Human autoreactive T-cell clones show flexibility in recognizing overlapping microepitopes, but inflexibility in T-cell receptor usage.
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Autoantigens↗
Biomedical subjects
M Londei
Publications and source records attributed to M Londei.
High level of interleukin-10 production by the activated T cell population within the rheumatoid synovial membrane.
OBJECTIVE: To characterize the cytokine profile of the activated T cell population derived from the synovial membrane of rheumatoid arthritis (RA) patients. METHODS: Interleukin-2 (IL-2) was used to select for in vivo-activated T cells from the synovial membrane of 2 patients with RA, and the cells were cloned nonspecifically. The cytokine production profile of these clones was compared with that of clones derived from peripheral blood monocytes (PBM) by stimulating all clones for 24 hours with immobilized anti-CD3 (coated at 10 micrograms/ml) or phorbol-12-myristate-13-acetate (10 ng/ml) plus soluble anti-CD3 (1 microgram/ml). Interferon-gamma (IFN gamma), IL-4, and IL-10, the cytokines that discriminate between Th1 and Th2 cells and are involved in immunoregulation, were assayed by enzyme-linked immunosorbent assay. RESULTS: There was a difference in the cytokines produced by the clones derived from the rheumatoid membranes compared with clones derived from the periphery. Clones derived from both membranes and PBM were mostly IFN gamma-producers, i.e., either a Th0 or a Th1 profile. There was a high proportion of IFN gamma/high IL-10-producing cells derived from the joint, but not from the periphery. Of clones derived from the synovial membrane of each of 2 RA patients, 100% and 50% produced both 1-10 ng/ml IFN gamma and > 7 ng/ml IL-10, compared with < 7% of clones derived from normal or RA peripheral blood. In addition, when autologous membrane and PBM were compared, the mean concentration of IL-10 produced by the clones derived from the synovial membrane sample was significantly different from those produced by clones derived from peripheral blood (P < 0.02). CONCLUSION: The cytokine profile of the T cell clones that were obtained from the RA joint after expansion with IL-2 is distinct from that of the T cells that are predominant in PBM. This supports the concept that the T cells subsets that accumulate in the joint are not a random sample. The high level of IL-10 production by clones derived from the synovium suggests that this cytokine may be a major contributor to the endogenous immunosuppression that occurs in RA.
Adult↗
Differential effects of interleukin-10 on the expression of HLA class II and CD1 molecules induced by granulocyte/macrophage colony-stimulating factor/interleukin-4.
Interleukin (IL)-10 down-regulates HLA class II molecules, whether constitutively expressed or up-regulated by interferon-gamma or IL-4 on monocytes but not on B lymphocytes. In this study we show that IL-10 does not inhibit HLA class II expression induced by the combination granulocyte/macrophage colony-stimulating factor and IL-4 on monocytes, although it simultaneously abrogates the expression of CD1 molecules induced by the same combination of cytokines. CD1 molecules can act as element of genetic restriction for CD4- CD8- T lymphocytes, and the suppression of CD1 expression by IL-10 abolished antigen presentation to CD1-restricted CD4- CD8- T cell receptor-positive T cells. Although HLA class II expression was not down-regulated by IL-10, the antigen specific proliferative response of CD4+ T cells was nevertheless decreased. This was not caused by down-regulation of known co-stimulatory molecules such as B7.1, B7.2 and ICAM-1. IL-10 decreased the antigen specific proliferative response further by directly influencing the T lymphocytes. Our results indicate that IL-10 exerts some of its immunoregulatory functions by differential modulation of antigen presenting molecules, induced by the same combination of cytokines.
Antigen Presentation↗
T-cell autoimmunity in multiple sclerosis.
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Animals↗
Comparison of peptide and superantigen-induced anergy in a peptide-specific polyclonal human T cell line.
T cells recognizing tetanus toxin peptide 'p2' (sequence 830-844) raised in HLA DR6 individuals preferentially express V beta 2 in the TCR. A p2-specific T cell line (60% V beta 2+) was used to compare peptide and superantigen [toxic shock syndrome toxin-1 (TSST-1)]-induced clonal anergy. Many experiments consistently revealed that the degree of 'tolerance' or 'clonal anergy' induced by peptide was greater than with the superantigen TSST-1. These results are of interest in a number of contexts. First they suggest that using superantigens or anti-V beta to delete the majority population of T cells may not be sufficient to diminish an autoimmune response. Secondly, the results indicate that induction of anergy of a large proportion of peptide-specific T cells does not lead to a suppressive bystander effect on the remaining responsive T cells. These results emphasize the need to define the dominant autoantigenic epitopes in human autoimmune diseases, since peptide based therapy such as the use of peptide analogues to induce anergy or a change in cytokine profile, is possibly more effective in controlling undesired immune responses than the use of non-antigen, TCR-directed approaches such as superantigens.
Amino Acid Sequence↗
Are CD4+ Th1 cells pro-inflammatory or anti-inflammatory? The ratio of IL-10 to IFN-gamma or IL-2 determines their function.
Human CD4+ T cells have, like their murine counterparts, been classified on the basis of their cytokine profile. Th1 cells produce IL-2 and IFN-gamma, but little or no IL-4. Th2 cells produce IL-4 but not IFN-gamma or IL-2, and Th0 produce IL-2, IL-4 and IFN-gamma. As IL-2 is the most potent T cell growth factor and IFN-gamma is the strongest activator of macrophages it is not surprising that CD4+ Th1 cells are considered to be pro-inflammatory. However, unlike results in the mouse, where IL-10 is only produced by Th2 cells, human IL-10 is produced by Th0, Th1 and Th2 cells. Hence some human Th1 cells are capable of producing both pro-inflammatory (IL-2, IFN-gamma) and anti-inflammatory (IL-10) cytokines, therefore the function of these cells may not be accurately encapsulated by the 'Th1' terminology. We thus investigated the correlation of cytokine production and function in human CD4+ Th1 clones. Cytokine production (IL-2, IFN-gamma, IL-10) was measured in supernatants by ELISA after stimulation with solid-phase anti-CD3. The capacity of these supernatants to activate or inhibit T cell proliferation or LPS induced TNF-alpha production by monocytes was assessed. The ratio of IL-2/IL-10 or IFN-gamma/IL-10 was of critical importance in determining the function of the supernatants. The inhibitory effects were verified to be due to IL-10, as they were neutralized by anti-IL-10 mAb.(ABSTRACT TRUNCATED AT 250 WORDS)
Antibodies, Monoclonal↗
Identification of thyroid stimulating hormone receptor-specific T cells in Graves' disease thyroid using autoantigen-transfected Epstein-Barr virus-transformed B cell lines.
The importance of thyrotropin receptor (TSHR) agonist antibodies in the manifestations of Graves' disease (GD) is recognized. There are, however, no convincing reports of TSHR-specific T cells. We have previously cloned T cells specific for thyroglobulin and thyroid peroxidase (TPO) from GD lymphoid infiltrates and used autologous EBV-transformed B cell lines (EBVL) transfected with an expression vector encoding TPO to efficiently detect TPO-specific T cells. Here we used EBVL transfected with TSHR to seek TSHR-specific T cells in the GD infiltrates, after cloning the in vivo activated T cells without antigen. 3 out of 30 clones responded vigorously and reproducibly to EBVL-TSHR, with a mean stimulation index > 7. Their release of IL-2, IL-4, and IL-10 after stimulation with soluble anti-CD3 and phorbol ester was indistinguishable from the other clones from this thyroid. However, they produced relatively little IFN gamma (median IL-4/IFN gamma ratio of 0.80) compared with the other clones (median IL-4/IFN gamma ratio 0.06). Thus, this new potent method of antigen presentation, using autoantigen-transfected EBVL, has permitted the first unequivocal identification of TSHR T cells in GD thyroid, with distinct Th0/Th2 characteristics, unlike previously cloned TPO-responsive cells which have Th1 characteristics.
Antigen Presentation↗
T cell repertoire usage in humans, from newborns to centenarians.
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Aged↗
Human CD4-CD8- alpha beta + T-cell receptor T cells recognize different mycobacteria strains in the context of CD1b.
Double-negative alpha beta+ T-cell receptor (TCR) human T cells have been reported to recognize antigen in the context of the HLA class I-like (Ib) CD1 complex. In particular, the CD1b molecule has been shown to act as the element of genetic restriction for antigens derived from Mycobacterium tuberculosis. The stenotopic nature of these major histocompatibility complex (MHC) class Ib molecules raised the question of whether the antigenic moiety recognized by CD4-CD8- alpha beta+ TCR T cells was shared by different mycobacteria. We demonstrate here that a CD4-CD8- alpha beta+ TCR T-cell line and three clones raised against M. tuberculosis proliferated following stimulation with soluble extracts from organisms of the M. tuberculosis complex, M. leprae and 10 out of 16 tested isolates of M. avium complex; however, four species of weakly or non-pathogenic mycobacteria were not stimulatory. Furthermore, the M. tuberculosis soluble extract (MTSE)-derived, recognized antigenic moiety proved to be proteinase K resistant and to have a molecular weight greater than 5000 MW, thus it differed from the reported antigenic moiety, recognized by CD4-CD8- gamma delta+ TCR cells. Our results suggest that a common antigenic moiety, presented by CD1b molecules to CD4-CD8- alpha beta+ TCR T cells, is shared by many mycobacterial species. Therefore they raise interest in the question of whether CD4-CD8- alpha beta+ TCR T cells, elicited by M. tuberculosis, may play a role in the natural history of other mycobacterial infections.
Antigens, Bacterial↗
Human alpha beta T-cell receptor CD4-CD8 T-cell clones are predominantly Th0-like.
The cytokine production profile, focusing on interleukin-4 (IL-4), interferon-gamma (IFN-gamma) and interleukin-10 (IL-10) of human CD4+, CD8+ and CD4- CD8- alpha beta T cells cloned from the peripheral blood of healthy individuals was compared. Solid-phase anti-CD3 stimulation of CD4- CD8- alpha beta T cell clones from one individual revealed a significantly increased frequency of IL-4-producing clones (81%), compared to CD4+ T cells (24%) or CD8+ (28%). All five CD4- CD8- alpha beta T-cell clones from two other individuals also produced IL-4. Clones that produced IFN-gamma with undetectable IL-4 production, thus being of the 'classical' Th1 phenotype, were infrequent in CD4- CD8- alpha beta T-cell clones (19%) compared to CD4+ (71%), and CD8+ clones (72%) cloned in OKT3, and CD4+ cells cloned in phytohaemaglutinin A (77%). Unlike previously reported studies with gamma delta cells, the alpha beta CD4- CD8- T cells produced IL-10 at appreciable frequency (38%) in PHA generated clones. The supernatants from anti-CD3 stimulated CD4- CD8- alpha beta T-cell clones contained sufficient IL-4 to activate B cells, enhancing CD23 and surface immunoglobulin M (IgM) expression and co-stimulating B-cell proliferation. These findings suggest that the function of CD4- CD8- alpha beta T cells is distinct from that of most CD4+ or CD8+ T cells.
B-Lymphocytes↗
Immunodominant epitopes of glutamic acid decarboxylase 65 and 67 in insulin-dependent diabetes mellitus.
To find the dominant epitopes of a major autoantigen in insulin-dependent diabetes mellitus (IDDM), glutamic acid decarboxylase (GAD), we studied the reactivity of peripheral blood T lymphocytes with peptides covering both major isoforms. A significant response to GAD 65 or GAD 67 peptides was detected in 13 of 15 IDDM patients and in 9 of 10 normal controls. Controls most frequently recognised the central region of GAD 65 (residues 161-243). IDDM patients preferentially recognised residues 473-555 (p < 0.03). T-cell responses to GAD 67 peptides were similar in IDDM patients and controls. T lymphocytes from IDDM patients recognise a distinct dominant epitope of GAD, which may be an important target for the disease process.
Adolescent↗
Transfection of thyroid autoantigens into EBV-transformed B cell lines. Recognition by Graves' disease thyroid T cells.
Autologous EBV-transformed B cell lines (EBVL) from a patient with Graves' disease (GD) were transfected with the expression vector pREP4 encoding thyroid peroxidase (TPO), the thyroid-stimulating hormone receptor (TSHR), or chloramphenicol acetyl transferase. The relevant proteins were expressed. The capacity of EBVL to present transfected Ag was assessed by using EBVL transfected with TPO (EBVL-TPO) to stimulate TPO-specific T cells cloned from autologous thyroid tissue; stimulation indices greater than 100 were consistently obtained, even at low APC:T cell ratios. EBVL-TPO and EBVL transfected with TSHR (EBVL-TSHR) were used to characterize the Ag specificity of five thyroid epithelial cell-specific T cell clones. Despite previous unresponsiveness to recombinant TPO or purified thyroglobulin, four of these clones responded vigorously to EBVL-TPO but none responded to EBVL-TSHR. Experiments were then performed to determine whether transfected EBVL were presenting endogenously synthesized TPO, or were taking up and reprocessing exogenous Ag shed from the surface of adjacent cells. The stimulatory capacity of EBVL-TPO was not transferable to untransfected EBVL by cell-free culture supernatant and to only a minor degree by EBVL-TPO-derived cell membrane fragments, indicating that the majority of the TPO presented by the transfected EBVL is endogenously synthesized. The clear and reproducible proliferative responses obtained illustrate the usefulness and potency of this method of Ag presentation to T cells. Because this system obviates the need for large numbers of autologous APC or purified recombinant Ag, it will be useful for obtaining autoantigen-reactive T cells and to help define the autoantigenic epitopes recognized.
Adult↗
Exploring myelin basic protein for HLA class I-binding sequences.
In view of the increasing evidence of the involvement of CD8+ T cells in the pathogenesis of multiple sclerosis (MS), we have scanned the sequence of the myelin basic protein (MBP), using 162 overlapping nonapeptides, for HLA-class I binding sites. Peptide binding was measured using the recently reported HLA class I alpha-chain-refolding assay, and the following HLA allelic products were analyzed: HLA-A2 (*0201, *0204), B27 (*2705), B35, B51 and B62. A considerable number of binding peptides were distinguished for each of the allelic products tested. In addition, three interesting points emerged. The first was the identification of several binding peptides which did not contain the known anchor motifs. The second was the evidence that several peptides showed a promiscuous binding profile, being able to bind to different HLA class I molecules that were either allelic or non allelic. The third was that in several cases two consecutive peptides could bind to the same HLA molecule.
Amino Acid Sequence↗
The peptide binding specificity of HLA-B27 subtypes.
Five HLA-B27 subtypes, B*2701, B*2703, B*2704, B*2705, and B*2706, were tested for direct binding with twenty-six synthetic nonapeptides carrying the primary anchor residue motifs (combination of amino residues at positions 2 and 9) relevant to B*2705. The peptide sequences were derived from human HSP89 alpha, P53 and MBP. The alpha chains were immunospecifically isolated from LH (B*2701), CH (B*2703), WE1 (B*2704), BTB (B*2705), and LIE (B*2706) cells and their peptide binding was measured by the HLA class I alpha chain refolding assay. The data obtained indicated that the B27 subtypes tested can bind a common set of peptides carrying several different anchor residue motifs. The motifs, R-K and R-R, reported for B*2705 and a new motif H-R were accepted by B*2703, B*2704, and B*2706, but not by B*2701. However, other motifs, including known B*2702 and/or B*2705 motifs, R-H, R-L, R-A, and R-F, and a new motif found here, R-G, were apparently accepted by all B27 subtypes tested. The observed cross-peptide binding in the B27 subgroup is compatible with the so-called arthritogenic peptide hypothesis in the pathogenesis of ankylosing spondylitis.
Amino Acid Sequence↗
Chronic exposure to tumor necrosis factor (TNF) in vitro impairs the activation of T cells through the T cell receptor/CD3 complex; reversal in vivo by anti-TNF antibodies in patients with rheumatoid arthritis.
Experiments were designed to test the hypothesis that chronic exposure to tumor necrosis factor alpha (TNF) alters the function of activated T lymphocytes. Pretreatment of tetanus toxoid-specific T cell clones with TNF for up to 16 d impaired rechallenge proliferative responses to antigen in a dose- and time-dependent fashion. IL-2 and PHA responses were preserved. Prolonged treatment with TNF impaired production of IL-2, IL-10, IFN gamma, TNF, and lymphotoxin (LT) following stimulation with immobilized OKT3, and resulted in suboptimal expression of the IL-2R alpha chain (Tac) but not CD3, CD4, or HLA-DR antigens, when compared to untreated control cells. By contrast, pretreatment of T cells for prolonged periods in vitro with neutralizing anti-TNF monoclonal antibodies (mAb) enhanced proliferative responses, increased lymphokine production, and upregulated Tac expression following stimulation with OKT3. To determine whether TNF exerts immunosuppressive effects on T cells in vivo, we studied cell-mediated immunity in patients with active rheumatoid arthritis (RA), before and after treatment with a chimeric anti-TNF mAb. Treatment with anti-TNF restored the diminished proliferative responses of PBMC to mitogens and recall antigens towards normal in all patients tested. These data demonstrate that persistent expression of TNF in vitro and in vivo impairs cell-mediated immune responses.
Antibodies, Monoclonal↗
Selective activation of blood T cells in children with newly diagnosed insulin dependent diabetes mellitus.
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Adolescent↗
IL-10 enhances expression of the IL-2 receptor alpha chain on T cells.
Interleukin-10 (IL-10) has various immunomodulatory actions depending on the target cell type. Some of these effects have been shown to be owing to its ability to down-regulate surface expression of markers, for example HLA-DR on macrophages and CD25 (IL-2 receptor alpha chain) on B cells. In this report we show that preincubation of IL-10 for 24 hr up-regulates expression of the activation marker CD25, but not HLA-DR on cloned T cells of various phenotypes such as CD4+, CD8+, CD4- CD8- alpha beta and gamma delta T-cell receptor (TCR)-expressing cells. This up-regulation of CD25 was accompanied by an increase in the T cells IL-2-dependent proliferative response in 63% of the CD4+ clones and 100% of the CD8+, CD4-, CD8- alpha beta and gamma delta TCR+ clones analysed. IL-10 was also shown to be at least partly responsible for the up-regulation of CD25 on mitogen-activated peripheral blood mononuclear cells, suggesting that IL-10 has this CD25 modulatory effect within a more physiological environment. Our data suggest that IL-10 can have a multitude of effects on human T cells, and should not be considered exclusively as an immunoinhibitory cytokine.
Adjuvants, Immunologic↗
T cells involved in human autoimmune disease are resistant to tolerance induction.
Deletion of potentially self-reactive T-cell clones during intrathymic development provides an important mechanism of preventing autoreactivity. However, some potentially damaging cells may escape this process. Recent evidence suggests that these cells may be rendered 'anergic', that is, nonresponsive to Ag, in the absence of cell death. Such a mechanism may be particularly important in maintaining tolerance to organ-specific self Ag that are not expressed in the thymus. If so, the emergence of T cells resistant to anergy induction might be expected to result in autoimmune disease. It has previously been shown that anergy can be induced in human T cells in vitro by exposure to specific target peptide or bacterial enterotoxins in the absence of Ag-presenting cells. We have recently defined the antigenic specificity of multiple T-cell clones present at the site of a human organ-specific autoimmune disease, Graves' thyroiditis (Graves disease). In the current work, thyroid-derived T cells recognizing residues 535-551 of the thyroid tissue-specific enzyme, TPO3 have been used to examine whether cells actively involved in the autoimmune process are resistant to anergy induction, as defined by anergy induction with in vitro systems. Two systems were used. First, supraimmunogenic concentrations of peptide 535-551 (up to 1 mg/ml) failed to significantly anergize these T cells in the absence of APC. In addition, the bacterial enterotoxin SED that could stimulate these T cells in the presence of APC, failed to induce anergy when APC were not present. T cells from the peripheral blood of the same individual, in contrast, were anergizable with bacterial enterotoxins, using the same protocol. These results suggest that thyroid-infiltrating autoantigen-reactive T cells are refractory to induction of anergy, and the possibility is raised that this deficiency may be of importance in the development of autoimmunity.
Amino Acid Sequence↗