bluuebunny/crossref_metadata_2025 huggingface dataset:
Recognition of E‐cadherin on epithelial cells by the mucosal T cell integrin αM290β7 (αEβ7)
Authors: Karecla Paula I. , Bowden Simon J. , Green Sally J. , Kilshaw Peter J.
Published: March 2007

The integrin αM290β7 on the surface of a T cell hybridoma, MTC‐1, mediated adhesion of these cells to the mouse epithelial cell line CMT93. This interaction was critically dependent on the presence of divalent cations; Mn2+ strongly promoted adhesion, Ca2+ was ineffective and Mg2+ gave intermediate results. Antibodies to molecules on the surface of CMT93 cells were tested for inhibition of adhesion. One monoclonal antibody (mAb) against E‐cadherin, ECCD‐2, was found to have significant inhibitory activity. Other mAb to E‐cadherin and antibodies to other molecules had no effect. To show that inhibition by ECCD‐2 was specific for adhesion mediated by αM290β7, MTC‐1 cells were induced to adhere to CMT93 via the LFA‐1/ICAM‐1 pathway. For this purpose, the epithelial cells were treated with interferon‐γ and tumor necrosis factor‐α to induce ICAM‐1 expression and, in addition, αM290β7 on MTC‐1 cells was down‐regulated by culturing the cells in the absence of transforming growth factor β. Under these circumstances adhesion of MTC‐1 cells to CMT93 was inhibited by an antibody to LFA‐1 but not by ECCD‐2. Transfection of mouse L cells with cDNA for mouse E‐cadherin enabled MTC‐1 cells to adhere to them through the αM290β7 integrin; this interaction was inhibited both by ECCD‐2 and by blocking antibody against the integrin. These data strongly suggest that E‐cadherin is a principal ligand for αM290β7.
CD40 ligand is functionally expressed on human eosinophils
Authors: Gauchat Jean‐François , Henchoz Sybille , Fattah Dilnya , Mazzei Gonzalo , Aubry Jean‐Pierre , Jomotte Therese , Dash Laura , Page Kevin , Solari Roberto , Aldebert Delphine , Capron Monique , Dahinden Clemens , Bonnefoy Jean‐Yves
Published: March 2007

CD40 ligand (CD40L), a surface molecule which can be expressed by T cells, mast cells and basophils, has been shown to be involved in the control of B cell proliferation, immunoglobulin class switching as well as in the activation of monocytes and T cells. We demonstrate that CD40L can also be expressed constitutively by eosinophils from an hypereosinophilic patient or, upon activation, by the eosinophilic cell line EOL‐3 and normal blood eosinophils. Eosinophils were able to induce, in conjunction with IL‐4, CD40L‐dependent B cell proliferation in vitro. These results suggest that CD40L could play a role in the inflammatory processes during which eosinophil infiltration and activation are observed.
More newly formed T than B lymphocytes leave the intestinal mucosa via lymphatics
Authors: Rothkötter Hermann J. , Hriesik Claudia , Pabst Reinhard
Published: March 2007

Many lymphocytes are produced in the intestinal mucosa, especially in the Peyer's patches. These newly formed lymphoid cells leave the gut wall, undergo further maturation and many reach the lamina propria of the intestinal mucosa where they function as effector and regulator cells of the intestinal immune response. However, the number and subset composition of these wall are not known. Therefore, the intestinal lymph duct was cannulated in eight minipigs, in which the mesenteric lymph nodes had been removed 3 months earlier. Thus, it was possible to obtain all lymphocytes leaving the intestinal mucosa including the Peyer's patches via lymphatics. The hourly output of lymphocyte subsets was examined over the course of 93 h. The percentage and the absolute numbers of newly formed T cells (CD2+, CD8+) and B cells (IgA+, IgM+) were determined by examining the incorporation of the DNA precursor bromodeoxyuridine. After a single i.v. bromodeoxyuridine injection 8.5% of the T, 55% of the IgA+ and 25% of the IgM+ cells were labeled. In absolute numbers (1.9 ± 0.7) × 106 newly formed T cells, (0.4 ± 0.3) × 106 IgA+ cells and (0.5 ± 0.4) × 106 IgM+ cells emigrated from the gut wall per hour. Both T and B lymphocyte subpopulations that are produced in the intestinal mucosa leave the gut wall via lymphatics; interestingly, the T cells outnumber the B cells. Obviously the induction and maintenance of mucosal immunity depend to a large extent on the function of newly formed T lymphocytes emigrating from the Peyer's patches and/or from the mucosa without Peyer's patches.
Gene structure and chromosomal localization of the mouse homologue of rat OX40 protein
Authors: Birkeland Marian L. , Copeland Neal G. , Gilbert Debra J. , Jenkins Nancy A. , Barclay A. Neil
Published: March 2007

The OX40 protein is expressed only on activated rat CD4+ T blasts and is a member of a superfamily of cell surface molecules which includes CD40, CD30, CD95 (Fas), CD27, 4‐1BB antigens and the receptors for tumor necrosis factor (TNF) and nerve growth factor (NGF). The proteins of this group are related to each other by having three to six repeats of a cysteine‐rich sequence in their extracellular domains. Members of this family of receptors have also been shown to bind to ligands which are structurally related to TNF.

The mouse homologue of the rat OX40 protein was cloned at the cDNA and genomic levels. The gene structure shows that there are several intron/exon borders shared between OX40 and CD27, CD40, TNF receptor type I, CD95 and 4‐1BB genes. This group of genes is less closely related structurally to the gene structure of the NGF receptor. The gene encoding murine OX40 has been placed on mouse chromosome 4, in an area which contains the genes for TNF receptor type II and 4‐1BB, and is syntenic with a region of human chromosome 1 which contains human TNF receptor type II, OX40, and CD30 genes.
Induction in mice of serum IgE levels after treatment with anti‐mouse IgD antibodies is preceded by differential modulation of tissue cytokine gene transcription
Authors: Kricek Franz , Ruf Christine , Žunić Melita , De Jong Gerard , Dukor Peter , Bahr George M.
Published: March 2007

Injection of mice with purified goat anti‐mouse IgD (GAMD) leads to an interleukin (IL)‐4‐dependent increase of serum IgE levels. Challenge of GAMD‐primed mice with goat IgG (GIG) initiates a secondary immune response with elevated serum IgE. In this report, kinetic cytokine transcript profiles of murine lymphoid tissues in response to primary i.v. GAMD treatment, as well as GIG challenge are presented. For the first time, gene transcription patterns of the recently described cytokines IL‐12 and IL‐13 are shown and compared with the corresponding patterns for other cytokine genes involved in IgE regulation, i.e. IL‐4, and interferon (IFN)‐γ. After GAMD injection, two groups of induction profiles were observed in spleen, mesenteric lymph nodes and Peyer's patches: while IL‐4 and IL‐12p35 gene transcription was strongly enhanced, IFN‐γ, IL‐12p40 and IL‐13 mRNA were only moderately induced. Generally, maximal mRNA induction was measured on days 3 to 4 after GAMD treatment. The data demonstrate a clear‐cut difference between the IL‐4 and IL‐13 response on the transcriptional level although both gene products show similar biological activities. The cytokine mRNA profiles support the assumption of IL‐4 playing the central role in generating an IgE response. However, they do not reflect a strict Th1 versus Th2 cytokine gene transcription pattern but rather point towards a concerted action of various, partially antagonizing cytokines with respect to the regulation of IgE synthesis. IL‐12 may, possibly via stimulation of IFN‐γ synthesis, represent a counterbalancing factor in the IL‐4‐mediated IgE response.
Surface expression of β2‐microglobulin‐associated thymus‐leukemia antigen is independent of TAP2
Authors: Rodgers John R. , Mehta Vidya , Cook Richard G.
Published: March 2007

Mouse thymus‐leukemia antigen (TL), like other major histocompatibility complex (MHC) class I‐b antigens, displays signs of a specialized function. It is normally expressed at high levels on immature thymocytes and at moderate levels on gut epithelium and activated mature T cells. A promoter/enhancer region unique among class I genes accounts for this narrow range of tissue distribution. Like most other class I molecules, TL is dependent upon endogenous β2‐microglobulin (β2m) for transport to the surface. However, here we show that unlike most other MHC class I molecules, TL is expressed efficiently in the absence of functional transporter associated with antigen processing subunit 2 (TAP2). A putative fourth TLa gene cloned from A.SL1 cells was expressed in RMA and RMA‐S cells. In bulk transformants, TL expression is higher in TAP2− RMA‐S cells than in wild‐type RMA cells, and is not elevated by incubation at reduced temperatures or exposure to exogenous β2m. Analysis of immunoprecipitasted molecules by polyacrylamide gel electrophoresis in the presence of sodium dodecyl sulfate indicates that TL is processed normally in RMA‐S cells and is associated with β2m both intracellularly and at the cell surface. However, TL heavy chains expressed on the cell surface in the absence of TAP2 are cleaved to a predominant 38 kDa fragment, presumably the result of an altered conformation that renders TL more susceptible to proteolysis. These results suggest that while TL may normally acquire TAP2‐dependent peptides, this class I‐b molecule does not require them for efficient export to, and stable expression at the cell surface.
Antibody response against poly (Glu60Ala30Tyr10) terpolymer and bacterial levan in ϰ‐deficient mice
Authors: Pricop Luminita , Hatakeyama Akira , Moran Tom , Bona Constantin
Published: March 2007

In murine species, the ϰ+‐bearing immunoglobulins‐dominate the antibody (Ab) repertoire with a ϰ/λ ratio of 95 : 5. The aim of the present study is to investigate the characteristics of the antibody response in ϰ‐deficient (K −/−) mice immunized with a T‐dependent synthetic antigen, poly (Glu60Ala30Tyr10) (GAT) and a T‐independent antigen, bacterial levan (BL). K −/− mice were obtained by targeted deletion of the JϰCϰ gene segments. In response to GAT, K −/− mice respond by producing increasing amounts of anti‐GAT Igλ1 and Igλ2 in the primary as well as secondary response, although anti‐GAT specific monoclonal antibodies (mAb) raised in K −/− mice are mostly of IgM isotype. The GAT public idiotype, GATIdX, present on all GAT‐specific Ab bearing x light chain, is not detected in the sera of K −/− mice or on any of the anti‐GATλ1 mAb. In response to BL, the amount of Igϰ1+ Ab in K −/− mice is comparable to the amount of Igϰ+ Ab in normal mice. However, λ2+ Ab are detected neither in wild‐type nor in K −/− mice. Like ϰ+ Ab, the majority of λ1+ mAb are specific for β2–6 fructosan present in BL and rye levan and, to some extent, express the BL‐specific idiotype, A48ld. Our results show that important compensatory mechanisms occur in ϰ‐deficient mice, restoring their ability to mount immune responses against a variety of T‐dependent and T‐independent antigens by the alternative usage of the clonally restricted λ repertoire.
Serial killing by cytotoxic T lymphocytes: T cell receptor triggers degranulation, re‐filling of the lytic granules and secretion of lytic proteins via a non‐granule pathway
Authors: Isaaz Sylvie , Baetz Kristin , Olsen Kristin , Podack Eckhard , Griffiths Gillian M.
Published: March 2007

CD8+ cytotoxic T lymphocyte (CTL) clones begin to synthesize the lytic proteins granzyme A, granzyme B and perforin after stimulation with allogeneic target cells. The lytic proteins are stored in the secretory granules which are released after cross‐linking of the T cell receptor (TcR) upon target cell recognition. During lytic granule biogenesis granzyme A protein synthesis can be detected between 2 and 10 days after allogeneic stimulation of the CTL. Although granzyme A is stored in the lytic granules over this period, the majority of granzyme A synthesized is secreted directly from the CTL.

TcR triggering of degranulation also results in new synthesis of the lytic proteins, which can be inhibited by cycloheximide (CHX). Some of the newly synthesized lytic proteins can be stored in the cell and refill the granules. But up to one third of granzymes A and B can be secreted directly from the CTL via the constitutive secretory pathway as shown by granzyme A enzymatic activity and immunoblots of secreted granzyme B, where one third of the protein fails to acquire the granule targeting signal. Perforin is also secreted via the constitutive pathway, both from the natural killer cell line, YT, and from CTL clones after TcR cross‐linking. Constitutive secretion of the lytic proteins can be blocked by both CHX and brefeldin A (BFA). While BFA does not affect the directional killing of recognized targets, it abrogates bystander killing, indicating that bystander killing arises from newly synthesized lytic proteins delivered via a non‐granule route.

These results demonstrate that the perforin/granzyme‐mediated lytic pathway can be maintained while CTL kill multiple targets. We show that CTL not only re‐fill their granules during killing, but also secrete lytic proteins via a non‐granule‐mediated pathway.
Expression and tissue localization of donor‐specific complement C3 synthesized in human renal allografts
Authors: Andrews Peter A. , Finn Jacqueline E. , Lloyd Clare M. , Zhou Wuding , Mathieson Peter W. , Sacks Steven H.
Published: March 2007

Recent evidence suggests that the third component of complement, C3, is synthesized in renal tissue, and that increased C3 synthesis occurs in allograft rejection and immune complex‐mediated nephritis. However, it is unclear whether intrinsic renal cells or migratory cells in the inflammatory infiltrate, possibly of recipient bone marrow origin, are the source of the C3 detected. This was investigated by determining the C3 allotypes of mRNA and protein produced by transplanted human kidney. Twenty donor‐recipient pairs were examined, of which nine pairs had C3 allotypes that were informatively mismatched at the C3 F/S locus. Reverse transcriptase polymerase chain reaction (RT‐PCR) followed by amplification refractory mutation system analysis showed intracellular donor‐specific mRNA expression in six of these nine cases, at up to 61 days post‐transplantation. Nested PCR reactions and the size of PCR products excluded contamination by genomic DNA. Allotype‐specific staining of frozen sections of renal cortex demonstrated donor‐derived C3 protein in both glomeruli and tubules of all biopsies examined, in a predominantly tubular distribution. These results imply that at least some of the pro‐inflammatory effects of complement arise from intrinsic tissue synthesis of donor C3, and that this may represent a previously unrecognized source of tissue injury. The occurrence of local synthesis of C3 of donor allotype may have functional implications related to C3 allotype, and may also be relevant to strategies to inhibit intrarenal complement‐mediated injury.
The protein defective in X‐linked agammaglobulinemia, Bruton's tyrosine kinase, shows increased autophosphorylation activity in vitro when isolated from cells in which the B cell receptor has been cross‐linked
Authors: Hinshelwood Steve , Lovering Ruth C. , Genevier Helen C. , Levinsky Roland J. , Kinnon Christine
Published: March 2007

X‐linked agammaglobulinemia is a primary inherited immunodeficiency resulting in a lack of or dramatic reduction in the number of mature B lymphocytes and, thus, greatly reduced levels of serum immunoglobulin. The defect results from mutations in the gene for Bruton's tyrosine kinase (Btk). Using rabbit antisera generated against Btk, we have demonstrated an increase in the level of in vitro kinase activity present in anti‐Btk immunoprecipitates from B cells following stimulation with anti‐immunoglobulin antibody. This increase in immune complex kinase activity is detectable 1 to 2 min following stimulation and remains elevated for over 30 min. A similar increase was not seen with two late pre‐B cell lines investigated in the same way. This stimulation of activity may suggest a role for Btk in signalling through the B cell receptor or associated proteins, in mature B cells.
Depletion of CD8+ T cells suppresses the development of experimental autoimmune myasthenia gravis in Lewis rats
Authors: Zhang Guang‐Xian , Ma Cun‐Gen , Xiao Bao‐Guo , Bakhiet Moiz , Link Hans , Olsson Tomas
Published: March 2007

To understand the role of CD8+ T cells in experimental autoimmune myasthenia gravis (EAMG), CD8+ T cells were depleted by injecting a monoclonal anti‐rat CD8 antibody (OX8) into Lewis rats immunized with Torpedo acetylcholine receptor (AChR) in complete Freund's adjuvant (CFA). CD8‐depleted EAMG rats showed strikingly less muscle weakness and lower anti‐AChR IgG antibody levels compared to Hy2‐15‐injected control EAMG rats. Moreover, the numbers of AChR‐specific IgG antibody‐secreting cells, AChR‐reactive interferony‐γ‐secreting T helper type 1‐like cells and lymphocyte proliferation to AChR were lower in the CD8‐depleted group than in control EAMG rats. These differences were significant among mononuclear cells from inguinal and popliteal lymph nodes, mesenteric lymph nodes and spleen, but not from thymus when examined 3, 5 and 7 weeks post‐immunization. We suggest that CD8+ T cells are involved in the induction and persistance of EAMG by directly or indirectly affecting AChR‐reactive T cells and anti‐AChR IgG antibody‐secreting cells.
Substitutions at the Glu62 residue of human interleukin‐2 differentially affect its binding to the α chain and the βγ complex of the interleukin‐2 receptor
Authors: Wang Zhiyong , Zheng Zhongcheng , Sun Lanyin , Liu Xinyuan
Published: March 2007

Human interleukin‐2 (IL‐2) α helix B is more conserved than the whole molecule, but has been less studied than other α helices of IL‐2. Using site‐directed mutagenesis, several IL‐2 mutants in this helix were obtained. We found that the IL‐2 mutant containing Leu at position 62 (Leu62‐IL‐2) loses its ability to bind IL‐2 receptor subunit α (IL‐2Rα), but retains binding affinity to IL‐2R subunit βγ as well as some bioactivity; nevertheless, another substitution at the same residue, Arg62 IL‐2, loses its binding ability to both IL‐2Rα and IL‐2Rβγ, and can no longer stimulate IL‐2‐dependent cell growth, showing that Glu62 not only takes part in IL‐2Rα binding, but can also affect IL‐2 binding to IL‐2Rβγ. In this regard, Glu62 may be a key site in the IL‐2/IL‐2Rα interaction, and can facilitate IL‐2R ternary‐complex formation, leading to IL‐2Rα‐mediated, IL‐2‐stimulated signal transduction.
Human T helper cells specific for HIV reverse transcriptase: possible role in intrastructural help for HIV envelope‐specific antibodies
Authors: Manca Fabrizio , Fenoglio Daniela , Valle Maria T. , Li Pira Giuseppina , Kunkl Annalisa , Balderas Robert S. , Baccala Roberto G. , Kono Dwight H. , Ferraris Anna , Saverino Daniele , Lancia Flavia , Lozzi Luisa , Theofilopoulos Argyrios N.
Published: March 2007

Cooperation between B cells specific for an antigen exposed on a viral structure and T helper (Th) cells specific for an internal antigen, as demonstrated with influenza, hepatitis B and rabies viruses, has been termed intrastructural help. Th cells specific for internal proteins of HIV, which are much less mutated than its exposed antigens, may be valuable in vaccine design against this virus. We investigated the human Th repertoire specific for the core HIV antigen reverse transcriptase (p66), and determined whether these cells could be candidate intrastructural T helpers. CD4+ T lines and clones were generated from non‐immune individuals by stimulation with p66‐pulsed antigen‐presenting cells (APC). Specific lines were obtained with p66 from 19 out of 21 (90%) of these individuals, vs. 7 out of 29 (24%) with gp120. Diverse epitopes were recognized by different individuals, and various Vβ genes were used by these clones. Clones using the same Vβ genes were of diverse origin, according to VDJ region sequence. Of these lines 45% responded to p66 in the context of HIV virions. Moreover, p66‐specific clones could respond to APC that had internalized HIV complexed with envelope‐specific monoclonal antibodies, suggesting that p66‐specific Th cells may participate in intrastructural help. These studies indicate that p66‐specific Th cells are detectable in vitro in most naive individuals and exhibit clonal heterogeneity, and that the majority recognize an HIV conserved antigen. They respond to p66 following processing of whole virions and are clearly candidates for intrastructural help. If confirmed in vivo, p66 should be included among vaccine candidates investigated to optimize the anti‐HIV Th response.
Signaling via major histocompatibility complex class II molecules and antigen receptors enhances the B cell response to gp39/CD40 ligand
Authors: Bishop Gail A. , Warren Wendy D. , Berton Michael T.
Published: March 2007

Activated T cells induce proliferation and differentiation of resting B cells in vitro through their CD40 molecules and lymphokine receptors. However, despite constitutive B cell expression of CD40 and lymphokine receptors, widespread nonspecific polyclonal B cell activation by activated T cells is seldom observed in vivo. The present study was designed to test the hypothesis that signals delivered via the B cell antigen (Ag) receptor (membrane immunoglobulin, mIg) and major histocompatibility complex (MHC) class II molecules enhance B cell responsiveness to CD40‐mediated signals, providing specificity to the Ag‐nonspecific, MHC‐unrestricted CD40 signal. To test this hypothesis, both an Ag‐specific mouse B cell clone CH12.LX, and freshly isolated resting splenic B cells were cultured with either soluble or membrane‐bound forms of the T cell ligand for CD40 (CD40L), in the presence or absence of additional signals provided by Ag or anti‐IgM, interleukin‐4, and class II‐specific monoclonal antibody (mAb). Differentiation of CH12.LX cells and proliferation of splenic B cells in response to both forms of CD40L was greatly enhanced by exposure to mIg‐mediated signals, with greatest enhancement seen when cells were cultured with Ag prior to receiving other signals. Response to CD40L was further enhanced by concurrent culture with class II‐specific, but not class I‐specific mAb. Enhancement was greatest at limiting concentrations of CD40L. The ability of class II MHC‐mediated signals to enhance Ag‐specific B cell responsiveness to CD40‐mediated signaling may selectively promote the activation of B cell clones capable of cognate interactions with helper T cells.
Modulation of CD4 lateral interaction with lymphocyte surface molecules induced by HIV‐1 gp120
Authors: Dianzani Umberto , Bragardo Manuela , Buonfiglio Donatella , Redoglia Valter , Funaro Ada , Portoles Pilar , Rojo José , Malavasi Fabio , Pileri Alessandro
Published: March 2007

CD4, a lymphocyte surface glycoprotein, serves as co‐receptor for antigen with the T cell receptor (TCR). It is also the lymphocyte receptor for HIV by binding the gp120 viral envelope protein. Interaction of gp120 with CD4 is crucial for viral infection, but is not sufficient to allow viral entry into cells. Recombinant gp120 alters CD4+ T cell responsiveness to activation stimuli. To express its co‐receptor function fully, CD4 must be laterally associated with the TCR and CD45 to form multi‐receptor complexes competent to transduce potent activation signals. Here, we examine the possibility that gp120/CD4 binding alters lateral associations of CD4 with other lymphocyte surface molecules, and that assembly of abnormal multi‐molecular complexes is involved in the gp120‐induced CD4+ T cell dysfunction and in viral entry. In the absence of gp120, CD4 displayed high association with CD3, CD5, CD45RC, CD25, CD28, CD44, and CD53; weak association with CD2, CD38, CD45RB, CD62L, and CD26; and no association with CD45RA, CD45RO, CD11b, CD11a, CD54, CD7, CD48, CD98, CD59, CD55, HLA class I and class II molecules. Treatment with gp120 significantly increased CD4 association with CD3, CD45RA, CD45RB, CD59, CD38, CD26, and HLA class I, and decreased that with CD45RC. Specificity of these results were assessed at various levels. First, gp120 did not influence lateral associations displayed by other molecules, such as HLA class II. Second, the Leu3 mAb, which binds CD4 on a site overlapping the gp120 binding site, did not elicit the same CD4 lateral associations as gp120, and finally, a direct gp120/CD4 interaction was needed to induce the lateral associations, as shown by the observation that blocking the gp120/CD4 binding by the Leu3 mAb inhibited the gp120‐induced associations. These results can be interpreted in several ways. gp120/CD4 interaction could trigger an inside‐out signal responsible for the associations, or gp120 could induce steric modifications of CD4 that increase its affinity for the associating molecules. Alternatively, these molecules may interact directly with gp120, bridging them with CD4. It is also possible that the associations may be mediated by additional components, interacting with both gp120 and the associating surface molecule. The last hypothesis is likely for CD59, whose gp120‐induced association with CD4 required the presence of serum in the co‐capping assay. Since both CD59 and gp120 bind complement, the observed association could be mediated by complement components.
Down‐regulation of class II major histocompatibility complex molecules on antigen presenting cells after interaction with helper T cells
Authors: Vidović Damir , Falcioni Fiorenza , Bolin David R. , Nagy Zoltan A.
Published: March 2007

The recognition of antigenic peptides by CD4+ helper T cells is demonstrated here to result in a dramatic (up to 90%) decrease in expression of major histocompatibility complex (MHC) class II molecules on the surface of antigen‐presenting cells (APC). The reduction is selective to the class II isotype presenting the antigen, but if affects both allelic forms of the same isotype in heterozygous APC. The observed MHC down‐regulation requires a specific T cell receptor‐peptide‐class II interaction, a direct contact between T cell and APC, and the involvement of CD2 molecules. These findings have important implications for the regulation of immune response, self tolerance, and autoimmunity.
Developmentally regulated expression of P‐glycoprotein (multidrug resistance) activity in mouse thymocytes
Authors: MacDonald H. Robson , Bommhardt Ursula , Cerottini Jean‐Charles
Published: March 2007

P‐glycoprotein (P‐gly) is the transmembrane efflux pump responsible for multidrug resistance in tumor cells. The activity of P‐gly in mature peripheral lymphocytes is lineage specific, with CD8+ T cells and natural killer (NK) cells expressing high levels as compared to CD4+ T cells and B cells. We have now investigated P‐gly activity in immature and mature subsets of mouse thymocytes. Our data indicate that P‐gly activity is undetectable in immature CD4−8− and CD4+8+ thymocyte subsets. Among mature thymocytes, P‐gly activity is absent in the CD4+ subset but present in the more mature (HSAlow) fraction of CD8+ cells. Furthermore, while thymic CD4−8− T cell receptor (TCR) γδ cells have little P‐gly activity, a minor subset of CD4−8− or CD4+ TCR αβ+ thymocytes bearing the NK1.1 surface marker expresses high levels of P‐gly activity. Collectively, our results indicate that P‐gly activity arises late during thymus development and is expressed in a lineage‐specific fashion.
Non‐exclusive Fas control and age dependence of viral superantigen‐induced clonal deletion in lupus‐prone mice
Authors: Papiernik Martine , Pontoux Christiane , Golstein Pierre
Published: March 2007

To investigate the role of Fas in the induction of tolerance by viral superantigen (SAG), we infected MRL‐+/+ and MRL‐lpr (Fas mutant) mice with mouse mammary tumor virus (MMTV) (SW), a virus encoding an SAG with the same specificity as endogenous Mtv‐7‐SAG. In normal mice, this infection has two distinct consequences on specific Vβ6+CD4+ T cells, consisting of activation followed by clonal deletion. MMTV (SW)‐SAG‐induced activation in vivo was identical in MRL‐+/+ and MRL‐lpr mice. In contrast, clonal deletion showed age‐dependent impairment. Early infection (5 weeks) led to identical clonal deletion of specific T cells in blood lymphocytes from MRL‐+/+ and MRL‐lpr mice, although clonal deletion was slightly impaired in the MRL‐lpr lymph nodes. Late infection (10 weeks) of MRL‐lpr mice led to markedly delayed and reduced clonal deletion. Vβ6+CD4+ T cells which escaped clonal deletion in aging MRL‐lpr mice were not anergized by interaction with SAG. These results show that peripheral clonal deletion induced by viral SAG in adult mice is controlled by Fas, but not exclusively so.
Induction of CTL in vivo by major histocompatibility complex class I‐peptide complexes covalently associated on the cell surface
Authors: Anjuère Fabienne , Horvath Clotilde , Cerottini Jean‐Charles , Luescher Immanuel F.
Published: March 2007

The identification of endogenously produced antigenic peptides presented by MHC class I molecules has opened the way to peptide‐based strategies for CTL induction in vivo. Here we demonstrate that the induction in vivo of CTL directed against naturally processed antigens can be triggered by injection of syngeneic cells expressing covalent major histocompatibility complex class I‐peptide complexes. In the model system used, the induction of HLA‐Cw3 specific cytotoxic T lymphocytes (CTL) in mice by cell surface‐associated, covalent H‐2Kd (Kd)‐Cw3 peptide complexes was investigated. The Kd‐restricted Cw3 peptide 170–179 (RYLKNGKETL), which mimics the major natural epitope recognized by Cw3‐specific CTL in H‐2d mice, was converted to a photoreactive derivative by replacing Arg‐170 with N‐β‐(4‐azidosalicyloyl)‐L‐2,3‐diaminopropionic acid. This peptide derivative was equivalent to the parental Cw3 peptide in terms of binding to Kd molecules and recognition by Cw3‐specific CTL clones and could be cross‐linked efficiently and selectively to Kd molecules on the surface of Con A‐stimulated spleen cells from H‐2d mice. Photocross‐linking prevented the rapid dissociation of Kd‐peptide derivative complexes that takes place under physiological conditions. Cultures of spleen cells or peritoneal exudate cells from mice inoculated i.p. with peptide‐pulsed and photocross‐linked cells developed a strong CTL response following antigenic stimulation in vitro. The cultured cells efficiently lysed not only target cells sensitized with the Cw3 170–179 peptide but also target cells transfected with the Cw3 gene. Moreover, their TCR preferentially expressed Vβ10 and JαpHDS58 segments as well as conserved junctional sequences, as has been observed previously in Cw3‐specific CTL responses. In contrast, no Cw3‐specific CTL response could be obtained in cultures derived from mice injected with Con A‐stimulated spleen cells pulsed with the peptide derivative without photocross‐linking.
Germ‐line transcription of the T cell receptor δ gene in mouse hematopoietic cell lines
Authors: Shimamura Michio , Ohta Sachiko
Published: March 2007

T cell receptor (TCR) genes are initially activated in hematopoietic stem cells that have been committed to the T cell lineage during ontogeny. We have established cell lines derived from hematopoietic organs which express truncated TCR δ mRNA (1.3 kbp). In this study, cDNA clones of this mRNA were analyzed to characterize TCR δ gene activation in early T cell development. Transcription started from an intron between Dδ2 and Jδ1, 72 bp upstream from Jδ1, and continued to Jδ1 and Cδ exons, which were directly spliced to Jδ1. Thus, the truncated mRNA expressed by the cell lines was determined to be a germ‐line transcript. The promoter activity of the DNA fragment between Dδ2 and Jδ was assessed by its ability to drive the transcription of a reporter gene linked to it in the cell line expressing the TCR δ germ‐line transcript. TCR δ germ‐line transcription was found not only in these cell lines but also in fetal liver and thymus cells. These findings indicate that the TCR δ germ‐line transcription is a naturally occurring event in developing T cells. The significance of germ‐line transcription of TCR δ genes is unknown, but it may be an event that follows T‐lineage commitment or, at least, may be closely associated with this commitment.
Interleukin‐4 and ‐10 exacerbate candidiasis in mice
Authors: Tonnetti Laura , Spaccapelo Roberta , Cenci Elio , Mencacci Antonella , Puccetti Paolo , Coffman Robert L. , Bistoni Francesco , Romani Luigina
Published: March 2007

Neutralization of endogenous interleukin (IL)‐4 or IL‐10 in mice with Candida albicans infection initiates or accelerates development of a T helper (Th)1‐associated protective response. Here, we report the effect of IL‐4 and IL‐10 administration on the course of systemic or gastrointestinal (GI) candidiasis and on the development of Th immunity using yeast/host combinations that result either in Th1‐associated self‐limiting infection (healer mice) or in Th2‐associated progressive disease (nonhealer mice). Treatment with IL‐4 or IL‐10 greatly exacerbated the course of systemic infection in nonhealer mice and rendered healer mice, inoculated with attenuated yeast cells, susceptible to infection. Under the latter conditions of yeast challenge and IL‐4/IL‐10 administration, the development of a fatal disease was associated with inhibition of IL‐12 production and detection of progressive Th2 cell dominance. In contrast, in healer mice allowed to resolve their infections and to develop long‐lived anti‐candidal resistance, the expression of this acquired resistance was not impaired by IL‐4 and/or IL‐10, as shown by the outcome of reinfection with virulent yeast cells. In the GI model of infection, both IL‐4 and IL‐10 were found to exacerbate the course of infection and to induce the appearance of CD4+ T cells producing high levels of IL‐4 and IL‐10 in Peyer's patches. These findings demonstrate that exogenous IL‐4 and IL‐10 may greatly affect the development of Th responses to C. albicans in vivo, but do not modify the expression of established and predominant Th1 cell reactivity.
T lymphocyte repertoire in Theiler's virus encephalomyelitis: the nonspecific infiltration of the central nervous system of infected SJL/J mice is associated with a selective local T cell expansion
Authors: Musette Philippe , Bureau Jean‐François , Gachelin Gabriel , Kourilsky Philippe , Brahic Michel
Published: March 2007

Theiler's virus causes, in the susceptible SJL/J mouse, a chronic demyelinating disease that resembles multiple sclerosis. Demyelination is at least in part immune mediated and coincides with the infiltration of the central nervous system by T lymphocytes. We analyzed the repertoire of the T cell receptor (TcR) β‐chain for each Vβ‐Jβ combination, in spinal cord and spleen T cells of infected animals. All Vβ families were detected in spinal cord as well as in spleen, as if the central nervous system were undiscriminantly invaded by T cells in the infected animals. Some T cells, defined by specific Vβ‐Jβ combinations, were expanded in the spinal cord but not in the spleen, most probably because of an antigen‐driven response. The TcR β‐chain repertoire of infiltrating T cells was the same at the onset of demyelination as when the disease was full‐blown. These results provide the first description of the repertoire of the T cells which infiltrate the central nervous system during the course of this disease.
Expression of Epstein‐Barr virus nuclear antigen‐2 (EBNA2) induces CD21/CR2 on B and T cell lines and shedding of soluble CD21
Authors: Larcher Clara , Kempkes Bettina , Kremmer Elisabeth , Prodinger Wolfgang M. , Pawlita Michael , Bornkamm Georg W. , Dierich Manfred P.
Published: March 2007

Stable transfection of Epstein‐Barr virus (EBV) nuclear antigen 2 (EBNA2) expressed as a fusion protein with the hormone‐binding domain of the estrogen receptor was used to study expression of CD21 and other surface markers in different cell lines. Special emphasis was placed on cell lines with a normally low expression of CD21, especially on T cell lines. After induction of EBNA2, a substantial increase in CD21 mRNA was observed, as well as increased production of membrane CD21. This was found not only in cell lines of B cell origin, but also in the T cell line Jurkat. The amount of CD21 was quantitated by means of a fluorescence immunoassay, and found to correlate with the presence of EBNA2 protein. A decrease in EBNA2 abundance was associated with complete loss of cell‐associated CD21. As we could also detect large amounts of soluble CD21 (sCD21) in the supernatant of the transfected cell lines, which exceeded the total amount contained in the respective cell lysates, this indicates considerable shedding of the newly synthesized receptor molecules induced by EBNA2, comparable to the situation described for CD23. It further provides an explanation of the recent findings of increased sCD21 levels in sera of patients with EBV‐associated disease, and suggests a possible additional function of EBNA2 in vivo.
The role of cellular competition in B cell survival and selection of B cell repertoires
Authors: Freitas António A. , Rosado Maria Manuela , Viale Anne‐Claire , Grandien Alf
Published: March 2007

We studied the competitive repopulation by different B cells of irradiated mice reconstituted with bone marrow from either congenic or Ig‐transgenic (TG) mice mixed at different ratios. We found that after reconstitution, the number of B cells recovered in the different chimeras is similar and independent of the ratio of injected cells. In chimeras hosting TG and non‐TG cells, the relative representation of the donor cell lineages diverges from the ratios present in the inoculum, i.e. at the periphery, non‐TG cells are preferentially selected. Selection of non‐TG cells only occurs when population growth plateaus, i.e. when resources become limiting and competition starts to operate. Selection of non‐TG cells depends on surface Ig expression, and they are selected because they have a longer survival. Finally, the life‐expectancy of the same B cell population differs depending upon the second population present. The present results show that the life‐span and the population size of each B cell clone can be altered (interfered with) by the presence of a second cell population, demonstrating the existence of cellular competition among B cells. Our findings establish the role of cellular competition in the selection of B cell repertoires and the existence of a hierarchy of B cell selection in the absence of antigenic stimulation. The implications of cellular competition on our understanding of the immune system are discussed.
gp120‐induced programmed cell death in recently activated T cells without subsequent ligation of the T cell receptor
Authors: Foster Serene , Beverley Peter , Aspinall Richard
Published: March 2007

In most individuals, HIV infection is characterized by a progressive decline in the number of peripheral blood CD4+ T lymphocytes, and while the number of CD4+ cells is within the normal range, defects in immune function are detectable. To date neither the decline in function nor the decline in cell number have been satisfactorily explained. Here we describe a mechanism which may contribute to the immunodeficiency and decline in CD4+ cell numbers in HIV‐infected individuals. We show that recently activated T cells are susceptible to apoptosis when exposed to HIV gp120 in the presence of anti‐gp120 antibody.
The glycosylphosphatidylinositol‐anchored CD59 protein stimulates both T cell receptor ζ/ZAP‐70‐dependent and ‐independent signaling pathways in T cells
Authors: Deckert Marcel , Ticchioni Michel , Mari Bernard , Mary Didier , Bernard Alain
Published: March 2007

The glycosylphosphatidylinositol (GPI)‐anchored CD59 protein (human protectin) protects cells against complement‐induced lysis, binds to CD2 and also transduces activation signals within T cells. We have further examined the biochemical signals transduced by CD59 and addressed its role in regard to the CD3‐mediated signaling cascade. We show here that CD59 cross‐linking induces a time‐dependent activation of p56lck and of p70zap (ZAP‐70) in CD3‐positive Jurkat cells, leading to the stimulation of the T cell receptor ζ/ZAP‐70 signaling cascade and interleukin‐2 (IL‐2) synthesis. Cross‐linking of CD59 on peripheral T cells and thymocytes induces tyrosine phosphorylations identical to those seen in Jurkat cells and this is followed by lymphokine production and proliferation. In contrast, only activation of CD59‐associated p56lck occurs in CD3‐negative Jurkat cells, while IL‐2 production is impaired, consistent with the lack of ZAP‐70 tyrosine phosphorylation observed in these cells. CD59 triggers activation events even in the absence of CD3/T cell receptor expression in Jurkat cells. CD59 cross‐linking synergizes with sub‐optimal doses of phorbol ester for activation of the protein kinase C and of the p42mapk, as shown by in vitro phosphorylation of histone HIIIS and myelin basic protein, respectively, and leads to CD25 but not CD69 expression. In conclusion, at least two signaling pathways are triggered through CD59, the first one involving ZAP‐70 activation and leading to IL‐2 secretion and a second pathway observed in the absence of ZAP‐70 activation leading to CD25 expression. These two pathways are likely to be involved in the modulation of T cell activation by CD59 protein.
Non random activation of endogenous interleukin‐2, (IL‐2), IL‐2 receptor α and IL‐2 receptor β genes after transfection of mouse fibroblasts with a cDNA for the α chain of the human IL‐2 receptor
Authors: Han Daishu , Plaisance Stephane , Rubinstein Eric , Alileche Abdelkrim , Pottin‐Clemenceau Corinne , Kim T. S. , Cohen Edward P. , Jasmin Claude , Azzarone Bruno
Published: March 2007

Mouse fibroblasts do not ordinarily express components for the interleukin‐2 receptor (IL‐2Rα, β, and γ). An analysis of these cells by reverse transcription followed by polymerase chain reaction, however, indicates the presence of transcripts specific for the IL‐2R β and γ genes. Transfection of the cDNA for the α chain of the human IL‐2R into LTK− mouse fibroblast cell line (L3 cells) leads, in long‐term cultures, to the formation of transcripts of endogenous mouse IL‐2, IL‐2Rα and β genes, as detected by Northern blotting. Based upon the results of the binding of 125I‐labeled IL‐2 to the transfected cells, three IL‐2‐binding proteins of 55 kDa, 65 kDa and 75 kDa were expressed by the transfected cells. The 65‐kDa and 75 kDa proteins bound IL‐2 in the presence of monoclonal antibodies for the IL‐2Rα chain. These polypeptides assembled to form high‐affinity IL‐2R, as shown by Scatchard binding analyses. The receptors were functionally active, since the expression of H‐2k major histocompatibility complex antigens on the surface membranes of L3 cells was enhanced by exposing the cells to IL‐2. Activation of the IL‐2 gene was also observed in long‐term cultures of Lαβ cells, another LTK− transfectant expressing the human IL‐2Rα chain. This type of gene activation was not observed in LTK− fibroblasts transfected with cDNA for human IL‐2 or IL‐2Rβ genes. In L3 and Lαβ cells, transcription of the endogenous IL‐2 gene was suppressed by cyclosporin A and enhanced by cycloheximide. These data may have implications for gene therapy of cancer cells.
Are CD8 T cells involved in control of African trypanosomiasis in a natural host environment?
Authors: Sileghem Maarten , Naessens Jan
Published: March 2007

Murine models have suggested that CD8 T cells might play a major parasite‐promoting role in African trypanosomiasis. To assess the role of these cells in a natural host environment, we have depleted CD8 cells from Boran cattle in vivo and subsequently infected these animals with Trypanosoma congolense by tsetse fly challenge. Following administration of a mouse monoclonal anti‐bovine CD8 antibody, we have been able to achieve a depletion of more than 99.9% in peripheral blood, spleen, prescapular lymph nodes, prefemoral lymph nodes, mesenteric lymph nodes and Peyer's patches. Depletion could be maintained over a 4–5‐week infection period. Despite the almost total absence of CD8 cells, no effect whatsoever was observed on parasitemia. In addition, anemia, which is the main factor determining the mean survival time in cattle was not affected by the CD8 depletion.
Proliferative response of human CD4+ T lymphocytes stimulated by the lectin jacalin
Authors: Blasco Elisabeth , Barra Anne , Nicolas Michel , Lecron Jean‐Claude , Wijdenes John , Preud'homme Jean‐Louis
Published: March 2007

The Galβ(1–3)GalNAc‐binding lectin jacalin is known to specifically induce the proliferation of human CD4+ T lymphocytes in the presence of autologous monocytes and to interact with the CD4 molecule and block HIV‐1 infection of CD4+ cells. We further show that jacalin‐induced proliferation is characterized by an unusual pattern of T cell activation and cytokine production by human peripheral blood mononuclear cells (PBMC). A cognate interaction between T cells and monocytes was critical for jacalin‐induced proliferation, and human recombinant interleukin (IL)‐1 and IL‐6 did not replace the co‐stimulatory activity of monocytes. Blocking studies using monoclonal antibodies (mAb) point out the possible importance of two molecular pathways of interaction, the CD2/LFA‐3 and LFA‐1/ICAM‐1 pathways. One out of two anti‐CD4 mAb abolished jacalin responsiveness. Jacalin induced interferon‐γ and high IL‐6 secretion, mostly by monocytes, and no detectable IL‐2 synthesis or secretion by PBMC. In contrast, jacalin‐stimulated Jurkat T cells secreted IL‐2. CD3− Jurkat cell variants failed to secrete IL‐2, suggesting the involvement of the T cell receptor/CD3 complex pathway in jacalin signaling. IL‐2 secretion by CD4− Jurkat variant cells was delayed and lowered. In addition to CD4, jacalin interacts with the CD5 molecule. Jacalin‐CD4 interaction and the proliferation of PBMC, as well as IL‐2 secretion by Jurkat cells were inhibited by specific jacalin‐competitive sugars.
α6 integrins participate in pro‐T cell homing to the thymus
Authors: Ruiz Patricia , Wiles Michael V. , Imhof Beat A.
Published: March 2007

During embryogenesis, colonization of the thymic rudiment by hemopoietic progenitor cells depends on the adhesion of these cells to the jugular endothelium. Previously, we showed that progenitor T cells (pro‐T cells) interact with α6 integrins present on vascular endothelium. Here, we demonstrate that anti‐α6 integrin antibodies reduced the number of thymocytes up to 80 % in a congenic mouse model for thymus colonization by pro‐T cells. In organotypic thymus cultures, the anti‐α6 integrin antibodies did not influence T cell development and proliferation. From this, we conclude that α6 integrin participates in thymus homing. During mouse thymus ontogeny, α6 integrin mRNA and protein expression was found as early as day 10 of development; at day 11, perithymic endothelial cells were α6 integrin positive. Two α6 integrin mRNA exist which are produced by alternative exon usage. The longer form, α6, integrin, predominates during early embryonic stages, while the shorter α6A form was present later during development. Although α6, integrins can be displayed by immature thymocytes, strongest expression was found on intra‐ and perithymic vascular endothelium. These data suggest that α6 integrins are involved in the homing of pro‐T cells to the developing thymus by mediating adhesion of pro‐T cells to the vascular endothelium.