bluuebunny/crossref_metadata_2025 huggingface dataset:
Interferon‐γ confers resistance to experimental allergic encephalomyelitis
Authors: Krakowski Michelle , Owens Trevor
Published: March 2007

In experimental allergic encephalomyelitis (EAE), T cells infiltrate the central nervous system (CNS) and induce inflammation. These CD4+ T cells secrete interferon (IFN)‐γ, levels of which correlate with disease severity, and which is proposed to play a key role in disease induction. Many strains of mice are resistant to EAE. We have studied the effect of deletion of IFN‐γ on the ability to induce EAE in resistant BALB/c‐backcrossed mice. As expected, only 0–6 % of BALB/c or BALB/c‐backcrossed mice developed EAE when immunized with myelin basic protein in adjuvant. Strikingly, abrogation of IFN‐γ expression by targeted disruption of the IFN‐γ gene (GKO mice) converted them to a susceptible phenotype. As many as 71 % of these IFN‐γ‐deficient mice developed EAE, a frequency comparable to that seen with the susceptible SJL/J strain. In addition, EAE was of unusually high severity in mice lacking IFN‐γ. Immunological characteristics of disease in IFN‐γ‐deficient mice were comparable to those seen in susceptible (SJL/J) mice with EAE, including perivascular infiltration in the CNS and order‐of‐magnitude increases for both CD3 γ chain and TNF‐α mRNA levels in the spinal cord. We thus demonstrate that lack of IFN‐γ converts an otherwise EAE‐resistant mouse strain to become susceptible to disease. Therefore, in BALB/c mice, IFN‐γ confers resistance to EAE.
Interferon‐γ‐inducing factor enhances T helper 1 cytokine production by stimulated human T cells: synergism with interleukin‐12 for interferon‐γ production
Authors: Micallef Mark J. , Ohtsuki Takashi , Kohno Keizo , Tanabe Fujimi , Ushio Shimpei , Namba Motoshi , Tanimoto Tadao , Torigoe Kakuji , Fujii Mitsukiyo , Ikeda Masao , Fukuda Shigeharu , Kurimoto Masashi
Published: March 2007

The novel cytokine interferon‐γ‐inducing factor (IGIF) augments natural killer (NK) cell activity in cultures of human peripheral blood mononuclear cells (PBMC), similarly to the structurally unrelated cytokine interleukin (IL)‐12. IGIF has been found to enhance the production of interferon‐γ (IFN‐γ) and granulocyte/macrophage colony‐stimulating factor (GM‐CSF) while inhibiting the production of IL‐10 in concanavalin A (Con A)‐stimulated PBMC. In this study, when anti‐CD3 monoclonal antibody (mAb)‐stimulated human enriched T cells were exposed to IGIF, the cytokine dose‐dependently enhanced the proliferation of the cells and this could be completely inhibited by a neutralizing antibody against IL‐2 at lower concentrations of IGIF. Neutralizing antibody against IFN‐γ had only insignificant inhibitory effects on T cell proliferation at higher concentrations of IGIF. Enzyme‐linked immunosorbent assays (ELISA) revealed that, like PBMC, T cells exposed to IGIF produced large amounts of IFN‐γ; however, changes in the production of IL‐4 and IL‐10 were minimal. IGIF, but not IL‐12, significantly enhanced IL‐2 and GM‐CSF production in T cell cultures, as determined by CTLL‐2 bioassay and ELISA, respectively; however, both IGIF and IL‐12 enhanced IFN‐γ production by the T cells. When T cells were exposed to a combination of IGIF and IL‐12, a synergistic effect was observed on the production of IFN‐γ, but not on production of IL‐2 and GM‐CSF. In conclusion, IGIF enhances T cell proliferation apparently through an IL‐2‐dependent pathway and enhances Th1 cytokine production in vitro and exhibits synergism when combined with IL‐12 in terms of enhanced IFN‐γ production but not IL‐2 and GM‐CSF production. Based on structural and functional differences from any known cytokines, it was recently proposed that this cytokine be designated interleukin‐18.
Interferon‐γ is required during the initiation of an organ‐specific autoimmune disease
Authors: Barrett Simon P. , Gleeson Paul A. , De Silva Harini , Toh Ban‐Hock , van Driel Ian R.
Published: March 2007

Autoimmune gastritis induced by neonatal thymectomy of mice is a CD4+ T cell‐mediated organ‐specific autoimmune disease. The characteristic features of autoimmune gastritis, which include a mononuclear infiltrate within the gastric mucosa, loss of parietal and chief cells and circulating autoantibodies to the gastric H+/K+ ATPase, appear 6–10 weeks after thymectomy. Here we have assessed the role of interferon‐γ (IFN‐γ) in the pathogenesis of the gastric lesion. Splenic T cells derived from mice with gastritis produced three‐ to tenfold more IFN‐γ than T cells from normal animals after stimulation with anti‐CD3 antibodies. Treatment of neonatally thymectomized mice at weekly intervals for 6 or 12 weeks with a neutralizing rat monoclonal antibody to mouse IFN‐γ abolished the production of anti‐gastric autoantibodies and decreased the incidence of gastric mononuclear infiltrates from the 69% observed in normal rat immunoglobulin (Ig)‐injected mice to 16%. Further, in mice treated with only a single dose of anti‐IFN‐γ immediately after thymectomy at 3 days after birth, the incidence of autoimmune gastritis was 1/19 compared to 8/19 in normal rat Ig‐injected mice. Prevention of autoimmunity by neutralization of IFN‐γ several weeks prior to the detection of a pathological lesion strongly suggests that IFN‐γ plays an essential role in the initiation of the gastric autoimmune response.
Increased expression of CD80, CD86 and CD70 on T cells from HIV‐infected individuals upon activation in vitro: regulation by CD4+ T cells
Authors: Wolthers Katja C. , Otto Sigrid A. , Lens Susanne M. A. , Kolbach Dinanda N. , van Lier René A. W. , Miedema Frank , Meyaard Linde
Published: March 2007

T cells express CD28 and CD27 which transduce co‐stimulatory signals after interaction with their ligands on antigen‐presenting cells (APC). These ligands, CD80, CD86 and CD70, are also expressed to some extent on activated T cells. Here, we show that in human immunodeficiency virus (HIV)‐infected individuals, CD28 and CD27 expression is decreased on CD8+ T cells. On the other hand, T cell stimulation in vitro induced high CD80, CD86 and CD70 expression on T cells from HIV‐infected individuals. It appeared that an inverted CD4:CD8 T cell ratio could explain this enhanced expression of co‐stimulatory ligands. Indeed, high expression levels of CD80, CD86 and CD70 were found on activated CD8+ T cells from HIV− individuals cultured in the absence of CD4+ T cells. Addition of CD4+ T cells prevented this up‐regulation. However, in HIV‐infected individuals, addition of excess autologous or healthy control CD4+ T cells did not completely counteract up‐regulation of co‐stimulatory ligand expression on CD8+ T cells. Thus, to some extent, CD8+ T cells in HIV‐infected individuals appeared to be refractory to CD4+ T cell‐mediated regulation of ligand expression in vitro. Activated T cells from HIV‐infected individuals and activated CD8+ T cells from healthy controls were able to act as accessory cells in CD3‐induced T cell proliferation, which was dependent on cell‐cell contact. Thus, we showed that T cells from HIV‐infected individuals express enhanced levels of co‐stimulatory ligands upon activation, which provides them with accessory cell properties. Enhanced stimulatory potential of these nonprofessional APC may contribute to persistently high levels of immune activation in HIV infection related to disease progression.
Identification of a tonsil IgD+ B cell subset with phenotypical and functional characteristics of germinal center B cells
Authors: Billian Gisèle , Bella Chantal , Mondière Paul , Defrance Thierry
Published: March 2007

We have identified and isolated a subpopulation of IgD+ B cells (IgD+ CD38+ B cells) from human tonsils which expresses the germinal center (GC)‐associated surface markers CD10, CD38, CD75, CD77 and CD95/Fas. The heterogeneity of expression of several markers on IgD+ CD38+ B cells suggests that this population can be further subdivided into two discrete subtypes. On a functional basis, IgD+ CD38+ B cells behave as GC B cells as they rapidly and spontaneously undergo apoptosis in vitro and cannot be stimulated to synthesize DNA upon cross‐linking of the antigen receptor. However, in contrast with most GC B cells, IgD+ CD38+ B cells have not completed Ig class switching since they predominantly secrete IgM following stimulation in vitro and lack surface expression of secondary isotypes. Immunoenzymatic staining performed on tonsil tissue sections revealed that IgD+ CD38+ B cells are located in two distinct histological structures: within the GC of a few classical secondary follicles, in which they appear as scattered cells, and within rare atypical GC, homogeneously constituted of IgD+ B cells. Taken together, our findings indicate that IgD+ CD38+ B cells constitute a novel subset of GC B cells. The possibility that these cells could represent an early stage of the follicular reaction or be generated in response to certain bacterial carbohydrate antigens is discussed.
Expression of an avian CD6 candidate is restricted to αβ T cells, splenic CD8+ γδ T cells and embryonic natural killer cells
Authors: Göbel Thomas W. F. , Chen Chen‐Lo H. , Cooper Max D.
Published: March 2007

A candidate avian CD6 homolog is identified by the S3 monoclonal antibody. The S3 antigen exists in a phosphorylated glycoprotein form of 130 kDa and a nonphosphorylated form of 110 kDa. Removal of phosphate groups and N‐linked carbohydrates indicates a 78‐kDa protein core. During thymocyte differentiation, the γδ T cells do not express S3, whereas mature CD4+ and CD8+ cells of αβ lineage acquire S3 antigen. All αβ T cells in the blood and spleen express the S3 antigen at relatively high levels. In contrast, only the CD8+ sub‐population of γδ T cells in the spleen expresses the antigen and neither αβ nor γδ T cells in the intestinal epithelium express the S3 antigen. The S3 antigen is also found on embryonic splenocytes with a phenotypic profile characteristic of avian natural killer cells. The biochemical characteristics and this cellular expression pattern imply that the S3 antigen is the chicken CD6 homolog.
Establishment of tissue‐specific tolerance is driven by regulatory T cells selected by thymic epithelium
Authors: Modigliani Yves , Coutinho Antonio , Pereira Pablo , Le Douarin Nicole , Thomas‐Vaslin Véronique , Burlen‐Defranoux Odile , Salaün Josselyne , Bandeira Antonio
Published: March 2007

Grafts of thymic epithelium (TE) rudiments restore T cell development and function in allogeneic athymic mice. These TE chimeras are specifically tolerant to grafts of peripheral tissues (e.g. skin and heart) from the TE donor strain, although they harbor peripheral immunocompetent T cells capable of rejecting those grafts. Initial analysis has shown that TE chimeras also harbor TE‐selected CD4 T lymphocytes that inhibit graft rejection by tissue‐reactive T cells in immunocompetent recipients. Peripheral tolerance in TE chimeras is thus maintained by dominant mechanisms dependent on regulatory CD4 T lymphocytes. Here we show that TE‐selected regulatory T cells recruit nontolerant tissue‐reactive CD4 and CD8 T cells to express similar regulatory functions. Only recent thymic emigrants, but not peripheral resident mature T cells are susceptible to this process of functional education, which also requires exposure to specific antigens and occurs entirely in the periphery. We propose that these mechanisms play a major role in establishing and maintaining natural self tolerance to tissue‐specific antigens.
A novel surface molecule homologous to the p58/p50 family of receptors is selectively expressed on a subset of human natural killer cells and induces both triggering of cell functions and proliferation
Authors: Bottino Cristina , Sivori Simona , Vitale Massimo , Cantoni Claudia , Falco Michela , Pende Daniela , Morelli Luigia , Augugliaro Raffaella , Semenzato Gianpietro , Biassoni Roberto , Moretta Lorenzo , Moretta Alessandro
Published: March 2007

Human natural killer (NK) cells express inhibitory (p58) or activatory (p50) receptors for HLA‐C alleles. Here, we describe a novel member of the p58/p50 family that is expressed by a subset of NK cells in about one third of donors. This molecule, termed p50.3, mediates NK cell triggering as revealed by the induction of intracellular free calcium mobilization, cytokine release and cytotoxicity. In addition, anti‐p50.3 monoclonal antibody (mAb) induced a selective, strong proliferation of p50.3+ NK cells in peripheral blood lymphocytes. Although p50.3 molecules do not appear to display an obvious HLA class I specificity, they are usually coexpressed with known inhibitory receptors for HLA class I alleles. mAb‐mediated cross‐linking of these receptors leads to inhibition of the anti‐p50.3 mAb‐induced NK cell activation and proliferation. Surface p50.3 molecules are glycoproteins of ∼ 55–58 kDa which, upon deglycosylation, display a relative molecular mass of 36 kDa, similar to that of deglycosylated (activatory) p50 receptors. Analysis of the two‐dimensional peptide maps of the 50.3 molecules revealed a high homology with the other HLA‐C‐specific p58/p50 receptors. The use of a set of oligodeoxynucleotide primers, previously shown to amplify the activatory (p50) forms of HLA‐C‐specific receptors, consistently amplified in p50.3+ clones a cDNA sequence termed KKA3. This sequence belongs to the p58/p50 multigene family, that encodes for a transmembrane protein specifically stained by anti‐p50.3 mAb in cell transfectants. Similar to p50 molecules, the KKA3‐encoded molecules are characterized by two extracellular immunoglobulin‐like domains, by the presence of a lysine in the transmembrane region and a short (39 amino acids) cytoplasmic tail which does not contain immune receptor tyrosine‐based activation motifs (ITAM)‐like sequences.
A biological function for the XP motif within the N terminus of major histocompatibility complex class II‐associated peptides
Authors: Breloer Minka , Ehrlich Svenja , Fleischer Bernhard , von Bonin Arne
Published: March 2007

A high proportion (up to 30%) of major histocompatibility complex (MHC) class II‐bound peptides in the mouse and humans contains a proline residue at the N‐terminal penultimate position (XP motif). We used a set of ovalbumin (OVA)‐specific and hen egg lysozyme (HEL)‐specific T cell hybridomas and asked whether the XP motif in MHC class II‐associated peptides might influence the stimulation of T cells. We created N‐terminally substituted variants of OVA323–339, an H2‐Ad restricted OVA epitope and of HEL50–63, a dominant epitope in the context of H2‐Ak. Our results show that the N‐terminal sequence of MHC class II‐bound peptides has a strong impact for the overall stimulation of specific T cells. Proline at the N terminus of antigenic peptides, in contrast to other amino acids, is tolerated or even enhances the recognition of MHC class II‐bound peptides significantly.
Distinct mechanisms mediate SHC association with the activated and resting B cell antigen receptor
Authors: D'Ambrosio Daniele , Hippen Keli L. , Cambier John C.
Published: March 2007

Ligation of the B cell antigen receptor (BCR) complex initiates tyrosine phosphorylation of the receptor's transducer components, Ig‐α and Ig‐β and tyrosine kinase‐dependent accumulation of GTP‐bound, activated p21ras. The mechanism of receptor coupling to p21ras activation and the roles of Ig‐α and Ig‐β are unknown. The results reported here indicate that the resting, nonphosphorylated BCR associates with the Grb‐2/Sos‐linker SHC via the Ig‐α immunoreceptor‐based tyrosine activation motif (ITAM). Ig‐α specificity of this interaction is determined by the sequence DCSM found in Ig‐α, but not Ig‐β. Tyrosine phosphorylation of Ig‐α and Ig‐β ITAM allows recruitment of SHC, which now binds directly to both Ig‐α and Ig‐β via a phosphotyrosine/SH2 interaction. In confirmation of recent studies by Saxton et al. (J. Immunol. 1994. 153: 623) receptor ligation leads to tyrosine phosphorylation of SHC and to the formation of a phospho‐SHC/Grb2/Sos complex. In view of previous studies which demonstrated p21ras co‐capping with ligated BCR, the data presented here suggest that Ig‐α/β‐ and SHC tyrosine phosphorylation‐dependent recruitment of the Grb2/Sos complex to the receptor can occur and may provide a mechanism by which the nucleotide exchange activity of Sos could mediate activation of BCR‐localized p21ras.
CD32 expression and signaling is down‐regulated by transforming growth factor‐β1 on human monocytes
Authors: Reterink Thomas J. F. , Klar‐Mohamad Ngaisah , Nibbering Peter H. , Van Es Leendert A. , Daha Mohamed R.
Published: March 2007

CD32 (FcγRII) is the most abundantly distributed class of IgG Fc receptors in the human body. In this study, we analyzed the effect of transforming growth factor (TGF)‐β1, a cytokine with strong immunosuppressive function, on the expression and function of CD32 on freshly isolated peripheral blood monocytes and three human monocytic cell lines, U937, THP‐1 and Mono mac‐6. We found that TGF‐β1 down‐regulates CD32 expression on monocytes and all monocytic cell lines in a dose‐ and time‐dependent fashion. A mean down‐regulation of CD32 expression on THP‐1 cells of 54 ± 3.2% after 24 h was found at a concentration of 1 ng/ml TGF‐β1. At the mRNA level, TGF‐β1 induced a twofold down‐regulation of CD32. Cross‐linking of CD32 induced an increase in the concentration of intracellular Ca2+, which was reduced by 50% by TGF‐β1, suggesting a decreased downstream signaling mediated by the receptor.
Triggering through CD40 promotes interleukin‐4‐induced CD23 production and enhanced soluble CD23 release in atopic disease
Authors: Paterson Robin L. K. , Lack Gideon , Domenico Joanne M. , Delespesse Guy , Leung Donald Y. M. , Finkel Terri H. , Gelfand Erwin W.
Published: March 2007

The pathogenesis of atopic disease is closely linked to the overproduction of IgE. CD23 and CD40 are two cellular receptors involved in the regulation of IgE production and both receptors are elevated in atopic disease. We have examined the role of CD40 in the regulation of CD23 and soluble CD23 production in healthy and atopic donors. Triggering of the B cell CD40 receptor directly enhances interleukin (IL)‐4‐mediated up‐regulation of CD23 at both the protein and the mRNA level. When atopic donors were studied, the synergistic effect of CD40 triggering on the IL‐4‐induced up‐regulation of CD23 and soluble CD23 (sCD23) was enhanced and there was a relative skewing toward production of sCD23. These studies implicate the CD40 receptor in the hyperproduction of CD23 and sCD23 in atopic disease and suggest that abnormalities may exist in the cellular pathways leading to sCD23 production.
Nerve growth factor signal transduction in human B lymphocytes is mediated by gp140trk
Authors: Melamed Isaac , Kelleher Colm A. , Franklin Richard A. , Brodie Chaya , Hempstead Barbara , Kaplan David , Gelfand Erwin W.
Published: March 2007

Nerve growth factor (NGF) plays an important role in the regulation of the immune system. Recent studies from this laboratory demonstrated the presence of functional NGF receptors on human B lymphocytes; in addition, NGF has been shown to enhance B lymphocyte proliferation. NGF caused both concentration‐ and time‐dependent increases in tyrosine phosphorylation of five proteins of 140, 110, 85, 60 and 42 kDa, which were identified as phospholipase C‐γ1, phosphatidylinositol‐3 kinase and mitogen‐activated protein kinase. To elucidate the contribution of the Trk family of tyrosine kinases to the phosphorylation events induced by NGF, we identified gp140trk in human B cells and in human B cell lines. Analysis of specific gp140trk immunoprecipitates indicated that addition of NGF to B cells induced a rapid increase in the tyrosine phosphorylation of gp140trk and inhibition of this phosphorylation prevented the tyrosine phosphorylation of other proteins. These data identify the central role of gp140trk in NGF signaling of human B lymphocytes.
Subepithelial B cells in the human palatine tonsil. II. Functional characterization
Authors: Dono Mariella , Zupo Simona , Augliera Adriano , Burgio Vito L. , Massara Rosanna , Melagrana Andrea , Costa Mauro , Grossi Carlo E. , Chiorazzi Nicholas , Ferrarini Manlio
Published: March 2007

This study investigates the main functional features of subepithelial (SE) B cells and compares them with those of purified germinal center (GC) and follicular mantle (FM) B cells isolated from the same tonsils. Unlike GC B cells, SE B cells failed to produce polyspecific antibodies in vitro; unlike GC B cells, SE B cells expressed high levels of Bcl‐2 and failed to undergo spontaneous apoptosis in vitro. The most striking function of SE B cells was their ability to produce IgM antibodies to T cell‐independent type‐2 (TI‐2) (but not to TI‐1) antigens (Ag). These antibodies could not be detected when both FM and GC B cells were stimulated with TI‐2 Ag in vitro. Moreover, B cells isolated from peripheral blood were unable to mount a response to TI‐2 Ag. The latter finding is consistent with the observation that B cells with the phenotypic features of SE B cells were virtually absent in the peripheral blood and emphasizes the notion that SE B cells belong to a subset of non‐recirculating B cells. SE B cells were by far superior to FM B cells in mixed lymphocyte reaction (MLR) stimulation of allogeneic T cells in vitro, although they were not as efficient as dendritic cells (DC). In order to stimulate T cells efficiently, SE B cells had to be exposed to anti‐μ antibody, a treatment which induced expression of activation markers such as CD80, CD86, CD69 and CD39, usually absent in resting SE B cells. CD80 and CD86 molecules expressed by SE B cells participated in the chain of events required to promote the proliferation of allogeneic T cells as demonstrated by inhibition tests with the appropriate mAb. The expression of CD80 and CD86 by anti‐μ‐treated SE B cells was not, however, the sole explanation for their good antigen presenting capacities since the exposure of FM B cells to anti‐μ antibody also induced expression of these surface structures. Nevertheless, these cells failed to become good MLR stimulators. Collectively, the above data contribute further to the characterization of a distinct subset of tonsillar B cells which resemble, both phenotypically and functionally, the B cells of the splenic marginal zone.
Modulation of integrin‐mediated intercellular adhesion during the interaction of thymocytes with stromal cells expressing VLA‐4 and LFA‐1 ligands
Authors: St‐Pierre Yves , Hugo Patrice , Legault Doris , Tremblay Pierre , Potworowski Edouard F.
Published: March 2007

Mature peripheral T cells closely regulate their intercellular interactions by modulating integrin adhesion functions. The ability of members of the integrin family to mediate intercellular adhesion is dependent on signals from within the cells (inside‐out signaling) that increase the avidity of integrins for their ligands. These changes in avidity are independent of the quantitative changes on the number of receptors, and there is evidence to suggest that phosphorylation events play a predominant role in the regulation of the avidity state of the integrins. Whether such regulatory mechanisms are operative during T cell development had hitherto been an opened question. In the present work, we have used an in vitro adhesion assay between thymocytes and target cells expressing VLA‐4 and LFA‐1 counter ligands to determine how thymocytes can discriminate between integrin‐specific signals during T cell development. Our findings are that VLA‐4, but not LFA‐1, is constitutively expressed in its high‐avidity state during the early stages of T cell development, and that the high‐avidity state of thymocytes for VCAM‐1‐expressing cells is closely regulated by signaling through protein kinase C and protein tyrosine kinase pathways. At later stages of development, mature thymocytes prior to leaving the thymus turn off both VLA‐4 and LFA‐1 adhesion functions. Our results show that the low‐affinity state of integrins on peripheral mature T cells is established before mature thymocytes leave the thymus. Only when mature T cells recognize antigenic peptides in the context of major histocompatibility complex in the periphery will they turn on the adhesion function of VLA‐4 and/or LFA‐1 integrins.
The p56lck SH2 domain mediates recruitment of CD8/p56lck to the activated T cell receptor/CD3/ζ complex
Authors: Thome Margot , Germain Valérie , Disanto James P. , Acuto Oreste
Published: March 2007

The CD4 or CD8 co‐receptors and the T cell receptor (TCR) are thought to interact with the same antigen‐presenting major histocompatibility complex molecule in a stable ternary complex. Therefore, the TCR and its co‐receptor need to come into close proximity on the surface of the T cell. We have previously shown that the interaction of the p56lck SH2 domain with ζ‐associated, tyrosine phosphorylated ZAP‐70 and Syk kinases leads to an enhanced association of CD4 with TCR/CD3/ζ complex after CD3 stimulation of Jurkat cells. In this report, we analyzed whether a similar mechanism can mediate recruitment of the CD8αβ and CD8αβ isoforms to the TCR. We demonstrate in vivo in association of CD8αα/p56lck with the tyrosine kinase ZAP‐70 after CD3 stimulation of Jurkat cells. A phosphopeptide competing in vitro for the binding of tyrosine phosphorylated proteins to the SH2 domain of p56lck specifically impedes the association of ZAP‐70 with CD8αα/p56lck without affecting the ζ/ZAP‐70 interaction. The same peptide is able to compete for the activation‐dependent association of the CD8αα or CD8αβ isoform with the TCR/CD3/ζ complex. Moreover, co‐precipitation of the TCR with both CD8 isoforms was observed after CD3 stimulation. These findings strongly suggest that the p56lck SH2 domain mediates recruitment of CD8/p56lck to the activated TCR/CD3/ζ complex.
Difference in antigen presentation pathways between cortical and medullary thymic epithelial cells
Authors: Kasai Michiyuki , Hirokawa Katsuiku , Kajino Ki‐Ichi , Ogasawara Kazumasa , Tatsumi Masashi , Hermel Evan , Monaco John J. , Mizuochi Toshiaki
Published: March 2007

Antigen presentation by thymic epithelial cells (TEC) to T cells that undergo maturation is one of the major events in the selection of the T cell repertoire. We have already reported that medullary TEC lines (mTEC) established from newborn C57BL/6 (H‐2b) mice are able to present a soluble antigen, ovalbumin (OVA), to OVA‐specific, I‐Ab restricted helper T cell lines but cortical TEC (cTEC) lines are not (Mizuochi, T. et al., J. Exp. Med. 1992. 175: 1601). In this report, to clarify the cause of this difference, we analyzed the biochemical nature as well as the distribution of both major histocompatibility complex (MHC) class II molecules and invariant chains (Ii) in both TEC by immunoprecipitation and laser confocal scanning microscopic analysis, as well as the expression of mRNA encoding H‐2Ma or H‐2Mb. Our results demonstrate that cTEC and mTEC are both able to present peptide antigens to peptide‐specific, I‐Ab‐restricted helper T cell hybridoma and are able to present class II MHC alloantigens to an I‐Ab‐specific T cell line, that mRNA for H‐2Ma and H‐2Mb are expressed in both TEC, that cTEC and mTEC apparently incorporate tetramethylrhodamine isothiocyanate‐labeled OVA in the same manner, and that the SDS‐stable MHC class II molecules, onto which peptides were loaded, are formed in both cTEC and mTEC. However, these molecules were more rapidly degraded in mTEC than in cTEC. In addition, two Ii‐derived polypeptides of approximately 21 kDa and 10 kDa were precipitated by the anti‐class II monoclonal antibody Y3P; 10‐kDa polypeptides were detected in the both TEC, while 21‐kDa polypeptides were detected only in cTEC. Finally, β chains of MHC class II with less sialylated oligosaccharides were precipitated from the cell surface of cTEC. Taken together, these results suggest that there are substantial differences in the antigen‐presenting pathways of cTEC and mTEC, and these difference might be responsible for T cell selection events in the thymus.
Orally induced, peptide‐specific γ/δ TCR+ cells suppress experimental autoimmune uveitis
Authors: Wildner Gerhild , Hünig Thomas , Thurau Stephan R.
Published: March 2007

We investigated the role of γ/δ TCR+ T cells in induction and suppression of the T cell‐mediated disease experimental autoimmune uveitis. Disease induction was studied in Lewis rats perinatally depleted of α/β or γ/δ TCR+ subpopulations. Depletion of α/δ TCR+ cells completely abrogated disease, whereas treatment with anti‐γ/δ antibodies had no influence on onset or intensity of uveitis. However, adoptively transferred γ/δ+ cells from orally tolerized rats could mediate suppression of uveitis in an antigen‐specific fashion. Uveitis induced by a peptide derived from the uveitogenic retinal soluble antigen (S‐Ag) was suppressed by γ/δ+ cells from rats orally tolerized with the same peptide as well as HLA peptide B27PD. This disease ameliorating effect could also be observed when rats were fed with the HLA peptide before immunization with S‐Ag peptide. Transfer of α/β+ T cells from the same donors as well as γ/δ+ or α/β+ cells from animals fed with control peptide had no ameliorating effect.
Cell surface expression of surrogate light chain (ΨL) in the absence of μ on human pro‐B cell lines and normal pro‐B cells
Authors: Meffre Eric , Fougereau Michel , Argenson Jean‐Noël , Aubaniac Jean‐Manuel , Schiff Claudine
Published: March 2007

Surrogate light chains (ΨL) encoded by λ‐like (λ5) and VpreB genes play a critical role in controlling the early steps of B cell differentiation. We prepared new anti‐VpreB monoclonal antibodies (mAb) (3C7/6F6) which preferentially recognize the VpreB epitope at the cell surface of human cell lines that do not express the μ chain. These mAb provide the first characterization of human pro‐B cell lines expressing surface ΨL. We demonstrate that surface ΨL expression is considerably enhanced upon interleukin‐7 stimulation and that the ΨL complex is formed independently of the Igα/Igβ heterodimer. Finally, using these antibodies, we confirm the existence of a normal pro‐B cell population in human adult bone marrow. These cells are CD34+ CD38+ ΨL+, do or do not express CD19, CD10, or both epitopes, and may represent the earliest cell population committed to B cell differentiation.
The fate of adoptively transferred antigen‐specific T cells in vivo
Authors: Zhang Li
Published: March 2007

In this study, anti‐major histocompatibility complex class I Ld T cell receptor (TCR)‐transgenic cells were adoptively transferred into severe‐combined immunodeficient (Scid) mice which express the Ld Ag on all nucleated cells, and the fate of transferred antigen‐specific T cells (ASTC) was followed in vivo. It was found that, after encountering antigen (Ag) in vivo, the number of ASTC increased 10–15‐fold followed by a decline in number to a value that was still above the starting value. The expansion of ASTC could be abrogated by blocking CD28 on the T cells prior to Ag stimulation. Using the technique which simultaneously stains ASTC surface markers and apoptotic cells, we have not only demonstrated directly that the ASTC that disappeared from the periphery died by activation‐induced apoptosis but also studied their kinetics in vivo. The remaining ASTC moderately down‐regulated both TCR and CD8 on their cell surface and were fully unresponsive when cultured with Ld+ cells even in the presence of exogenous interleukin (IL)‐2 and IL‐4. However, they were still susceptible to apoptosis when transfereed into a secondary host that provided a new source of Ag and antigen‐presenting cells. These studies indicate that peripheral T cell tolerance can be induced by multiple mechanisms in which activation‐induced ASTC apoptosis plays an important role.
Induction of sensitivity to activation‐induced death in primary CD4+ cells: A role for interleukin‐2 in the negative regulation of responses by mature CD4+ T cells
Authors: Wang Ruduan , Rogers Amy M. , Rush Brian J. , Russell John H.
Published: March 2007

We examine the requirements for converting naive, mature CD4+ cells from an activation‐induced death (AID)‐resistant to a ‐sensitive phenotype. Priming for sensitivity to AID can be divided into two steps. The first is a mitogen/CD3‐dependent, cyclosporin‐sensitive signal and the second a cytokine‐dependent, cyclosporin‐insensitive one. Under these conditions, interleukin (IL)‐2, but not IL‐4, IL‐7 or IL‐15, the receptors of which share a common receptor γ chain, is capable of providing the cytokine signal for inducing sensitivity to AID. Increased expression of the low‐affinity IL‐2Rα chain (CD25) is associated with acquisition of AID sensitivity and antibodies to CD25 block acquisition of AID sensitivity in the presence of IL‐2. As with T cell hybridomas, AID is dependent on both CD95 and CD95 ligand (CD95L) expression, but unlike hybridomas, the sensitive and resistant phenotypes of primary CD4+ cells cannot be distinguished by levels of CD95 expression, functional CD95L nor the fraction of cells in cycle. The results suggest that the unique function of IL‐2 is to regulate proteins, either not important or constitutively regulated in T cell hybridomas, that are essential for cell‐autonomous suicide of activated CD4+ cells. These experiments provide a mechanism for the recent observations of chronic lymphoproliferation and autoimmune disease in mice with null mutations in IL‐2 or CD25.
Human B7‐1 is more efficient than B7‐2 in providing co‐stimulation for alloantigen‐specific T cells
Authors: van Dijk Astrid M. C. , Otten Henny G. , Vercauteren Suzanne M. , Kessler Floortje L. , de Boer Mark , Yerdonck Leo F. , de Gast Gijsbert C.
Published: March 2007

Besides a signal via the T cell receptor/CD3 complex, an additional co‐stimulatory signal is required for optimal T cell activation. This signal can be delivered by interaction of either B7‐1 or B7‐2 expressed by antigen‐presenting cells with CD28 on the T cells. Comparison of the function of B7‐1 and B7‐2 in different experimental animal systems generated conflicting data on the roles for the co‐stimulatory molecules. We therefore investigated whether there are differences between B7‐1 and B7‐2‐mediated co‐stimulation in an alloantigen‐specific primary T cell response induced by B7‐transfected human cell lines of epithelial origin. Both transfected keratinocyte cell lines efficiently induce T cell proliferation and the ratios of stimulator versus responder cells are similar. The kinetics of proliferation and interleukin (IL)‐2, IL‐4 and interferon‐γ production are also comparable between both transfectant lines. However, despite equal B7 expression levels, it is consistently found that the magnitude of the B7‐1‐induced T cell proliferation was higher than that of B7‐2. Comparison of precursor frequencies of helper T lymphocytes responsive with either B7‐1 or B7‐2 revealed that the frequency of B7‐1‐responsive T cells was higher than that of B7‐2, and that the frequency of cells activated by a combination of B7‐1 and B7‐2 did not differ significantly from that of B7‐1 alone. We therefore conclude that the B7‐2‐responsive T cells are part of the B7‐1‐responsive population, and that B7‐1 on keratinocytes is more efficient in providing co‐stimulation for alloantigen‐specific T cells.
Prevention and amelioration of collagen‐induced arthritis by blockade of the CD28 co‐stimulatory pathway: requirement for both B7‐1 and B7‐2
Authors: Webb Louise M. C. , Walmsley Marita J. , Feldmann Marc
Published: March 2007

Collagen type II‐induced arthritis (CIA) is an experimental model of arthritis that has been successfully used to dissect the pathogenesis of human rheumatoid arthritis and to identify potential therapeutic targets. We have used this model to evaluate the role of T cell co‐stimulation in both disease development and progression. T cell co‐stimulation is provided by ligation of CD28 with either B7‐1 or B7‐2 present on antigen‐presenting cells and can be prevented by a soluble form of CTLA‐4 (CTLA‐4Ig) which binds with high affinity to both B7‐1 and B7‐2. We found that administration of CTLA‐4Ig at the time of immunization prevented the development of CIA and was associated with lack of lymphocyte expansion within the draining lymph node and failure to produce anti‐collagen IgG1 or IgG2a antibodies. To determine which CD28 ligand plays a more dominant role in CIA, we treated mice with monoclonal antibodies (mAb) against either B7‐1 or B7‐2. Neither anti‐B7‐1 nor anti‐B7‐2 had any effect on the course of CIA when given alone, but resulted in reduced incidence and clinical scores when given together. Interestingly, when treatment was delayed until after the onset of clinical disease, both CTLA‐4Ig or anti‐B7‐1 plus anti‐B7‐2 mAb still ameliorated disease. Effective treatment was associated with a reduction in interferon‐γ production by lymph node cells following stimulation in vitro, suggesting that Th1 responses were diminished. This study points to a critical role of CD28 co‐stimulation in the development and perpetuation of CIA in DBA/1 mice. Interestingly, it demonstrates an active role for T cells in the later stages of this disease and implicates both B7‐1 and B7‐2‐mediated co‐stimulation in the pathogenesis of CIA.
A novel nucleotide‐containing antigen for human blood γδ T lymphocytes
Authors: Poquet Yannick , Constant Patricia , Halary Franck , Peyrat Marie‐Alix , Gilleron Martine , Davodeau François , Bonneville Marc , Fournié Jean‐Jacques
Published: March 2007

The stimulation of human γδ T cells by mycobacteria occurs through recognition of four distinct nonpeptide phosphorylated antigens termed TUBag1–4. Among these latter, TUBag4 has already been biochemically characterized as a γ‐X derivative of 5′‐deoxythymidine triphosphate (Constant, P., Davodeau, F., Peyrat, M. A., Poquet, Y., Puzo, G., Bonneville, M. and Fournié, J.‐J., Science 1994. 264: 267). However, despite chemical synthesis of weakly stimulatory nucleotide‐containing analogs, these mycobacterial compounds remained the sole nucleotide‐containing antigens actually isolated from natural sources. Here, we present the complete isolation of the TUBag3 antigen from Mycobacterium fortuitum and demonstrate that this nonpeptide molecule contains a 5′‐UTP nucleotide moiety. On selected Vγ9/Vδ2 clones, T cell responses can be triggered with nanomolar concentrations of TUBag3. Like crude mycobacterial extracts, this purified nucleotide conjugate elicits a strong polyclonal response of γδ PBL from healthy donors. Furthermore, we present evidence that this compound is distinct from the recently synthesized γ‐isopentenyl 5′‐UTP, a nucleotide conjugate of isopentenyl pyrophosphate that was found to be stimulatory for human γδ T cells (Tanaka, Y., Morita, C. T., Tanaka, Y., Nieves, E., Brenner, M. B. and Bloom, B. R., Nature 1995. 375: 155). Since it appears that both mycobacterial nucleotide antigens are molecules structurally related to peculiar precursors of nucleic acid synthesis, we propose that TUBag‐reactive T cells might be specifically devoted to surveillance of proliferating cells.
Raf regulates positive selection
Authors: O'Shea Clodagh C. , Crompton Tessa , Rosewell Ian R. , Hayday Adrian C. , Owen Michael J.
Published: March 2007

T cell development is regulated by extracellular signals that mediate cellular proliferation and differentiation via specific signal transduction pathways. To determine the importance of the mitogen‐activated protein kinase (MAP kinase) pathway in thymocyte development, we analyzed transgenic mice expressing dominant negative Raf (DN Raf) and a constitutively active v‐Raf under the control of the p56lck proximal promoter. DN Raf had a profound effect on T cell receptor (TCR)‐mediated signaling events as assessed by the inhibition of mitogen‐induced proliferation of thymocytes in vitro. Overall thymocyte numbers were decreased by at most twofold from nontransgenic littermates. Positive selection was inhibited in DN Raf transgenic mice, as evidenced by both reduced numbers of mature thymocytes and a decrease in CD8+ thymocytes in female mice doubly transgenic for DN‐Raf and a class I‐restricted H‐Y TCR. In contrast, the differentiation of double‐positive thymocytes to single‐positive thymocytes was enhanced in H‐YTCR transgenic mice expressing constitutively active Raf (v‐Raf). Thus, Raf regulates positive selection in the thymus.
Identification and cellular distribution of the rat interleukin‐2 receptor β chain: induction of the IL‐2Rα−β+ phenotype by major histocompatibility complex class I recognition during T cell development in vivo and by T cell receptor stimulation of CD4+8+ immature thymocytes in vitro
Authors: Park Jung‐Hyun , Hanke Thomas , Hünig Thomas
Published: March 2007

A mouse monoclonal antibody (mAb) to the rat interleukin‐2 receptor β (IL‐2Rβ) chain was generated using IL‐2Rβ cDNA‐transfected mouse L929 cells for immunization and differential screening. This antibody, called L316, detects a cell surface protein with an apparent molecular mass of about 80 kDa. In peripheral lymphoid organs of young adult rats, IL‐2Rβ expression is restricted to T and natural killer (NK) cells, and less than 10% of IL‐2Rβ+ cells co‐express the IL‐2Rα chain. IL‐2Rβ was detected on all NKRP‐1hi (NK−) and NKRP‐1lo cells (T‐lineage cells of unknown function), most peripheral γδ T cells and on 30–40% of CD8 and 10% of CD4 αβ T cells. In the adult rat thymus, mAb L316 detects a small subset (about 1%) of predominantly IL‐2Rα− cells which express cell surface markers characteristic of mature T lymphocytes and contain a high proportion of CD4−8− and CD4−8+ αβ T cell receptor (TCR)+ thymocytes. TCR‐V usage suggests that major histocompatibility complex (MHC) class I plays a more important role than MHC class II in the selection of these cells. On immature CD4+8+ rat thymocytes, IL‐2Rβ cell surface expression is readily induced by TCR stimulation in vitro, supporting the idea that in vivo, the IL‐2Rβ+ phenotype is the result of TCR engagement during thymic selection.
Th1/Th2 cytokine gene expression after mercuric chloride in susceptible and resistant rat strains
Authors: Gillespie Kathleen M. , Saoudi Abdelhadi , Kuhn Joëlle , Whittle Catheryne J. , Druet Philippe , Bellon Blanche , Mathieson Peter W.
Published: March 2007

Mercuric chloride (HgCl2) has contrasting effects on different rat strains: susceptible strains, e.g. Brown Norway (BN) develop polyclonal B cell activation, multiple autoantibodies and widespread tissue injury. Lewis (LEW) rats are resistant: no autoimmune response occurs after HgCl2; instead, there is immunosuppression. We have previously shown, by fully quantitative polymerase chain reaction (PCR), up‐regulation of interleukin‐4 (IL‐4) gene expression in HgCl2‐treated BN rats, implicating Th2 cells in the autoimmune syndrome. Involvement of the reciprocal Th1 subset, producing interferon‐γ (IFN‐γ), in resistance of LEW rats to HgCl2 has been suggested. We now report extensive analysis of Th1 and Th2 cytokine gene expression in spleen and lymph nodes of susceptible (BN) and resistant (LEW) rats after HgCl2. IL‐4 and IFN‐γ were analyzed by quantitative PCR, other cytokines were assessed using semiquantitative PCR: the relative merits of these two techniques are discussed. We show pronounced up‐regulation of IL‐4 and more modest up‐regulation of IFN‐γ in BN rats, but no up‐regulation of either in LEW rats. Baseline levels of IFN‐γ were higher in LEW rats. Semi‐quantitative PCR showed increased expression of IL‐2, IL‐6 and IL‐10 in BN; in LEW rats only IL‐10 was increased. There was no marked change in IL‐5, IL‐13 or transforming growth factor‐β (TGF‐β) in either strain. These data further support the key role of IL‐4 in HgCl2‐induced autoimmunity, and suggest that failure of up‐regulation of IL‐4, together with higher baseline IFN‐γ expression, accounts for resistance of LEW rats to HgCl2. However, neither IFN‐γ nor TGF‐β can be implicated in HgCl2‐induced immunosuppression in the LEW rat in vivo: our data suggest a role for IL‐10 in this phenomenon.
Phenotype, and migration properties of three major subsets of tissue homing T cells in sheep
Authors: Mackay Charles R. , Andrew David P. , Briskin Michael , Ringler Douglas J. , Butcher Eugene C.
Published: March 2007

T cells show a bias in their migration pathways: some T cells preferentially migrate to peripheral lymph nodes (LN), some to mucosal tissues, and some to peripheral tissues such as skin. These recirculation pathways were examined in sheep by collecting lymph draining into and out of peripheral and intestinal LN, and using fluorescent dyes to trace the recirculation of the lymph cells. Monoclonal antibodies (mAb) to α4, β1, and β7 integrins, and L‐selectin, were used to define three major populations of recirculating T cells. Naive‐type T cells (L‐selectin+, α4β1lo β7lo) migrated preferentially through peripheral LN. Two memory populations could be defined: α4β1hi β7− and α4β7hiβ1lo. α4β1hi β7− T cells were present in lymph draining from the skin. T cells migrating preferentially through intestinal LN were α4β7hi β1lo. Consistent with this migration pattern, the endothelial receptor for α4β7, mucosal addressin cell adhesion molecule‐1 (MAdCAM‐1) was detected on high endothelial venules within intestinal LN and Peyer's patches, but only weakly on high endothelial venules within peripheral LN. Thus, there are at least three easily definable subsets of T cells, based on integrin expression, which show distinct migration preferences.
Comparison of rheumatoid factors of rheumatoid arthritis patients, of individuals with mycobacterial infections and of normal controls: evidence for maturation in the absence of an autoimmune response
Authors: Djavad Nargues , Bas Sylvette , Shi Xiaowen , Schwager Joseph , Jeannet Michel , Vischer Thomas , Roosnek Eddy
Published: March 2007

We analyzed the rheumatoid factors (RF) produced by Epstein‐Barr virus‐transformed monoclonal B cells established from four patients with rheumatoid arthritis (RA), three individuals with a history of Mycobacterium tuberculosis (TB) and four normal controls (NI). Fifty‐eight RF were analyzed for specific activity (international units‐RF/μg) for the Fc part of IgG and their interaction with tetanus toxoid (TT) and DNA (polyspecificity). Furthermore, we sequenced the V‐D‐J heavy chain region of 16 (9TB‐/7RA‐) RF. Significant differences were observed between the NI‐RF and the TB‐ and RA‐RF. While the RF repertoire of normal individuals comprised of low‐avidity RF of which the majority (15/17) were polyspecific, more than half of the TB‐ and RA‐RF were monoreactive. Furthermore, the monospecific TB‐ and RA‐RF were of significantly higher avidity than the NI‐RF (RA > TB ≫ NI). With respect to poly‐specificity, the RF in the three groups were comparable: the interaction with DNA, TT as well as with Fc was inhibited either by an increase of the ionic strength to 0.3–0.5 M NaCl or by addition of the polyanion dextran sulfate, indicating that the antibodies interacted with similar anionic epitopes shared by the three antigens. Analysis of the V‐D‐J heavy chain regions showed significant differences between the respective RF. The salt‐sensitive binding was highly correlated with the presence of arginine in the complementarity‐determining region 3 (CDR3). Furthermore, whereas the polyspecific RF consisted predominantly of germ‐line encoded antibodies, the genes of the monospecific RA/TB‐RF were somatically mutated (RA > TB). It is therefore likely that maturation of RF can be initiated by chronic infections and that monospecific, somatically mutated RF are not a unique characteristic of autoimmune diseases.