bluuebunny/crossref_metadata_2025 huggingface dataset:
Isotype switching in anti‐immunoglobulin‐activated B lymphoblasts: differential requirements for interleukin 4 and other lymphokines to elicit membrane vs. secreted IgG1
Authors: Purkerson Jeffrey M. , Isakson Peter C.
Published: March 2007

Anti‐immunoglobulin (Ig)‐activated B lymphoblasts, prepared by culturing high‐density B cells with anti‐Ig‐Sepharose for 48 h, can be induced to secrete IgM and IgG1 by a mixture of T cell‐derived lymphokines containing interleukin (IL) 4. In this study we have examined the conditions required for lymphokine‐mediated induction of IgG1 secretion and membrane (m)IgG1 expression in B lymphoblasts. Resting B cells exposed to IL 4 (10–100 U/ml) during anti‐Ig‐mediated blast transformation did not secrete IgG1 upon subsequent culture with lipopolysaccharide (LPS) regardless of whether IL 4 was present or absent during the secondary culture. In contrast, B lymphoblasts previously exposed to IL 4 did secrete IgG1 in response to T cell‐derived lymphokines [EL 4 supernatant depleted of IL 4; (D)EL 4 SN]. However, optimal IgG1 secretion was obtained when B lymphoblasts were simultaneously exposed to IL 4 and other lymphokines. Pre‐exposure to (D)EL 4 SN, which contains IL 5 and IL 2, failed to prepare anti‐Ig blasts to secrete IgG1 in response to LPS and IL 4. Inhibition of IL 5 and IL 2 activity in EL 4 SN suppressed IL 4‐mediated IgG1 secretion. Together, these data indicate that B lymphoblasts require IL 5 and IL 2 in addition to IL 4 to secrete IgG1, and that the IL 4 signal(s) must precede or accompany those provided by the other lymphokines. As a measure of the fraction of cells capable of switching to IgG1, we assessed expression of mIgG1 on B lymphoblasts by fluorescence flow cytometry. B lymphoblasts cultured for 3 days with (D)EL 4 SN and IL 4 (10–100 U/ml) were 8% to 20% mIgG1+; in the absence of IL 4 blasts did not express detectable mIgG1. Although anti‐Ig blasts treated with LPS and IL 4 did not secrete appreciable IgG1, a substantial fraction of B lymphoblasts (4%–19%) cultured with LPS and IL 4, but not LPS alone, expressed mIgG1. These results suggest that LPS and IL 4 are sufficient to commit B lymphoblasts to mIgG1 expression, but that additional signals provided by T cell‐derived lymphokines are required to elicit IgG1 secretion.
Isoforms of the E2 molecule: D44 monoclonal antibody defines an epitope on E2 and reacts differentially with T cell subsets
Authors: Gelin Catherine , Zoccola Didier , Valentin Hélène , Raynal Brigitte , Bernard Alain
Published: March 2007

Human T cell rosetting with erythrocytes is clearly dependent on the CD2‐CD58 interaction. We have previously demonstrated that other T cell molecules are involved in the rosette phenomenon, including the E2 molecule, a 32‐kDa transmembrane glycoprotein. In this report we show that the D44 monoclonal antibody (mAb), previously shown to subdivide T cells into subpopulations with distinct functional repertoires and to identify 70% of lymphocytes from bronchoalveolar lavage from HIV+ patients, defined a new epitope on the E2 molecule. This was illustrated by the reactivity of the D44 mAb in Western blot experiments performed with the immunoaffinity purified E2 molecule. Moreover, double‐labeling experiments showed that the E2 molecule exhibited varying epitopes when expressed in different cell types. The D44 and 12E7 epitopes were restricted to distinct subpopulations of T cells and, more importantly, the D44 expression was limited to the CD29+ population including the memory subset. The O662 and L129 epitopes are present on all T cells. Thus, the E2 molecule has both common and variable epitopes in its extracellular domain, and only the common epitopes seem to be involved in T cell adhesion processes.
A lectin‐binding soluble factor released by CD8+CD57+ lymphocytes from AIDS patients inhibits T cell cytotoxicity
Authors: Sadat‐Sowti Behazine , Debré Patrice , Guillon Jean‐Marcel , Hadida Fabienne , Autran Brigitte , Idziorek Thierry , Okzenhendler Eric , Katlama Christine , Mayaud Charles
Published: March 2007

CD8+CD57+ T cells, expanded in peripheral blood lymphocytes of AIDS patients, inhibit the effector phase of HLA‐specific cytotoxic T lymphocytes, natural killer and lymphocyte‐activated killer cells in a 4‐h chromium‐release assay. This inhibitory activity present in supernatants of purified sorted CD8+CD57+ cells is mediated by a non‐antigen‐specific inhibitory factor which is distinct from prostaglandin E2, T cell growth factor (TGF)‐β, latent‐TGF‐β, tumor necrosis factor (TNF)‐α and TNF‐β. Partial biochemical characterization demonstrates that the CD8+CD57+ inhibitory activity (a) is heat, trypsin and acid resistant, (b) binds to concanavalin A columns, indicating its glycosylation state and (c) is mediated by a 20–30‐kDa soluble molecule.
Cytosolic targeting of hen egg lysozyme gives rise to a short‐lived protein presented by class I but not class II major histocompatibility complex molecules
Authors: Calin‐Laurens Veronique , Forquet Frédérique , Gerlier Denis , Rabourdin‐Combe Chantal , Mottez Estelle , Kanellopoulos Jean , Godeau François , Kourilsky Philippe
Published: March 2007

A way to study the role of intracellular trafficking of an antigen in its presentation to T cells is to target the antigen to various cell compartments of the antigen‐presenting cells (APC) and compare the nature of the complexes associating major histocompatibility complex (MHC) molecules and antigenic peptides, expressed on the cell surface. MHC class I+ and MHC class II+ mouse L fibroblasts secreting hen egg lysozyme (HELs cells) or expressing HEL in their cytosol (HELc cells) were obtained after transfection with HEL cDNA and signal sequence‐deleted HEL cDNA, respectively. HEL was evidenced in both HELs‐and HELc‐transfected cells and the former type of transfectant secreted a large amount of HEL. However, HEL produced in the cytosol exhibited a short half‐life of less than 5 min. HEL‐derived peptides could not be shown biochemically either in HELc‐ nor in HELs‐transfected cells. We then studied the capacity of these cells to present HEL to HEL‐specific class I‐ and class II‐restricted T cells. Both cell types could be recognized by the HEL‐specific MHC class I‐restricted CTL clones. In contrast, MHC class II‐HEL peptide complexes, recognized by HEL‐specific helper T cell hybridomas, could be detected on MHC class II+ HELs‐ but not HELc‐transfected cells. In vivo experiments showed, however, that HELc‐transfected cells could provide host APC with HELc‐derived peptides able to associate with MHC class II molecules. This was inferred from the capacity of MHC class II− HELc‐transfected cells, unable by themselves to elicit any anti‐HEL antibody response, to prime syngeneic and allogeneic mice against HEL. The priming was revealed by the induction of an antibody response after a boost with an amount of HEL unable itself to elicit an antibody response.
Human complement factor H: two factor H proteins are derived from alternatively spliced transcripts
Authors: Estaller Cornelia , Weiss Elisabeth H. , Schwaeble Wilhelm , Dierich Manfred
Published: March 2007

The human complement factor H is an important component in the control of the alternative pathway of complement activation. We have previously shown that at least three factor H homologous mRNA species of 4.3 kb, 1.8 kb and 1.4 kb in length are constitutively expressed in human liver. In addition, several factor H‐related proteins have been detected in human sera using antibodies directed against the classical human factor H glycoprotein of 150 kDa. The structure of the additional polypeptides has not been shown so far. Circumstantial evidence suggests that the 1.8‐kb mRNA might encode the 43‐kDa factor H‐like polypeptide. Here we report the isolation, characterization and eukaryotic expression of the first full‐length cDNA representing the major 4.3‐kb mRNA and the 1.8‐kb mRNA of human factor H. We show that the 4.3‐kb transcript encodes the 150‐kDa‐factor H glycoprotein and the 1.8‐kb mRNA the 43‐kDa factor H polypeptide. The identity of the two cDNA in a region of 1400 nucleotides suggests that the two factor H‐related transcripts are derived from one gene by a process of alternative splicing.
A transcriptional enhancer of the mouse T cell receptor δ gene locus
Authors: Gill Laura Lee , Karjalainen Klaus , Zaninetta Daniela
Published: March 2007

T cells express receptors for antigen on their surface (TcR) which consist either of α/β or of γ/δ polypeptide chains. Since the TcR δ chain gene is located within the TcR α chain gene locus, strict regulation of expression of the region must operate to ensure that the two loci are not concomitantly expressed in T cells bearing either α/β or γ/δ TcR. In this report it is demonstrated that elements within the mouse TcR δ gene locus, located between the Jδ2 and Cδ exons, enhance transcription from a heterologous promoter five‐ to tenfold in a T cell hybridoma expressing a TcR γ/δ, whereas enhancement was only twofold in an α/β‐bearing T cell hybridoma. No enhancement of expression was observed in a B cell hybridoma. The sequences responsible for this enhancing activity are largely confined to a 766‐bp Hae III DNA restriction fragment. A region within this DNA segment shows significant homology (73% identify) to a recently identified enhancer of the human TcR δ gene locus.
Evidence for a restricted idiotypic and epitopic specificity of anti‐thyroglobulin autoantibodies in patients with autoimmune thyroiditis
Authors: Dietrich Gilles , Kazatchkine Michel D. , Piechaczyk Martine , Pau Bernard
Published: March 2007

Anti‐thyroglobulin (TG) autoantibodies from patients with autoimmune thyroiditis express a cross‐reactive α idiotype (Id) termed T44 which is not expressed by IgG from normal individuals. The present study demonstrates that the expression of the T44 Id is strongly associated with the recognition by anti‐TG autoantibodies of a specific epitopic cluster on human TG. The epitopic reactivity of anti‐TG autoantibodies was determined using a competitive inhibition assay with a panel of 15 monoclonal antibodies that define six antigenic clusters on TG. All T44+ autoantibodies from patients recognized cluster II, whereas no anti‐TG IgG from healthy individuals reacted with this region. Affinity columns of Sepharose‐bound intravenous therapeutic immunoglobulins which contain anti‐T44 activity, retained both T44 Id‐expressing antibodies and a subset of region II‐specific anti‐TG autoantibodies from patients with Hashimoto's disease. Restricted idiotypic and epitopic specificity may demarcate disease‐associated from natural anti‐TG autoantibodies, suggesting that qualitative rather than quantitative criteria should be used to identify pathological autoantibodies.
Biased expression of individual T cell receptor V gene segments in CD4+ and CD8+ human peripheral blood T lymphocytes
Authors: Grunewald Johan , Janson Carl Harald , Wigzell Hans
Published: March 2007

The expression of seven different α and β gene segments of the T cell receptor on normal human CD4+ and CD8+ T lymphocytes, respectively, was examined using V gene‐specific monoclonal antibodies. We found a statistically significant (p < 0.001) bias of the expression of four V gene products towards the CD4+ subpopulation. In every individual analyzed, the Vβ5.1 gene segment was expressed to a higher degree among CD4+ compared to CD8+ cells, with a median value of 4.8% among CD4+ cells and 1.5% among CD8+ cells. There was also a statistically significant skewness in the usage of the Vβ6.7, Vβ8 and Vβ12 gene segments towards the CD4+ T cell population, but not as dominating as for the Vβ5.1. Lymphocytes from umbilical cord blood showed similar skewed reactivities for the Vβ5.1, Vβ6.7 and Vβ12‐specific monoclonal antibodies. Aspects of positive and negative selection, as possible explanations for these findings, are discussed.
Expression of lineage‐restricted protein tyrosine kinase genes in human natural killer cells
Authors: Biondi Andrea , Rossi Vincenzo , Benvestito Serena , Paganin Carla , Allavena Paola , Mantovani Alberto , Perlmutter Roger M.
Published: March 2007

The hematopoietic lineage derivation, recognition structures and associated signal transduction pathways of CD3− natural killer (NK) cells have not been identified. Protein tyrosine kinases (PTK) structurally related to the product of the c‐src protooncogene are differentially expressed in distinct hematopoietic differentiation lineages and may participate in specific signal transduction pathways.

The present study was aimed at characterizing the expression of src‐related PTK genes in normal human NK cells and in cells from patients with CD3− granular lymphocyte proliferative disease. CD3− normal NK cells had high levels of transcripts of the lck gene, which is highly expressed in T cells. CD8+ and CD8− NK cells expressed similarly high levels of Ick mRNA.

In contrast, NK cells expressed very low levels (25‐80 times less than monocytes) of mRNA encoding the myelomonocytic PTK hck.

NK cells also expressed fyn transcripts (p59fyn reportedly associates with the T cell receptor in T cells) and fgr transcripts, the latter observation confirming a previous report. The pattern of expression of the lineage‐restricted PTKs lck and hck in NK cells is consistent with the hypothesis of an ontogenic relationship of this population with the lymphocytic rather than myelocytic differentiation pathway. PTK expressed in NK cells may participate in signal transduction pathways in this cell population.
Dominant expression of the T cell receptor BALB invariant δ (BID) chain in resident pulmonary lymphocytes is due to selection
Authors: Sim Gek‐Kee , Augustin Andrei
Published: March 2007

During ontogeny, the T cell receptor BALB invariant δ (BID) chain (Sim, G.‐K. and Augustin, A., Cell 1990. 61: 397) is generated in C57BL/6 fetal thymocytes at levels similar to those detected in BALB/c mice. Hence, the dominance of BID observed among resident pulmonary lymphocytes of BALB/c and of (BALB/c x C57BL/6) F1 and its absence in C57BL/6 mice is due to positive selection and peripheral expansion rather than to the inability of C57BL/6 mice to generate this particular rearrangement.
Anti‐HLA antigen class I heavy chain monoclonal antibodies inhibit human immunodeficiency virus production by peripheral blood mononuclear cells
Authors: Corbeau Pierre , Olive Daniel , Devaux Christian
Published: March 2007

We have examined the capacity of monoclonal antibodies (mAb) specific for HLA class I heavy chain to interfere with the human immunodeficiency virus (HIV) replicative cycle in human T cells. Among six anti‐HLA class I heavy chain‐specific mAb assayed, two mAb, RL4‐24‐6 and W6/32, were able to delay HIV1 and HIV2 cytopathic effect on MT4 cells, a human T cell leukemia virus type I (HTLVI) immortalized T cell line. mAb RL4‐24‐6, chosen for further studies, also inhibited HIV1 production by peripheral blood mononuclear cells (PBMC), and this inhibition was dose dependent. However, no effect was observed when mAb treatment was performed with either the CEM or Jurkat T cell lines. Our investigation of how RL4‐24‐6 interferes with the HIV replicative cycle revealed that: (a) incubation of PBMC with RL4‐24‐6 prior to HIV exposure did not change the susceptibility of these cells to HIV infection, (b) syncytia formation between CD4+ MT4 cells and HIV chronically infected PBMC was not affected by RL4‐24‐6 and (c) treatment of freshly infected PBMC with RL4‐24‐6, however, inhibited viral production. These data, together with those we previously reported using anti‐β2‐microglobulin (β2m) mAb, suggest that anti‐HLA class I/β2m complex mAb can modify an early step of the HIV replicative cycle without affecting the viral entry.
The presence of eosinophil‐activating mediators in sera from individuals with Schistosoma mansoni infections
Authors: Mazza Graziella , Thorne Kareen J. I. , Richardson Brian A. , Butterworth Anthony E.
Published: March 2007

IgG antibodies and eosinophils kill schistosomula of Schistosoma mansoni in vitro, and there is now evidence to suggest that the main factor that contributes to the expression of purified IgG effector function is the degree of activation of the donor's eosinophils. This study was designed to identify serum‐derived activating factors in sera from individuals infected with S. mansoni. Such activating factors may be responsible for enhancing eosinophil cytotoxicity against schistosomula. Serum‐borne mediators were prepared by fractionation of sera from infected individuals by gel filtration high‐performance liquid chromatography. The eosinophil‐stimulating activity of these mediators was assayed by a new method which depends on the increased expression of the CR3 α chain (CD11b) on the surface of activated eosinophils. Sera from infected individuals exhibited different levels of eosinophil activation, and activation appeared to be due to several serum factors, including interleukin 5. In conclusion, our results suggest that eosinophil‐activating factors present in infection sera may not only be responsible for enhancing eosinophil cytotoxicity but also be necessary for its expression.
Major histocompatibility complex class II hyperexpression on B cells in interleukin 4‐transgenic mice does not lead to B cell proliferation and hypergammaglobulinemia
Authors: Müller Werner , Kühn Ralf , Rajewsky Klaus
Published: March 2007

The murine interleukin 4 (IL 4)‐encoding cDNA expressed under the control of the immunoglobulin heavy chain enhancer/promoter was inserted into the mouse germ line. All B cells present in the IL 4 transgenic mice show a marked increase in expression of class II histocompatibility (Ia) antigens compared to B cells from control mice. In vivo 5‐bromo‐2′‐deoxyuridine incorporation and cell cycle analysis revealed no significant increase in B cell proliferation in spleens of IL 4 transgenic mice compared to littermate controls. Likewise, serum immunoglobulin concentrations in IL4 transgenic mice were not increased. From these experiments we conclude that Ia hyperexpression on B cells per se does not induce activation of autoreactive T cells leading to B cell proliferation or hypergammaglobulinemia.
A role for adhesion molecules in contact‐dependent T help for B cells
Authors: Owens Trevor
Published: March 2007

The role of cell contact in T‐dependent B cell activation was examined. Small resting B cells from C57BL/6 mice were cultured with CBA‐derived, non‐alloreactive cloned T helper cells in anti‐T cell receptor Vβ8‐coated microwells. This induced polyclonal B cell activation to enter cell cycle (as measured by thymidine incorporation at 2 days) and to secrete immunoglobulin (as measured by an enzyme‐linked immunoassay detecting high‐rate Ig secretion at 5 days). The inclusion of monoclonal antibodies against LFA‐1, ICAM‐1 and CD4 in these cultures strongly inhibited antibody responses, although proliferative responses were only inhibited to about 50%. Inhibitory monoclonal antibodies did not significantly affect lipopolysaccharide‐induced responses, T cell activation to interleukin (IL) 3 secretion, nor did they inhibit the formation of multicellular clusters containing T and B cells. There was no correlation between the level of expression of adhesion molecules by T cells and their ability to induce B cell responses. Anti‐LFA‐1 abrogated T‐dependent responses to IL 2 which were inducible after 2 days in culture, but did not inhibit the induction of this IL 2 responsiveness. These results suggest that continued cell contact involving adhesion/accessory molecules induces B cells to proliferate and to respond to T cell lymphokines. A signaling role for cell interaction molecules on B cells is proposed, similar to the role of these and analogous molecules on T cells.
Expression of mouse IgA by transgenic mice, pigs and sheep
Authors: Lo David , Pursel Vernon , Linton Phyllis J. , Sandgren Eric , Behringer Richard , Rexroad Caird , Palmiter Richard D. , Brinster Ralph L.
Published: March 2007

The development of transgenic animal technology allows the introduction of desired traits into the germ line of mice and other animals. Since the production of antibody to various polysaccharide antigens can be protective against pathogenic bacteria, we generated transgenic mice, sheep and pigs carrying genes encoding the mouse α and χ chains for antibodies against phosphorylcholine (PC) to determine whether transgene antibody might be used to influence susceptibility to disease. High serum levels of mouse IgA were detected in transgenic mice and pigs but not in transgenic sheep. It has been noted that transgene immunoglobulin expression can suppress endogenous immunoglobulin gene rearrangement and expression, and indeed, in one mouse line, expression of the transgene resulted in < 10% of spleen B cells expressing endogenous IgM. Despite this suppression, significant levels of endogenous IgM were still secreted into the serum. Suppression of endogenous IgM expression was not seen in other mouse lines, nor was it seen in transgenic pigs. In the transgenic pigs, the mouse IgA was detected in the serum despite the absence of an intact mouse χ transgene, so the secreted antibody presumably included pig light chains. Little if any of the mouse IgA in these sera showed binding specificity for PC. In one of the founder sheep, mouse IgA was detectable in peripheral lymphocytes but not in serum. Mouse χ expression was not detected in the transgenic sheep harboring an intact χ transgene. These results illustrate the potential of introducing beneficial traits such as germ‐line‐encoded immunity into large mammalian species.
The role of oil and agalactosyl IgG in the induction of arthritis in rodent models
Authors: Rook Graham , Thompson Steve , Buckley Marion , Elson Chris , Brealey Richard , Lambert Claude , White Tony , Rademacher Thomas
Published: March 2007

The proportion of agalactosyl IgG [Gal(O)] is raised in human rheumatoid arthritis and tuberculosis. We report here that injection of pristane into the peritoneal cavities of mice on days 0 and 50, which is known to induce plasmacytomas and arthritis, also induced a rise in the proportion of Gal(O), correlating with a simultaneous rise in the level of IgG antibody binding to the 65‐kDa heat‐shock protein of Mycobacterium bovis (hsp65). Arthritis developed in a proportion of those CBA/Igb mice with the highest percentage of Gal(O). Pretreatment with 50 μg of recombinant mycobacterial hsp65 intraperitoneal (i.p.) on day —10, or with 500 rad irradiation on day ‐ 2 before the first of the two injections of pristane reduced the incidence of arthritis from 24% in control animals, to 5.3% and 0.4%, respectively. The reduced incidence of disease correlated with smaller rises in the % Gal(O) at 50–75 days, although levels at 150–200 days were not affected.

The arthritogenic effect of oil was not confined to the pristane model, since a single i.p. injection of oil 21 days before immunizing DBA/1 mice with type II collagen reduced the mean day of onset of this arthritis, [which we have previously shown to correlate with raised % Gal(O)], from 38 to 15 days (p < 0.001). One interpretation is that an autoimmunogenic stimulus, given when % Gal (O) is raised, is more likely to evoke disease.

Since oil granulomata are known to secrete interleukin 6, which has B cell‐regulatory properties and is secreted by rheumatoid synovial cells, we tested sera from interleukin 6‐transgenic mice, and found a strikingly raised percentage of Gal(O). We suggest, therefore, that the role of oil in the induction of arthritis is the dysregulation of cytokine release of which a raised percentage of Gal(O) may be a direct or indirect consequence, associated with an increased susceptibility to autoimmunogenic stimuli.
Induction of interleukin 2 (IL2) and interferon‐γ and enhancement of IL2 receptor expression by a CD26 monoclonal antibody
Authors: Plana Montserrat , Viñas Odette , de la Calle‐Martin Oscar , Lozano Francisco , Inglés‐Esteve Julia , Romero Matilde , Alberola‐Ila José , Yagüe Jordi , Vilella Ramön , Vives Jordi
Published: March 2007

The ability of the 134‐2C2 monoclonal antibody (mAb; CD26) to transmit an activation signal and to affect T cell proliferation has been studied. The 134‐2C2 mAb, although not being mitogenic by itself, is able to increase the proliferation of purified T cells in the presence of exogenous interleukin 2 (IL2) or phorbol 12‐myristate 13‐acetate (PMA). No effect of our mAb was observed on the proliferation of T cells induced by other stimuli such as Sepharose‐bound CD3 mAb, phytohemagglutinin or calcium ionophore. Since the co‐stimulatory effect of 134‐2C2 mAb on PMA‐induced T cell proliferation was strongly inhibited by an anti‐Tac antibody, its involvement on the IL2/IL2 receptor pathway was investigated. An increased IL2 secretion in T cells cultured with PMA plus 134‐2C2 mAb was observed and Northern blot analysis showed that the mAb 134‐2C2 acts synergistically with PMA favoring the induction of both IL2 and interferon‐γ mRNA synthesis, as well as the enhancement of IL2 receptor and transferrin receptor mRNA expression. Studies on mechanisms implicated in signal transduction showed that 134‐2C2 mAb modifies neither intracellular calcium levels nor phosphoinositide breakdown. Additionally, no effect was exerted on protein kinase C translocation. These data suggest that the CD26 antigen is involved in T cell activation in an IL2/IL2 receptor‐dependent pathway.
Surface expression by mononuclear phagocytes of an epitope shared with mycobacterial heat shock protein 60
Authors: Wand‐Württenberger Angela , Schoel Bernd , Ivanyi Juraj , Kaufmann Stefan H. E.
Published: March 2007

Bone marrow‐derived macrophages were stained with a variety of monoclonal antibodies (mAb) with specificity for the mycobacterial heat shock protein (hsp) 60 or with specific rabbit antiserum raised against mycobacterial hsp 60. The mAb ML30 as well as the specific antiserum brightly stained bone marrow‐derived macrophages whereas all the other mAb as well as normal rabbit antiserum gave no or negligible reactivity. Surface expression of the shared epitope is observed by day 3 of in vitro cultivation and markedly increased by interferon‐γ stimulation on day 9. Although hsp 60 is thought to be restricted to the mitochondrial compartment, antibody responses to this molecule have been implicated in autoimmunity. Our finding that bone marrow‐derived macrophages express a cross‐reactive epitope recognized by an mAb with specificity for amino acids 275 to 295 of the mycobacterial hsp 60 is consistent with such a role.
Recombinant 25‐kDa CD23 and interleukin 1α promote the survival of germinal center B cells: evidence for bifurcation in the development of centrocytes rescued from apoptosis
Authors: Liu Yong‐Jun , Cairns Jennifer A. , Holder Michelle J. , Abbot Sandra D. , Jansen Katherin U. , Bonnefoy Jean‐Yves , Gordon John , Maclennan Ian C. M.
Published: March 2007

Germinal centers contain a proliferating pool of centroblasts which give rise to non‐dividing centrocytes. Centrocytes are programmed to die by apoptosis unless they receive a positive signal for rescue. Rescue, in vivo, is likely to be dependent, initially, on interaction with antigen held on follicular dendritic cells (FDC). A subset of FDC located in that part of the germinal center furthest from centroblasts is particularly rich in CD23. Supernatants containing high levels of soluble CD23 were found not only to encourage the survival of germinal center B cells but also to promote their differentiation toward a plasmacytoid morphology; these activities were diminished following removal of CD23 from the supernatants. Recombinant 25‐kDa CD23 was initially found to be incapable of providing the signal for germinal center cell development but on the addition of interleukin 1α which, by itself, was inactive, rescue and differentiation of germinal center B cells were now achieved. Apoptosis in germinal center cells could also be prevented by the ligation of surface CD40 with monoclonal antibody: however, rescue via this pathway was not accompanied by plasmacytoid differentiation. These findings provide a functional rationale to the high level expression of CD23 found within a discrete subset of FDC and indicate a bifurcation in the development of germinal center B cells following their rescue from apoptosis.
Phenotypic and genetic characterization of a unique B lymphocyte deficiency in strain A/WySnJ mice
Authors: Miller David J. , Hayes Colleen E.
Published: March 2007

The elucidation of B lymphocyte development has been partially achieved through genetic models such as the X‐linked immunodeficiency (xid) mutation. We discovered a unique B lymphocyte developmental defect in strain A/WySnJ mice. We used single‐ and two‐color flow cytometry to analyze lymphocytes from A/J and A/WySnJ mice. Adult A/WySnJ mice had a severe B cell deficiency, which was apparent in the spleen, lymph nodes, peritoneum and peripheral blood, compared to adult A/J mice. An ontogeny study revealed that a developmental defect, inhibiting B lymphocyte maturation or differentiation but not B lymphopoiesis, was responsible for the deficiency. This maturational defect blocked the production of B220hi/Iahi/surface IgMlo B cells, and was manifested in the adult bone marrow and neonatal spleen, but not the adult spleen. Neither Ly‐1 B cells nor peripheral T cells were apparently affected by the A/WySnJ defect. The B cell immunodeficiency segregated as an autosomal co‐dominant trait in F1 and F2 mice. We propose that A/WySnJ mice have a novel genetic defect arresting B cell differentiation in adult bone marrow and neonatal spleen.
Anti‐α/β T cell receptor monoclonal antibody provides an efficient therapy for autoimmune diabetes in nonobese diabetic (NOD) mice
Authors: Sempé Pascal , Bédossa Pierre , Richard Marie‐Francoise , Villà Maria‐Carme , Bach Jean‐Francois , Boitard Christian
Published: March 2007

The nonobese diabetic (NOD) mouse is a relevant model for studying human insulin‐dependent diabetes mellitus (IDDM). The selective destruction of insulin‐secreting cells in this model is subsequent to an autoimmune reaction directed towards the β cells inside the islets of L angerhans of the pancreas. Given the key role played by T cells in the development of IDDM, we investigated a model of IDDM prevention in NOD mice by administration of a monoclonal antibody to the α/β dimer of the T cell receptor for antigen. Our data provide evidence that aiming at the T cell receptor protects against both spontaneous and cyclophosphamide‐induced diabetes in the NOD mouse. Interestingly, potencial clinical application is suggested by the efficient and durable reversal of recent onset diabetes in mice treated with anti‐α/β monoclonal antibody within 1 week following the clinical discovery of IDDM.
Major histocompatibility complex binding and T cell recognition of a viral nonapeptide containing a minimal tetrapeptide
Authors: Schulz Manfred , Aichele Peter , Schneider Reto , Hansen Ted H. , Zinkernagel Rolf M. , Hengartner Hans
Published: March 2007

The primary immune response of cytotoxic T lymphocytes in H‐2d and H‐2q mice to infection with lymphocytic choriomeningitis virus is directed mostly towards the common major T cell epitope of amino acids 112–132 on the viral nucleoprotein (NP). The molecules responsible for presentation of the T cell epitope NP112‐132 are in both haplotypes the MHC class I L antigens (Ld, Lq). Truncations of the amino and carboxy termini of the NP 112–132 sequence revealed the nonapeptide RPQASGVYM (NP118‐126) as a most effective peptide antigen, but even the tetrapeptide GVYM was recognized by CTL of both haplotypes in a class I antigen‐restricted specificity. When tyrosine (Y) or methionine (M) were substituted with alanine, CTL recognition of the altered nonamer required 106 to 108 times higher peptide concentrations and in one case (Y → A on Ld) the peptide was not recognized at all. Up‐modulation of the expression of Ld and Lq class I antigens as measured by flow cytometry correlated with the ability to present the peptide antigens. The only exception was peptide NP118‐126 (M → A), which was recognized by T cells on L‐Ld and L‐Lq target cells but failed to up‐regulate Ld and Lq antigens.
Molecular cloning of rat interleukin 4 cDNA and analysis of the cytokine repertoire of subsets of CD4+ T cells
Authors: McKnight Andrew J. , Barclay A. Neil , Mason Don W.
Published: March 2007

Rat peripheral CD4+ T cells may be subdivided into two functionally distinct subpopulations (OX‐22highCD4+ and OX‐22lowCD4+) on the basis of their reactivity with the monoclonal antibody MRCOX‐22, which recognizes a restricted epitope on the leukocyte common antigen (LCA, CD45). Previous studies have demonstrated the increased activity of the OX‐22highCD4+ subset in assays of cell‐mediated reactivity, whereas the reciprocal OX‐22lowCD4+ subset provides the majority of help for B cells in secondary antibody responses. Analyses of in vivo function have subsequently shown that the autoreactive activity associated with the OX‐22highCD4+ subset can be inhibited through a suppressor activity within the OX‐22lowCD4+ subset, indicating a further immunoregulatory role for these cells. Since the differential production of lymphokines such as interferon‐γ (IFN‐γ) and interleukin 4 (IL4) is believed to regulate alternative effector responses to a particular antigen, we have compared the lymphokine mRNA profiles of activated OX‐22highCD4+ and OX‐22lowCD4+ subsets using nucleic acid probes specific for rat IL 2, IFN‐γ and IL4, the latter of which has been isolated by a polymerase chain reaction cloning technique and its sequence is described. A higher frequency of cells expressed IL 2 mRNA in the OX‐22high subset, in accordance with the relative levels of IL 2 protein produced. In contrast, more IFN‐γ mRNA was detected in the OX‐22lowCD4+ subset 24 h after mitogenic stimulation although these cells have consistently been shown to produce less IFN‐γ protein than the OX‐22highCD4+ subset. This apparent paradox was resolved by the finding that the IFN‐γ mRNA levels in the OX‐22lowCD4+ subset declined rapidly after 24 h while the levels continued to rise in the OX‐22highCD4+ population such that at 48 h the relative levels were reversed. We have also demonstrated a higher level of IL 4 mRNA expression within the OX‐22lowCD4+ subset, which is undoubtably involved in the increased B cell helper activity mediated by this subpopulation and may be responsible, in part, for their active suppression of cell‐mediated immune responses.
A myeloma M104E private idiotope is represented predominantly among anti‐dextran, peritoneal cavity Ly‐1 precursor B cells
Authors: Lehmann Hans‐Peter , Lehle Georg
Published: March 2007

Public and private idiotopes (Id) had been defined in the BALB/c anti‐dextran B1355 fraction S (Dex) response by means of syngeneic monoclonal anti‐M 104E Id antibodies. Most of the primary anti‐Dex antibodies shared the public Id‐1 while the private Id‐5 constituted a comparatively low, but highly variable proportion of humoral anti‐Dex antibodies of individual mice. In the present work we have analyzed by limiting dilution the expression of these two Id by anti‐Dex B cell precursors from the spleen and peritoneal cavity of BALB/c mice. Testing spleen cells we found about 65% of anti‐Dex precursors to be Id‐1+ and only a minority to be Id‐5+. Treatment with anti‐Ly‐1 and complement was without any effect on anti‐Dex precursors from spleen. Anti‐Dex B cell precursors among peritoneal cavity lymphocytes differed from those found in the spleen in two ways. First, on average, 50% of them were Id‐5+; second, about 80% carried the Ly‐1 marker. Both markers are, therefore, expressed on at least 30% of anti‐Dex B cell precursors from the peritoneal cavity but rarely on anti‐Dex precursors from the spleen.
Molecular analysis of VxIII variable regions of polyclonal rheumatoid factors during rheumatoid arthritis
Authors: Blaison Gilles , Kuntz Jean‐Louis , Pasquali Jean‐Louis
Published: March 2007

We report the first molecular characterization of Vx regions of the main human autoantibodies occurring during rheumatoid arthritis, the polyclonal rheumatoid factors. Using two sets of polymerase chain reactions in order to amplify the cDNA derived from both peripheral blood and synovial fluid rheumatoid factor‐secreting cells, nucleotide analysis of the VxIII family usage shows the following: (a) at least three different VxIII genes are used to encode polyclonal rheumatoid factors in a single patient, (b) each one of these genes seems more or less somatically mutated (from 1 to 14 mutations), (c) the mutation process preferentially affects the complementarity determining regions suggesting a selective pressure of antigen and (d) there is no clear difference between the mutation rates affecting the synovial fluid and peripheral blood rheumatoid factor‐secreting cells. These results are able to explain some of the known idiotypic differences between monoclonal and polyclonal rheumatoid factors in humans. They also provide evidence that polyclonal autoantibodies arising during an autoimmune disease can be the products of multiple somatically mutated genes and suggest that this process is antigen driven, whether this antigen is the Fc piece of IgG or another unknown antigen.
Transient depletion of T cells with high LFA‐1 expression from peripheral circulation during acute Plasmodium falciparum malaria
Authors: Hviid Lars , Theander Thor G. , Abdulhadi Nasreldin H. , Abu‐Zeid Yousif A. , Bayoumi Riad A. , Jensen James B.
Published: March 2007

Acute P. falciparum malaria is associated with loss of in vitro T cell responsiveness to antigenic stimulation, and with high plasma levels of soluble interleukin 2 receptor (IL 2R). In the present study peripheral T cells from acute P. falciparum malaria patients from a malaria‐endemic area of Sudan were analyzed for expression of cell surface antigens associated with T lymphocyte adhesion, activation and maturation. The results were compared to results from T cells obtained from the same donors either before the attack, or during convalescence. Most donors showed a remarkable loss of T cells with high expression of the surface marker LFA‐1 (CD11a/CD18) during the clinical episode, in addition to the functional changes described above. Two donors that did not show phenotypic changes were furthermore characterized by having an unabated proliferative response and normal plasma IL2R levels. All peripheral CD3+ T lymphocytes expressed LFA‐1, which had a clearly bimodal distribution on these cells. The T cell subpopulation having high LFA‐1 expression (LFA‐1++) was composed of both memory and unprimed T cells, according to their expression of CD45RA and CD45R0. Analysis of expression of membrane‐bound IL 2R (CD25) and ICAM‐1 (CD54) did not reveal in vivo activated T cells in the peripheral blood of the patients. Taken together, these data suggest that circulating T cells recognizing parasite antigens are temporarily withdrawn from peripheral circulation during P. falciparum malaria.
Antigen presentation of hen egg‐white lysozyme but not of ribonuclease A is augmented by the major histocompatibility complex class II‐associated invariant chain
Authors: Nadimi Farsin , Moreno José , Momburg Frank , Heuser Andreas , Fuchs Serge , Adorini Luciano , Hämmerling Günter J.
Published: March 2007

The influence of the class II‐associated invariant chain (Ii) on the presentation of the protein antigens hen egg‐white lysozyme (HEL) and ribonuclease A (RNase) was investigated. For this purpose the Ii− rat‐2 fibroblasts were transfected with I‐Ak genes with or without Ii. Transfectants expressing Ii were superior in the presentation of the complete HEL protein to a panel of I‐Ak‐restricted T hybridomas characterized by distinct specificities for different HEL peptides and by different sensitivities to antigen concentration. There appeared to be a correlation between the antigen‐presenting capacity and the amount of Ii, in that transfectants expressing large amounts of Ii were the best antigen presentors. The presentation of synthetic HEL peptides was not infuenced by Ii.

In contrast to the findings with HEL, the presentation of RNase by the same set of transfectants was clearly independent of Ii. Both antigens, HEL and RNase, required processing in the chloroquine‐sensitive compartment. However, only the presentation of HEL but not of RNase could be efficiently blocked by brefeldin A. These data confirm that presentation of HEL depends on de novo synthesized class II molecules, whereas the presentation of RNase seems to be predominantly mediated by a pool of pre‐existing class II molecules whose interaction with endocytosed antigen does not depend on Ii. These results suggest different mechanisms for the presentation of HEL and RNase and they raise the possibility that different antigens intersect the class II pathway at distinct intracellular locations.
Human eosinophils from hypereosinophilic patients spontaneously express the p55 but not the p75 interleukin 2 receptor subunit
Authors: Plumas Joël , Gruart Valérie , Aldebert Delphine , Truong Marie José , Capron Monique , Capron Audré , Prin Lionel
Published: March 2007

The expression of interleukin 2 receptor (IL 2R) on eosinophils was investigated in patients with hypereosinophilia. Hypodense activated eosinophils have been described in various diseases such as parasitic or allergic diseases, hypereosinophilic syndrome (HES) associated in some cases to myeloproliferative markers, and more recently described in patients undergoing recombinant IL 2 treatment. The presence of p55 α chain of IL 2R (CD25) on purified eosinophils collected from blood of hypereosinophilic patients was detected by flow cytometry. In 10 out of 19 cases, more than 10% of eosinophils were CD25+. Cross‐linking studies on enriched eosinophils showed one 64–75‐kDa band, consisting of IL 2 (15 kDa) cross‐linked to the IL2R p55 subunit. In Northern blot analysis the two messenger mRNA (3.5 and 1.5 kb) encoding the IL2R p55 subunit were identified after hybridization with a CD25 cDNA probe. In contrast, the presence of the IL2R p75 subunit was not detected. These data provide preliminary evidence for the expression of a low‐affinity receptor for IL 2 on in vivo activated eosinophils and raise the question of the role played by this cytokine in eosinophil differentiation and activation.
Calcium signals in single T cells on activation by lectin
Authors: Tregear Richard T. , Hoyland John , Mason William T.
Published: March 2007

Succinylation of concanavalin A (Con A) reduces its oligomer size while retaining its mitogenicity, and provides a probe of T cell activation. We have observed responses of cytosolic ionized calcium to succinyl Con A in suspensions of Jurkat and rat lymph node (LN) cells, using a fluorimeter, and in single cells settled on glass, using a dual wavelength video imaging system. In the fluorimeter a mitogenic level of succinyl Con A (30 μg/ml) produced only a 15–30 nM rise in average cell calcium in the suspended Jurkat or rat cells whereas the use of quantitative video imaging produced asynchronous 250–1000 nM pulses of free calcium in 35% of Jurkat cells and 300–850 nM pulses in 45% of rat LN cells. In Jurkat cells these pulses were sometimes repetitive, giving rise to apparent oscillations. In the fluorimeter 30 μg/ml of native Con A (a supra‐mitogenic concentration) produced a 300 nM rise in average cell calcium in suspended Jurkat cells, and a 100 nM rise in rat LN cells; when major histocompatibility complex class II‐bearing cells were removed the response rose. Mitogenic Con A (3 μg/ml) produced a much lower rise in calcium. With video imaging the response seen was greater. Levels > 30 μg/ml Con A caused 700–5000 nM pulses synchronously in 94% of Jurkat cells and 250–1000 nM pulses in 73% of rat LN cells. At 3 μg/ml Con A produced asynchronous 300–1100 nM pulses in 36% of rat LN cells. We conclude that the absence of a calcium signal in the fluorimeter can conceal asynchronous calcium responses in individual cells and that brief asynchronous cytosolic calcium pulses are sufficient for lectin to activate rat T cells.
T cell reactivity to an epitope of the mycobacterial 65‐kDa heat‐shock protein (hsp 65) corresponds with arthritis susceptibility in rats and is regulated by hsp 65‐specific cellular responses
Authors: Hogervorst Els J. M. , Boog Claire J. P. , Wagenaar Josée P. A. , Wauben Marca H. M. , van der Zee Ruurd , van Eden Willem
Published: March 2007

Adjuvant arthritis (AA) can be induced in genetically susceptible rats by immunization with heat‐killed mycobacteria suspended in mineral oil. From our analysis of arthritogenic T cell clone A2b, obtained from an arthritic Lewis rat and specific for the 180–188 epitope of mycobacterial 65‐kDa heat‐shock protein (hsp 65), the possible origin of AA was explained by the existence of a molecular mimicry of the 180–188 epitope with a cartilage‐associated self antigen. We now have shown that Lewis rats respond to the 180–188 epitope after Mycobacterium tuberculosis immunization and that arthritis‐resistant Fisher and (Lewis × Fisher)F1 rats, although major histocompatibility complex class II identical with Lewis, do not respond to this epitope. However, in rare cases of arthritis in Fisher rats, responses to the epitope were seen. We obtained no evidence for a defect at the level of antigen processing and presentation or for suppression in Fisher rats. Thus, non‐responsiveness in Fisher rats was likely due to a difference at the level of the T cell repertoire.

Previously, we have reported that pretreatment with hsp 65 in experimental arthritis, and not only in AA, caused resistance to arthritis induction. We now present evidence that immunization with hsp 65 or in vitro stimulation with hsp 65 may lead to inhibition of responses specific for epitope 180–188. Thus the hsp 65‐induced resistance to arthritis is probably caused by the induction of regulatory control specifically targeted at the 180–188 epitope. Especially in rats that tend to focus their responses on the critical 180–188 sequence, such as Lewis, regulation seems to develop following immunization with hsp 65. Since recent evidence suggests that hsp 65 and also the 180–188 epitope have a role in human arthritic conditions, the present findings are expected to contribute to further experimentation directed at exploiting hsp 65 or its epitopes for the development of new therapeutical approaches in humans.
VH gene usage in humans: biased usage of the VH6 gene in immature B lymphoid cells
Authors: Berman Jeffrey E. , Nickerson Katherine G. , Pollock Roberta R. , Barth Jeremy E. , Schuurman Ruud K. B. , Knowles Daniel M. , Chess Leonard , Alt Frederick W.
Published: March 2007

Preferential usage of JH‐proximal VH genes has been demonstrated in immature murine B cell repertoires. To determine whether this phenomenon is also evident in human repertoires, we studied utilization of VH6, the most JH‐proximal human VH gene. Examination of VH gene usage in a panel of precursor B cell acute lymphoblastic leukemia samples indicated that 15% of the IgH rearrangements utilized VH6. VH6 is a single‐member family in a total repertoire of 100–200 VH genes; thus, if usage were purely random, one would expect VH6 rearrangement frequency to be < 1%. Analysis of VH gene usage in normal lymphoid tissues also revealed biased usage of VH6. VH6 was preferentially utilized in 16‐ to 24‐week‐old fetal liver as compared to adult peripheral blood mononuclear cells or spleen. Possible implications of the conservation of preferential usage of JH‐proximal genes in both immature murine and human repertoires are discussed.