bluuebunny/crossref_metadata_2025 huggingface dataset:
Mechanisms of peripheral T cell deletion: anergized T cells are Fas resistant but undergo proliferation‐associated apoptosis
Authors: Miethke Thomas , Vabulas Ramunas , Bittlingmaier Renate , Heeg Klaus , Wagner Hermann
Published: March 2007

The complementary receptor pair Fas ligand: Fas controls apoptosis during activation‐induced cell death (AICD) of peripheral T cells sensitized for the Fas signal pathway by interleukin‐2 (IL‐2). In the present study, we used the bacterial superantigen staphylococcal enterotoxin B (SEB) to anergize ligand‐reactive peripheral T cells in wild‐type and Fas‐defective lpr mice. In a second step, we investigated whether apoptosis in anergized and thus operationally IL‐2‐defective peripheral T cells is triggered via the Fas signal pathway. We report here that SEB‐driven anergy induction and deletion of anergized peripheral Vβ8+ T cells is similar in wild‐type and healthy C3H/lpr mice. In monitoring SEB‐driven Vβ8+ T cell apoptosis in situ, we observe in both wild‐type and lpr mice an intimate association between proliferation and apoptosis of anergized Vβ8+ T cells. We further show that Vβ8+ T cells activated in vitro from wild‐type mice express a Fas‐sensitive phenotype determined by Fas cross‐linking which causes apoptosis. In contrast, Vβ8+ T cells anergized in vivo from wild‐type mice are Fas resistant. As expected, T cells from lpr mice activated in vitro or anergized in vivo are Fas resistant. Taken together, these data indicate that both in wild‐type and Fas‐defective C3H/lpr mice, anergized T cells become deleted via a Fas‐independent, proliferation‐associated apoptosis signal pathway.
Relationship between Th1/Th2 cytokine patterns and the arthritogenic response in collagen‐induced arthritis
Authors: Mauri Claudia , Williams Richard O. , Walmsley Marita , Feldmann Marc
Published: March 2007

It is hypothesized that the balance of cytokines produced by Th1/Th2 subsets of T helper cells plays an important role in the development of autoimmune diseases. Murine collagen‐induced arthritis (CIA) is an example of an autoimmune disease in which immunization with cartilage‐derived type II collagen induces, firstly, a T cell response to type II collagen and, secondly, the manifestation of a destructive inflammatory response in affected joints. We have investigated the role of Th1/Th2 responses in the development of CIA by monitoring levels of interferon (IFN)‐γ (a Th1 cytokine) and interleukin (IL)‐4 and IL‐10 (Th2 cytokines), and IL‐1β and tumor necrosis factor (TNF) (pro‐inflammatory cytokines) produced by cultured draining lymph node cells (LNC) from collagen‐immunized DBA/1 mice during the induction phase of arthritis and throughout the time of clinical manifestation and subsequent remission of the disease. Although a transient increase in IL‐10 was detected 3 days after immunization, Th2 cytokine production was found to be almost completely suppressed 6 days after immunization. In contrast, IFN‐γ was detected in LNC cultures as early as 6 days after immunization and the addition of type II collagen to the culture medium resulted in an approximately 10‐fold increase in IFN‐γ production, indicating that a predominantly Th1 response had become established by this time. IFN‐γ production by LNC was found to be further increased at the time of clinical manifestation of arthritis and could be up‐regulated by co‐culture with type II collagen. IL‐10 was not detected in LNC cultures at the onset of arthritis and IL‐4, although present, was found to be markedly suppressed in LNC cultures containing type II collagen. These findings indicate that Th1 responses are predominant at the time of onset of arthritis and that the activation of collagen‐specific Th1 cells may result in suppression of Th2 activity. IFN‐γ production declined progressively during the progression and subsequent remission of arthritis whereas levels of IL‐10 increased and low, though persistent, levels of IL‐4 were detected throughout this period. High levels of IL‐1β and TNF‐α production were detected at the onset of the disease. The role of Th1 responses in the development of CIA was further emphasized by the observation that immunization of mice with type II collagen in incomplete Freund's adjuvant, which normally fails to induce arthritis, resulted in a predominantly Th2 cytokine profile.
Accessory signaling by CD40 for T cell activation: induction of Th1 and Th2 cytokines and synergy with interleukin‐12 for interferon‐γ production
Authors: Peng Xiaohui , Kasran Ahmad , Warmerdam Petra A. M. , De Boer Mark , Ceuppens Jan L.
Published: March 2007

The interaction of CD40 ligand (CD40L) on activated T cells with CD40 on B cells, monocytes and dendritic cells is essential for humoral immunity and for up‐regulation of antigen‐presenting cell (APC) functions, as a result of signaling through CD40. There are also some indications that after interaction with CD40, CD40L can directly signal T cells. In this study we demonstrate that upon stimulation of human peripheral blood T cells through the T cell receptor (TCR)/CD3 complex, CD40/CD40L interaction strongly enhances the production of Th1 cytokines such as interleukin (IL)‐2 and interferon (IFN)‐γ and Th2 cytokines such as IL‐4, IL‐5 and IL‐10 by a direct effect on T cells. Furthermore, CD40/CD40L interaction synergizes with IL‐12 in selectively enhancing IFN‐γ production by purified anti‐CD3‐stimulated T cells. These effects were observed at both the protein and the mRNA level. Both CD4+ and CD8+ T cells were able to produce IFN‐γ in the presence of helper signals from IL‐12 and CD40, although CD8+ T cells were less active. Since CD40/CD40L interaction also up‐regulates IL‐12 production and B7 expression by APC, our results suggest that CD40/CD40L interaction is bidirectional, and promotes activation of both APC and T cells.
Cell type‐specific processing of the I‐Ed‐restricted hen egg lysozyme determinant 107–116
Authors: Robadey Chantal , Wallny Hans‐Joachim , Demotz Stéphane
Published: March 2007

Class II major histocompatibility complex heterodimers present to T cells determinants as sets of antigen fragments with ragged N and C termini. It is not yet elucidated whether different types of antigen‐presenting cells generate identical sets of peptides containing the same determinant. Taking advantage of recombinant I‐Ed molecules produced by insect cells as empty heterodimers, a sensitive T cell stimulation assay was developed to analyze naturally processed hen egg lysozyme (HEL) peptides. I‐Ed preparations were isolated from antigen‐presenting cells cultured with HEL. Following acid treatment, peptides eluted from I‐Ed were chromatographed and the fractions incubated at acidic pH with purified recombinant I‐Ed molecules, conditions which favor peptide binding. The stimulatory capacity of the reconstituted peptide‐I‐Ed complexes adsorbed on the well surface of cell culture plates was then evaluated by measuring interleukin‐2 secreted by an HEL 107–116‐specific, I‐Ed‐restricted T cell hybridoma. We found that the B lymphoma A20 and an I‐Ed‐transfected fibroblast cell line generated distinct sets of peptides containing the HEL sequence 107–116. Our results suggest the possibility that presentation of one determinant by different types of antigen‐presenting cells stimulates populations of T cells with distinct fine antigen specificities.
Induction of class II major histocompatibility complex blockade as well as T cell tolerance by peptides administered in soluble form
Authors: Nihira Shin‐Ichi , Falcioni Fiorenza , Juretic Antonio , Bolin David , Nagy Zoltan A.
Published: March 2007

Peptides binding to a particular class II major histocompatibility complex (MHC) molecule can inhibit the activation of T cells by other peptides binding to the same molecule, a phenomenon termed class II MHC blockade. All class II‐binding peptides exert MHC blockade in vivo in depot form with adjuvant, and some also retain their blocking properties in soluble form. We demonstrate here that soluble peptides, when used at doses causing short‐term MHC blockade, can also induce long‐term antigen‐specific T cell tolerance to themselves. The tolerogenicity of soluble peptides correlates with their antigenicity in adjuvant, but it is not necessarily related to their capacity to act as class II blockers in vivo. The tolerant state is manifested in a decreased production of both T helper cell 1 (Th1)‐type and Th2‐type lymphokines, and it cannot be reversed by interleukin‐2. Once T cells are primed with a peptide in complete Freund's adjuvant, they are resistant to tolerization with the same peptide applied in soluble form. Tolerance induction is partially impaired in B cell‐deficient μMT−/− mice, suggesting a role for B cell antigen presentation in this process. The results suggest that the potential immunogenicity of class II MHC blockers could be circumvented by choosing a tolerogenic mode of application.
A point mutation in the human transporter associated with antigen processing (TAP2) alters the peptide transport specificity
Authors: Armandola Elena A. , Momburg Frank , Nijenhuis Marga , Bulbuc Nadja , Früh Klaus , Hämmerling Günter J.
Published: March 2007

The heterodimeric transporter associated with antigen processing (TAP1/TAP2) translocates peptides from the cytosol into the endoplasmic reticulum where loading of major histocompatibility complex class I molecules takes place. TAP transporters from different species are known to exhibit distinct transport specificities with regard to the C‐terminal amino acid (aa) of peptides. Thus, human TAP (hTAP), and rat TAP (rTAP) containing the rTAP2a allele are rather promiscuous, whereas mouse TAP (mTAP), and rTAP containing the rTAP2u allele are restrictive and select against peptides with C‐terminal small polar/hydrophobic or positively charged aa. The structural basis for this selectivity is not clear. To assess the relative contribution of the TAP1 and TAP2 subunits to transport specificity, we have constructed and analyzed interspecies TAP hybrids and point mutants of hTAP2 expressed in Sf9 insect cells and in TAP‐deficient T2 cells. Transport assays with 20 C‐terminal variants of the peptide RYWA‐NATRSX showed that: first, transport specificity with regard to C‐terminal aa is mainly influenced by TAP2, but TAP1 can also contribute. Second, the selective transport of peptides with C‐terminal positively charged aa is critically controlled by the amino‐terminal region (1–361) on the TAP2 chain, while transport of peptides with C‐terminal small polar/hydrophobic aa is determined by residues located within as well as outside the region 1–361. Third, a single point mutation in hTAP2 (374A → D) resulted in a drastic alteration of the transport pattern. These results indicate that both TAP1 and TAP2 contribute to efficient peptide transport and that single point mutations in hTAP2 are able to alter the peptide transport specificity. This opens the possibility that naturally occurring mutations in one of the hTAP subunits may alter epitope selection in vivo.
Redirecting the complete T cell receptor/CD3 signaling machinery towards native antigen via modified T cell receptor
Authors: Brocker Thomas , Riedinger Mireille , Karjalainen Klaus
Published: March 2007

We show that a chimeric T cell receptor (TCR) β chain consisting of a single‐chain Fv portion derived from a monoclonal antibody and the full TCR β chain is able to assemble functionally with endogenous TCR/CD3 components and transfer the antibody specificity as well as the TCR specificity into TCRβ− as well as into TCRβ+ T cells. This allows the incorporation new non‐major histocompatibility complex‐restricted ligand specificities into the intact TCR/CD3 complex which can exploit the full range of biological activities of the endogenous TCR signaling machinery. This approach can provide wider opportunities to redirect T cells to virus or tumor antigen‐bearing cells.
Evidence for intact CD28 signaling in T cell hyporesponsiveness induced by the HIV‐1 nef gene
Authors: Collette Yves , Mawas Claude , Olive Daniel
Published: March 2007

Infection by human immunodeficiency virus (HIV)‐1 is associated with quantitative and qualitative T cell alterations that severely impair the host's immune defense system. The molecular basis for this immunosuppression remains unclear. Peripheral blood mononuclear cells (PBMC) isolated from patients show markedly decreased interleukin (IL)‐2 secretion but unaffected or even increased T helper (Th)2 cytokine production. T cell functional defects were recently reported to correlate more with T cell receptor (TcR) signaling, whereas signals provided by ligation of co‐receptors CD27 and CD28 appeared to be preserved. Among the various mechanisms proposed to be involved in HIV‐1‐induced T cell dysfunction, we and others have reported that the nef gene product exhibited significant immunosuppressive activity. By using an inducible stably integrated nef gene, we demonstrated that Nef specifically down‐regulated IL‐2 and interferon (IFN)‐γ produced upon TcR triggering. Here, using the same experimental system, we extended our initial observations to additional mitogenic signals, and investigated the co‐stimulatory function of CD28. Nef down‐regulated IL‐2, but not IL‐4 produced upon induction by combinations of mitogens that mimicked TcR signals together with CD28 mAb or CD28's natural ligand (CD80 and CD86). However, the co‐signals provided by CD28 to up‐regulate IL‐2 induction were unaffected by Nef, since IL‐2 produced by nef‐transfected cells was proportionally enhanced to the same extent as that of control cells, either upon stimulation by the CD28 mAb or CD80 and CD86. In addition, phosphatidylinositol‐3 kinase recruitement induced upon CD28 triggering was also found to be unaltered by nef expression. Together with the observation that similar levels of the Nef protein were detected in nef‐transfected cells and upon infection of PBMC, these data suggest a selective immunosuppression induced by nef in human T cells by altering TcR signaling without detectable impact on CD28 co‐receptor function. These data agree with the T cell defects observed in PBMC isolated from HIV‐infected individuals.
Molecular Ig gene analysis reveals that monocytoid B cell lymphoma is a malignancy of mature B cells carrying somatically mutated V region genes and suggests that rearrangement of the kappa‐deleting element (resulting in deletion of the Ig kappa enhancers) abolishes somatic hypermutation in the human
Authors: Küppers Ralf , Hajadi Mande , Plank Lukas , Rajewsky Klaus , Hansmann Martin‐Leo
Published: March 2007

Five cases of monocytoid B cell lymphoma (MBCL) were analyzed for somatic mutations in the rearranged V region genes. Somatic mutations were found in four of the five cases, whereas one unusual CD5+ lymphoma harbored unmutated V region genes. Since somatic mutations are introduced into V regiongenes of antigen‐activated B cells in the course of T cell‐dependent immune responses, these results suggest a derivation of the tumor B cells in MBCL from antigen‐experienced mature B cells. An analysis of the ϰ‐deleting element in two of the cases in which mutated VH but unmutated and nonfunctional Vϰ gene rearrangements were found suggests that somatic hypermutation does not take place in human rearranged Vϰ region genes when the Cϰ gene and the ϰ enhancers have been deleted in cis by rearrangement of the ϰ‐deleting element.
Sequential antigen‐specific growth of T cells in the T zones and follicles in response to pigeon cytochrome c
Authors: Gulbranson‐Judge Adam , Maclennan Ian
Published: March 2007

The sites of accumulation and growth of antigen‐specific T cells was assessed during the Vα11/Vβ3 T cell receptor‐restricted response of IEk+ mice to pigeon cytochrome c. In the T zone, the response was early but transient; Vα11/Vβ3+ T cell numbers fell to background levels as germinal centers developed. The follicles were a major site of specific T cell growth, but Vα11/Vβ3+ T cells were not obvious in foci of extrafollicular B cell growth in the red pulp. As germinal centers waned, Vα11/Vβ3+ T cells in the T zones again rose above background levels and some persisted in the follicles. After the initial stage of T cell priming, specific T cell growth seems to occur where specific interaction can occur with B cells that are presenting processed antigen.
Chemically induced sarcomas from nude mice are more immunogenic than similar sarcomas from congenic normal mice
Authors: Svane Inge Marie , Engel Anne‐Marie , Nielsen Mai‐Britt , Ljunggren Hans‐Gustaf , Rygaard Jørgen , Werdelin Ole
Published: March 2007

To detect possible differences in immunogenicity between tumors induced in T cell‐deficient mice and phenotypically normal congenic mice, 16 sarcomas, 8 having developed in nude BALB/c mice and 8 having developed in congenic normal (nu/+) mice, were transplanted to normal BALB/c recipients and the rates of rejection or acceptance were registered. The 16 tumors were chosen randomly from a panel of 39 sarcomas induced with 0.5% or 0.1% 3‐methylcholanthrene and maintained as cell lines in culture. Out of the tumors originating from nude mice, 66% were rejected by the normal BALB/c recipients, while only 30% of the tumors originating from normal mice were rejected. Tumors with short induction times from normal mice were more readily accepted than tumors with long induction times. Tumors originating from nude mice had significantly longer mean latency times after transplantation to both normal and nude recipients than tumors originating from normal mice. Contrary to what has been reported by others, there was no correlation between the rejection rates of the individual tumors and their Kd, Dd or Ld major histocompatibility complex (MHC) class I surface expression as measured by flow cytometric analysis of cultured tumor cells. The Kd, Dd and Ld proteins of the transplanted tumor lines were analyzed by isoelectric focusing for the occurrence of mutations resulting in altered charge of the MHC protein. No such mutations were found, ruling out MHC mutations of that kind as the source of immunogenicity in the cell lines used in these experiments. Our results suggest the existence of a T cell‐mediated selection in the original tumor cell mass of tumors induced in normal mice, adapting the tumor to growth in a host with a functional T cell system, but apparently there is no connection between this loss of immunogenicity and loss of MHC class I expression.
Heterogeneous effects of B7‐1 and B7‐2 in the induction of both protective and therapeutic anti‐tumor immunity against different mouse tumors
Authors: Martin‐Fontecha Alfonso , Cavallo Federica , Bellone Matteo , Heltai Silvia , Iezzi Giandomenica , Tornaghi Paola , Nabavi Nasrin , Forni Guido , Dellabona Paolo , Casorati Giulia
Published: March 2007

Experimental mouse tumors are classified as intrinsically immunogenic when, after a single injection into syngeneic mice as nonreplicating cell vaccines, they elicit a protective immune response against a subsequent lethal challenge. Tumors that do not retain this residual immunogenicity are defined as poorly immunogenic or nonimmunogenic. The expression of the B7‐1 co‐stimulatory molecule on immunogenic tumors can further increase their capacity to induce a T cell‐dependent anti‐tumor immunity, whereas it has limited effects on nonimmunogenic tumors. Recently, B7‐2, a second molecule with an apparently similar co‐stimulatory activity, has been cloned. In this report, we compare the efficiency of nonreplicating cells from one immunogenic and two nonimmunogenic mouse tumors transfected with B7‐1 or B7‐2 in the induction of protective and curative anti‐tumor immunity. Immunogenic lymphoma cells expressing B7‐1 or B7‐2 are equally effective in both protecting against a subsequent challenge and curing established tumors. By contrast, nonimmunogenic adenocarcinoma and melanoma cells expressing B7‐2 provide superior protective immunity, and only B7‐2+ adenocarcinoma cells induce an efficient curative immunity. CD8+ and polymorphonuclear cells, but not CD4+ T cells, are critically involved in the rejection of the adenocarcinoma elicited by both B7‐1+ and B7‐2+ vaccines. These data indicate that B7‐1 and B7‐2 are not redundant co‐stimulatory molecules and that, in these experimental models, B7‐2 is superior to B7‐1 in the induction of an efficient immunity when the immunogenicity of a tumor is a limiting factor.
Restricted onset of T cell receptor α gene rearrangement in fetal and neonatal thymocytes
Authors: Rytkönen Marja A. , Hurwitz Julia L. , Thompson Susan D. , Pelkonen Jukka
Published: March 2007

The initial T cell receptor (TCR) α gene rearrangements were analyzed in fetal and neonatal thymocyte hybridomas by Southern blotting. Interestingly, in 30% of all thymocyte hybridomas and in all fetal day 16 thymocyte hybridomas the most proximal Jα50 (ΨJα) gene was rearranged. This rearrangement was found on one chromosome only and mostly in association with a δ rearrangement on the homologous chromosome. Jα50 was rearranged to multiple target genes based on the variable size of the restriction fragments. In addition, δ rearrangement was found with a concomitant α rearrangement in the majority of hybridomas and it was not only associated with Jα50 but with several other rearranged Jα genes as well. Our results clearly demonstrate that T cell precursors are not pre‐committed to either δ or α rearrangement but a flexible progenitor responds to multiple regulatory signals during T cell differentiation and they do not support the notion that δrec‐ΨJα rearrangement is required for cell commitment to TCR α gene rearrangement.
T lymphocyte development in the absence of Fcϵ receptor Iγ subunit: analysis of thymic‐dependent and independent αβ and γδ pathways
Authors: Heiken Hans , Schulz Ralf‐Joachim , Ravetch Jeffrey V. , Reinherz Ellis L. , Koyasu Shigeo
Published: March 2007

During fetal development, early thymocyte progenitors transiently express low affinity Fc receptors for IgG (FcγR) of both FcγRII and III isoforms. Only the FcγRIII isoform requires association of an FcγRIII (CD16) α subunit with an FcϵRIγ homodimer for surface expression. To address the role of FcγR in ontogeny, we studied thymic development in FcϵRIγ−/− mice. We find that day 14.5 CD4−CD8− double‐negative (DN) fetal thymocytes of FcϵRIγ−/− mice express mRNA of both FcγRIIb1 and FcγRIII. Surface expression of FcγRII/III is readily detected on these cells. It appears that FcγRIIb1, whose surface expression is FcϵRIγ independent, replaces FcγRIII during thymic development in these animals. Moreover, subsequent development into CD4+CD8+ double‐positive and CD4+CD8− and CD4−CD8+ single‐positive subsets appears normal even in the absence of FcϵRIγ. However, alterations were noted in adult animals among the DN αβ TCR+ thymocytes and peripheral splenic DN T cells as well as CD8αα+ intestinal intraepithelial lymphocytes (iIEL). In contrast to conventional T lymphocytes, which do not express either FcγRIII or FcϵRIγ, DN αβ TCR+ thymocytes and extrathymically derived αβ TCR+ and γδ TCR+ CD8αα+β− iIEL express TCR which incorporate FcϵRIγ as one of their subunits. Consistent with this, the TCR levels of these cells are lower than the TCR levels on cells from wild‐type C57BL/6 mice. Despite the reduction in the level of surface TCR, the development of these cells was unaltered by the absence of FcϵRIγ. Thus, we observed alterations in adult DN αβ TCR+ thymocytes, splenic DN αβ TCR+ and DN γδ TCR+ large granular lymphocytes (LGL), and αβ TCR+ and γδ TCR+ CD8αα+β− iIEL, but no detectable changes in their major fetal thymic developmental pathways. Cultivation of peripheral DN αβ TCR+ and DN γδ TCR+ cells from FcϵRIγ−/− mice with interleukin‐2 generates LGL which mediate natural killer activity. Unlike LGL from wild‐type C57BL/6 mice, LGL from FcϵRIγ−/− mice lack FcγRIII expression and could not mediate antibody‐dependent cellular cytotoxicity through FcγRIII.
Threonine 80 on HLA‐B27 confers protection against lysis by a group of natural killer clones
Authors: Luque Isabel , Solana Rafael , Galiani M. Dolores , González Rafael , García Fernando , López de Castro José A. , Peña José
Published: March 2007

Recognition of major histocompatibility complex (MHC) class I molecules on target cells by natural killer (NK) cells inhibits NK cell‐mediated lysis. Although it is known that this inhibitory effect is regulated by MHC polymorphism, the precise structural determinants remain undefined. Based on the capacity of different HLA‐C and HLA‐B motifs specifically to inhibit cytotoxicity of some NK clones, three different NK cell specificities (NK1, NK2 and NK3) have been described. In this study, the recognition of HLA‐B27 by NK clones has been analyzed using C1R cells transfected with different HLA‐B27 subtypes as target cells. Cytotoxicity was inhibited by the HLA‐B*2705, ‐B*2701 ‐B*2703, ‐B*2704 and ‐B*2706 alleles, but not by ‐B*2702. This subtype is distinguished from the other B27 subtypes by the presence of isoleucine instead of threonine at position 80. Direct involvement of this residue was assessed by showing that site‐directed mutagenesis of Thr80 to Ile80 in HLA‐B*2705 reverted the NK protective effect of HLA‐B*2705. Based on these data, we suggest that Thr80 could act as a single residue conferring target cell protection from lysis by a group of NK clones, tentatively designated NK4.
Selective stimulation of T helper 2 cytokine responses by the anti‐psoriasis agent monomethylfumarate
Authors: de Jong Rolien , Bezemer Anja C. , Zomerdijk Timo P. L. , van de Pouw‐Kraan Tineke , Ottenhoff Tom H. M. , Nibbering Peter H.
Published: March 2007

Type 2 cytokines are thought to have a protective role in psoriasis vulgaris by dampening the activity of T helper 1 (Th1) lymphocytes. The aim of the present study was to determine the effect of monomethylfumarate (MMF), the most active metabolite of the new anti‐psoriatic drug Fumaderm®, on the production of cytokines and the development of Th subsets. MMF was found to enhance interleukin (IL)‐4 and IL‐5 production by CD2/CD8 monoclonal antibody‐stimulated peripheral blood mononuclear cells (PBMC) in a dose‐dependent manner. Maximal effects of MMF were found at a concentration of 200 μM and resulted in tenfold enhanced levels of IL‐4 and IL‐5 production. MMF did not affect the levels of IL‐2 production, interferon (IFN)‐γ production or proliferative T cell responses in these cultures. Similar effects of MMF were observed in cultures of purified peripheral blood T cells indicating that this compound can act directly on T cells. MMF did not influence cytokine production by purified CD4+CD45RA+ (unprimed) T cells, but greatly enhanced IL‐4 and IL‐5 production without affecting IFN‐γ production by purified CD4+CD45R0+ (primed) T cells. Furthermore, MMF also augmented IL‐4 and IL‐5 production in established Th1/Th0 clones that were stimulated with CD2/CD28 monoclonal antibody. Finally, when PBMC were challenged with Mycobacterium tuberculosis that typically induces Th1 recall responses with strong IFN‐γ secretion, MMF again appeared to induce high levels of IL‐4 and IL‐5 secretion while IFN‐γ production was unaffected. These results may be relevant for the development of therapeutic regimens designed to correct inappropriate Th1 subset development in immunopathologic conditions.
TNF2, a polymorphism of the tumour necrosis‐α gene promoter, is a component of the celiac disease major histocompatibility complex haplotype
Authors: Mc Manus Ross , Wilson Anthony G. , Mansfield John , Weir Donald G. , Duff Gordon W. , Kelleher Dermot
Published: March 2007

Celiac disease (CD) is an immune disease triggered by the cereal antigen gliadin, resulting in villous atrophy in the small intestine. Susceptibility to the development of CD is strongly influenced by genes in the major histocompatibility complex, in particular alleles of the DQ genes in the class II region. However recent evidence has suggested that the major histocompatibility complex (MHC) class III region may be linked to celiac disease independently of the class II region. Among the genes located in this area is TNF‐α, which encodes the cytokine tumor necrosis factor‐α which has a broad range of pro‐inflammatory, immunomodulatory and catabolic activities. Therefore, aberrant expression of TNF‐α could be important in the pathogenesis of MHC‐associated immune disorders. A TNF‐α variant with a polymorphism in its promoter region has been described and designated TNF2. TNF2 has been associated with a variety of MHC‐linked diseases, including systemic lupus erythematosus, dermatitis herpetiformis and insulin‐dependent diabetes mellitus (IDDM), as well as parasitic infections. TNF2 has previously been shown to be associated with the MHC haplotype HLAA1‐B8‐DR3‐DQ2, which confers susceptibility to CD. We have analyzed the distribution of TNF2 alleles in a group of celiac patients (n = 52) compared to controls (n = 52) in an effort to evaluate its role, if any, in susceptibility to the condition. TNF2 has a frequency of 0.5000 (SE ± 0.0490) in CD, compared to 0.1635 (± 0.0362) in a control sample (p < 10−6). Of 52 patients, 44 carried one or more TNF2 alleles. Analysis indicates that the distribution of TNF2 is best explained by assuming 100% allelic association between it and HLA‐DQB1*0201 (frequency = 0.7791 ± 0.0447). However, the number of TNF2 heterozygotes significantly exceeds expectations and measurements of linkage disequilibrium confirm that allelic associations spanning the DQ and TNF regions are strongly maintained in CD. Taken together, these results indicate that TNF2 may have a role in the pathogenesis of CD; however, since it is not an independent association, the possibility that TNF2 constitutes a passive component of the CD haplotype cannot be excluded.
Opposite effects of interleukin‐13 and interleukin‐12 on the release of inflammatory cytokines, cytokine inhibitors and prostaglandin E from synovial fibroblasts and blood mononuclear cells
Authors: Seitz Michael , Loetscher Pius , Dewald Beatrice , Towbin Harry , Baggiolini Marco
Published: March 2007

We examined the effects of interleukin‐12 (IL‐12) and interleukin‐13 (IL‐13) on cytokine, cytokine inhibitor and prostaglandin E (PGE) release from synovial fibroblasts and blood mononuclear cells (MNC). In resting synovial fibroblasts, we found that IL‐13 is an inhibitor of IL‐8 and PGE release. A significant decrease of PGE synthesis caused by IL‐13 was also observed in tumor necrosis factor (TNF)‐α‐stimulated synovial fibroblasts, whereas IL‐12 had no regulatory effects on these cells. In resting and cytokine‐stimulated MNC, IL‐13 markedly inhibited IL‐1β, IL‐8 and monocyte chemoattractant protein‐1 (MCP‐1) release and potently stimulated interleukin‐1 receptor antagonist (IL‐1ra) synthesis. In contrast, IL‐12 stimulated the production of IL‐1β and MCP‐1 in TNF‐α‐stimulated MNC and inhibited IL‐1ra synthesis in cytokine‐stimulated cells. These findings identify novel biological actions of IL‐12 and IL‐13 on connective tissue and on blood mononuclear cells which indicate their regulatory functions as enhancer and suppressor of inflammatory processes, respectively.
Neurons induced to express major histocompatibility complex class I antigen are killed via the perforin and not the Fas (APO‐1/CD95) pathway
Authors: Rensing‐Ehl Anne , Malipiero Ursula , Irmler Martin , Tschopp Jürg , Constam Daniel , Fontana Adriano
Published: March 2007

Cytotoxic T lymphocytes (CTL) kill target cells by perforin‐mediated pore formation, induction of apoptosis by the Fas ligand, or both. It has been demonstrated that depolarized neurons can be induced to express major histocompatibility complex (MHC) class I antigens by interferon‐γ. Evidence for antigen‐dependent CTL‐mediated killing was obtained by transfecting neurons with MHC class I cDNA. The present study was designed to investigate the mechanisms of killing of cerebellar granule neurons depolarized by high K+ concentrations and thereby inducible for MHC class I antigen expression. We found that neurons express only low levels of Fas (APO‐1/CD95) and are resistant to Fas ligand‐mediated killing even when pretreated with cytokines. However, granules extracted from CTL as well as purified perform induce almost complete lysis of neurons. These data suggest that CTL‐mediated elimination of neurons involves the perform, but not the Fas pathway of target cell killing.
The outer surface lipoprotein OspA of Borrelia burgdorferi provides co‐stimulatory signals to normal human peripheral CD4+ and CD8+ T lymphocytes
Authors: Knigge Heidrun , Simon Markus M. , Meuer Stefan C. , Kramer Michael D. , Wallich Reinhard
Published: March 2007

Studies in man and mice have indicated that T cells induced during Borrelia burgdorferi infection are involved in the pathogenesis of the disease. We analyzed the ability of B. burgdorferi to provide co‐stimulatory signals to highly enriched normal human CD2+ T lymphocytes in the presence of suboptimal concentrations of immobilized anti‐CD3 antibodies. Here we show that the lipid‐containing recombinant outer surface lipoprotein A (rlip‐OspA) of B. burgdorferi but not its delipidated derivative rNS1‐OspA augmented CD3‐induced T cell proliferation in a dose‐dependent manner and at levels similar to that obtained with anti‐CD28 antibodies. Lipopolysaccharide had no effect in this system at any concentration tested, suggesting that the active principle of co‐stimulation is associated with the lipid moiety of rlip‐OspA and distinct from conventional lipid A. Furthermore, incubation of CD2+ T cells or selected CD4+ as well as CD8+ subpopulations with rlip‐OspA, but not with rNS1‐OspA led to the production of interferon (IFN)‐γ, interleukin (IL)‐6 and tumor necrosis factor (TNF)‐α, but not IL‐4. In contrast, co‐stimulation of the respective T cell populations with anti‐CD28 antibodies resulted in the generation of IFN‐γ, IL‐4 and TNF‐α, but not IL‐6. This indicated that the signal transduction pathway induced by rlip‐OspA is distinct from that elicited via the CD28 receptor. Co‐stimulation of T cells with rlip‐OspA also resulted in the development of cytolytic effector cells. In light of the fact that inflamed tissues of B. burgdorferi‐infected hosts contain blood leukocytes together with spirochetes, their degradation products, or both, these results suggest that infiltrating CD4+ and CD8+ T cells of any specificities, including spirochetes, autoantigens, or both, participate in the pathogenesis of Lyme disease.
Antigen‐specific targeting of CD28‐mediated T cell co‐stimulation using chimeric single‐chain antibody variable fragment‐CD28 receptors
Authors: Alvarez‐Vallina Luis , Hawkins Robert E.
Published: March 2007

T cells require two distinct signals for optimal activation. One is an antigen‐specific signal and is provided by engagement of the T cell receptor (TCR). The second is an antigen‐independent signal mediated by engagement of the T cell surface molecule CD28 with members of the B7 family. To endow CD28 molecules with antibody‐type recognition, we have constructed chimeric single‐chain antibody variable fragment (scFv)‐CD28 molecules; following transfection of the genes encoding such constructs into the Jurkat human T cell line we show that they are stably expressed as functional cell surface receptors. These chimeric molecules have no apparent negative effects on the expression and signaling ability of the wild‐type CD28 and TCR/CD3 molecules. When combined with signaling via the TCR/CD3 complex, these antigen‐specific scFv‐CD28 chimeric molecules provide signals similar to those elicited by the cross‐linking of the unmodified CD28 molecules. Furthermore, the generation of double transfectants simultaneously expressing scFv‐CD28 and scFv‐CD3ζ chimeras demonstrates that antigen‐specific co‐stimulatory signals can also synergize with signals mediated through chimeric CD3ζ chains to secrete maximal levels of interleukin‐2. Overall, our results suggest that optimal, predefined antigen‐specific activation of T cells directed by the specificity of the scFv should be possible.
Tyrosine kinase and cAMP‐dependent protein kinase activities in CD40‐activated human B lymphocytes
Authors: Lapointe Réjean , Lemieux Réal , Olivier Martin , Darveau André
Published: March 2007

In vitro, human B lymphocytes undergo long‐term proliferation when activated through CD40, a protein expressed on their cell surface. The nature of CD40‐dependent signals in proliferating fresh human Epstein‐Barr virus‐negative B lymphocytes is currently unknown. In this study, a CD40‐dependent B cell culture system was used to examine the role of different signal transduction elements. Protein kinase C (PKC) depletion generated by a long‐term phorbol 12‐myristate 13‐acetate treatment had weak effects on proliferation. Rather, tyrosine phosphorylation was shown to be directly involved in mediating CD40‐dependent signals. The use of the protein tyrosine kinase (PTK)‐specific inhibitor herbimycin A dramatically decreased cellular proliferation without altering the activity of the human immunodeficiency virus‐1 long terminal repeat (HIV‐1 LTR), a promoter largely dependent on the binding of nuclear factor ϰB (NF‐ϰB). In contrast, the cAMP‐dependent protein kinase specific inhibitor H‐89 totally inhibited HIV‐1 LTR activity at a concentration as low as 100 nM without affecting cellular proliferation. Electrophoretic mobility shift assay (EMSA) and supershift assay using an NF‐ϰB binding sequence from the ϰ light chain as a probe, revealed that both p65 (RelA) and c‐Rel were present in CD40‐stimulated B cells. While PKC depletion did not alter the NF‐ϰB level, treatment of B lymphocytes with H‐89 or herbimycin A provoked a decrease in the NF‐ϰB level. These observations establish the importance of different signal transducing pathways leading to CD40 activation of B lymphocytes.
Chronic relapsing experimental autoimmune encephalomyelitis (CREAE) in mice: enhancement by monoclonal antibodies against interferon‐γ
Authors: Heremans Hubertine , Dillen Chris , Groenen Marleen , Martens Eric , Billiau Alfons
Published: March 2007

Experimental autoimmune encephalomyelitis (EAE) is a T cell‐mediated inflammatory and demyelinating disorder of the central nervous system. Depending on the experimental conditions, it takes an acute monophasic or a chronic relapsing‐remitting course. We have previously reported that the incidence and severity of acute EAE in mice are reduced by administration of interferon (IFN)‐γ and augmented by treatment with neutralizing antibodies against IFN‐γ. Here, we investigated the role of IFN‐γ in chronic relapsing models of EAE (CREAE) in SJL/J and Biozzi ABH mice. Spontaneous relapses in Biozzi mice as well as induced relapses in SJL/J mice were facilitated by administration of neutralizing monoclonal antibody (mAb) against IFN‐γ in the disease‐free interval. The enhancing effect of anti‐IFN‐γ mAb given before and during the primary attack did not carry over to the relapses. However, early administration of IFN‐γ in Biozzi mice, which developed spontaneous relapses in a high proportion, provided partial protection not only against the first attack, but also against subsequent relapses. Administration of exogenous IFN‐γ during the remission phase provided some protection against subsequent relapses. These results indicate that in both types of relapses, IFN‐γ is produced and does provide a certain degree of protection against disease progression.
Characterization of T cell receptor single‐chain Fv fragments secreted by myeloma cells
Authors: Grégoire Claude , Malissen Bernard , Mazza Gilbert
Published: March 2007

Myeloma cells have been used to produce milligram quantities of soluble αβ T cell receptor (TCR) molecules as single‐chain polypeptides in which the TCR variable (V) domains are connected by a peptide linker (TCR scFv). Unlike most TCR scFv produced in bacteria, the purified TCR scFv were stable and showed no tendency to aggregate when kept at concentrations up to 10 mg/ml. Circular dichroism analyses of the TCR scFv indicated that they contained a high proportion of β‐pleated sheet structures. Since the Vα subunits present in the TCR scFv contained their own signal sequences, they provided the opportunity to determine by N‐terminal amino acid sequencing the position of the signal cleavage of three distinct mouse Vα. Two of the experimentally determined signal cleavage sites differed from those previously predicted on the basis of biochemical and statistical criteria. The expression approach outlined in this report has been applicable to three distinct αβ TCR and should contribute to the large scale production of soluble TCR amenable to structural studies.
Purified human peripheral blood basophils release interleukin‐13 and preformed interleukin‐4 following immunological activation
Authors: Gibbs Bernhard F. , Haas Helmut , Falcone Franco H. , Albrecht Claudia , Vollrath Ilka B. , Noll Tobias , Wolff Helmut H. , Amon Ulrich
Published: March 2007

Recent studies have shown that human basophils, like mast cells, generate interleukin (IL)‐4 following immunological activation and may thus participate in late‐phase allergic and inflammatory processes. Here, we report the capacity of human basophils to release IL‐13 within 24 h following stimulation with anti‐IgE. Additionally, in 14 out of 31 experiments, we observed that basophils rapidly release preformed IL‐4 within 5–10 min, as well as newly generated IL‐4, which was released 4 h following stimulation of the cells with anti‐IgE. In contrast to the biphasic release of IL‐4 from the cells, no preformed IL‐13 was detected at earlier times (5–30 min). Preformed IL‐4 and IL‐4 and IL‐13 generated de novo were also released after stimulation of the cells with IL‐3; an enhanced production of these cytokines was observed using a combination of IL‐3 and anti‐IgE. We conclude from these data that, by releasing preformed IL‐4 and IL‐4 and IL‐13 generated de novo, human basophils may be centrally involved in the orchestration of allergic inflammation by providing a trigger to IL‐4‐mediated T helper 2 lymphocyte activation, B cell IgE switching, and increased vascular adhesion molecule expression.
Negative regulation of rat natural killer cell activity by major histocompatibility complex class I recognition
Authors: Kraus Elmar , Lambracht Doris , Wonigeit Kurt , Hünig Thomas
Published: March 2007

The cytolytic activity of human and mouse natural killer (NK) cells is negatively regulated by self major histocompatibility complex (MHC) class I molecules on potential target cells. In the rat, protection by RT1 class I gene products has so far not been formally shown although the complex effects of foreign and self RT1 genes on polyclonal NK cell activity suggest that MHC recognition can have both stimulatory and inhibitory effects. Here we report that the expression of self‐MHC class I molecules on target cells strongly inhibits lysis by a long term NK cell line derived from LEW (RT1l) rats and by LEW NK cells activated by short‐term culture in the presence of interleukin‐2. This was demonstrated with mouse‐rat hybridoma target cells expressing different rat MHC alleles and with mouse tumor target cells transfected with classical (RT1.Al) and nonclassical (RT1.Cl) rat MHC class I genes. With hybridoma target cells, the strongest reduction in lysis as compared to the parental mouse myeloma line was observed when “self” (LEW) MHC was expressed, while hybridomas expressing other MHC alleles showed less and variable reduction. Transfection of RT1.Al protected both L‐929 fibroblasts and P815 mastocytoma cells from lysis by the NK cell line, while RT1.Cl only protected P815 cells, indicating that additional target cell properties regulate rat NK cell activity.
Dendritic cells process exogenous viral proteins and virus‐like particles for class I presentation to CD8+ cytotoxic T lymphocytes
Authors: Bachmann Martin F. , Lutz Manfre D. B. , Layton Guy T. , Harris Stephen J. , Fehr Thomas , Rescigno Maria , Ricciardi‐Castagnoli Paola
Published: March 2007

Previous reports have indicated that both dendritic cells and macrophages have the ability to induce cytotoxic T lymphocyte (CTL) and T helper (Th) cell responses in vivo. Dendritic cells process exogenous antigens conventionally for presentation on major histocompatibility complex (MHC) class II molecules. However, unconventional processing of exogenous antigens in vitro for presentation on MHC class I molecules is still an open question. In this study, we report that a cloned dendritic cell line (D2SC/1) is able to present cell debris‐associated exogenous viral proteins to MHC class I‐restricted CTL in vitro The dendritic cell line was very efficient in processing recombinant lymphocytic choriomeningitis virus nucleoprotein (LCMV NP) and presenting the class I‐restricted epitope to CTL primed in vivo. Peritoneal macrophages could also process the recombinant LCMV NP for subsequent MHC class I presentation, but were less efficient compared to the dendritic cells. Furthermore, recombinant yeast‐derived virus‐like particles carrying the HIV‐1 V3 loop (V3‐VLP), which are protenaceous and do not contain any lipid, were also found to be efficiently processed by the dendritic cell line for presentation of the class I‐restricted epitope. These results clearly indicate that viral proteins, in particulate form or associated with cell debris, are processed by dendritic cells for CTL induction.
Regulation of interleukin‐10 production by β‐adrenergic agonists
Authors: Suberville Sidonie , Bellocq Agnès , Fouqueray Bruno , Philippe Carole , Lantz Olivier , Perez Joëlle , Baud Laurent
Published: March 2007

Catecholamines have been shown to inhibit some aspects of macrophage activation through a β receptor‐dependent mechanism. This study was undertaken to analyze the effects of isoproterenol, a specific β‐adrenergic agonist, on the synthesis of interleukin‐10 (IL‐10), a major macrophage‐deactivating factor. Isoproterenol increased IL‐10 release from lipopolysaccharide‐(LPS)‐activated mouse peritoneal macrophages in a dose‐dependent manner. A significant effect was already observed with 1 μM isoproterenol, while a 4.5‐fold increase was achieved with 10 μM. This increase was observed only if macrophages were exposed to isoproterenol for at least 2 h before LPS challenge. It was apparent within 0.5 h and persisted through 24 h at all the LPS concentrations used. A similar increase was observed at the IL‐10 mRNA level, as judged by enzyme‐linked immunosorbent assay‐polymerase chain reaction. The macrophage response to isoproterenol that led to cyclic AMP accumulation was markedly inhibited by H‐89, a potent inhibitor of protein kinase A. These data suggest the involvement of cyclic AMP in the regulation of IL‐10 synthesis by isoproterenol. IL‐10 was in turn partly responsible for a reduction in tumor necrosis factor‐α synthesis. In vivo, the administration of oxprenolol, a β‐receptor antagonist, significantly reduced serum IL‐10 levels 90 min after LPS challenge. Thus, the present study provides the first evidence that endogenous catecholamines are of critical importance in determining the magnitude of the IL‐10 response in experimental endotoxemia.
A wild‐type p53 cytotoxic T cell epitope is presented by mouse hepatocarcinoma cells
Authors: Lacabanne Valérie , Viguier Mireille , Guillet Jean‐Gérard , Choppin Jeannine
Published: March 2007

The possibility to identify epitopes presented by tumor cells to cytotoxic T lymphocytes (CTL) has given rise to new fields in tumor immunology. The tumor suppressor gene product p53 is a good candidate antigen because it is involved in the tumorigenesis of many cancers. It accumulates in an inactivated form due to mutation or formation of heterodimers with an oncogene product. Epitopes from the mutant or wild‐type p53 proteins are thought to be presented by tumor cells and to induce a tumor‐specific CTL response. To identify such epitopes, mouse wild‐type p53 peptides encompassing the H‐2 Db anchoring motif were tested for their association with the Db molecule. Positive peptides were assayed for their ability to induce CTL in C57BL/6 mice. CTL specific for one wild‐type p53 peptide, p232–240, were isolated and found to lyse hepatocarcinoma cell lines established from mice transgenic for simian virus 40 large T antigen which overexpress p53. These results show that the p232–240 epitope from wild‐type p53 is naturally processed and presented in H‐2b tumor cells.