摘要:

AbstractWe have previously reported that human T cell receptors (TcR) selected in the class II‐restricted (HLA‐DRB1*1302) response to a tetanus toxin peptide (tt830‐843) frequently used the Vβ2 germ‐line segment which paired with several Vα segments and that the putative CDR3 of both α and β chains showed remarkable heterogeneity. To analyze the structural basis for recognition of the tt830‐843/DR complex, five of these TcR were reconstituted into a murine T cell hybridoma, 58 α−β−, by expressing the human α and β variable regions joined to the mouse α and β constant regions, respectively. The chimeric TcR, expressing the same Vβ germ‐line segment (Vβ2), two expressing Vα21.1, twoVα17.1 and one Vα8.1 were shown to have the expected antigen specificity and DR restriction.Two lines of evidence suggested that the putative CDR3, although not conserved in these TcR, played a key role in recognition. First, two TcR with identical V germ‐line segments but distinct CDR3 showed large differences in their capacity to react with the ligand. Second, interchanging the α and β chains from tt830‐843/DR1302‐specific TcR which differed in their CDR3 sequences invariably led to loss of recognition. We also asked whether germ‐line Vα17.1 could functionally replace Vα21.1, as they appear to be related in their primary sequence. However, as in the case of CDR3 exchanges, Vα replacement abrogated TcR reactivity. Taken together, these data underline the fine interdependence of the structural components of the TcR binding site in defining a given specificity.Four of the TcR studied displaying promiscuous recognition were also tested against different DR alleles and site‐directed mutants. The results of these experiments suggested that, in spite of their structural heterogeneity, anti‐tt830‐843 TcR may have a similar orientation with respect to the peptide/DR complex.The reconstitution system described herein should represent a valuable t

ISSN:0014-2980    

DOI:10.1002/eji.1830231203

出版商:WILEY‐VCH Verlag GmbH    

年代:1993

数据来源: WILEY

摘要:

AbstractInterleukin‐4 (IL‐4) has important regulatory functions in the immune system, particularly in the generation of immunoglobulin E, the principal mediator of allergic responses. The molecular basis of IL‐4 action has remained elusive so far. Here we report on a novel human transcription factor, termed nuclear factor IL‐4 (NF‐IL4), which is posttranslationally activated by IL‐4 in lymphoid and monocytic cells. Homologous binding sequences for NF‐IL4 were identified in the promoter regions of the IL‐4 controlled CD23b and Iϵ genes. We defined a palindromic 9‐bp consensus sequence (5′‐TYCYRRGAA‐3′) as IL‐4‐responsive element (IL‐4RE). Point mutation analysis of the CD23b promoter showed that binding of NF‐IL4 to the IL‐4RE is essential for the initiation of gene transcription in response to IL‐4. NF‐IL4 was not activated by Ca2+ionophore, phorbol ester and cAMP either alone or in combination suggesting a non classi

ISSN:0014-2980    

DOI:10.1002/eji.1830231204

出版商:WILEY‐VCH Verlag GmbH    

年代:1993

数据来源: WILEY

摘要:

AbstractStimulation of human peripheral blood lymphocytes via T cell receptor/CD3 complex resulted in a bimodal activation of protein kinase(s) C (PKC). Within 10 min of stimulation PKC‐α was translocated to, and thus activated in, the plasma membranes of human lymphocytes, followed by a fast dissociation of this isotype from the plasma membrane. This short term activation and translocation PKC‐α proved to be cyclosporin A (CsA) insensitive. After 90 min of stimulation PKC‐β was translocated to and remained bound to the plasma membranes for up to 4 h. Preincubation of human lymphocytes with 200 ng/ml CsA specifically and completely abolished the sustained activation of PKC‐β. Neither the phorbol ester‐induced direct activation of PKC nor the specific activity of the plasma membrane‐bound enzyme was influenced by CsA, suggesting that a signal transduction pathway leading to sustained activation of PKC‐β was influenced by the immunosuppressive agent. In fact, CsA inhibited, in a concentration‐dependent manner, the activation of lysophosphatid acyltransferase‐catalyzed elevated incorporation of cis‐polyunsaturated fatty acids into plasma membrane phospholipids. While interleukin‐2 (IL‐2) synthesis and cellular proliferation were completely inhibited by 200 ng/ml CsA in BMA 030‐ or BMA 031‐stimulated cells, expression of high‐affinity IL‐2 receptors was not influenced by the immunosuppressive drug. These results suggest that synthesis and expression of high‐affinity IL‐2 receptors might be regulated by a signal‐transducing pathway involving activation and translocation of PKC‐α. Lysophosphatid acyltransferase‐catalyzed incorporation of cis‐polyunsaturated fatty acids might represent another mechanism of signal transduction implicated in the activation and translocation of PKC‐β, which is specifically inhibited by CsA. Neutralization of PKC‐β by introducing anti‐PKC‐β antibodies prevented IL‐2 synthesis and proliferation in stimulated human lymphocytes. The results suggest a possible link between activation of PKC‐β and regulation of IL‐2 synthesis in activated human lymphocytes. Thus, inhibition of the activation and translocation of PKC‐β by CsA

ISSN:0014-2980    

DOI:10.1002/eji.1830231205

出版商:WILEY‐VCH Verlag GmbH    

年代:1993

数据来源: WILEY

摘要:

AbstractThe protein products of the Tap (Transporter associated with antigen processing) 1 and 2 genes are presumed to deliver peptides across the endoplasmic reticulum (ER) for assembly with major histocompatibility complex (MHC) class I molecules. The antigen processing‐defective cell line RMA‐S (H‐2b) has a premature stop in the Tap 2 gene and probably therefore fails to deliver peptides into the ER, which leads to a low level of cell surface MHC class I molecules. Transfection of a Tap 2 gene restores to RMA‐S both MHC class I molecule expression and the ability to present influenza viral antigens. We investigated the ability of RMA‐S cells transfected with a Tap 2 gene to process and present alloantigens, Sendai and Rauscher viral antigens to allogeneic and virus‐specific cytotoxic T lymphocytes. We found that allogeneic peptides as well as Rauscher and Sendai viral peptides can be processed and presented by RMA‐S but at reduced levels. Transfection of a Tap 2 gene of mouse (BALB/c, H‐2d) or rat origin into RMA‐S increased the presentation of Sendai viral antigens and partially restored the presentation of allogeneic antigens. The already low level of Rauscher viral peptides presented by RMA‐S is not elevated by transfection of either Tap 2 gene into RMA‐S. This indicates a differential effect of transfection of a Tap 2 gene of rat or allogeneic mouse origin into RMA‐S on v

ISSN:0014-2980    

DOI:10.1002/eji.1830231206

出版商:WILEY‐VCH Verlag GmbH    

年代:1993

数据来源: WILEY

摘要:

AbstractPolymorphism of the major histocompatibility complex (MHC) influences susceptibility to experimental autoimmune encephalomyelitis (EAE) induced by myelin basic protein (MBP) in rats. Current concepts relate such influences to the capacity of class II molecules to present relevant peptides to autoreactive T cells. We have here analyzed the MHC influence on the immune response and the development of EAE after immunization with the immunodominant peptide MBP‐63–88. Analysis of MHC‐congenic LEWIS strains showed that RT1a, RT1cand RT1lhaplotypes are permissive for disease induction, whereas RT1dand RT1uare resistant. All EAE responding strains showed peptide‐specific proliferation and interferon (IFN)‐γ secretion, but no early significant tendency to express interleukin (IL‐4) or transforming growth factor (TGF)‐β mRNA in lymphocytes in response to the MBP 63–88, 7 days post immunization (p.i.). Later, 14 days p.i., peptide‐specific induction of IL‐4 and TGF‐β occurred in RT1lrats. Among the EAE non‐responders strains, only the RT1urats showed an immune response to MBP 63‐88. This response, however, was qualitatively different from the immune response in the EAE‐susceptible strains. Thus, there was no proliferation and only moderate IFN‐γ production in response to peptide, but in contrast, a significant and early peptide‐induced IL‐4 and TGF‐β response was observed. The data suggest that the MHC‐associated susceptibility to EAE is partly related to the ability to mount a TH1‐like immune response while the MHC‐associated EAE resistance may either be related to MBP peptide non‐responsiveness or to peptide recognition and induction of a qualitatively diffe

ISSN:0014-2980    

DOI:10.1002/eji.1830231207

出版商:WILEY‐VCH Verlag GmbH    

年代:1993

数据来源: WILEY

摘要:

AbstractMercuric chloride (HgCl2) injections protect (Lewis x Brown‐Norway) F1 (F1) rats against experimental autoimmune uveoretinitis (EAU) induced by immunization with the retinal S antigen (S‐Ag); in contrast HgCl2‐injected F1 rats develop EAU following transfer of lymph node (LN) cells from rats immunized with S‐Ag alone. In the present study we demonstrate that the ability of LN cells from rats protected against EAU to transfer the disease into naive F1 rats was considerably reduced. These LN cells neither produced interleukin (IL)‐2 nor (interferon (IFN)‐γ but exhibited mRNA for IL‐4. In contrast, LN cells from diseased rats easily transferred EAU into naive F1 rats, produced significant IL‐2 and IFN‐γ levels but barely exhibited mRNA for IL‐4. Furthermore protected rats predominantly produced IgG2b anti‐S‐Ag antibodies, while diseased rats produced IgG2b anti‐S‐Ag antibodies and the increase in expression of MHC class II molecules on B cells was higher in protected rats than in diseased rats. These data suggest that (1) to exert a protective effect, HgCl2must act at an early stage of differentiation of precursors of S‐Ag specific T cells, and (2) this effect is related to the preferential activation of TH2 cells to the detriment of uveitogenic TH1 cells. Finally, these results indicate that activation of TH2 cells protect from a T

ISSN:0014-2980    

DOI:10.1002/eji.1830231208

出版商:WILEY‐VCH Verlag GmbH    

年代:1993

数据来源: WILEY

摘要:

AbstractThree‐color immunofluorescence and flow cytometric analysis showed that the vast majority of normal human T‐lamina propria lymphocytes (LPL) expressed high levels of the early activation antigen CD69, together with CD45R0, irrespective of their CD4, CD8 or γ/δ‐TcR phenotype, indicating that they are continuously stimulatedin vivo.Importantly, measurement of cytoplasmic [Ca2+]ishowed that T‐LPL had significantly higher basal [Ca2+]ilevels, compared to autologous peripheral blood lymphocytes (PBL). Both cytoplasmic [Ca2+]ielevation and inositol (1, 4, 5) trisphosphate generation following CD3 cross‐linking by monoclonal antibodiesin vitrowere essentially abolished in T‐LPL, as compared to autologous T‐PBL. Moreover, freshly isolated LPL could be induced to proliferate by CD2‐ or CD28‐mediated signals, but not by CD3‐mediated signals. Surprisingly however, impairment in TcR/CD3‐mediated early signaling and proliferation in T‐LPL could be completely reversed by 24 h incubation of the cells at 37 °C in culture medium, a condition which allowed basal intracellular [Ca2+]ito return to levels comparable to peripheral T cells.Our data suggest that selective hyporesponsiveness to TcR/CD3‐mediated signaling may represent a transient event during continuousin vivoactiv

ISSN:0014-2980    

DOI:10.1002/eji.1830231209

出版商:WILEY‐VCH Verlag GmbH    

年代:1993

数据来源: WILEY

摘要:

AbstractX‐linked agammaglobulinemia (XLA) is an immunodeficiency disease in man, resulting from an arrest in early B cell differentiation. The gene defective in XLA has recently been identified and encodes a cytoplasmic protein tyrosine kinase, named Bruton's tyrosine kinase(btk), essential for cell differentiation and proliferation at the transition from pre‐B to later B cell stages. In this study we investigatedbtkexpression by Northern blotting experiments in a series of human (precursor‐) B cell lines, acute lymphoblastic leukemias and plasmacytomas,btkwas found to be already expressed in very early stages of B cell differentiation, even prior to immunoglobulin (Ig) heavy (H) or light (L) chain gene rearrangements. Transcripts were also detected at the pre‐B cell stage and in mature B cells, irrespective of the Ig H chain class expressed. Approximately at the transition from mature B cells to plasma cells, expression of thebtkgene is down‐regulated. In addition, thebtkgene was found to be expressed in myeloid cell lines and acute myeloid leukemias.btkexpression in myeloid cells is probably not a prerequisite for myeloid differentiation, since myeloid cells in XL A patients seem not to be affected. Nobtkexpression was found in T‐lineage cells. Thebtkexpression profile,i.e.from early precursor‐B cell stages preceding Ig rearrangement up to mature B cells, supports the hypothesis that the XLA defect resides in a critical step of B cell development which is independent of the Ig gene recombinat

ISSN:0014-2980    

DOI:10.1002/eji.1830231210

出版商:WILEY‐VCH Verlag GmbH    

年代:1993

数据来源: WILEY

摘要:

AbstractTargeted disruption of β2‐microgobulin gene results in deficient major histocompatibility complex class I expression and failure to develop CD4−8+T cells. Despite this, β2M−/−mice reject skin grafts and cope with most viral infections tested. We asked whether CD4+8−cytotoxic T cells could play a role in compensating for the defect in CD4−8+cytotoxic T cell function. We found that the cytotoxic activity against class II+targets is significantly higher among CD4+8−T cells of β2M−/−than among those of β2M+/+mice. In the limiting dilution experiment, we showed that the precursor frequency for the cytotoxic, CD4+8−, class II‐specific T cells is at least fivefold higher in β2M−/−than in β2M+/+mice. These results suggest that CD4+8−cytotoxic T cells could play a major role in carrying out

ISSN:0014-2980    

DOI:10.1002/eji.1830231211

出版商:WILEY‐VCH Verlag GmbH    

年代:1993

数据来源: WILEY

摘要:

AbstractIt has been extensively shown that when T cells are co‐stimulated with B7‐CD28 interaction, a strong proliferative as well as cytolytic T cell response can be induced. In contrast, there exists only indirect evidence that the B7‐CD28 interaction is of importance for the induction of T cell helper functions in B cell responses. Here we have used mouse fibroblasts transfected with the human Fcγ receptor type II and human B7 to address this issue. We found that T cells, when activated through the T cell receptor (TcR)/CD3 complex with monoclonal antibodies and co‐stimulated by B7‐CD28 interaction, can provide efficient help for the induction of both IgM and IgG production by resting B cells. This helper activity is, at least in part, mediated by the interaction between the CD40 ligand on the T cells and CD40 on the B cells. We also demonstrate that more than one signal to the T cell is required for the induction of the CD40 ligand, one being delivered through the TcR/CD3 complex and the second by ligation of CD28 with the B7 molecule. In addition to the induction of cognate T helper function, we provide evidence that co‐stimulation of T cells with B7‐CD28 interaction can result in the secretion of both Th1‐ and Th2

ISSN:0014-2980    

DOI:10.1002/eji.1830231212

出版商:WILEY‐VCH Verlag GmbH    

年代:1993

数据来源: WILEY