CD45RA+ cells (p = 0.015 and 0.023). The CD45R0+ subset contained a higher proportion of cells expressing activation markers, such as CD25, CD71 and major histocompatibility complex class II, when compared to the CD45RA+ subset. Our results suggest that the elevated basal DAG levels observed in the CD45R0+ subset may reflect the recent activation of these cells. Both the higher basal DAG and CD3-induced elevation in intracellular calcium observed in the CD45R0+ cells may contribute to the greater PKC activation signals triggered by CD3 mAb in this subset. These findings elucidate the greater response of CD45R0+ T cells to mitogenic stimuli compared to CD45RA+ cells.

  • Selective coupling of the T cell Antigen receptor to phosphoinositide‐derived diacylglycerol production in HPB‐ALL T cells correlates with CD45‐regulated p59fyn activity
    European journal of immunology, 1993
    Co-Authors: Mark Biffen, Masahiro Shiroo, Denis R. Alexander
    Abstract:

    In HPB-ALL T-cells the p59fyn tyrosine kinase is regulated by the CD45 phosphotyrosine phosphatase and plays a critical role in coupling the T cell receptor (TCR) to the generation of intracellular signals which include diacylglycerol (DAG) production and protein kinase C activation. The aim of this study was to determine the phospholipid pools from which the DAG is generated and to identify which phospholipase activities are regulated by the TCR. When CD45+ cells were pre-labeled with [3H]arachidonic acid, CD3-Antigen cross-linking stimulated negligible increases in both [3H]DAG and [3H]phosphatidic acid (PA). However, CD3 monoclonal antibody (mAb) induced an increase of 300% in [3H]PA when the cells were permeabilized with streptolysin-O, and this correlated with increased levels of protein tyrosine phosphorylation. Stimulation of [3H]PA production upon CD3 cross-linking was 77% lower in permeabilized CD45- cells than in CD45+ cells, consistent with the reduced activity of p59fyn in CD45- cells. The stimulated production of PA was not mediated by activation of phospholipase D (PLD), although the presence of a G-protein-regulated PLD activity was established. The CD3-induced increase in total inositol phosphates (InsP) in permeabilized cells was similar to the stimulated production of [3H]PA production in both CD45+ and CD45- cells. Dose-response curves for InsP and PA production triggered by CD3 mAb were super-imposable and the production of InsP and PA over a range of Ca2+ concentrations was comparable. Differential labeling of phospholipids with 3H-labeled fatty acids revealed that CD3-induced PA production reflected incorporation of label into the phosphatidylinositol pool. Our data suggest that in HPB-ALL cells the production of DAG following CD3-Antigen cross-linking can be fully accounted for by the selective coupling of the TCR to breakdown of phosphatidylinositol-(4,5)-bisphosphate as the result of phospholipase C gamma 1 activation. This event correlates with the activity of the CD45-regulated TCR-associated tyrosine kinase, p59fyn.

  • Does co-aggregation of the CD45 and CD3 Antigens inhibit T cell Antigen receptor complex-mediated activation of phospholipase C and protein kinase C?
    European journal of immunology, 1992
    Co-Authors: Emer Shivnan, Mark Biffen, Masahiro Shiroo, Edward Pratt, Martin J. Glennie, Denis R. Alexander
    Abstract:

    The binding of agonistic monoclonal antibodies (mAb) to the CD3 Antigen in T cells induces a rapid increase in tyrosine phosphorylation, inositive phosphate (IP) production, a rise in intracellular calcium and protein kinase C (PKC) activation. These intracellular signals have been implicated in the control of interleukin-2 and interleukin-2R receptor gene expression, thereby regulating T cell proliferation. Previous studies have shown that co-ligation of the CD45 and CD3 Antigens inhibits CD3-induced tyrosine phosphorylation, IP production, calcium signals and T cell proliferation. It has therefore been suggested that the CD45 Antigen uncouples the T cell receptor (TcR) from mitogenic signal pathways. In this study co-ligation of the CD3 and CD45 Antigens with precisely constructed bispecific mAb did not inhibit CD3-induced T cell proliferation, IP production, calcium signals, diacylglycerol production or PKC activation. Furthermore, co-ligation of CD3 and CD45 Antigens already cross-linked with IgM mAb did not lead to inhibition of CD3-induced calcium signals. Inhibitions of CD3-induced intracellular signals were observed following co-ligation of IgG CD45 and CD3 mAb with anti-IgG F(ab')2 fragments. However, comparable inhibitions were also noted following co-ligation of CD3 with other abundant cell-surface Antigens such as CD5 and LFA-1, and inhibitions were only observed when the CD3 mAb used required cross-linking to induce signals. These results suggested that the inhibitory effects of CD45 IgG mAb were not specific and were caused by the prevention of CD3-CD3 cross-linking following CD3 Antigen co-ligation with other cell surface molecules. These findings are inconsistent with a specific inhibitory role for the CD45 phosphotyrosine phosphatase in uncoupling the TcR from mitogenic signal pathways.

Pilar Portolés - One of the best experts on this subject based on the ideXlab platform.

  • Characteristics of TCR/CD3 complex CD3ɛ chains of regulatory CD4+ T (Treg) lymphocytes: role in Treg differentiation in vitro and impact on Treg in vivo
    Journal of leukocyte biology, 2013
    Co-Authors: José M. Rojo, Gloria Ojeda, Yenny Y. Acosta, María Montes-casado, Gabriel Criado, Pilar Portolés
    Abstract:

    Tregs are anergic CD4(+)CD25(+)Foxp3(+) T lymphocytes exerting active suppression to control immune and autoimmune responses. However, the factors in TCR recognition underlying Treg differentiation are unclear. Based on our previous data, we hypothesized that Treg TCR/CD3 Antigen receptor complexes might differ from those of CD4(+)CD25(-) Tconv. Expression levels of TCR/CD3, CD3e,ζ chains, or other molecules involved in Antigen signaling and the characteristics of CD3e chains were analyzed in thymus or spleen Treg cells from normal mice. Tregs had quantitative and qualitatively distinct TCR/CD3 complexes and CD3e chains. They expressed significantly lower levels of the TCR/CD3 Antigen receptor, CD3e chains, TCR-ζ chain, or the CD4 coreceptor than Tconv. Levels of kinases, adaptor molecules involved in TCR signaling, and early downstream activation pathways were also lower in Tregs than in Tconv. Furthermore, TCR/CD3 complexes in Tregs were enriched in CD3e chains conserving their N-terminal, negatively charged amino acid residues; this trait is linked to a higher activation threshold. Transfection of mutant CD3e chains lacking these residues inhibited the differentiation of mature CD4(+)Foxp3(-) T lymphocytes into CD4(+)Foxp3(+) Tregs, and differences in CD3e chain recognition by antibodies could be used to enrich for Tregs in vivo. Our results show quantitative and qualitative differences in the TCR/CD3 complex, supporting the hyporesponsive phenotype of Tregs concerning TCR/CD3 signals. These differences might reconcile avidity and flexible threshold models of Treg differentiation and be used to implement therapeutic approaches involving Treg manipulation.

Jai Dev Dasgupta - One of the best experts on this subject based on the ideXlab platform.

  • Defective signal transduction via T-cell receptor-CD3 structure in T cells from rheumatoid arthritis patients.
    Human immunology, 1993
    Co-Authors: Valerie Relias, Edmond J. Yunis, William N. Pachas, Jai Dev Dasgupta
    Abstract:

    Abstract T cells from patients with active RA are known to produce low levels of IL-2 and proliferate poorly in response to various mitogenic stimuli. The present work shows that cross-linking of CD3 Antigen on patients' T- cell surface induces two-to threefold lower Ca 2+ response than in T cells from age-matched controls. Immunofluorescence studies indicate that the attenuated signal may be due to the suppressed expression of CD3 and/or CD45 molecules on patients' T cells. In the majority of the patients, the level of CD45 expression is reduced by 60%–70% as compared with that in the control T cells. Therefore, the poor mitogenic response of patient cells is apparently due to a deffect in early stages of signal transduction through the T-cell receptor (TCR-CD3).

Michael J. Crumpton - One of the best experts on this subject based on the ideXlab platform.

  • Physical association of CD5 and the T cell receptor/CD3 Antigen complex on the surface of human T lymphocytes.
    European journal of immunology, 1993
    Co-Authors: Narin Osman, Andrew I. Lazarovits, Michael J. Crumpton
    Abstract:

    The physical association of CD5 and the T cell Antigen receptor (TcR)/CD3 complex on the surface of intact human lymphocytes was investigated using co-capping experiments and fluorescence resonance energy transfer (FRET) analyses. Antibody-induced capping of CD5 or CD3 and double indirect immunofluorescence labeling revealed a specific co-localization of a significant fraction of CD3 and CD5 molecules on the T cell surface. By means of FRET measurements we studied further the physical proximity of CD5 and the TcR/CD3 complex at the surface of normal lymphocytes. Significant fluorescence energy transfer was measured between CD5 and CD3 molecules indicating that the associated molecules were within 10 nm of one another. No energy transfer was observed between the integrin alpha 4 beta 7 and CD3 or CD5. The close physical proximity measured between CD5 and CD3 correlates with our co-capping data and taken together the results show that the association of CD5 and the TcR/CD3 complex first detected by immunoprecipitation occurs on the surface of human T cells under physiologically relevant conditions.

  • CD3 and CD2 Antigen-mediated CD3 gamma-chain phosphorylation in permeabilized human T cells. Regulation by cytosolic phosphatases.
    Biochemical Journal, 1992
    Co-Authors: D R Alexander, Michael J. Crumpton, M H Brown, A L Tutt, E Shivnan
    Abstract:

    The role of cytosolic and membrane-associated phosphatases in regulating dephosphorylation of the CD3 Antigen gamma-chain has been investigated using streptolysin-O-permeabilized T lymphoblasts and Jurkat T leukaemia cells. Permeabilization of T cells caused a rapid extrusion of cytosolic type 2A phosphatases, but a membrane-associated phosphorylase phosphatase activity remained inside the cells. This activity had the properties characteristic of type 2A phosphatases, being resistant to inhibition by type 1 phosphatase inhibitors, though it was inhibited in a time-dependent manner by ATP or by non-hydrolysable ATP analogues, but not by GTP, CTP, ITP or PPi. The membrane-associated type 2A phosphatase in permeabilized cells did not dephosphorylate the CD3 Antigen gamma-chain, suggesting that cytosolic phosphatases dephosphorylate the gamma-chain in situ. Cross-linking the CD2 and CD3 Antigens with a bivalent monoclonal antibody in the absence of cytosolic phosphatases induced marked phosphorylation of the CD3 gamma-chain, immunoprecipitated using a novel gamma-chain peptide analogue directed antiserum (TG1). Phosphorylation was inhibited by a protein kinase C (PKC) pseudosubstrate inhibitor, indicating that CD2/CD3-induced gamma-chain phosphorylation is a PKC-mediated event. Activation of T cells either with phorbol 12,13-dibutyrate or by CD2-CD3 cross-linking caused [32P]Pi incorporation into the same gamma-chain Ser residues. The site-mapping data suggested that PKC in situ may incorporate phosphate at the CD3 gamma-chain Ser-123 and Ser-126 residues, but that phosphate is rapidly lost from Ser-123 by cytosolic phosphatase action. Our findings underline the importance of the dual actions of kinases and phosphatases as potential regulators of T cell Antigen-receptor complex function.

  • evidence for an association between the t cell receptor CD3 Antigen complex and the cd5 Antigen in human t lymphocytes
    European Journal of Immunology, 1992
    Co-Authors: Narin Osman, Steven C. Ley, Michael J. Crumpton
    Abstract:

    In this work we report that CD5, a T cell accessory activation Antigen and receptor for the B cell surface protein CD72, is associated with the T cell Antigen receptor (TcR)/CD3 complex in human T lymphocytes. In vitro phosphorylation of either CD3 or CD5 immunoprecipitates prepared from CD3-stimulated Jurkat and peripheral blood T cells in the presence of the detergent polyoxyethylene 10 oleyl ether (Brij96) showed, unexpectedly, an identical pattern of five phosphopolypeptides of 70, 59, 56, 21 and 18 kDa, respectively. Peptide mapping of the five bands demonstrated that the same protein kinase substrates co-precipitated with both CD3 and CD5 and that the majority of the protein phosphorylation occurred on tyrosine residues. These data suggested that the TcR/CD3 complex and and the CD5 Antigen might be associated in T cells. Evidence to support this hypothesis was obtained from analysis of immunoprecipitates prepared from surface-iodinated T cells. Bands characteristic of the TcR and CD3 Antigens were identified in CD5 immunoprecipitates and conversely, CD5 was identified in CD3 immunoprecipitates. Conformation that CD3 and CD5 co-precipitated in the presence of Brij96 was obtained by Western blotting. Quantitative immunodepletion demonstrated that between 10 % –20 % of cell surface CD5 was associated with the TcR/CD3 complex in Brij96 detergent lysates of human T cells and, furthermore, that this association was independent of T cell activation. The association of these two receptors provides a possible physical basis for the accessory role of the CD5 Antigen in T cell activation.

  • Evidence for an association between the T cell receptor/CD3 Antigen complex and the CD5 Antigen in human T lymphocytes
    European journal of immunology, 1992
    Co-Authors: Narin Osman, Steven C. Ley, Michael J. Crumpton
    Abstract:

    In this work we report that CD5, a T cell accessory activation Antigen and receptor for the B cell surface protein CD72, is associated with the T cell Antigen receptor (TcR)/CD3 complex in human T lymphocytes. In vitro phosphorylation of either CD3 or CD5 immunoprecipitates prepared from CD3-stimulated Jurkat and peripheral blood T cells in the presence of the detergent polyoxyethylene 10 oleyl ether (Brij96) showed, unexpectedly, an identical pattern of five phosphopolypeptides of 70, 59, 56, 21 and 18 kDa, respectively. Peptide mapping of the five bands demonstrated that the same protein kinase substrates co-precipitated with both CD3 and CD5 and that the majority of the protein phosphorylation occurred on tyrosine residues. These data suggested that the TcR/CD3 complex and and the CD5 Antigen might be associated in T cells. Evidence to support this hypothesis was obtained from analysis of immunoprecipitates prepared from surface-iodinated T cells. Bands characteristic of the TcR and CD3 Antigens were identified in CD5 immunoprecipitates and conversely, CD5 was identified in CD3 immunoprecipitates. Conformation that CD3 and CD5 co-precipitated in the presence of Brij96 was obtained by Western blotting. Quantitative immunodepletion demonstrated that between 10 % –20 % of cell surface CD5 was associated with the TcR/CD3 complex in Brij96 detergent lysates of human T cells and, furthermore, that this association was independent of T cell activation. The association of these two receptors provides a possible physical basis for the accessory role of the CD5 Antigen in T cell activation.