bluuebunny/crossref_metadata_2025 huggingface dataset:
Predominance of Th1‐type T cells in synovial fluid of patients with Yersinia‐induced reactive arthritis
Authors: Schlaak Jörg , Hermann Elisabeth , Ringhoffer Mark , Probst Peter , Gallati Harald , zum Büschenfelde Karl‐Hermann Meyer , Fleischer Bernhard
Published: March 2007

The pathogenetic mechanisms underlying the development of reactive arthritis and the functional capacities of synovial T cells specific forYersinia enterocoliticaare still unclear. In this study we have determined the cytokine secretion patterns of 24 CD4+synovial fluid (SF)‐derived T cell clones from 2 patients with Yersinia‐induced reactive arthritis, 16 clones specific for different Yersinia antigens and 8 clones as controls. The clones specific for Yersinia antigens predominantly belong to the T helper cell 1 (Th1) subset with production of interferon (IFN)‐γ and interleukin (IL)‐2, but no IL‐4, whereas SF T cells not reactive with Yersinia antigens produce IL‐2, IL‐4 and IFN‐γ and thus belonged to the Th0 subset. Moreover, short‐term T cell lines established from SF and peripheral blood showed the same pattern. To further analyze the functional relevance of these data we investigated the influence of IFN‐γ and IL‐4 on the intracellular killing of Yersinia in a human glioblastoma cell line. Our data show that the Th1 cytokine IFN‐γ promotes intracellular killing of Yersinia, whereas this effect is antagonized by the Th2 cytokine IL‐4. Furthermore, the Th2 cytokine IL‐10 inhibited the antigen‐specific proliferative response and IFN‐γ and IL‐2 production by the Th1 cells. These results provide insight into the antibacterial mechanisms at work in reactive arthritis after infection withYersinia enterocoliticaand, for the first time, reveal the cross‐regulatory properties of cytokines derived from Th1 and Th2 cells in a human immune response to bacterial antigens.
Differential thymus dependence of rat CD8 isoform expression
Authors: Torres‐Nagel Nora , Kraus Elmar , Brown Marion H. , Tiefenthaler Georg , Mitnacht Rita , Williams Alan F. , Hünig Thomas
Published: March 2007

Expression of the rat CD8 molecule was studied using five novel monoclonal antibodies (mAb), four of which are specific for the V‐like domain of CD8α, whereas one reacts either with the β chain or with a determinant only expressed on the CD8 α/β heterodimer. mAb to both chains effectively blocked purified lymph node CD8 T cells in mixed lymphocyte reaction and in cell‐mediated cytotoxicity. Flow cytometric analysis showed that CD8 T cells from lymph nodes or spleen of normal rats almost exclusively express the α/β isoform, regardless of the T cell receptor isotype (α/β or γ/δ). In contrast, natural killer (NK) cells carry only CD8α chains. This CD8α+β− phenotype was also prominent among CD8 T cells from athymic rats and from intestinal epithelium of normal rats. CD8α homodimers can also be expressed as a result of activation, as shown by analysis of CD4 CD8 double‐positive T cells obtained from highly purified lymph node CD4 T cells by in vitrok stimulation. Such CD4+CD8α+β− cells also represent a major subset among adult intestinal intraepithelial lymphocytes (IEL), suggesting local activation. Taken together, the difference in CD8 isoform expression among T cells from athymic rats, NK cells, and gut IEL versus CD8 T cells from peripheral lymphatic organs of euthymic animals suggests that like in mice, expression of the CD8 heterodimer is more dependent on intrathymic maturation than that of the homodimer. Since the more stringent thymus dependence of CD8α+β+ T cells may be due to a requirement for thymic selection on self major histocompatibility complex class I antigens, the virtually exclusive CD8α+β+ phenotype of peripheral rat γ/δ T cells could mean that antigen recognition by this subset is also restricted by MHC class I molecules.
In vivo administration of interleukin‐2 turns on anergic self‐reactive T cells and leads to autoimmune disease
Authors: Gutierrez‐Ramos José C. , De Alboran Ignacio Moreno , Martinez‐A Carlos
Published: March 2007

One major mechanism of self tolerance involves the deletion of T cell clones in the thymus. In athymic mice, tolerance to self antigens must be generated extrathymically. T cells with self‐reactive receptors undergo either peripheral clonal deletion or become unresponsive (i.e. anergic). The unresponsive state of human and mouse T cell clones in vitro can be reversed by the addition of exogenous interleukin (IL)‐2, thus transforming anergic T cells to an activated state. Here it is shown that the in vivo delivery of IL‐2 to athymic BALB/c nu/nu mice abrogates the anergic state of self‐reactive Vβ3+ and Vβ11+ T cells [which are normally deleted in the minor lymphocyte stimulatory (Mls)‐1b‐, I‐E+‐expressing euthymic counterparts]. Thus, Vβ3+ and Vβ11+ T cells from IL‐2‐treated nude mice proliferate in response to T cell receptor cross‐linking and acquire effector functions as measured by their ability to deliver aid to B cells upon specific stimulation. This activation correlates with the development of autoimmune manifestations (DNA autoantibodies, rheumatoid factors, erythroleukopenia and minimal change nephritis) in these IL‐2‐treated mice.
Interferon‐γ induced lethality in the late phase of Plasmodium vinckei malaria despite effective parasite clearance by chloroquine
Authors: Kremsner Peter G. , Neifer Stefan , Chaves Mair F. , Rudolph Roland , Bienzle Ulrich
Published: March 2007

A combination therapy was tested consisting of chloroquine and interferon‐γ (IFN‐γ) in the late phase of blood‐stage Plasmodium vinckei malaria in BALB/c mice. When mice were treated with three times 300 μg chloroquine at 24‐h intervals starting at a parasitemia of 30%‐50%, only 5 of 14 mice (36%) died 2–4 days after initiation of therapy. However, when infected mice received chloroquine plus 1 μg IFN‐γ at the same time, 14 of 18 mice (78%) died 0.5–3 days after start of therapy (p < 0.05) despite clearance of parasitemia. The histopathology from mice dying after combination therapy revealed interstitial leukocyte infiltration of lung tissue, severe liver cell necrosis and kidney tubular necrosis.

Pretreatment of P. vinckei‐infected mice with pentoxifylline, a phosphodiesterase inhibitor, led to a significant decrease of IFN‐γ‐induced lethality (p < 0.05). In contrast, pretreatment with neutralizing antibodies to tumor necrosis factor or with L‐N‐monomethyl arginine, the latter an inhibitor of the nitric oxide synthase, significantly increased lethality (p < 0.05).
Heat shock prevents lipopolysaccharide‐induced tumor necrosis factor‐α synthesis by rat mononuclear phagocytes
Authors: Fouqueray Bruno , Philippe Carole , Amrani Abdelaziz , Perez Joëlle , Baud Laurent
Published: March 2007

Tumor necrosis factor‐α (TNF‐α), a mononuclear phagocyte‐derived peptide is known to participate in the pathogenesis of fever. To determine whether a feedback mechanism exists by which elevated temperatures influence TNF‐α generation, we have examined the effects of heat shock on the in vitro synthesis of TNF‐α by rat glomeruli, inflammatory peritoneal macrophages and blood monocytes. Preexposure of peritoneal macrophages to elevated temperatures for 20 min decreased the subsequent lipopolysaccharide‐induced release of TNF‐α bioactivity. The mean reductions were 11.9 ± 86.3 ± 12.0, and 95.2 ± 3.5% after pretreatment at 39, 41 and 43°C, respectively. Reductions, that were transient, were maximum when lipopolysaccharide was added 0–2 h after heat shock. They correlated with the decreased release of immunoreactive TNF‐α and the decreased expression of both cell‐associated TNF‐α molecule and TNF‐α mRNA. Heat shock‐induced inhibition of TNF‐α release was independent of variations of prostaglandin synthesis, but was possibly related to the induction of heat‐shock proteins since (a) macrophages exposed to heat shock synthesized the major 70‐ and 90‐kDa heat‐shock proteins, and (b) chemical inducers of the heat‐shock response were also effective inhibitors of TNF‐α release. The mean reduction of TNF‐α release after pretreatment at 41°C was found to be identical in glomerular tissue (82.0 ± 7.5%), but significantly less in blood monocytes (43.9 ± 10.9%). This supports the hypothesis that a negative‐feedback mechanism exists between elevated temperature and lipopolysaccharide‐induced TNF‐α synthesis, and suggests that this regulation is less active in blood monocytes than in tissue macrophages.
Evidence for a role of CD28RE as a response element for distinct mitogenic T cell activation signals
Authors: Civil Azem , Geerts Marlieke , Aarden Lucien A. , Verweij Cornelis L.
Published: March 2007

T lymphocytes can be induced to produce interleukin (IL)‐2 and proliferate upon T cell receptor (TcR) occupancy together with a CD28‐induced co‐stimulatory signal. The T cell surface molecule CD28 is believed to function as a regulator in T cell activation at both the level of lymphokine mRNA stabilization and gene transcription. Activation of IL‐2 gene transcription via CD28 has been shown to be mediated through a xB‐like sequence, called CD28RE. DNA binding analysis revealed that the CD28‐induced signal is involved in the induction of CD28RE binding activity. Here, we demonstrate that the induction of CD28RE binding activity is not specific for the CD28‐induced signal. Our data indicate that distinct mitogenic T cell activation signals converge on the induction of CD28RE binding activity, and suggest a crucial role for this activity in the IL‐2 enhancer responsiveness to different modes of T cell activation.
Polyreactive autoantibodies to negatively charged epitopes following Trypanosoma cruzi infection
Authors: Kahn Stuart , Kahn Maria , Eisen Harvey
Published: March 2007

During the course of many human autoimmune diseases, antibodies which recognize negatively charged epitopes on self antigens are detected. Trypanosoma cruzi, an intracellular protozoan parasite capable of infecting a wide variety of vertebrates, is the cause of Chagas disease in humans. Infection with the parasite frequently results in autoimmune and inflammatory pathology. We report here on an affinity‐purified population of antibodies that bind to a broad class of antigens that contain runs of acidic amino acids, including tubulin. Although these antibodies can be isolated from both uninfected and T. cruzi chronically infected C3H/He mice, the antibodies from the normal mice (the natural autoantibodies) bind to tubulin poorly at physiological pH, whereas the antibodies isolated from the infected animals bind well at physiological pH. We propose that similar processes may occur in humans following other infections accounting for the detection of antibodies to negatively charged epitopes in a variety of autoimmune diseases.
Vaccination or tolerance to prevent diabetes
Authors: Oehen Stephan , Ohashi Pamela S. , Aichele Peter , Bürki Kurt , Hengartner Hans , Zinkernagel Rolf M.
Published: March 2007

Experiments with transgenic mice expressing the glycoprotein (GP) of lymphocytic choriomeningitis virus (LCMV) under the control of the rat insulin promoter (RIP) have demonstrated that potentially self‐reactive T cells that normally ignore self peptides may nevertheless be induced by self peptides or “cross‐reactive” foreign (e.g. viral) peptides that arise in the host in an immunogenic form; once activated these potentially self‐reactive T cells may cause autoaggressive diseases (e.g. diabetes). The possibility of vaccinating against such T cell‐mediated immunopathological diseases was evaluated in the RIP‐GP transgenic mouse line Bln. Any attempt to vaccinate with the self antigen itself (e.g. recombinant vaccinia virus expressing LCMV‐GP) failed to protect mice from disease. However, immunization with a recombinant vaccinia virus expressing LCMV‐nucleoprotein (vacc‐NP) as a non‐GP LCMV vaccine was able to modulate the immune response and prevented autoaggressive disease in a MHC‐dependent fashion. In contrast, tolerance induction neonatally or, more generally applicable, by lethal irradiation and reconstitution with neo‐self antigen‐expressing bone marrow cells always resulted in prevention of virally induced diabetes in this model situation.
An activated murine B cell lymphoma line (A‐20) produces a factor‐like activity which is functionally related to human natural killer cell stimulatory factor
Authors: Mengel José , Daré Leopoldo , Daré Gustavo Marcelo , Delgado Maristela , Nomizo Auro , Silva João S. , Campos‐Neto Antonio
Published: March 2007

In the present article we show that supernatants derived from lipopolysaccharide (LPS) or phorbol 12‐myristate 13‐acetate (PMA)‐stimulated A‐20 B cell lymphoma are able to induce polyclonal immunoglobulin (Ig) secretion by normal B cells in a T cell‐dependent manner. This activity could be blocked by neutralizing monoclonal antibodies against interferon‐γ, but not by monoclonal antibodies against interleukin (IL)‐2, IL‐4, IL‐5, IL‐6, IL‐10 and granulocyte macrophage colony‐stimulating factor (GM‐CSF) or even a polyclonal antibody against tumor necrosis factor (TNF)‐α. Furthermore, A‐20 supernatants induced the production of measurable amounts of interferon‐γ by normal murine spleen cells and activates natural killer (NK) cells. Fractionation of factor‐rich supernatants on a Sephacryl S‐200 column revealed that the factor activity is located in the fractions corresponding to a molecular mass of 160–150 kDa and 80–70 kDa. The biological activities found in the A‐20 supernatant are very similar to the ones described for the recently cloned human IL‐12/NK cell stimulatory factor. These results suggest the existence of a murine analogous factor for the human IL‐12 produced by A‐20 B cell lymphoma.
Inflammation activates self hsp60‐specific T cells
Authors: Anderton Stephen M. , Zee Ruurd Van Der , Goodacre John A.
Published: March 2007

Injection of incomplete Freund's adjuvant (IFA) into the footpads of BALB/c mice induced an acute inflammation. Draining popliteal lymph nodes showed major histocompatibility complex (MHC) class II‐restricted proliferation when challenged in vitro with recombinant Mycobacterium bovis 65‐kDa heat shock protein (hsp65). αβ Tcell receptor‐positive, CD4+, hsp65‐specific T cell lines and clones were generated from these lymph nodes, and 87% of clones responded to a P galactosidase fusion protein containing residues 238–573 of human hsp60. Seventy percent of these hsp60‐responsive clones also responded to a synthetic peptide corresponding to residues 412–423 of the mouse hsp60. This peptide also induced significant responses in IFA‐primed lymph node cells but not in lymphoid cells from unimmunized mice.

These results demonstrate that T cells specific for epitopes in self hsp60 are activated during inflammatory responses induced in the absence of exogenous bacterial hsp65. The findings of this study may provide a basis for understanding the often reported isolation of mycobacterial hsp65‐responsive T cells from inflammatory sites of arthritis patients, and the protective effects of preimmunization with hsp65 in experimental models of arthritis.
Antigen and helper T lymphocytes activate B lymphocytes by distinct signaling pathways
Authors: Kawakami Kazuyoshi , Parker David C.
Published: March 2007

Resting murine B lymphocytes can be induced to proliferate by cross‐linking membrane immunoglobulin, the antigen receptor, or by contact with activated helper T lymphocytes in the absence of a signal through membrane immunoglobulin. Little is known about the molecular nature of contact‐dependent T cell help. To determine whether helper T cells activate B cells through different signal transduction and second messenger pathways from those used by membrane immunoglobulin, the effects of drugs which block activation of B cells through membrane immunoglobulin were measured on B cell activation by contact with anti‐CD3‐activated and fixed T helper cells. Cyclosporin A, phorbol esters added at the time of activation, and cAMP agonists all block activation of B cells through membrane immunoglobulin at concentrations at least 100‐fold lower than those necessary to block B cell activation by contact with activated Th1 or Th2 helper T cells. Depletion of protein kinase C by pretreatment of B cells with phorbol ester inhibits the proliferative response to anti‐immunoglobulin but not the response to contact with activated T cells. The B cell response to lipopolysaccharide is intermediate in sensitivity to cyclosporin A and cAMP agonists, and resembles the response to activated T cells in resistance to phorbol esters and protein kinase C depletion. Various protein kinase inhibitors did not distinguish among these B cell activation pathways, except for the tyrosine kinase inhibitor, herbimycin A, which inhibited anti‐immunoglobulin responses at 3‐ to 5‐fold lower concentrations.
H‐2 I‐E molecules isolated from MIs1a stimulatory cells do not activate Mls1a‐responsive T cells but do present exogenous staphylococcal enterotoxins
Authors: Macphail Stuart , Stutman Osias
Published: March 2007

The T cell response to allogeneic murine Mls determinants is not H‐2 restricted but is dependent on H‐2 class II molecules on the Mls‐expressing stimulator cells. We have tested planar membranes containing H‐2 class II I‐E molecules alone or with I‐A molecules for their ability to activate a panel of Mls1a‐specific T hybrids. Despite the ability of the planar membranes to activate an alloreactive T hybrid and to present staphylococcal enterotoxins or an antigenic peptide to appropriately responsive T hybrids, they failed to stimulate the Mls1a‐specific T hybrids. These findings, in the light of the various controls demonstrating sufficiency of the I‐E molecules in the planar membranes, indicate that Mls1a determinants are not covalently bound to I‐E molecules; the two molecular species are thus either not physically associated or are linked by a relatively weak interaction. In addition, our experiments show that isolated I‐E molecules but not I‐A molecules present staphylococcal enterotoxins A and B to two independently derived T hybrids expressing T cell receptor Vβ1, Vβ8.2 and Vβ6 elements.
Bursa‐dependent subpopulations of peripheral B lymphocytes in chicken blood
Authors: Paramithiotis Eustache , Ratcliffe Michael J. H.
Published: March 2007

By selective labeling of juvenile chicken bursal cells with colloidal fluorescein isothiocyanate in situ, the emigration rate of bursal lymphocytes to the periphery was estimated at approximately 0.84% and 0.96% of the peripheral blood lymphocyte (PBL) and splenic B cell pool per hour, respectively. Emigrant bursal cells were found primarily in blood and spleen, with very small numbers migrating to thymus, bone marrow, and gut‐associated lymphoid tissues. Emigrant bursal cells expressed high levels of both major histocompatibility complex class II antigen and the Ov alloantigen, a phenotype found on a population comprising approximately 4% of bursal cells from which the bursal emigrants may be derived. Surgical bursectomy at 3 weeks of age revealed that peripheral blood B cells could be divided into three distinct populations. Specifically, 60% of the peripheral blood B cells were short lived with a half‐life of about 30 h in the blood. These cells accounted for the great majority of emigrants from the bursa to the peripheral blood. Approximately 35 % of PBL B cells had a half‐life of 12 days following bursectomy and comprised cells which did not divide in the periphery. Consequently, we propose that physiological differences between this population and the majority of bursal emigrants are established intrabursally. The remaining PBL B cells, whose relative proportion increases with age from about 5 % of PBL B cells at 2‐3 weeks of age, are short lived and are being continually produced from (a) post‐bursal site(s) of B cell production.
Epitope mapping of anti‐rat CD53 monoclonal antibodies. Implications for the membrane orientation of the Transmembrane 4 Superfamily
Authors: Tomlinson Michael G. , Williams Alan F. , Wright Mark D.
Published: March 2007

CD53 is a pan‐leukocyte glycoprotein which is a member of the recently described Transmembrane 4 Superfamily (TM4SF) of membrane proteins that are predicted to span the lipid bilayer four times. The major hydrophilic region of murine CD53 was expressed as a glutathione‐S‐transferase fusion protein, and the epitopes of four mouse anti‐rat CD53 monoclonal antibodies (mAb) (OX‐44, 2D1, 6E2 and 7D2) were mapped to this region using mouse/rat chimeric fusion proteins. The epitopes of OX‐44,6E2 and 7D2 are restored by the substitution of a single isoleucine residue for threonine at position 154 in the mouse protein. The 2D 1 epitope is non‐linear and appears to require the juxtaposition of isoleucine at position 154 with one or more of the amino acids arginine (132), methionine (133) and serine (140). All of these epitopes are shown to be sensitive to reduction, thus indicating the importance of disulfide bonding in the correct folding of the CD53 hydrophilic domain. Moreover, as these four mAb recognize CD53 at the cell surface, the data provide direct molecular evidence for the proposed membrane orientation of the TM4SF.
Leishmania mexicana promastigotes induce cytotoxic T lymphocytes in vivo that do not recognize infected macrophages
Authors: López José Alejandro , Lebowitz Jonathan H. , Beverley Stephen M. , Rammensee Hans‐Georg , Overath Peter
Published: March 2007

The question is addressed whether antigens of Leishmania, a parasite residing in the endosomal compartment of macrophages, can be presented in the context of major histocompatibility complex class I molecules. We used E. coli β‐galactosidase as a model antigen which can be expressed in high levels in L. mexicana promastigotes (L. mexicana‐gal). Infection of BALB/c mice with L. mexicanagal induces β‐galactosidase‐specific cytotoxic T cells (CTL), which can be isolated using a β‐galactosidase‐expressing mastocytoma line as an antigen‐presenting cell. These CTL recognize epitopes of β‐galactosidase in the context of H‐2Kd; however, they do not recognize L. mexicanagal‐infected macrophages even after killing of the intracellular amastigotes by drug treatment or macrophage activation by lymphokines, although class I‐peptide interaction and the presentation of endogenously produced antigens is normal. It is concluded that parasite antigens can induce a CTL response in vivo but that these CTL cannot recognize infected macrophages because the relevant epitopes cannot gain access to class I molecules. The effect of priming in vivo may be explained by the well‐known but ill‐understood phenomenon of cross‐priming.
The effects of a monoclonal antibody to interferon‐γ on experimental autoimmune thyroiditis (EAT): prevention of disease and decrease of EAT‐specific T cells
Authors: Tang Haiwen , Mignon‐Godefroy Karine , Meroni Pier Luigi , Garotta Gianni , Charreire Jeannine , Nicoletti Ferdinando
Published: March 2007

CBA/J mice immunized with thyreoglobulin (Tg) develop an experimental autoimmune thyroiditis (EAT) with lymphocytic infiltration of the thyroid glands, autoantibodies to Tg and occurrence of EAT‐specific T cells. When these mice were treated for 4 weeks after immunization with 1 mg/week of a monoclonal antibody (mAb) that neutralizes the activity of interferon‐γ (IFN) a beneficial effect on the onset of EAT was observed. Characteristic features of EAT were significantly reduced, including the lymphocytic infiltrations of the thyroid glands and the serum levels of autoantibodies to Tg. Moreover, in lymphoid organs, mAb to IFN‐γ significantly reduced the percentages of Tg‐specific CD8+ cells, labeled by the anti‐clonotypic mAb AG7. These Tg‐specific T cells seem responsible for thyroid damages and disease development, since EAT was simultaneously abrogated.

These results show that IFN‐γ plays an essential role in the pathophysiology of EAT and suggest the possibility to treat autoimmune thyroid diseases with mAb to IFN‐γ or drugs able to antagonize the production and/or the action of this cytokine.
Effects of cyclosporin A, FK 506, and mycalamide A on the activation of murine CD4+ T cells by the murine B7 antigen
Authors: Galvin Frances , Freeman Gordon J. , Razi‐Wolf Ziba , Benacerraf Baruj , Nadler Lee , Reiser Hans
Published: March 2007

The murine B7 (mB7) protein is a potent co‐stimulatory molecule for the activation of murine CD4+ T cells. We have previously shown that stable mB7‐transfected Chinese hamster ovary (CHO) cells synergize with either anti‐CD3 monoclonal antibodies (mAb) or concanavalin A to stimulate T cell activation. In addition, mB7 can synergize with phorbol 12‐myristate 13‐acetate (PMA) to induce T cell activation in an alternative pathway. In the present report we describe the effects of three immunosuppressive drugs, cyclosporin A (CsA), FK 506, and mycalamide A on mB7‐mediated T cell activation. The immuno‐philin ligands CsA and FK 506 block activation of murine CD4+ T cells by the combination of anti‐CD3 mAb and CHO‐mB7 cells but do not affect activation by CHO‐mB7 cells and PMA. These results support the relevance of pharmacological studies of immunosuppressive compounds in the murine model prior to their application in man. In contrast to the effects of CsA and FK 506, mycalamide A blocks activation of murine CD4+ T cells by anti‐CD3 mAb and mB7 as well as activation by mB7 and PMA. On a molar basis, mycalamide A appears to be at least 10‐fold more potent than FK 506 which is 100‐fold more potent than CsA. Because of its effects on multiple activation pathways and because of its potency, mycalamide A could have considerable therapeutic potential.
Serum modulates mast cell responses to IgE antigen stimulation
Authors: Baranes Danny , Lewin Iris , Razin Ehud
Published: March 2007

Serum induces the expression of the fos and jun gene families, which encode the transcription factor AP‐1. Since we previously found that activation of mast cells by IgE‐antigen (Ag) induces the mRNA accumulation of c‐fos, c‐jun, junB and junD proto‐oncogenes, we were prompted to investigate whether serum could affect such accumulation in these cells. In addition, we investigated whether serum could modulate inhibition of DNA synthesis in immunologically stimulated mast cells.

Mast cells, which were cultured in the presence of fetal calf serum (FCS), were characterized by a high proliferation rate and high accumulation of the mRNA of c‐fos, junB and junD proto‐oncogenes. After sustained FCS deprivation both DNA synthesis and the level of c‐fos mRNA were significantly decreased, as expected, whereas the level of c‐jun, junB and junD mRNA were not affected.

As opposed to mast cells which were cultured in the presence of FCS, immunological stimulation of FCS‐deprived cells resulted in DNA synthesis inhibition and an increase in c‐fos expression. The results also show that the level of c‐fos mRNA was increased by either IgE‐Ag or FCS up to a similar level, while these two triggers could not act synergistically to enhance this expression further. Thus, changes in DNA synthesis, induced by FCS, block the ability of the immunological challenge to inhibit mast cell growth and to enhance c‐fos mRNA accumulation.
T cell regulation of collagen‐induced arthritis in mice. II. Immunomodulation of arthritis by cytotoxic T cell hybridomas specific for type II collagen
Authors: Chiocchia Gilles , Boissier Marie‐Christophe , Manoury Bénédicte , Fouriner Catherine
Published: March 2007

Injection of native type II collagen (CII) to susceptible strains of mice (H‐2q) induces a rheumatoid arthritis‐like disease. To study the role of CD8+ T cells in the collagen‐induced arthritis (CIA),we generated CII‐specific T cell hybridomas by fusion of cells from arthritic C3H.Q mice and an AKR thymoma. Two hybrid clones (P3G8 and P2D9) were selected for their ability to lyse syngeneic CH‐pulsed macrophages and recognize different antigenic epitopes in association with Kq molecules. When these T cell clones were irradiated and inoculated into (C3H.Q × AKR)F1 mice 21 days prior to priming with native CII/ complete Freund's adjuvant, the incidence and the duration of CIA were significantly reduced in comparison to groups receiving saline or control T cell hybridoma. Furthermore, both anti‐CII T cell hybridomas were able to attenuate CIA in highly susceptible inbred strains of mice and this suppression was antigen and disease specific. The protective activity seems to require intact cells as neither membrane fractions nor cytosolic preparations of the hybridoma T cells retained the vaccinating activity. Most importantly, one of the hybrid clones (P3G8) had a therapeutic effect on CIA since its administration to arthritic DBA/1 mice on day 30 after priming down‐regulated the ongoing disease. Taken together, these findings suggest that anti‐CII cytotoxic T cell clones can vaccinate against CIA and even reverse the disease.
Dissection of human Langerhans cells' allostimulatory function: The need for an activation step for full development of accessory function
Authors: Pèguet‐Navarro Josette , Dalbiez‐Gauthier Catherine , Dezutter‐Dambuyant Colette , Schmitt Daniel
Published: March 2007

In the present study, we analyzed the mechanism by which human Langerhans cells (LC), the dendritic cells (DC) from epidermis, support the induction of a primary allogeneic T cell response. We reported that paraformaldehyde (PF) fixation completely abrogated the stimulatory property of freshly isolated LC, although the level of major histocompatibility complex (MHC) class II antigen (Ag) expression was unaltered by the fixative. Addition of either interleukin (IL)‐ip and/or IL‐6, during the mixed epidermal cell lymphocyte reaction, failed to restore the proliferative response. By contrast, when human LC were incubated for 3 days in culture medium before fixation, they retained a low but significant allostimulatory capacity. Trypsin treatment of incubated LC before fixation did not impair their function, suggesting that stimulatory activity by fixed incubated LC did not merely reflect a repair of LC membrane after trypsin trauma suffered during epidermal cell (EC) isolation. More interestingly, we found that addition of interferon‐y during LC incubation mediated an enhanced allostimulatory activity by the PF‐fixed LC. Acquisition of allostimulatory property by in vitro activated and fixed LC did not correlate with increased MHC class II Ag expression at the cell surface. By contrast, we showed that ICAM‐1 Ag expression by human LC is involved in this maturation process. Finally, we found that once human LC have been activated, IL‐1β but not IL‐6, could serve as a costimulatory factor in the primary allogeneic T cell response. In conclusion, the data suggest that human LC accessory function is not constitutive but requires an activation step which can be provided by interferon‐y during LC‐T cell interaction.
The generation of a humanized, non‐mitogenic CD3 monoclonal antibody which retains in vitro immunosuppressive properties
Authors: Bolt Sarah , Routledge Edward , Lloyd Irene , Chatenoud Lucienne , Pope Heather , Gorman Scott D. , Clark Mike , Waldmann Herman
Published: March 2007

CD3 antibodies are proven immunosuppressants capable of reversing transplant rejection episodes. Their general application has been limited both by their immunogenicity and, in particular, by the “first‐dose” cytokine‐release syndrome experienced by patients after the initial administration of antibody. We have produced a set of variants of the humanized YTH 12.5 CD3 monoclonal antibody (mAb) (Routledge et al., Eur. J. Immunol. 1991. 21: 2717) bearing different human heavy (H) chain constant regions, with the intention of finding a form of the antibody that is not able to activate T cells. Comparison of the variants having γl, γ2, γ3 and γ4 H chains in a competitive binding assay showed that antibody avidity was not affected by IgG subclass. Using a sensitive indicator of FcR binding activity (the capacity of the CD3 mAb to redirect cytotoxic T cells to kill the monocytic cell line U‐937) we demonstrated a functional hierarchy of γl = γ4 α2 = γ3 mb > γ2. An aglycosyl version of the γl CD3 mAb, produced by site‐directed mutagenesis (Asn297 to Ala), still had considerable activity in this assay (intermediate to the γl and α2 CD3 mAb), albeit at a level approximately 10‐fold lower than that of the parental γl form. When we tested their ablity to stimulate T cell proliferation in vitro in the presence of 5% human serum, all of the wild‐type immunoglobulin isotypes were found to be active, although there were T cell donor‐dependent variations in the extent of the responses. The aglycosyl γl mAb was, however, completely non‐mitogenic in all of ten donors tested, unless the assay was performed in IgG‐free medium. Despite being non‐stimulatory, this mAb was also able to inhibit the mixed lymphocyte reaction responses of both naive and primed T cells. Comparison of the yl and aglycosyl γ1 mAb in an experimental mouse model for CD3 mAb‐induced cytokine release indicated that removal of the carbohydrate moiety from the γl constant region reduced the in vivo tumor necrosis factor‐a response by a factor of at least 16‐fold. These data suggest that the aglycosyl γl CD3 mAb is a promising candidate for immunosuppressive therapy without “first dose” side effects.
CD2/LFA‐3 or LFA‐l/ICAM‐1 but not CD28/B7 interactions can augment cytotoxicity by virus‐specific CD8+ cytotoxic T lymphocytes
Authors: De Waal Malefyt Rene , Verma Sunita , Bejarano Maria‐Theresa , Ranes‐Goldberg Monica , Hill Marc , Spits Hergen
Published: March 2007

It is well established that adhesion molecules are required for interaction between cytotoxic T lymphocytes (CTL) and target cells. Two adhesion pathways, CD2/LFA‐3 and LFA‐l/ICAM‐1 can support cytotoxicity by allospecific CD8+ CTL. In this study, it was investigated whether these adhesion pathways can be utilized independently by influenza virus‐specific HLA‐A2‐restricted CTL clones. It was furthermore examined whether the CD28/B7 pathway can augment virus‐specific CTL activity. To this end, seven CD8+ CTL clones were established that were specific for a peptide encompassing positions 59 to 68 (p[59‐68]) of the influenza virus matrix protein. These seven clones apparently originated from different precursors, as they utilized different Vα and Vβ or Jα gene segments. Six of seven clones were able to lyse mouse L cells co‐transfected with HLA‐A2 and either LFA‐3 (LA2/LFA‐3) or ICAM‐1 (LA2/ICAM‐1) in the presence of p[59‐68] but did not lyse Lcells that expressed only HLA‐A2 and peptide. Three of the most cytotoxic clones were selected for further analysis. The cytotoxicity of the clones against LA2/LFA‐3 cells was blocked by anti‐LFA‐3 and anti‐CD2 monoclonal antibodies (mAb), while these antibodies did not affect cytotoxicity against LA2/ICAM‐1 cells. Likewise, the activity against LA2/ICAM‐1 was blocked only by anti‐LFA‐1 and ICAM‐1 mAb. These clones were unable to lyse Lcells co‐transfected with HLA‐A2 and B7, the counter structure of CD28, despite the fact that these clones expressed CD28. These data indicate that CD8+ virus‐specific CTL can utilize either the CD2/LFA‐3 or the LFA‐l/ICAM‐1 adhesion pathway. The CD28/B7 pathway seems not to be required for cytotoxicity mediated by activated virus‐specific CTL.
The set of naturally processed peptides displayed by DR molecules is tuned by polymorphism of residue 86
Authors: Demotz Stéphane , Barbey Catherine , Corradin Giampietro , Amoroso Antonio , Lanzavecchia Antonio
Published: March 2007

The response to tetanus toxoid (TT) was studied in immune donors that carry two alleles of DR5 that differ only at DRβ residue 86: DRB1*1101 (G86, abbreviated 1101) and DRB1*1104 (V86, abbreviated 1104). A large number of TT‐specific T cell clones was isolated and the epitopes recognized in association with 1101 and 1104 were mapped. We found that two epitopes (p2 and p32) can be recognized in association with both 1101 and 1104 while three epitopes (p23, p27 and p30) are recognized in association with 1101, but never in association with 1104.

The sets of naturally processed self peptides displayed by 1101 and 1104 were characterized using alloreactive T cell clones. We found that all 1104 alloreactive clones recognize both 1104 and 1101, while ˜30% of the alloreactive 1101 clones fail to recognize 1104. Thus it is apparent that both naturally processed TTand self peptides displayed on 1104 molecules represent a fraction of those displayed on 1101 molecules.

The mechanism responsible for this differential presentation was investigated by comparing the capacity of 1101 and 1104 antigen‐presenting cells to present TTor synthetic peptides to specific T cells and by measuring the binding of these peptides to DR molecules. Three sets of results suggest that V86 acts as a constraint to the binding of naturally processed peptides: (i) all 1104‐restricted or alloreactive T cell clones recognize TT‐ or allo‐epitopes presented by 1101 as well, thus ruling out a major effect of V86 as a residue determining T cell restriction specificity; (ii) presentation of naturally processed peptides correlates in general with the capacity of long synthetic peptides to bind to 1101 or 1104 and (iii) while the naturally processed p30 epitope discriminates between 1101 and 1104, a short synthetic peptide binds equally well to and is comparably recognized in association with both 1101 and 1104.

Taken together these results suggest that the natural polymorphism at residue 86 might be a molecular switch that finely tunes the complexity of the peptide collection presented on DR molecules.
Protective immunity induced in rat schistosomiasis by a single dose of the Sm28GST recombinant antigen: Effector mechanisms involving IgE and IgA antibodies
Authors: Grezel Delphine , Capron Monique , Grzych Jean‐Marie , Fontaine Josette , Lecocq Jean‐Pierre , Capron Andre
Published: March 2007

Rats immunized by a single dose of the recombinant Sm28GST antigen, using either aluminium hydroxide or Bacillus Calmette‐GuSrin adjuvant, were significantly protected (up to 59% reduction in worm burden) against a challenge infection with Schistosoma mansoni cercariae. A follow‐up study of the humoral response revealed the presence of high levels of IgE and IgA antibodies together with specific IgG. Sera from once Sm28GST‐immunized rats induced a cytotoxic response for schistosomula targets in the presence of normal rat eosinophils, similar to the one induced by sera from twice immunized rats. Depletion or competition studies indicated the participation of both IgE and IgA antibodies in eosinophil‐dependent cytotoxicity mechanisms.

These results suggest the existence, in immunized rats exhibiting protection against schistosomiasis, of an original effector mechanism implying eosinophils and IgA antibodies, together with documented effector mechanisms involving IgE and eosinophils. In addition, they raise questions concerning the role of IgA antibodies in schistosomiasis.
Cross‐linking of surface IgM, but not surface IgD receptors, by soluble monoclonal antibodies primes murine B cells to secrete immunoglobulin in response to lymphokines
Authors: Phillips Catherine , Klaus Gerry G. B.
Published: March 2007

We have previously reported the development of a two‐step culture system in which soluble anti‐μ monoclonal antibodies prime small resting murine B cells to secrete immunoglobulin (Ig) in response to restimulation with a mixture of interleukin‐4 (IL‐4) and IL‐5. Here we have extended these studies to investigate the effects of engaging surface IgD (sIgD).We find that, unlike anti‐μ, three different anti‐5 monoclonal antibodies did not prime B cells to secrete Ig. In addition, these anti‐δ antibodies inhibited anti‐μ‐stimulated priming for Ig secretion, while enhancing DNA synthesis in response to anti‐μ,. Furthermore, anti‐δ antibodies still inhibited anti‐μ‐induced priming when added 24‐48 h after anti‐μ. These results therefore suggest that triggering of sIgD on B cells induces a dominant inhibitory signal which is not necessarily dependent upon co‐ligation of sIgM and sIgD receptors. In addition, these findings raise the possibility that ligating sIgM or sIgD receptors on mature B cells in the absence of Tcell help, may produce different downstream effects.
The T cell receptor αβ V‐J shuffling shows lack of autonomy between the combining site and the constant domain of the receptor chains
Authors: Casorati Giulia , Traunecker André , Karjalainen Klaus
Published: March 2007

In order to assess the structural independence of the T cell receptor (TCR) combining site from the rest of the molecule we have generated two recombinant chains consisting of aTCR V‐J α region linked to the Cβ and aTCR V‐J β linked to the Cα. If the V and C domains of the TCR form independent domains, as has been shown for the Ig molecules, we would expect to obtain a functional chimeric TCR. Interestingly, it was found that the shuffled molecules are produced intracellularly in T cell hybridomas, but are not expressed on the cell surface. To explain this failure of the shuffled molecules we propose that the TCR has a more compact structure, compared to the Ig, and that it is indispensable to keep a longitudinal inter‐domain contact between the V‐J and C portion to have a functional molecule.
A promoter element of the human serine esterase granzyme B gene controls specific transcription in activated T cells
Authors: Haddad Patrick , Wargnier Alain , Bourge Jean‐François , Sasportes Maryline , Paul Pascale
Published: March 2007

The human granzyme B gene encodes a serine protease expressed specifically in cytoplasmic granules of cytotoxic T lymphocytes, released upon effector‐target cell interaction. Previous studies have shown that granzyme B mRNA was induced in T lymphocytes after antigenic or mitogenic stimulation. To study the regulation of human granzyme B gene expression during lymphocyte activation we analyzed its 5′ flanking region using chloramphenicol acetyl transferase (CAT) reporter gene constructs. We show that a 208‐bp fragment (‐148 to +60) containing an NF‐AT (nuclear factor of activated T cells)‐binding site promotes CAT expression in phytohemagglutinin‐activated T lymphocytes, in immobilized monoclonal anti‐CD3 antibody‐activated Jurkat T cell line while it is inactive in unstimulated PEER and Jurkat T cells lines or B Epstein‐Barr virus‐transformed cell lines.
Expression and control of the natural autoreactive IgG repertoire in normal human serum
Authors: Hurez Vincent , Kaveri Srinivas‐ V. , Kazatchkine Michel D.
Published: March 2007

We have investigated the autoreactive repertoire expressed by serum IgG of healthy individuals of various age groups using a large panel of self antigens. Natural IgG autoantibodies against all self antigens of the panel were found in the purified IgG fraction of the serum of all donors that were tested. The mean binding activity to self antigens of IgG of pregnant women was higher than that of IgG purified from the serum of infants, young adults and aged individuals. No increase in IgG autoreactivity was observed with aging neither in the purified IgG fraction of serum nor in whole serum. Whereas autoantibody activity was easily detectable in purified IgG, it was low in serum. No difference was observed, however, between the binding activity of purified IgG and of IgG in serum in the case of foreign antigens nor in the case of anti‐thyroglobulin autoantibodies of patients with Hashimoto's thyroiditis. Purified IgM from normal serum bound to F(ab')2 fragments of autologous IgG in a dose‐dependent fashion and inhibited the binding of autologous IgG to self antigens. Our results thus indicate that autologous IgM contributes to regulate expression of the natural IgG autoreactive repertoire through V region‐dependent interactions, resulting in low levels of IgG autoreactivity in serum under physiological conditions.
Normal T cell receptor Vβ usage in a primary immunodeficiency associated with HLA class II deficiency
Authors: Rieux‐Laucat Frédéric , le Deist Franchise , Selz Franchise , And Alain Fischer , de Villartay Jean‐Pierre
Published: March 2007

The human Tcell receptor was studied using an anchored‐polymerase chain reaction (A‐PCR) and hybridization with Vβ‐specific oligonucleotide probes, together with the few anti‐Vβ monoclonal antibodies (mAb) available. After confirming the semiquantitative and reproducible nature of the A‐PCR technique, we assessed the complete Vβ repertoire in sorted CD4+ and CD8+ lymphocyte populations from three normal donors. These experiments confirmed the absence of Vβ‐restricted deletions in human peripheral cells, in contrast to several mouse strains. This feature makes it difficult to study negative selection in man, given the apparent absence of an endogenous superantigen corresponding to the Mis system in the mouse. To investigate human Vβ repertoire shaping, we studied VP usage in CD4+ and CD8+ T cells from children with an inherited immunodeficiency characterized by defective expression of human leukocyte antigen class II molecules. An initial study using anti‐Vβ monoclonal antibodies failed to show significant abnormalities in Vβ usage. Four patients analyzed using the A‐PCR method all had a polyclonal Vβ repertoire, suggesting normal positive selection and raising questions as to the importance of VP major histocompatibility complex (MHC) interactions and the role of thymic MHC density in shaping the Vβ repertoire.
Cloning and expression of murine CD27: comparison with 4‐1BB, another lymphocyte‐specific member of the nerve growth factor receptor family
Authors: Gravestein Loes A. , Blom Bianca , Nolten Linda A. , de Vries Evert , Horst Gerda Van Der , Ossendorp Ferry , And Jannie Borst , Loenen Wil A. M.
Published: March 2007

CD27 is a member of the nerve growth factor receptor family, that includes two types of tumor necrosis factor receptor, CD40 and Fas/Apo‐1. Human CD27 has been found only on lymphocytes. In T cells, its expression strongly increases in a transient fashion upon antigenic stimulation, suggesting that CD27 plays a role during T cell activation. To analyze the function of CD27, we have identified the murine CD27 at the cDNA and protein level. Murine CD27 shows an identity of 65% compared with human CD27. The amino‐terminal cysteine‐rich region, i.e. the putative ligand‐binding domain, and the carboxy‐terminal part of the cytoplasmic domain are approximately 80% identical in man and mouse. Murine CD27 has 29% identity to 4‐IBB, another lymphocyte‐specific member of the receptor family defined only at the cDNA level. Murine CD27 and 4‐1BB have 39% homology in the cysteine‐rich domain and share a conserved region in the cytoplasmic tail. Expression studies identified murine CD27 mRNA in thymus and spleen, but not in non‐lymphoid tissues, while 4‐IBB mRNA was not detected in any tissue tested. In resting T cells, only murine CD27 mRNA was found, while in activated T cells murine CD27 as well as 4‐1BB were present at high levels. Murine CD27 and 4‐1BB mRNA are expressed with different kinetics during T cell activation, suggesting that these molecules play different roles in this process. Peptide antisera identified murine CD27 as a 45‐kDa protein on thymocytes and activated T cells, while 4‐1BB was precipitated as a 35‐40‐kDa protein from activated T cells.