bluuebunny/crossref_metadata_2025_split huggingface dataset:
Cassette baculovirus vectors for the production of chimeric, humanized, or human antibodies in insect cells
Authors: Poul Marie‐Alix , Cerutti Martine , Chaabihi Hassan , Ticchioni Michel , Deramoudt François‐Xavier , Bernard Alain , Devauchelle Gérard , Kaczorek Michel , Lefranc Marie‐Paule 
Published: March 2007

AbstractPlasmid cassette‐transfer vectors pBHuCϰ and pBHuCy1 have been designed which enable the construction of recombinant baculoviruses directing the co‐expression of complete immunoglobulin in insect cells. We describe the application of these vectors for the expression of a human/mouse chimeric monoclonal antibody of potential immunosuppressive clinical value derived from a mouse anti‐human CD29 monoclonal antibody (Mu‐K20). The chimeric K20 light and heavy chains produced in sf9 insect cells were correctly processed and assembled into a normal immunoglobulin which is secreted into the culture medium of infected cells. The chimeric mAb Ch‐K20‐sf9 reproduces in vitro the functional properties of the parental mouse K20, including affinity and inhibition of lymphocyte proliferation. These results demonstrate that the baculovirus/insect cell expression system is suitable for the expression of fully active monoclonal antibodies of therapeutic value. Our generic cassette approach makes this system a very flexible and convenient one for the rapid production of either chimeric, humanized or human mAb with heavy and light chains of any isotype.
Cell cycle regulation of immunoglobulin class switch recombination and germ‐line transcription: potential role of Ets family members
Authors: Lundgren Mats , Ström Lena , Bergquist Lars‐Olof , Skog Sven , Heiden Thomas , Stavnezer Janet , Severinson Eva 
Published: March 2007

AbstractPrevious studies have indicated that transcription of germ‐line (GL) CH genes is necessary to obtain immunoglobulin (Ig) class switching. We report here a correlation between proliferation, switching and GL transcripts. Sμ‐Sγ1 switch recombination in lipopolysaccharide (LPS) + interleukin‐4 (IL‐4)‐activated mouse B cells was assayed by a digestion‐circularization polymerase chain reaction. Switching to γ1 is reduced upon inhibition of DNA synthesis with hydroxyurea (HU) or aphidicholin (AC). Incubation of activated B cells with HU severely reduces steady‐state levels of GL γ1 and ϵ RNA. By utilizing elutriation to synchronize B cell blasts in different phases of the cell cycle, it was found that GL γ1 transcripts are mainly expressed in G1 and S phases, but not in G0. Using the electrophoretic mobility shift assay, we characterized two major LPS‐induced complexes, which bind to the GL γ1 promoter and are expressed at levels which correlate with the amount of LPS‐induced DNA synthesis. Furthermore, the intensity of the complexes is reduced when cells are arrested with the DNA synthesis inhibitors HU or AC. Elutriation experiments revealed that the complexes are expressed in G1 and S, but not in G0. They bind to an Ets consensus element near the major initiation sites used in proliferating cells. The possible implications of these findings for Ig isotype switching are discussed.
B7.2 provides co‐stimulatory functions in vivo in response to staphylococcal enterotoxin B
Authors: Muraille Eric , de Smedt Thibaut , Urbain Jacques , Moser Muriel , Leo Oberdan 
Published: March 2007

AbstractExcessive T cell activation induced by bacterial superantigens plays an important role in the pathology associated with Gram‐positive bacteremia. To gain insight into the early phases of T cell activation by bacterial enterotoxins in vivo, we investigated the ability of antibodies to well‐defined co‐stimulatory molecules to inhibit T cell activation and the subsequent toxic shock syndrome induced in BALB/c mice following the injection of staphylococcal enterotoxin B (SEB). We demonstrate here that a single dose of anti‐B7.2 antibodies, but not anti‐B7.1 antibodies, significantly inhibits T cell activation, as judged by lower systemic IL‐2 release, blastogenesis and IL‐2 receptor expression, and reduces the lethal effect of SEB in D‐galactosamine‐sensitized mice. These results demonstrate that co‐stimulation through the B7.2 molecule plays an important role in the activation of T cells in response to SEB in vivo and suggest alternative therapies for septic shock caused by bacterial enterotoxins based on blocking antibodies to co‐stimulatory molecules.
Regulation of murine B cell growth and differentiation by CD30 ligand
Authors: Shanebeck Kurt D. , Maliszewski Charles R. , Kennedy Mary K. , Picha Kathleen S. , Smith Craig A. , Goodwin Ray G. , Grabstein Kenneth H. 
Published: March 2007

AbstractA ligand for CD30 has been recently cloned, and has been shown to have sequence homology with the tumor necrosis factor family of cytokines. CD30 ligand (CD30L) was found to be induced on helper T cell clones, and its receptor was expressed on freshly isolated and activated murine B cells. Recombinant murine CD30L was found to share many functional properties with CD40 ligand (CD40L) in the regulation of murine B cell growth and differentiation in vitro. CD30L stimulated B cell proliferation, antigen‐specific antibody production, and polyclonal immunoglobulin secretion in a cytokine‐dependent manner. In particular, the stimulation of B cell proliferation by CD30L required interleukin (IL)‐4 and IL‐5, induction of anti‐sheep red blood cell antibody‐secreting B cells by CD30L required IL‐2 and IL‐5, and optimal induction of polyclonal immunoglobulin secretion required IL‐4 and IL‐5. Under these conditions, the polyclonal secretion of IgGl, IgA, IgG3 and IgE was induced. The induction of immunoglobulin secretion by CD30L was independent of CD40L, as B cells from CD40L deficient‐mice responded normally to CD30L treatment. We conclude that CD30L is a potent mediator of B cell growth and differentiation in vitro and may play a role in cognate T cell‐B cell interactions.
Antigen receptor‐initiated growth inhibition is blocked in CD45‐loss variants of a mature B lymphoma, with limited effects on apoptosis
Authors: Ogimoto Mami , Katagiri Tatsuo , Mashima Keisuke , Hasegawa Kiminori , Mizuno Kazuya , Yakura Hidetaka 
Published: March 2007

AbstractRole of CD45 in B cell antigen receptor (BcR)‐mediated signaling events in mature B cells was examined using BAL‐17 and its CD45‐negative clones. In the CD45‐negative clones, BcR stimulation induced tyrosine phosphorylation almost identical to the parental cells, with a few exceptions of reduced phosphorylation, especially of a protein of about 60 kDa. BcR‐induced calcium responses were reduced in the CD45‐negative clones, but the kinetics were similar to the parent. BcR stimulation led to growth inhibition in the parental cells, but signals for growth inhibition were completely blocked in the CD45‐negative clones. Interestingly, the same stimulation induced low, but significant levels of apoptosis both in the parent and in the CD45‐negative clones. Thus, in mature BAL‐17 cells, CD45 subtly mediates early signaling events (tyrosine phosphorylation and Ca2+ mobilization), and is absolutely required for the signaling pathway leading to growth regulation, but has limited effects on apoptosis.
Antigen receptor‐triggered apoptosis in immature B cell lines is associated with the binding of a 44‐kDa phosphoprotein to the PTP1C tyrosine phosphatase
Authors: Wu Yongjian , Pani Giovanni , Siminovitch Katherine A. , Hozumi Nobumichi 
Published: March 2007

AbstractWhile cross‐linking of the membrane IgM (mIgM) molecules expressed on WEHI 231 lymphoma cells induces these cells to undergo apoptosis, we have previously observed that ligation of the mIgD expressed on IgD‐transfected WEHI 231 (Wδ) cells is not associated with induction of cell death. Thus mIgM+IgD+ Wδ cells provide a valuable reagent for delineating the molecular events which modulate the physiologic outcome of B cell antigen receptor (BCR) engagement. In view of recent data implicating the cytosolic phosphotyrosine phosphatase PTP1C in the regulation of BCR signaling capacity, we used Wδ cells to investigate the potential role for PTP1C in modulating the cell response to BCR activation. The results of this analysis revealed PTP1C to undergo rapid tyrosine phosphorylation following mIgM or mIgD cross‐linking and to associate with a number of other phosphoproteins in stimulated Wδ cells. Among these latter phosphoproteins, one prominent species of about 44 kDa (pp44) which co‐precipitated with PTP1C in mIgM‐ligated cells was not detected in PTP1C immunoprecipitates from mIgD‐ligated cells. The association of PTP1C with this 44‐kDa phosphoprotein following mIgM cross‐linking was also observed in two additional B cell lines representing an immature state of differentiation, but was not detected after BCR engagement in two representative mature B cell lines or in splenic B cells. Initial data concerning the identity of pp44 indicate that this molecule does not represent the Shc, MAPK or Ig‐β proteins and may, therefore, constitute a previously unidentified signaling effector. While the structural and biochemical properties of pp44 require further definition, the findings suggest that BCR‐triggered interactions of PTP1C with pp44 occur only in the context of an immature state of cellular differentiation and the induction of apoptosis. These data therefore suggest that PTP1C interactions with pp44 may be relevant to the transduction of BCR signals which evoke cell death.
Characteristics of antigen‐independent and antigen‐dependent interaction of dendritic cells with CD4+ T cells
Authors: Hauss Pascale , Selz Françoise , Cavazzana‐Calvo Marina , Fischer Alain 
Published: March 2007

AbstractDendritic cells (DC) are the main antigen‐presenting cells for the initiation of primary T cell‐mediated immune responses. In the first stage of activation, T cells bind to DC in an antigen‐independent manner. We studied the adhesion characteristics of human CD4+ T cells to DC generated from CD34+ hematopoietic progenitors following 12 to 13 days of culture in the presence of granulo‐cyte/macrophage colony‐stimulating factor and tumor necrosis factor‐α. A majority of these cells had the morphology, phenotype and functions of DC. CD4+ T/DC adhesion was measured by means of fluorescence microscopy and flow cytometry. Four independent receptor/ligand pathways, LFA‐1/ICAM, ICAM/LFA‐1, CD2/LFA‐3 and CD28/CD80, were involved in the transient adhesion of DC to CD4+ T cells in antigen‐independent and specific alloantigen‐dependent situations, as shown by blocking experiments using monoclonal antibodies. The antibodies also blocked a primary mixed lymphocyte reaction (MLR) in which DC were used as stimulatory cells. Adhesion of alloreactive CD4+ T cells to antigen‐presenting DC was stronger than that of resting CD4+ T cells, while peak adhesion occurred after 5 and 20 min, respectively. The LFA‐1 ligands involved in adhesion of resting CD4 T cells to DC and alloreactive CD4+ T cells to specific DC differed in part, since ICAM‐3 on resting T cells and ICAM‐1 on alloreactive T lymphocytes preferentially bound LFA‐1. Studies of interactions between DC and phorbol ester‐activated T cells expressing the CD40 ligand revealed a fifth independent adhesion pathway, CD40/CD40 ligand. CD4‐mediated regulation of CD4+ T/DC adhesion was suggested by the observation that preincubation of CD4+ T cells and DC individually with anti‐CD4 antibodies inhibited adhesion. In addition, antibodies specific for HLA class II molecules inhibited adhesion when used to pretreat DC but not alloactivated CD4+ T cells.
Control of thymus‐independent intestinal intraepithelial lymphocytes by β2‐microglobulin
Authors: Neuhaus Oliver , Emoto Masashi , Blum Carmen , Yamamoto Shigeki , Kaufmann Stefan H. E. 
Published: March 2007

AbstractMurine intestinal intraepithelial lymphocytes (i‐IEL) comprise thymusdependent cells such as T cell receptor (TcR) α/β CD8α/β+ i‐IEL, as well as thymus‐independent ones such as TcRα/β CD8α/α+ and TcRγ/δ CD8α/α+ i‐IEL. Whilst the development of the CD8α/β expressing i‐IEL is strictly contingent on major histocompatibility complex (MHC) class I surface expression, that of CD8α/α i‐IEL appears largely MHC class I independent. We have used β2‐microglobulin (β2m)−/− mutant mice lacking surface‐expressed MHC class I and TcRα/β CD8α/β+ i‐IEL to analyze the potential impact of MHC class I on regional activation of thymus‐independent i‐IEL. To analyze the role of TcRγ/δ i‐IEL in regional cell interactions, these mice were treated with the anti‐TcRγ/δ mAb, GL3. Whilst numbers of TcRα/β CD8α/α i‐IEL were markedly reduced in βm+/− mice, those of TcRγ/δ i‐IEL were elevated. Administration of GL3 in vivo caused TcR down‐modulation and functional inactivation of TcRγ/δ i‐IEL in β2m+/− mice. In contrast, TcR expression and functional activities of TcRγ/δ i‐IEL from β2m−/− mice were not impaired by GL3 treatment. The TcRα/β CD8β i‐IEL from β2m−/− mice were expanded and functionally activated as a consequence of TcRγ/δ engagement. The TcRγ/δ i‐IEL and TcRα/β CD8α/α+ i‐IEL from athymic nu/nu mice which express MHC class I, but lack TcRα/β CD8α/β+ i‐IEL, responded to TcRγ/δ engagement as those from the β2m+/− controls. In addition, the TcRγ/δ i‐IEL from TcRβ−/− and TcRβ+/− mutants were equally affected by GL3. We conclude that the absence of β2m renders TcRγ/δ i‐IEL resistant to TcR‐mediated inactivation and promotes activation of TcRα/β CD8β− i‐IEL. The activation of TcRγ/δ i‐IEL seems to be directly controlled by β2m/MHC class I expression and independent from TcRα/β CD8β+ i‐IEL. Regulation of self‐reactive thymus‐independent i‐IEL through β2m/MHC class I may contribute to control of autoreactive immune responses in the intestine.
CD34‐positive early human thymocytes: T cell receptor and cytokine receptor gene expression
Authors: Ktorza Sandra , Sarun Sukhéna , Rieux‐Laucat Frédéric , de Villartay Jean‐Pierre , Debré Patrice , Schmitt Christian 
Published: March 2007

AbstractCD34, a stem cell marker, has been shown to be expressed on human CD3−CD4−CD8− (triple‐negative; TN) thymocytes. Phenotypic and functional analyses suggest the following differentiation sequence: CD34+ 1−3−4−8− → CD34+ 1+3−4→8− → CD34−1++3−4+8 → CD34−1++3−4+8+. In this report, we examined cytokine receptor gene expression on these subsets by reverse transcription‐polymerase chain reaction analysis (RT‐PCR). We were able to detect interleukin‐7 receptor (IL‐7R), c‐kit and IL‐2Rγ in all CD34+ thymocyte subsets, consistent with previous functional studies. We found IL‐1R, granulocyte/macrophage colony‐stimulating factor receptor‐α and IL‐4R transcripts in CD3− and CD34+ subsets. Secondly, we investigated T cell receptor (TCR)‐δ and ‐β gene rearrangement and transcription in CD34+ thymocytes. Our results show that a full‐length TCR‐δ transcript and the recombination activating genes RAG‐1 and RAG‐2 mRNA were already expressed in the CD34+1− subset. Mature Vβ‐containing TCR transcripts were also detected in the CD34+ 1+ subset, but not in the CD1− fraction. Furthermore, PCR analysis of D‐Jβ gene rearrangements showed that ≥ 70% of CD34+ 1− cells are in a TCRβ germ‐line configuration, although D‐Jβ recombination had already started in this population.
T cell receptor diversity in alloreactive responses against HLA‐B27 (B*2705) is limited by multiple‐level restrictions in both α and β chains
Authors: Barber Domingo F. , López Daniel , de Castro José A. López 
Published: March 2007

AbstractThe T cell receptors (TCR) in HLA‐B27 (B*2705) alloreactivity were analyzed in cytotoxic T lymphocytes (CTL) from two individuals. Non‐random usage was found in Vβ, N+Dβ, Vα, and Jα, but not in Jβ segments or Nα‐regions. Vβ segments from homology subgroup 4 were predominant and not associated to a particular donor or fine specificity, suggesting involvement in recognizing the HLA‐B27 molecule. In contrast, preferential Vα usage was associated with particular individuals and fine specificities, indicating distinct Vβ and Vα recruitment and contribution to allorecognition. Recurrent N+Dβ motifs and Jα segments, even from different donors, limited junctional diversity, suggesting that CDR3 usage was determined by the alloantigenic epitope independently of individuals. TCR were selected differently at various levels, as indicated by the following findings. Four clonotypes with similar fine specificity had identical β and unrelated α chains. Similar α were associated with unrelated β chains, and vice versa. CTL using Vβ subgroup 4 did not globally show concomitant predominance of other TCR elements. Vα7, one of the preferred Vα segments, was always associated with Vβ subgroups other than 4. Sometimes, a TCR showed homology in elements of one chain to a second TCR or group of TCR, and to another in the other chain. These results are best explained by differential selection of TCR elements by different epitopes, providing a key to the inner structure of allospecific TCR repertoires.
Superantigens initiate cognate CD4+ T cell/ B cell interactions leading to early activation and proliferation of B cells
Authors: Al‐Daccak Reem , Damaj Bassam , Naccache Paul H. , Mourad Walid 
Published: March 2007

AbstractDimerization or even multimerization of various receptors is commonly required for signal transduction. We report here that clustering of major histocompatibility complex class II molecules in human B cells by biotinylated staphylococcal enterotoxin A (SEA) cross‐linked with avidin induces an increase in the level of intracellular calcium. This response was abolished by prior treatment with protein tyrosine kinase (PTK) inhibitors, suggesting that SEA‐triggered calcium mobilization in B cells is probably dependent on the activation of PTK. The implication of PTK in SEA‐induced early B cell activation was then confirmed by demonstrating that cross‐linked SEA induces a significant increase in the level of tyrosine phosphorylation in B cells. The requirement of biotinavidin cross‐linking in SEA‐induced calcium mobilization in B cells can be fulfilled by the addition CD4+ T cells, suggesting a role for CD4 molecules. Using the murine CD4− T cell hybridoma 3DT, or its derivative 11B3 transfected with human CD4 that both express SEA‐specific TCR, we confirmed the CD4 requirement for B cell calcium mobilization and that both specific TCR and CD4 molecules are required in early events of B cell activation induced by SEA. The role of CD4 in SEA‐induced B cell proliferation was then investigated. SEA‐stimulated B cells proliferated in the presence of CD4+ T cells, whereas no response was observed in the presence of CD8+ T cells. The addition of clone 11B3 CD4+ T cells failed to fulfill the requirement of CD4+ T cells in SEA‐induced B cell proliferation, indicating the possible involvement of other CD4+ T cell surface molecules in this response. This issue is currently under investigation.
Sialyl LewisX carbohydrate is expressed differentially during avian lymphoid cell development
Authors: Palojoki Eeva , Jalkanen Sirpa , Toivanen Paavo 
Published: March 2007

AbstractSialyl LewisX is a carbohydrate moiety involved in the regulation of white blood cell adhesion to endothelial cells. In this work, we have studied the expression, localization and function of sialyl LewisX carbohydrate on maturing B and T cells and the stroma of avian bursa, thymus and spleen as well as the role of sialyl LewisX in the generation of immune response and formation of germinal centers in the spleen. The expression of sialyl LewisX on bursal B cells decreases during the embryonic period, while on T cells it remains at a stable low level throughout their thymic development. Immediately after hatching, by which time the expression of sialyl LewisX on bursal B cells has already decreased, the stromal microenvironment of the bursa starts to express it. After hatching, sialyl LewisX is localized exclusively in the medullae of the follicles and the number of sialyl LewisX‐positive follicles declines gradually during the following weeks. Thymic stroma does not express sialyl LewisX at any stage of development. In spleen, sialyl LewisX is expressed on a number of B and T cells as well as on other cell types in a way that seems not to be restricted to any structurally defined area. The cells or stroma of the germinal centers, however, do not express sialyl LewisX. When given simultaneously with an immunizing agent, anti‐sialyl LewisX monoclonal antibody reduced the formation of germinal centers and suppressed antibody response in young birds. These results demonstrate that sialyl LewisX‐bearing cell surface molecules are required during the generation of immune responses. They also suggest a role for sialyl LewisX in B cell maturation.
Fetal VDJ gene repertoire in rabbit: evidence for preferential rearrangement of VH1
Authors: Tunyaplin Chainarong , Knight Katherine L. 
Published: March 2007

AbstractB cells in rabbit preferentially utilize the 3'‐most VH gene, VH1, in VDJ genes. To determine whether the preferential utilization of VH1 results from preferential rearrangement, we examined VH gene usage in nonproductive VDJ gene rearrangements, genes that would not be influenced by antigen selection. Since nonproductive VDJ gene rearrangements are found infrequently in neonatal and adult rabbits, we examined the VDJ gene rearrangements during early fetal development. According to nested‐primer polymerase chain reaction analyses, VDJ genes in fetal liver appeared around day 14 of gestation and nucleotide sequence analyses of VDJ genes cloned from 14‐ to 28‐day‐old fetuses showed that many of the genes were nonproductive. We found that most of these non‐productive genes had utilized VH1 indicating to us that the preferential utilization of VH1 is due to preferential rearrangement. We compared the nucleotide sequence of the promoter region of VH1 with sequences of several other unutilized and infrequently utilized VH genes and found that the core transcriptional factor motifs were similar in all VH promoters examined. We suggest that transcriptional activity of the VH1 promoter region does not explain the preferential rearrangement of VH1.