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Biomedical subjects

B Fleischer

Publications and source records attributed to B Fleischer.

At least 199 records · Page 11Linked to original sources
PubMed1989

Response of synovial fluid T cell clones to Yersinia enterocolitica antigens in patients with reactive Yersinia arthritis.

From synovial fluids of two patients with reactive arthritis following Yersinia enterocolitica infection, T lymphocyte clones were obtained that showed proliferative responses to Y. enterocolitica. The responses required autologous T-cell-depleted peripheral blood mononuclear cells as antigen presenting cells. Three clones were studied in detail; two of them showed a marked and specific response to Yersinia antigens alone, the other one recognized both Yersinia and Salmonella typhimurium antigens. The antigen-specific proliferation of the clones could be completely blocked by monoclonal antibodies to HLA-DR. These experiments show that synovial T lymphocytes presumably involved in the in situ immune response to microbial antigens triggering reactive arthritis can be cloned directly from the site of inflammation.

Adult↗
PubMed1988

A T cell receptor-associated GTP-binding protein triggers T cell receptor-mediated granule exocytosis in cytotoxic T lymphocytes.

To study the subcellular events occurring after T cell activation we used cloned human CTL permeabilized with alpha-toxin of Staphylococcus aureus. This method of permeabilization leads to stable transmembrane channels that permit the introduction of small molecules into the cell but preserves the cellular structures and macromolecular contents of the CTL. We used the exocytosis of CTL-specific serine esterases as a marker of T cell activation. The TCR-activated exocytosis is functioning in such permeabilized CTL. Introduction of the membrane impermeable guanosine nucleotide-binding protein (G-protein) activating GTP-analog GTP gamma S into CTL triggers exocytosis if Ca2+ is present. For optimal exocytosis ATP is required. The G-protein inactivating GDP-analog GDP beta S inhibited exocytosis triggered via the TCR-CD3 complex but not that triggered by activating the protein kinase C. If the protein kinase C was depleted in CTL by overnight incubation with phorbolester, the response to GTP-gamma S was reduced by more than 50%. These experiments demonstrate the presence of a G-protein involved in TCR-mediated CTL triggering. In the sequence of signaling steps this G-protein is localized after TCR-triggering but before the formation of the protein kinase C-activating phosphoinositol breakdown product diacylglycerol in the sequence of signaling steps.

Bacterial Toxins↗
PubMed1988

Triggering of cytotoxic T lymphocytes and NK cells via the Tp103 pathway is dependent on the expression of the T cell receptor/CD3 complex.

The expression and function of the T cell activation molecule Tp103 on human cloned cytotoxic CD3+ and CD3- cells were studied. All in vitro growing CD3+ and CD3- clones expressed Tp103 regardless of their phenotype and the expression of a CD3-associated TCR complex. Whereas the CD2 pathway was functional in all these clones, only CD3-expressing clones could be triggered via Tp103 to kill target cells. In contrast, both CD2 and Tp103 pathways were suppressed after modulation of the TCR complex with anti-CD3 mAb. This indicates that the function of Tp103 but not of CD2 is dependent on the expression of a functional Ag receptor on cytotoxic T cells. Furthermore, modulation of the Ag receptor induces a state of unresponsiveness in cytotoxic T cells that cannot be attributed to just the removal of the CD3/TCR complex from the cell membrane.

Antigens, Differentiation, T-Lymphocyte↗
PubMed1988

Inactivation of a T cell receptor-associated GTP-binding protein by antibody-induced modulation of the T cell receptor/CD3 complex.

TCR modulation induced by anti-TCR or anti-CD3 mAbs leads to a transient state of refractoriness of the T cell to all signals given via cell surface structures. To investigate the underlying mechanisms, we have used human CTL permeabilized with the alpha toxin of S. aureus. This method of permeabilization allows manipulation of the interior milieu of the cell, but maintains its functional and structural integrity. Introduction of the G protein activator GTP gamma S into permeabilized CTL leads to triggering of granule exocytosis. The G protein inactivator GDP beta S inhibited exocytosis induced by TCR triggering but not that induced by activation of protein kinase C. This indicates that the G protein that triggers exocytosis is localized after CD3 triggering but before formation of the polyphosphoinositol breakdown product diacylglycerol. In TCR-modulated CTL, GTP gamma S is no longer able to activate exocytosis. Such CTL, however, still respond to PKC activators. This demonstrates that a TCR-associated G protein has been functionally inactivated by TCR modulation.

Antigen-Antibody Complex↗
PubMed1988

A regulatory role for the CD4 and CD8 molecules in T cell activation.

The role of the CD4 and CD8 molecules in T cell activation is presently a matter of controversy. Although their role as associative binding elements to MHC class II or class I is well documented, their influence on the triggering process in unclear. Because antibodies to CD4 or CD8 block T cell activation in the absence of their respective ligands, a negative signaling by these molecules has been suggested. However, recent experimental evidence argues against a negative regulatory effect of these molecules, since, e.g., simultaneous cross-linking of TCR and CD4 leads to enhanced T cell activation. Therefore, a current model suggests that the association of TCR and CD4 in the membrane gives a positive signal essential for triggering. In this report we present evidence that this model is likely to be too simple. Anti-CD4 and CD8 antibodies inhibit alternative, nonreceptor pathways of T cell triggering via Tp103 and Tp44 in the absence of class II positive accessory or target cells. These antibodies also inhibit bypass activation of T cells by phorbol ester and calcium ionophore in an accessory cell-free system. Furthermore, if the CD4 or CD8 molecules are removed from the cell surface by antibody-induced modulation, the proliferative and cytotoxic response of T cell clones is enhanced. This enhancement is also observed if resting peripheral blood T cells are used as responder cells. These data show that the CD4 or CD8 molecules have a complex regulatory function in T cell activation beyond the requirement for co-cross-linking with the TCR.

Adjuvants, Immunologic↗
PubMed1988

Non-specific propagation of human antigen-dependent T lymphocyte clones.

Most T lymphocyte clones require periodical restimulation with specific antigen in addition to growth factors to maintain long-term growth in vitro. Since in many cases the specific antigen for a given T cell clone is not known or not available, alternative methods of stimulation are required. In this report a method for a non-antigen-specific stimulation of antigen-dependent proliferative or cytotoxic human T cell clones is described. Introduction of aldehydes into galactose residues on stimulator cells leads to the stimulation of T cell clones by modified stimulator cells in the absence of a mitogen. This system is as efficient as the use of mitogenic antibodies or lectins but avoids the disadvantages of these polyclonal T cell activators.

Antigens↗
PubMed1988

T cell stimulation by staphylococcal enterotoxins. Clonally variable response and requirement for major histocompatibility complex class II molecules on accessory or target cells.

Staphylococcal enterotoxins (SE) are the most potent mitogens for T lymphocytes known; concentrations of less than 10(-9) M are sufficient for T cell activation. The mechanism of T cell activation by SE is unknown. We have used cloned human cytotoxic and proliferative T lymphocytes to dissect the molecular mechanism of T cell activation by SE. With rare exceptions, all TCR alpha/beta chain-expressing T cell clones of CD4+ or CD8+ phenotype, as well as CD4-8- TCR alpha/beta chain negative chain-expressing T lymphocyte clones, respond with proliferation and/or cytotoxicity to SE. For triggering of all these clones, the presence of autologous or allogeneic MHC class II molecules on accessory or target cells is necessary. This requirement for class II antigens is not due to an immunological recognition of processed SE, since inhibition of antigen processing has no influence on the T cell response to SE. SE acts on the T cells directly since (a) they stimulate a rise in intracellular calcium concentration in T cell lines or purified T cells, and (b) accessory cells can be replaced by phorbolesters in the proliferative activation of resting T cells by SE. Furthermore, the T cell response to SE shows extensive clonal heterogeneity. These results suggest that SE are functionally bivalent mitogens binding highly selectively to HLA class II molecules and the TCR. Thus, compared with other polyclonal T cell activating agents, activation with SE most closely mimicks the physiological way of MHC-restricted antigen recognition by T lymphocytes.

Antigen-Presenting Cells↗
PubMed1988

Clonal analysis of human T cell activation by the Mycoplasma arthritidis mitogen (MAS).

Mycoplasma arthritidis produces an as yet undefined soluble molecule (MAS) that has a potent mitogenic effect on T cells of several species. We have used cloned human cytotoxic and proliferative T lymphocytes to dissect the molecular mechanism of T cell activation by this mitogen. Reactivity to MAS is clonally expressed among T cell receptor (TcR) alpha/beta chain-expressing T cell clones of CD4+ or CD8+ phenotype, as well as CD4-8- TcR alpha/beta chain-negative T lymphocyte clones expressing the CD3-associated TcR gamma chain. MAS is able to induce cytotoxicity and/or proliferation in these T cell clones. For triggering of these T cells, regardless of their phenotype of specificity, the presence of autologous, allogeneic or xenogeneic major histocompatibility complex (MHC) class II molecules on accessory cells or target cells is necessary. However, T cells do not immunologically recognize MAS on class II molecules, since a direct action of MAS on the T cells themselves can be demonstrated. Triggering of T cells by MAS can be blocked by monoclonal antibodies against CD2, CD3 and the TcR alpha/beta chain dimer. We discuss as a possible explanation that MAS is a functionally bivalent molecule cross-linking TcR and MHC class II molecules. Thus, the mechanism of T cell activation by MAS has striking similarities to the mechanisms by which Staphylococcal enterotoxins activate T cells. It is intriguing that a similar mitogenic principle has been developed by two evolutionary distinct pathogenic microorganisms.

Antigen-Antibody Reactions↗
PubMed1987

Mitogenic activity of staphylococcal protein A is due to contaminating staphylococcal enterotoxins.

Soluble protein A from S. aureus is widely used as a polyclonal activator of human T cells. However, recombinant protein A produced in E. coli does not show any mitogenic properties, although its IgG-binding activity is identical to protein A purified from S. aureus. Antisera against the staphylococcal enterotoxins A and B are able to specifically inhibit the response of human T lymphocytes to protein A from S. aureus. Therefore, the mitogenic principle of this extensively used T cells activator is due to minute contaminations by enterotoxins that are active in picomolar concentrations.

Cells, Cultured↗
PubMed1987

T lymphocyte control of human eosinophilic granulopoiesis. Clonal analysis in an idiopathic hypereosinophilic syndrome.

The idiopathic hypereosinophilic syndrome (HES) comprises a heterogeneous group of disorders with unknown pathogenesis characterized by persistent peripheral blood and bone marrow eosinophilia and eosinophilic infiltrates of multiple organs leading to severe organ dysfunction. In the present study, T lymphocyte clones were randomly established from the blood of a patient with HES and propagated in culture with mitogen and interleukin 2. Whereas 28 of 29 clones were able to stimulate myeloid colony formation when co-cultured with normal bone marrow cells in a double-layer micro-agar culture system, one third of these clones preferentially stimulated pure eosinophil colonies (up to 98% of all colonies). This pattern differed markedly (p less than 0.001) from the pattern of release of hemopoietic factors by 126 T cell clones established from four other individuals. Eosinophil colony stimulation was due to the release of a lineage-specific eosinophilic colony-stimulating factor (Eo-CSF) by these clones after appropriate stimulation. Production of Eo-CSF in vitro was inhibited by hydrocortisone or cyclosporin A. All Eo-CSF-producing clones had the T4+8-phenotype and were capable of producing in addition interleukin 2 and interferon-gamma. Southern blot analysis of the T cell receptor beta-chain rearrangement of the Eo-CSF-producing clones showed a different rearrangement pattern for each clone. These studies suggest a reactive T cell-mediated eosinophilia as the pathogenetic mechanism in this case of HES and, for the first time, point to a biologic relevance of a lymphokine-induced stimulation of hemopoiesis.

Adult↗
PubMed1987

A novel pathway of human T cell activation via a 103 kD T cell activation antigen.

A novel triggering signal for human proliferating and cytotoxic T lymphocytes defined by a 103 kD T cell-specific activation antigen (Tp103) is described. Tp103 is expressed on all proliferating normal T cells but is not present, or present only in low amounts, on resting peripheral blood T lymphocytes. Cross-linking of T cell and Fc receptor-positive accessory or target cells by an antibody against Tp103 leads to activation of the T cell. The proliferative response is due to an autocrine IL 2-dependent mechanism and can be inhibited by antibodies against the IL 2 receptor or by Cyclosporin A. Resting Tp103-positive T cells also respond to anti-Tp103. Although Tp103 is not linked to the antigen receptor/T3 complex, triggering via Tp103 can be inhibited by modulation of the T3 molecule. Thus, Tp103 defines a new antigen-independent pathway of T cell activation that can be regulated via other T cell surface structures.

Antigens, Differentiation, T-Lymphocyte↗
PubMed1987

Antigen-presenting T cells. II. Clonal responses of alloreactive and virus-specific self-restricted human cytotoxic T cell responses stimulated by T lymphoblasts.

The capacity of various stimulator cell types to present alloantigens or viral antigens to resting human CD8+ cytotoxic lymphocyte precursors (CLP) was analyzed in a limiting dilution culture system. Cell sorter-separated T lymphoblasts of both CD4+ and CD8+ phenotypes but not resting T cells were found to efficiently stimulate the clonal development of allogeneic CD8+ CLP. Thus, 5000 CD4+ T lymphoblasts activated as many (one out of 200 to one out of 300) allogeneic CLP as 50,000 peripheral blood mononuclear stimulator cells. This potent stimulator activity was found in CD4+ and CD8+ T lymphoblasts activated by mitogen, anti-T3 monoclonal antibody, or mixed leukocyte reactions. Cytotoxic T cells generated in this system were highly specific for HLA class I antigens. Furthermore, T lymphoblasts infected with mumps virus efficiently induced development of autologous CLP into CTL clones that were virus specific and self-HLA restricted, as shown by split-well analysis. The possible in vivo significance of antigen-presenting T lymphoblasts is discussed.

Antigen-Presenting Cells↗
PubMed1987

Human mumps virus-specific cytotoxic T lymphocytes: quantitative analysis of HLA restriction.

To obtain quantitative information about the use of HLA antigens as restriction element by antiviral cytotoxic T lymphocytes (CTL), we have analyzed precursors of human mumps virus-specific CTL by limiting dilution. CTL generated by restimulation of peripheral blood T lymphocytes with autologous mumps virus (MV)-infected stimulator cells were restricted by autologous HLA class I antigens, and derived from the T4-8+ population. They were specific for MV and did not lyse autologous target cells infected with other viruses. Frequencies of MV-specific CTL precursors ranged from 1/500 to 1/8000. HLA restriction was analyzed by split-well analysis of individual CTL colonies. CTL recognizing HLA-A or B antigens were unequally distributed: HLA-B7, -B13, and -B27 were found to function as predominant, in some cases as exclusive, restriction elements, whereas other antigens such as HLA-A24 were never or rarely used. In several combinations, there was no evidence for antigenic variants of HLA molecules as reason for the failure to be recognized. The proportion of CTL precursors recognizing HLA-A2 and -B8 seemed to be dependent on the presence or absence of "dominant" restriction elements. We conclude that CTL precursors recognizing certain virus-HLA combinations are preferentially expanded during an infection, but that low responsiveness to a given combination is not necessarily absolute.

Antigens, Viral↗
PubMed1986

Characterization of polyisoprenyl phosphate phosphatase activity in rat liver.

The polyisoprenyl phosphate dephosphorylating activity of rat liver has been investigated with regard to substrate specificity, subcellular distribution, and transmembrane orientation. Total liver microsomes were employed as a source of enzymatic activity against a variety of 32P-labeled substrates. Susceptibility to dephosphorylation followed the order solanesyl phosphate greater than alpha-cis-polyprenyl 19-phosphate = alpha-trans-polyprenyl 19-phosphate = dihydrosolanesyl phosphate greater than (S)-dolichyl 19-phosphate = (R)-dolichyl 19-phosphate = (R,S)-dolichyl 11-phosphate. There appeared to be no major effect of chain length from 11 to 20 isoprenes. Data obtained from inhibition studies using solanesyl [32P]phosphate as substrate were consistent with the substrate specificity studies and suggested that a single activity is responsible. With dolichyl [32P]phosphate as substrate, the phosphatase specific activity of the subcellular fractions prepared from rat liver was found to follow the sequence Golgi = smooth endoplasmic reticulum greater than plasma membrane greater than lysosomes = rough endoplasmic reticulum greater than nuclei greater than mitochondria. Transmembrane topography studies, using enzyme latency as a criterion, were consistent with an orientation of the active site facing the cytoplasm.

Animals↗