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Sample records for b-cell activation antigen

  1. Antigen-specific monoclonal antibodies isolated from B cells expressing constitutively active STAT5.

    Directory of Open Access Journals (Sweden)

    Ferenc A Scheeren

    Full Text Available BACKGROUND: Fully human monoclonal antibodies directed against specific pathogens have a high therapeutic potential, but are difficult to generate. METHODOLOGY/PRINCIPAL FINDINGS: Memory B cells were immortalized by expressing an inducible active mutant of the transcription factor Signal Transducer and Activator of Transcription 5 (STAT5. Active STAT5 inhibits the differentiation of B cells while increasing their replicative life span. We obtained cloned B cell lines, which produced antibodies in the presence of interleukin 21 after turning off STAT5. We used this method to obtain monoclonal antibodies against the model antigen tetanus toxin. CONCLUSIONS/SIGNIFICANCE: Here we describe a novel and relatively simple method of immortalizing antigen-specific human B cells for isolation of human monoclonal antibodies. These results show that STAT5 overexpression can be employed to isolate antigen specific antibodies from human memory B cells.

  2. Biochemical basis of synergy between antigen and T-helper (Th) cell-mediated activation of resting human B cells.

    OpenAIRE

    Chartash, E K; Crow, M K; Friedman, S M

    1989-01-01

    We have utilized CD23 expression as a marker for B cell activation in order to investigate the biochemical basis for synergy between antigen and T helper (Th) cells in the activation of resting human B cells. Our results confirm that while ligation of surface immunoglobulin (sIg) receptors by antigen analogues (e.g., F(ab')2 goat anti-human IgM) does not lead to CD23 expression, this stimulus markedly enhances CD23 expression induced during antigen specific Th-B cell interaction or by rIL-4. ...

  3. Epstein-Barr virus nuclear antigen 2 specifically induces expression of the B-cell activation antigen CD23

    International Nuclear Information System (INIS)

    Epstein-Barr virus (EBV) infection of EBV-negative Burkitt lymphoma (BL) cells includes some changes similar to those seen in normal B lymphocytes that have been growth transformed by EBV. The role of individual EBV genes in this process was evaluated by introducing each of the viral genes that are normally expressed in EBV growth-transformed and latently infected lymphoblasts into an EBV-negative BL cell line, using recombinant retrovirus-mediated transfer. Clones of cells were derived that stably express the EBV nuclear antigen 1 (EBNA-1), EBNA-2, EBNA-3, EBNA-leader protein, or EBV latent membrane protein (LMP). These were compared with control clones infected with the retrovirus vector. All 10 clones converted to EBNA-2 expression differed from control clones or clones expressing other EBV proteins by growth in tight clumps and by markedly increased expression of one particular surface marker of B-cell activation, CD23. Other activation antigens were unaffected by EBNA-2 expression, as were markers already expressed on the parent BL cell line. The results indicate that EBNA-2 is a specific direct or indirect trans-activator of CD23. This establishes a link between an EBV gene and cell gene expression. Since CD23 has been implicated in the transduction of B-cell growth signals, its specific induction by EBNA-2 could be important in EBV induction of B-lymphocyte transformation

  4. Germinal center B cells recognize antigen through a specialized immune synapse architecture.

    Science.gov (United States)

    Nowosad, Carla R; Spillane, Katelyn M; Tolar, Pavel

    2016-07-01

    B cell activation is regulated by B cell antigen receptor (BCR) signaling and antigen internalization in immune synapses. Using large-scale imaging across B cell subsets, we found that, in contrast with naive and memory B cells, which gathered antigen toward the synapse center before internalization, germinal center (GC) B cells extracted antigen by a distinct pathway using small peripheral clusters. Both naive and GC B cell synapses required proximal BCR signaling, but GC cells signaled less through the protein kinase C-β-NF-κB pathway and produced stronger tugging forces on the BCR, thereby more stringently regulating antigen binding. Consequently, GC B cells extracted antigen with better affinity discrimination than naive B cells, suggesting that specialized biomechanical patterns in B cell synapses regulate T cell-dependent selection of high-affinity B cells in GCs. PMID:27183103

  5. Trafficking of B cell antigen in lymph nodes

    DEFF Research Database (Denmark)

    Gonzalez, Santiago F.; Degn, Søren Egedal; Pitcher, Lisa A.; Woodruff, Matthew; Heesters, Balthasar A.; Carroll, Michael C.

    2011-01-01

    The clonal selection theory first proposed by Macfarlane Burnet is a cornerstone of immunology ( 1 ). At the time, it revolutionized the thinking of immunologists because it provided a simple explanation for lymphocyte specificity, immunological memory, and elimination of self-reactive clones ( 2...... microscopy ( 4, 5 ) have provided new insights into the trafficking of B cells and their antigen. In this review, we summarize these advances in the context of our current view of B cell circulation and activation....

  6. B cells pulsed with Helicobacter pylori antigen efficiently activate memory CD8+ T cells from H. pylori-infected individuals.

    Science.gov (United States)

    Azem, Josef; Svennerholm, Ann-Mari; Lundin, B Samuel

    2006-01-01

    Helicobacter pylori infection causes chronic gastritis that may progress to peptic ulcers or gastric adenocarcinoma and thereby cause major world-wide health problems. Previous studies have shown that CD4+ T cells are important in the immune response to H. pylori in humans, but the role of CD8+ T cells is less clear. In order to study the CD8+ T cell response to H. pylori in greater detail, we have evaluated efficient conditions for activation of CD8+ T cells in vitro. We show that H. pylori-reactive CD8+ T cells can be activated most efficiently by B cells or dendritic cells pulsed with H. pylori antigens. We further show that the majority of CD8+ T cells in H. pylori-infected gastric mucosa are memory cells, and that memory CD8+ T cells sorted from peripheral blood of H. pylori-infected individuals respond 15-fold more to H. pylori urease compared to memory cells from uninfected subjects. We conclude that CD8+ T cells do participate in the immune response to H. pylori, and this may have implications for the development of more severe disease outcomes in H. pylori-infected subjects. PMID:16324887

  7. Presentation of antigen by B cells subsets. Pt. 1. Lyb-5+ and Lyb-5- B cells differ in ability to stimulate specific T cells

    International Nuclear Information System (INIS)

    We have examined the antigen presenting cell (APC) function of different B cells. Resident, peritoneal B cells from normal mice were more efficient than splenic B cells in presenting antigen to CD4+ T cell lines. Peritoneal B cells from X-linked immunodeficient (Xid) mice, by contrast, stimulated no detectable responses. Xid splenic B cells were much less efficient APC than normal splenic B cells. B cells from neonatal mice also were very poor APC until the mice were 3 to 4 weeks old. Xid B cells presented antigen to T cell hybridomas as well as normal B cells showing that they process antigen normally. Thus, the defect is most likely in providing secondary signals. The ability of B cells to present antigen efficiency correlates with the percentage of B cells reported to express the Lyb-5 antigen. Anti-Lyb-5 serum and complement abrogated the APC activity of B cells suggesting that Lyb-5+, but not Lyb-5- cells are efficient APC. We also found that activated and resting normal splenic B cells, separated by buoyant density, presented antigen equally. Both populations also contained Lyb-5+ B cells although they were a larger fraction of the activated cells. Lyb-5 is now thought to be an activation antigen rather than a differentiation antigen. If this idea is correct, then our data indicate that anti-Lyb-5 more cleanly separates activated and resting B cells than buoyant density techniques. (author). 38 refs, 7 figs, 1 tab

  8. B cell antigen receptor-induced activation of an IRAK4-dependent signaling pathway revealed by a MALT1-IRAK4 double knockout mouse model

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    Dufner Almut

    2011-03-01

    Full Text Available Abstract Background The B cell antigen receptor (BCR and pathogen recognition receptors, such as Toll-like receptor 4 (TLR4, act in concert to control adaptive B cell responses. However, little is known about the signaling pathways that integrate BCR activation with intrinsic TLR4 stimulation. Antigen receptors initialize activation of the inducible transcription factor nuclear factor-κB (NF-κB via recruitment of the membrane-associated guanylate kinase caspase recruitment domain protein 11 (CARD11, the adapter molecule B cell CLL/lymphoma 10 (BCL10, and the "paracaspase" mucosa-associated lymphoid tissue lymphoma translocation gene 1 (MALT1 into lipid rafts. Upon BCR triggering, this activation strictly depends on BCL10, but not on MALT1, leading to the hypothesis that a MALT1-independent NF-κB activation pathway contributes to BCR-induced NF-κB activation downstream of BCL10. The identity of this pathway has remained elusive. Results Using genetic and biochemical approaches, we demonstrate that the IRAK4- and IRAK1-dependent TLR signaling branch is activated upon BCR triggering to induce partial NF-κB activation. BCR-induced MALT1-independent IκB degradation and B cell proliferation were inhibited in MALT1/IRAK4 double knockout B cells. Moreover, IRAK1 was recruited into lipid rafts upon BCR stimulation and activated following transient recruitment of IRAK4. Conclusion We propose that the observed crosstalk between BCR and TLR signaling components may contribute to the discrimination of signals that emanate from single and dual receptor engagement to control adaptive B cell responses.

  9. Recruitment of Cbl-b to B cell antigen receptor couples antigen recognition to Toll-like receptor 9 activation in late endosomes.

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    Margaret Veselits

    Full Text Available Casitas B-lineage lymphoma-b (Cbl-b is a ubiquitin ligase (E3 that modulates signaling by tagging molecules for degradation. It is a complex protein with multiple domains and binding partners that are not involved in ubiquitinating substrates. Herein, we demonstrate that Cbl-b, but not c-Cbl, is recruited to the clustered B cell antigen receptor (BCR and that Cbl-b is required for entry of endocytosed BCRs into late endosomes. The E3 activity of Cbl-b is not necessary for BCR endocytic trafficking. Rather, the ubiquitin associated (UBA domain is required. Furthermore, the Cbl-b UBA domain is sufficient to confer the receptor trafficking functions of Cbl-b on c-Cbl. Cbl-b is also required for entry of the Toll-like receptor 9 (TLR9 into late endosomes and for the in vitro activation of TLR9 by BCR-captured ligands. These data indicate that Cbl-b acts as a scaffolding molecule to coordinate the delivery of the BCR and TLR9 into subcellular compartments required for productively delivering BCR-captured ligands to TLR9.

  10. KLF2--a negative regulator of pre-B cell clonal expansion and B cell activation.

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    Rebecca Winkelmann

    Full Text Available Maturation as well as antigen-dependent activation of B cells is accompanied by alternating phases of proliferation and quiescence. We and others have previously shown that Krüppel-like factor 2 (KLF2, a regulator of T cell quiescence and migration, is upregulated in small resting precursor (pre-B cells after assembly of the immature pre-B cell receptor (pre-BCR and is downregulated upon antigen-induced proliferation of mature B cells. These findings suggest that KLF2, besides its function in maintaining follicular B cell identity, peripheral B cell homeostasis and homing of antigen-specific plasma cells to the bone marrow, also controls clonal expansion phases in the B cell lineage. Here, we demonstrate that enforced expression of KLF2 in primary pre-B cells results in a severe block of pre-BCR-induced proliferation, upregulation of the cell cycle inhibitors p21 and p27 and downregulation of c-myc. Furthermore, retroviral KLF2 transduction of primary B cells impairs LPS-induced activation, favors apoptosis and results in reduced abundance of factors, such as AID, IRF4 and BLIMP1, that control the antigen-dependent phase of B cell activation and plasma cell differentiation. Hence, we conclude that KLF2 is not only a key player in terminating pre-B cell clonal expansion but also a potent suppressor of B cell activation.

  11. Antigen Processing by Autoreactive B Cells Promotes Determinant Spreading

    Institute of Scientific and Technical Information of China (English)

    Yang D.Dai; George Carayanniotis; Eli Sercarz

    2005-01-01

    Acute primary immune responses tend to focus on few immunodominant determinants using a very limited number of T cell clones for expansion, whereas chronic inflammatory responses generally recruit a large number of different T cell clones to attack a broader range of determinants of the invading pathogens or the inflamed tissues.In T cell-mediated organ-specific autoimmune disease, a transition from the acute to the chronic phase contributes to pathogenesis, and the broadening process is called determinant spreading. The cellular components catalyzing the spreading reaction are not identified. It has been suggested that autoreactive B cells may play a central role in diversifying autoreactive T cell responses, possibly through affecting antigen processing and presentation. The clonal identity and diversity of the B cells and antibodies seem critical in regulating T cell activity and subsequent tissue damage or repair. Here, we use two autoimmune animal models, experimental autoimmune thyroiditis (EAT)and type 1 diabetes (T1D), to discuss how autoreactive B cells or antibodies alter the processing and presentation of autoantigens to regulate specific T cell response.

  12. Tolerogenicity of resting and activated B cells

    OpenAIRE

    1994-01-01

    Antigen presentation by resting splenic B cells has been shown previously to induce T helper 1 cell (Th1) anergy. In contrast to expectations, it was found here that B cells treated with F(ab')2 goat anti-mouse immunoglobulin (IgM) for 24 or 48 h also presented antigen (Ag) to Th1 cells in a manner that induced dramatic Ag-specific proliferative inactivation. The tolerogenicity of the anti-Ig-treated B cells was consistent with the observation that these B cells were only slightly more effici...

  13. The Functional Response of B Cells to Antigenic Stimulation: A Preliminary Report of Latent Tuberculosis

    Science.gov (United States)

    du Plessis, Willem J.; Kleynhans, Léanie; du Plessis, Nelita; Stanley, Kim; Malherbe, Stephanus T.; Maasdorp, Elizna; Ronacher, Katharina; Chegou, Novel N.; Walzl, Gerhard; Loxton, Andre G.

    2016-01-01

    Mycobacterium tuberculosis (M.tb) remains a successful pathogen, causing tuberculosis disease numbers to constantly increase. Although great progress has been made in delineating the disease, the host-pathogen interaction is incompletely described. B cells have shown to function as both effectors and regulators of immunity via non-humoral methods in both innate and adaptive immune settings. Here we assessed specific B cell functional interaction following stimulation with a broad range of antigens within the LTBI milieu. Our results indicate that B cells readily produce pro- and anti-inflammatory cytokines (including IL-1β, IL-10, IL-17, IL-21 and TNF-α) in response to stimulation. TLR4 and TLR9 based stimulations achieved the greatest secreted cytokine-production response and BCG stimulation displayed a clear preference for inducing IL-1β production. We also show that the cytokines produced by B cells are implicated strongly in cell-mediated communication and that plasma (memory) B cells (CD19+CD27+CD138+) is the subset with the greatest contribution to cytokine production. Collectively our data provides insight into B cell responses, where they are implicated in and quantifies responses from specific B cell phenotypes. These findings warrant further functional B cell research with a focus on specific B cell phenotypes under conditions of active TB disease to further our knowledge about the contribution of various cell subsets which could have implications for future vaccine development or refined B cell orientated treatment in the health setting. PMID:27050308

  14. OCA-B integrates B cell antigen receptor-, CD40L- and IL 4-mediated signals for the germinal center pathway of B cell development.

    OpenAIRE

    Qin, X F; Reichlin, A; Luo, Y.; Roeder, R. G.; Nussenzweig, M.C.

    1998-01-01

    Many of the key decisions in lymphocyte differentiation and activation are dependent on integration of antigen receptor and co-receptor signals. Although there is significant understanding of these receptors and their signaling pathways, little is known about the molecular requirements for signal integration at the level of activation of gene expression. Here we show that in primary B cells, expression of the B-cell specific transcription coactivator OCA-B (also known as OBF-1 or Bob-1) is re...

  15. The B cell antigen receptor and overexpression of MYC can cooperate in the genesis of B cell lymphomas.

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    Yosef Refaeli

    2008-06-01

    Full Text Available A variety of circumstantial evidence from humans has implicated the B cell antigen receptor (BCR in the genesis of B cell lymphomas. We generated mouse models designed to test this possibility directly, and we found that both the constitutive and antigen-stimulated state of a clonal BCR affected the rate and outcome of lymphomagenesis initiated by the proto-oncogene MYC. The tumors that arose in the presence of constitutive BCR differed from those initiated by MYC alone and resembled chronic B cell lymphocytic leukemia/lymphoma (B-CLL, whereas those that arose in response to antigen stimulation resembled large B-cell lymphomas, particularly Burkitt lymphoma (BL. We linked the genesis of the BL-like tumors to antigen stimulus in three ways. First, in reconstruction experiments, stimulation of B cells by an autoantigen in the presence of overexpressed MYC gave rise to BL-like tumors that were, in turn, dependent on both MYC and the antigen for survival and proliferation. Second, genetic disruption of the pathway that mediates signaling from the BCR promptly killed cells of the BL-like tumors as well as the tumors resembling B-CLL. And third, growth of the murine BL could be inhibited by any of three distinctive immunosuppressants, in accord with the dependence of the tumors on antigen-induced signaling. Together, our results provide direct evidence that antigenic stimulation can participate in lymphomagenesis, point to a potential role for the constitutive BCR as well, and sustain the view that the constitutive BCR gives rise to signals different from those elicited by antigen. The mouse models described here should be useful in exploring further the pathogenesis of lymphomas, and in preclinical testing of new therapeutics.

  16. Transgelin-2 in B-Cells Controls T-Cell Activation by Stabilizing T Cell - B Cell Conjugates

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    Chae, Myoung-Won; Kim, Hye-Ran; Kim, Chang-Hyun; Jun, Chang-Duk; Park, Zee-Yong

    2016-01-01

    The immunological synapse (IS), a dynamic and organized junction between T-cells and antigen presenting cells (APCs), is critical for initiating adaptive immunity. The actin cytoskeleton plays a major role in T-cell reorganization during IS formation, and we previously reported that transgelin-2, an actin-binding protein expressed in T-cells, stabilizes cortical F-actin, promoting T-cell activation in response to antigen stimulation. Transgelin-2 is also highly expressed in B-cells, although no specific function has been reported. In this study, we found that deficiency in transgelin-2 (TAGLN2-/-) in B-cells had little effect on B-cell development and activation, as measured by the expression of CD69, MHC class II molecules, and CD80/86. Nevertheless, in B-cells, transgelin-2 accumulated in the IS during the interaction with T-cells. These results led us to hypothesize that transgelin-2 may also be involved in IS stability in B-cells, thereby influencing T-cell function. Notably, we found that transgelin-2 deficiency in B-cells reduced T-cell activation, as determined by the release of IL-2 and interferon-γ and the expression of CD69. Furthermore, the reduced T-cell activation was correlated with reduced B-cell–T-cell conjugate formation. Collectively, these results suggest that actin stability in B-cells during IS formation is critical for the initiation of adaptive T-cell immunity. PMID:27232882

  17. Identification of a Coccidioides immitis antigen 2 domain that expresses B-cell-reactive epitopes.

    OpenAIRE

    Zhu, Y; Tryon, V; Magee, D M; Cox, R. A.

    1997-01-01

    Antigen 2 (Ag2), a major immunoreactive component of Coccidioides immitis mycelium- and spherule-phase cell walls, was recently cloned in our laboratory and was shown to elicit T-cell responses in Coccidioides-immune mice. In this investigation, we evaluated recombinant Ag2 (rAg2) and PCR-generated Ag2 truncations for expression of B-cell-reactive epitopes in enzyme-linked immunosorbent and immunoblot assays with sera from patients with active coccidioidomycosis, a hyperimmune goat anti-Ag2 s...

  18. Role of Calcium Signaling in B Cell Activation and Biology.

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    Baba, Yoshihiro; Kurosaki, Tomohiro

    2016-01-01

    Increase in intracellular levels of calcium ions (Ca2+) is one of the key triggering signals for the development of B cell response to the antigen. The diverse Ca2+ signals finely controlled by multiple factors participate in the regulation of gene expression, B cell development, and effector functions. B cell receptor (BCR)-initiated Ca2+ mobilization is sourced from two pathways: one is the release of Ca2+ from the intracellular stores, endoplasmic reticulum (ER), and other is the prolonged influx of extracellular Ca2+ induced by depleting the stores via store-operated calcium entry (SOCE) and calcium release-activated calcium (CRAC) channels. The identification of stromal interaction molecule 1(STIM1), the ER Ca2+ sensor, and Orai1, a key subunit of the CRAC channel pore, has now provided the tools to understand the mode of Ca2+ influx regulation and physiological relevance. Herein, we discuss our current understanding of the molecular mechanisms underlying BCR-triggered Ca2+ signaling as well as its contribution to the B cell biological processes and diseases. PMID:26369772

  19. Relationship between Fc receptors, antigen-binding sites on T and B cells, and H-2 complex-associated determinants.

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    Basten, A; Miller, J F; Abraham, R

    1975-03-01

    The relationship between H-2 complex-associated determinants, Fc receptors, and specific antigen-recognition sites on T and B cells was examined by binding and functional assays. The Fc receptor was detected by radiolabeled immune complexes or aggregated human IgG. Both these reagents selectively bound to B cells, not to T cells. When spleen cells, from mice primed to several antigens, were exposed to highly substituted radioactive aggregates, their capacity to transfer both a direct and indirect plaque-forming cell response to these antigens was abrogated. Addition of B cells, but not of T cells, restored responsiveness. Complexed Ig binding to Fc receptors was prevented by pretreatment of mixed lymphoid cell populations with antisera directed against membrane components on the same cell (e.g., H-2) and on other cells (e.g., theta). The lack of specificity of inhibition was thought to be due to the formation on cell surfaces of antigen-antibody complexes which would then attach to the Fc receptor during the incubation precedure. Specific blockade of the Fc receptor during the incubation procedure. Specific blockade of the Fc receptor however occurred when B cells were pretreated with the Fab fragments of anti-H-2 antibody. This was demonstrated autoradiographically and by inhibition of aggregate-induced suicide. The blocking activity of ante-H-2 Fab was removed by absorption with spleen cells from thymectomized irradiated mice but not with thymus cells of appropriate specificity. This suggested that the antibodies involved had specificity for determinants on the B-cell membrane distinct from those coded by the K or D end of the H-2 complex, and either absent from, or poorly represented on, thymus cells. Specific antigen-induced suicide of B cells was achieved simply by incubating the cells with radioactive antigen in the cold. T-cell suicide on the other hand required that the 125I-labeled antigen be presented to the T cells at 37 degrees-C on the surface of

  20. Invariant chain induces B cell maturation in a process that is independent of its chaperonic activity

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    Matza, Didi; Lantner, Frida; Bogoch, Yoel; Flaishon, Liat; Hershkoviz, Rami; Shachar, Idit

    2002-01-01

    Early stages of B cell development take place in the bone marrow, resulting in formation of immature B cells, which migrate to the spleen for their final differentiation into mature cells. This final maturation step is essential for B cells to become responsive to antigens and to participate in the immune response. Previously, we showed that the MHC class II chaperone, invariant chain (Ii), controls the differentiation of B cells from the immature to the mature stage. In this study, by generating transgenic mice expressing truncated Ii lacking its luminal domain, we could dissect the chaperonin activity of Ii from its role in B cell maturation. We demonstrate in vivo that Ii N-terminal domain is directly involved in the maturation of B cells and is sufficient to promote B cell differentiation. PMID:11867743

  1. Autoantibodies in autoimmune thyroid disease promote immune complex formation with self antigens and increase B cell and CD4+ T cell proliferation in response to self antigens

    DEFF Research Database (Denmark)

    Nielsen, Claus Henrik; Hegedüs, Laszlo; Leslie, Robert Graham Quinton

    2004-01-01

    B cells are centrally involved as antigen-presenting cells in certain autoimmune diseases. To establish whether autoantibodies form immune complexes (IC) with self-antigens in autoimmune thyroid disease (AITD) and promote B cell uptake of self-antigen, sera from patients with Hashimoto's thyroidi......B cells are centrally involved as antigen-presenting cells in certain autoimmune diseases. To establish whether autoantibodies form immune complexes (IC) with self-antigens in autoimmune thyroid disease (AITD) and promote B cell uptake of self-antigen, sera from patients with Hashimoto...

  2. Docking of B-cell epitope antigen to specific hepatitis B antibody

    Indian Academy of Sciences (India)

    R Rajkannan; E J Padma Malar

    2007-09-01

    The interaction of pres1 region of hepatitis B virus B-cell epitope antigen with specific hepatitis B neutralizing monoclonal antibody was examined by docking study. We modelled the 3D complex structure of B-cell epitope antigen residues CTTPAQGNSMFPSCCCTKPTDGNCY by homology modelling and docked it with the crystal structure of monoclonal antibody specific for the pres1 region of the hepatitis B virus. At the optimized docked conformation, the interactions between the amino acids of antigen and antibody were examined. It is found that the docked complex is stabilized by 59.3 kcal/mol. The stability of the docked antigen-antibody complex is due to hydrogen bonding and van der Waals interactions. The amino acids of the antigen and antibody responsible for the interaction were identified.

  3. Persistent Polyclonal B Cell Lymphocytosis B Cells Can Be Activated through CD40-CD154 Interaction

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    Emmanuelle Dugas-Bourdages

    2014-01-01

    Full Text Available Persistent polyclonal B cell lymphocytosis (PPBL is a rare disorder, diagnosed primarily in adult female smokers and characterized by an expansion of CD19+CD27+IgM+ memory B cells, by the presence of binucleated lymphocytes, and by a moderate elevation of serum IgM. The clinical course is usually benign, but it is not known whether or not PPBL might be part of a process leading to the emergence of a malignant proliferative disorder. In this study we sought to investigate the functional response of B cells from patients with PPBL by use of an optimal memory B cell culture model based on the CD40-CD154 interaction. We found that the proliferation of PPBL B cells was almost as important as that of B cells from normal controls, resulting in high immunoglobulin secretion with in vitro isotypic switching. We conclude that the CD40-CD154 activation pathway is functional in the memory B cell population of PPBL patients, suggesting that the disorder may be due to either a dysfunction of other cells in the microenvironment or a possible defect in another B cell activation pathway.

  4. B cell infection and activation by rabies virus-based vaccines.

    Science.gov (United States)

    Lytle, Andrew G; Norton, James E; Dorfmeier, Corin L; Shen, Shixue; McGettigan, James P

    2013-08-01

    Replication-deficient rabies viruses (RABV) are promising rabies postexposure vaccines due to their prompt and potent stimulation of protective virus neutralizing antibody titers, which are produced in mice by both T-dependent and T-independent mechanisms. To promote such early and robust B cell stimulation, we hypothesized that live RABV-based vaccines directly infect B cells, thereby activating a large pool of antigen-presenting cells (APCs) capable of providing early priming and costimulation to CD4(+) T cells. In this report, we show that live RABV-based vaccine vectors efficiently infect naive primary murine and human B cells ex vivo. Infection of B cells resulted in the significant upregulation of early markers of B cell activation and antigen presentation, including CD69, major histocompatibility complex class II (MHC-II), and CD40 in murine B cells or HLA-DR and CD40 in human B cells compared to mock-infected cells or cells treated with an inactivated RABV-based vaccine. Furthermore, primary B cells infected with a live RABV expressing ovalbumin were able to prime and stimulate naive CD4(+) OT-II T cells to proliferate and to secrete interleukin-2 (IL-2), demonstrating a functional consequence of B cell infection and activation by live RABV-based vaccine vectors. We propose that this direct B cell stimulation by live RABV-based vaccines is a potential mechanism underlying their induction of early protective T cell-dependent B cell responses, and that designing live RABV-based vaccines to infect and activate B cells represents a promising strategy to develop a single-dose postexposure rabies vaccine where the generation of early protective antibody titers is critical. PMID:23760241

  5. Kinetics of antigen specific and non-specific polyclonal B-cell responses during lethal Plasmodium yoelii malaria

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    Laurence Rolland

    1992-06-01

    Full Text Available In order to study the kinetics and composition of the polyclonal B-cell activation associated to malaria infection, antigen-specific and non-specific B-cell responses were evaluated in the spleens of mice infected with Plasmodium yoelii 17 XL or injected with lysed erythrocytes or plasma from P. yoelii infected mice or with P. falciparum culture supernatants. Spleen/body weigth ratio, numbers of nucleated spleen cells and Immunoglobulin-containing and Immunoglobulin-secreting cells increased progressively during the course of infection,in parallel to the parasitemia. A different pattern of kinetics was observed when anti-sheep red blood cell and anti-trinitrophenylated-sheep red blood cell plaque forming cells response were studied: maximum values were observed at early stages of infection, whereas the number of total Immunoglobulin-containing and Immunoglobulin-secreting cells were not yet altered. Conversely, at the end of infection, when these latter values reached their maximum, the anti-sheep red blood cell and anti-trinitrophenylated-sheep red blood cell specific responses were normal or even infranormal. In mice injected with Plasmodium-derived material, a higher increase in antigen-specific PFC was observed, as compared to the increase of Immunoglobulin-containing and Immunoglobulin-secreting cell numbers. This suggested a "preferential" (antigen-plus mitogen-induced stimulation of antigen-specific cells rather than a generalized non-specific (mitogen-induced triggering of B-lymphocytes. On the basis of these and previous results, it is suggested that polyclonal B-cell activation that takes place during the course of infection appears as a result of successive waves of antigen-specific B-cell activation.

  6. Targeting of antigens to B cells augments antigen-specific T-cell responses and breaks immune tolerance to tumor-associated antigen MUC1

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    Ding, Chuanlin; Wang, Li; Marroquin, Jose

    2008-01-01

    B cells are antibody (Ab)–secreting cells as well as potent antigen (Ag)–presenting cells that prime T-cell activation, which evokes great interest in their use for vaccine development. Here, we targeted ovalbumin (OVA) to B cells via CD19 and found that a single low dose of anti–CD19-OVA conjugates, but not isotype mAb-OVA, stimulated augmented CD4 and CD8 T-cell proliferation and expansion. Administration of TLR9 agonist CpG could significantly enhance long-term T-cell survival. Similar results were obtained when the tumor-associated Ag MUC1 was delivered to B cells. MUC1 transgenic (Tg) mice were previously found to lack effective T-cell help and produce low-titer of anti-MUC1 Abs after vaccination. Targeting MUC1 to B cells elicited high titer of anti-MUC1 Abs with different isotypes, predominantly IgG2a and IgG2b, in MUC1 Tg mice. The isotype switching of anti-MUC1 Ab was CD4 dependent. In addition, IFN-γ–producing CD8 T cells and in vivo cytolytic activity were significantly increased in these mice. The mice also showed significant resistance to MUC1+ lymphoma cell challenge both in the prophylactic and therapeutic settings. We conclude that Ags targeting to B cells stimulate CD4 and CD8 T-cell responses as well as Th-dependent humoral immune responses. PMID:18669871

  7. Cutaneous lymphocyte antigen expression on human effector B cells depends on the site and on the nature of antigen encounter.

    Science.gov (United States)

    Kantele, Anu; Savilahti, Erkki; Tiimonen, Heidi; Iikkanen, Katja; Autio, Soile; Kantele, Jussi M

    2003-12-01

    In contrast to T cells, information on skin-homing B cells expressing the cutaneous lymphocyte antigen (CLA) is sparse. CLA expression on human B cells was investigated among circulating immunoglobulin-secreting cells (ISC) and among antigen-specific antibody-secreting cells (ASC) elicited by parenteral, oral or rectal primary immunization, or by parenteral or oral secondary immunization with Salmonella typhi Ty21a. CLA expression was examined by combining cell sorting with an enzyme-linked immunospot assay. Among all ISC, the proportion of CLA(+) cells was 13-21%. Parenteral immunization induced antigen-specific ASC of which 13% were CLA(+), while oral and rectal immunizations were followed by only 1% of CLA(+) ASC (p<0.001). Oral re-immunization was followed by an up-regulation of CLA (34-48%) regardless of the route of priming. Parenteral re-immunization elicited ASC of which 9-14% were CLA(+). In conclusion, the expression of CLA on human effector B cells depends on the site of antigen encounter: intestinal stimulation elicits cells with no CLA, while parenteral encounter elicits significant numbers of CLA(+) cells. Even though primary antigen encounter in the intestine failed to stimulate CLA expression, up-regulation of CLA was found upon intestinal antigen re-encounter. These findings may be of relevance in the pathogenesis of some cutaneous disorders. PMID:14635035

  8. Improved method for linear B-cell epitope prediction using antigen's primary sequence.

    Directory of Open Access Journals (Sweden)

    Harinder Singh

    Full Text Available One of the major challenges in designing a peptide-based vaccine is the identification of antigenic regions in an antigen that can stimulate B-cell's response, also called B-cell epitopes. In the past, several methods have been developed for the prediction of conformational and linear (or continuous B-cell epitopes. However, the existing methods for predicting linear B-cell epitopes are far from perfection. In this study, an attempt has been made to develop an improved method for predicting linear B-cell epitopes. We have retrieved experimentally validated B-cell epitopes as well as non B-cell epitopes from Immune Epitope Database and derived two types of datasets called Lbtope_Variable and Lbtope_Fixed length datasets. The Lbtope_Variable dataset contains 14876 B-cell epitope and 23321 non-epitopes of variable length where as Lbtope_Fixed length dataset contains 12063 B-cell epitopes and 20589 non-epitopes of fixed length. We also evaluated the performance of models on above datasets after removing highly identical peptides from the datasets. In addition, we have derived third dataset Lbtope_Confirm having 1042 epitopes and 1795 non-epitopes where each epitope or non-epitope has been experimentally validated in at least two studies. A number of models have been developed to discriminate epitopes and non-epitopes using different machine-learning techniques like Support Vector Machine, and K-Nearest Neighbor. We achieved accuracy from ∼54% to 86% using diverse s features like binary profile, dipeptide composition, AAP (amino acid pair profile. In this study, for the first time experimentally validated non B-cell epitopes have been used for developing method for predicting linear B-cell epitopes. In previous studies, random peptides have been used as non B-cell epitopes. In order to provide service to scientific community, a web server LBtope has been developed for predicting and designing B-cell epitopes (http://crdd.osdd.net/raghava/lbtope/.

  9. T Cells Expressing CD19/CD20 Bispecific Chimeric Antigen Receptors Prevent Antigen Escape by Malignant B Cells.

    Science.gov (United States)

    Zah, Eugenia; Lin, Meng-Yin; Silva-Benedict, Anne; Jensen, Michael C; Chen, Yvonne Y

    2016-06-01

    The adoptive transfer of T cells expressing anti-CD19 chimeric antigen receptors (CARs) has shown remarkable curative potential against advanced B-cell malignancies, but multiple trials have also reported patient relapses due to the emergence of CD19-negative leukemic cells. Here, we report the design and optimization of single-chain, bispecific CARs that trigger robust cytotoxicity against target cells expressing either CD19 or CD20, two clinically validated targets for B-cell malignancies. We determined the structural parameters required for efficient dual-antigen recognition, and we demonstrate that optimized bispecific CARs can control both wild-type B-cell lymphoma and CD19(-) mutants with equal efficiency in vivo To our knowledge, this is the first bispecific CAR capable of preventing antigen escape by performing true OR-gate signal computation on a clinically relevant pair of tumor-associated antigens. The CD19-OR-CD20 CAR is fully compatible with existing T-cell manufacturing procedures and implementable by current clinical protocols. These results present an effective solution to the challenge of antigen escape in CD19 CAR T-cell therapy, and they highlight the utility of structure-based rational design in the development of receptors with higher-level complexity. Cancer Immunol Res; 4(6); 498-508. ©2016 AACRSee related Spotlight by Sadelain, p. 473. PMID:27059623

  10. B Cell Infection and Activation by Rabies Virus-Based Vaccines

    OpenAIRE

    Lytle, Andrew G.; Norton, James E.; Dorfmeier, Corin L.; Shen, Shixue; McGettigan, James P.

    2013-01-01

    Replication-deficient rabies viruses (RABV) are promising rabies postexposure vaccines due to their prompt and potent stimulation of protective virus neutralizing antibody titers, which are produced in mice by both T-dependent and T-independent mechanisms. To promote such early and robust B cell stimulation, we hypothesized that live RABV-based vaccines directly infect B cells, thereby activating a large pool of antigen-presenting cells (APCs) capable of providing early priming and costimulat...

  11. Chimeric Antigen Receptor Therapy for B-cell Malignancies

    Directory of Open Access Journals (Sweden)

    David L Porter, Michael Kalos, Zhaohui Zheng, Bruce Levine, Carl June

    2011-01-01

    Full Text Available We presented data showing that the CART-19 cells expressing the 4-1BB signaling domain can have unprecedented and massive in-vivo expansion, traffic to tumor sites, persist long term in vivo, and induce rapid and potent anti-tumor activity in chemotherapy refractory CLL patients.

  12. Chimeric Antigen Receptor Therapy for B-cell Malignancies

    OpenAIRE

    Porter, David L.; Kalos, Michael; Zheng, Zhaohui; Levine, Bruce; June, Carl

    2011-01-01

    We presented data showing that the CART-19 cells expressing the 4-1BB signaling domain can have unprecedented and massive in-vivo expansion, traffic to tumor sites, persist long term in vivo, and induce rapid and potent anti-tumor activity in chemotherapy refractory CLL patients.

  13. An antigen-specific, four-color, B-cell FluoroSpot assay utilizing tagged antigens for detection.

    Science.gov (United States)

    Jahnmatz, Peter; Bengtsson, Theresa; Zuber, Bartek; Färnert, Anna; Ahlborg, Niklas

    2016-06-01

    The FluoroSpot assay, a variant of ELISpot utilizing fluorescent detection, has so far been used primarily for assessment of T cells, where simultaneous detection of several cytokines has allowed a more qualitative analysis of functionally distinct T cells. The potential to measure multiple analytes also presents several advantages when analyzing B cells. Our aim was to develop a B-cell FluoroSpot assay adaptable to studies of a variety of antigens. The assay utilizes anti-IgG antibodies immobilized in 96-well filter membrane plates. During cell culture, IgG antibodies secreted by antibody-secreting cells (ASCs) are captured in the vicinity of each of these cells and the specificity of single ASCs is defined using antigens for detection. The antigens were labeled with biotin or peptide tags enabling secondary detection with fluorophore-conjugated streptavidin or tag-specific antibodies. The assay, utilizing up to four different tag systems and fluorophores simultaneously, was evaluated using hybridomas and immunized splenocytes as ASCs. Assay variants were developed that could: i) identify multiple ASCs with different antigen specificities; ii) detect ASCs showing cross-reactivity with different but related antigens; and iii) define the antigen-specificity and, by including anti-IgG subclass detection reagents, simultaneously determine the IgG subclass of antibodies secreted by ASCs. As demonstrated here, the B-cell FluoroSpot assay using tag-based detection systems provides a versatile and powerful tool to investigate antibody responses by individual cells that can be readily adapted to studies of a variety of antigen-specific ASCs. PMID:26930550

  14. Generation of Recombinant Monoclonal Antibodies from Immunised Mice and Rabbits via Flow Cytometry and Sorting of Antigen-Specific IgG+ Memory B Cells.

    Directory of Open Access Journals (Sweden)

    Dale O Starkie

    Full Text Available Single B cell screening strategies, which avoid both hybridoma fusion and combinatorial display, have emerged as important technologies for efficiently sampling the natural antibody repertoire of immunized animals and humans. Having access to a range of methods to interrogate different B cell subsets provides an attractive option to ensure large and diverse panels of high quality antibody are produced. The generation of multiple antibodies and having the ability to find rare B cell clones producing IgG with unique and desirable characteristics facilitates the identification of fit-for-purpose molecules that can be developed into therapeutic agents or research reagents. Here, we describe a multi-parameter flow cytometry single-cell sorting technique for the generation of antigen-specific recombinant monoclonal antibodies from single IgG+ memory B cells. Both mouse splenocytes and rabbit PBMC from immunised animals were used as a source of B cells. Reagents staining both B cells and other unwanted cell types enabled efficient identification of class-switched IgG+ memory B cells. Concurrent staining with antigen labelled separately with two spectrally-distinct fluorophores enabled antigen-specific B cells to be identified, i.e. those which bind to both antigen conjugates (double-positive. These cells were then typically sorted at one cell per well using FACS directly into a 96-well plate containing reverse transcriptase reaction mix. Following production of cDNA, PCR was performed to amplify cognate heavy and light chain variable region genes and generate transcriptionally-active PCR (TAP fragments. These linear expression cassettes were then used directly in a mammalian cell transfection to generate recombinant antibody for further testing. We were able to successfully generate antigen-specific recombinant antibodies from both the rabbit and mouse IgG+ memory B cell subset within one week. This included the generation of an anti-TNFR2 blocking

  15. Innate response activator B cells: origins and functions.

    Science.gov (United States)

    Chousterman, Benjamin G; Swirski, Filip K

    2015-10-01

    Innate response activator (IRA) B cells are a subset of B-1a derived B cells that produce the growth factors granulocyte macrophage colony stimulating factor and IL-3. In mouse models of sepsis and pneumonia, B-1a B cells residing in serosal sites recognize bacteria, migrate to the spleen or lung, and differentiate to IRA B cells that then contribute to the host response by amplifying inflammation and producing polyreactive IgM. In atherosclerosis, IRA B cells accumulate in the spleen, where they promote extramedullary hematopoiesis and activate classical dendritic cells. In this review, we focus on the ontogeny and function of IRA B cells in acute and chronic inflammation. PMID:25957266

  16. Expression of MHC class II antigens in human B-cell leukaemia and non-Hodgkin's lymphoma.

    OpenAIRE

    Guy, K.; Krajewski, A S; Dewar, A E

    1986-01-01

    In this review we have summarized our experiences of serological analysis of MHC class II antigen expression in human B cell malignant disease. Cells from a large number of cases of B-cell chronic lymphocytic leukaemia (CLL) and non-Hodgkin's lymphoma (NHL) have been examined for expression of class II antigens. Using a number of monoclonal antibodies which in some cases are specific for class II subregion products (DP, DQ and DR), MHC class II antigens were detected by indirect immunofluores...

  17. γδ T cells recognize a microbial encoded B cell antigen to initiate a rapid antigen specific Interleukin 17 response

    Science.gov (United States)

    Zeng, Xun; Wei, Yu-ling; Huang, Jun; Newell, Evan W.; Yu, Hongxiang; Kidd, Brian A.; Kuhns, Michael S.; Waters, Ray W.; Davis, Mark M.; Weaver, Casey T.; Chien, Yueh-hsiu

    2012-01-01

    Summary γδ T cells contribute uniquely to host immune defense. However, how they function remains an enigma. Although it is unclear what most γδ T cells recognize, common dogma asserts that they recognize self-antigens. While they are the major initial Interleukin-17 (IL-17) producers in infections, it is unclear what is required to trigger these cells to act. Here, we report that a noted B cell antigen, the algae protein-phycoerythrin (PE) is an antigen for murine and human γδ T cells. PE also stained specific bovine γδ T cells. Employing this specificity, we demonstrated that antigen recognition, but not extensive clonal expansion, was required to activate naïve γδ T cells to make IL-17. In this activated state, γδ T cells gained the ability to respond to cytokine signals that perpetuated the IL-17 production. These results underscore the adaptability of lymphocyte antigen receptors and suggest a previously unrecognized antigen-driven rapid response in protective immunity prior to the maturation of classical adaptive immunity. PMID:22960222

  18. Non-CpG Oligonucleotides Exert Adjuvant Effects by Enhancing Cognate B Cell-T Cell Interactions, Leading to B Cell Activation, Differentiation, and Isotype Switching

    Directory of Open Access Journals (Sweden)

    Melinda Herbáth

    2015-01-01

    Full Text Available Natural and synthetic nucleic acids are known to exert immunomodulatory properties. Notably, nucleic acids are known to modulate immune function via several different pathways and various cell types, necessitating a complex interpretation of their effects. In this study we set out to compare the effects of a CpG motif containing oligodeoxynucleotide (ODN with those of a control and an inhibitory non-CpG ODN during cognate B cell-T cell interactions. We employed an antigen presentation system using splenocytes from TCR transgenic DO11.10 mice and the ovalbumin peptide recognized by the TCR as model antigen. We followed early activation events by measuring CD69 expression, late activation by MHC class II expression, cell division and antibody production of switched, and nonswitched isotypes. We found that both of the tested non-CpG ODN exerted significant immunomodulatory effects on early T cell and on late B cell activation events. Importantly, a synergism between non-CpG effects and T cell help acting on B cells was observed, resulting in enhanced IgG production following cognate T cell-B cell interactions. We propose that non-CpG ODN may perform as better adjuvants when a strong antigen-independent immune activation, elicited by CpG ODNs, is undesirable.

  19. CD83 Modulates B Cell Activation and Germinal Center Responses.

    Science.gov (United States)

    Krzyzak, Lena; Seitz, Christine; Urbat, Anne; Hutzler, Stefan; Ostalecki, Christian; Gläsner, Joachim; Hiergeist, Andreas; Gessner, André; Winkler, Thomas H; Steinkasserer, Alexander; Nitschke, Lars

    2016-05-01

    CD83 is a maturation marker for dendritic cells. In the B cell lineage, CD83 is expressed especially on activated B cells and on light zone B cells during the germinal center (GC) reaction. The function of CD83 during GC responses is unclear. CD83(-/-) mice have a strong reduction of CD4(+) T cells, which makes it difficult to analyze a functional role of CD83 on B cells during GC responses. Therefore, in the present study we generated a B cell-specific CD83 conditional knockout (CD83 B-cKO) model. CD83 B-cKO B cells show defective upregulation of MHC class II and CD86 expression and impaired proliferation after different stimuli. Analyses of GC responses after immunization with various Ags revealed a characteristic shift in dark zone and light zone B cell numbers, with an increase of B cells in the dark zone of CD83 B-cKO mice. This effect was not accompanied by alterations in the level of IgG immune responses or by major differences in affinity maturation. However, an enhanced IgE response was observed in CD83 B-cKO mice. Additionally, we observed a strong competitive disadvantage of CD83-cKO B cells in GC responses in mixed bone marrow chimeras. Furthermore, infection of mice with Borrelia burgdorferi revealed a defect in bacterial clearance of CD83 B-cKO mice with a shift toward a Th2 response, indicated by a strong increase in IgE titers. Taken together, our results show that CD83 is important for B cell activation and modulates GC composition and IgE Ab responses in vivo. PMID:26983787

  20. B-cell exposure to self-antigen induces IL-10 producing B cells as well as IL-6- and TNF-α-producing B-cell subsets in healthy humans

    DEFF Research Database (Denmark)

    Langkjær, Anina; Kristensen, Birte; Hansen, Bjarke E;

    2012-01-01

    Human B cells are able to secrete IL-10 after stimulation with mitogens, but their ability to produce IL-10 and regulate T-cell responses after stimulation with self-antigens is unclear. We co-cultured thyroglobulin-pulsed B cells from healthy donors with autologous T cells and observed production...... of IL-10 and TGF-β, in addition to TNF-α and IL-6. Pulsing with foreign antigen, tetanus toxoid (TT), induced a Th1-response with minimal IL-10 production. After thyroglobulin-pulsing, 1.10±0.50% of B cells and 1.00±0.20% of CD4(+) T cells produced IL-10, compared to 0.29±0.19% of B cells (P=0.......01) and 0.13±0.15% of CD4(+) T cells (P=0.006) following TT-pulsing. Thyroglobulin-stimulated, IL-10-secreting B cells were enriched within CD5(+) and CD24(high) cells. While thyroglobulin-pulsed B cells induced only modest proliferation of CD4(+) T cells, B cells pulsed with TT induced vigorous...

  1. γδ T cells recognize a microbial encoded B cell antigen to initiate a rapid antigen-specific interleukin-17 response.

    Science.gov (United States)

    Zeng, Xun; Wei, Yu-Ling; Huang, Jun; Newell, Evan W; Yu, Hongxiang; Kidd, Brian A; Kuhns, Michael S; Waters, Ray W; Davis, Mark M; Weaver, Casey T; Chien, Yueh-hsiu

    2012-09-21

    γδ T cells contribute uniquely to immune competence. Nevertheless, how they function remains an enigma. It is unclear what most γδ T cells recognize, what is required for them to mount an immune response, and how the γδ T cell response is integrated into host immune defense. Here, we report that a noted B cell antigen, the algae protein phycoerythrin (PE), is a murine and human γδ T cell antigen. Employing this specificity, we demonstrated that antigen recognition activated naive γδ T cells to make interleukin-17 and respond to cytokine signals that perpetuate the response. High frequencies of antigen-specific γδ T cells in naive animals and their ability to mount effector response without extensive clonal expansion allow γδ T cells to initiate a swift, substantial response. These results underscore the adaptability of lymphocyte antigen receptors and suggest an antigen-driven rapid response in protective immunity prior to the maturation of classical adaptive immunity. PMID:22960222

  2. Primary cardiac diffuse large B-cell lymphoma with activated B-cell-like phenotype

    Directory of Open Access Journals (Sweden)

    Vijaya Gadage

    2011-01-01

    Full Text Available Primary cardiac lymphoma (PCL is a rare and fatal disorder. It may often mimic other common cardiac tumors like cardiac myxoma because of similarities in the clinical presentation. We report a case of PCL of diffuse large B-cell type, in a 38-year-old, immunocompetent male who presented with superior vena cava syndrome that was excised as a myxoma. Histology revealed a large cell population diffusely and strongly expressing CD45, CD20, MUM1/IRF4 and FOXP1 hinting at an activated B-cell (ABC-like phenotype. After four cycles of Rituximab with CHOP (cyclophosphamide, hydroxydaunorubicin, Oncovin, and prednisolone the tumor regressed completely but the patient had a relapse and subsequently succumbed to the disease confirming the aggressive nature. The aggressive behavior of PCL may be possibly linked to its ABC-like origin.

  3. Single molecule analysis of B cell receptor motion during signaling activation

    Science.gov (United States)

    Rey Suarez, Ivan; Koo, Peter; Mochrie, Simon; Song, Wenxia; Upadhyaya, Arpita

    B cells are an essential part of the adaptive immune system. They patrol the body looking for signs of infection in the form of antigen on the surface of antigen presenting cells. The binding of the B cell receptor (BCR) to antigen induces signaling cascades that lead to B cell activation and eventual production of high affinity antibodies. During activation, BCR organize into signaling microclusters, which are platforms for signal amplification. The physical processes underlying receptor movement and aggregation are not well understood. Here we study the dynamics of single BCRs on activated murine primary B cells using TIRF imaging and single particle tracking. The tracks obtained are analyzed using perturbation expectation-maximization (pEM) a systems-level analysis that allows the identification of different short-time diffusive states from a set of single particle tracks. We identified five different diffusive states on wild type cells, which correspond to different molecular states of the BCR. By using actin polymerization inhibitors and mutant cells lacking important actin regulators we were able to identify the BCR molecule configuration associated with each diffusive state.

  4. Antigen-specific T8+ human clone of cells with a nonspecific augmenting function on the T4 cell-B cell helper interaction

    International Nuclear Information System (INIS)

    The authors isolated a T8+ T3+ Ia+ clone of cells from the peripheral blood mononuclear cells of a healthy subject. The clone was expanded and maintained with autologous feed cells, interleukin 2, and a streptococcal antigen. The T8+ clone of cells responded specifically to the streptococcal antigen, in the absence of accessory cells,and released a soluble factor. Both the cloned cells and the corresponding soluble factor expressed augmenting helper but not suppressor activity. The augmenting helper activity for B cell antibody synthesis was demonstrable only in the presence of autologous T 4 cells. Radioimmunoassay was used to measure antibodies. Although stimulation of the T8+ cloned cells was antigen-specific, the resulting soluble factor elicited nonspecific antibody synthesis in the presence of T4 and B cells. The T8+ cloned cell-derived factor was adsorbed by B cells but not by T4 cells. Preliminary studies suggest that the factor has the properties of a B cell growth factor. They suggest that the T8+ population consists of functionally heterogeneous cell subsets, some that have suppressor function and others that augment the T4+ helper-inducer activity in B cell antibody synthesis

  5. CD40 signaling synergizes with TLR-2 in the BCR independent activation of resting B cells.

    Science.gov (United States)

    Jain, Shweta; Chodisetti, Sathi Babu; Agrewala, Javed N

    2011-01-01

    Conventionally, signaling through BCR initiates sequence of events necessary for activation and differentiation of B cells. We report an alternative approach, independent of BCR, for stimulating resting B (RB) cells, by involving TLR-2 and CD40--molecules crucial for innate and adaptive immunity. CD40 triggering of TLR-2 stimulated RB cells significantly augments their activation, proliferation and differentiation. It also substantially ameliorates the calcium flux, antigen uptake capacity and ability of B cells to activate T cells. The survival of RB cells was improved and it increases the number of cells expressing activation induced deaminase (AID), signifying class switch recombination (CSR). Further, we also observed increased activation rate and decreased threshold period required for optimum stimulation of RB cells. These results corroborate well with microarray gene expression data. This study provides novel insights into coordination between the molecules of innate and adaptive immunity in activating B cells, in a BCR independent manner. This strategy can be exploited to design vaccines to bolster B cell activation and antigen presenting efficiency, leading to faster and better immune response. PMID:21674065

  6. CD40 signaling synergizes with TLR-2 in the BCR independent activation of resting B cells.

    Directory of Open Access Journals (Sweden)

    Shweta Jain

    Full Text Available Conventionally, signaling through BCR initiates sequence of events necessary for activation and differentiation of B cells. We report an alternative approach, independent of BCR, for stimulating resting B (RB cells, by involving TLR-2 and CD40--molecules crucial for innate and adaptive immunity. CD40 triggering of TLR-2 stimulated RB cells significantly augments their activation, proliferation and differentiation. It also substantially ameliorates the calcium flux, antigen uptake capacity and ability of B cells to activate T cells. The survival of RB cells was improved and it increases the number of cells expressing activation induced deaminase (AID, signifying class switch recombination (CSR. Further, we also observed increased activation rate and decreased threshold period required for optimum stimulation of RB cells. These results corroborate well with microarray gene expression data. This study provides novel insights into coordination between the molecules of innate and adaptive immunity in activating B cells, in a BCR independent manner. This strategy can be exploited to design vaccines to bolster B cell activation and antigen presenting efficiency, leading to faster and better immune response.

  7. Protective Antigen-Specific Memory B Cells Persist Years after Anthrax Vaccination and Correlate with Humoral Immunity

    Directory of Open Access Journals (Sweden)

    Lori Garman

    2014-08-01

    Full Text Available Anthrax Vaccine Adsorbed (AVA generates short-lived protective antigen (PA specific IgG that correlates with in vitro toxin neutralization and protection from Bacillus anthracis challenge. Animal studies suggest that when PA-specific IgG has waned, survival after spore challenge correlates with an activation of PA-specific memory B cells. Here, we characterize the quantity and the longevity of AVA-induced memory B cell responses in humans. Peripheral blood mononuclear cells (PBMCs from individuals vaccinated ≥3 times with AVA (n = 50 were collected early (3–6 months, n = 27 or late after their last vaccination (2–5 years, n = 23, pan-stimulated, and assayed by ELISPOT for total and PA-specific memory B cells differentiated into antibody secreting cells (ASCs. PA-specific ASC percentages ranged from 0.02% to 6.25% (median: 1.57% and did not differ between early and late post-vaccination individuals. PA-specific ASC percentages correlated with plasma PA-specific IgG (r = 0.42, p = 0.03 and toxin neutralization (r = 0.52, p = 0.003 early post vaccination. PA-specific ASC percentages correlated with supernatant anti-PA both early (r = 0.60, p = 0.001 and late post vaccination (r = 0.71, p < 0.0001. These data suggest PA-specific memory B cell responses are long-lived and can be estimated after recent vaccination by the magnitude and neutralization capacity of the humoral response.

  8. CD80 and CD86 Differentially Regulate Mechanical Interactions of T-Cells with Antigen-Presenting Dendritic Cells and B-Cells

    OpenAIRE

    Tong Seng Lim; James Kang Hao Goh; Alessandra Mortellaro; Chwee Teck Lim; Hämmerling, Günter J.; Paola Ricciardi-Castagnoli

    2012-01-01

    Functional T-cell responses are initiated by physical interactions between T-cells and antigen-presenting cells (APCs), including dendritic cells (DCs) and B-cells. T-cells are activated more effectively by DCs than by B-cells, but little is known about the key molecular mechanisms that underpin the particular potency of DC in triggering T-cell responses. To better understand the influence of physical intercellular interactions on APC efficacy in activating T-cells, we used single cell force ...

  9. The cell biology of T-dependent B cell activation

    DEFF Research Database (Denmark)

    Owens, T; Zeine, R

    1989-01-01

    The requirement that CD4+ helper T cells recognize antigen in association with class II Major Histocompatibility Complex (MHC) encoded molecules constrains T cells to activation through intercellular interaction. The cell biology of the interactions between CD4+ T cells and antigen-presenting cells...

  10. Virus-Like Display of a Neo-Self Antigen Reverses B Cell Anergy in a B Cell Receptor Transgenic Mouse Model1

    Science.gov (United States)

    Chackerian, Bryce; Durfee, Marisa R.; Schiller, John T.

    2012-01-01

    The ability to distinguish between self and foreign Ags is a central feature of immune recognition. For B cells, however, immune tolerance is not absolute, and factors that include Ag valency, the availability of T help, and polyclonal B cell stimuli can influence the induction of autoantibody responses. Here, we evaluated whether multivalent virus-like particle (VLP)-based immunogens could induce autoantibody responses in well-characterized transgenic (Tg) mice that express a soluble form of hen egg lysozyme (HEL) and in which B cell tolerance to HEL is maintained by anergy. Immunization with multivalent VLP-arrayed HEL, but not a trivalent form of HEL, induced high-titer Ab responses against HEL in both soluble HEL Tg mice and double Tg mice that also express a monoclonal HEL-specific BCR. Induction of autoantibodies against HEL was not dependent on coadministration of strong adjuvants, such as CFA. In contrast to previous data showing the T-independent induction of Abs to foreign epitopes on VLPs, the ability of HEL-conjugated VLPs to induce anti-HEL Abs in tolerant mice was dependent on the presence of CD4+ Th cells, and could be enhanced by the presence of pre-existing cognate T cells. In in vitro studies, VLP-conjugated HEL was more potent than trivalent HEL in up-regulating surface activation markers on purified anergic B cells. Moreover, immunization with VLP-HEL reversed B cell anergy in vivo in an adoptive transfer model. Thus, Ag multivalency and T help cooperate to reverse B cell anergy, a major mechanism of B cell tolerance. PMID:18424700

  11. The phosphoinositide 3-kinase signaling pathway in normal and malignant B cells: activation mechanisms, regulation and impact on cellular functions.

    Science.gov (United States)

    Pauls, Samantha D; Lafarge, Sandrine T; Landego, Ivan; Zhang, Tingting; Marshall, Aaron J

    2012-01-01

    The phosphoinositide 3-kinase (PI3K) pathway is a central signal transduction axis controlling normal B cell homeostasis and activation in humoral immunity. The p110δ PI3K catalytic subunit has emerged as a critical mediator of multiple B cell functions. The activity of this pathway is regulated at multiple levels, with inositol phosphatases PTEN and SHIP both playing critical roles. When deregulated, the PI3K pathway can contribute to B cell malignancies and autoantibody production. This review summarizes current knowledge on key mechanisms that activate and regulate the PI3K pathway and influence normal B cell functional responses including the development of B cell subsets, antigen presentation, immunoglobulin isotype switch, germinal center responses, and maintenance of B cell anergy. We also discuss PI3K pathway alterations reported in select B cell malignancies and highlight studies indicating the functional significance of this pathway in malignant B cell survival and growth within tissue microenvironments. Finally, we comment on early clinical trial results, which support PI3K inhibition as a promising treatment of chronic lymphocytic leukemia. PMID:22908014

  12. Refining the Neuberger Model: uracil processing by activated B cells

    OpenAIRE

    Maul, Robert W.; Gearhart, Patricia J.

    2014-01-01

    During the immune response, B cells undergo a programmed mutagenic cascade to promote increased affinity and expanded antibody function. The two processes, somatic hypermutation (SHM) and class switch recombination (CSR), are initiated by the protein activation-induced deaminase (AID), which converts cytosine to uracil in the immunoglobulin loci. The presence of uracil in DNA promotes DNA mutagenesis though a subset of DNA repair proteins. Two distinct mechanisms have been proposed to control...

  13. The role of antigen in the development of B-cell chronic lymphocytic leukemia

    OpenAIRE

    Hoogeboom, R.

    2013-01-01

    These studies strongly suggest that MALT-lymphomas and M-CLL in majority are highly selected for single extrinsic antigens and that these antigens can be both self-antigens and exo-antigens. Our finding that primary CLL cells are responsive to stimulation with their cognate antigen suggests that antigen-dependent BCR signaling may drive CLL expansion in vivo.

  14. The Correlation between the Virus- and Brain Antigen-Specific B Cell Response in the Blood of Patients with Multiple Sclerosis

    Directory of Open Access Journals (Sweden)

    Marie Wunsch

    2016-04-01

    Full Text Available There is a largely divergent body of literature regarding the relationship between Epstein-Barr virus (EBV infection and brain inflammation in multiple sclerosis (MS. Here, we tested MS patients during relapse (n = 11 and in remission (n = 19 in addition to n = 22 healthy controls to study the correlation between the EBV- and brain-specific B cell response in the blood by enzyme-linked immunospot (ELISPOT and enzyme-linked immunosorbent assay (ELISA. Cytomegalovirus (CMV was used as a control antigen tested in n = 16 MS patients during relapse and in n = 35 patients in remission. Over the course of the study, n = 16 patients were untreated, while n = 33 patients received immunomodulatory therapy. The data show that there was a moderate correlation between the frequencies of EBV- and brain-reactive B cells in MS patients in remission. In addition we could detect a correlation between the B cell response to EBV and disease activity. There was no evidence of an EBV reactivation. Interestingly, there was also a correlation between the frequencies of CMV- and brain-specific B cells in MS patients experiencing an acute relapse and an elevated B cell response to CMV was associated with higher disease activity. The trend remained when excluding seronegative subjects but was non-significant. These data underline that viral infections might impact the immunopathology of MS, but the exact link between the two entities remains subject of controversy.

  15. A Computational Study of the Effects of Syk Activity on B Cell Receptor Signaling Dynamics

    Directory of Open Access Journals (Sweden)

    Reginald L. McGee

    2015-02-01

    Full Text Available The kinase Syk is intricately involved in early signaling events in B cells and isrequired for proper response when antigens bind to B cell receptors (BCRs. Experimentsusing an analog-sensitive version of Syk (Syk-AQL have better elucidated its role, buthave not completely characterized its behavior. We present a computational model for BCRsignaling, using dynamical systems, which incorporates both wild-type Syk and Syk-AQL.Following the use of sensitivity analysis to identify significant reaction parameters, we screenfor parameter vectors that produced graded responses to BCR stimulation as is observedexperimentally. We demonstrate qualitative agreement between the model and dose responsedata for both mutant and wild-type kinases. Analysis of our model suggests that the level of NF-KB activation, which is reduced in Syk-AQL cells relative to wild-type, is more sensitiveto small reductions in kinase activity than Erkp activation, which is essentially unchanged.Since this profile of high Erkp and reduced NF-KB is consistent with anergy, this implies thatanergy is particularly sensitive to small changes in catalytic activity. Also, under a range offorward and reverse ligand binding rates, our model of Erkp and NF-KB activation displaysa dependence on a power law affinity: the ratio of the forward rate to a non-unit power of thereverse rate. This dependence implies that B cells may respond to certain details of bindingand unbinding rates for ligands rather than simple affinity alone.

  16. IgE production after antigen-specific and cognate activation of HLA-DPw4-restricted T-cell clones, by 78% of randomly selected B-cell donors

    NARCIS (Netherlands)

    Baselmans, PJ; Pollabauer, EM; van Reijsen, FC; Heystek, HC; Hren, A; Stumptner, P; Tilanus, MGJ; Vooijs, WC; Mudde, GC

    2000-01-01

    The frequency of expression of the MHC class II antigen, HLA-DPw4, in the caucasoid population is approximately 78%, and is unmatched by phenotypic frequencies of other HLA class II molecules. Here we describe three human Der-P1-specific T-cell clones (TCC), restricted by the HLA-DPw4-variant HLA-DP

  17. Synthetic Peptide Ligands of the Antigen Binding Receptor Induce Programmed Cell Death in a Human B-Cell Lymphoma

    Science.gov (United States)

    Renschler, Markus F.; Bhatt, Ramesh R.; Dower, William J.; Levy, Ronald

    1994-04-01

    Peptide ligands for the antigen binding site of the surface immunoglobulin receptor of a human B-cell lymphoma cell line were identified with the use of filamentous phage libraries displaying random 8- and 12-amino acid peptides. Corresponding synthetic peptides bound specifically to the antigen binding site of this immunoglobulin receptor and blocked the binding of an anti-idiotype antibody. The ligands, when conjugated to form dimers or tetramers, induced cell death by apoptosis in vitro with an IC50 between 40 and 200 nM. This effect was associated with specific stimulation of intracellular protein tyrosine phosphorylation.

  18. Affinity isolation of antigen-specific circulating B cells for generation of phage display-derived human monoclonal antibodies

    DEFF Research Database (Denmark)

    Ditzel, Henrik

    2009-01-01

    A method is described for affinity isolation of antigen-specific circulating B cells of interest for subsequent generation of immune antibody phage display libraries. This approach should overcome the problem of low yields of monoclonal antibodies of interest in the libraries generated from...... the frequency of antibody phage particles of interest in the library and allow for efficient isolation monoclonal antibodies with the predefined specificity....

  19. Semiallogenic fusions of MSI+ tumor cells and activated B cells induce MSI-specific T cell responses

    Directory of Open Access Journals (Sweden)

    Klier Ulrike

    2011-09-01

    Full Text Available Abstract Background Various strategies have been developed to transfer tumor-specific antigens into antigen presenting cells in order to induce cytotoxic T cell responses against tumor cells. One approach uses cellular vaccines based on fusions of autologous antigen presenting cells and allogeneic tumor cells. The fusion cells combine antigenicity of the tumor cell with optimal immunostimulatory capacity of the antigen presenting cells. Microsatellite instability caused by mutational inactivation of DNA mismatch repair genes results in translational frameshifts when affecting coding regions. It has been shown by us and others that these mutant proteins lead to the presentation of immunogenic frameshift peptides that are - in principle - recognized by a multiplicity of effector T cells. Methods We chose microsatellite instability-induced frameshift antigens as ideal to test for induction of tumor specific T cell responses by semiallogenic fusions of microsatellite instable carcinoma cells with CD40-activated B cells. Two fusion clones of HCT116 with activated B cells were selected for stimulation of T cells autologous to the B cell fusion partner. Outgrowing T cells were phenotyped and tested in functional assays. Results The fusion clones expressed frameshift antigens as well as high amounts of MHC and costimulatory molecules. Autologous T cells stimulated with these fusions were predominantly CD4+, activated, and reacted specifically against the fusion clones and also against the tumor cell fusion partner. Interestingly, a response toward 6 frameshift-derived peptides (of 14 tested could be observed. Conclusion Cellular fusions of MSI+ carcinoma cells and activated B cells combine the antigen-presenting capacity of the B cell with the antigenic repertoire of the carcinoma cell. They present frameshift-derived peptides and can induce specific and fully functional T cells recognizing not only fusion cells but also the carcinoma cells. These

  20. Semiallogenic fusions of MSI+ tumor cells and activated B cells induce MSI-specific T cell responses

    International Nuclear Information System (INIS)

    Various strategies have been developed to transfer tumor-specific antigens into antigen presenting cells in order to induce cytotoxic T cell responses against tumor cells. One approach uses cellular vaccines based on fusions of autologous antigen presenting cells and allogeneic tumor cells. The fusion cells combine antigenicity of the tumor cell with optimal immunostimulatory capacity of the antigen presenting cells. Microsatellite instability caused by mutational inactivation of DNA mismatch repair genes results in translational frameshifts when affecting coding regions. It has been shown by us and others that these mutant proteins lead to the presentation of immunogenic frameshift peptides that are - in principle - recognized by a multiplicity of effector T cells. We chose microsatellite instability-induced frameshift antigens as ideal to test for induction of tumor specific T cell responses by semiallogenic fusions of microsatellite instable carcinoma cells with CD40-activated B cells. Two fusion clones of HCT116 with activated B cells were selected for stimulation of T cells autologous to the B cell fusion partner. Outgrowing T cells were phenotyped and tested in functional assays. The fusion clones expressed frameshift antigens as well as high amounts of MHC and costimulatory molecules. Autologous T cells stimulated with these fusions were predominantly CD4+, activated, and reacted specifically against the fusion clones and also against the tumor cell fusion partner. Interestingly, a response toward 6 frameshift-derived peptides (of 14 tested) could be observed. Cellular fusions of MSI+ carcinoma cells and activated B cells combine the antigen-presenting capacity of the B cell with the antigenic repertoire of the carcinoma cell. They present frameshift-derived peptides and can induce specific and fully functional T cells recognizing not only fusion cells but also the carcinoma cells. These hybrid cells may have great potential for cellular immunotherapy and

  1. CD80 and CD86 differentially regulate mechanical interactions of T-cells with antigen-presenting dendritic cells and B-cells.

    Science.gov (United States)

    Lim, Tong Seng; Goh, James Kang Hao; Mortellaro, Alessandra; Lim, Chwee Teck; Hämmerling, Günter J; Ricciardi-Castagnoli, Paola

    2012-01-01

    Functional T-cell responses are initiated by physical interactions between T-cells and antigen-presenting cells (APCs), including dendritic cells (DCs) and B-cells. T-cells are activated more effectively by DCs than by B-cells, but little is known about the key molecular mechanisms that underpin the particular potency of DC in triggering T-cell responses. To better understand the influence of physical intercellular interactions on APC efficacy in activating T-cells, we used single cell force spectroscopy to characterize and compare the mechanical forces of interactions between DC:T-cells and B:T-cells. Following antigen stimulation, intercellular interactions of DC:T-cell conjugates were stronger than B:T-cell interactions. DCs induced higher levels of T-cell calcium mobilization and production of IL-2 and IFNγ than were elicited by B-cells, thus suggesting that tight intercellular contacts are important in providing mechanically stable environment to initiate T-cell activation. Blocking antibodies targeting surface co-stimulatory molecules CD80 or CD86 weakened intercellular interactions and dampen T-cell activation, highlighting the amplificatory roles of CD80/86 in regulating APC:T-cell interactions and T-cell functional activation. The variable strength of mechanical forces between DC:T-cells and B:T-cell interactions were not solely dependent on differential APC expression of CD80/86, since DCs were superior to B-cells in promoting strong interactions with T-cells even when CD80 and CD86 were inhibited. These data provide mechanical insights into the effects of co-stimulatory molecules in regulating APC:T-cell interactions. PMID:23024807

  2. CD80 and CD86 differentially regulate mechanical interactions of T-cells with antigen-presenting dendritic cells and B-cells.

    Directory of Open Access Journals (Sweden)

    Tong Seng Lim

    Full Text Available Functional T-cell responses are initiated by physical interactions between T-cells and antigen-presenting cells (APCs, including dendritic cells (DCs and B-cells. T-cells are activated more effectively by DCs than by B-cells, but little is known about the key molecular mechanisms that underpin the particular potency of DC in triggering T-cell responses. To better understand the influence of physical intercellular interactions on APC efficacy in activating T-cells, we used single cell force spectroscopy to characterize and compare the mechanical forces of interactions between DC:T-cells and B:T-cells. Following antigen stimulation, intercellular interactions of DC:T-cell conjugates were stronger than B:T-cell interactions. DCs induced higher levels of T-cell calcium mobilization and production of IL-2 and IFNγ than were elicited by B-cells, thus suggesting that tight intercellular contacts are important in providing mechanically stable environment to initiate T-cell activation. Blocking antibodies targeting surface co-stimulatory molecules CD80 or CD86 weakened intercellular interactions and dampen T-cell activation, highlighting the amplificatory roles of CD80/86 in regulating APC:T-cell interactions and T-cell functional activation. The variable strength of mechanical forces between DC:T-cells and B:T-cell interactions were not solely dependent on differential APC expression of CD80/86, since DCs were superior to B-cells in promoting strong interactions with T-cells even when CD80 and CD86 were inhibited. These data provide mechanical insights into the effects of co-stimulatory molecules in regulating APC:T-cell interactions.

  3. Differential Effects of Tacrolimus versus Sirolimus on the Proliferation, Activation and Differentiation of Human B Cells

    OpenAIRE

    Traitanon, Opas; Mathew, James M.; La Monica, Giovanna; Xu, Luting; Mas, Valeria; Gallon, Lorenzo

    2015-01-01

    The direct effect of immunosuppressive drugs calcineurin inhibitor (Tacrolimus, TAC) and mTOR inhibitor (Sirolimus, SRL) on B cell activation, differentiation and proliferation is not well documented. Purified human B cells from healthy volunteers were stimulated through the B Cell Receptor with Anti-IgM + anti-CD40 + IL21 in the absence / presence of TAC or SRL. A variety of parameters of B cell activity including activation, differentiation, cytokine productions and proliferation were monit...

  4. Impairment of pneumococcal antigen specific isotype-switched Igg memory B-cell immunity in HIV infected Malawian adults.

    Directory of Open Access Journals (Sweden)

    Oluwadamilola H Iwajomo

    Full Text Available Pneumococcal disease is associated with a particularly high morbidity and mortality amongst adults in HIV endemic countries. Our previous findings implicating a B-cell defect in HIV-infected children from the same population led us to comprehensively characterize B-cell subsets in minimally symptomatic HIV-infected Malawian adults and investigate the isotype-switched IgG memory B-cell immune response to the pneumococcus. We show that similar to vertically acquired HIV-infected Malawian children, horizontally acquired HIV infection in these adults is associated with IgM memory B-cell (CD19(+ CD27(+ IgM(+ IgD(+ depletion, B-cell activation and impairment of specific IgG B-cell memory to a range of pneumococcal proteins. Our data suggest that HIV infection affects both T-cell independent and T-cell dependent B-cell maturation, potentially leading to impairment of humoral responses to extracellular pathogens such as the pneumococcus, and thus leaving this population susceptible to invasive disease.

  5. A Recombinant Bispecific CD20×CD95 Antibody With Superior Activity Against Normal and Malignant B-cells.

    Science.gov (United States)

    Nalivaiko, Kristina; Hofmann, Martin; Kober, Karina; Teichweyde, Nadine; Krammer, Peter H; Rammensee, Hans-Georg; Grosse-Hovest, Ludger; Jung, Gundram

    2016-02-01

    Monoclonal antibodies directed to the B-cell-specific CD20-antigen are successfully used for the treatment of lymphomas and autoimmune diseases. Here, we compare the anti-B-cell activity of three different antibodies directed to CD20: (i) a chimeric, monospecific antibody, (ii) an Fc-optimized variant thereof, and (iii) a bispecific CD20×CD95-antibody in a newly developed recombinant format, termed Fabsc. The bispecific antibody specifically triggers the CD95 death receptor on malignant, as well as activated, normal B-cells. We found that the capability of this antibody to suppress the growth of malignant B-cells in vitro and in vivo and to specifically deplete normal, activated B-cells from peripheral blood mononuclear cell (PBMC) cultures was superior to that of the Fc-optimized monospecific antibody. This antibody in turn was more effective than its nonoptimized variant. Moreover, the bispecific antibody was the only reagent capable of significantly suppressing antibody production in vitro. Our findings imply that the bispecific CD20×CD95-antibody might become a new, prototypical reagent for the treatment of B-cell-mediated autoimmune disease. PMID:26581163

  6. Antibody Reactivity of B Cells in Lupus Patients with Increased Disease Activity and ARID3a Expression

    Directory of Open Access Journals (Sweden)

    Julie M. Ward

    2015-11-01

    Full Text Available Earlier studies showed that the DNA-binding protein, Bright/ARID3a bound to a subset of human and mouse immunoglobulin heavy chain promoters where it enhanced expression. Indeed, mice with transgenic expression of ARID3a in all B lymphocytes have expanded MZ B cells and produce anti-nuclear antibodies (ANAs. Consistent with our findings in mice, we observed that human systemic lupus erythematosus (SLE patients had expanded numbers of peripheral blood ARID3a+ B cells that were associated with increased disease activity (p = 0.0038. We hypothesized that ARID3a+ naïve B cells would eventually produce autoantibodies, explaining associations between ARID3a expression and disease activity in lupus. Unlike healthy controls, ARID3a was expressed in the naïve B cell population in SLE patients, and we hypothesized that these might represent expansions of autoreactive cells. Therefore, monoclonal antibodies were generated from single-sorted naïve B cells derived from patients with normal (ARID3aN and high (ARID3aH numbers of ARID3a+ B cells. We found that ARID3a expression did not correlate with autoantibody expression. Furthermore, measures of antigen specificities of autoreactive antibodies did not reveal skewing toward particular proteins. These data suggest that the association of increased disease activity in SLE with numbers of ARID3a+ B lymphocytes may be mediated by an antibody-independent mechanism.

  7. Evidence for a new segregant series of B cell antigens that are encoded in the HLA-D region and that stimulate secondary allogenic proliferative and cytotoxic responses

    OpenAIRE

    1980-01-01

    Five new histocompatibility antigens, designated secondary B cell or (SB) antigens, have been identified by secondary allogeneic proliferative and cytotoxic responses. The reagents used to define the SB antigents are lymphocytes primed between donors matched for all known HLA antigens. The SB antigens stimulate weak primary allogeneic proliferative responses (a mean relative response of 8%) but strong secondary proliferative responses. Strong secondary cell-mediated cytotoxicity is generated ...

  8. Switch-mediated activation and retargeting of CAR-T cells for B-cell malignancies.

    Science.gov (United States)

    Rodgers, David T; Mazagova, Magdalena; Hampton, Eric N; Cao, Yu; Ramadoss, Nitya S; Hardy, Ian R; Schulman, Andrew; Du, Juanjuan; Wang, Feng; Singer, Oded; Ma, Jennifer; Nunez, Vanessa; Shen, Jiayin; Woods, Ashley K; Wright, Timothy M; Schultz, Peter G; Kim, Chan Hyuk; Young, Travis S

    2016-01-26

    Chimeric antigen receptor T (CAR-T) cell therapy has produced impressive results in clinical trials for B-cell malignancies. However, safety concerns related to the inability to control CAR-T cells once infused into the patient remain a significant challenge. Here we report the engineering of recombinant antibody-based bifunctional switches that consist of a tumor antigen-specific Fab molecule engrafted with a peptide neo-epitope, which is bound exclusively by a peptide-specific switchable CAR-T cell (sCAR-T). The switch redirects the activity of the bio-orthogonal sCAR-T cells through the selective formation of immunological synapses, in which the sCAR-T cell, switch, and target cell interact in a structurally defined and temporally controlled manner. Optimized switches specific for CD19 controlled the activity, tissue-homing, cytokine release, and phenotype of sCAR-T cells in a dose-titratable manner in a Nalm-6 xenograft rodent model of B-cell leukemia. The sCAR-T-cell dosing regimen could be tuned to provide efficacy comparable to the corresponding conventional CART-19, but with lower cytokine levels, thereby offering a method of mitigating cytokine release syndrome in clinical translation. Furthermore, we demonstrate that this methodology is readily adaptable to targeting CD20 on cancer cells using the same sCAR-T cell, suggesting that this approach may be broadly applicable to heterogeneous and resistant tumor populations, as well as other liquid and solid tumor antigens. PMID:26759369

  9. Aging affects B-cell antigen receptor repertoire diversity in primary and secondary lymphoid tissues.

    Science.gov (United States)

    Tabibian-Keissar, Hilla; Hazanov, Lena; Schiby, Ginette; Rosenthal, Noemie; Rakovsky, Aviya; Michaeli, Miri; Shahaf, Gitit Lavy; Pickman, Yishai; Rosenblatt, Kinneret; Melamed, Doron; Dunn-Walters, Deborah; Mehr, Ramit; Barshack, Iris

    2016-02-01

    The elderly immune system is characterized by reduced responses to infections and vaccines, and an increase in the incidence of autoimmune diseases and cancer. Age-related deficits in the immune system may be caused by peripheral homeostatic pressures that limit bone marrow B-cell production or migration to the peripheral lymphoid tissues. Studies of peripheral blood B-cell receptor spectratypes have shown that those of the elderly are characterized by reduced diversity, which is correlated with poor health status. In the present study, we performed for the first time high-throughput sequencing of immunoglobulin genes from archived biopsy samples of primary and secondary lymphoid tissues in old (74 ± 7 years old, range 61-89) versus young (24 ± 5 years old, range 18-45) individuals, analyzed repertoire diversities and compared these to results in peripheral blood. We found reduced repertoire diversity in peripheral blood and lymph node repertoires from old people, while in the old spleen samples the diversity was larger than in the young. There were no differences in somatic hypermutation characteristics between age groups. These results support the hypothesis that age-related immune frailty stems from altered B-cell homeostasis leading to narrower memory B-cell repertoires, rather than changes in somatic hypermutation mechanisms. PMID:26614343

  10. B cell depletion with rituximab in patients with rheumatoid arthritis: Multiplex bead array reveals the kinetics of IgG and IgA antibodies to citrullinated antigens.

    Science.gov (United States)

    Cambridge, Geraldine; Leandro, Maria J; Lahey, Lauren J; Fairhead, Thomas; Robinson, William H; Sokolove, Jeremy

    2016-06-01

    The serology of patients with Rheumatoid arthritis (RA) is characterized by persistently raised levels of autoantibodies: Rheumatoid Factors (RhF) against Fc of IgG, and to citrullinated (Cit) protein/peptide sequences: ACPA, recognizing multiple Cit-sequences. B cell depletion therapy based on rituximab delivers good clinical responses in RA patients, particularly in the seropositive group, with responses sometimes lasting beyond the phase of B cell reconstitution. In general, ACPA levels fall following rituximab, but fluctuations with respect to predicting relapse have proved disappointing. In order to identify possible immunodominant specificities within either IgG- or IgA-ACPA we used a Multiplex bead-based array consisting of 30 Cit-peptides/proteins and 22 corresponding native sequences. The kinetics of the serum ACPA response to individual specificities was measured at key points (Baseline, B cell depletion phase, Relapse) within an initial cycle of rituximab therapy in 16 consecutive patients with severe, active RA. All had achieved significant decreases in Disease Activity Scores-28 and maintained B cell depletion in the peripheral blood (<5 CD19+cells/μl) for at least 3 months. At Baseline, mean fluorescence intensity shown by individual IgG- and IgA-ACPA were strongly correlated (R(2) = 0.75; p < 0.0001) but IgA-ACPA were approximately 10-fold lower. Data were Z-normalised in order to compare serial results and antibody classes. At Baseline, a total of 68 IgG- and 51 IgA-ACPA had Z-scores ≥ 1 (above population mean) were identified, with at least one Cit-antigen identified in each serum. ACPA to individual specificities subsequently fluctuated with 3 different patterns. Most 51/68 (75%) IgG- and 48/51 IgA-ACPA (94%) fell between Baseline and Depletion, of which 57% IgG- and 65% IgA-ACPA rebounded pre-Relapse. Interestingly, 17/68 IgG-ACPA (25%) and some IgA-ACPA (3/51; 6%) transiently increased from Baseline, subsequently falling pre

  11. Induction of polyclonal B cell activation and differentiation by the AIDS retrovirus (HTLV-III/LAV)

    International Nuclear Information System (INIS)

    The immune systems of individuals infected with HTLV-III/LAV are characterized by a profound defect in cellular immunity together with paradoxical polyclonal B cell activation. The present study examined the direct effects of HTLV-III/LAV on B lymphocytes. Peripheral blood B cells from healthy donors were incubated with a variety of HTLV-III/LAV isolates for 1 h and 3H-thymidine incorporation was measured at multiple time points. Responses ranged from 9000-28,000 cpm and peaked on day 4. This B cell activation was not enhanced by the addition of interleukin-2 to culture, was not synergistic with Staphylococcus aureus Cowan I, was not modulated by the addition of T lymphocytes to culture, and was not associated with B cell transformation. Supernatant Ig could first be detected in virus-activated cultures at day 4, plateaued by day 8, and yielded a mean of 12,500 ng IgG+IgM/ml/50,000 B cells. Thus, HTLV-III/LAV is a potent T cell independent B cell mitogen capable of inducing B cell activation, proliferation, and differentiation comparable in magnitude to that of the most potent B cell activators. This biological property of HTLV-III/LAV may help explain the profound polyclonal B cell activation observed in patients with AIDS and may provide investigators with another probe for investigating the mechanisms of B cell activation

  12. Enhanced T cell responses to antigenic peptides targeted to B cell surface Ig, Ia, or class I molecules

    OpenAIRE

    1988-01-01

    The helper T cell recognition of soluble globular protein antigens requires that the proteins be processed by an APC, releasing a peptide that is transported to and held on the APC surface where it is recognized by the specific T cell in conjunction with Ia. When cellular processing functions are blocked, APC lose their ability to present native antigens while retaining the capacity to activate T cells when provided with a cognate peptide fragment that contains the T cell antigenic determinan...

  13. T-dependent B-cell activation is signalled by an early increase in potassium influx

    DEFF Research Database (Denmark)

    Owens, T; Kaplan, J G

    1982-01-01

    (previously demonstrated when B and T lymphocytes were separately stimulated) also occurs when B cells are stimulated through cooperation with mitogen-activated T cells, and is also detectable early in culture. T-dependent activation of B cells is therefore detectable considerably earlier than by conventional...

  14. Simultaneous Detection of Antigen-Specific IgG- and IgM-Secreting Cells with a B Cell Fluorospot Assay

    OpenAIRE

    M Anthony Moody; Mattia Bonsignori

    2012-01-01

    The traditional enzyme-linked immunospot (ELISpot) assay is the gold standard for the enumeration of antigen-specific B cells. Since B cell availability from biological samples is often limited, either because of sample size/volume or the need of performing multiple analyses on the same sample, the implementation of ELISpot assay formats that allow the simultaneous detection of multiple antibody types is desirable. While dual-color ELISpot assays have been described, technical complexities ha...

  15. Evolutionary and functional relationships of B cells from fish and mammals: insights into their novel roles in phagocytosis and presentation of particulate antigen.

    Science.gov (United States)

    Sunyer, J Oriol

    2012-06-01

    been shown that phagocytic B-1 B cells have a potent ability to present particulate antigen to CD4+ T cells. Thus, studies carried out originally on fish B cells have lead to the discovery of new innate and adaptive roles of B cells in mammals. This review will concentrate on the evolutionary and functional relationships of fish and mammalian B cells, focusing mainly on the newly discovered roles of these cells in phagocytosis, intracellular killing and presentation of particulate antigen. PMID:22394174

  16. Innate B cells: oxymoron or validated concept?

    OpenAIRE

    Ware, Carl F.; Chris Benedict

    2012-01-01

    B lymphocytes promote the initial innate interferon response to viral pathogens without the need for antigen receptor activation. B cell dependent IFN production requires the cytokine, lymphotoxin-β. The LTβ pathway is well known to regulate lymphoid organogenesis and homeostasis by differentiating stromal cells and macrophages. However, in response to viral pathogens these same B cell-regulated populations rapidly produce type 1 interferons. Thus, B cells act as innate effector cells via LTβ...

  17. Exosomes released from Mycoplasma infected tumor cells activate inhibitory B cells.

    Directory of Open Access Journals (Sweden)

    Chenjie Yang

    Full Text Available Mycoplasmas cause numerous human diseases and are common opportunistic pathogens in cancer patients and immunocompromised individuals. Mycoplasma infection elicits various host immune responses. Here we demonstrate that mycoplasma-infected tumor cells release exosomes (myco+ exosomes that specifically activate splenic B cells and induce splenocytes cytokine production. Induction of cytokines, including the proinflammatory IFN-γ and the anti-inflammatory IL-10, was largely dependent on the presence of B cells. B cells were the major IL-10 producers. In splenocytes from B cell deficient μMT mice, induction of IFN-γ+ T cells by myco+ exosomes was greatly increased compared with wild type splenocytes. In addition, anti-CD3-stimulated T cell proliferation was greatly inhibited in the presence of myco+ exosome-treated B cells. Also, anti-CD3-stimulated T cell signaling was impaired by myco+ exosome treatment. Proteomic analysis identified mycoplasma proteins in exosomes that potentially contribute to the effects. Our results demonstrate that mycoplasma-infected tumor cells release exosomes carrying mycoplasma components that preferentially activate B cells, which in turn, are able to inhibit T cell activity. These results suggest that mycoplasmas infecting tumor cells can exploit the exosome pathway to disseminate their own components and modulate the activity of immune cells, in particular, activate B cells with inhibitory activity.

  18. Differential Effects of Tacrolimus versus Sirolimus on the Proliferation, Activation and Differentiation of Human B Cells.

    Directory of Open Access Journals (Sweden)

    Opas Traitanon

    Full Text Available The direct effect of immunosuppressive drugs calcineurin inhibitor (Tacrolimus, TAC and mTOR inhibitor (Sirolimus, SRL on B cell activation, differentiation and proliferation is not well documented. Purified human B cells from healthy volunteers were stimulated through the B Cell Receptor with Anti-IgM + anti-CD40 + IL21 in the absence / presence of TAC or SRL. A variety of parameters of B cell activity including activation, differentiation, cytokine productions and proliferation were monitored by flow cytometry. SRL at clinically relevant concentrations (6 ng/ml profoundly inhibited CD19(+ B cell proliferation compared to controls whereas TAC at similar concentrations had a minimal effect. CD27(+ memory B cells were affected more by SRL than naïve CD27- B cells. SRL effectively blocked B cell differentiation into plasma cells (CD19(+CD138(+ and Blimp1(+/Pax5(low cells even at low dose (2 ng/ml, and totally eliminated them at 6 ng/ml. SRL decreased absolute B cell counts, but the residual responding cells acquired an activated phenotype (CD25(+/CD69(+ and increased the expression of HLA-DR. SRL-treated stimulated B cells on a per cell basis were able to enhance the proliferation of allogeneic CD4(+CD25(- T cells and induce a shift toward the Th1 phenotype. Thus, SRL and TAC have different effects on B lymphocytes. These data may provide insights into the clinical use of these two agents in recipients of solid organ transplants.

  19. Cbl Suppresses B Cell Receptor–Mediated Phospholipase C (Plc)-γ2 Activation by Regulating B Cell Linker Protein–Plc-γ2 Binding

    OpenAIRE

    Yasuda, Tomoharu; Maeda, Akito; Kurosaki, Mari; Tezuka, Tohru; Hironaka, Katsunori; Yamamoto, Tadashi; Kurosaki, Tomohiro

    2000-01-01

    Accumulating evidence indicates that the Cbl protein plays a negative role in immune receptor signaling; however, the mode of Cbl action in B cell receptor (BCR) signaling still remains unclear. DT40 B cells deficient in Cbl showed enhanced BCR-mediated phospholipase C (PLC)-γ2 activation, thereby leading to increased apoptosis. A possible explanation for the involvement of Cbl in PLC-γ2 activation was provided by findings that Cbl interacts via its Src homology 2 (SH2) domain with B cell lin...

  20. Human leukocyte antigen-DO regulates surface presentation of human leukocyte antigen class II-restricted antigens on B cell malignancies

    NARCIS (Netherlands)

    Kremer, A.N.; Meijden, E.D. van der; Honders, M.W.; Pont, M.J.; Goeman, J.J.; Falkenburg, J.H.F.; Griffioen, M.

    2014-01-01

    Hematological malignancies often express surface HLA class II, making them attractive targets for CD4+ T cell therapy. We previously demonstrated that HLA class II ligands can be divided into DM-resistant and DM-sensitive antigens. In contrast to presentation of DM-resistant antigens, presentation o

  1. Antigen presentation by non-immune B-cell hybridoma clones: presentation of synthetic antigenic sites reveals clones that exhibit no specificity and clones that present only one epitope

    Science.gov (United States)

    Cohly, H. H.; Morrison, D. R.; Atassi, M. Z.

    1989-01-01

    Recently, we reported the preparation and antigen-presenting properties of hybridoma B-cell clones obtained after fusing non-secreting, non-antigen presenting Balb/c 653-myeloma cells with non-immune SJL spleen cells. It was found that antigen presentation at the clonal level can be specific or non-specific, depending on the particular B-cell clone. In the present work, one specific and one general presenter B-cell clones were tested for their epitope presentation ability to SJL T-cells that were specific to lysozyme or myoglobin. B-cell clone A1G12, a general presenter which presented both lysozyme and myoglobin to their respective T-cell lines, was found to present all five myoglobin epitopes while clone A1L16, a lysozyme specific presenter presented only one of the three epitopes of lysozyme. The latter reveals a hitherto unknown submolecular specificity (to a given epitope within a protein) for antigen presenting cells at the clonal level. Therefore, the specificity of T-cell recognition does not only derive from the T-cell but may also be dependent on the epitope specificity of the antigen-presenting B-cell.

  2. B-Cell Activation and Non-Hodgkin Lymphoma Risk in an HIV Positive Population

    OpenAIRE

    Chang, Po-Yin

    2013-01-01

    Background: B-cell non-Hodgkin lymphoma (NHL) in HIV populations (AIDS-NHL) has become the leading cause of AIDS-defining cancers. Studies suggested that genetic or serum markers of B-cell activation are related to AIDS-NHL. However, associations between HIV viral load and AIDS-NHL risk have not been explicitly explored with consideration of B-cell activation markers. Furthermore, associations of hepatitis C virus (HCV) infection to AIDS-NHL risk are inconclusive. Methods: We used two nested ...

  3. Activation of human B cells by the agonist CD40 antibody CP-870,893 and augmentation with simultaneous toll-like receptor 9 stimulation

    Directory of Open Access Journals (Sweden)

    Rüter Jens

    2009-11-01

    Full Text Available Abstract Background CD40 activation of antigen presenting cells (APC such as dendritic cells (DC and B cells plays an important role in immunological licensing of T cell immunity. Agonist CD40 antibodies have been previously shown in murine models to activate APC and enhance tumor immunity; in humans, CD40-activated DC and B cells induce tumor-specific T cells in vitro. Although clinical translation of these findings for patients with cancer has been previously limited due to the lack of a suitable and available drug, promising clinical results are now emerging from phase I studies of the agonist CD40 monoclonal antibody CP-870,893. The most prominent pharmacodynamic effect of CP-870,893 infusion is peripheral B cell modulation, but direct evidence of CP-870,893-mediated B cell activation and the potential impact on T cell reactivity has not been reported, despite increasing evidence that B cells, like DC, regulate cellular immunity. Methods Purified total CD19+ B cells, CD19+ CD27+ memory, or CD19+ CD27neg subsets from peripheral blood were stimulated in vitro with CP-870,893, in the presence or absence of the toll like receptor 9 (TLR9 ligand CpG oligodeoxynucleotide (ODN. B cell surface molecule expression and cytokine secretion were evaluated using flow cytometry. Activated B cells were used as stimulators in mixed lymphocyte reactions to evaluate their ability to induce allogeneic T cell responses. Results Incubation with CP-870,893 activated B cells, including both memory and naïve B cells, as demonstrated by upregulation of CD86, CD70, CD40, and MHC class I and II. CP-870,893-activated B cells induced T cell proliferation and T cell secretion of effector cytokines including IFN-gamma and IL-2. These effects were increased by TLR9 co-stimulation via a CpG ODN identical in sequence to a well-studied clinical grade reagent. Conclusion The CD40 mAb CP-870,893 activates both memory and naïve B cells and triggers their T cell stimulatory

  4. Evolution of B-cell malignancy; Pre-B-cell leukemia resulting from MYC activation in a B-cell neoplasm with a rearranged BCL2 gene

    International Nuclear Information System (INIS)

    The authors have analyzed the molecular genetics of the breakpoints involved in the t(8;14) and t(14;18) translocations of an acute pre-B-cell leukemia from a patient with a history of follicular lymphoma. In this patient's leukemic cells, the breakpoint of the t(14;18) translocation occurred in the major breakpoint-cluster region of the BCL2 gene and became linked to the JH4 joining-region gene segment of the immunoglobulin heavy-chain locus on the 14q+ chromosome as previously observed in follicular lymphoma. An N region and heptamer and nonamer signal sequences indicated that this translocation occurred as a mistake in VH-DH-JH joining (where VH and DH are the variable and diversity segments). In the t(8;14) translocation, the breakpoint was located immediately 5' of the first exon of the MYC protooncogene, which was juxtaposed with the Cγ2 constant gene segment of the second 14q+ chromosome. The finding of repeated sequences typical of switch regions suggested that this translocation occurred during heavy-chain isotype switching, resulting in progression to pre-B-cell leukemia with both the 5(8;14) and the t(14;18) translocations. The terminal deoxynucleotidyltransferase-positive phenotype of the patient's leukemic cells further suggests that the pre-B-cell leukemia was derived from a pre-B cell carrying a t(14;18) translocation in the original follicular lymphoma. The polymerase chain reaction method was then used to identify cancer cells in the bone marrow of the patient

  5. Targeting Syk-activated B cells in murine and human chronic graft-versus-host disease.

    Science.gov (United States)

    Flynn, Ryan; Allen, Jessica L; Luznik, Leo; MacDonald, Kelli P; Paz, Katelyn; Alexander, Kylie A; Vulic, Ante; Du, Jing; Panoskaltsis-Mortari, Angela; Taylor, Patricia A; Poe, Jonathan C; Serody, Jonathan S; Murphy, William J; Hill, Geoffrey R; Maillard, Ivan; Koreth, John; Cutler, Corey S; Soiffer, Robert J; Antin, Joseph H; Ritz, Jerome; Chao, Nelson J; Clynes, Raphael A; Sarantopoulos, Stefanie; Blazar, Bruce R

    2015-06-25

    Novel therapies for chronic graft-versus-host disease (cGVHD) are needed. Aberrant B-cell activation has been demonstrated in mice and humans with cGVHD. Having previously found that human cGVHD B cells are activated and primed for survival, we sought to further evaluate the role of the spleen tyrosine kinase (Syk) in cGVHD in multiple murine models and human peripheral blood cells. In a murine model of multiorgan system, nonsclerodermatous disease with bronchiolitis obliterans where cGVHD is dependent on antibody and germinal center (GC) B cells, we found that activation of Syk was necessary in donor B cells, but not T cells, for disease progression. Bone marrow-specific Syk deletion in vivo was effective in treating established cGVHD, as was a small-molecule inhibitor of Syk, fostamatinib, which normalized GC formation and decreased activated CD80/86(+) dendritic cells. In multiple distinct models of sclerodermatous cGVHD, clinical and pathological disease manifestations were not eliminated when mice were therapeutically treated with fostamatinib, though both clinical and immunologic effects could be observed in one of these scleroderma models. We further demonstrated that Syk inhibition was effective at inducing apoptosis of human cGVHD B cells. Together, these data demonstrate a therapeutic potential of targeting B-cell Syk signaling in cGVHD. PMID:25852057

  6. An adaptive immune response driven by mature, antigen-experienced T and B cells within the microenvironment of oral squamous cell carcinoma.

    Science.gov (United States)

    Quan, Hongzhi; Fang, Liangjuan; Pan, Hao; Deng, Zhiyuan; Gao, Shan; Liu, Ousheng; Wang, Yuehong; Hu, Yanjia; Fang, Xiaodan; Yao, Zhigang; Guo, Feng; Lu, Ruohuang; Xia, Kun; Tang, Zhangui

    2016-06-15

    Lymphocyte infiltrates have been observed in the microenvironment of oral cancer; however, little is known about whether the immune response of the lymphocyte infiltrate affects tumor biology. For a deeper understanding of the role of the infiltrating-lymphocytes in oral squamous cell carcinoma (OSCC), we characterized the lymphocyte infiltrate repertoires and defined their features. Immunohistochemistry revealed considerable T and B cell infiltrates and lymphoid follicles with germinal center-like structures within the tumor microenvironment. Flow cytometry demonstrated that populations of antigen-experienced CD4+ and CD8+ cells were present, as well as an enrichment of regulatory T cells; and T cells expressing programmed death-1 (PD-1) and T cell Ig and mucin protein-3 (Tim-3), indicative of exhaustion, within the tumor microenvironment. Characterization of tumor-infiltrating B cells revealed clear evidence of antigen exposure, in that the cardinal features of an antigen-driven B cell response were present, including somatic mutation, clonal expansion, intraclonal variation and isotype switching. Collectively, our results point to an adaptive immune response occurring within the OSCC microenvironment, which may be sustained by the expression of specific antigens in the tumor. PMID:26815146

  7. Murine retroviruses activate B cells via interaction with toll-like receptor 4

    OpenAIRE

    Rassa, John C.; Meyers, Jennifer L.; Zhang, Yuanming; Kudaravalli, Rama; Susan R Ross

    2002-01-01

    Although most retroviruses require activated cells as their targets for infection, it is not known how this is achieved in vivo. A candidate protein for the activation of B cells by either mouse mammary tumor virus (MMTV) or murine leukemia virus is the toll-like receptor 4 (TLR4), a component of the innate immune system. MMTV caused B cell activation in C3H/HeN mice but not in C3H/HeJ or BALB/c (C.C3H Tlr4lps-d) congenic mice, both of which have a mutant TLR4 gene. This activation was indepe...

  8. Comparison of human B cell activation by TLR7 and TLR9 agonists

    Directory of Open Access Journals (Sweden)

    Neys Lori

    2008-07-01

    Full Text Available Abstract Background Human B cells and plasmacytoid dendritic cells (pDC are the only cells known to express both TLR7 and TLR9. Plasmacytoid dendritic cells are the primary IFN-α producing cells in response to TLR7 and TLR9 agonists. The direct effects of TLR7 stimulation on human B cells is less understood. The objective of this study was to compare the effects of TLR7 and TLR9 stimulation on human B cell function. Results Gene expression and protein production of cytokines, chemokines, various B cell activation markers, and immunoglobulins were evaluated. Purified human CD19+ B cells (99.9%, containing both naïve and memory populations from peripheral blood were stimulated with a TLR7-selective agonist (852A, TLR7/8 agonist (3M-003, or TLR9 selective agonist CpG ODN (CpG2006. TLR7 and TLR9 agonists similarly modulated the expression of cytokine and chemokine genes (IL-6, MIP1 alpha, MIP1 beta, TNF alpha and LTA, co-stimulatory molecules (CD80, CD40 and CD58, Fc receptors (CD23, CD32, anti-apoptotic genes (BCL2L1, certain transcription factors (MYC, TCFL5, and genes critical for B cell proliferation and differentiation (CD72, IL21R. Both agonists also induced protein expression of the above cytokines and chemokines. Additionally, TLR7 and TLR9 agonists induced the production of IgM and IgG. A TLR8-selective agonist was comparatively ineffective at stimulating purified human B cells. Conclusion These results demonstrate that despite their molecular differences, the TLR7 and TLR9 agonists induce similar genes and proteins in purified human B cells.

  9. Adoptive transfer of syngeneic T cells transduced with a chimeric antigen receptor that recognizes murine CD19 can eradicate lymphoma and normal B cells

    OpenAIRE

    Kochenderfer, James N.; Yu, Zhiya; Frasheri, Dorina; Restifo, Nicholas P; Rosenberg, Steven A.

    2010-01-01

    Adoptive T-cell therapy with anti-CD19 chimeric antigen receptor (CAR)–expressing T cells is a new approach for treating advanced B-cell malignancies. To evaluate anti-CD19–CAR-transduced T cells in a murine model of adoptive T-cell therapy, we developed a CAR that specifically recognized murine CD19. We used T cells that were retrovirally transduced with this CAR to treat mice bearing a syngeneic lymphoma that naturally expressed the self-antigen murine CD19. One infusion of anti-CD19–CAR-tr...

  10. Molecular evolution of a central region containing B cell epitopes in the gene encoding the p67 sporozoite antigen within a field population of Theileria parva

    OpenAIRE

    Obara, Isaiah; Ulrike, Seitzer; Musoke, Tony; Paul R Spooner; Jabbar, Ahmed; Odongo, David; Kemp, Stephen; Silva, Joana C.; Bishop, Richard P.

    2015-01-01

    Protective immunity induced by the infective sporozoite stage of Theileria parva indicates a potential role for antibodies directed against conserved serologically reactive regions of the major sporozoite surface antigen p67 in vaccination to control the parasite. We have examined the allelic variation and determined the extent of B cell epitope polymorphism of the gene encoding p67 among field isolates originating from cattle exposed to infected ticks in the Marula area of the rift valley in...

  11. Epstein-Barr nuclear antigen 1 induces expression of the cellular microRNA hsa-miR-127 and impairing B-cell differentiation in EBV-infected memory B cells. New insights into the pathogenesis of Burkitt lymphoma

    International Nuclear Information System (INIS)

    Epstein-Barr Virus (EBV) is a γ-herpesvirus that infects >90% of the human population. Although EBV persists in its latent form in healthy carriers, the virus is also associated with several human cancers. EBV is strongly associated with Burkitt lymphoma (BL), even though there is still no satisfactory explanation of how EBV participates in BL pathogenesis. However, new insights into the interplay between viruses and microRNAs (miRNAs) have recently been proposed. In particular, it has been shown that B-cell differentiation in EBV-positive BL is impaired at the post-transcriptional level by altered expression of hsa-miR-127. Here, we show that the overexpression of hsa-miR-127 is due to the presence of the EBV-encoded nuclear antigen 1 (EBNA1) and give evidence of a novel mechanism of direct regulation of the human miRNA by this viral product. Finally, we show that the combinatorial expression of EBNA1 and hsa-miR-127 affects the expression of master B-cell regulators in human memory B cells, confirming the scenario previously observed in EBV-positive BL primary tumors and cell lines. A good understanding of these mechanisms will help to clarify the complex regulatory networks between host and pathogen, and favor the design of more specific treatments for EBV-associated malignancies

  12. Blockade of B-cell-activating factor suppresses lupus-like syndrome in autoimmune BXSB mice

    OpenAIRE

    Ding, Hanlu; Wang, Li; Wu, Xiongfei; Yan, Jun; He, Yani; Ni, Bing; Gao, Wenda; Zhong, Xuemei

    2009-01-01

    Abstract B-cell-activating factor (BAFF), a member of the tumour necrosis factor superfamily, plays a critical role in the maturation, homeostasis and function of B cells. In this study, we demonstrated the biological outcome of BAFF blockade in BXSB murine lupus model, using a soluble fusion protein consisting of human BAFF-R and human mutant IgG4 Fc. Mutation of Leu235 to Glu in IgG4 Fc eliminated antibody-dependent cell cytotoxicity (ADCC) and complement lysis activity, and generated a pro...

  13. A conditional form of Bruton's tyrosine kinase is sufficient to activate multiple downstream signaling pathways via PLC Gamma 2 in B cells

    Directory of Open Access Journals (Sweden)

    Witte Owen N

    2001-06-01

    Full Text Available Abstract Background Bruton's tyrosine kinase (Btk is essential for B cell development and function. Mutations of Btk elicit X-linked agammaglobulinemia in humans and X-linked immunodeficiency in the mouse. Btk has been proposed to participate in B cell antigen receptor-induced signaling events leading to activation of phospholipase C-γ2 (PLCγ2 and calcium mobilization. However it is unclear whether Btk activation is alone sufficient for these signaling events, and whether Btk can activate additional pathways that do not involve PLCγ2. To address such issues we have generated Btk:ER, a conditionally active form of the kinase, and expressed it in the PLCγ2-deficient DT40 B cell line. Results Activation of Btk:ER was sufficient to induce multiple B cell signaling pathways in PLCγ2-sufficient DT40 cells. These included tyrosine phosphorylation of PLCγ2, mobilization of intracellular calcium, activation of extracellular signal-regulated kinase (ERK and c-Jun NH2-terminal kinase (JNK mitogen-activated protein kinase (MAPK pathways, and apoptosis. In DT40 B cells deficient for PLCγ2, Btk:ER activation failed to induce the signaling events described above with the consequence that the cells failed to undergo apoptosis. Conclusions These data suggest that Btk:ER regulates downstream signaling pathways primarily via PLCγ2 in B cells. While it is not known whether activated Btk:ER precisely mimics activated Btk, this conditional system will likely facilitate the dissection of the role of Btk and its family members in a variety of biological processes in many different cell types.

  14. Elucidation of tonic and activated B-cell receptor signaling in Burkitt's lymphoma provides insights into regulation of cell survival.

    Science.gov (United States)

    Corso, Jasmin; Pan, Kuan-Ting; Walter, Roland; Doebele, Carmen; Mohr, Sebastian; Bohnenberger, Hanibal; Ströbel, Philipp; Lenz, Christof; Slabicki, Mikolaj; Hüllein, Jennifer; Comoglio, Federico; Rieger, Michael A; Zenz, Thorsten; Wienands, Jürgen; Engelke, Michael; Serve, Hubert; Urlaub, Henning; Oellerich, Thomas

    2016-05-17

    Burkitt's lymphoma (BL) is a highly proliferative B-cell neoplasm and is treated with intensive chemotherapy that, because of its toxicity, is often not suitable for the elderly or for patients with endemic BL in developing countries. BL cell survival relies on signals transduced by B-cell antigen receptors (BCRs). However, tonic as well as activated BCR signaling networks and their relevance for targeted therapies in BL remain elusive. We have systematically characterized and compared tonic and activated BCR signaling in BL by quantitative phosphoproteomics to identify novel BCR effectors and potential drug targets. We identified and quantified ∼16,000 phospho-sites in BL cells. Among these sites, 909 were related to tonic BCR signaling, whereas 984 phospho-sites were regulated upon BCR engagement. The majority of the identified BCR signaling effectors have not been described in the context of B cells or lymphomas yet. Most of these newly identified BCR effectors are predicted to be involved in the regulation of kinases, transcription, and cytoskeleton dynamics. Although tonic and activated BCR signaling shared a considerable number of effector proteins, we identified distinct phosphorylation events in tonic BCR signaling. We investigated the functional relevance of some newly identified BCR effectors and show that ACTN4 and ARFGEF2, which have been described as regulators of membrane-trafficking and cytoskeleton-related processes, respectively, are crucial for BL cell survival. Thus, this study provides a comprehensive dataset for tonic and activated BCR signaling and identifies effector proteins that may be relevant for BL cell survival and thus may help to develop new BL treatments. PMID:27155012

  15. Activation of B cells by 1 microm particulate lysozyme or peptides: a Th-dependent pathway requiring CD40-CD40 ligand interaction.

    Science.gov (United States)

    Sedlik, C; Rojas, M; Leclerc, C

    1998-08-01

    Many antigens encountered by the immune system are included in complex structures such as bacteria or parasites. We previously developed an in vivo model to study the immunogenicity of particulate antigens, based on covalent linkage of proteins or peptides to 1 microm latex particles and showed that these antigens cannot be presented to MHC class II-restricted specific T cells by B cells. However, they induce strong CD4+ T cell responses when injected to mice without adjuvant. The present study demonstrates that four out of the five proteins tested did not stimulate antibody synthesis when linked to 1 microm microparticles, although a strong IgG production was induced by the same proteins administered in soluble form with adjuvant. In contrast, lysozyme and two synthetic peptides containing B and T cell viral epitopes induced strong and long-lasting specific antibody responses when linked to 1 micrometer synthetic beads. The isotypic pattern of antibodies induced by particulate lysozyme was similar to that induced by the soluble protein in alum. Studies using CD4+ T cell-depleted mice revealed that the induction of antibodies by particulate lysozyme strictly required Th cell activity. Moreover, the T-B cell cooperation involved in B cell activation by antigens linked to beads required CD40-CD40 ligand interaction. Thus, these particulate antigens provide a useful tool to study the mechanisms of induction of antibody response against complex bacterial or parasitic antigens. Moreover, they may represent attractive candidates to elaborate efficient new vaccines using short synthetic peptides. PMID:9723697

  16. Interleukin 2 receptors on human B cells. Implications for the role of interleukin 2 in human B cell function

    OpenAIRE

    1985-01-01

    In the present study, we examined the expression of interleukin 2 (IL- 2) receptors on normal human B cells as well as established B cell lines. Anti-Tac monoclonal antibody did not bind to freshly separated normal human B cells. Unexpectedly, with the appropriate activation of the normal B cells by anti-mu antibody, phorbol myristate acetate, or Staphylococcus aureus Cowan I (SAC), Tac antigen was induced on the activated B cells. Anti-Tac antibody showed consistent reactivity with two B cel...

  17. Genomic regions associated with multiple sclerosis are active in B cells.

    Directory of Open Access Journals (Sweden)

    Giulio Disanto

    Full Text Available More than 50 genomic regions have now been shown to influence the risk of multiple sclerosis (MS. However, the mechanisms of action, and the cell types in which these associated variants act at the molecular level remain largely unknown. This is especially true for associated regions containing no known genes. Given the evidence for a role for B cells in MS, we hypothesized that MS associated genomic regions co-localized with regions which are functionally active in B cells. We used publicly available data on 1 MS associated regions and single nucleotide polymorphisms (SNPs and 2 chromatin profiling in B cells as well as three additional cell types thought to be unrelated to MS (hepatocytes, fibroblasts and keratinocytes. Genomic intervals and SNPs were tested for overlap using the Genomic Hyperbrowser. We found that MS associated regions are significantly enriched in strong enhancer, active promoter and strong transcribed regions (p = 0.00005 and that this overlap is significantly higher in B cells than control cells. In addition, MS associated SNPs also land in active promoter (p = 0.00005 and enhancer regions more than expected by chance (strong enhancer p = 0.0006; weak enhancer p = 0.00005. These results confirm the important role of the immune system and specifically B cells in MS and suggest that MS risk variants exert a gene regulatory role. Previous studies assessing MS risk variants in T cells may be missing important effects in B cells. Similar analyses in other immunological cell types relevant to MS and functional studies are necessary to fully elucidate how genes contribute to MS pathogenesis.

  18. Plasmacytoid dendritic cells regulate autoreactive B cell activation via soluble factors and in a cell-to-cell contact manner.

    Science.gov (United States)

    Ding, Chuanlin; Cai, Yihua; Marroquin, Jose; Ildstad, Suzanne T; Yan, Jun

    2009-12-01

    Plasmacytoid dendritic cells (pDCs) are specialized type I IFN producers, which play an important role in pathogenesis of autoimmune disorders. Dysregulated autoreactive B cell activation is a hallmark in most autoimmune diseases. This study was undertaken to investigate interactions between pDCs and autoreactive B cells. After coculture of autoreactive B cells that recognize self-Ag small nuclear ribonucleoprotein particles with activated pDCs, we found that pDCs significantly enhance autoreactive B cell proliferation, autoantibody production, and survival in response to TLR and BCR stimulation. Neutralization of IFN-alpha/beta and IL-6 abrogated partially pDC-mediated enhancement of autoreactive B cell activation. Transwell studies demonstrated that pDCs could provide activation signals to autoreactive B cells via a cell-to-cell contact manner. The involvement of the ICAM-1-LFA-1 pathway was revealed as contributing to this effect. This in vitro enhancement effect was further demonstrated by an in vivo B cell adoptive transfer experiment, which showed that autoreactive B cell proliferation and activation were significantly decreased in MyD88-deficient mice compared with wild-type mice. These data suggest the dynamic interplay between pDCs and B cells is required for full activation of autoreactive B cells upon TLR or BCR stimulation. PMID:19890051

  19. B-Cell Activating Factor (BAFF) is elevated in Chronic Granulomatous Disease

    OpenAIRE

    Matharu, Kabir; Zarember, Kol A.; Marciano, Beatriz E.; Kuhns, Douglas B; Spalding, Christine; Garofalo, Mary; DiMaggio, Thomas; Estwick, Tyra; Huang, Chiung-yu; Fink, Danielle; Priel, Debra L.; Fleisher, Thomas A.; Holland, Steven M.; Malech, Harry L.; Gallin, John I.

    2013-01-01

    Chronic Granulomatous Disease (CGD) is an inherited defect in superoxide production leading to life-threatening infections, granulomas, and, possibly, abnormal immunoglobulin concentrations. We investigated whether factors controlling antibody production, such as B-cell activating factor (BAFF), were altered in CGD. CGD subjects had significantly increased mean (2.3-fold, p

  20. Rainbow trout CK9, a CCL25-like ancient chemokine that attracts and regulates B cells and macrophages, the main antigen presenting cells in fish

    Science.gov (United States)

    Aquilino, Carolina; Granja, Aitor G.; Castro, Rosario; Wang, Tiehui; Abos, Beatriz; Parra, David; Secombes, Christopher J.; Tafalla, Carolina

    2016-01-01

    CK9 is a rainbow trout (Oncorhynchus mykiss) CC chemokine phylogenetically related to mammalian CCL25. Although CK9 is known to be transcriptionally regulated in response to inflammation particularly in mucosal tissues, its functionality has never been revealed. In the current work, we have demonstrated that CK9 is chemoattractant for antigen presenting cells (APCs) expressing major histocompatibility complex class II (MHC II) on the cell surface. Among these APCs, CK9 has a strong chemotactic capacity for both B cells (IgM+ and IgT+) and macrophages. Along with its chemotactic capacities, CK9 modulated the MHC II turnover of B lymphocytes and up-regulated the phagocytic capacity of both IgM+ cells and macrophages. Although CK9 had no lymphoproliferative effects, it increased the survival of IgT+ lymphocytes. Furthermore, we have established that the chemoattractant capacity of CK9 is strongly increased after pre-incubation of leukocytes with a T-independent antigen, whereas B cell receptor (BCR) cross-linking strongly abrogated their capacity to migrate to CK9, indicating that CK9 preferentially attracts B cells at the steady state or under BCR-independent stimulation. These results point to CK9 being a key regulator of B lymphocyte trafficking in rainbow trout, able to modulate innate functions of teleost B lymphocytes and macrophages. PMID:27003360

  1. High-throughput combinatorial screening identifies drugs that cooperate with ibrutinib to kill activated B-cell-like diffuse large B-cell lymphoma cells.

    Science.gov (United States)

    Mathews Griner, Lesley A; Guha, Rajarshi; Shinn, Paul; Young, Ryan M; Keller, Jonathan M; Liu, Dongbo; Goldlust, Ian S; Yasgar, Adam; McKnight, Crystal; Boxer, Matthew B; Duveau, Damien Y; Jiang, Jian-Kang; Michael, Sam; Mierzwa, Tim; Huang, Wenwei; Walsh, Martin J; Mott, Bryan T; Patel, Paresma; Leister, William; Maloney, David J; Leclair, Christopher A; Rai, Ganesha; Jadhav, Ajit; Peyser, Brian D; Austin, Christopher P; Martin, Scott E; Simeonov, Anton; Ferrer, Marc; Staudt, Louis M; Thomas, Craig J

    2014-02-11

    The clinical development of drug combinations is typically achieved through trial-and-error or via insight gained through a detailed molecular understanding of dysregulated signaling pathways in a specific cancer type. Unbiased small-molecule combination (matrix) screening represents a high-throughput means to explore hundreds and even thousands of drug-drug pairs for potential investigation and translation. Here, we describe a high-throughput screening platform capable of testing compounds in pairwise matrix blocks for the rapid and systematic identification of synergistic, additive, and antagonistic drug combinations. We use this platform to define potential therapeutic combinations for the activated B-cell-like subtype (ABC) of diffuse large B-cell lymphoma (DLBCL). We identify drugs with synergy, additivity, and antagonism with the Bruton's tyrosine kinase inhibitor ibrutinib, which targets the chronic active B-cell receptor signaling that characterizes ABC DLBCL. Ibrutinib interacted favorably with a wide range of compounds, including inhibitors of the PI3K-AKT-mammalian target of rapamycin signaling cascade, other B-cell receptor pathway inhibitors, Bcl-2 family inhibitors, and several components of chemotherapy that is the standard of care for DLBCL. PMID:24469833

  2. Eicosapentaenoic and docosahexaenoic acid ethyl esters differentially enhance B-cell activity in murine obesity[S

    OpenAIRE

    Teague, Heather; Harris, Mitchel; Fenton, Jenifer; Lallemand, Perrine; Shewchuk, Brian M.; Shaikh, Saame Raza

    2014-01-01

    EPA and DHA are not biologically equivalent; however, their individual activity on B cells is unknown. We previously reported fish oil enhanced murine B-cell activity in obesity. To distinguish between the effects of EPA and DHA, we studied the ethyl esters of EPA and DHA on murine B-cell function as a function of time. We first demonstrate that EPA and DHA maintained the obese phenotype, with no improvements in fat mass, adipose inflammatory cytokines, fasting insulin, or glucose clearance. ...

  3. MYC/BCL2 protein coexpression contributes to the inferior survival of activated B-cell subtype of diffuse large B-cell lymphoma and demonstrates high-risk gene expression signatures

    DEFF Research Database (Denmark)

    Hu, Shimin; Xu-Monette, Zijun Y; Tzankov, Alexander;

    2013-01-01

    Diffuse large B-cell lymphoma (DLBCL) is stratified into prognostically favorable germinal center B-cell (GCB)-like and unfavorable activated B-cell (ABC)-like subtypes based on gene expression signatures. In this study, we analyzed 893 de novo DLBCL patients treated with R-CHOP (rituximab, cyclo...

  4. Induction of N-glycosylation activity in B-cells by PMA and ionomycin

    Energy Technology Data Exchange (ETDEWEB)

    Rush, J.S.; Snow, E.C.; Waechter, C.J.

    1987-05-01

    Although N-glycosylation activity in murine splenic B-lymphocytes (B-cells) was not stimulated by phorbol 12-myristate 13-acetate (PMA) or ionomycin added separately, the combination of PMA plus ionomycin produced a dramatic increase in the incorporation of (TH)mannose into dolichol-linked oligosaccharides and glycoprotein. Maximal stimulation of N-glycosylation activity was observed after exposure of B-cells to 1 M ionomycin in the presence of 1-10 ng/ml PMA for 36 hr. At these concentrations of PMA and ionomycin, RNA and DNA synthesis, which required 1% and 5% fetal calf serum, respectively, peaked at 20-24 hr. 1-(5-Isoquinolinylsulfonyl)-2-methylpiperazine (H-7), a potent inhibitor of protein kinase C, prevented the stimulation of N-glycosylation activity, RNA synthesis and DNA synthesis. N-(2-(Methylamino)ethyl)-5-isoquinolinesulfonamide (H-8), an effective inhibitor of cyclic nucleotide-dependent protein kinases, reduced N-glycosylation activity, but required a higher concentration than H-7. These results support the conclusion that protein kinase C plays a key role in the activation of B-cells. Current work is aimed at determining if any enzymes in the dolichol pathway are activated directly by protein kinase C, or if the stimulation in N-glycosylation activity is a secondary effect of the primary activation process.

  5. Analysis by Flow Cytometry of B-Cell Activation and Antibody Responses Induced by Toll-Like Receptors.

    Science.gov (United States)

    Pone, Egest J

    2016-01-01

    Toll-like receptors (TLRs) are expressed in B lymphocytes and contribute to B-cell activation, antibody responses, and their maturation. TLR stimulation of mouse B cells induces class switch DNA recombination (CSR) to isotypes specified by cytokines, and also induces formation of IgM(+) as well as class-switched plasma cells. B-cell receptor (BCR) signaling, while on its own inducing limited B-cell proliferation and no CSR, can enhance CSR driven by TLRs. Particular synergistic or antagonistic interactions among TLR pathways, BCR, and cytokine signaling can have important consequences for B-cell activation, CSR, and plasma cell formation. This chapter outlines protocols for the induction and analysis of B-cell activation and antibody production by TLRs with or without other stimuli. PMID:26803633

  6. A Rapid Method to Characterize Mouse IgG Antibodies and Isolate Native Antigen Binding IgG B Cell Hybridomas.

    Directory of Open Access Journals (Sweden)

    Haolin Liu

    Full Text Available B cell hybridomas are an important source of monoclonal antibodies. In this paper, we developed a high-throughput method to characterize mouse IgG antibodies using surface plasmon resonance technology. This assay rapidly determines their sub-isotypes, whether they bind native antigen and their approximate affinities for the antigen using only 50 μl of hybridoma cell culture supernatant. Moreover, we found that mouse hybridomas secreting IgG antibodies also have membrane form IgG expression without Igα. Based on this surface IgG, we used flow cytometry to isolate rare γ2a isotype switched variants from a γ2b antibody secreting hybridoma cell line. Also, we used fluorescent antigen to single cell sort antigen binding hybridoma cells from bulk mixture of fused hybridoma cells instead of the traditional multi-microwell plate screening and limiting dilution sub-cloning thus saving time and labor. The IgG monoclonal antibodies specific for the native antigen identified with these methods are suitable for in vivo therapeutic uses, but also for sandwich ELISA assays, histology, flow cytometry, immune precipitation and x-ray crystallography.

  7. Chimeric antigen receptor-engineered cytokine-induced killer cells overcome treatment resistance of pre-B-cell acute lymphoblastic leukemia and enhance survival.

    Science.gov (United States)

    Oelsner, Sarah; Wagner, Juliane; Friede, Miriam E; Pfirrmann, Verena; Genßler, Sabrina; Rettinger, Eva; Buchholz, Christian J; Pfeifer, Heike; Schubert, Ralf; Ottmann, Oliver G; Ullrich, Evelyn; Bader, Peter; Wels, Winfried S

    2016-10-15

    Pre-emptive cancer immunotherapy by donor lymphocyte infusion (DLI) using cytokine-induced killer (CIK) cells may be beneficial to prevent relapse with a reduced risk of causing graft-versus-host-disease. CIK cells are a heterogeneous effector cell population including T cells (CD3(+) CD56(-) ), natural killer (NK) cells (CD3(-) CD56(+) ) and natural killer T (T-NK) cells (CD3(+) CD56(+) ) that exhibit non-major histocompatibility complex (MHC)-restricted cytotoxicity and are generated by ex vivo expansion of peripheral blood mononuclear cells in the presence of interferon (IFN)-γ, anti-CD3 antibody, interleukin-2 (IL-2) and interleukin-15 (IL-15). To facilitate selective target-cell recognition and enhance specific cytotoxicity against B-cell acute lymphoblastic leukemia (B-ALL), we transduced CIK cells with a lentiviral vector encoding a chimeric antigen receptor (CAR) that carries a composite CD28-CD3ζ domain for signaling and a CD19-specific scFv antibody fragment for cell binding (CAR 63.28.z). In vitro analysis revealed high and specific cell killing activity of CD19-targeted CIK/63.28.z cells against otherwise CIK-resistant cancer cell lines and primary B-ALL blasts, which was dependent on CD19 expression and CAR signaling. In a xenograft model in immunodeficient mice, treatment with CIK/63.28.z cells in contrast to therapy with unmodified CIK cells resulted in complete and durable molecular remissions of established primary pre-B-ALL. Our results demonstrate potent antileukemic activity of CAR-engineered CIK cells in vitro and in vivo, and suggest this strategy as a promising approach for adoptive immunotherapy of refractory pre-B-ALL. PMID:27253354

  8. Human Leukocyte Antigen Class I and II Alleles and Overall Survival in Diffuse Large B-Cell Lymphoma and Follicular Lymphoma

    OpenAIRE

    Yani Lu; Amr M Abdou; Cerhan, James R.; Morton, Lindsay M.; Richard K Severson; Scott Davis; Wendy Cozen; Nathaniel Rothman; Leslie Bernstein; Stephen Chanock; Patricia Hartge; Wang, Sophia S.

    2011-01-01

    Genetic variation in the 6p21 chromosomal region, including human leukocyte antigen (HLA) genes and tumor necrosis factor (TNF), has been linked to both etiology and clinical outcomes of lymphomas. We estimated the effects of HLA class I (A, B, and C), class II DRB1 alleles, and the ancestral haplotype (AH) 8.1 (HLAA*01-B*08-DRB1*03-TNF-308A) on overall survival (OS) among patients with diffuse large B-cell lymphoma (DLBCL) and follicular lymphoma (FL) in a population-based study of non-Hodgk...

  9. Insights into the Molecular Pathogenesis of Activated B-Cell-like Diffuse Large B-Cell Lymphoma and Its Therapeutic Implications

    Directory of Open Access Journals (Sweden)

    Georg Lenz

    2015-05-01

    Full Text Available Within the last couple of years, the understanding of the molecular mechanisms that drive the pathogenesis of diffuse large B-cell lymphoma (DLBCL has significantly improved. Large-scale gene expression profiling studies have led to the discovery of several molecularly defined subtypes that are characterized by specific oncogene addictions and significant differences in their outcome. Next generation sequencing efforts combined with RNA interference screens frequently identify crucial oncogenes that lead to constitutive activation of various signaling pathways that drive lymphomagenesis. This review summarizes our current understanding of the molecular pathogenesis of the activated B-cell-like (ABC DLBCL subtype that is characterized by poor prognosis. A special emphasis is put on findings that might impact therapeutic strategies of affected patients.

  10. Insights into the Molecular Pathogenesis of Activated B-Cell-like Diffuse Large B-Cell Lymphoma and Its Therapeutic Implications

    Energy Technology Data Exchange (ETDEWEB)

    Lenz, Georg [Translational Oncology, Department of Medicine A, Albert-Schweitzer Campus 1, University Hospital Münster, 48149 Münster (Germany); Cluster of Excellence EXC 1003, Cells in Motion, 48149 Münster (Germany)

    2015-05-22

    Within the last couple of years, the understanding of the molecular mechanisms that drive the pathogenesis of diffuse large B-cell lymphoma (DLBCL) has significantly improved. Large-scale gene expression profiling studies have led to the discovery of several molecularly defined subtypes that are characterized by specific oncogene addictions and significant differences in their outcome. Next generation sequencing efforts combined with RNA interference screens frequently identify crucial oncogenes that lead to constitutive activation of various signaling pathways that drive lymphomagenesis. This review summarizes our current understanding of the molecular pathogenesis of the activated B-cell-like (ABC) DLBCL subtype that is characterized by poor prognosis. A special emphasis is put on findings that might impact therapeutic strategies of affected patients.

  11. G0 B cells activated by anti-μ acquire the ability to proliferate in response to B cell-activating factors independently from entry into G1 phase

    International Nuclear Information System (INIS)

    The early steps in B cell activation were investigated and it was determined that activation could be separated from entry into cell cycle. Purified B cells from BALB/c nu/nu mice, activated under controlled conditions by affinity-purified rabbit IgG anti-μ or F(ab')2 fragments prepared from the same antibodies, were sorted according to size by flow cytometry. Approximately 80% of the B cells were small and were shown to be in G0 state by quantitative RNA analysis and by [3H]uridine and [3H]thymidine incorporation studies. The sorted G0 B cells, when again incubated in serum-free medium with the appropriate anti-μ preparations, remained in G0. However, G0 B cells in the presence of anti-μ have undergone significant change(s), in as much as they were now able to proliferate in response to soluble mediators. The ability to develop a proliferative response to B cell-activating factor(s), acquired independently from entry into cell cycle, characterizes another important step in early B cell activation and also indicates that G0 B cells characterized on the basis of cell size, density, and/or RNA content may be in an activated state

  12. Deficiency of the B Cell-Activating Factor Receptor Results in Limited CD169+ Macrophage Function during Viral Infection

    OpenAIRE

    Xu, Haifeng C.; Huang, Jun; Khairnar, Vishal; Duhan, Vikas; Pandyra, Aleksandra A; Grusdat, Melanie; Shinde, Prashant; McIlwain, David R.; Maney, Sathish Kumar; Gommerman, Jennifer; Löhning, Max; Ohashi, Pamela S.; Mak, Tak W; Pieper, Kathrin; Sic, Heiko

    2015-01-01

    The B cell-activating factor (BAFF) is critical for B cell development and humoral immunity in mice and humans. While the role of BAFF in B cells has been widely described, its role in innate immunity remains unknown. Using BAFF receptor (BAFFR)-deficient mice, we characterized BAFFR-related innate and adaptive immune functions following infection with vesicular stomatitis virus (VSV) and lymphocytic choriomeningitis virus (LCMV). We identified a critical role for BAFFR signaling in the gener...

  13. The BCL6 RD2 domain governs commitment of activated B cells to form germinal centers.

    Science.gov (United States)

    Huang, Chuanxin; Gonzalez, David G; Cote, Christine M; Jiang, Yanwen; Hatzi, Katerina; Teater, Matt; Dai, Kezhi; Hla, Timothy; Haberman, Ann M; Melnick, Ari

    2014-09-11

    To understand how the Bcl6 transcriptional repressor functions in the immune system, we disrupted its RD2 repression domain in mice. Bcl6RD2(MUT) mice exhibit a complete loss of germinal center (GC) formation but retain normal extrafollicular responses. Bcl6RD2(MUT) antigen-engaged B cells migrate to the interfollicular zone and interact with cognate T helper cells. However, these cells fail to complete early GC-commitment differentiation and coalesce as nascent GC aggregates. Bcl6 directly binds and represses trafficking receptors S1pr1 and Gpr183 by recruiting Hdac2 through the RD2 domain. Deregulation of these genes impairs B cell migration and may contribute to GC failure in Bcl6RD2(MUT) mice. The development of functional GC-TFH cells was partially impaired in Bcl6RD2(MUT) mice. In contrast to Bcl6(-/-) mice, Bcl6RD2(MUT) animals experience no inflammatory disease or macrophage deregulation. These results reveal an essential role for RD2 repression in early GC commitment and striking biochemical specificity in Bcl6 control of humoral and innate immune-cell phenotypes. PMID:25176650

  14. The BCL6 RD2 Domain Governs Commitment of Activated B Cells to Form Germinal Centers

    Directory of Open Access Journals (Sweden)

    Chuanxin Huang

    2014-09-01

    Full Text Available To understand how the Bcl6 transcriptional repressor functions in the immune system, we disrupted its RD2 repression domain in mice. Bcl6RD2MUT mice exhibit a complete loss of germinal center (GC formation but retain normal extrafollicular responses. Bcl6RD2MUT antigen-engaged B cells migrate to the interfollicular zone and interact with cognate T helper cells. However, these cells fail to complete early GC-commitment differentiation and coalesce as nascent GC aggregates. Bcl6 directly binds and represses trafficking receptors S1pr1 and Gpr183 by recruiting Hdac2 through the RD2 domain. Deregulation of these genes impairs B cell migration and may contribute to GC failure in Bcl6RD2MUT mice. The development of functional GC-TFH cells was partially impaired in Bcl6RD2MUT mice. In contrast to Bcl6−/− mice, Bcl6RD2MUT animals experience no inflammatory disease or macrophage deregulation. These results reveal an essential role for RD2 repression in early GC commitment and striking biochemical specificity in Bcl6 control of humoral and innate immune-cell phenotypes.

  15. B-cell responses to pregnancy-restricted and -unrestricted Plasmodium falciparum erythrocyte membrane protein 1 antigens in Ghanaian women naturally exposed to malaria parasites

    DEFF Research Database (Denmark)

    Ampomah, Paulina; Stevenson, Liz; Ofori, Michael F;

    2014-01-01

    Protective immunity to Plasmodium falciparum malaria acquired after natural exposure is largely antibody mediated. IgG-specific P. falciparum EMP1 (PfEMP1) proteins on the infected erythrocyte surface are particularly important. The transient antibody responses and the slowly acquired protective...... immunity probably reflect the clonal antigenic variation and allelic polymorphism of PfEMP1. However, it is likely that other immune-evasive mechanisms are also involved, such as interference with formation and maintenance of immunological memory. We measured PfEMP1-specific antibody levels by enzyme......-linked immunosorbent assay (ELISA) and memory B-cell frequencies by enzyme-linked immunosorbent spot (ELISPOT) assay in a cohort of P. falciparum-exposed nonpregnant Ghanaian women. The antigens used were a VAR2CSA-type PfEMP1 (IT4VAR04) with expression restricted to parasites infecting the placenta, as well as two...

  16. LRRK1 is critical in the regulation of B-cell responses and CARMA1-dependent NF-κB activation

    Science.gov (United States)

    Morimoto, Keiko; Baba, Yoshihiro; Shinohara, Hisaaki; Kang, Sujin; Nojima, Satoshi; Kimura, Tetsuya; Ito, Daisuke; Yoshida, Yuji; Maeda, Yohei; Sarashina-Kida, Hana; Nishide, Masayuki; Hosokawa, Takashi; Kato, Yasuhiro; Hayama, Yoshitomo; Kinehara, Yuhei; Okuno, Tatsusada; Takamatsu, Hyota; Hirano, Toru; Shima, Yoshihito; Narazaki, Masashi; Kurosaki, Tomohiro; Toyofuku, Toshihiko; Kumanogoh, Atsushi

    2016-01-01

    B-cell receptor (BCR) signaling plays a critical role in B-cell activation and humoral immunity. In this study, we discovered a critical function of leucine-rich repeat kinase 1 (LRRK1) in BCR-mediated immune responses. Lrrk1−/− mice exhibited altered B1a-cell development and basal immunoglobulin production. In addition, these mice failed to produce IgG3 antibody in response to T cell–independent type 2 antigen due to defects in IgG3 class-switch recombination. Concomitantly, B cells lacking LRRK1 exhibited a profound defect in proliferation and survival upon BCR stimulation, which correlated with impaired BCR-mediated NF-κB activation and reduced expression of NF-κB target genes including Bcl-xL, cyclin D2, and NFATc1/αA. Furthermore, LRRK1 physically interacted and potently synergized with CARMA1 to enhance NF-κB activation. Our results reveal a critical role of LRRK1 in NF-κB signaling in B cells and the humoral immune response. PMID:27166870

  17. Simultaneous Detection of Antigen-Specific IgG- and IgM-Secreting Cells with a B Cell Fluorospot Assay

    Directory of Open Access Journals (Sweden)

    M. Anthony Moody

    2012-03-01

    Full Text Available The traditional enzyme-linked immunospot (ELISpot assay is the gold standard for the enumeration of antigen-specific B cells. Since B cell availability from biological samples is often limited, either because of sample size/volume or the need of performing multiple analyses on the same sample, the implementation of ELISpot assay formats that allow the simultaneous detection of multiple antibody types is desirable. While dual-color ELISpot assays have been described, technical complexities have so far prevented their wide utilization as well as further expansion of their multicolor capability. An attractive solution is to replace the chromogenic reaction of the traditional ELISpot assay with a fluorescent detection system (fluorospot assay. Fluorospot assays using fluorophore-conjugated secondary antibodies in conjunction with fluorescence enhancers, FITC/anti-FITC and biotin/avidin amplification systems and dedicated equipment for spot detection have been developed to enumerate T-cells secreting two or three specific cytokines and, more recently, IgG and IgA antibody-secreting cells (ASCs. We hereby report a method for a multiplex B cell fluorospot assay that utilizes quantum-dot nanocrystals as reporters without further amplification systems or need of dedicated equipment. With this method we simultaneously enumerated HIV-1 gp41 envelope glycoprotein-specific IgG and IgM antibody-secreting cells with sensitivity comparable to that of the traditional ELISpot assay.

  18. Deposition of idiotype-anti-idiotype immune complexes in renal glomeruli after polyclonal B cell activation

    International Nuclear Information System (INIS)

    We investigated the possible role of idiotypic interactions in the pathogenesis of the glomerular lesions observed in mice undergoing polyclonal B cell activation. BALB/c mice were studied for the presence of renal deposits of T15 idiotype-anti-T15 idiotype-immune complexes (IC) after injection of bacterial lipopolysaccharides (LPS). The T15 idiotype is the major idiotype of BALB/c mice anti-phosphorylcholine (PC) antibodies, which are cross-reactive with the idiotype of the TEPC-15 myeloma protein. This model was used because T15 idiotype-anti-T15 idiotype IC have been detected in the circulation of BALB/c mice after polyclonal B cell activation. First, an idiotype-specific immunofluorescence technique allowed us to detect T15 idiotype-bearing immunoglobulins in glomeruli from day 6 to day 28 after LPS injection. Second, fluorescein isothiocyanate-conjugated TEPC-15 myeloma protein was found to localize in the glomeruli after in vivo injection 18 d after LPS administration. This renal localization was shown to be idiotype-specific and could be quantified in a trace-labeling experiment. Third, kidney-deposited immunoglobulins of mice injected with LPS were eluted, radiolabeled, and analyzed by radioimmunoassay. Both T15 idiotype-bearing immunoglobulins and anti-T15 idiotype antibodies were detected in the eluates, providing further evidence for a renal deposition of T15 idiotype-anti-T15 idiotype IC. Polyclonal B cell activation is likely to result in a simultaneous triggering of many idiotypic clones and of corresponding anti-idiotypic clones represented in the B cell repertoire. This could lead to the formation of a variety of idiotype-anti-idiotype IC that could participate in the development of glomerular lesions

  19. Pro-Apoptotic Activity of New Honokiol/Triphenylmethane Analogues in B-Cell Lymphoid Malignancies.

    Science.gov (United States)

    Mędra, Aleksandra; Witkowska, Magdalena; Majchrzak, Agata; Cebula-Obrzut, Barbara; Bonner, Michael Y; Robak, Tadeusz; Arbiser, Jack L; Smolewski, Piotr

    2016-01-01

    Honokiol and triphenylmethanes are small molecules with anti-tumor properties. Recently, we synthesized new honokiol analogues (HAs) that possess common features of both groups. We assessed the anti-tumor effectiveness of HAs in B-cell leukemia/lymphoma cells, namely in chronic lymphocytic leukemia (CLL) cells ex vivo and in pre-B-cell acute lymphoblastic leukemia (Nalm-6), Burkitt lymphoma (BL; Raji), diffuse large B-cell lymphoma (DLBCL; Toledo) and multiple myeloma (MM; RPMI 8226) cell lines. Four of these compounds appeared to be significantly active against the majority of cells examined, with no significant impact on healthy lymphocytes. These active HAs induced caspase-dependent apoptosis, causing significant deregulation of several apoptosis-regulating proteins. Overall, these compounds downregulated Bcl-2 and XIAP and upregulated Bax, Bak and survivin proteins. In conclusion, some of the HAs are potent tumor-selective inducers of apoptosis in ex vivo CLL and in BL, DLBCL and MM cells in vitro. Further preclinical studies of these agents are recommended. PMID:27483232

  20. Antigen Specificity of Clonally Expanded and Receptor Edited Cerebrospinal Fluid B cells from Patients with Relapsing Remitting MS

    OpenAIRE

    Lambracht-Washington, Doris; O’Connor, Kevin C.; Cameron, Elizabeth; Jowdry, Andrea; Ward, E. Sally; Frohman, Elliot; Racke, Michael K.; Monson, Nancy L.

    2007-01-01

    We re-engineered the immunoglobulin rearrangements from clonally expanded CSF B cells of three Multiple Sclerosis patients as Fab fragments, and used three methods to test for their Ag-specificity. Nine out of ten Fab fragments were reactive to Myelin Basic Protein (MBP). The one Fab that did not react to MBP was a product of receptor editing. Two of the nine MBP-reactive Fabs were also reactive to GFAP and CNPase, indicating that these clones were polyreactive. Targeting the mechanisms that ...

  1. Construction and characterization of an HCV-derived multi-epitope peptide antigen containing B-cell HVR1 mimotopes and T-cell conserved epitopes

    Institute of Scientific and Technical Information of China (English)

    GAO; Jun; GONG; Yuping; ZHAO; Ping; ZHU; Qing; YANG; Xiaoping; QI; Zhongtian

    2006-01-01

    Hepatitis C (HCV) genome is highly variable, particularly in the hypervariable region 1(HVR1) of its E2 envelope gene. The variability of HCV genome has been a major obstacle for developing HCV vaccines. Due to B-cell HVR1 mimotopes mimicking the antigenicity of natural HVR1 epitopes and some T-cell epitopes from the consensus sequence of HCV genes conserving among the different HCV genotypes, we synthesized an minigene of HCV-derived multi-epitope peptide antigen (CMEP), which contains 9 B-cell HVR1 mimotopes in E2, 2 conserved CTL epitopes in C, 1conserved CTL epitope in NS3 and 1 conserved Th epitope in NS3. This minigene was cloned into a GST expression vector to generate a fusion protein GST-CMER The immunogenic properties of CEMP were characterized by HCV infected patients' sera, and found that the reactivity frequency reached 75%. The cross reactivity of anti-CEMP antibody with different natural HVR1 variants was up to 90%. Meanwhile, we constructed an HCV DNA vaccine candidate, plasmid pVAX1.0-st-CMEP carrying the recombinant gene (st) of a secretion signal peptide and PADRE universal Th cell epitope sequence in front of the CMEP minigene. Immunization of rabbits with pVAX1.0-st-CMEP resulted in the production of antibody, which was of the same cross reactivity as the fusion protein GST-CMEP.Our findings indicate that the HCV-derived multi-epitope peptide antigen in some degree possessed the characteristics of neutralizing HCV epitopes, and would be of the value as a candidate for the development of HCV vaccines.

  2. Auto-transporter A protein of Neisseria meningitidis: a potent CD4+ T-cell and B-cell stimulating antigen detected by expression cloning.

    Science.gov (United States)

    Ait-Tahar, K; Wooldridge, K G; Turner, D P; Atta, M; Todd, I; Ala'Aldeen, D A

    2000-09-01

    A meningococcal genomic expression library was screened for potent CD4+ T-cell antigens, using patients' peripheral blood lymphocytes (PBLs). One of the most promising positive clones was fully characterized. The recombinant meningococcal DNA contained a single, incomplete, open reading frame (ORF), which was fully reconstructed with reference to available genomic sequence data. The gene was designated autA (auto-transporter A) as its peptide sequence shares molecular characteristics of the auto-transporter family of proteins. Only a single copy of this gene was detected in the meningococcal, and none in the gonococcal, genomic sequence databases. The complete autA gene, when cloned into an expression vector, expressed a protein of approximately 68 kDa. Purified rAutA recalled strong secondary T-cell responses in PBLs of patients and some healthy donors, and induced strong primary T-cell responses in healthy donors. The human B-cell immunogenicity and cross-reactivity of AutA, purified under native conditions, was confirmed in dot immunoblot experiments. Immunoblots with rabbit polyclonal antibodies to rAutA demonstrated the conserved nature, antigenicity and cross-reactivity of AutA amongst meningococci of different serogroups and strains representing different hypervirulent lineages. AutA showed homology with another meningococcal and gonococcal ORF (designated AutB). AutB was cloned and expressed and used to raise an autB-specific antiserum. Immunoblot experiments indicated that AutB is not expressed in meningococci and does not cross-react with AutA. Thus, AutA, being a potent CD4+ T-cell and B-cell-stimulating antigen, which is highly conserved, deserves further investigation as a potential vaccine candidate. PMID:10972828

  3. Murine retroviruses activate B cells via interaction with toll-like receptor 4

    Science.gov (United States)

    Rassa, John C.; Meyers, Jennifer L.; Zhang, Yuanming; Kudaravalli, Rama; Ross, Susan R.

    2002-01-01

    Although most retroviruses require activated cells as their targets for infection, it is not known how this is achieved in vivo. A candidate protein for the activation of B cells by either mouse mammary tumor virus (MMTV) or murine leukemia virus is the toll-like receptor 4 (TLR4), a component of the innate immune system. MMTV caused B cell activation in C3H/HeN mice but not in C3H/HeJ or BALB/c (C.C3H Tlr4lps-d) congenic mice, both of which have a mutant TLR4 gene. This activation was independent of viral gene expression, because it occurred after treatment of MMTV with ultraviolet light or 2,2′-dithiodipyridine and in azidothymidine-treated mice. Nuclear extracts prepared from the lymphocytes of MMTV-injected C3H/HeN but not C3H/HeJ mice showed increased nuclear factor κB activity. Additionally, the MMTV- and Moloney murine leukemia virus envelope proteins coimmunoprecipitated with TLR4 when expressed in 293T cells. The MMTV receptor failed to coimmunoprecipitate with TLR4, suggesting that MMTV/TLR4 interaction is independent of virus attachment and fusion. These results identify retroviral proteins that interact with a mammalian toll receptor and show that direct activation by such viruses may initiate in vivo infection pathways. PMID:11854525

  4. Overexpression of Fc receptor-like 1 associated with B-cell activation during hepatitis B virus infection

    International Nuclear Information System (INIS)

    The role of B cells in the pathogenesis of hepatitis B virus (HBV) infection has not been explored in depth. In the present study, the activation status of B cells from peripheral blood of healthy controls (N = 20) and patients with acute hepatitis B (AHB, N = 15) or chronic hepatitis B (CHB, N = 30) was evaluated by measuring the expression levels of B-cell activation markers CD69 and CD86, using quantitative real-time PCR and flow cytometry. Moreover, the potential mechanism underlying B-cell activation during HBV infection was further investigated by analyzing the expression profile of FCRL1, an intrinsic activation molecule of B cells. An elevation in the levels of B-cell activation markers including CD69 and CD86 was observed in the AHB patients (44.31 ± 9.27, 27.64 ± 9.26%) compared to CHB patients (30.35 ± 11.27, 18.41 ± 6.56%, P < 0.05), which was still higher than healthy controls (12.23 ± 7.84, 8.22 ± 3.43%, P < 0.05). Furthermore, the expression of FCRL1 was found to be similar to B-cell activation markers, which was highest in AHB patients (70.15 ± 17.11%), lowest in healthy donors (36.32 ± 9.98%, P < 0.05) and half-way between these levels in patients with CHB (55.17 ± 12.03%, P < 0.05). The results were positively associated with aberrant B-cell activation. These data suggest that B cells can play a role in HBV infection, and therefore more effort should be devoted to exploring their functions

  5. Overexpression of Fc receptor-like 1 associated with B-cell activation during hepatitis B virus infection

    Energy Technology Data Exchange (ETDEWEB)

    Wang, Ke [Key Laboratory of Nuclear Medicine, Ministry of Health, Jiangsu Key Laboratory of Molecular Nuclear Medicine, Jiangsu Institute of Nuclear Medicine, Wuxi, Jiangsu Province (China); Pei, Hao [Wuxi Hospital of Infectious Disease, Wuxi, Jiangsu Province (China); Huang, Biao; Yang, Run-Lin [Key Laboratory of Nuclear Medicine, Ministry of Health, Jiangsu Key Laboratory of Molecular Nuclear Medicine, Jiangsu Institute of Nuclear Medicine, Wuxi, Jiangsu Province (China); Wu, Hang-Yuan [Wuxi Hospital of Infectious Disease, Wuxi, Jiangsu Province (China); Zhu, Xue; Zhu, Lan [Key Laboratory of Nuclear Medicine, Ministry of Health, Jiangsu Key Laboratory of Molecular Nuclear Medicine, Jiangsu Institute of Nuclear Medicine, Wuxi, Jiangsu Province (China)

    2012-08-17

    The role of B cells in the pathogenesis of hepatitis B virus (HBV) infection has not been explored in depth. In the present study, the activation status of B cells from peripheral blood of healthy controls (N = 20) and patients with acute hepatitis B (AHB, N = 15) or chronic hepatitis B (CHB, N = 30) was evaluated by measuring the expression levels of B-cell activation markers CD69 and CD86, using quantitative real-time PCR and flow cytometry. Moreover, the potential mechanism underlying B-cell activation during HBV infection was further investigated by analyzing the expression profile of FCRL1, an intrinsic activation molecule of B cells. An elevation in the levels of B-cell activation markers including CD69 and CD86 was observed in the AHB patients (44.31 ± 9.27, 27.64 ± 9.26%) compared to CHB patients (30.35 ± 11.27, 18.41 ± 6.56%, P < 0.05), which was still higher than healthy controls (12.23 ± 7.84, 8.22 ± 3.43%, P < 0.05). Furthermore, the expression of FCRL1 was found to be similar to B-cell activation markers, which was highest in AHB patients (70.15 ± 17.11%), lowest in healthy donors (36.32 ± 9.98%, P < 0.05) and half-way between these levels in patients with CHB (55.17 ± 12.03%, P < 0.05). The results were positively associated with aberrant B-cell activation. These data suggest that B cells can play a role in HBV infection, and therefore more effort should be devoted to exploring their functions.

  6. BAFF promotes regulatory T-cell apoptosis and blocks cytokine production by activating B cells in primary biliary cirrhosis

    International Nuclear Information System (INIS)

    Primary biliary cirrhosis (PBC) is a chronic and slowly progressive cholestatic liver disease of autoimmune etiology. A number of questions regarding its etiology are unclear. CD4+CD25+ regulatory T cells (Tregs) play a critical role in self-tolerance and, for unknown reasons, their relative number is reduced in PBC patients. B-cell-activating factor (BAFF) is a key survival factor during B-cell maturation and its concentration is increased in peripheral blood of PBC patients. It has been reported that activated B cells inhibit Treg cell proliferation and there are no BAFF receptors on Tregs. Therefore, we speculated that excessive BAFF may result in Treg reduction via B cells. To prove our hypothesis, we isolated Tregs and B cells from PBC and healthy donors. BAFF and IgM concentrations were then analyzed by ELISA and CD40, CD80, CD86, IL-10, and TGF-β expression in B cells and Tregs were measured by flow cytometry. BAFF up-regulated CD40, CD80, CD86, and IgM expression in B cells. However, BAFF had no direct effect on Treg cell apoptosis and cytokine secretion. Nonetheless, we observed that BAFF-activated B cells could induce Treg cell apoptosis and reduce IL-10 and TGF-β expression. We also showed that BAFF-activated CD4+ T cells had no effect on Treg apoptosis. Furthermore, we verified that bezafibrate, a hypolipidemic drug, can inhibit BAFF-induced Treg cell apoptosis. In conclusion, BAFF promotes Treg cell apoptosis and inhibits cytokine production by activating B cells in PBC patients. The results of this study suggest that inhibition of BAFF activation is a strategy for PBC treatment

  7. Self DNA from lymphocytes that have undergone activation-induced cell death enhances murine B cell proliferation and antibody production.

    Directory of Open Access Journals (Sweden)

    Qing Lu

    Full Text Available Systemic lupus erythematosus (SLE is characterized by prominent autoinflammatory tissue damage associated with impaired removal of dying cells and DNA. Self DNA-containing immune complexes are able to activate both innate and adaptive immune responses and play an important role in the maintenance and exacerbation of autoimmunity in SLE. In this study, we used DNA from lymphocytes that have undergone activation-induced cell death (ALD-DNA and analyzed its role on the activation and differentiation of B cells from normal BALB/c mice as well as lupus-prone MRL+/+ and MRL/lpr mice. We found that ALD-DNA directly increased the expression of costimulatory molecules and the survival of naïve B cells in vitro. Although ALD-DNA alone had little effect on the proliferation of naïve B cells, it enhanced LPS-activated B cell proliferation in vitro and in vivo. In addition, ALD-DNA increased plasma cell numbers and IgG production in LPS-stimulated cultures of naïve B cells, in part via enhancing IL-6 production. Importantly, B cells from lupus mice were hyperresponsive to ALD-DNA and/or LPS relative to normal control B cells in terminal plasma cell differentiation, as evidenced by increases in CD138+ cell numbers, IgM production, and mRNA levels of B lymphocyte-induced maturation protein-1 (Blimp-1 and the X-box binding protein 1 (XBP1. Furthermore, ALD-DNA enhanced CD40-activated naïve B cell proliferation. Collectively, these data indicate that self DNA can serve as a DAMP (damage-associated molecular pattern that cooperates with signals from both innate and adaptive immunity to promote polyclonal B cell activation, a common characteristic of autoimmune diseases.

  8. BAFF promotes regulatory T-cell apoptosis and blocks cytokine production by activating B cells in primary biliary cirrhosis

    Directory of Open Access Journals (Sweden)

    Bo Zhang

    2013-10-01

    Full Text Available Primary biliary cirrhosis (PBC is a chronic and slowly progressive cholestatic liver disease of autoimmune etiology. A number of questions regarding its etiology are unclear. CD4+CD25+ regulatory T cells (Tregs play a critical role in self-tolerance and, for unknown reasons, their relative number is reduced in PBC patients. B-cell-activating factor (BAFF is a key survival factor during B-cell maturation and its concentration is increased in peripheral blood of PBC patients. It has been reported that activated B cells inhibit Treg cell proliferation and there are no BAFF receptors on Tregs. Therefore, we speculated that excessive BAFF may result in Treg reduction via B cells. To prove our hypothesis, we isolated Tregs and B cells from PBC and healthy donors. BAFF and IgM concentrations were then analyzed by ELISA and CD40, CD80, CD86, IL-10, and TGF-β expression in B cells and Tregs were measured by flow cytometry. BAFF up-regulated CD40, CD80, CD86, and IgM expression in B cells. However, BAFF had no direct effect on Treg cell apoptosis and cytokine secretion. Nonetheless, we observed that BAFF-activated B cells could induce Treg cell apoptosis and reduce IL-10 and TGF-β expression. We also showed that BAFF-activated CD4+ T cells had no effect on Treg apoptosis. Furthermore, we verified that bezafibrate, a hypolipidemic drug, can inhibit BAFF-induced Treg cell apoptosis. In conclusion, BAFF promotes Treg cell apoptosis and inhibits cytokine production by activating B cells in PBC patients. The results of this study suggest that inhibition of BAFF activation is a strategy for PBC treatment.

  9. BAFF promotes regulatory T-cell apoptosis and blocks cytokine production by activating B cells in primary biliary cirrhosis

    Energy Technology Data Exchange (ETDEWEB)

    Zhang, Bo; Hu, Mintao [Department of Hepatology, Wuxi Infectious Diseases Hospital, Wuxi, Jiangsu (China); Zhang, Peng [Nanjing Medical University, Nanjing, Jiangsu (China); Cao, Hong [Department of Hepatology, Wuxi Infectious Diseases Hospital, Wuxi, Jiangsu (China); Wang, Yongzhen [The Second Hospital of Nanjing, Nanjing, Jiangsu (China); Wang, Zheng; Su, Tingting [Department of Hepatology, Wuxi Infectious Diseases Hospital, Wuxi, Jiangsu (China)

    2013-05-10

    Primary biliary cirrhosis (PBC) is a chronic and slowly progressive cholestatic liver disease of autoimmune etiology. A number of questions regarding its etiology are unclear. CD4+CD25+ regulatory T cells (Tregs) play a critical role in self-tolerance and, for unknown reasons, their relative number is reduced in PBC patients. B-cell-activating factor (BAFF) is a key survival factor during B-cell maturation and its concentration is increased in peripheral blood of PBC patients. It has been reported that activated B cells inhibit Treg cell proliferation and there are no BAFF receptors on Tregs. Therefore, we speculated that excessive BAFF may result in Treg reduction via B cells. To prove our hypothesis, we isolated Tregs and B cells from PBC and healthy donors. BAFF and IgM concentrations were then analyzed by ELISA and CD40, CD80, CD86, IL-10, and TGF-β expression in B cells and Tregs were measured by flow cytometry. BAFF up-regulated CD40, CD80, CD86, and IgM expression in B cells. However, BAFF had no direct effect on Treg cell apoptosis and cytokine secretion. Nonetheless, we observed that BAFF-activated B cells could induce Treg cell apoptosis and reduce IL-10 and TGF-β expression. We also showed that BAFF-activated CD4+ T cells had no effect on Treg apoptosis. Furthermore, we verified that bezafibrate, a hypolipidemic drug, can inhibit BAFF-induced Treg cell apoptosis. In conclusion, BAFF promotes Treg cell apoptosis and inhibits cytokine production by activating B cells in PBC patients. The results of this study suggest that inhibition of BAFF activation is a strategy for PBC treatment.

  10. BAFF promotes regulatory T-cell apoptosis and blocks cytokine production by activating B cells in primary biliary cirrhosis

    Directory of Open Access Journals (Sweden)

    Bo Zhang

    2013-05-01

    Full Text Available Primary biliary cirrhosis (PBC is a chronic and slowly progressive cholestatic liver disease of autoimmune etiology. A number of questions regarding its etiology are unclear. CD4+CD25+ regulatory T cells (Tregs play a critical role in self-tolerance and, for unknown reasons, their relative number is reduced in PBC patients. B-cell-activating factor (BAFF is a key survival factor during B-cell maturation and its concentration is increased in peripheral blood of PBC patients. It has been reported that activated B cells inhibit Treg cell proliferation and there are no BAFF receptors on Tregs. Therefore, we speculated that excessive BAFF may result in Treg reduction via B cells. To prove our hypothesis, we isolated Tregs and B cells from PBC and healthy donors. BAFF and IgM concentrations were then analyzed by ELISA and CD40, CD80, CD86, IL-10, and TGF-β expression in B cells and Tregs were measured by flow cytometry. BAFF up-regulated CD40, CD80, CD86, and IgM expression in B cells. However, BAFF had no direct effect on Treg cell apoptosis and cytokine secretion. Nonetheless, we observed that BAFF-activated B cells could induce Treg cell apoptosis and reduce IL-10 and TGF-β expression. We also showed that BAFF-activated CD4+ T cells had no effect on Treg apoptosis. Furthermore, we verified that bezafibrate, a hypolipidemic drug, can inhibit BAFF-induced Treg cell apoptosis. In conclusion, BAFF promotes Treg cell apoptosis and inhibits cytokine production by activating B cells in PBC patients. The results of this study suggest that inhibition of BAFF activation is a strategy for PBC treatment.

  11. Global gene expression changes of in vitro stimulated human transformed germinal centre B cells as surrogate for oncogenic pathway activation in individual aggressive B cell lymphomas

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    Schrader Alexandra

    2012-12-01

    Full Text Available Abstract Background Aggressive Non-Hodgkin lymphomas (NHL are a group of lymphomas derived from germinal centre B cells which display a heterogeneous pattern of oncogenic pathway activation. We postulate that specific immune response associated signalling, affecting gene transcription networks, may be associated with the activation of different oncogenic pathways in aggressive Non-Hodgkin lymphomas (NHL. Methodology The B cell receptor (BCR, CD40, B-cell activating factor (BAFF-receptors and Interleukin (IL 21 receptor and Toll like receptor 4 (TLR4 were stimulated in human transformed germinal centre B cells by treatment with anti IgM F(ab2-fragments, CD40L, BAFF, IL21 and LPS respectively. The changes in gene expression following the activation of Jak/STAT, NF-кB, MAPK, Ca2+ and PI3K signalling triggered by these stimuli was assessed using microarray analysis. The expression of top 100 genes which had a change in gene expression following stimulation was investigated in gene expression profiles of patients with Aggressive non-Hodgkin Lymphoma (NHL. Results αIgM stimulation led to the largest number of changes in gene expression, affecting overall 6596 genes. While CD40L stimulation changed the expression of 1194 genes and IL21 stimulation affected 902 genes, only 283 and 129 genes were modulated by lipopolysaccharide or BAFF receptor stimulation, respectively. Interestingly, genes associated with a Burkitt-like phenotype, such as MYC, BCL6 or LEF1, were affected by αIgM. Unique and shared gene expression was delineated. NHL-patients were sorted according to their similarity in the expression of TOP100 affected genes to stimulated transformed germinal centre B cells The αIgM gene module discriminated individual DLBCL in a similar manner to CD40L or IL21 gene modules. DLBCLs with low module activation often carry chromosomal MYC aberrations. DLBCLs with high module activation show strong expression of genes involved in cell

  12. Serologic markers of effective tumor immunity against chronic lymphocytic leukemia include non-mutated B cell antigens

    OpenAIRE

    Marina, Ovidiu; Hainz, Ursula; Biernacki, Melinda A.; Zhang, Wandi; Cai, Ann; Duke-Cohan, Jonathan S.; Liu, Fenglong; Brusic, Vladimir; Neuberg, Donna; Kutok, Jeffery L.; Alyea, Edwin P.; Canning, Christine M.; Soiffer, Robert J.; Ritz, Jerome; Wu, Catherine J.

    2010-01-01

    Patients with chronic lymphocytic leukemia (CLL) who relapse after allogeneic transplant may achieve durable remission following donor lymphocytes infusion (DLI), demonstrating the potency of donor-derived immunity in eradicating tumor. We sought to elucidate the antigenic basis of the effective graft-versus-leukemia (GvL) responses associated with DLI for the treatment of CLL by analyzing the specificity of plasma antibody responses developing in two DLI-treated patients who achieved long-te...

  13. B-cell activating factor in the pathophysiology of multiple myeloma: a target for therapy?

    International Nuclear Information System (INIS)

    Multiple myeloma (MM) is a currently incurable malignancy of plasma cells. Malignant myeloma cells (MMCs) are heavily dependent upon the bone marrow (BM) microenvironment for their survival. One component of this tumor microenvironment, B-Cell Activating Factor (BAFF), has been implicated as a key player in this interaction. This review discusses the role of BAFF in the pathophysiology of MM, and the potential of BAFF-inhibitory therapy for the treatment of MM. Multiple studies have shown that BAFF functions as a survival factor for MMCs. Furthermore, MMCs express several BAFF-binding receptors. Of these, only Transmembrane Activator and CAML Interactor (TACI) correlates with the MMC's capability to ligate BAFF. Additionally, the level of expression of TACI correlates with the level of the MMC's BM dependency. Ligation of BAFF receptors on MMCs causes activation of the Nuclear Factor of κ-B (NF-κB) pathway, a crucial pathway for the pathogenesis of many B-cell malignancies. Serum BAFF levels are significantly elevated in MM patients when compared to healthy controls, and correlate inversely with overall survival. BAFF signaling is thus an interesting target for the treatment of MM. Several BAFF-inhibitory drugs are currently under evaluation for the treatment of MM. These include BAFF-monoclonal antibodies (tabalumab) and antibody-drug conjugates (GSK2857916)

  14. Outer membrane protein A (OmpA of Shigella flexneri 2a induces TLR2-mediated activation of B cells: involvement of protein tyrosine kinase, ERK and NF-κB.

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    Rajsekhar Bhowmick

    Full Text Available B cells are critically important in combating bacterial infections and their differentiation into plasma cells and memory cells aids bacterial clearance and long-lasting immunity conferred by essentially all vaccines. Outer membrane protein A (OmpA of Shigella flexneri 2a has been demonstrated to induce the production of IgG and IgA in vivo following immunization of mice through intranasal route, but the direct involvement of B cells in OmpA-mediated immune regulation was not determined. Consequently, we investigated whether OmpA can modulate B cell functions and identified the molecular events involved in OmpA-induced B cell immune response in vitro. We show that OmpA of S. flexneri 2a activates B cells to produce protective cytokines, IL-6 and IL-10 as well as facilitates their differentiation into antibody secreting cells (ASCs. The immunostimulatory properties of OmpA are attributed to the increased surface expression of MHCII and CD86 on B cells. We also report here that B cell activation by OmpA is mediated strictly through recognition by TLR2, resulting in initiation of cascades of signal transduction events, involving increased phosphorylation of protein tyrosine kinases (PTKs, ERK and IκBα, leading to nuclear translocation of NF-κB. Importantly, a TLR2 antibody diminishes OmpA-induced upregulation of MHCII and CD86 on B cell surface as well as significantly inhibits B cell differentiation and cytokine secretion. Furthermore, we illustrate that B cell differentiation into ASCs and induction of cytokine secretion by OmpA are dependent on PTKs activity. Moreover, we identify that OmpA-induced B cell differentiation is entirely dependent on ERK pathway, whereas both NF-κB and ERK are essential for cytokine secretion by B cells. Overall, our data demonstrate that OmpA of S. flexneri 2a amplifies TLR signaling in B cells and triggers B cell immune response, which is critical for the development of an effective adaptive immunity to an

  15. Removing N-terminal sequences in pre-S1 domain enhanced antibody and B-cell responses by an HBV large surface antigen DNA vaccine.

    Science.gov (United States)

    Ge, Guohong; Wang, Shixia; Han, Yaping; Zhang, Chunhua; Lu, Shan; Huang, Zuhu

    2012-01-01

    Although the use of recombinant hepatitis B virus surface (HBsAg) protein vaccine has successfully reduced global hepatitis B infection, there are still a number of vaccine recipients who do not develop detectable antibody responses. Various novel vaccination approaches, including DNA vaccines, have been used to further improve the coverage of vaccine protection. Our previous studies demonstrated that HBsAg-based DNA vaccines could induce both humoral and CMI responses in experimental animal models. However, one form of the the HBsAg antigen, the large S antigen (HBs-L), expressed by DNA vaccine, was not sufficiently immunogenic in eliciting antibody responses. In the current study, we produced a modified large S antigen DNA vaccine, HBs-L(T), which has a truncated N-terminal sequence in the pre-S1 region. Compared to the original HBs-L DNA vaccine, the HBs-L(T) DNA vaccine improved secretion in cultured mammalian cells and generated significantly enhanced HBsAg-specific antibody and B cell responses. Furthermore, this improved HBsL DNA vaccine, along with other HBsAg-expressing DNA vaccines, was able to maintain predominantly Th1 type antibody responses while recombinant HBsAg protein vaccines produced in either yeast or CHO cells elicited mostly Th2 type antibody responses. Our data indicate that HBsAg DNA vaccines with improved immunogenicity offer a useful alternative choice to recombinant protein-based HBV vaccines, particularly for therapeutic purposes against chronic hepatitis infection where immune tolerance led to poor antibody responses to S antigens. PMID:22844502

  16. Removing N-terminal sequences in pre-S1 domain enhanced antibody and B-cell responses by an HBV large surface antigen DNA vaccine.

    Directory of Open Access Journals (Sweden)

    Guohong Ge

    Full Text Available Although the use of recombinant hepatitis B virus surface (HBsAg protein vaccine has successfully reduced global hepatitis B infection, there are still a number of vaccine recipients who do not develop detectable antibody responses. Various novel vaccination approaches, including DNA vaccines, have been used to further improve the coverage of vaccine protection. Our previous studies demonstrated that HBsAg-based DNA vaccines could induce both humoral and CMI responses in experimental animal models. However, one form of the the HBsAg antigen, the large S antigen (HBs-L, expressed by DNA vaccine, was not sufficiently immunogenic in eliciting antibody responses. In the current study, we produced a modified large S antigen DNA vaccine, HBs-L(T, which has a truncated N-terminal sequence in the pre-S1 region. Compared to the original HBs-L DNA vaccine, the HBs-L(T DNA vaccine improved secretion in cultured mammalian cells and generated significantly enhanced HBsAg-specific antibody and B cell responses. Furthermore, this improved HBsL DNA vaccine, along with other HBsAg-expressing DNA vaccines, was able to maintain predominantly Th1 type antibody responses while recombinant HBsAg protein vaccines produced in either yeast or CHO cells elicited mostly Th2 type antibody responses. Our data indicate that HBsAg DNA vaccines with improved immunogenicity offer a useful alternative choice to recombinant protein-based HBV vaccines, particularly for therapeutic purposes against chronic hepatitis infection where immune tolerance led to poor antibody responses to S antigens.

  17. Absence of missense mutations in activated c-myc genes in avian leukosis virus-induced B-cell lymphomas

    International Nuclear Information System (INIS)

    The authors determined the nucleotide sequences of two independent DNA clones which contained the activated c-myc genes from avian leukosis virus-induced B-cell lymphomas. Neither of these c-myce genes contained missense mutations. This strongly supports the notion that the c-myc photo-oncogene in avian leukosis virus-induced B-cell lymphomas can be oncogenically activated by altered expression of the gene without a change in the primary structure of the gene product

  18. Absence of missense mutations in activated c-myc genes in avian leukosis virus-induced B-cell lymphomas.

    OpenAIRE

    Hahn, M; Hayward, W S

    1988-01-01

    We have determined the nucleotide sequences of two independent DNA clones which contained the activated c-myc genes from avian leukosis virus-induced B-cell lymphomas. Neither of these c-myc genes contained missense mutations. This strongly supports the notion that the c-myc proto-oncogene in avian leukosis virus-induced B-cell lymphomas can be oncogenically activated by altered expression of the gene without a change in the primary structure of the gene product.

  19. Absence of missense mutations in activated c-myc genes in avian leukosis virus-induced B-cell lymphomas

    Energy Technology Data Exchange (ETDEWEB)

    Hahn, M.; Hayward, W.S.

    1988-06-01

    The authors determined the nucleotide sequences of two independent DNA clones which contained the activated c-myc genes from avian leukosis virus-induced B-cell lymphomas. Neither of these c-myce genes contained missense mutations. This strongly supports the notion that the c-myc photo-oncogene in avian leukosis virus-induced B-cell lymphomas can be oncogenically activated by altered expression of the gene without a change in the primary structure of the gene product.

  20. Soluble suppressor supernatants elaborated by concanavalin A-activated human mononuclear cells. Characterization of a soluble suppressor of B cell immunoglobulin production

    International Nuclear Information System (INIS)

    Human peripheral blood mononuclear cells (PBMC) activated with the mitogenic lectin concanavalin A (Con A) elaborate a soluble immune suppressor supernatant (SISS) that contains at least 2 distinct suppressor factors. One of these, SISS-B, inhibits polyclonal B cell immunoglobulin production, whereas the other, SISS-T, suppresses T cell proliferation to both mitogens and antigens. The latter mediator is discussed in the companion paper. Characteristics of the human soluble suppressor of B cell immunoglobulin production (SISS-B) include: 1) inhibition by a noncytotoxic mechanism, 2) loss of activity in the presence of the monosaccharide L-rhamnose, 3) appearance within 8 to 16 hr after the addition of Con A, 4) elaboration by cells irradiated with 500 or 2000 rads, 5) production by highly purified T cells, 6) stability at pH 2.5 but instability at 56/sup o/C, and 7) m.w. of 60 to 80,000. These data indicate that after Con A activation, selected T cells not only become potent suppressor cells, but also generate a soluble saccharide-specific factor(s) that inhibits polyclonal immunoglobulin production by human B cells

  1. ICAM-1-based rabies virus vaccine shows increased infection and activation of primary murine B cells in vitro and enhanced antibody titers in-vivo.

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    James E Norton

    Full Text Available We have previously shown that live-attenuated rabies virus (RABV-based vaccines infect and directly activate murine and human primary B cells in-vitro, which we propose can be exploited to help develop a single-dose RABV-based vaccine. Here we report on a novel approach to utilize the binding of Intracellular Adhesion Molecule-1 (ICAM-1 to its binding partner, Lymphocyte Function-associated Antigen-1 (LFA-1, on B cells to enhance B cell activation and RABV-specific antibody responses. We used a reverse genetics approach to clone, recover, and characterize a live-attenuated recombinant RABV-based vaccine expressing the murine Icam1 gene (rRABV-mICAM-1. We show that the murine ICAM-1 gene product is incorporated into virus particles, potentially exposing ICAM-1 to extracellular binding partners. While rRABV-mICAM-1 showed 10-100-fold decrease in viral titers on baby hamster kidney cells compared to the parental virus (rRABV, rRABV-mICAM-1 infected and activated primary murine B cells in-vitro more efficiently than rRABV, as indicated by significant upregulation of CD69, CD40, and MHCII on the surface of infected B cells. ICAM-1 expression on the virus surface was responsible for enhanced B cell infection since pre-treating rRABV-mICAM-1 with a neutralizing anti-ICAM-1 antibody reduced B cell infection to levels observed with rRABV alone. Furthermore, 100-fold less rRABV-mICAM-1 was needed to induce antibody titers in immunized mice equivalent to antibody titers observed in rRABV-immunized mice. Of note, only 10(3 focus forming units (ffu/mouse of rRABV-mICAM-1 was needed to induce significant anti-RABV antibody titers as early as five days post-immunization. As both speed and potency of antibody responses are important in controlling human RABV infection in a post-exposure setting, these data show that expression of Icam1 from the RABV genome, which is then incorporated into the virus particle, is a promising strategy for the development of a

  2. Gamma delta T cells recognize a microbial encoded B Cell antigen to initiate a rapid antigen-specific Interleukin-17 response

    Science.gov (United States)

    Gamma delta T cells contribute uniquely to host immune defense, but the way in which they do so remains an enigma. Here we show that an algae protein, phycoerythrin (PE) is recognized by gamma delta T cells from mice, bovine and humans and binds directly to specific gamma delta T cell antigen recept...

  3. 1,25-dihydroxyvitamin D3 impairs NF-κB activation in human naive B cells

    International Nuclear Information System (INIS)

    Highlights: → In naive B cells, VDR activation by calcitriol results in reduced NF-κB p105 and p50 protein expression. → Ligating the VDR with calcitriol causes reduced nuclear translocation of NF-κB p65. → Reduced nuclear amount of p65 after calcitriol incubation results in reduced binding of p65 on the p105 promoter. → Thus, vitamin D receptor signaling may reduce or prevent activation of B cells and unwanted immune responses, e.g. in IgE dependent diseases such as allergic asthma. -- Abstract: 1α,25-dihydroxyvitamin D3 (calcitriol), the bioactive metabolite of vitamin D, modulates the activation and inhibits IgE production of anti-CD40 and IL-4 stimulated human peripheral B cells. Engagement of CD40 results in NF-κB p50 activation, which is essential for the class switch to IgE. Herein, we investigated by which mechanism calcitriol modulates NF-κB mediated activation of human naive B cells. Naive B cells were predominantly targeted by calcitriol in comparison with memory B cells as shown by pronounced induction of the VDR target gene cyp24a1. Vitamin D receptor activation resulted in a strongly reduced p105/p50 protein and mRNA expression in human naive B cells. This effect is mediated by impaired nuclear translocation of p65 and consequently reduced binding of p65 to its binding site in the p105 promoter. Our data indicate that the vitamin D receptor reduces NF-κB activation by interference with NF-κB p65 and p105. Thus, the vitamin D receptor inhibits costimulatory signal transduction in naive B cells, namely by reducing CD40 signaling.

  4. 1,25-dihydroxyvitamin D{sub 3} impairs NF-{kappa}B activation in human naive B cells

    Energy Technology Data Exchange (ETDEWEB)

    Geldmeyer-Hilt, Kerstin, E-mail: kerstin.hilt@charite.de [Allergie-Centrum-Charite, CCM, Klinik fuer Dermatologie und Allergologie, Charite - Universitaetsmedizin Berlin, Chariteplatz 1, 10117 Berlin (Germany); Heine, Guido, E-mail: guido.heine@charite.de [Allergie-Centrum-Charite, CCM, Klinik fuer Dermatologie und Allergologie, Charite - Universitaetsmedizin Berlin, Chariteplatz 1, 10117 Berlin (Germany); Deutsches Rheuma-Forschungszentrum Berlin, Chariteplatz 1, 10117 Berlin (Germany); Hartmann, Bjoern, E-mail: bjoern.hartmann@charite.de [Allergie-Centrum-Charite, CCM, Klinik fuer Dermatologie und Allergologie, Charite - Universitaetsmedizin Berlin, Chariteplatz 1, 10117 Berlin (Germany); Baumgrass, Ria, E-mail: baumgrass@drfz.de [Deutsches Rheuma-Forschungszentrum Berlin, Chariteplatz 1, 10117 Berlin (Germany); Radbruch, Andreas, E-mail: radbruch@drfz.de [Deutsches Rheuma-Forschungszentrum Berlin, Chariteplatz 1, 10117 Berlin (Germany); Worm, Margitta, E-mail: margitta.worm@charite.de [Allergie-Centrum-Charite, CCM, Klinik fuer Dermatologie und Allergologie, Charite - Universitaetsmedizin Berlin, Chariteplatz 1, 10117 Berlin (Germany)

    2011-04-22

    Highlights: {yields} In naive B cells, VDR activation by calcitriol results in reduced NF-{kappa}B p105 and p50 protein expression. {yields} Ligating the VDR with calcitriol causes reduced nuclear translocation of NF-{kappa}B p65. {yields} Reduced nuclear amount of p65 after calcitriol incubation results in reduced binding of p65 on the p105 promoter. {yields} Thus, vitamin D receptor signaling may reduce or prevent activation of B cells and unwanted immune responses, e.g. in IgE dependent diseases such as allergic asthma. -- Abstract: 1{alpha},25-dihydroxyvitamin D{sub 3} (calcitriol), the bioactive metabolite of vitamin D, modulates the activation and inhibits IgE production of anti-CD40 and IL-4 stimulated human peripheral B cells. Engagement of CD40 results in NF-{kappa}B p50 activation, which is essential for the class switch to IgE. Herein, we investigated by which mechanism calcitriol modulates NF-{kappa}B mediated activation of human naive B cells. Naive B cells were predominantly targeted by calcitriol in comparison with memory B cells as shown by pronounced induction of the VDR target gene cyp24a1. Vitamin D receptor activation resulted in a strongly reduced p105/p50 protein and mRNA expression in human naive B cells. This effect is mediated by impaired nuclear translocation of p65 and consequently reduced binding of p65 to its binding site in the p105 promoter. Our data indicate that the vitamin D receptor reduces NF-{kappa}B activation by interference with NF-{kappa}B p65 and p105. Thus, the vitamin D receptor inhibits costimulatory signal transduction in naive B cells, namely by reducing CD40 signaling.

  5. Radioprotective activity of shigella antigens

    International Nuclear Information System (INIS)

    The possibility of using experimental microbe antigenous preparation out of Flexner and Zonne shigellas as a protector and a remedy in the case of gamma irradiation, is investigated. The experiments are carried out on mice of both sexes immunized before or after irradiation by two methods: subcutaneously and enerally. It is found that in most cases investigated, the introduction of the experimental preparation 3, 5, 7 and 10 days before irradiation increases the survivability of animals

  6. Retrotransposon derepression leads to activation of the unfolded protein response and apoptosis in pro-B cells.

    Science.gov (United States)

    Pasquarella, Alessandra; Ebert, Anja; Pereira de Almeida, Gustavo; Hinterberger, Maria; Kazerani, Maryam; Nuber, Alexander; Ellwart, Joachim; Klein, Ludger; Busslinger, Meinrad; Schotta, Gunnar

    2016-05-15

    The H3K9me3-specific histone methyltransferase Setdb1 impacts on transcriptional regulation by repressing both developmental genes and retrotransposons. How impaired retrotransposon silencing may lead to developmental phenotypes is currently unclear. Here, we show that loss of Setdb1 in pro-B cells completely abrogates B cell development. In pro-B cells, Setdb1 is dispensable for silencing of lineage-inappropriate developmental genes. Instead, we detect strong derepression of endogenous murine leukemia virus (MLV) copies. This activation coincides with an unusual change in chromatin structure, with only partial loss of H3K9me3 and unchanged DNA methylation, but strongly increased H3K4me3. Production of MLV proteins leads to activation of the unfolded protein response pathway and apoptosis. Thus, our data demonstrate that B cell development depends on the proper repression of retrotransposon sequences through Setdb1. PMID:27013243

  7. Sulfhydryl-based tumor antigen-carrier protein conjugates stimulate superior antitumor immunity against B cell lymphomas.

    Science.gov (United States)

    Betting, David J; Kafi, Kamran; Abdollahi-Fard, Alireza; Hurvitz, Sara A; Timmerman, John M

    2008-09-15

    Therapeutic vaccination of B cell lymphoma patients with tumor-specific Ig (idiotype, or Id) chemically coupled to the immunogenic foreign carrier protein keyhole limpet hemocyanin (KLH) using glutaraldehyde has shown promising results in early clinical trials, and phase III trials are underway. However, glutaraldehyde Id-KLH vaccines fail to elicit anti-Id immune and clinical responses in many patients, possibly because glutaraldehyde reacts with lysine, cysteine, tyrosine, and histidine residues, damaging critical immunogenic epitopes. A sulfhydryl-based tumor Ag-carrier protein conjugation system using maleimide chemistry was used to enhance the efficacy of Id-KLH vaccines. Maleimide Id-KLH conjugates eradicated A20 lymphoma from most tumor-bearing mice, whereas glutaraldehyde Id-KLH had little efficacy. Maleimide Id-KLH elicited tumor-specific IgG Abs and T cells, with CD8(+) T cells being the major effectors of antilymphoma immunity. Maleimide Id-KLH vaccines also demonstrated superior efficacy in 38C13 and BCL-1 lymphoma models, where Abs were shown to be critical for protection. Importantly, standard glutaraldehyde Id-KLH conjugation procedures could result in "overconjugation" of the tumor Ag, leading to decreased efficacy, whereas the heterobifunctional maleimide-based conjugation yielded potent vaccine product regardless of conjugation duration. Under lysosomal processing conditions, the Id-carrier protein linkage was cleavable only after maleimide conjugation. Maleimide KLH conjugation was easily performed with human Igs analogous to those used in Id-KLH clinical trials. These data support the evaluation of sulfhydryl-based Id-KLH vaccines in lymphoma clinical trials and possibly the use of tumor Ag-carrier protein vaccines for other cancers. PMID:18768870

  8. Intratracheal administration of fullerene nanoparticles activates splenic CD11b+ cells

    International Nuclear Information System (INIS)

    Highlights: → Fullerene administration triggered splenic responses. → Splenic responses occurred at different time-points than in the lung tissue. → CD11b+ cells were demonstrated to function as responder cells to fullerene. - Abstract: Fullerene nanoparticles ('Fullerenes'), which are now widely used materials in daily life, have been demonstrated to induce elevated pulmonary inflammation in several animal models; however, the effects of fullerenes on the immune system are not fully understood. In the present study, mice received fullerenes intratracheally and were sacrificed at days 1, 6 and 42. Mice that received fullerenes exhibited increased proliferation of splenocytes and increased splenic production of IL-2 and TNF-α. Changes in the spleen in response to fullerene treatment occurred at different time-points than in the lung tissue. Furthermore, fullerenes induced CDK2 expression and activated NF-κB and NFAT in splenocytes at 6 days post-administration. Finally, CD11b+ cells were demonstrated to function as responder cells to fullerene administration in the splenic inflammatory process. Taken together, in addition to the effects on pulmonary responses, fullerenes also modulate the immune system.

  9. Specifically modified Env immunogens activate B-cell precursors of broadly neutralizing HIV-1 antibodies in transgenic mice.

    Science.gov (United States)

    McGuire, Andrew T; Gray, Matthew D; Dosenovic, Pia; Gitlin, Alexander D; Freund, Natalia T; Petersen, John; Correnti, Colin; Johnsen, William; Kegel, Robert; Stuart, Andrew B; Glenn, Jolene; Seaman, Michael S; Schief, William R; Strong, Roland K; Nussenzweig, Michel C; Stamatatos, Leonidas

    2016-01-01

    VRC01-class broadly neutralizing HIV-1 antibodies protect animals from experimental infection and could contribute to an effective vaccine response. Their predicted germline forms (gl) bind Env inefficiently, which may explain why they are not elicited by HIV-1 Env-immunization. Here we show that an optimized Env immunogen can engage multiple glVRC01-class antibodies. Furthermore, this immunogen activates naive B cells expressing the human germline heavy chain of 3BNC60, paired with endogenous mouse light chains in vivo. To address whether it activates B cells expressing the fully humanized gl3BNC60 B-cell receptor (BCR), we immunized mice carrying both the heavy and light chains of gl3BNC60. B cells expressing this BCR display an autoreactive phenotype and fail to respond efficiently to soluble forms of the optimized immunogen, unless it is highly multimerized. Thus, specifically designed Env immunogens can activate naive B cells expressing human BCRs corresponding to precursors of broadly neutralizing HIV-1 antibodies even when the B cells display an autoreactive phenotype. PMID:26907590

  10. Regulation of germinal center B-cell differentiation.

    Science.gov (United States)

    Zhang, Yang; Garcia-Ibanez, Laura; Toellner, Kai-Michael

    2016-03-01

    Germinal centers (GC) are the main sites where antigen-activated B-cell clones expand and undergo immunoglobulin gene hypermutation and selection. Iterations of this process will lead to affinity maturation, replicating Darwinian evolution on the cellular level. GC B-cell selection can lead to four different outcomes: further expansion and evolution, apoptosis (non-selection), or output from the GC with differentiation into memory B cells or plasma cells. T-helper cells in GC have been shown to have a central role in regulating B-cell selection by sensing the density of major histocompatibility complex (MHC):peptide antigen complexes. Antigen is provided on follicular dendritic cells in the form of immune complex. Antibody on these immune complexes regulates antigen accessibility by shielding antigen from B-cell receptor access. Replacement of antibody on immune complexes by antibody generated from GC-derived plasma cell output will gradually reduce the availability of antigen. This antibody feedback can lead to a situation where a slow rise in selection stringency caused by a changing environment leads to directional evolution toward higher affinity antibody. PMID:26864101

  11. CNS accumulation of regulatory B cells is VLA-4-dependent

    Science.gov (United States)

    Lehmann-Horn, Klaus; Sagan, Sharon A.; Winger, Ryan C.; Spencer, Collin M.; Bernard, Claude C.A.; Sobel, Raymond A.

    2016-01-01

    Objective: To investigate the role of very late antigen-4 (VLA-4) on regulatory B cells (Breg) in CNS autoimmune disease. Methods: Experimental autoimmune encephalomyelitis (EAE) was induced in mice selectively deficient for VLA-4 on B cells (CD19cre/α4f/f) by immunization with myelin oligodendrocyte glycoprotein (MOG) peptide (p)35–55 or recombinant human (rh) MOG protein. B-cell and T-cell populations were examined by flow cytometry and immunohistochemistry. Breg were evaluated by intracellular IL-10 staining of B cells and, secondly, by coexpression of CD1d and CD5. Results: As previously reported, EAE was less severe in B-cell VLA-4-deficient vs control CD19cre mice when induced by rhMOG, a model that is B-cell-dependent and leads to efficient B-cell activation and antibody production. Paradoxically, B-cell VLA-4-deficient mice developed more severe clinical disease than control mice when EAE was induced with MOG p35-55, a B-cell-independent encephalitogen that does not efficiently activate B cells. Peripheral T-cell and humoral immune responses were not altered in B-cell VLA-4-deficient mice. In MOG p35-55-induced EAE, B-cell VLA-4 deficiency reduced CNS accumulation of B but not T cells. Breg were detected in the CNS of control mice with MOG p35-55-induced EAE. However, more severe EAE in B-cell VLA-4-deficient mice was associated with virtual absence of CNS Breg. Conclusions: Our results demonstrate that CNS accumulation of Breg is VLA-4-dependent and suggest that Breg may contribute to regulation of CNS autoimmunity in situ. These observations underscore the need to choose the appropriate encephalitogen when studying how B cells contribute to pathogenesis or regulation of CNS autoimmunity. PMID:27027096

  12. Recombinant human interleukin 2 acts as a B cell growth and differentiation promoting factor

    OpenAIRE

    Emmrich, F.; Moll, Heidrun; Markus M Simon

    2009-01-01

    Human B cells appropriately activated by a B cell mitogen are rendered susceptible to human Interleukin 2 (IL-2) as demonstrated with recombinant human IL-2 (rec. h IL-2). They show increased proliferation and drastically enhanced immunoglobulin secretion. Susceptibility to IL-2 is accompanied with the expression of the IL-2 receptor (Tac antigen) on B cells. The data suggest that IL-2 is one of the lymphokines directly involved in the activation of B lymphocytes.

  13. Dominant B-cell epitopes from cancer/stem cell antigen SOX2 recognized by serum samples from cancer patients

    OpenAIRE

    Shih, Julia; Rahman, Munira; Luong, Quang T; Lomeli, Shirley H.; Riss, Joseph; Prins, Robert M.; Gure, Ali O.; Zeng, Gang

    2014-01-01

    Human sex determining region Y-box 2 (SOX2) is an important transcriptional factor involved in the pluripotency and stemness of human embryonic stem cells. SOX2 plays important roles in maintaining cancer stem cell activities of melanoma and cancers of the brain, prostate, breast, and lung. SOX2 is also a lineage survival oncogene for squamous cell carcinoma of the lung and esophagus. Spontaneous cellular and humoral immune responses against SOX2 present in cancer patients classify it as a tu...

  14. Requirement for noncognate interaction with T cells for the activation of B cell immunoglobulin secretion by IL-2

    DEFF Research Database (Denmark)

    Owens, T

    1991-01-01

    23.1+ TH1 clone E9.D4 in F23.1 (anti-T cell receptor V-beta 8)-coated microwells. This induced polyclonal B cell activation to enter cell cycle (thymidine incorporation) at 2 days and to secrete immunoglobulin at 5 days. An anti-IL-2 mAb (S4B6) inhibited antibody production completely. Anti-IL-2 did...... not inhibit either LPS-induced B cell responses, or T cell activation (measured as IL-3 secretion). Anti-IL-2 receptor (anti-Tac) mAbs also inhibited T-dependent B cell responses, without affecting LPS responses. An anti-IFN-gamma mAb partially inhibited Ig secretion, without affecting entry into...

  15. A Recently Established Murine Model of Nasal Polyps Demonstrates Activation of B Cells, as Occurs in Human Nasal Polyps.

    Science.gov (United States)

    Kim, Dong-Young; Lee, Sun Hye; Carter, Roderick G; Kato, Atsushi; Schleimer, Robert P; Cho, Seong H

    2016-08-01

    Animal model systems are invaluable for examining human diseases. Our laboratory recently established a mouse model of nasal polyps (NPs) and investigated similarities and differences between this mouse model and human NPs. We especially focus on the hypothesis that B cell activation occurs during NP generation in the murine model. After induction of ovalbumin-induced allergic rhinosinusitis, 6% ovalbumin and Staphylococcus aureus enterotoxin B (10 ng) were instilled into the nasal cavity of mice three times per week for 8 weeks. The development of structures that somewhat resemble NPs (which we will refer to as NPs) was confirmed by hematoxylin and eosin staining. The mRNA and protein levels of various inflammatory cell markers and mediators were measured by real-time PCR in nasal tissue and by ELISA in nasal lavage fluid (NLF), respectively. Total Ig isotype levels in NLF were also quantitated using the Mouse Ig Isotyping Multiplex kit (EMD Millipore, Billerica, MA) on a Luminex 200 instrument (Life Technologies, Grand Island, NY). Similar to human NPs, there were significant increases in gene expression of inflammatory cell markers, such as CD19, CD138, CD11c, and mast cell protease-6 in nasal tissue samples of the NP group compared with those of the control group. In further investigations of B cell activation, mRNA expressions of B cell activating factor and a proliferation-inducing ligand were found to be significantly increased in mouse NP tissue. B cell-activating factor protein concentration and IgA and IgG1 levels in NLF were significantly higher in the NP group compared with the control group. In this study, the NP mouse model demonstrated enhanced B cell responses, which are reminiscent of B cell responses in human NPs. PMID:27163839

  16. B cells as a target of immune modulation

    Directory of Open Access Journals (Sweden)

    Hawker Kathleen

    2009-01-01

    Full Text Available B cells have recently been identified as an integral component of the immune system; they play a part in autoimmunity through antigen presentation, antibody secretion, and complement activation. Animal models of multiple sclerosis (MS suggest that myelin destruction is partly mediated through B cell activation (and plasmablasts. MS patients with evidence of B cell involvement, as compared to those without, tend to have a worse prognosis. Finally, the significant decrease in new gadolinium-enhancing lesions, new T2 lesions, and relapses in MS patients treated with rituximab (a monoclonal antibody against CD20 on B cells leads us to the conclusion that B cells play an important role in MS and that immune modulation of these cells may ameliorate the disease. This article will explore the role of B cells in MS and the rationale for the development of B cell-targeted therapeutics. MS is an immune-mediated disease that affects over 2 million people worldwide and is the number one cause of disability in young patients. Most therapeutic targets have focused on T cells; however, recently, the focus has shifted to the role of B cells in the pathogenesis of MS and the potential of B cells as a therapeutic target.

  17. Human dendritic cells activated via dectin-1 are efficient at priming Th17, cytotoxic CD8 T and B cell responses.

    Directory of Open Access Journals (Sweden)

    Sudhanshu Agrawal

    Full Text Available BACKGROUND: Dendritic cells capture antigens through PRRs and modulate adaptive immune responses. The type of adaptive immune T cell response generated is dependent upon the type of PRR activated by the microbes. Dectin-1 is a C-type lectin receptor present on dendritic cells. METHODOLOGY/PRINCIPAL FINDINGS: Here we show that selective dectin-1 agonist Curdlan can activate human DCs and induce the secretion of large amounts of IL-23, IL-1β, IL-6 and low levels of IL-12p70 as determined by ELISA. The Curdlan-stimulated DCs are efficient at priming naïve CD4 cells to differentiate into Th17 and Th1 cells. Furthermore, these CD4 T cells induce differentiation of B cells to secrete IgG and IgA. In addition, Curdlan-stimulated DCs promote the expansion and differentiation of Granzyme and perforin expressing cytotoxic T lymphocyte that display high cytolytic activity against target tumor cells in vitro. CONCLUSIONS/SIGNIFICANCE: These data demonstrate that DCs stimulated through Dectin-1 can generate efficient Th, CTL and B cell responses and can therefore be used as effective mucosal and systemic adjuvants in humans.

  18. 2,3,7,8-Tetrachlorodibenzo-p-dioxin-mediated disruption of the CD40 ligand-induced activation of primary human B cells

    International Nuclear Information System (INIS)

    Suppression of the primary antibody response is particularly sensitive to suppression by 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) in mice; however, surprisingly little is known concerning the effects of TCDD on humoral immunity or B cell function in humans. Results from a limited number of previous studies, primarily employing in vitro activation models, suggested that human B cell effector function is suppressed by TCDD. The present study sought to extend these findings by investigating, in primary human B cells, the effects of TCDD on several critical stages leading to antibody secretion including activation and plasmacytic differentiation using an in vitro CD40 ligand activation model. These studies revealed important differences in the response of human and mouse B cells to TCDD, the most striking being altered expression of plasmacytic differentiation regulators, B lymphocyte-induced maturation protein 1 and paired box protein 5, in mouse but not human B cells. The activation of human B cells was profoundly impaired by TCDD, as evidenced by decreased expression of activation markers CD80, CD86, and CD69. The impaired activation correlated with decreased cell viability, which prevented the progression of human B cells toward plasmacytic differentiation. TCDD treatment also attenuated the early activation of mitogen-activated protein kinases (MAPK) and Akt signaling in human B cells. Collectively, the present study provided experimental evidence for novel mechanisms by which TCDD impairs the effector function of primary human B cells. - Highlights: → In this study primary human and mouse B cell toxicity to TCDD was compared. → TCDD altered the expression of Blimp-1 and Pax5 in mouse but not human B cells. → TCDD markedly suppressed human B cell activation as characterized by CD80, CD86 and CD69 expression. → TCDD inhibited ERK, p38, and Akt phosphorylation in human B cells.

  19. Serum Concentrations of Antibodies against Outer Membrane Protein P6, Protein D, and T- and B-Cell Combined Antigenic Epitopes of Nontypeable Haemophilus influenzae in Children and Adults of Different Ages.

    Science.gov (United States)

    Hua, Chun-Zhen; Hu, Wei-Lin; Shang, Shi-Qiang; Li, Jian-Ping; Hong, Li-Quan; Yan, Jie

    2016-02-01

    Nontypeable Haemophilus influenzae (NTHi) is one of the most common etiologies of acute otitis media, rhinosinusitis, and pneumonia. Outer membrane proteins (OMPs) are the main focus in new vaccine development against NTHi, as the H. influenzae type b (Hib) vaccine does not cover noncapsulated NTHi. The OMPs P6 and protein D are the most promising candidate antigens for an NTHi vaccine, and low antibody levels against them in serum may be correlated with infection caused by NTHi. In the current study, we measured the antibody titers against P6, protein D, and their T- and B-cell combined peptide epitopes in healthy individuals of different ages. We found that children B-cell combined antigenic epitopes. Antibody titers increased at ages 1 to 6 months, peaked at 7 months to 3 years, and remained high at 4 to 6 years. The antibody titers started to decrease after 6 years and were the lowest in the 21- to 30-year group. The geometric mean titers (GMTs) of T- and B-cell combined antigenic epitopes in P6 and protein D were positively correlated with those of the protein antigens. Among 12 peptides tested, P6-61, P6-123, and protein D-167 epitopes were better recognized than others in human serum. These findings might contribute to the development of an effective serotype-independent vaccine for H. influenzae. PMID:26677200

  20. Plasmacytoid Dendritic Cells Regulate Autoreactive B Cell Activation via Soluble Factors and in a Cell-to-Cell Contact Manner1

    Science.gov (United States)

    Ding, Chuanlin; Cai, Yihua; Marroquin, Jose; Ildstad, Suzanne T.; Yan, Jun

    2012-01-01

    Plasmacytoid dendritic cells (pDCs) are specialized type I IFN producers, which play an important role in pathogenesis of autoimmune disorders. Dysregulated autoreactive B cell activation is a hallmark in most autoimmune diseases. This study was undertaken to investigate interactions between pDCs and autoreactive B cells. After co-culture of autoreactive B cells that recognize self-Ag small nuclear ribonucleoprotein particles with activated pDCs, we found that pDCs significantly enhance autoreactive B cell proliferation, autoAb production, and survival in response to toll-like receptor (TLR) and BCR stimulation. Neutralization of IFN-α/β and IL-6 abrogated partially pDC-mediated enhancement of autoreactive B cell activation. Transwell studies demonstrated that pDCs could provide activation signals to autoreactive B cells via cell-to-cell contact manner. The involvement of the ICAM-1-LFA-1 pathway was revealed as contributing to this effect. This in vitro enhancement effect was further demonstrated by an in vivo B cell adoptive transfer experiment, which showed that autoreactive B cell proliferation and activation were significantly decreased in MyD-88-deficient mice compared to WT mice. These data suggest the dynamic interplay between pDCs and B cells is required for full activation of autoreactive B cells upon TLR or BCR stimulation. PMID:19890051

  1. Antibody repertoire development in fetal and neonatal piglets. XX. B cell lymphogenesis is absent in the ileal Peyer´s patches, their repertoire development is antigen dependent, and they are not required for B cell maintenance

    Czech Academy of Sciences Publication Activity Database

    Butler, J.; Santiago-Mateo, K.; Sun, X.-Z.; Wertz, N.; Šinkora, Marek; Francis, D.

    2011-01-01

    Roč. 187, č. 10 (2011), s. 5141-5149. ISSN 0022-1767 R&D Projects: GA MŠk ME09089 Institutional research plan: CEZ:AV0Z50200510 Keywords : neonatal piglets * Peyer’s patches * b cell Subject RIV: EC - Immunology Impact factor: 5.788, year: 2011

  2. Regulation of Anti-DNA B Cells in Recombination-activating Gene–deficient Mice

    OpenAIRE

    Xu, Hui; Li, Hui; Suri-Payer, Elisabeth; Hardy, Richard R.; Weigert, Martin

    1998-01-01

    Anti-DNA antibodies are regulated in normal individuals but are found in high concentration in the serum of systemic lupus erythematosus (SLE) patients and the MRL lpr/lpr mouse model of SLE. We previously studied the regulation of anti–double-stranded (ds)DNA and anti–single-stranded (ss)DNA B cells in a nonautoimmune background by generating mice carrying immunoglobulin transgenes coding for anti-DNAs derived from MRL lpr/lpr. Anti-dsDNA B cells undergo receptor editing, but anti-ssDNA B ce...

  3. miR290-5p/292-5p activate the immunoglobulin kappa locus in B cell development.

    Directory of Open Access Journals (Sweden)

    Patty B Garcia

    Full Text Available Regulated expression of miRNAs influences development in a wide variety of contexts. We report here that miR290-5p (100049710 and miR292-5p (100049711 are induced at the pre-B stage of murine B cell development and that they influence assembly of the Igκ light chain gene (243469 by contributing to the activation of germline Igκ transcription (κGT. We found that upon forced over-expression of miR290-5p/292-5p in Abelson Murine Leukemia Virus (AMuLV transformed pro-B cells, two known activators of κGT, E2A (21423 and NF-κB (19697, show increased chromosomal binding to the kappa intronic enhancer. Conversely, knockdown of miR290-5p/292-5p in AMuLV pro-B cells blunts drug-induced activation of κGT. Furthermore, miR290-5p/292-5p knockdown also diminishes κGT activation, but not Rag1/2 (19373, 19374 expression, in an IL-7 dependent primary pro-B cell culture system. In addition, we identified a deficiency in κGT induction in miR290 cluster knockout mice. We hypothesize that increased expression of miR290-5p and miR292-5p contributes to the induction of κGT at the pre-B stage of B cell development through increased binding of NF-κB and E2A to kappa locus regulatory sequences.

  4. miR290-5p/292-5p activate the immunoglobulin kappa locus in B cell development.

    Science.gov (United States)

    Garcia, Patty B; Cai, Amie; Bates, Jamie G; Nolla, Hector; Schlissel, Mark S

    2012-01-01

    Regulated expression of miRNAs influences development in a wide variety of contexts. We report here that miR290-5p (100049710) and miR292-5p (100049711) are induced at the pre-B stage of murine B cell development and that they influence assembly of the Igκ light chain gene (243469) by contributing to the activation of germline Igκ transcription (κGT). We found that upon forced over-expression of miR290-5p/292-5p in Abelson Murine Leukemia Virus (AMuLV) transformed pro-B cells, two known activators of κGT, E2A (21423) and NF-κB (19697), show increased chromosomal binding to the kappa intronic enhancer. Conversely, knockdown of miR290-5p/292-5p in AMuLV pro-B cells blunts drug-induced activation of κGT. Furthermore, miR290-5p/292-5p knockdown also diminishes κGT activation, but not Rag1/2 (19373, 19374) expression, in an IL-7 dependent primary pro-B cell culture system. In addition, we identified a deficiency in κGT induction in miR290 cluster knockout mice. We hypothesize that increased expression of miR290-5p and miR292-5p contributes to the induction of κGT at the pre-B stage of B cell development through increased binding of NF-κB and E2A to kappa locus regulatory sequences. PMID:22928038

  5. T-Independent Activation-Induced Cytidine Deaminase Expression, Class-Switch Recombination, and Antibody Production by Immature/Transitional 1 B Cells1

    OpenAIRE

    Ueda, Yoshihiro; Liao, Dongmei; Yang, Kaiyong; Patel, Anjali; Kelsoe, Garnett

    2007-01-01

    Inflammation elicits a splenic lymphopoiesis of unknown physiologic significance but one that juxtaposes developing B cells and exogenous Ag. We show that immature and transitional 1 (immature/T1) B cells constitutively express activation-induced cytidine deaminase and B lymphocyte-induced maturation protein 1 in amounts that support accelerated plasmacytic differentiation and limited class-switch recombination. In vivo, activation of immature/T1 B cells by TLR ligands or bacterial vaccine ra...

  6. B-Cell Activating Factor as a Cancer Biomarker and Its Implications in Cancer-Related Cachexia

    Directory of Open Access Journals (Sweden)

    Michal Rihacek

    2015-01-01

    Full Text Available B-cell activating factor (BAFF is a cytokine and adipokine of the TNF ligand superfamily. The main biological function of BAFF in maintaining the maturation of B-cells to plasma cells has recently made it a target of the first FDA-approved selective BAFF antibody, belimumab, for the therapy of systemic lupus erythematosus. Concomitantly, the role of BAFF in cancer has been a subject of research since its discovery. Here we review BAFF as a biomarker of malignant disease activity and prognostic factor in B-cell derived malignancies such as multiple myeloma. Moreover, anti-BAFF therapy seems to be a promising approach in treatment of B-cell derived leukemias/lymphomas. In nonhematologic solid tumors, BAFF may contribute to cancer progression by mechanisms both dependent on and independent of BAFF’s proinflammatory role. We also describe ongoing research into the pathophysiological link between BAFF and cancer-related cachexia. BAFF has been shown to contribute to inflammation and insulin resistance which are known to worsen cancer cachexia syndrome. Taking all the above together, BAFF is emerging as a biomarker of several malignancies and a possible hallmark of cancer cachexia.

  7. B cells actively develop in the thymus by different means of selection

    Czech Academy of Sciences Publication Activity Database

    Šinkora, Marek; Šinkorová, M.; Holtmeier, W.; Butler, J. E.

    Québec City, 2004, s. 200. [International Veterinary Immunology Symposium /7./. Québec City (CA), 25.07.2004-30.07.2004] Institutional research plan: CEZ:AV0Z5020903 Keywords : b cell * cdr3 * pcr Subject RIV: EE - Microbiology, Virology

  8. An activation antigen on a subpopulation of B lymphocytes identified by the monoclonal antibody CMRF-17.

    Science.gov (United States)

    Peach, S F; Davidson, S E; McKenzie, J L; Nimmo, J C; Hart, D N

    1989-07-01

    The identification of membrane molecules expressed on subpopulations of B lymphocytes is of potential significance because these molecules may be candidates for regulating the activation, proliferation and differentiation of B cells. A new monoclonal antibody, CMRF-17, which reacts with a subpopulation of tonsil B lymphocytes has been produced. The antibody did not react with T lymphocytes in tonsil or peripheral blood nor most peripheral blood B lymphocytes but did label erythrocytes and some platelets. In tonsil, the germinal centre cells, cells in the interfollicular region and endothelial cells were positive, but mantle zone B cells were negative. Double labelling experiments showed that CMRF-17 reacted with activated tonsillar lymphocytes. The antigen recognized by CMRF-17 was heat stable, resistant to treatment with proteolytic enzymes and neuraminidase and was shown to be a carbohydrate determinant on one or more glycolipids. These characteristics of the antigen recognized by CMRF-17 and its pattern of reactivity distinguish this antibody from other monoclonal antibodies recognizing B-cell activation markers. It was notable that of the B-lymphoid malignancies tested to date, including those of probable follicular origin, few stained with CMRF-17. PMID:2474491

  9. Therapeutic strategies targeting B-cells in multiple sclerosis.

    Science.gov (United States)

    Milo, Ron

    2016-07-01

    Multiple sclerosis (MS) is a chronic inflammatory and demyelinating disease of the central nervous system (CNS) that traditionally has been considered to be mediated primarily by T-cells. Increasing evidence, however, suggests the fundamental role of B-cells in the pathogenesis of the disease. Recent strategies targeting B-cells in MS have demonstrated impressive and sometimes surprising results: B-cell depletion by monoclonal antibodies targeting the B-cell surface antigen CD20 (e.g. rituximab, ocrelizumab, ofatumumab) was shown to exert profound anti-inflammatory effect in MS with favorable risk-benefit ratio, with ocrelizumab demonstrating efficacy in both relapsing-remitting (RR) and primary-progressive (PP) MS in phase III clinical trials. Depletion of CD52 expressing T- and B-cells and monocytes by alemtuzumab resulted in impressive and durable suppression of disease activity in RRMS patients. On the other hand, strategies targeting B-cell cytokines such as atacicept resulted in increased disease activity. As our understanding of the biology of B-cells in MS is increasing, new compounds that target B-cells continue to be developed which promise to further expand the armamentarium of MS therapies and allow for more individualized therapy for patients with this complex disease. PMID:26970489

  10. Active suppression of major histocompatibility complex class II gene expression during differentiation from B cells to plasma cells

    International Nuclear Information System (INIS)

    Constitutive expression of major histocompatibility complex class II genes is acquired very early in B-cell ontogeny and is maintained up to the B-cell blast stage. Terminal differentiation in plasma cells is, however, accompanied by a loss of class II gene expression. In B cells this gene system is under the control of several loci encoding transacting factors with activator function, one of which, the aIr-1 gene product, operates across species barriers. In this report human class II gene expression is shown to be extinguished in somatic cell hybrids between the human class II-positive B-cell line Raji and the mouse class-II negative plasmacytoma cell line P3-U1. Since all murine chromosomes are retained in these hybrids and no preferential segregation of a specific human chromosome is observed, the results are compatible with the presence of suppressor factors of mouse origin, operating across species barriers and inhibiting class II gene expression. Suppression seems to act at the level of transcription or accumulation of class II-specific mRNA, since no human, and very few murine, class II transcripts are detectable in the hybrids

  11. B Cells Promote Th1- Skewed NKT Cell Response by CD1d-TCR Interaction

    OpenAIRE

    Shin, Jung Hoon; Park, Se-Ho

    2013-01-01

    CD1d expressing dendritic cells (DCs) are good glyco-lipid antigen presenting cells for NKT cells. However, resting B cells are very weak stimulators for NKT cells. Although α-galactosylceramide (α-GalCer) loaded B cells can activate NKT cells, it is not well defined whether B cells interfere NKT cell stimulating activity of DCs. Unexpectedly, we found in this study that B cells can promote Th1-skewed NKT cell response, which means a increased level of IFN-γ by NKT cells, concomitant with a d...

  12. Alterations in peripheral blood memory B cells in patients with active rheumatoid arthritis are dependent on the action of tumour necrosis factor

    OpenAIRE

    Souto-Carneiro, M. Margarida; Mahadevan, Vijayabhanu; Takada, Kazuki; Fritsch-Stork, Ruth; Nanki, Toshihiro; Brown, Margaret; Fleisher, Thomas A.; Wilson, Mildred; Goldbach-Mansky, Raphaela; Lipsky, Peter E

    2009-01-01

    INTRODUCTION: Disturbances in peripheral blood memory B cell subpopulations have been observed in various autoimmune diseases, but have not been fully delineated in rheumatoid arthritis (RA). Additionally, the possible role of tumour necrosis factor (TNF) in regulating changes in specific peripheral blood memory B cell subsets in RA is still unclear. METHODS: The frequency and distribution of B cell subsets in the peripheral blood and synovial membrane of active RA patients with long-standing...

  13. PEG10 Activation by Co-Stimulation of CXCR5 and CCR7 Essentially Contributes to Resistance to Apoptosis in CD19+CD34+ B Cells from Patients with B Cell Lineage Acute and Chronic Lymphocytic Leukemia

    Institute of Scientific and Technical Information of China (English)

    ChunsongHu; JeiXiong; LinjeiZhang; BaojunHuang; QiupingZhang; QunLi; MingzhenYang; YaouWu; QunWu; QianShen; QingpingGao; KejianZhang; ZhiminSun; JunyanLin; YouxinJin

    2004-01-01

    We investigated CD19+CD34+ and CD19+CD34 B cells from cord blood (CB) and typical patients with B cell lineage acute and chronic lymphocytic leukemia (B-ALL and B-CLL) in terms of expression and functions of CXCR5/CXCL13 and CCR7/CCL19. CXCR5 and CCR7 were selectively frequent expressed on B-ALL, B-CLL and CB CD19+CD34+ B cells, but not on CD19+CD34- B cells. Instead of induction of impressive chemotactic responsiveness, CXCL13 and CCL19 together induced significant resistance to TNF-α-mediated apoptosis in B-ALL and B-CLL but not CB CD19+CD34+ B cells. B-ALL and B-CLL CD19+CD34+ B cells expressed elevatedlevel of Paternally Expressed Gene 10 (PEG10), and CXCL13 and CCL19 together significantly up-regulated PEG10 expression in the cells. We found that CXCL13 and CCL19 together by means of activation of CXCR5 and CCR7 up-regulated PEG10 expression and function, subsequent stabilized caspase-3 and caspase-8 in B-ALL and B-CLL CD19+CD34+ B cells, and rescued the cells from TNF-α-mediated apoptosis. We suggested that normal lymphocytes, especially naive B and T cells, utilized CXCR5/CXCL13 and CCR7/CCL19 for migration, homing, maturation, and cell homeostasis as well as secondary lymphoid tissues organogenesis. Meanwhile certain malignant cells took advantages of CXCR5/CXCL13 and CCR7/CCL19 for infiltration, resistance to apoptosis, and inappropriate proliferation. Cellular & Molecular Immunology.

  14. PEG10 Activation by Co-Stimulation of CXCR5 and CCR7 Essentially Contributes to Resistance to Apoptosis in CD19+CD34+ B Cells from Patients with B Cell Lineage Acute and Chronic Lymphocytic Leukemia

    Institute of Scientific and Technical Information of China (English)

    Chunsong Hu; Qian Shen; Qingping Gao; Kejian Zhang; Zhimin Sun; Junyan Liu; Youxin Jin; Jinquan Tan; Jei Xiong; Linjei zhang; Baojun Huang; Qiuping Zhang; Qun Li; Mingzhen Yang; Yaou Wu; Qun Wu

    2004-01-01

    We investigated CD19+CD34+ and CD19+CD34- B cells from cord blood (CB) and typical patients with B cell lineage acute and chronic lymphocytic leukemia (B-ALL and B-CLL) in terms of expression and functions of CXCR5/CXCL13 and CCR7/CCL19. CXCR5 and CCR7 were selectively frequent expressed on B-ALL, B-CLL and CB CD19+CD34+ B cells, but not on CD19+CD34- B cells. Instead of induction of impressive chemotactic responsiveness, CXCL13 and CCL19 together induced significant resistance to TNF-α-mediated apoptosis in B-ALL and B-CLL but not CB CD19+CD34+ B cells. B-ALL and B-CLL CD19+CD34+ B cells expressed elevated level of Paternally Expressed Gene 10 (PEG10), and CXCL13 and CCL19 together significantly up-regulated PEG10 expression in the cells. We found that CXCL13 and CCL19 together by means of activation of CXCR5 and CCR7 up-regulated PEG10 expression and function, subsequent stabilized caspase-3 and caspase-8 in B-ALL and B-CLL CD19+CD34+ B cells, and rescued the cells from TNF-α-mediated apoptosis. We suggested that normal lymphocytes, especially na(I)ve B and T cells, utilized CXCR5/CXCL13 and CCR7/CCL19 for migration, homing, maturation, and cell homeostasis as well as secondary lymphoid tissues organogenesis.Meanwhile certain malignant cells took advantages of CXCR5/CXCL13 and CCR7/CCL19 for infiltration,resistance to apoptosis, and inappropriate proliferation.

  15. The BCL6 RD2 domain governs commitment of activated B-cells to form germinal centers

    Science.gov (United States)

    Huang, Chuanxin; Gonzalez, David G.; Cote, Christine M.; Jiang, Yanwen; Hatzi, Katerina; Teater, Matt; Dai, Kezhi; Hla, Timothy; Haberman, Ann M.; Melnick, Ari

    2014-01-01

    Summary To understand how the Bcl6 transcriptional repressor functions in the immune system we disrupted its RD2 repression domain in mice. Bcl6RD2MUT mice exhibit a complete loss of GC formation but retain normal extrafollicular responses. Bcl6RD2MUT antigen-engaged B-cells migrate to the interfollicular zone and interact with cognate T helper cells. However, these cells fail to complete early GC-commitment differentiation and coalesce as nascent GC aggregates. Bcl6 directly binds and represses trafficking receptors S1pr1 and Gpr183 by recruiting Hdac2 through the RD2 domain. Deregulation of these genes impairs B-cell migration and may contribute to GC failure in Bcl6RD2MUT mice. The development of functional GC-TFH cells was partially impaired in Bcl6RD2MUT mice. In contrast to Bcl6−/− mice, Bcl6RD2MUT animals experience no inflammatory disease or macrophage deregulation. These results reveal an essential role for RD2 repression in early GC commitment and striking biochemical specificity in Bcl6 control of humoral and innate immune-cell phenotypes. PMID:25176650

  16. Programming of marginal zone B-cell fate by basic Kruppel-like factor (BKLF/KLF3).

    Science.gov (United States)

    Turchinovich, Gleb; Vu, Thi Thanh; Frommer, Friederike; Kranich, Jan; Schmid, Sonja; Alles, Melanie; Loubert, Jean-Baptiste; Goulet, Jean-Philippe; Zimber-Strobl, Ursula; Schneider, Pascal; Bachl, Jürgen; Pearson, Richard; Crossley, Merlin; Agenès, Fabien; Kirberg, Jörg

    2011-04-01

    Splenic marginal zone (MZ) B cells are a lineage distinct from follicular and peritoneal B1 B cells. They are located next to the marginal sinus where blood is released. Here they pick up antigens and shuttle the load onto follicular dendritic cells inside the follicle. On activation, MZ B cells rapidly differentiate into plasmablasts secreting antibodies, thereby mediating humoral immune responses against blood-borne type 2 T-independent antigens. As Krüppel-like factors are implicated in cell differentiation/function in various tissues, we studied the function of basic Krüppel-like factor (BKLF/KLF3) in B cells. Whereas B-cell development in the bone marrow of KLF3-transgenic mice was unaffected, MZ B-cell numbers in spleen were increased considerably. As revealed in chimeric mice, this occurred cell autonomously, increasing both MZ and peritoneal B1 B-cell subsets. Comparing KLF3-transgenic and nontransgenic follicular B cells by RNA-microarray revealed that KLF3 regulates a subset of genes that was similarly up-regulated/down-regulated on normal MZ B-cell differentiation. Indeed, KLF3 expression overcame the lack of MZ B cells caused by different genetic alterations, such as CD19-deficiency or blockade of B-cell activating factor-receptor signaling, indicating that KLF3 may complement alternative nuclear factor-κB signaling. Thus, KLF3 is a driving force toward MZ B-cell maturation. PMID:21297003

  17. Active Akt and functional p53 modulate apoptosis in Abelson virus-transformed pre-B cells.

    Science.gov (United States)

    Gong, Li; Unnikrishnan, Indira; Raghavan, Anuradha; Parmar, Kalindi; Rosenberg, Naomi

    2004-02-01

    Suppression of apoptosis is an important feature of the Abelson murine leukemia virus (Ab-MLV) transformation process. During multistep transformation, Ab-MLV-infected pre-B cells undergo p53-dependent apoptosis during the crisis phase of transformation. Even once cells are fully transformed, an active v-Abl protein tyrosine kinase is required to suppress apoptosis because cells transformed by temperature-sensitive (ts) kinase mutants undergo rapid apoptosis after a shift to the nonpermissive temperature. However, inactivation of the v-Abl protein by a temperature shift interrupts signals transmitted via multiple pathways, making it difficult to identify those that are critically important for the suppression of apoptosis. To begin to dissect these pathways, we tested the ability of an SH2 domain Ab-MLV mutant, P120/R273K, to rescue aspects of the ts phenotype of pre-B cells transformed by the conditional kinase domain mutant. The P120/R273K mutant suppressed apoptosis at the nonpermissive temperature, a phenotype correlated with its ability to activate Akt. Apoptosis also was suppressed at the nonpermissive temperature by constitutively active Akt and in p53-null pre-B cells transformed with the ts kinase domain mutant. These data indicate that an intact Src homology 2 (SH2) domain is not critical for apoptosis suppression and suggest that signals transmitted through Akt and p53 play an important role in the response. PMID:14747529

  18. Deconvoluting post-transplant immunity: cell subset-specific mapping reveals pathways for activation and expansion of memory T, monocytes and B cells.

    Directory of Open Access Journals (Sweden)

    Yevgeniy A Grigoryev

    Full Text Available A major challenge for the field of transplantation is the lack of understanding of genomic and molecular drivers of early post-transplant immunity. The early immune response creates a complex milieu that determines the course of ensuing immune events and the ultimate outcome of the transplant. The objective of the current study was to mechanistically deconvolute the early immune response by purifying and profiling the constituent cell subsets of the peripheral blood. We employed genome-wide profiling of whole blood and purified CD4, CD8, B cells and monocytes in tandem with high-throughput laser-scanning cytometry in 10 kidney transplants sampled serially pre-transplant, 1, 2, 4, 8 and 12 weeks. Cytometry confirmed early cell subset depletion by antibody induction and immunosuppression. Multiple markers revealed the activation and proliferative expansion of CD45RO(+CD62L(- effector memory CD4/CD8 T cells as well as progressive activation of monocytes and B cells. Next, we mechanistically deconvoluted early post-transplant immunity by serial monitoring of whole blood using DNA microarrays. Parallel analysis of cell subset-specific gene expression revealed a unique spectrum of time-dependent changes and functional pathways. Gene expression profiling results were validated with 157 different probesets matching all 65 antigens detected by cytometry. Thus, serial blood cell monitoring reflects the profound changes in blood cell composition and immune activation early post-transplant. Each cell subset reveals distinct pathways and functional programs. These changes illuminate a complex, early phase of immunity and inflammation that includes activation and proliferative expansion of the memory effector and regulatory cells that may determine the phenotype and outcome of the kidney transplant.

  19. Prolactin Rescues Immature B-Cells from Apoptosis Induced by B-Cell Receptor Cross-Linking

    Science.gov (United States)

    Flores-Fernández, Rocio; Blanco-Favela, Francisco; Fuentes-Pananá, Ezequiel M.; Chávez-Sánchez, Luis; Gorocica-Rosete, Patricia; Pizaña-Venegas, Alberto; Chávez-Rueda, Adriana Karina

    2016-01-01

    Prolactin has an immunomodulatory effect and has been associated with B-cell-triggered autoimmune diseases, such as systemic lupus erythematosus (SLE). In mice that develop SLE, the PRL receptor is expressed in early bone marrow B-cells, and increased levels of PRL hasten disease manifestations, which are correlated with a reduction in the absolute number of immature B-cells. The aim of this work was to determine the effect of PRL in an in vitro system of B-cell tolerance using WEHI-231 cells and immature B-cells from lupus prone MRL/lpr mice. WEHI-231 cells express the long isoform of the PRL receptor, and PRL rescued the cells from cell death by decreasing the apoptosis induced by the cross-linking of the B-cell antigen receptor (BCR) as measured by Annexin V and active caspase-3. This decrease in apoptosis may have been due to the PRL and receptor interaction, which increased the relative expression of antiapoptotic Bcl-xL and decreased the relative expression of proapoptotic Bad. In immature B-cells from MRL/lpr mice, PRL increased the viability and decreased the apoptosis induced by the cross-linking of BCR, which may favor the maturation of self-reactive B-cells and contribute to the onset of disease. PMID:27314053

  20. Comparison of hematological alterations and markers of B-cell activation in workers exposed to benzene, formaldehyde and trichloroethylene.

    Science.gov (United States)

    Bassig, Bryan A; Zhang, Luoping; Vermeulen, Roel; Tang, Xiaojiang; Li, Guilan; Hu, Wei; Guo, Weihong; Purdue, Mark P; Yin, Songnian; Rappaport, Stephen M; Shen, Min; Ji, Zhiying; Qiu, Chuangyi; Ge, Yichen; Hosgood, H Dean; Reiss, Boris; Wu, Banghua; Xie, Yuxuan; Li, Laiyu; Yue, Fei; Freeman, Laura E Beane; Blair, Aaron; Hayes, Richard B; Huang, Hanlin; Smith, Martyn T; Rothman, Nathaniel; Lan, Qing

    2016-07-01

    Benzene, formaldehyde (FA) and trichloroethylene (TCE) are ubiquitous chemicals in workplaces and the general environment. Benzene is an established myeloid leukemogen and probable lymphomagen. FA is classified as a myeloid leukemogen but has not been associated with non-Hodgkin lymphoma (NHL), whereas TCE has been associated with NHL but not myeloid leukemia. Epidemiologic associations between FA and myeloid leukemia, and between benzene, TCE and NHL are, however, still debated. Previously, we showed that these chemicals are associated with hematotoxicity in cross-sectional studies of factory workers in China, which included extensive personal monitoring and biological sample collection. Here, we compare and contrast patterns of hematotoxicity, monosomy 7 in myeloid progenitor cells (MPCs), and B-cell activation biomarkers across these studies to further evaluate possible mechanisms of action and consistency of effects with observed hematologic cancer risks. Workers exposed to benzene or FA, but not TCE, showed declines in cell types derived from MPCs, including granulocytes and platelets. Alterations in lymphoid cell types, including B cells and CD4+ T cells, and B-cell activation markers were apparent in workers exposed to benzene or TCE. Given that alterations in myeloid and lymphoid cell types are associated with hematological malignancies, our data provide biologic insight into the epidemiological evidence linking benzene and FA exposure with myeloid leukemia risk, and TCE and benzene exposure with NHL risk. PMID:27207665

  1. Ibrutinib for B cell malignancies

    OpenAIRE

    Novero, Aileen; Ravella, Pavan M; Chen, Yamei; Dous, George; Liu, Delong

    2014-01-01

    Research over the role of Bruton’s agammaglobulinemia tyrosine kinase (BTK) in B-lymphocyte development, differentiation, signaling and survival has led to better understanding of the pathogenesis of B-cell malignancies. Down-regulation of BTK activity is an attractive novel strategy for treating patients with B-cell malignancies. Ibrutinib (PCI-32765), a potent inhibitor of BTK induces impressive responses in B-cell malignancies through irreversible bond with cysteine-481 in the active site ...

  2. E47 activates the Ig-heavy chain and TdT loci in non-B cells.

    OpenAIRE

    Choi, J K; Shen, C. P.; Radomska, H.S.; Eckhardt, L A; Kadesch, T

    1996-01-01

    The E2A proteins, E12 and E47, are basic helix-loop-helix (bHLH) proteins essential for the B-cell lineage. Initially identified as immunoglobulin enhancer-binding proteins, they have also been shown to activate immunoglobulin enhancer-based reporters in transient transfection assays. Here, we examine the relationship between E2A DNA binding activity and activation of the endogenous, chromosomal immunoglobulin heavy chain (IgH) locus. Using sterile I(mu) transcription as an indicator of IgH e...

  3. Typing of HLA class II and class I antigens using PHA-activated, IL-2-propagated T lymphocytes.

    Science.gov (United States)

    Leshem, B; Cohen, I; Sherman, L; Brautbar, C; Kedar, E

    1988-06-28

    We describe here a simple procedure, by which HLA class II antigens can be accurately and reliably identified in those patients where there is minimal or absent expression of HLA-DR,DQw antigens on B cells, or when the total number of leukocytes recovered from the patients do not permit reliable typing. Ficoll-Hypaque-separated peripheral blood mononuclear leukocytes, fresh or cryopreserved, were activated by PHA and then propagated in IL-2-containing medium until enough cells for typing were obtained (usually 7-14 days). At this stage, the cultured cells were shown to be primarily T cells (greater than 90% CD3+). Since the activated T cells propagate in the presence of IL-2, even a small number (10(4] of fresh or cryopreserved patients' cells suffice for this protocol. To date we have been able to successfully HLA-DR,DQw type 34/34 bone marrow transplantation candidates and 12/12 long-term dialysis patients, who were untypable using fresh cells. HLA-DR,DQw antigens on activated T cells from normal individuals were identical to those found on their uncultured B cells. In addition, class I antigens that were undetectable on the uncultured cells of one patient could be identified on activated T cells. The HLA antigens identified on the patients' activated T cells were confirmed by phenotypic analysis of cells from family members. PMID:3260612

  4. A Stochastic Model of the Germinal Center Integrating Local Antigen Competition, Individualistic T-B Interactions, and B Cell Receptor Signaling.

    Science.gov (United States)

    Wang, Peng; Shih, Chang-Ming; Qi, Hai; Lan, Yue-Heng

    2016-08-15

    The germinal center (GC) reaction underlies productive humoral immunity by orchestrating competition-based affinity maturation to produce plasma cells and memory B cells. T cells are limiting in this process. How B cells integrate signals from T cells and BCRs to make fate decisions while subjected to a cyclic selection process is not clear. In this article, we present a spatiotemporally resolved stochastic model that describes cell behaviors as rate-limited stochastic reactions. We hypothesize a signal integrator protein integrates follicular helper T (Tfh)- and Ag-derived signals to drive different B cell fates in a probabilistic manner and a dedicated module of Tfh interaction promoting factors control the efficiency of contact-dependent Tfh help delivery to B cells. Without assuming deterministic affinity-based decisions or temporal event sequence, this model recapitulates GC characteristics, highlights the importance of efficient T cell help delivery during individual contacts with B cells and intercellular positive feedback for affinity maturation, reveals the possibility that antagonism between BCR signaling and T cell help accelerates affinity maturation, and suggests that the dichotomy between affinity and magnitude of GC reaction can be avoided by tuning the efficiency of contact-dependent help delivery during reiterative T-B interactions. PMID:27421481

  5. Innate lymphoid cells integrate stromal and immunological signals to enhance antibody production by splenic marginal zone B cells

    OpenAIRE

    Magri, Giuliana; Miyajima, Michio; Bascones, Sabrina; Mortha, Arthur; Puga, Irene; Cassis, Linda; Barra, Carolina M; Comerma, Laura; Chudnovskiy, Aleksey; Gentile, Maurizio; Llige, David; Cols, Montserrat; Serrano, Sergi; Aróstegui, Juan Ignacio; Juan, Manel

    2014-01-01

    Innate lymphoid cells (ILCs) regulate stromal, epithelial and immune cells, but their impact on B cells remains unclear. We identified RORγt+ ILCs nearby the marginal zone (MZ), a splenic compartment containing innate-like B cells that respond to circulating T cell-independent (TI) antigens. Spenic ILCs established a bidirectional crosstalk with MAdCAM-1+ marginal reticular cells by providing tumor necrosis factor (TNF) and lymphotoxin, and activated MZ B cells via BAFF, CD40 ligand and the N...

  6. Similar disturbances in B cell activity and regulatory T cell function in Henoch-Schonlein purpura and systemic lupus erythematosus

    International Nuclear Information System (INIS)

    The immunoglobulin synthesizing activities of peripheral mononuclear cells (MNC) from five patients with Henoch-Schonlein purpura (HSP) and eight patients with active systemic lupus erythematosus (SLE) were compared. Cumulative amounts of IgM, IgG, and IgA synthesized and secreted by unstimulated and PWM-stimulated patient cells over a 12-day period were determied in a solid-phase radioimmunoassay. In unstimulated control cultures mean rates of IgM, IgG, and IgA synthesis were less than 250 ng/ml. The synthetic activities of patient MNC were markedly increased. In HSP cultures IgA was the major immunoglobulin class produced (2810 x/divide 1.33 ng/ml) followed by IgG (1754 x/divide 1.32 ng/ml) and IgM (404 x/divide 1.16 ng/ml). In SLE cultures IgA and IgG syntheses were equally elevated (4427 x/divide 1.20 and 4438 x/divide 1.49 ng/ml, respectively) whereas IgM synthesis averaged 967 x/divide 1.66 ng/ml. PWM stimulation of pateient MNC caused a sharp decline in the synthesis of all three immunoglobulin classes. After T cell depletion B cell-enriched fractions from HSP and SLE patients maintained high levels of IgA and IgG synthesis that were inhibited by PWM and by normal allogeneic but not autologous T cells. In PWM-stimulted co-cultures, patient T cells nonspecifically suppressed the synthetic activities of autologous and control B cells. in contrast patient B cells achieved normal levels of immunoglobulin synthesis when cultured with control T cells plus PWM. In longitudinal studies patient B and T cell disturbances persisted despite clinical improvement

  7. Interleukin 4 (B cell stimulatory factor 1) can mediate the induction of lymphokine-activated killer cell activity directed against fresh tumor cells

    OpenAIRE

    1987-01-01

    Interleukin 4 (IL-4) expresses multiple biologic activities, including B cell, mast cell, and T cell stimulation. We showed that the incubation of resting splenocytes from C57BL/6 mice solely in purified native or recombinant mouse IL-4 results in the generation of lymphokine-activated killer (LAK) activity directed against fresh, syngeneic sarcoma cells. The precursor activated by IL-4 expresses surface asialo-GM1. In addition, IL-4 is capable of amplifying the splenic LAK activity induced b...

  8. FOXP1 suppresses immune response signatures and MHC class II expression in activated B-cell-like diffuse large B-cell lymphomas

    DEFF Research Database (Denmark)

    Brown, P J; Wong, K K; Felce, S L;

    2016-01-01

    )-treated DLBCL patients (n=150), reduced HLA-DRA (<90% frequency) expression correlated with inferior overall survival (P=0.0003) and progression-free survival (P=0.0012) and with non-GCB subtype stratified by the Hans, Choi or Visco-Young algorithms (all P<0.01). In non-GCB DLBCL cases with <90% HLA-DRA, there......, including human leukocyte antigen DR alpha chain (HLA-DRA), were inversely correlated with FOXP1 transcript expression (P<0.05). FOXP1 knockdown in ABC-DLBCL cells led to increased cell-surface expression of HLA-DRA and CD74. In R-CHOP (rituximab, cyclophosphamide, doxorubicin, vincristine and prednisone...... was an inverse correlation with the frequency (P=0.0456) and intensity (P=0.0349) of FOXP1 expression. We propose that FOXP1 represents a novel regulator of genes targeted by the class II MHC transactivator CIITA (MHC II and CD74) and therapeutically targeting the FOXP1 pathway may improve antigen...

  9. Physical and functional association of the cbl protooncogen product with an src-family protein tyrosine kinase, p53/56lyn, in the B cell antigen receptor-mediated signaling.

    Science.gov (United States)

    Tezuka, T; Umemori, H; Fusaki, N; Yagi, T; Takata, M; Kurosaki, T; Yamamoto, T

    1996-02-01

    To identify novel signal transducers involved in signaling mediated by the Src-family protein tyrosine kinases (PTKs), we used a yeast two-hybrid system with a probe corresponding to the regulatory region of p56lyn, a member of Src-family PTKs. One of the isolated clones contained the COOH-terminal 470 amino acid residues of p120c-cbl, the product of the cellular homologue of the v-cbl retroviral oncogene. p120c-cbl is a cytoplasmic protein with nuclear protein-like motifs. Here we show in vivo association of p120c-cbl with p53/56lyn. After stimulation of the B cell antigen receptor (BCR), p120c-cbl was rapidly tyrosine phosphorylated. Studies with lyn- or syk-negative chicken B cells demonstrated that p53/56lyn, but not p72syk, was crucial for tyrosine phosphorylation of p120c-cbl upon stimulation of the BCR. We also show the importance of p59fyn in tyrosine phosphorylation of p120c-cbl in the T-cell receptor-mediated signaling using fyn-overexpressing T cell hybridomas and splenic T cells from fyn-deficient mice. These results suggest that p120c-cbl is an important substrate of Src-family PTKs in the intracellular signaling mediated by the antigen receptors PMID:8627181

  10. Differential Cytokine Production and Bystander Activation of Autoreactive B Cells in Response to CpG-A and CpG-B ODNs1

    OpenAIRE

    Avalos, Ana M.; Latz, Eicke; Mousseau, Betty; Christensen, Sean R.; Shlomchik, Mark J.; Lund, Frances,; Marshak-Rothstein, Ann

    2009-01-01

    Synthetic oligonucleotides containing CpG motifs (CpG ODNs) have been shown to induce proliferation, differentiation and cytokine production in B cells, macrophages and DCs through a TLR9-dependent mechanism. A class (CpG-A) and B class (CpG-B) ODNs display distinct physical properties. CpG-A, but not CpG-B, can multimerize to form exceedingly large lattices. CpG-A cannot effectively activate B cells but does induce pDCs to produce high levels of IFNα, while CpG-B is a potent B cell mitogen. ...

  11. 嵌合抗原受体T细胞在B细胞淋巴瘤中的研究进展%Research progress of chimeric antigen receptor-modified T cell in B cell lymphoma

    Institute of Scientific and Technical Information of China (English)

    应志涛

    2014-01-01

    B细胞淋巴瘤的预后在利妥昔单抗出现之后有了明显改善,但仍有一部分患者会复发进展.嵌合抗原受体T细胞(CAR T细胞)是通过基因修饰的方法获得的针对肿瘤细胞表面特定抗原的特异性T细胞,在复发难治B细胞淋巴瘤的治疗中取得了很好疗效,目前研究最多的是针对B细胞淋巴瘤表面的CD19抗原.文章综述了抗CD19 CAR T细胞在复发难治B细胞淋巴瘤中的疗效、不良反应及目前存在的问题.%The prognosis of B cell lymphoma has been significantly improved in the rituximab era,but there are also patients who are refractory or relapsed after rituximab-containing treatment.Chimeric antigen receptor T (CAR T) cell is a kind of T cell which is genetically modified to target specific antigen expressed on the lymphoma cell surface.The response rate of CAR T cell in relapsed or refractory B cell lymphoma is inspiring.Anti-CD19 CAR T cell is the most popular cell being tested in clinical studies.This article discussed the present situation of CAR T cell in B cell lymphoma,including the efficacy,toxicity,side effects and open questions existing in this treatment.

  12. Spontaneous complement activation on human B cells results in localized membrane depolarization and the clustering of complement receptor type 2 and C3 fragments

    DEFF Research Database (Denmark)

    Pedersen, Morten Løbner; Leslie, Robert G Q; Prodinger, Wolfgang M;

    2009-01-01

    While our previous studies have demonstrated that complement activation induced by complement receptors type 2 (CR2/CD21) and 1 (CR1/CD35) results in C3-fragment deposition and membrane attack complex (MAC) formation in human B cells, the consequences of these events for B-cell functions remain...... requires activation of complement via the alternative pathway, as indicated by total inhibition upon neutralization of factor D, and is abrogated by combined blockade of CR1 and CR2, but not of either receptor alone. The membrane depolarization is not associated with the apoptosis of B cells, as examined...... by co-staining with APO-2.7 or by the TdT-mediated biotin-dUTP nick-end labelling (TUNEL) assay. Confocal microscopy revealed that depolarization and C3 deposition, unlike MAC deposition, are limited to restricted areas on the B-cell surface. Double staining revealed a close association between the C...

  13. Early activation of natural killer and B cells in response to primary dengue virus infection in A/J mice

    International Nuclear Information System (INIS)

    Dengue virus (DEN) causes the most prevalent arthropod-borne viral illness in humans worldwide. Immune mechanisms that are involved in protection and pathogenesis of DEN infection have not been fully elucidated due largely to the lack of an adequate animal model. Therefore, as a first step, we characterized the primary immune response in immunocompetent inbred A/J mice that were infected intravenously with a non-mouse-adapted DEN type 2 (DEN2) strain. A subset (55%) of infected mice developed paralysis by 14 days post-infection (p.i.), harbored infectious DEN in the central nervous system (CNS), and had an elevated hematocrit and a decreased white blood cell (WBC) count. Immunologic studies detected (i) increased numbers of CD69+ splenic natural killer (NK) and B cells at day 3 p.i., (ii) DEN-specific IgM and IgG responses by days 3 and 7 p.i., respectively, and (iii) splenocyte production of IFNγ at day 14 p.i. We conclude that the early activities of NK cells, B cells and IgM, and later actions of IFNγ and IgG likely play a role in the defense against DEN infection

  14. The tumor antigen N-glycolyl-GM3 is a human CD1d ligand capable of mediating B cell and natural killer T cell interaction.

    Science.gov (United States)

    Gentilini, M Virginia; Pérez, M Eugenia; Fernández, Pablo Mariano; Fainboim, Leonardo; Arana, Eloísa

    2016-05-01

    The expression of N-glycolyl-monosialodihexosyl-ganglioside (NGcGM3) in humans is restricted to cancer cells; therefore, it is a tumor antigen. There are measurable quantities of circulating anti-NGcGM3 antibodies (aNGcGM3 Abs) in human serum. Interestingly, some people have circulating Ag-specific immunoglobulins G (IgGs) that are capable of complement mediated cytotoxicity against NGcGM3 positive cells, which is relevant for tumor surveillance. In light of the chemical nature of Ag, we postulated it as a candidate ligand for CD1d. Furthermore, we hypothesize that the immune mechanism involved in the generation of these Abs entails cross talk between B lymphocytes (Bc) and invariant natural killer T cells (iNKT). Combining cellular techniques, such as flow cytometry and biochemical assays, we demonstrated that CD1d binds to NGcGM3 and that human Bc present NGcGM3 in a CD1d context according to two alternative strategies. We also showed that paraformaldehyde treatment of cells expressing CD1d affects the presentation. Finally, by co-culturing primary human Bc with iNKT and measuring Ki-67 expression, we detected a reproducible increment in the proliferation of the iNKT population when Ag was on the medium. Our findings identify a novel, endogenous, human CD1d ligand, which is sufficiently competent to stimulate iNKT. We postulate that CD1d-restricted Bc presentation of NGcGM3 drives effective iNKT activation, an immunological mechanism that has not been previously described for humans, which may contribute to understanding aNGcGM3 occurrence. PMID:26969612

  15. Antigen

    Science.gov (United States)

    An antigen is any substance that causes your immune system to produce antibodies against it. This means your immune ... and is trying to fight it off. An antigen may be a substance from the environment, such ...

  16. The Mertk receptor tyrosine kinase promotes T-B interaction stimulated by IgD B-cell receptor cross-linking.

    Science.gov (United States)

    Shao, Wen-Hai; Zhen, Yuxuan; Finkelman, Fred D; Cohen, Philip L

    2014-09-01

    The Mertk receptor tyrosine kinase facilitates macrophage and DC apoptotic-cell clearance and regulates immune tolerance. Mertk may also contribute to B-cell activation, because Mertk-KO mice fail to develop autoantibodies when allo-activated by T cells. We investigated this possibility with a well-characterized model in which injection of mice with goat anti-IgD antibody causes membrane IgD cross-linking that induces T-independent B cell activation and antigen presentation to T cells. Goat anti-mouse IgD antibody-injected C57BL/6 Mertk-KO mice had normal initial B cell activation and proliferation, but significantly lower T cell activation and proliferation, as well as lower IgE and IgG anti-goat IgG responses, as compared to C57BL/6 WT controls. B cell antigen processing, analyzed by evaluating B cell fluorescence following injection of monoclonal anti-IgD antibody labeled with biotin or FITC, was comparable between Mertk-KO mice and WT mice. IgD Ab primed B cells from Mertk-KO mice exhibited significantly lower ability in activating memory T cells isolated from WT mice injected with the same antigen 10 days before. These observations suggest that Mertk expression is required for optimal B-cell antigen presentation, which is, in turn, required in this model for optimal T cell activation and subsequent T cell-dependent B cell differentiation. PMID:24768065

  17. Suppressive effect on polyclonal B-cell activation of a synthetic peptide homologous to a transmembrane component of oncogenic retroviruses

    International Nuclear Information System (INIS)

    Purified feline leukemia virus, UV light-inactivated feline leukemia virus, and a synthetic peptide (CKS-17) homologous to a well-conserved region of the transmembrane components of several human and animal retroviruses were each studied for their effect on IgG production by feline peripheral blood lymphocytes. Using a reverse hemolytic plaque assay, both the viable virus and the UV-inactivated feline leukemia virus, but not the CKS-17, activated B lymphocytes to secrete IgG. When staphylococcal protein A, a polyclonal B-cell activator, was used to stimulate IgG synthesis by feline lymphocytes, the viable virus, the UV-inactivated virus, and the CKS-17 peptide each strongly suppressed IgG secretion without compromising viability of the lymphocytes. These finding suggest that the immunosuppressive influences of feline leukemia virus on immunoglobulin synthesis may reside in a conserved portion of the envelope glycoprotein that includes the region homologous to CKS-17

  18. TIM-1 signaling in B cells regulates antibody production

    International Nuclear Information System (INIS)

    Highlights: → TIM-1 is highly expressed on anti-IgM + anti-CD40-stimulated B cells. → Anti-TIM-1 mAb enhanced proliferation and Ig production on activated B cell in vitro. → TIM-1 signaling regulates Ab production by response to TI-2 and TD antigens in vivo. -- Abstract: Members of the T cell Ig and mucin (TIM) family have recently been implicated in the control of T cell-mediated immune responses. In this study, we found TIM-1 expression on anti-IgM- or anti-CD40-stimulated splenic B cells, which was further up-regulated by the combination of anti-IgM and anti-CD40 Abs. On the other hand, TIM-1 ligand was constitutively expressed on B cells and inducible on anti-CD3+ anti-CD28-stimulated CD4+ T cells. In vitro stimulation of activated B cells by anti-TIM-1 mAb enhanced proliferation and expression of a plasma cell marker syndecan-1 (CD138). We further examined the effect of TIM-1 signaling on antibody production in vitro and in vivo. Higher levels of IgG2b and IgG3 secretion were detected in the culture supernatants of the anti-TIM-1-stimulated B cells as compared with the control IgG-stimulated B cells. When immunized with T-independent antigen TNP-Ficoll, TNP-specific IgG1, IgG2b, and IgG3 Abs were slightly increased in the anti-TIM-1-treated mice. When immunized with T-dependent antigen OVA, serum levels of OVA-specific IgG2b, IgG3, and IgE Abs were significantly increased in the anti-TIM-1-treated mice as compared with the control IgG-treated mice. These results suggest that TIM-1 signaling in B cells augments antibody production by enhancing B cell proliferation and differentiation.

  19. Ran GTPase-activating protein 1 is a therapeutic target in diffuse large B-cell lymphoma.

    Directory of Open Access Journals (Sweden)

    Kung-Chao Chang

    Full Text Available Lymphoma-specific biomarkers contribute to therapeutic strategies and the study of tumorigenesis. Diffuse large B-cell lymphoma (DLBCL is the most common type of malignant lymphoma. However, only 50% of patients experience long-term survival after current treatment; therefore, developing novel therapeutic strategies is warranted. Comparative proteomic analysis of two DLBCL lines with a B-lymphoblastoid cell line (LCL showed differential expression of Ran GTPase-activating protein 1 (RanGAP1 between them, which was confirmed using immunoblotting. Immunostaining showed that the majority of DLBCLs (92%, 46/50 were RanGAP1(+, while reactive lymphoid hyperplasia (n = 12 was RanGAP1(+ predominantly in germinal centers. RanGAP1 was also highly expressed in other B-cell lymphomas (BCL, n = 180 with brisk mitotic activity (B-lymphoblastic lymphoma/leukemia: 93%, and Burkitt lymphoma: 95% or cell-cycle dysregulation (mantle cell lymphoma: 83%, and Hodgkin's lymphoma 91%. Interestingly, serum RanGAP1 level was higher in patients with high-grade BCL (1.71 ± 2.28 ng/mL, n = 62 than in low-grade BCL (0.75 ± 2.12 ng/mL, n = 52 and healthy controls (0.55 ± 1.58 ng/mL, n = 75 (high-grade BCL vs. low-grade BCL, p = 0.002; high-grade BCL vs. control, p < 0.001, Mann-Whitney U test. In vitro, RNA interference of RanGAP1 showed no effect on LCL but enhanced DLBCL cell death (41% vs. 60%; p = 0.035 and cell-cycle arrest (G0/G1: 39% vs. 49%, G2/M: 19.0% vs. 7.5%; p = 0.030 along with decreased expression of TPX2 and Aurora kinases, the central regulators of mitotic cell division. Furthermore, ON 01910.Na (Estybon, a multikinase inhibitor induced cell death, mitotic cell arrest, and hyperphosphorylation of RanGAP1 in DLBCL cell lines but no effects in normal B and T cells. Therefore, RanGAP1 is a promising marker and therapeutic target for aggressive B-cell lymphoma, especially DLBCL.

  20. N-wasp is essential for the negative regulation of B cell receptor signaling.

    Directory of Open Access Journals (Sweden)

    Chaohong Liu

    2013-11-01

    Full Text Available Negative regulation of receptor signaling is essential for controlling cell activation and differentiation. In B-lymphocytes, the down-regulation of B-cell antigen receptor (BCR signaling is critical for suppressing the activation of self-reactive B cells; however, the mechanism underlying the negative regulation of signaling remains elusive. Using genetically manipulated mouse models and total internal reflection fluorescence microscopy, we demonstrate that neuronal Wiskott-Aldrich syndrome protein (N-WASP, which is coexpressed with WASP in all immune cells, is a critical negative regulator of B-cell signaling. B-cell-specific N-WASP gene deletion causes enhanced and prolonged BCR signaling and elevated levels of autoantibodies in the mouse serum. The increased signaling in N-WASP knockout B cells is concurrent with increased accumulation of F-actin at the B-cell surface, enhanced B-cell spreading on the antigen-presenting membrane, delayed B-cell contraction, inhibition in the merger of signaling active BCR microclusters into signaling inactive central clusters, and a blockage of BCR internalization. Upon BCR activation, WASP is activated first, followed by N-WASP in mouse and human primary B cells. The activation of N-WASP is suppressed by Bruton's tyrosine kinase-induced WASP activation, and is restored by the activation of SH2 domain-containing inositol 5-phosphatase that inhibits WASP activation. Our results reveal a new mechanism for the negative regulation of BCR signaling and broadly suggest an actin-mediated mechanism for signaling down-regulation.

  1. B-cell-specific enhancer activity of conserved upstream elements of the class II major histocompatibility complex DQB gene

    Energy Technology Data Exchange (ETDEWEB)

    Sakurai, M.; Strominger, J.L.

    1988-09-01

    A 95-base-pair immediate upstream sequence of the human class II major histocompatibility complex DQB gene containing the conserved X and Y elements showed enhancer activity in a transient expression assay. An enhancer test plasmid harboring the bacterial chloramphenicol acetyltransferase gene under the control of a truncated simian virus 40 enhancerless early promoter was employed. The DQB sequence inserted into this plasmid was active as an enhancer in Raji cells (human Burkitt lymphoma cells) but not active in Jurkat cells (human T-cell leukemia cells) or in HeLa cells (human cervical carcinoma cells). This cell-type specificity suggests that this enhancer activity may be involved in the tissue specificity of the DQB gene that is normally expressed only in mature B cells, macrophages, and thymic epithelial cells. Deletion analysis showed that both X and Y box sequences are essential for the full activity of the enhancer sequence and that these two sequences may function in a cooperative manner as cis-acting elements. Further deletions were used to define the 5' border of the X element. These results suggest that previously characterized protein factors that bind to X and Y include transcription factors involved in the cell-type specificity of this enhancer activity.

  2. Intestinal B cell hyperactivity in AIDS is controlled by highly active antiretroviral therapy

    OpenAIRE

    Nilssen, D E; Øktedalen, O; Brandtzaeg, P

    2004-01-01

    Background: It is well documented that highly active antiretroviral therapy (HAART) restores systemic immunity to human immunodeficiency virus (HIV) but the effect of this treatment on the mucosal immune system is less clear.

  3. Carbohydrate-functionalized nanovaccines preserve HIV-1 antigen stability and activate antigen presenting cells.

    Science.gov (United States)

    Vela Ramirez, J E; Roychoudhury, R; Habte, H H; Cho, M W; Pohl, N L B; Narasimhan, B

    2014-01-01

    The functionalization of polymeric nanoparticles with ligands that target specific receptors on immune cells offers the opportunity to tailor adjuvant properties by conferring pathogen mimicking attributes to the particles. Polyanhydride nanoparticles are promising vaccine adjuvants with desirable characteristics such as immunomodulation, sustained antigen release, activation of antigen presenting cells (APCs), and stabilization of protein antigens. These capabilities can be exploited to design nanovaccines against viral pathogens, such as HIV-1, due to the important role of dendritic cells (DCs) and macrophages in viral spread. In this work, an optimized process was developed for carbohydrate functionalization of HIV-1 antigen-loaded polyanhydride nanoparticles. The carbohydrate-functionalized nanoparticles preserved antigenic properties upon release and also enabled sustained antigen release kinetics. Particle internalization was observed to be chemistry-dependent with positively charged nanoparticles being taken up more efficiently by DCs. Up-regulation of the activation makers CD40 and CD206 was demonstrated with carboxymethyl-α-d-mannopyranosyl-(1,2)-d-mannopyranoside functionalized nanoparticles. The secretion of the cytokines IL-6 and TNF-α was shown to be chemistry-dependent upon stimulation with carbohydrate-functionalized nanoparticles. These results offer important new insights upon the interactions between carbohydrate-functionalized nanoparticles and APCs and provide foundational information for the rational design of targeted nanovaccines against HIV-1. PMID:25068589

  4. Subcellular localization of PUMA regulates its pro-apoptotic activity in Burkitt's lymphoma B cells

    OpenAIRE

    Ambroise, Gorbatchev; Portier, Alain; Roders, Nathalie; Arnoult, Damien; Vazquez, Aimé

    2015-01-01

    The BH3-only protein PUMA (p53-upregulated modulator of apoptosis) is a major regulator of apoptosis. It belongs to the Bcl-2 family of proteins responsible for maintaining mitochondrial outer membrane integrity by controlling the intrinsic (mitochondrial) apoptotic pathway. We describe here a new pathway regulating PUMA activation through the control of its subcellular distribution. Surprisingly, neither PUMA upregulation in normal activated human B lymphocytes nor high levels of PUMA in Bur...

  5. T-cell independent, B-cell receptor-mediated induction of telomerase activity differs among IGHV mutation-based subgroups of chronic lymphocytic leukemia patients

    OpenAIRE

    Damle, Rajendra N.; Temburni, Sonal; Banapour, Taraneh; Paul, Santanu; Mongini, Patricia K. A.; Allen, Steven L.; Kolitz, Jonathan E.; Rai, Kanti R; Chiorazzi, Nicholas

    2012-01-01

    Although B-cell chronic lymphocytic leukemia (B-CLL) clones with unmutated IGHV genes (U-CLL) exhibit greater telomerase activity than those with mutated IGHV genes (M-CLL), the extent to which B-cell receptor (BCR) triggering contributes to telomerase up-regulation is not known. Therefore, we studied the effect of BCR stimulation on modulating telomerase activity. The multivalent BCR ligand, dextran conjugated anti-μ mAb HB57 (HB57-dex), increased telomerase activity and promoted cell surviv...

  6. GILT expression in B cells diminishes cathepsin S steady-state protein expression and activity

    OpenAIRE

    Phipps-Yonas, Hannah; Semik, Vikki; Hastings, Karen Taraszka

    2012-01-01

    MHC class II-restricted Ag processing requires protein degradation in the endocytic pathway for the activation of CD4+ T cells. Gamma-interferon-inducible lysosomal thiol reductase (GILT) facilitates Ag processing by reducing protein disulfide bonds in this compartment. Lysosomal cysteine protease cathepsin S (CatS) contains disulfide bonds and mediates essential steps in MHC class II-restricted processing, including proteolysis of large polypeptides and cleavage of the invariant chain. We so...

  7. The Fab fragment of a directly activating monoclonal antibody that precipitates a disulfide-linked heterodimer from a helper T cell clone blocks activation by either allogeneic Ia or antigen and self-Ia

    OpenAIRE

    1984-01-01

    We characterize a monoclonal antibody directed against the antigen/Ia receptor of a cloned helper T cell line that induced T cell clone proliferation and T cell clone-dependent B cell proliferation at antibody concentrations as low as 10(-11) M. A Fab fragment of this antibody was not stimulatory, implicating cross-linking of antigen receptors as the primary signal for T cell activation. The Fab fragment inhibited activation of this clone by both allogeneic Ia and antigen plus self-Ia, but no...

  8. Impact of adjuvants on CD4(+) T cell and B cell responses to a protein antigen vaccine: Results from a phase II, randomized, multicenter trial.

    Science.gov (United States)

    Leroux-Roels, Geert; Marchant, Arnaud; Levy, Jack; Van Damme, Pierre; Schwarz, Tino F; Horsmans, Yves; Jilg, Wolfgang; Kremsner, Peter G; Haelterman, Edwige; Clément, Frédéric; Gabor, Julian J; Esen, Meral; Hens, Annick; Carletti, Isabelle; Fissette, Laurence; Tavares Da Silva, Fernanda; Burny, Wivine; Janssens, Michel; Moris, Philippe; Didierlaurent, Arnaud M; Van Der Most, Robbert; Garçon, Nathalie; Van Belle, Pascale; Van Mechelen, Marcelle

    2016-08-01

    Immunogenicity and safety of different adjuvants combined with a model antigen (HBsAg) were compared. Healthy HBV-naïve adults were randomized to receive HBs adjuvanted with alum or Adjuvant Systems AS01B, AS01E, AS03A or AS04 at Days 0 and 30. Different frequencies of HBs-specific CD4+ T cells 14days post dose 2 but similar polyfunctionality profiles were induced by the different adjuvants with frequencies significantly higher in the AS01B and AS01E groups than in the other groups. Antibody concentrations 30days post-dose 2 were significantly higher in AS01B, AS01E and AS03A than in other groups. Limited correlations were observed between HBs-specific CD4+ T cell and antibody responses. Injection site pain was the most common solicited local symptom and was more frequent in AS groups than in alum group. Different adjuvants formulated with the same antigen induced different adaptive immune responses and reactogenicity patterns in healthy naïve adults. The results summary for this study (GSK study number 112115 - NCT# NCT00805389) is available on the GSK Clinical Study Register and can be accessed at www.gsk-clinicalstudyregister.com. PMID:27236001

  9. Pathogenic role of B-cell receptor signaling and canonical NF-κB activation in mantle cell lymphoma.

    Science.gov (United States)

    Saba, Nakhle S; Liu, Delong; Herman, Sarah E M; Underbayev, Chingiz; Tian, Xin; Behrend, David; Weniger, Marc A; Skarzynski, Martin; Gyamfi, Jennifer; Fontan, Lorena; Melnick, Ari; Grant, Cliona; Roschewski, Mark; Navarro, Alba; Beà, Sílvia; Pittaluga, Stefania; Dunleavy, Kieron; Wilson, Wyndham H; Wiestner, Adrian

    2016-07-01

    To interrogate signaling pathways activated in mantle cell lymphoma (MCL) in vivo, we contrasted gene expression profiles of 55 tumor samples isolated from blood and lymph nodes from 43 previously untreated patients with active disease. In addition to lymph nodes, MCL often involves blood, bone marrow, and spleen and is incurable for most patients. Recently, the Bruton tyrosine kinase (BTK) inhibitor ibrutinib demonstrated important clinical activity in MCL. However, the role of specific signaling pathways in the lymphomagenesis of MCL and the biologic basis for ibrutinib sensitivity of these tumors are unknown. Here, we demonstrate activation of B-cell receptor (BCR) and canonical NF-κB signaling specifically in MCL cells in the lymph node. Quantification of BCR signaling strength, reflected in the expression of BCR regulated genes, identified a subset of patients with inferior survival after cytotoxic therapy. Tumor proliferation was highest in the lymph node and correlated with the degree of BCR activation. A subset of leukemic tumors showed active BCR and NF-κB signaling apparently independent of microenvironmental support. In one of these samples, we identified a novel somatic mutation in RELA (E39Q). This sample was resistant to ibrutinib-mediated inhibition of NF-κB and apoptosis. In addition, we identified germ line variants in genes encoding regulators of the BCR and NF-κB pathway previously implicated in lymphomagenesis. In conclusion, BCR signaling, activated in the lymph node microenvironment in vivo, appears to promote tumor proliferation and survival and may explain the sensitivity of this lymphoma to BTK inhibitors. PMID:27127301

  10. Cutting Edge: IL-4, IL-21, and IFN-γ Interact To Govern T-bet and CD11c Expression in TLR-Activated B Cells.

    Science.gov (United States)

    Naradikian, Martin S; Myles, Arpita; Beiting, Daniel P; Roberts, Kenneth J; Dawson, Lucas; Herati, Ramin Sedaghat; Bengsch, Bertram; Linderman, Susanne L; Stelekati, Erietta; Spolski, Rosanne; Wherry, E John; Hunter, Christopher; Hensley, Scott E; Leonard, Warren J; Cancro, Michael P

    2016-08-15

    T-bet and CD11c expression in B cells is linked with IgG2c isotype switching, virus-specific immune responses, and humoral autoimmunity. However, the activation requisites and regulatory cues governing T-bet and CD11c expression in B cells remain poorly defined. In this article, we reveal a relationship among TLR engagement, IL-4, IL-21, and IFN-γ that regulates T-bet expression in B cells. We find that IL-21 or IFN-γ directly promote T-bet expression in the context of TLR engagement. Further, IL-4 antagonizes T-bet induction. Finally, IL-21, but not IFN-γ, promotes CD11c expression independent of T-bet. Using influenza virus and Heligmosomoides polygyrus infections, we show that these interactions function in vivo to determine whether T-bet(+) and CD11c(+) B cells are formed. These findings suggest that T-bet(+) B cells seen in health and disease share the common initiating features of TLR-driven activation within this circumscribed cytokine milieu. PMID:27430719

  11. Interleukin-24 mediates apoptosis in human B-cells through early activation of cell cycle arrest followed by late induction of the mitochondrial apoptosis pathway.

    Science.gov (United States)

    Hadife, Nader; Nemos, Christophe; Frippiat, Jean-Pol; Hamadé, Tala; Perrot, Aurore; Dalloul, Ali

    2013-03-01

    Interleukin (IL)-24 has death-promoting effects on various proliferating cells including B-cells from chronic lymphocytic leukemia (CLL) and germinal center B-cells, but its molecular mechanisms are poorly understood. Using a B-cell differentiation model and mRNA profiling, we found that recombinant (r)IL-24 stimulated genes of the mitochondrial apoptotic pathway (Bax, Bid, Casp8, COX6C, COX7B) after 36 h, whereas the transcription of genes involved in DNA replication and metabolism was inhibited within 6 h. Unexpectedly, insulin-like growth factor 1 (IGF1), a hormone known to promote cell growth, was stimulated by IL-24. Activated B-cells express receptor for IGF1, to which they become sensitized and undergo apoptosis, a mechanism similar in this respect to IL-24-induced cell death. Furthermore, inhibition of the IGF1 pathway reversed the effects of IL-24. IL-24-mediated apoptosis was also antagonized by pifithrin-alpha, an inhibitor of p53 transactivation. Altogether, these results disclose sequential molecular signals generated by IL-24 in activated B-cells. PMID:22860893

  12. Identification and characterization of TspA, a major CD4(+) T-cell- and B-cell-stimulating Neisseria-specific antigen.

    Science.gov (United States)

    Kizil, G; Todd, I; Atta, M; Borriello, S P; Ait-Tahar, K; Ala'Aldeen, D A

    1999-07-01

    In search for novel T-cell immunogens involved in protection against invasive meningococcal disease, we screened fractionated proteins of Neisseria meningitidis (strain SD, B:15:P1.16) by using peripheral blood mononuclear cells (PBMCs) and specific T-cell lines obtained from normal individuals and patients convalescing from N. meningitidis infection. Proteins of iron-depleted meningococci produced higher PBMC proliferation indices than proteins of iron-replete organisms, indicating that iron-regulated proteins are T-cell immunogens. Insoluble proteins of the iron-depleted cells, which produced better T-cell stimulation than soluble ones, were fractionated by using sodium dodecyl sulfate-polyacrylamide gels and recovered as five fractions (F1 to F5) corresponding to decreasing molecular weight ranges. The proteins were purified (by elution and precipitation) or electroblotted onto nitrocellulose membranes (dissolved and precipitated) before use in further T-cell proliferation assays. One of the fractions (F1), containing high-molecular-mass proteins (>130 kDa), consistently showed the strongest T-cell proliferation responses in all of the T-cell lines examined. F1 proteins were subdivided into four smaller fractions (F1A to F1D) which were reexamined in T-cell proliferation assays, and F1C induced the strongest responses in patients' T-cell lines. Rabbit polyclonal antibodies to F1C components were used to screen a genomic expression library of N. meningitidis. Two major clones (C1 and C24) of recombinant meningococcal DNA were identified and fully sequenced. Sequence analysis showed that C24 (1,874 bp) consisted of a single open reading frame (ORF), which was included in clone C1 (2, 778 bp). The strong CD4(+) T-cell-stimulating effect of the polypeptide product of this ORF (named TspA) was confirmed, using a patient T-cell line. Immunogenicity for B cells was confirmed by showing that convalescent patients' serum antibodies recognized TspA on Western blots

  13. Resource Competition Determines Selection of B Cell Repertoires

    OpenAIRE

    de Boer, R J; Freitas, A. A.; Perelson, A. S.

    2001-01-01

    Previous experiments with mouse chimeras demonstrated that cellular competition for antigen- specific survival signals plays a crucial role in the maintenance of the naive B cell repertoire. Transgenic (Tg) B cell populations in these chimeras have a shortened lifespan and poor competitive abilities as compared to more diverse non-Tg populations in the same mice. We develop a mathematical model to investigate the mechanism of B cell competition. The model allows for various B cell clones, gen...

  14. Patients with Tuberculosis Have a Dysfunctional Circulating B-Cell Compartment, Which Normalizes following Successful Treatment

    Science.gov (United States)

    del Nonno, Franca; Baiocchini, Andrea; Petrone, Linda; Vanini, Valentina; Smits, Hermelijn H.; Palmieri, Fabrizio; Goletti, Delia; Ottenhoff, Tom H. M.

    2016-01-01

    B-cells not only produce immunoglobulins and present antigens to T-cells, but also additional key roles in the immune system. Current knowledge on the role of B-cells in infections caused by intracellular bacteria is fragmentary and contradictory. We therefore analysed the phenotypical and functional properties of B-cells during infection and disease caused by Mycobacterium tuberculosis (Mtb), the bacillus causing tuberculosis (TB), and included individuals with latent TB infection (LTBI), active TB, individuals treated successfully for TB, and healthy controls. Patients with active or treated TB disease had an increased proportion of antibodies reactive with mycobacteria. Patients with active TB had reduced circulating B-cell frequencies, whereas only minor increases in B-cells were detected in the lungs of individuals deceased from TB. Both active TB patients and individuals with LTBI had increased relative fractions of B-cells with an atypical phenotype. Importantly, these B-cells displayed impaired proliferation, immunoglobulin- and cytokine- production. These defects disappeared upon successful treatment. Moreover, T-cell activity was strongest in individuals successfully treated for TB, compared to active TB patients and LTBI subjects, and was dependent on the presence of functionally competent B-cells as shown by cellular depletion experiments. Thus, our results reveal that general B-cell function is impaired during active TB and LTBI, and that this B-cell dysfunction compromises cellular host immunity during Mtb infection. These new insights may provide novel strategies for correcting Mtb infection-induced immune dysfunction towards restored protective immunity. PMID:27304615

  15. RAG-mediated DNA double-strand breaks activate a cell type-specific checkpoint to inhibit pre-B cell receptor signals.

    Science.gov (United States)

    Bednarski, Jeffrey J; Pandey, Ruchi; Schulte, Emily; White, Lynn S; Chen, Bo-Ruei; Sandoval, Gabriel J; Kohyama, Masako; Haldar, Malay; Nickless, Andrew; Trott, Amanda; Cheng, Genhong; Murphy, Kenneth M; Bassing, Craig H; Payton, Jacqueline E; Sleckman, Barry P

    2016-02-01

    DNA double-strand breaks (DSBs) activate a canonical DNA damage response, including highly conserved cell cycle checkpoint pathways that prevent cells with DSBs from progressing through the cell cycle. In developing B cells, pre-B cell receptor (pre-BCR) signals initiate immunoglobulin light (Igl) chain gene assembly, leading to RAG-mediated DNA DSBs. The pre-BCR also promotes cell cycle entry, which could cause aberrant DSB repair and genome instability in pre-B cells. Here, we show that RAG DSBs inhibit pre-BCR signals through the ATM- and NF-κB2-dependent induction of SPIC, a hematopoietic-specific transcriptional repressor. SPIC inhibits expression of the SYK tyrosine kinase and BLNK adaptor, resulting in suppression of pre-BCR signaling. This regulatory circuit prevents the pre-BCR from inducing additional Igl chain gene rearrangements and driving pre-B cells with RAG DSBs into cycle. We propose that pre-B cells toggle between pre-BCR signals and a RAG DSB-dependent checkpoint to maintain genome stability while iteratively assembling Igl chain genes. PMID:26834154

  16. CD137 is induced by the CD40 signal on chronic lymphocytic leukemia B cells and transduces the survival signal via NF-κB activation.

    Directory of Open Access Journals (Sweden)

    Yukana Nakaima

    Full Text Available CD137 is a member of the tumor necrosis factor receptor family that is expressed on activated T cells. This molecule provides a co-stimulatory signal that enhances the survival, and differentiation of cells, and has a crucial role in the development of CD8 cytotoxic T cells and anti-tumor immunity. Here we report that CD137 expression is also induced on normal or malignant human B cells by CD40 ligation by its ligand CD154. This CD137 induction was more prominent in chronic lymphocytic leukemia (CLL cells than in other types of B cells. CD137 stimulation on B cells by its ligand induced the nuclear translocation of p52 (a non-canonical NF-κB factor. In agreement with this finding, expression of the survival factor BCL-XL was upregulated. Consequently, the CD137 signal augmented the survival of CD154-stimulated CLL B cells in vitro. This unexpected induction of CD137 on B cells by CD40 signal may influence the clinical course of CLL.

  17. B Cell Receptor Activation Predominantly Regulates AKT-mTORC1/2 Substrates Functionally Related to RNA Processing

    Science.gov (United States)

    Mohammad, Dara K.; Ali, Raja H.; Turunen, Janne J.; Nore, Beston F.; Smith, C. I. Edvard

    2016-01-01

    Protein kinase B (AKT) phosphorylates numerous substrates on the consensus motif RXRXXpS/T, a docking site for 14-3-3 interactions. To identify novel AKT-induced phosphorylation events following B cell receptor (BCR) activation, we performed proteomics, biochemical and bioinformatics analyses. Phosphorylated consensus motif-specific antibody enrichment, followed by tandem mass spectrometry, identified 446 proteins, containing 186 novel phosphorylation events. Moreover, we found 85 proteins with up regulated phosphorylation, while in 277 it was down regulated following stimulation. Up regulation was mainly in proteins involved in ribosomal and translational regulation, DNA binding and transcription regulation. Conversely, down regulation was preferentially in RNA binding, mRNA splicing and mRNP export proteins. Immunoblotting of two identified RNA regulatory proteins, RBM25 and MEF-2D, confirmed the proteomics data. Consistent with these findings, the AKT-inhibitor (MK-2206) dramatically reduced, while the mTORC-inhibitor PP242 totally blocked phosphorylation on the RXRXXpS/T motif. This demonstrates that this motif, previously suggested as an AKT target sequence, also is a substrate for mTORC1/2. Proteins with PDZ, PH and/or SH3 domains contained the consensus motif, whereas in those with an HMG-box, H15 domains and/or NF-X1-zinc-fingers, the motif was absent. Proteins carrying the consensus motif were found in all eukaryotic clades indicating that they regulate a phylogenetically conserved set of proteins. PMID:27487157

  18. IgA production requires B cell interaction with subepithelial dendritic cells in Peyer's patches.

    Science.gov (United States)

    Reboldi, Andrea; Arnon, Tal I; Rodda, Lauren B; Atakilit, Amha; Sheppard, Dean; Cyster, Jason G

    2016-05-13

    Immunoglobulin A (IgA) induction primarily occurs in intestinal Peyer's patches (PPs). However, the cellular interactions necessary for IgA class switching are poorly defined. Here we show that in mice, activated B cells use the chemokine receptor CCR6 to access the subepithelial dome (SED) of PPs. There, B cells undergo prolonged interactions with SED dendritic cells (DCs). PP IgA class switching requires innate lymphoid cells, which promote lymphotoxin-β receptor (LTβR)-dependent maintenance of DCs. PP DCs augment IgA production by integrin αvβ8-mediated activation of transforming growth factor-β (TGFβ). In mice where B cells cannot access the SED, IgA responses against oral antigen and gut commensals are impaired. These studies establish the PP SED as a niche supporting DC-B cell interactions needed for TGFβ activation and induction of mucosal IgA responses. PMID:27174992

  19. B Cells and Autoantibodies in Multiple Sclerosis

    Directory of Open Access Journals (Sweden)

    Anne-Katrin Pröbstel

    2015-07-01

    Full Text Available While over the past decades T cells have been considered key players in the pathogenesis of multiple sclerosis (MS, it has only recently become evident that B cells have a major contributing role. Our understanding of the role of B cells has evolved substantially following the clinical success of B cell-targeting therapies and increasing experimental evidence for significant B cell involvement. Rather than mere antibody-producing cells, it is becoming clear that they are team players with the capacity to prime and regulate T cells, and function both as pro- and anti-inflammatory mediators. However, despite tremendous efforts, the target antigen(s of B cells in MS have yet to be identified. The first part of this review summarizes the clinical evidence and results from animal studies pointing to the relevance of B cells in the pathogenesis of MS. The second part gives an overview of the currently known potential autoantigen targets. The third part recapitulates and critically appraises the currently available B cell-directed therapies.

  20. Overcoming antigen masking of anti-amyloidbeta antibodies reveals breaking of B cell tolerance by virus-like particles in amyloidbeta immunized amyloid precursor protein transgenic mice

    Directory of Open Access Journals (Sweden)

    Ugen Kenneth E

    2004-06-01

    Full Text Available Abstract Background In prior work we detected reduced anti-Aβ antibody titers in Aβ-vaccinated transgenic mice expressing the human amyloid precursor protein (APP compared to nontransgenic littermates. We investigated this observation further by vaccinating APP and nontransgenic mice with either the wild-type human Aβ peptide, an Aβ peptide containing the "Dutch Mutation", E22Q, or a wild-type Aβ peptide conjugated to papillomavirus virus-like particles (VLPs. Results Anti-Aβ antibody titers were lower in vaccinated APP than nontransgenic mice even when vaccinated with the highly immunogenic Aβ E22Q. One concern was that human Aβ derived from the APP transgene might mask anti-Aβ antibodies in APP mice. To test this possibility, we dissociated antigen-antibody complexes by incubation at low pH. The low pH incubation increased the anti-Aβ antibody titers 20–40 fold in APP mice but had no effect in sera from nontransgenic mice. However, even after dissociation, the anti-Aβ titers were still lower in transgenic mice vaccinated with wild-type Aβ or E22Q Aβ relative to non-transgenic mice. Importantly, the dissociated anti-Aβ titers were equivalent in nontransgenic and APP mice after VLP-based vaccination. Control experiments demonstrated that after acid-dissociation, the increased antibody titer did not cross react with bovine serum albumin nor alpha-synuclein, and addition of Aβ back to the dissociated serum blocked the increase in antibody titers. Conclusions Circulating human Aβ can interfere with ELISA assay measurements of anti-Aβ titers. The E22Q Aβ peptide vaccine is more immunogenic than the wild-type peptide. Unlike peptide vaccines, VLP-based vaccines against Aβ abrogate the effects of Aβ self-tolerance.

  1. CD19 chimeric antigen receptor (CD19 CAR)-redirected adoptive T-cell immunotherapy for the treatment of relapsed or refractory B-cell Non-Hodgkin’s Lymphomas

    Science.gov (United States)

    Onea, Alexandra S; Jazirehi, Ali R

    2016-01-01

    Recovery rates for B-cell Non-Hodgkin’s Lymphoma (NHL) are up to 70% with current standard-of-care treatments including rituximab (chimeric anti-CD20 monoclonal antibody) in combination with chemotherapy (R-CHOP). However, patients who do not respond to first-line treatment or develop resistance have a very poor prognosis. This signifies the need for the development of an optimal treatment approach for relapsed/refractory B-NHL. Novel CD19- chimeric antigen receptor (CAR) T-cell redirected immunotherapy is an attractive option for this subset of patients. Anti-CD19 CAR T-cell therapy has already had remarkable efficacy in various leukemias as well as encouraging outcomes in phase I clinical trials of relapsed/refractory NHL. In going forward with additional clinical trials, complementary treatments that may circumvent potential resistance mechanisms should be used alongside anti-CD19 T-cells in order to prevent relapse with resistant strains of disease. Some such supplementary tactics include conditioning with lymphodepletion agents, sensitizing with kinase inhibitors and Bcl-2 inhibitors, enhancing function with multispecific CAR T-cells and CD40 ligand-expressing CAR T-cells, and safeguarding with lymphoma stem cell-targeted treatments. A therapy regimen involving anti-CD19 CAR T-cells and one or more auxiliary treatments could dramatically improve prognoses for patients with relapsed/refractory B-cell NHL. This approach has the potential to revolutionize B-NHL salvage therapy in much the same way rituximab did for first-line treatments. PMID:27186412

  2. Suppression by Δ9-tetrahydrocannabinol of the primary immunoglobulin M response by human peripheral blood B cells is associated with impaired STAT3 activation

    International Nuclear Information System (INIS)

    This study was undertaken to gain insights into the mechanism for Δ9-tetrahydrocannabinol (Δ9-THC)-mediated suppression of primary immunoglobulin M (IgM) responses in humans. An in vitro activation model, which employs cell surface-expressed CD40 ligand (CD40L) and recombinant cytokines (interleukin (IL)-2, -6, and -10), was used to differentiate human peripheral blood (HPB) naïve B cells into IgM secreting cells. Pretreatment with Δ9-THC significantly decreased the number of IgM secreting cells as determined by ELISPOT. The attenuation of IgM secretion by Δ9-THC involved, at least in part, the impairment of plasma cell differentiation as evidenced by suppression of immunoglobulin joining chain (IgJ) mRNA expression. The analysis at each of two different stages critically involved in plasma cell differentiation indicates that Δ9-THC impaired both the primary activation stage and proliferation of B cells. Interestingly, Δ9-THC selectively suppressed the surface expression of CD80, but not other measured B-cell activation markers (CD69, CD86, and ICAM1). Furthermore, pretreatment with Δ9-THC was accompanied by a robust decrease of STAT3 phosphorylation, whereas the phosphorylation of the p65 NFκB subunit was not affected. Collectively, these data provide new insights into the mechanisms for impaired B cell function by Δ9-THC

  3. Early B-cell factor 1 (EBF1) is critical for transcriptional control of SLAMF1 gene in human B cells.

    Science.gov (United States)

    Schwartz, Anton M; Putlyaeva, Lidia V; Covich, Milica; Klepikova, Anna V; Akulich, Kseniya A; Vorontsov, Ilya E; Korneev, Kirill V; Dmitriev, Sergey E; Polanovsky, Oleg L; Sidorenko, Svetlana P; Kulakovskiy, Ivan V; Kuprash, Dmitry V

    2016-10-01

    Signaling lymphocytic activation molecule family member 1 (SLAMF1)/CD150 is a co-stimulatory receptor expressed on a variety of hematopoietic cells, in particular on mature lymphocytes activated by specific antigen, costimulation and cytokines. Changes in CD150 expression level have been reported in association with autoimmunity and with B-cell chronic lymphocytic leukemia. We characterized the core promoter for SLAMF1 gene in human B-cell lines and explored binding sites for a number of transcription factors involved in B cell differentiation and activation. Mutations of SP1, STAT6, IRF4, NF-kB, ELF1, TCF3, and SPI1/PU.1 sites resulted in significantly decreased promoter activity of varying magnitude, depending on the cell line tested. The most profound effect on the promoter strength was observed upon mutation of the binding site for Early B-cell factor 1 (EBF1). This mutation produced a 10-20 fold drop in promoter activity and pinpointed EBF1 as the master regulator of human SLAMF1 gene in B cells. We also identified three potent transcriptional enhancers in human SLAMF1 locus, each containing functional EBF1 binding sites. Thus, EBF1 interacts with specific binding sites located both in the promoter and in the enhancer regions of the SLAMF1 gene and is critical for its expression in human B cells. PMID:27424222

  4. C/EBPα Activates Pre-existing and De Novo Macrophage Enhancers during Induced Pre-B Cell Transdifferentiation and Myelopoiesis.

    Science.gov (United States)

    van Oevelen, Chris; Collombet, Samuel; Vicent, Guillermo; Hoogenkamp, Maarten; Lepoivre, Cyrille; Badeaux, Aimee; Bussmann, Lars; Sardina, Jose Luis; Thieffry, Denis; Beato, Miguel; Shi, Yang; Bonifer, Constanze; Graf, Thomas

    2015-08-11

    Transcription-factor-induced somatic cell conversions are highly relevant for both basic and clinical research yet their mechanism is not fully understood and it is unclear whether they reflect normal differentiation processes. Here we show that during pre-B-cell-to-macrophage transdifferentiation, C/EBPα binds to two types of myeloid enhancers in B cells: pre-existing enhancers that are bound by PU.1, providing a platform for incoming C/EBPα; and de novo enhancers that are targeted by C/EBPα, acting as a pioneer factor for subsequent binding by PU.1. The order of factor binding dictates the upregulation kinetics of nearby genes. Pre-existing enhancers are broadly active throughout the hematopoietic lineage tree, including B cells. In contrast, de novo enhancers are silent in most cell types except in myeloid cells where they become activated by C/EBP factors. Our data suggest that C/EBPα recapitulates physiological developmental processes by short-circuiting two macrophage enhancer pathways in pre-B cells. PMID:26235892

  5. C/EBPα Activates Pre-existing and De Novo Macrophage Enhancers during Induced Pre-B Cell Transdifferentiation and Myelopoiesis

    Directory of Open Access Journals (Sweden)

    Chris van Oevelen

    2015-08-01

    Full Text Available Transcription-factor-induced somatic cell conversions are highly relevant for both basic and clinical research yet their mechanism is not fully understood and it is unclear whether they reflect normal differentiation processes. Here we show that during pre-B-cell-to-macrophage transdifferentiation, C/EBPα binds to two types of myeloid enhancers in B cells: pre-existing enhancers that are bound by PU.1, providing a platform for incoming C/EBPα; and de novo enhancers that are targeted by C/EBPα, acting as a pioneer factor for subsequent binding by PU.1. The order of factor binding dictates the upregulation kinetics of nearby genes. Pre-existing enhancers are broadly active throughout the hematopoietic lineage tree, including B cells. In contrast, de novo enhancers are silent in most cell types except in myeloid cells where they become activated by C/EBP factors. Our data suggest that C/EBPα recapitulates physiological developmental processes by short-circuiting two macrophage enhancer pathways in pre-B cells.

  6. Friend or foe: emerging role of nuclear factor kappa-light-chain-enhancer of activated B cells in cell senescence

    Directory of Open Access Journals (Sweden)

    Mowla SN

    2013-09-01

    Full Text Available Sophia N Mowla,1 Neil D Perkins,2 Parmjit S Jat11Department of Neurodegenerative Disease and MRC Prion Unit, UCL Institute of Neurology, Queen Square, London, UK; 2Institute for Cell and Molecular Biosciences, Faculty of Medical Sciences, Newcastle University, Newcastle upon Tyne, UKAbstract: The nuclear factor kappa-light-chain-enhancer of activated B cells (NF-κB proteins are a family of ubiquitously expressed transcription factors that regulate the response to cellular stress. They mediate innate and adaptive immunity through the initiation of an inflammatory response to pro-inflammatory signals. The role of persistent inflammation in aiding tumor development has led to the NF-κB family of transcription factors being strongly implicated in promoting cancer. However, recent studies have now revealed that NF-κB can also function as a tumor suppressor through the induction of cellular senescence. Cellular senescence is a stable cell cycle arrest that normal cells undergo in response to a variety of intrinsic and extrinsic stimuli including: progressive telomere shortening, changes in telomeric structure, or other forms of genotoxic stress. Senescence can compromise tissue repair and regeneration, contributing to tissue and organismal aging via the accumulation of senescent cells, depletion of stem/progenitor cells and secretion of an array of inflammatory cytokines, chemokines, and matrix metalloproteinases. Senescence can also lead to the removal of potentially cancerous cells, thereby acting as a potent tumor suppressor mechanism. Herein, we review the evidence indicating a role for NF-κB in tumor suppression via cellular senescence and suggest that depending upon the subunit expressed, the biological context, and the type and intensity of the signal, NF-κB can indeed promote senescence growth arrest.Keywords: cell senescence, growth arrest, NF-κB, SASP

  7. Assembly and Function of the Precursor B-Cell Receptor.

    Science.gov (United States)

    Übelhart, Rudolf; Werner, Markus; Jumaa, Hassan

    2016-01-01

    During early stages of development, precursor B lymphocytes express a characteristic type of antigen receptor known as the pre-B-cell receptor (pre-BCR). This receptor differs from conventional BCRs in that it possesses a germ line-encoded surrogate light chain (SLC), which is associated with the signal transduction machinery via heavy chain (HC) proteins that have been generated by productive rearrangement of the immunoglobulin HC genes. The pre-BCR marks a key step of B-cell commitment, as it activates the B-cell-specific signaling cascade and mediates the selection, expansion, and differentiation of cells expressing a productively rearranged HC protein. Another difference between the pre-BCR and conventional BCR might be the initial event that triggers receptor activation, as the pre-BCR is activated in the absence of external ligands, while conventional BCRs require antigen for activation. Nonetheless, the pre-BCR downstream signaling cascade is largely similar to that of the BCR suggesting that the characteristic LC of the pre-BCR mediates important receptor interactions thereby providing distinctive, germ line-encoded features to the pre-BCR. In fact, the SLC enables the pre-BCR to act as a surrogate autoreactive receptor. Here, we outline the structure and function of the pre-BCR and how the autonomous signaling capacity might be a direct consequence of pre-BCR assembly. In addition to its role in early B-cell development, we discuss how the ordered activation of downstream signaling cascades enables the pre-BCR to activate seemingly opposing cellular programs such as proliferation and differentiation. PMID:26415650

  8. Research progress of chimeric antigen receptor-transduced-T cells in B-cell malignancies%嵌合抗原受体T细胞在B细胞性血液肿瘤中的研究进展

    Institute of Scientific and Technical Information of China (English)

    徐丽丽; 王扬

    2014-01-01

    Chimeric antigen receptor-transduced-T (CAR-T) cells shows strong cytotoxicity to the tumor with specific antigen by CTL effects and cytokines releasing without MHC-restriction.As multiple clinical trials from cancer centers around the West countries reported recent years,anti-CD19-CAR T cells can not only induce complete remission of B-cell malignancies resistant,but also eliminate the minimal residual disease (MRD) at the molecular level.It is believed that the obstacles from bench to clinic will be cleared and CAR will become one of the main cancer therapies with breakthroughs in comprehensive treatment of tumors.%嵌合抗原受体(CAR)转染的T细胞通过释放细胞因子及细胞毒性T淋巴细胞(CTL)效应可高效杀伤靶抗原阳性的肿瘤细胞,且不受相容性复合体(MHC)的限制.近年欧美多个癌症中心的临床试验表明,CD19靶向的CAR-T细胞输注不仅可使复发状态的B细胞急性淋巴细胞白血病(B-ALL)患者达完全缓解,且分子水平的微小残留病(MRD)也可转阴.其在慢性淋巴细胞白血病、B细胞性恶性淋巴瘤患者中也取得了显著疗效,相信不久的将来该技术必将成为肿瘤综合治疗的重要手段.

  9. Prevalence and chemotherapy-induced reactivation of occult hepatitis B virus among hepatitis B surface antigen negative patients with diffuse large B-cell lymphoma: Significance of hepatitis B core antibodies screening

    International Nuclear Information System (INIS)

    Background: Occult hepatitis B infection (OBI) is characterized by negative hepatitis B surface antigen (HBsAg) and detectable hepatitis B virus (HBV)-DNA in the liver and/or serum, with or without hepatitis B core antibody (anti-HBc). Anti-HBc is the most sensitive marker of previous HBV. HBV reactivation in patients under immunosuppressive treatment is life-threatening, occurring in both overt and occult HBV especially in hematological malignancies. Aim of the work: To evaluate the prevalence and chemotherapy-induced reactivation of OBI among hepatitis B surface antigen negative patients with diffuse large B-cell lymphoma (DLBCL) patients and to determine the significance of anti-HBc screening among this group of patients before receiving chemotherapy. Patients and methods: This cross-sectional study included 72 DLBCL patients negative for HBsAg, HBsAb and hepatitis C virus antibodies (anti-HCV). Patients were subjected to investigations including anti-HBc. All patients underwent alanine transaminase (ALT) monitoring before each cycle of chemotherapy and monthly for 12 months after the end of chemotherapy. Patients with suspected OBI were tested for HBV-DNA using real-time polymerase chain reaction (PCR). Results: Anti-HBc was detected in 10 of 72 HBsAg negative sera (13.89%) (95% confidence interval 6.9-22.2%). Five of the 10 anti-HBc positive patients in this study had OBI reactivation. Conclusion: The study concluded that anti-HBc screening is mandatory before chemotherapy. HBsAg-negative/anti-HBc-positive patients should be closely observed for signs of HBV reactivation through the regular monitoring of ALT. Prophylaxis lamivudine is recommended for anti-HBc positive patients before chemotherapy.

  10. Characterization of xenoantiserum produced against B cell acute lymphoblastic leukemia cell line

    Directory of Open Access Journals (Sweden)

    Akagi,Tadaatsu

    1982-10-01

    Full Text Available Antiserum was produced in white rabbit by intravenously injecting living cells of a B cell acute lymphoblastic leukemia (ALL line (BALL-1. The reactivity of the antiserum against various lymphoid cell lines was examined by membrane immunofluorescence after appropriate absorption. Serum absorbed with non-T, non-B (NALL-1 and T-ALL (TALL-1 cells recognized B cell antigens distinct from Ia-like antigens on both normal and neoplastic B cells. After further absorption with tonsillar cells or normal B cell line (KO-HL-3, it reacted only with BALL-1 cells and did not react with other leukemia/lymphoma and normal B cell lines. The serum absorbed with tonsillar cells reacted only with BALL-1 and some B cell lines. Thus we were able to obtain antisera with specificity to B cell antigen, B-ALL antigen, and B cell line antigen.

  11. Analysis of human B cell response to recombinantLeishmaniaLPG3

    Institute of Scientific and Technical Information of China (English)

    Mostafa Haji Fatahaliha; Maryam Hosseini; Sanaz Rasolzadeh; Dariush Shane Bandi; Behzad Baradaran; Farhad Jadidi-Niaragh; Mehdi Yousefi

    2015-01-01

    Objective:To evaluate the capability of recombinant Leishmania LPG3 and its fragments in the activation of B cells.Methods:In the present study, human B cells were purified from peripheral blood of 10 adult healthy subjects using magnetic-activated cell sorting technique. Subsequently, purified B cells were treated with recombinant LPG3, and itsN-terminal and C-terminal fragments at different concentrations (2, 10 and 20 μg/mL). B cell activation was assessed through expression of CD69 molecule by flowcytometry and secretion of IL-6, TNF-αα and IL-10 cytokines via enzyme-linked immunosorbent assay following treatment with recombinant antigens.Results:Our results showed that while the recombinant LPG-3 could significantly increase the production of IL-6 and TNF-α (P<0.05) in B cells, it had no effect on the secretion of IL-10 by B cells.Conclusions: Our study indicated that recombinant LPG-3 and especially itsN-terminal fragment could stimulate B cell response as an important immune response component against leishmaniasis. Thus, it seems that it can be considered as an effective adjuvant in vaccine developments against leishmaniasis.

  12. Interleukin-21 stimulates B-cell immunoglobulin E synthesis in human beings concomitantly with activation-induced cytidine deaminase expression and differentiation into plasma cells.

    Science.gov (United States)

    Kobayashi, Shinji; Haruo, Nagumo; Sugane, Kazuo; Ochs, Hans D; Agematsu, Kazunaga

    2009-01-01

    Interleukin (IL)-21 downregulates immunoglobulin E (IgE) production in murine systems by inhibiting germline epsilon transcription in IL-4-stimulated B cells. We here sought to clarify the function of IL-21 in human B-cell IgE synthesis. IL-21 dramatically enhanced IgE production by human mononuclear cells, or purified total, naive, or memory B cells in the presence of IL-4 plus anti-CD40 mAb cross-linked with CD32-transfectants, and the production was strengthened with further addition of IL-10. It was concomitant to the enhancement of activation-induced cytidine deaminase (AID) mRNA expression, but no increase of germline epsilon transcription. We also observed that IL-21 promoted B-cell differentiation into plasma cells with increase of B-lymphocyte-induced maturation protein-1 (Blimp-1), but not X-box binding protein 1 (XBP-1), which was further accentuated by co-stimulation with IL-4 plus CD40 signaling. Thus, IL-21 is a strong inducer of IgE production in human beings concomitantly with AID expression and the differentiation into plasma cells. Our data suggest that IL-21 plays an important role in occurrence and the treatment of allergic disorders. PMID:19026702

  13. T-cell independent, B-cell receptor-mediated induction of telomerase activity differs among IGHV mutation-based subgroups of chronic lymphocytic leukemia patients.

    Science.gov (United States)

    Damle, Rajendra N; Temburni, Sonal; Banapour, Taraneh; Paul, Santanu; Mongini, Patricia K A; Allen, Steven L; Kolitz, Jonathan E; Rai, Kanti R; Chiorazzi, Nicholas

    2012-09-20

    Although B-cell chronic lymphocytic leukemia (B-CLL) clones with unmutated IGHV genes (U-CLL) exhibit greater telomerase activity than those with mutated IGHV genes (M-CLL), the extent to which B-cell receptor (BCR) triggering contributes to telomerase up-regulation is not known. Therefore, we studied the effect of BCR stimulation on modulating telomerase activity. The multivalent BCR ligand, dextran conjugated anti-μ mAb HB57 (HB57-dex), increased telomerase activity and promoted cell survival and proliferation preferentially in U-CLL cases, whereas the PI3K/Akt inhibitor LY294002 blocked HB57-dex induced telomerase activation. Although both U-CLL and M-CLL clones exhibited similar membrane proximal signaling responses to HB57-dex, telomerase activity and cell proliferation, when inducible in M-CLL, differed. B-CLL cells stimulated using bivalent F(ab')(2) -goat anti-μ antibody (goat anti-μ) exhibited higher membrane proximal response in U-CLL than M-CLL cells, whereas telomerase activity, cell survival, and proliferation were induced to lower levels than those induced by HB57-dex. In normal B lymphocytes, HB57-dex induced less protein phosphorylation but more cell proliferation and survival than goat anti-μ. Although both anti-BCR stimuli induced comparable telomerase activity, normal CD5(+) B cells preferentially exhibited higher hTERT positivity than their CD5(-) counterparts. These findings provide an understanding of how BCR-mediated signals impact telomerase modulation in IGHV mutation-based subgroups of B-CLL and normal B cells. PMID:22875913

  14. B cells contribute to MS pathogenesis through antibody-dependent and antibody-independent mechanisms

    Directory of Open Access Journals (Sweden)

    Wilson HL

    2012-05-01

    Full Text Available Heather L Wilson1,21Vaccine and Infectious Disease Organization-International Vaccine Center, 2Department of Biochemistry, University of Saskatchewan, Saskatoon, Saskatchewan, CanadaAbstract: For many years, central dogma defined multiple sclerosis (MS as a T cell-driven autoimmune disorder; however, over the past decade there has been a burgeoning recognition that B cells contribute to the pathogenesis of certain MS disease subtypes. B cells may contribute to MS pathogenesis through production of autoantibodies (or antibodies directed at foreign bodies, which unfortunately cross-react with self-antigens, through promotion of T cell activation via antigen presentation, or through production of cytokines. This review highlights evidence for antibody-dependent and antibody-independent B cell involvement in MS pathogenesis.Keywords: autoantibodies, antibody targets, clinically isolated MS, primary progressive MS, secondary progressive MS, relapsing and remitting MS, T cells, T regulatory cells

  15. Cloning,expression and purification of B cell epitope antigen peptide of EGFR dimerization%EGFR二聚化B细胞表位抗原肤基因克隆、表达与纯化

    Institute of Scientific and Technical Information of China (English)

    朱磊; 吴梅芝; 赵林; 黄绮玲; 梁梅; 李黄金

    2011-01-01

    目的 利用基因工程手段建立表皮生长因子受体(EGFR)二聚化B细胞表位抗原肽MVF-ER的高效制备方法.方法 采用重叠延伸PCR扩增MVF-ER后克隆于表达载体pET32a,于大肠杆菌BL21(DE3)进行表达,产物采用肠激酶酶切和镍离子鳌合亲和层析法纯化.结果 通过10对引物5步重叠延伸得到273bp的扩增产物,目的基因与硫氧环蛋白(Trx)融合后可高效表达,经酶切和层析后得到纯度为95%的抗原肤.结论 成功建立抗原肽"MVF-ER"的高效制备方法,为进一步研究EGFR过表达恶性肿瘤的治疗性疫苗打下了坚实基础.%We aim to establish a high performance preparation system for B cell epitope antigen peptide of EGFR dimerization for further research on therapeutic vaccine of EGFR over-expression tumors. The MVF-ER gene of 273 bp was amplified by splicing overlap extension PCR (SOE-PCR), then cloned into the vector of pET32a, and expressed in E.coli BL21 (DE3) in the form of fusion with Trx. The expression product was purified by Ni+ affinity chromatography. The antigen peptide was recovered from the fusion protein by enterokinase and purified to a purity of 95% confirmed by SDS-PAGE. The results above showed that the preparation method of MVF-ER antigen peptide was successfully established, which will facilitate further study of the therapeutic cancer vaccine.

  16. Characterization and functional studies of forkhead box protein 3(-) lymphocyte activation gene 3(+) CD4(+) regulatory T cells induced by mucosal B cells.

    Science.gov (United States)

    Chu, K-H; Chiang, B-L

    2015-05-01

    The induction of mucosal tolerance has been demonstrated to be an effective therapeutic approach for the treatment of allergic diseases. Our previous study demonstrated that Peyer's patch B cells could convert naive T cells into regulatory T cells (so-called Treg -of-B(P) cells); however, it is important to characterize this particular subset of Treg -of-B cells for future applications. This study aimed to investigate the role of lymphocyte activating gene 3 (LAG3) in mediating the regulatory function of Treg -of-B(P) cells induced by mucosal follicular B (FOB) cells. Microarray analysis and real-time polymerase chain reaction (PCR) were used to assess the gene expression pattern of Treg -of-B(P) cells. To evaluate the role of LAG3, the in-vitro suppressive function and the alleviation of airway inflammation in a murine model of asthma was assessed. Our data indicated that FOB cells isolated from Peyer's patches had the ability to generate more suppressive Treg -of-B cells with LAG3 expression, compared with CD23(lo) CD21(lo) B cells. LAG3 is not only a marker for Treg -of-B(P) cells, but also participate in the suppressive ability. Moreover, CCR4 and CCR6 could be detected on the LAG3(+) , not LAG3(-) , Treg -of-B(P) cells and would help cells homing to allergic lung. In the murine model of asthma, the adoptive transfer of LAG3(+) Treg -of-B(P) cells was able to sufficiently suppress T helper type 2 (Th2) cytokine production, eosinophil infiltration and alleviate asthmatic symptoms. LAG3 was expressed in Treg -of-B(P) cells and was also involved in the function of Treg -of-B(P) cells. In the future, this particular subset of Treg -of-B cells might be used to alleviate allergic symptoms. PMID:25581421

  17. Inhibition of COP9-signalosome (CSN) deneddylating activity and tumor growth of diffuse large B-cell lymphomas by doxycycline

    OpenAIRE

    Pulvino, Mary; Chen, Luojing; Oleksyn, David; Li, Jing; Compitello, George; Rossi, Randy; Spence, Stephen; Balakrishnan, Vijaya; Jordan, Craig; Poligone, Brian; Casulo, Carla; Burack, Richard; Shapiro, Joel L.; Bernstein, Steven; Friedberg, Jonathan W.

    2015-01-01

    In searching for small-molecule compounds that inhibit proliferation and survival of diffuse large B-cell lymphoma (DLBCL) cells and may, therefore, be exploited as potential therapeutic agents for this disease, we identified the commonly used and well-tolerated antibiotic doxycycline as a strong candidate. Here, we demonstrate that doxycycline inhibits the growth of DLBCL cells both in vitro and in mouse xenograft models. In addition, we show that doxycycline accumulates in DLBCL cells to hi...

  18. Platelet-activating factor induces phospholipid turnover, calcium flux, arachidonic acid liberation, eicosanoid generation, and oncogene expression in a human B cell line

    International Nuclear Information System (INIS)

    Platelet-activating factor is a potent mediator of the inflammatory response. Studies of the actions of platelet-activating factor have centered mainly around neutrophils, monocytes, and platelets. In this report we begin to uncover the influence of platelet-activating factor on B lymphocytes. Employing the EBV-transformed human B cell line SKW6.4, we demonstrate that platelet-activating factor significantly alters membrane phospholipid metabolism indicated by the incorporation of 32P into phosphatidylcholine, phosphatidylinositol, and phosphatidic acid but not significantly into phosphatidylethanolamine at concentrations ranging from 10(-9) to 10(-6) M. The inactive precursor, lyso-platelet-activating factor, at a concentration as high as 10(-7) M had no effect on any of the membrane phospholipids. We also show that platelet-activating factor from 10(-12) to 10(-6) M induced rapid and significant elevation in intracellular calcium levels, whereas lyso-platelet-activating factor was again ineffective. We further demonstrate the impact of platelet-activating factor binding to B cells by measuring platelet-activating factor induced arachidonic acid release and 5-hydroxyeicosatetraenoic acid production. Moreover, platelet-activating factor was capable of inducing transcription of the nuclear proto-oncogenes c-fos and c-jun. Finally we explored the possible role of 5-hydroxyeicosatetraenoic acid as a regulator of arachidonic acid liberation demonstrating that endogenous 5-lipoxygenase activity modulates platelet-activating factor induced arachidonic acid release perhaps acting at the level of phospholipase A2. In summary, platelet-activating factor is shown here to have a direct and profound effect on a pure B cell line

  19. Primary cutaneous b-cell lymphoma successfully treated with highly active antiretroviral therapy alone: A case report and review of the literature

    Directory of Open Access Journals (Sweden)

    María F Villafañe

    2011-01-01

    Full Text Available Cutaneous B-cell lymphoma (CBCL is an unusual skin neoplasm with a great range of clinical presentations. Here, we report a case of CBCL in an AIDS patient presented as a single and nodular/ulcerative lesion in the perianal area. The patient was started on highly active antiretroviral therapy alone with a good clinical and oncological response. Two years later, the patient is asymptomatic with undetectable viral load and immune reconstitution.

  20. Immunological Activities of Purified Preparations of Enterobacterial Common Antigen

    OpenAIRE

    Gannon, Patrick J.; Jacobs, Diane M.; Marx, Arthur; Mayer, Hubert; Romanowska, Elzbieta; Neter, Erwin

    1982-01-01

    The immunological activities of three purified preparations of enterobacterial common antigen (ECA) obtained by different procedures were studied. ECA-Ma (method of A. Marx) was from Salmonella typhimurium TV149 (Ra mutant), ECA-My (method of H. Mayer) was from S. montevideo, and ECA-Ro (method of E. Romanowska) was from Shigella sonnei phase I. These preparations, on a weight basis, neutralized similar amounts of ECA antibodies, indicating that the serological activities were comparable. Nei...

  1. Impact of Gastrointestinal Bacillus anthracis Infection on Hepatic B Cells

    Directory of Open Access Journals (Sweden)

    Natacha Colliou

    2015-09-01

    Full Text Available Ingestion of Bacillus anthracis results in rapid gastrointestinal (GI infection, known as GI anthrax. We previously showed that during GI anthrax, there is swift deterioration of intestinal barrier function leading to translocation of gut-associated bacteria into systemic circulation. Additionally, we described dysfunction in colonic B cells. In concordance with our previous studies, here, we report early migration of the Sterne strain of B. anthracis along with other gut-resident bacteria into the infected murine liver. Additionally, despite a global decrease in the B cell population, we observed an increase in both B-1a and marginal zone (MZ-like B cells. Both of these cell types are capable of producing immunoglobulins against common pathogens and commensals, which act as a general antibody barrier before an antigen-specific antibody response. Accumulation of these cells in the liver was associated with an increase in chemokine expression. These data suggest that the presence of Sterne and other commensals in the liver trigger migration of MZ-like B cells from the spleen to the liver to neutralize systemic spread. Further research is required to evaluate the possible cause of their failure to clear the infection within the liver, including the potential role of dysfunctional mitogen-activated protein kinase (MAPK signaling.

  2. Activation of TAK1 by MYD88 L265P drives malignant B-cell Growth in non-Hodgkin lymphoma

    International Nuclear Information System (INIS)

    Massively parallel sequencing analyses have revealed a common mutation within the MYD88 gene (MYD88L265P) occurring at high frequencies in many non-Hodgkin lymphomas (NHLs) including the rare lymphoplasmacytic lymphoma, Waldenström's macroglobulinemia (WM). Using whole-exome sequencing, Sanger sequencing and allele-specific PCR, we validate the initial studies and detect the MYD88L265P mutation in the tumor genome of 97% of WM patients analyzed (n=39). Due to the high frequency of MYD88 mutation in WM and other NHL, and its known effects on malignant B-cell survival, therapeutic targeting of MYD88 signaling pathways may be clinically useful. However, we are lacking a thorough characterization of the role of intermediary signaling proteins on the biology of MYD88L265P-expressing B cells. We report here that MYD88L265P signaling is constitutively active in both WM and diffuse large B-cell lymphoma cells leading to heightened MYD88L265P, IRAK and TRAF6 oligomerization and NF-κB activation. Furthermore, we have identified the signaling protein, TAK1, to be an essential mediator of MYD88L265P-driven signaling, cellular proliferation and cytokine secretion in malignant B cells. Our studies highlight the biological significance of MYD88L265P in NHL and reveal TAK1 inhibition to be a potential therapeutic strategy for the treatment of WM and other diseases characterized by MYD88L265P

  3. Requirement of 8-mercaptoguanosine as a costimulus for IL-4-dependent μ to γ1 class switch recombination in CD38-activated B cells

    International Nuclear Information System (INIS)

    Mature B-2 cells expressing surface IgM and IgD proliferate upon stimulation by CD38, CD40 or lipopolysaccharide (LPS) and differentiate into IgG1-producing plasma cells in the presence of cytokines. The process of class switch recombination (CSR) from IgM to other isotypes is highly regulated by cytokines and activation-induced cytidine deaminase (AID). Blimp-1 and XBP-1 play an essential role in the terminal differentiation of switched B-2 cells to Ig-producing plasma cells. IL-5 induces AID and Blimp-1 expression in CD38- and CD40-activated B-2 cells, leading to μ to γ1 CSR at DNA level and IgG1 production. IL-4, a well-known IgG1-inducing factor, does not induce μ to γ1 CSR in CD38-activated B-2 cells or Blimp-1, while IL-4 induces μ to γ1 CSR, XBP-1 expression, and IgG1 production expression in CD40-activated B-2 cells. Interestingly, the addition of 8-mercaptoguanosine (8-SGuo) with IL-4 to the culture of CD38-activated B cells can induce μ to γ1 CSR, Blimp-1 expression, and IgG1 production. Intriguingly, 8-SGuo by itself induces AID expression in CD38-activated B cells. However, it does not induce μ to γ1 CSR. These results imply that the mode of B-cell activation for extracellular stimulation affects the outcome of cytokine stimulation with respect to the efficiency and direction of CSR, and the requirements of the transcriptional regulator and the generation of antibody-secreting cells. Furthermore, our data suggest the requirement of additional molecules in addition to AID for CSR

  4. Engagement of CD22 on B cells with the monoclonal antibody epratuzumab stimulates the phosphorylation of upstream inhibitory signals of the B cell receptor.

    Science.gov (United States)

    Lumb, Simon; Fleischer, Sarah J; Wiedemann, Annika; Daridon, Capucine; Maloney, Alison; Shock, Anthony; Dörner, Thomas

    2016-06-01

    The binding of antigen to the B cell receptor (BCR) results in a cascade of signalling events that ultimately drive B cell activation. Uncontrolled B cell activation is regulated by negative feedback loops that involve inhibitory co-receptors such as CD22 and CD32B that exert their functions following phosphorylation of immunoreceptor tyrosine-based inhibition motifs (ITIMs). The CD22-targeted antibody epratuzumab has previously been shown to inhibit BCR-driven signalling events, but its effects on ITIM phosphorylation of CD22 and CD32B have not been properly evaluated. The present study therefore employed both immunoprecipitation and flow cytometry approaches to elucidate the effects of epratuzumab on direct phosphorylation of key tyrosine (Tyr) residues on both these proteins, using both transformed B cell lines and primary human B cells. Epratuzumab induced the phosphorylation of Tyr(822) on CD22 and enhanced its co-localisation with SHP-1. Additionally, in spite of high basal phosphorylation of other key ITIMs on CD22, in primary human B cells epratuzumab also enhanced phosphorylation of Tyr(807), a residue involved in the recruitment of Grb2. Such initiation events could explain the effects of epratuzumab on downstream signalling in B cells. Finally, we were able to demonstrate that epratuzumab stimulated the phosphorylation of Tyr(292) on the low affinity inhibitory Fc receptor CD32B which would further attenuate BCR-induced signalling. Together, these data demonstrate that engagement of CD22 with epratuzumab leads to the direct phosphorylation of key upstream inhibitory receptors of BCR signalling and may help to explain how this antibody modulates B cell function. PMID:27125377

  5. Progesterone and estradiol exert an inhibitory effect on the production of anti-inflammatory cytokine IL-10 by activated MZ B cells.

    Science.gov (United States)

    Bommer, I; Muzzio, D O; Zygmunt, M; Jensen, F

    2016-08-01

    The main message of this work is the fact that female sex hormones, progesterone and estradiol, whose levels significantly rise during pregnancy, inhibit the production of anti-inflammatory cytokine IL-10 with no apparent effect on pro-inflammatory cytokine TNF-α by activated MZ B cells. This is an important piece of information and helps to better understand how the maternal immune system controls the balance between immune tolerance and immune activation during pregnancy leading to the simultaneously acceptance of the semi-allogeneic fetus and the proper defense of the mother against pathogens during this critical period of time. PMID:27317920

  6. Zinc-mediated regulation of caspases activity: dose-dependent inhibition or activation of caspase-3 in the human Burkitt lymphoma B cells (Ramos).

    Science.gov (United States)

    Schrantz, N; Auffredou, M T; Bourgeade, M F; Besnault, L; Leca, G; Vazquez, A

    2001-02-01

    Divalent cations, including Zinc and Manganese ions, are important modulators of cell activation. We investigated the ability of these two divalent cations to modulate apoptosis in human Burkitt lymphoma B cells line (Ramos). We found that Zinc (from 10 to 50 microM) inhibited Manganese-induced caspase-3 activation and apoptosis of Ramos cells. Higher concentration of Zinc (50 to 100 microM) did not prevent Manganese-mediated apoptosis but rather increased cell death among Ramos cells. This Zinc-mediated cell death was associated with apoptotic features such as cell shrinkage, the presence of phosphatidylserine residues on the outer leaflet of the cells, chromatin condensation, DNA fragmentation and decrease of mitochondrial transmembrane potential. Zinc-mediated apoptosis was associated with caspase-9 and caspase-3 activation as revealed by the appearance of active p35 fragment of caspase-9 and p19 and p17 of caspase-3 as well as in vivo cleavage of PARP and of a cell-permeable fluorogenic caspase-3 substrate (Phiphilux-G(1)D(2)). Both Zinc-mediated apoptosis and caspase-3 activation were prevented by the cell-permeable, broad-spectrum inhibitor of caspases (zVAD-fmk) or overexpression of bcl-2. In addition, we show that Zinc-induced loss of transmembrane mitochondrial potential is a caspase-independent event, since it is not modified by the presence of zVAD-fmk, which is inhibited by overexpression of bcl-2. These results indicate that depending on its concentration, Zinc can exert opposite effects on caspase-3 activation and apoptosis in human B lymphoma cells: concentrations below 50 microM inhibit caspase-3 activation and apoptosis whereas higher concentrations of Zinc activate a death pathway associated with apoptotic-like features and caspase-3 activation. PMID:11313717

  7. Caspase-1 Dependent IL-1β Secretion and Antigen-Specific T-Cell Activation by the Novel Adjuvant, PCEP

    Directory of Open Access Journals (Sweden)

    Sunita Awate

    2014-06-01

    Full Text Available The potent adjuvant activity of the novel adjuvant, poly[di(sodiumcarboxylatoethylphenoxyphosphazene] (PCEP, with various antigens has been reported previously. However, very little is known about its mechanisms of action. We have recently reported that intramuscular injection of PCEP induces NLRP3, an inflammasome receptor gene, and inflammatory cytokines, including IL-1β and IL-18, in mouse muscle tissue. Caspase-1 is required for the processing of pro-forms of IL-1β and IL-18 into mature forms and is a critical constituent of the NLRP3 inflammasome. Hence, in the present study, we investigated the role of caspase-1 in the secretion of IL-1β and IL-18 in PCEP-stimulated splenic dendritic cells (DCs. Caspase inhibitor YVAD-fmk-treated splenic DCs showed significantly reduced IL-1β and IL-18 secretion in response to PCEP stimulation. Further, PCEP had no effect on the expression of MHC class II or co-stimulatory molecules, CD86 and CD40, suggesting that PCEP does not induce DC maturation. However, PCEP directly activated B-cells to induce significant production of IgM. In addition, PCEP+ovalbumin (OVA immunized mice showed significantly increased production of antigen-specific IFN-γ by CD4+ and CD8+ T-cells. We conclude that PCEP activates innate immunity, leading to increased antigen-specific T-cell responses.

  8. Sleeping Beauty transposon screen identifies signaling modules that cooperate with STAT5 activation to induce B-cell acute lymphoblastic leukemia.

    Science.gov (United States)

    Heltemes-Harris, L M; Larson, J D; Starr, T K; Hubbard, G K; Sarver, A L; Largaespada, D A; Farrar, M A

    2016-06-30

    Signal transducer and activator of transcription 5 (STAT5) activation occurs frequently in human progenitor B-cell acute lymphoblastic leukemia (B-ALL). To identify gene alterations that cooperate with STAT5 activation to initiate leukemia, we crossed mice expressing a constitutively active form of STAT5 (Stat5b-CA) with mice in which a mutagenic Sleeping Beauty transposon (T2/Onc) was mobilized only in B cells. Stat5b-CA mice typically do not develop B-ALL (<2% penetrance); in contrast, 89% of Stat5b-CA mice in which the T2/Onc transposon had been mobilized died of B-ALL by 3 months of age. High-throughput sequencing approaches were used to identify genes frequently targeted by the T2/Onc transposon; these included Sos1 (74%), Kdm2a (35%), Jak1 (26%), Bmi1 (19%), Prdm14 or Ncoa2 (13%), Cdkn2a (10%), Ikzf1 (8%), Caap1 (6%) and Klf3 (6%). Collectively, these mutations target three major cellular processes: (i) the Janus kinase/STAT5 pathway (ii) progenitor B-cell differentiation and (iii) the CDKN2A tumor-suppressor pathway. Transposon insertions typically resulted in altered expression of these genes, as well as downstream pathways including STAT5, extracellular signal-regulated kinase (Erk) and p38. Importantly, expression of Sos1 and Kdm2a, and activation of p38, correlated with survival, further underscoring the role these genes and associated pathways have in B-ALL. PMID:26500062

  9. Increase in intracellular calcium and activation of protein kinase C in the expression of lymphokine receptors and the proliferation of murine B-cells

    Energy Technology Data Exchange (ETDEWEB)

    Proust, J.; Filburn, C.; Chrest, F.; Buchholz, M.; Nordin, A.

    1986-03-01

    BCGF dependent B cell proliferation requires costimulation by anti-..mu.. whereas IL2 dependent B cell proliferation requires preactivation by both anti-..mu.. and LPS. These studies show that in the BCGF dependent pathway the activation signal delivered by anti-..mu.. can be mimicked by an increase in intracellular calcium ((Ca/sup 2 +/)i) induced by the calcium ionophore, Ionomycin. In the IL2 dependent pathway, the additional signal delivered by LPS that leads to IL2 receptors expression and cell proliferation, can be replaced by the phorbol ester PMA as well as the diacylglycerol sn-1,2-dioctanoyl glycerol (DiC8), both shown to induce protein kinase C (PKc) activation. In agreement with these observations they show that LPS activates PKc with no concomitant increase in intracellular calcium ((Ca/sup 2 +/)i), while anti-..mu.. induces an increase in (Ca/sup 2 +/)i but no PKc activation. (Ca/sup 2 +/)i increase and PKc activation are both critical for the expression of IL2 receptor and the subsequent IL2 proliferation whereas an increase in (Ca/sup 2 +/)i is sufficient to induce BCGF dependent proliferation. These results suggest that the activation signals leading to BCGF and IL2 dependent proliferation proceed through 2 different biochemical pathways.

  10. Changes in Blood B Cell-Activating Factor (BAFF Levels in Multiple Sclerosis: A Sign of Treatment Outcome.

    Directory of Open Access Journals (Sweden)

    Karin Kannel

    Full Text Available Multiple sclerosis (MS is mediated primarily by autoreactive T cells. However, evidence suggesting the involvement of humoral immunity in brain diseases has increased interest in the role of B cells and their products during MS pathogenesis. The major survival factor for B cells, BAFF has been shown to play a role in several autoimmune conditions. Elevated BAFF levels have been reported in MS animal model and during MS relapse in patients. Moreover, disease-modifying treatments (DMT reportedly influence blood BAFF levels in MS patients, but the significance of these changes remains unclear. The present study addresses how blood BAFF levels are associated with the clinical course of relapsing-remitting MS and the effectiveness of DMT and short-term steroid treatment. During a prospective longitudinal follow-up of 2.3 years, BAFF was measured in the blood of 170 MS patients in the stable phase and within 186 relapses. BAFF levels were significantly higher in MS patients compared to healthy controls. However, stable MS patients without relapses exhibited significantly higher BAFF levels than relapsing patients. Treatment with interferon-β and immunosuppressants raised BAFF blood levels. Interestingly, a similar effect was not seen in patients treated with glatiramer acetate. Short-term treatment with high doses of intravenous methylprednisolone did not significantly alter plasma BAFF levels in 65% of relapsing-remitting MS patients. BAFF were correlated weakly but significantly with monocyte and basophil counts, but not with other blood cell types (neutrophils, lymphocytes, or eosinophils or inflammatory biomarkers. To our knowledge, this is the first report demonstrating that higher blood BAFF levels may reflect a more stable and effective MS treatment outcome. These results challenge hypotheses suggesting that elevated blood BAFF levels are associated with more severe disease presentation and could explain the recent failure of pharmaceutical

  11. Ectopic T Cell Receptor-α Locus Control Region Activity in B Cells Is Suppressed by Direct Linkage to Two Flanking Genes at Once

    Science.gov (United States)

    Andino, Blanca E.; Harrow, Faith; Erhard, Karl F.; Kovalovsky, Damian; Sant'Angelo, Derek B.; Ortiz, Benjamin D.

    2010-01-01

    The molecular mechanisms regulating the activity of the TCRα gene are required for the production of the circulating T cell repertoire. Elements of the mouse TCRα locus control region (LCR) play a role in these processes. We previously reported that TCRα LCR DNA supports a gene expression pattern that mimics proper thymus-stage, TCRα gene-like developmental regulation. It also produces transcription of linked reporter genes in peripheral T cells. However, TCRα LCR-driven transgenes display ectopic transcription in B cells in multiple reporter gene systems. The reasons for this important deviation from the normal TCRα gene regulation pattern are unclear. In its natural locus, two genes flank the TCRα LCR, TCRα (upstream) and Dad1 (downstream). We investigated the significance of this gene arrangement to TCRα LCR activity by examining transgenic mice bearing a construct where the LCR was flanked by two separate reporter genes. Surprisingly, the presence of a second, distinct, reporter gene downstream of the LCR virtually eliminated the ectopic B cell expression of the upstream reporter observed in earlier studies. Downstream reporter gene activity was unaffected by the presence of a second gene upstream of the LCR. Our findings indicate that a gene arrangement in which the TCRα LCR is flanked by two distinct transcription units helps to restrict its activity, selectively, on its 5′-flanking gene, the natural TCRα gene position with respect to the LCR. Consistent with these findings, a TCRα/Dad1 locus bacterial artificial chromosome dual-reporter construct did not display the ectopic upstream (TCRα) reporter expression in B cells previously reported for single TCRα transgenes. PMID:21124935

  12. Ectopic T cell receptor-α locus control region activity in B cells is suppressed by direct linkage to two flanking genes at once.

    Directory of Open Access Journals (Sweden)

    Stefan Knirr

    Full Text Available The molecular mechanisms regulating the activity of the TCRα gene are required for the production of the circulating T cell repertoire. Elements of the mouse TCRα locus control region (LCR play a role in these processes. We previously reported that TCRα LCR DNA supports a gene expression pattern that mimics proper thymus-stage, TCRα gene-like developmental regulation. It also produces transcription of linked reporter genes in peripheral T cells. However, TCRα LCR-driven transgenes display ectopic transcription in B cells in multiple reporter gene systems. The reasons for this important deviation from the normal TCRα gene regulation pattern are unclear. In its natural locus, two genes flank the TCRα LCR, TCRα (upstream and Dad1 (downstream. We investigated the significance of this gene arrangement to TCRα LCR activity by examining transgenic mice bearing a construct where the LCR was flanked by two separate reporter genes. Surprisingly, the presence of a second, distinct, reporter gene downstream of the LCR virtually eliminated the ectopic B cell expression of the upstream reporter observed in earlier studies. Downstream reporter gene activity was unaffected by the presence of a second gene upstream of the LCR. Our findings indicate that a gene arrangement in which the TCRα LCR is flanked by two distinct transcription units helps to restrict its activity, selectively, on its 5'-flanking gene, the natural TCRα gene position with respect to the LCR. Consistent with these findings, a TCRα/Dad1 locus bacterial artificial chromosome dual-reporter construct did not display the ectopic upstream (TCRα reporter expression in B cells previously reported for single TCRα transgenes.

  13. Network-of-queues approach to B-cell-receptor affinity discrimination

    Science.gov (United States)

    Felizzi, Federico; Comoglio, Federico

    2012-06-01

    The immune system is one of the most complex signal processing machineries in biology. The adaptive immune system, consisting of B and T lymphocytes, is activated in response to a large spectrum of pathogen antigens. B cells recognize and bind the antigen through B-cell receptors (BCRs) and this is fundamental for B-cell activation. However, the system response is dependent on BCR-antigen affinity values that span several orders of magnitude. Moreover, the ability of the BCR to discriminate between affinities at the high end (e.g., 109M-1-1010M-1) challenges the formulation of a mathematical model able to robustly separate these affinity-dependent responses. Queuing theory enables the analysis of many related processes, such as those resulting from the stochasticity of protein binding and unbinding events. Here we define a network of queues, consisting of BCR early signaling states and transition rates related to the propensity of molecular aggregates to form or disassemble. By considering the family of marginal distributions of BCRs in a given signaling state, we report a significant separation (measured as Jensen-Shannon divergence) that arises from a broad spectrum of antigen affinities.

  14. Proliferation of resting B cells is modulated by CR2 and CR1.

    Science.gov (United States)

    Fingeroth, J D; Heath, M E; Ambrosino, D M

    1989-06-15

    Absence of the third component of complement, C3, is associated with impaired ability to synthesize antibody, particularly in the presence of limiting antigen [1-9]. The mature B lymphocyte bearing the surface immunoglobulin receptor transduces signals for proliferation and differentiation upon binding of specific antigen. This mature B cell also bears two related membrane proteins, CR2 (the C3d/Epstein-Barr virus receptor) (CD35) [15], which can mediate the binding of ligands to which appropriate cleavage fragments of C3 have become attached [16]. It has been suggested that these receptors play a direct role(s) in B cell activation [17-25]. In light of previous in vivo observations we decided to assess the function of CR2 and CR1 in relation to B cell activation through the membrane IgM receptor. Highly purified splenic B cells were prepared. No contaminating T cells or macrophages were detected by flow cytometric analysis and no proliferative activity was present upon PHA or ConA stimulation of the purified cells. The B cells were separated into low (activated), medium (preactivated) and high density (resting) fractions by Percoll gradient density centrifugation [26]. The responses of the B cell subpopulations to various concentrations of anti mu (DA4.4 monoclonal antibody) [27] were examined for proliferation at 72 h and for IgM/IgG production at 7 days. Low density B cells were maximally stimulated and no concentration of anti-mu was effective in enhancing their responses. High density B cells proliferated to anti-mu in a concentration dependent manner. When substimulatory concentrations of anti-mu were employed, concomitant crosslinking of CR2 (with either of 2 distinct monoclonal antibodies HB-5 [28] or OKB7 [17]) resulted in a 45% enhancement of B cell proliferation above that observed by crosslinking of SIgM alone. In these studies, total IgM and IgG did not increase in the absence of T cells or T cell factors, indicating that terminal differentiation did

  15. The strength of the antibody response to the nematode Ascaris lumbricoides inversely correlates with levels of B-Cell Activating Factor (BAFF)

    OpenAIRE

    Bornacelly, Adriana; Mercado, Dilia; Acevedo, Nathalie; Caraballo, Luis

    2014-01-01

    Background B-Cell Activating Factor (BAFF) is a cytokine regulating antibody production. Polymorphisms in the gene encoding BAFF were associated with the antibody response to Ascaris but not to mite allergens. In the present study we evaluated the relationship between BAFF and specific antibodies against Ascaris and mites in 448 controls and 448 asthmatics. Soluble BAFF was measured by ELISA and BAFF mRNA by qPCR. Surface expression of BAFF and its receptor (BAFF-R) was analyzed by flow cytom...

  16. Adaptive Response of T and B Cells in Atherosclerosis.

    Science.gov (United States)

    Ketelhuth, Daniel F J; Hansson, Göran K

    2016-02-19

    Atherosclerosis is a chronic inflammatory disease that is initiated by the retention and accumulation of cholesterol-containing lipoproteins, particularly low-density lipoprotein, in the artery wall. In the arterial intima, lipoprotein components that are generated through oxidative, lipolytic, and proteolytic activities lead to the formation of several danger-associated molecular patterns, which can activate innate immune cells as well as vascular cells. Moreover, self- and non-self-antigens, such as apolipoprotein B-100 and heat shock proteins, can contribute to vascular inflammation by triggering the response of T and B cells locally. This process can influence the initiation, progression, and stability of plaques. Substantial clinical and experimental data support that the modulation of adaptive immune system may be used for treating and preventing atherosclerosis. This may lead to the development of more selective and less harmful interventions, while keeping host defense mechanisms against infections and tumors intact. Approaches such as vaccination might become a realistic option for cardiovascular disease, especially if they can elicit regulatory T and B cells and the secretion of atheroprotective antibodies. Nevertheless, difficulties in translating certain experimental data into new clinical therapies remain a challenge. In this review, we discuss important studies on the function of T- and B-cell immunity in atherosclerosis and their manipulation to develop novel therapeutic strategies against cardiovascular disease. PMID:26892965

  17. Down-regulation of Sonic hedgehog signaling pathway activity is involved in 5-fluorouracil-induced apoptosis and motility inhibition in Hep3B cells

    Institute of Scientific and Technical Information of China (English)

    Qiyu Wang; Shuhong Huang; Ling Yang; Ling Zhao; Yuxia Yin; Zhongzhen Liu; Zheyu Chen; Hongwei Zhang

    2008-01-01

    The Sonic hedgehog (SHh) pathway plays a critical role in normal embryogenesis and carcinogenesis, but its function in cancer cells treated with 5-fluorouracil (5-FU) remains unknown. We examined the expression of a subset of SHh signaling pathway genes, including SHh, SMO, PTC1, Su(Fu) and HIP in human hepatocellular carcinoma (HCC) cell lines,Hep3B and HepG2, treated with 5-FU by reverse transcriptionpolymerase chain reaction. Using trypan blue analysis,3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide assay and terminal deoxynucleotidyl transferase-mediated digoxigenin-dUTP nick-end labeling assay, we also detected the apoptosis of Hep3B cells resulting from the transfection of pCS2-Gli1 expression vector combined with 5-FU treatment.The motility of the cells was detected by scratch wound closure assay. The expression and subcellular location of PTC1 protein in Hep3B cells treated by 5-FU were also investigated by Western blot analysis and immunofluorescent microscopy. The results indicated that the expression of SHh pathway target molecules at both messenger RNA and protein levels are evidently down-regulated in Hep3B cells treated with 5-FU. The overexpression of Gli1 restores cell viability and, to some extent, the migration abilities inhibited by 5-FU.Furthermore, 5-FU treatment affects the subcellular localization of PTC1 protein, a key member in SHh signaling pathway. Our data showed that the down-regulation of SHh signaling pathway activity was involved in 5-FU-induced apoptosis and the inhibition of motility in hedgehog-activated HCC cell lines. This implies that the combination of SHh signaling pathway inhibitor and 5-FU-based chemotherapy might represent a more promising strategy against HCC.

  18. Congenital B cell lymphocytosis explained by novel germline CARD11 mutations.

    Science.gov (United States)

    Snow, Andrew L; Xiao, Wenming; Stinson, Jeffrey R; Lu, Wei; Chaigne-Delalande, Benjamin; Zheng, Lixin; Pittaluga, Stefania; Matthews, Helen F; Schmitz, Roland; Jhavar, Sameer; Kuchen, Stefan; Kardava, Lela; Wang, Wei; Lamborn, Ian T; Jing, Huie; Raffeld, Mark; Moir, Susan; Fleisher, Thomas A; Staudt, Louis M; Su, Helen C; Lenardo, Michael J

    2012-11-19

    Nuclear factor-κB (NF-κB) controls genes involved in normal lymphocyte functions, but constitutive NF-κB activation is often associated with B cell malignancy. Using high-throughput whole transcriptome sequencing, we investigated a unique family with hereditary polyclonal B cell lymphocytosis. We found a novel germline heterozygous missense mutation (E127G) in affected patients in the gene encoding CARD11, a scaffolding protein required for antigen receptor (AgR)-induced NF-κB activation in both B and T lymphocytes. We subsequently identified a second germline mutation (G116S) in an unrelated, phenotypically similar patient, confirming mutations in CARD11 drive disease. Like somatic, gain-of-function CARD11 mutations described in B cell lymphoma, these germline CARD11 mutants spontaneously aggregate and drive constitutive NF-κB activation. However, these CARD11 mutants rendered patient T cells less responsive to AgR-induced activation. By reexamining this rare genetic disorder first reported four decades ago, our findings provide new insight into why activating CARD11 mutations may induce B cell expansion and preferentially predispose to B cell malignancy without dramatically perturbing T cell homeostasis. PMID:23129749

  19. Suppression of unprimed T and B cells in antibody responses by irradiation-resistant and plastic-adherent suppressor cells in Toxoplasma gondii-infected mice

    International Nuclear Information System (INIS)

    In the acute phase of Toxoplasma infection, the function of both helper T and B cells was suppressed in primary antibody responses to dinitrophenol (DNP)-conjugated protein antigens. During the course of infection, the suppressive effect on T cells seems to continue longer than that on B cells, since suppression in responses to sheep erythrocytes, a T-dependent antigen, persisted longer than those to DNP-Ficoll, a T-independent antigen. Plastic-adherent cells from the spleens of Toxoplasma-infected and X-irradiated (400 rads) mice had strong suppressor activity in primary anti-sheep erythrocyte antibody responses of normal mouse spleen cells in vitro. These data suggest that the activation of irradiation-resistant and plastic-adherent suppressor cells causes the suppression of both T and B cells in Toxoplasma-infected mice

  20. TLR9-induced miR-155 and Ets-1 decrease expression of CD1d on B cells in SLE.

    Science.gov (United States)

    Liu, Fei; Fan, Hongye; Ren, Deshan; Dong, Guanjun; Hu, Erling; Ji, Jianjian; Hou, Yayi

    2015-07-01

    B cells present lipid antigens to CD1d-restricted invariant natural killer T (iNKT) cells to maintain autoimmune tolerance, and this process is disrupted in systemic lupus erythematosus (SLE). Inflammation may inhibit CD1d expression to exacerbate the pathology of lupus. However, how inflammation regulates CD1d expression on B cells is unclear in SLE. In the present study, we showed that the surface expression of CD1d on B cells from SLE mice was decreased and that stimulation of inflammatory responses through TLR9 decreased the membrane and total CD1d levels of CD1d on B cells. Moreover, inflammation-related microRNA-155 (miR-155) negatively correlated with the expression of CD1d in B cells. miR-155 directly targeted the 3'-untranslated region (3'-UTR) of CD1d upon TLR9 activation in both humans and mice. The inhibitory effects of miR-155 on CD1d expression in B cells impaired their antigen-presenting capacity to iNKT cells. In addition, Ets-1, a susceptibility gene of SLE, also directly regulated the expression of the CD1d gene at the transcriptional level. These findings provide new insight into the mechanism underlying decreased CD1d expression on B cells in SLE, suggesting that inhibition of inflammation may increase CD1d expression in B cells to ameliorate SLE via modulating iNKT cells. PMID:25929465

  1. Fluctuations in anti-nRNP levels in patients with mixed connective tissue disease are related to disease activity as part of a polyclonal B cell response.

    OpenAIRE

    Houtman, P M; Kallenberg, C. G.; Limburg, P C; Leeuwen, M.A. van; van Rijswijk, M H; The, T H

    1986-01-01

    In a follow up study of 11 patients with mixed connective tissue disease the levels of antibodies to nuclear ribonucleoprotein (nRNP) as measured by an enzyme linked immunosorbent assay (ELISA) were related to clinical activity of disease. To assess the relation between anti-nRNP levels and disease activity the levels of total immunoglobulin G, IgM rheumatoid factor (IgM RF), and antibodies to an unrelated antigen (tetanus toxoid) were determined simultaneously. No significant changes in anti...

  2. B-Cell Response during Protozoan Parasite Infections

    OpenAIRE

    Amezcua Vesely, María C; Bermejo, Daniela A.; Montes, Carolina L.; Acosta-Rodríguez, Eva V.; Adriana Gruppi

    2012-01-01

    In this review, we discuss how protozoan parasites alter immature and mature B cell compartment. B1 and marginal zone (MZ) B cells, considered innate like B cells, are activated during protozoan parasite infections, and they generate short lived plasma cells providing a prompt antibody source. In addition, protozoan infections induce massive B cell response with polyclonal activation that leads to hypergammaglobulnemia with serum antibodies specific for the parasites and self and/or non relat...

  3. Polyanhydride Nanovaccines Induce Germinal Center B Cell Formation and Sustained Serum Antibody Responses.

    Science.gov (United States)

    Vela Ramirez, Julia E; Tygrett, Lorraine T; Hao, Jihua; Habte, Habtom H; Cho, Michael W; Greenspan, Neil S; Waldschmidt, Thomas J; Narasimhan, Balaji

    2016-06-01

    Biodegradable polymeric nanoparticle-based subunit vaccines have shown promising characteristics by enhancing antigen presentation and inducing protective immune responses when compared with soluble protein. Specifically, polyanhydride nanoparticle-based vaccines (i.e., nanovaccines) have been shown to successfully encapsulate and release antigens, activate B and T cells, and induce both antibody- and cell-mediated immunity towards a variety of immunogens. One of the characteristics of strong thymus-dependent antibody responses is the formation of germinal centers (GC) and the generation of GC B cells, which is part of the T helper cell driven cellular response. In order to further understand the role of nanovaccines in the induction of antigen-specific immune responses, their ability to induce germinal center B cell formation and isotype switching and the effects thereof on serum antibody responses were investigated in these studies. Polyanhydride nanovaccines based on 1,6-bis(p-carboxyphenoxy)hexane and 1,8-bis(p-carboxyphenoxy)-3,6-dioxaoctane were used to subcutaneously administer a viral antigen. GC B cell formation and serum antibody responses induced by the nanovaccines were compared to that induced by alum-based vaccine formulations. It was demonstrated that a single dose of polyanhydride nanovaccines resulted in the formation of robust GCs and serum antibody in comparison to that induced by the alum-based formulation. This was attributed to the sustained release of antigen provided by the nanovaccines. When administered in a multiple dose regimen, the highest post-immunization titer and GC B cell number was enhanced, and the immune response induced by the nanovaccines was further sustained. These studies provide foundational information on the mechanism of action of polyanhydride nanovaccines. PMID:27319223

  4. Murine T-lymphocyte activation by mycobacterial antigens

    International Nuclear Information System (INIS)

    There has been renewed interest in the diagnosis of tuberculosis and other mycobacterial infections in the United States. Effective immunity to mycobacterial infections, as well as diagnosis by the skin test, involves T-cells rather than antibodies. Studies currently underway use the new technologies of monoclonal antibodies and recombinant DNA to define better mycobacterial antigens for T-cell activation, in the hope of identifying species specific antigens. Lymph node cells from mice sensitized to Mycobacterium intracellulare and Mycobacterium avium were assayed for activation by mycobacterial fractions, and cell lines and clones were generated. Comparing BALB/c and B10 mice indicated better responses to M. avium sonicate by B10 mice. A recombinant gene product containing a M. intracellulare peptide was assayed with lymph node cells and indicated excellent T-cell stimulation in BALB/c lymph node cells and cell lines. However, assays using B10 T-cell clones have yet to detect responders to the recombinant protein. Future studies using synthetic epitopes produced by recombinant DNA techniques and defined by monoclonal antibodies are necessary for the identification of reactive T-cell epitopes that are potentially species specific. 4 refs, 7 figs, 1 tab

  5. Innate B cells: oxymoron or validated concept? [v1; ref status: indexed, http://f1000r.es/T4CAVP

    Directory of Open Access Journals (Sweden)

    Carl F Ware

    2012-08-01

    Full Text Available B lymphocytes promote the initial innate interferon response to viral pathogens without the need for antigen receptor activation. B cell dependent IFN production requires the cytokine, lymphotoxin-β. The LTβ pathway is well known to regulate lymphoid organogenesis and homeostasis by differentiating stromal cells and macrophages. However, in response to viral pathogens these same B cell-regulated populations rapidly produce type 1 interferons. Thus, B cells act as innate effector cells via LTβ homeostatic pathways, which serve as innate host barriers to viral pathogens.

  6. File list: ALL.Bld.10.AllAg.Pro-B_cells [Chip-atlas[Archive

    Lifescience Database Archive (English)

    Full Text Available ALL.Bld.10.AllAg.Pro-B_cells mm9 All antigens Blood Pro-B cells SRX1553109,SRX15531...9,SRX1143902 http://dbarchive.biosciencedbc.jp/kyushu-u/mm9/assembled/ALL.Bld.10.AllAg.Pro-B_cells.bed ...

  7. The Relationship between B-cell Epitope and Mimotope Sequences.

    Science.gov (United States)

    Zhang, Chunhua; Li, Yunyun; Tang, Weina; Zhou, Zhiguo; Sun, Pingping; Ma, Zhiqiang

    2016-01-01

    B-cell epitope is a group of residues which is on the surface of an antigen. It invokes humoral responses. Locating B-cell epitope is important for effective vaccine design, and the development of diagnostic reagents. Mimotope-based B-cell epitope prediction method is a kind of conformational B-cell epitope prediction, and the core idea of the method is mapping the mimotope sequences which are obtained from a random phage display library. However, current mimotope-based B-cell epitope prediction methods cannot maintain a high degree of satisfaction in the circumstances of employing only mimotope sequences. In this study, we did a multi-perspective analysis on parameters for conformational B-cell epitopes and characteristics between epitope and mimotope on a benchmark datasets which contains 67 mimotope sets, corresponding to 40 unique complex structures. In these 67 cases, there are 25 antigen-antibody complexes and 42 protein-protein interactions. We analyzed the two parts separately. The results showed the mimotope sequences do have some epitope features, but there are also some epitope properties that mimotope sequences do not contain. In addition, the numbers of epitope segments with different lengths were obviously different between the antigen-antibody complexes and the protein-protein interactions. This study reflects how similar do mimotope sequence and genuine epitopes have; and evaluates existing mimotope-based B-cell epitope prediction methods from a novel viewpoint. PMID:26715528

  8. Activity profile of the CA125 antigen towards human red blood cells

    Directory of Open Access Journals (Sweden)

    Mitić N.

    2010-01-01

    Full Text Available Starting from the mucin nature of the CA125 antigen and conditions associated with high serum concentrations, this study is an attempt to gain insight into its activity profile towards human erythrocytes. Carcinomaassociated and pregnancy-associated CA125 antigens were tested in agglutination/aggregation, adhesion and hemolysis assays. The results obtained indicated that CA125 antigens increased agglutination/aggregation and inhibited erythrocyte adhesion, but differed in their effective concentrations. Galectin-1 slightly modulated the effects observed. CA125 antigens had no effect on hemolysis. The activity profile of the CA125 antigen towards erythrocytes may have biomedical consequences in different microenvironments in relevant physiological and pathophysiological conditions.

  9. Cadmium induces carcinogenesis in BEAS-2B cells through ROS-dependent activation of PI3K/AKT/GSK-3β/β-catenin signaling

    Energy Technology Data Exchange (ETDEWEB)

    Son, Young-Ok; Wang, Lei; Poyil, Pratheeshkumar; Budhraja, Amit; Hitron, J. Andrew; Zhang, Zhuo [Graduate Center for Toxicology, College of Medicine, University of Kentucky, Lexington, KY (United States); Lee, Jeong-Chae [Graduate Center for Toxicology, College of Medicine, University of Kentucky, Lexington, KY (United States); School of Dentistry and Institute of Oral Biosciences (BK21 program), Research Center of Bioactive Materials, Chonbuk National University, Jeonju 561-756 (Korea, Republic of); Shi, Xianglin, E-mail: xshi5@email.uky.edu [Graduate Center for Toxicology, College of Medicine, University of Kentucky, Lexington, KY (United States)

    2012-10-15

    Cadmium has been widely used in industry and is known to be carcinogenic to humans. Although it is widely accepted that chronic exposure to cadmium increases the incidence of cancer, the mechanisms underlying cadmium-induced carcinogenesis are unclear. The main aim of this study was to investigate the role of reactive oxygen species (ROS) in cadmium-induced carcinogenesis and the signal transduction pathways involved. Chronic exposure of human bronchial epithelial BEAS-2B cells to cadmium induced cell transformation, as evidenced by anchorage-independent growth in soft agar and clonogenic assays. Chronic cadmium treatment also increased the potential of these cells to invade and migrate. Injection of cadmium-stimulated cells into nude mice resulted in the formation of tumors. In contrast, the cadmium-mediated increases in colony formation, cell invasion and migration were prevented by transfection with catalase, superoxide dismutase-1 (SOD1), or SOD2. In particular, chronic cadmium exposure led to activation of signaling cascades involving PI3K, AKT, GSK-3β, and β-catenin and transfection with each of the above antioxidant enzymes markedly inhibited cadmium-mediated activation of these signaling proteins. Inhibitors specific for AKT or β-catenin almost completely suppressed the cadmium-mediated increase in total and active β-catenin proteins and colony formation. Moreover, there was a marked induction of AKT, GSK-3β, β-catenin, and carcinogenic markers in tumor tissues formed in mice after injection with cadmium-stimulated cells. Collectively, our findings suggest a direct involvement of ROS in cadmium-induced carcinogenesis and implicate a role of AKT/GSK-3β/β-catenin signaling in this process. -- Highlights: ► Chronic exposure to cadmium induces carcinogenic properties in BEAS-2B cells. ► ROS involved in cadmium-induced tumorigenicity of BEAS-2B cells. ► Cadmium activates ROS-dependent AKT/GSK-3β/β-catenin-mediated signaling. ► ROS

  10. Cadmium induces carcinogenesis in BEAS-2B cells through ROS-dependent activation of PI3K/AKT/GSK-3β/β-catenin signaling

    International Nuclear Information System (INIS)

    Cadmium has been widely used in industry and is known to be carcinogenic to humans. Although it is widely accepted that chronic exposure to cadmium increases the incidence of cancer, the mechanisms underlying cadmium-induced carcinogenesis are unclear. The main aim of this study was to investigate the role of reactive oxygen species (ROS) in cadmium-induced carcinogenesis and the signal transduction pathways involved. Chronic exposure of human bronchial epithelial BEAS-2B cells to cadmium induced cell transformation, as evidenced by anchorage-independent growth in soft agar and clonogenic assays. Chronic cadmium treatment also increased the potential of these cells to invade and migrate. Injection of cadmium-stimulated cells into nude mice resulted in the formation of tumors. In contrast, the cadmium-mediated increases in colony formation, cell invasion and migration were prevented by transfection with catalase, superoxide dismutase-1 (SOD1), or SOD2. In particular, chronic cadmium exposure led to activation of signaling cascades involving PI3K, AKT, GSK-3β, and β-catenin and transfection with each of the above antioxidant enzymes markedly inhibited cadmium-mediated activation of these signaling proteins. Inhibitors specific for AKT or β-catenin almost completely suppressed the cadmium-mediated increase in total and active β-catenin proteins and colony formation. Moreover, there was a marked induction of AKT, GSK-3β, β-catenin, and carcinogenic markers in tumor tissues formed in mice after injection with cadmium-stimulated cells. Collectively, our findings suggest a direct involvement of ROS in cadmium-induced carcinogenesis and implicate a role of AKT/GSK-3β/β-catenin signaling in this process. -- Highlights: ► Chronic exposure to cadmium induces carcinogenic properties in BEAS-2B cells. ► ROS involved in cadmium-induced tumorigenicity of BEAS-2B cells. ► Cadmium activates ROS-dependent AKT/GSK-3β/β-catenin-mediated signaling. ► ROS

  11. Polyclonal B-cell activation by Neisseria meningitidis capsular polysaccharides elicit antibodies protective against Trypanosoma cruzi infection in vitro.

    Science.gov (United States)

    Oliveira, T G; Milani, S R; Travassos, L R

    1996-01-01

    A hyperimmune rabbit antiserum against group C Neisseria meningitidis agglutinated and lysed Trypanosoma cruzi metacyclic trypomastigotes in a complement-mediated reaction. Immunization of rabbits with the purified polysaccharide C from N. meningitidis and of human volunteers with the AC-polysaccharide vaccine against meningitis also resulted in antibody production cross-reactive with T. cruzi infective forms. The rabbit antibodies bound to parasites, lysed metacyclic forms, and recognized several components from lysates of cell-derived trypomastigotes. The sera from six human volunteers reacted with cell-cultured trypomastigotes in vitro, lysed these forms, and recognized glycoconjugates migrating diffusely on the top of immunoblots. One serum also reacted with the isolated mucin-like glycoconjugate carrying the Ssp-3 epitope from cell-derived trypomastigotes, but treatment with sialidase did not abolish this reactivity. The anti-AC human antiserum also protected against HeLa cell infection and markedly decreased the number of parasites liberated after cell burst. The polyclonal response that resulted from human immunization with N. meningitidis polysaccharides A and C comprised trypanolytic antibodies that recognized nonsialylated epitopes expressed on infective forms of the parasite. It is suggested that human AC vaccination could be potentially helpful as an adjuvant to a specific immunotherapy of Chagas disease, developed with native or recombinant antigens of the parasite. PMID:8811466

  12. Multiple antigen glycopeptides (MAGs) with Tn tumour antigens and incorporated adjuvant: synthesis and immunobiological activities

    Czech Academy of Sciences Publication Activity Database

    Ježek, Jan; Kelkar, Shripad; Vepřek, Pavel; Hajdůch, M.; Sejbal, J.; Trnka, T.

    Napoli : Edizioni Ziino, 2002 - (Benedetti, E.; Pedone, C.), s. 524-525 ISBN 88-900948-1-8. [Peptides 2002. European Peptide Symposium /27./. Sorrento (IT), 31.08.2002-06.09.2002] R&D Projects: GA ČR GA303/01/0690 Institutional research plan: CEZ:AV0Z4055905 Keywords : Tn antigen * multiple antigen glycopeptide * synthetic vaccine Subject RIV: CE - Biochemistry

  13. B cells and immunological tolerance.

    Science.gov (United States)

    Manjarrez-Orduño, Nataly; Quách, Tâm D; Sanz, Iñaki

    2009-02-01

    Work from multiple groups continues to provide additional evidence for the powerful and highly diverse roles, both protective and pathogenic, that B cells play in autoimmune diseases. Similarly, it has become abundantly clear that antibody-independent functions may account for the opposing influences that B cells exercise over other arms of the immune response and ultimately over autoimmunity itself. Finally, it is becoming apparent that the clinical impact of B-cell depletion therapy may be, to a large extent, determined by the functional balance between different B-cell subsets that may be generated by this therapeutic intervention. In this review, we postulate that our perspective of B-cell tolerance and our experimental approach to its understanding are fundamentally changed by this view of B cells. Accordingly, we first discuss current knowledge of B-cell tolerance conventionally defined as the censoring of autoantibody-producing B cells (with an emphasis on human B cells). Therefore, we discuss a different model that contemplates B cells not only as targets of tolerance but also as mediators of tolerance. This model is based on the notion that the onset of clinical autoimmune disease may require a B-cell gain-of-pathogenic function (or a B-cell loss-of-regulatory-function) and that accordingly, disease remission may depend on the restoration of the physiological balance between B-cell pathogenic and protective functions. PMID:19148217

  14. Irreversible dual inhibitory mode: the novel Btk inhibitor PLS-123 demonstrates promising anti-tumor activity in human B-cell lymphoma

    OpenAIRE

    Ding, Ning; Li, Xitao; Shi, Yunfei; Ping, Lingyan; Wu, Lina; Fu, Kai; Feng, Lixia; Zheng, Xiaohui; Song, Yuqin; Pan, Zhengying; Zhu, Jun

    2015-01-01

    The B-cell receptor (BCR) signaling pathway has gained significant attention as a therapeutic target in B-cell malignancies. Recently, several drugs that target the BCR signaling pathway, especially the Btk inhibitor ibrutinib, have demonstrated notable therapeutic effects in relapsed/refractory patients, which indicates that pharmacological inhibition of BCR pathway holds promise in B-cell lymphoma treatment. Here we present a novel covalent irreversible Btk inhibitor PLS-123 with more poten...

  15. Mode of Bioenergetic Metabolism during B Cell Differentiation in the Intestine Determines the Distinct Requirement for Vitamin B1

    Directory of Open Access Journals (Sweden)

    Jun Kunisawa

    2015-10-01

    Full Text Available Bioenergetic metabolism varies during cell differentiation, but details of B cell metabolism remain unclear. Here, we show the metabolic changes during B cell differentiation in the intestine, where B cells differentiate into IgA+ plasma cells (PCs. Naive B cells in the Peyer’s patches (PPs and IgA+ PCs in the intestinal lamina propria (iLP both used the tricarboxylic acid (TCA cycle, but only IgA+ PCs underwent glycolysis. These metabolic differences reflected their dependencies on vitamin B1, an essential cofactor for the TCA cycle. Indeed, the diminished activity of the TCA cycle after dietary vitamin B1 depletion decreased the number of naive B cells in PPs without affecting IgA+ PCs in the iLP. The maintenance of naive B cells by dietary vitamin B1 was required to induce—but not maintain—intestinal IgA responses against oral antigens. These findings reveal the diet-mediated maintenance of B cell immunometabolism in organized and diffuse intestinal tissues.

  16. Marginal zone B-cells, a gatekeeper of innate immunity.

    Science.gov (United States)

    Zouali, Moncef; Richard, Yolande

    2011-01-01

    To maintain the integrity of an organism constantly challenged by pathogens, the immune system is endowed with a variety of cell types. B lymphocytes were initially thought to only play a role in the adaptive branch of immunity. However, a number of converging observations revealed that two B-cell subsets, marginal zone (MZ) and B1 cells, exhibit unique developmental and functional characteristics, and can contribute to innate immune responses. In addition to their capacity to mount a local antibody response against type-2 T-cell-independent (TI-2) antigens, MZ B-cells can participate to T-cell-dependent (TD) immune responses through the capture and import of blood-borne antigens to follicular areas of the spleen. Here, we discuss the multiple roles of MZ B-cells in humans, non-human primates, and rodents. We also summarize studies - performed in transgenic mice expressing fully human antibodies on their B-cells and in macaques whose infection with Simian immunodeficiency virus (SIV) represents a suitable model for HIV-1 infection in humans - showing that infectious agents have developed strategies to subvert MZ B-cell functions. In these two experimental models, we observed that two microbial superantigens for B-cells (protein A from Staphylococcus aureus and protein L from Peptostreptococcus magnus) as well as inactivated AT-2 virions of HIV-1 and infectious SIV preferentially deplete innate-like B-cells - MZ B-cells and/or B1 B-cells - with different consequences on TI and TD antibody responses. These data revealed that viruses and bacteria have developed strategies to deplete innate-like B-cells during the acute phase of infection and to impair the antibody response. Unraveling the intimate mechanisms responsible for targeting MZ B-cells in humans will be important for understanding disease pathogenesis and for designing novel vaccine strategies. PMID:22566852

  17. Characterization of xenoantiserum produced against B cell acute lymphoblastic leukemia cell line

    OpenAIRE

    Akagi,Tadaatsu; Sonobe, Hiroshi; Miyoshi, Isao; Yoshimoto,Shizuo

    1982-01-01

    Antiserum was produced in white rabbit by intravenously injecting living cells of a B cell acute lymphoblastic leukemia (ALL) line (BALL-1). The reactivity of the antiserum against various lymphoid cell lines was examined by membrane immunofluorescence after appropriate absorption. Serum absorbed with non-T, non-B (NALL-1) and T-ALL (TALL-1) cells recognized B cell antigens distinct from Ia-like antigens on both normal and neoplastic B cells. After further absorption with tonsillar cells or n...

  18. Fingolimod therapy modulates circulating B cell composition, increases B regulatory subsets and production of IL-10 and TGFβ in patients with Multiple Sclerosis.

    Science.gov (United States)

    Blumenfeld, Shiri; Staun-Ram, Elsebeth; Miller, Ariel

    2016-06-01

    Fingolimod, an oral therapeutic agent approved for patients with relapsing-remitting Multiple Sclerosis (MS), has been shown to prevent lymphocyte egress from secondary lymphoid tissues; however the specific drug effect on B cells in fingolimod-treated patients remains to be fully elucidated. We present here a comprehensive analysis on the proportions of B cell subsets in the periphery, and the levels of activation, functional surface markers and cytokine profile of B cells in MS patients, following initiation of fingolimod therapy, using flow cytometry and cytokine bead array. Fingolimod therapy increased the ratio of naïve to memory cells, elevated the percentage of plasma cells and highly increased the proportion of transitional B cells as well as additional regulatory subsets, including: IL10(+), CD25(+) and CD5(+) B cells. The percentage of activated CD69(+) cells was highly elevated in the remaining circulating B cells, which produced increased levels of IL10, TGFβ, IL6, IL4, LTα, TNFα and IFNγ cytokines, with an overall increased ratio of TGFβ to pro-inflammatory cytokines. Furthermore, fingolimod therapy reduced ICAM-1(+) cells, suggesting a possible reduction in antigen-presenting capacity. Phosphorylated-fingolimod was shown in vitro to reduce S1PR1 RNA and protein, to slightly increase viability and to activate anti-apoptotic Bcl2 in transformed B cells of patients with MS. In conclusion, fingolimod therapy modulates significantly the composition of circulating B cells, promoting regulatory subsets and an anti-inflammatory cytokine repertoire. PMID:27055778

  19. Ibrutinib Before and After Stem Cell Transplant in Treating Patients With Relapsed or Refractory Diffuse Large B-cell Lymphoma

    Science.gov (United States)

    2016-09-01

    Activated B-Cell-Like Diffuse Large B-Cell Lymphoma; B-Cell Lymphoma, Unclassifiable, With Features Intermediate Between Diffuse Large B-Cell Lymphoma and Burkitt Lymphoma; Recurrent Diffuse Large B-Cell Lymphoma; Refractory Diffuse Large B-Cell Lymphoma

  20. Do natural Treg become activated to antigen specific Treg in transplantation and in autoimmunity?

    OpenAIRE

    Bruce Milne Hall; Tran, Giang T.; Nirupama eVerma; Karren ePlain; Catherine M. Robinson; Masaru eNomura; Hodgkinson, Suzanne J.

    2013-01-01

    Antigen specific Treg are often CD4+CD25+FoxP3+T cells, with a phenotype similar to natural Treg (nTreg). It is assumed that nTreg cannot develop into an antigen specific Treg as repeated culture with IL-2 and a specific antigen does not increase the capacity or potency of nTreg to promote immune tolerance or suppress in vitro. This has led to an assumption that antigen specific Treg mainly develop from CD4+CD25-FoxP3- T cells, by activation with antigen and TGF-beta in the absence of infla...

  1. Nbs1 ChIP-Seq Identifies Off-Target DNA Double-Strand Breaks Induced by AID in Activated Splenic B Cells.

    Directory of Open Access Journals (Sweden)

    Lyne Khair

    2015-08-01

    Full Text Available Activation-induced cytidine deaminase (AID is required for initiation of Ig class switch recombination (CSR and somatic hypermutation (SHM of antibody genes during immune responses. AID has also been shown to induce chromosomal translocations, mutations, and DNA double-strand breaks (DSBs involving non-Ig genes in activated B cells. To determine what makes a DNA site a target for AID-induced DSBs, we identify off-target DSBs induced by AID by performing chromatin immunoprecipitation (ChIP for Nbs1, a protein that binds DSBs, followed by deep sequencing (ChIP-Seq. We detect and characterize hundreds of off-target AID-dependent DSBs. Two types of tandem repeats are highly enriched within the Nbs1-binding sites: long CA repeats, which can form Z-DNA, and tandem pentamers containing the AID target hotspot WGCW. These tandem repeats are not nearly as enriched at AID-independent DSBs, which we also identified. Msh2, a component of the mismatch repair pathway and important for genome stability, increases off-target DSBs, similar to its effect on Ig switch region DSBs, which are required intermediates during CSR. Most of the off-target DSBs are two-ended, consistent with generation during G1 phase, similar to DSBs in Ig switch regions. However, a minority are one-ended, presumably due to conversion of single-strand breaks to DSBs during replication. One-ended DSBs are repaired by processes involving homologous recombination, including break-induced replication repair, which can lead to genome instability. Off-target DSBs, especially those present during S phase, can lead to chromosomal translocations, deletions and gene amplifications, resulting in the high frequency of B cell lymphomas derived from cells that express or have expressed AID.

  2. Methamphetamine activates nuclear factor kappa-light-chain-enhancer of activated B cells (NF-κB) and induces human immunodeficiency virus (HIV) transcription in human microglial cells

    Science.gov (United States)

    Wires, Emily S.; Alvarez, David; Dobrowolski, Curtis; Wang, Yun; Morales, Marisela; Karn, Jonathan; Harvey, Brandon K.

    2012-01-01

    Human immunodeficiency virus (HIV) primarily infects glial cells in the central nervous system (CNS). Recent evidence suggests that HIV-infected individuals who abuse drugs such as methamphetamine (METH) have higher viral loads and experience more severe neurological complications than HIV-infected individuals who do not abuse drugs. The aim of this study was to determine the effect of METH on HIV expression from the HIV long terminal repeats (LTR) promoter and on an HIV integrated provirus in microglial cells, the primary host cells for HIV in the CNS. Primary human microglial cells immortalized with SV40 T-antigen (CHME-5 cells) were co-transfected with an HIV LTR reporter and the HIV Tat gene, a key regulator of viral replication and gene expression, and exposed to METH. Our results demonstrate that METH treatment induced LTR activation, an effect potentiated in the presence of Tat. We also found that METH increased the nuclear translocation of the nuclear factor kappa B (NF-κB), a key cellular transcriptional regulator of the LTR promoter, and the activity of an NF-κB-specific reporter plasmid in CHME-5 cells. The presence of a dominant-negative regulator of NF-κB blocked METH-related activation of the HIV LTR. Furthermore, treatment of HIV-latently infected CHME-5 (CHME-5/HIV) cells with METH induced HIV expression in a dose-dependent manner, and nuclear translocation of the p65 subunit of NF-κB. These results suggest that METH can stimulate HIV gene expression in microglia cells through activation of the NF-κB signaling pathway. This mechanism may outline the initial biochemical events leading to the observed increased neurodegeneration in HIV-positive individuals who use METH. PMID:22618514

  3. The role of class I histocompatibility antigens in the regulation of T-cell activation.

    OpenAIRE

    Dasgupta, J D; Cemach, K; Dubey, D P; Yunis, E J; Amos, D. B.

    1987-01-01

    Class I major histocompatibility antigens in humans (HLA antigens) were found to participate in the regulation of T-cell activation and proliferation induced by phytohemagglutinin. W6/32, a monomorphic antibody directed against class I HLA-A,B,C antigens, significantly inhibited the phytohemagglutinin-induced cell proliferation of peripheral blood lymphocytes. Almost complete suppression of cell activation was achieved on a subfraction of peripheral blood lymphocytes enriched in Mo1+ monocyte...

  4. IL-2 and GM-CSF are regulated by DNA demethylation during activation of T cells, B cells and macrophages

    International Nuclear Information System (INIS)

    Highlights: ► DNA methylation is dynamic and flexible and changes rapidly upon cell activation. ► DNA methylation controls the inducible gene expression in a given cell type. ► Some enzymes are involved in maintaining the methylation profile of immune cells. -- Abstract: DNA demethylation has been found to occur at the promoters of a number of actively expressed cytokines and is believed to play a critical role in transcriptional regulation. While many DNA demethylation studies have focused on T cell activation, proliferation and differentiation, changes in DNA methylation in other types of immune cells are less well studied. We found that the expression of two cytokines (IL-2 and GM-CSF) responded differently to activation in three types of immune cells: EL4, A20 and RAW264.7 cells. Using the McrBC and MeDIP approaches, we observed decreases in DNA methylation at a genome-wide level and at the promoters of the genes of these cytokines. The expression of several potential enzymes/co-enzymes involved in the DNA demethylation pathways seemed to be associated with immune cell activation.

  5. NFκB1 is essential to prevent the development of multiorgan autoimmunity by limiting IL-6 production in follicular B cells.

    Science.gov (United States)

    de Valle, Elisha; Grigoriadis, George; O'Reilly, Lorraine A; Willis, Simon N; Maxwell, Mhairi J; Corcoran, Lynn M; Tsantikos, Evelyn; Cornish, Jasper K S; Fairfax, Kirsten A; Vasanthakumar, Ajithkumar; Febbraio, Mark A; Hibbs, Margaret L; Pellegrini, Marc; Banerjee, Ashish; Hodgkin, Philip D; Kallies, Axel; Mackay, Fabienne; Strasser, Andreas; Gerondakis, Steve; Gugasyan, Raffi

    2016-04-01

    We examined the role of NFκB1 in the homeostasis and function of peripheral follicular (Fo) B cells. Aging mice lacking NFκB1 (Nfκb1(-/-)) develop lymphoproliferative and multiorgan autoimmune disease attributed in large part to the deregulated activity of Nfκb1(-/-)Fo B cells that produce excessive levels of the proinflammatory cytokine interleukin 6 (IL-6). Despite enhanced germinal center (GC) B cell differentiation, the formation of GC structures was severely disrupted in the Nfκb1(-/-)mice. Bone marrow chimeric mice revealed that the Fo B cell-intrinsic loss of NFκB1 led to the spontaneous generation of GC B cells. This was primarily the result of an increase in IL-6 levels, which promotes the differentiation of Fo helper CD4(+)T cells and acts in an autocrine manner to reduce antigen receptor and toll-like receptor activation thresholds in a population of proliferating IgM(+)Nfκb1(-/-)Fo B cells. We demonstrate that p50-NFκB1 represses Il-6 transcription in Fo B cells, with the loss of NFκB1 also resulting in the uncontrolled RELA-driven transcription of Il-6.Collectively, our findings identify a previously unrecognized role for NFκB1 in preventing multiorgan autoimmunity through its negative regulation of Il-6 gene expression in Fo B cells. PMID:27022143

  6. Disodium cromoglycate inhibits S mu-->S epsilon deletional switch recombination and IgE synthesis in human B cells

    OpenAIRE

    1994-01-01

    IgE synthesis requires interleukin 4 (IL-4) and a T-B cell interaction that involves the B cell antigen CD40 and its ligand expressed on activated T cells. IL-4 induces epsilon germline transcription whereas ligation of CD40 results in deletional S mu-->S epsilon switch recombination, expression of mature epsilon transcripts, and IgE synthesis and secretion. We demonstrate that disodium cromoglycate (DSCG), a drug commonly used for the prophylactic treatment of allergic disease, inhibits T ce...

  7. Antiproliferative activity and phenotypic modification induced by selected Peruvian medicinal plants on human hepatocellular carcinoma Hep3B cells

    OpenAIRE

    Carraz, Maëlle; Lavergne, C.; Jullian, Valérie; Wright, M.; Gairin, J. E.; de la Cruz, M. G.; Bourdy, Geneviève

    2015-01-01

    Ethnopharmacological relevance: The high incidence of human hepatocellular carcinoma (HCC) in Peru and the wide use of medicinal plants in this country led us to study the activity against HCC cells in vitro of somes species used locally against liver and digestive disorders. Materials and methods: Ethnopharmacological survey: Medicinal plant species with a strong convergence of use for liver and digestive diseases were collected fresh in the wild or on markets, in two places of Peru: Chiclay...

  8. Mutations in the HLA class II genes leading to loss of expression of HLA-DR and HLA-DQ in diffuse large B-cell lymphoma

    NARCIS (Netherlands)

    Jordanova, ES; Philippo, K; Giphart, MJ; Schuuring, E; Kluin, PM

    2003-01-01

    Loss of expression of human leukocyte antigen (HLA) class II molecules on tumor cells affects the onset and modulation of the immune response through lack of activation of CD4(+) T lymphocytes. Previously, we showed that the frequent loss of expression of HLA class II in diffuse large B-cell lymphom

  9. Mapping of B-Cell Epitopes on a Novel 11.5-Kilodalton Fasciola hepatica-Schistosoma mansoni Cross-Reactive Antigen Belonging to a Member of the F. hepatica Saposin-Like Protein Family

    OpenAIRE

    Torres, Daricel; Espino, Ana M.

    2006-01-01

    FhSAP-2 is a novel member of the Fasciola hepatica saposin-like protein family that induces protection in rabbits against a challenge infection. We investigated the presence of lineal B-cell epitopes within this protein using a set of overlapping synthetic peptides. Peptides were tested in enzyme-linked immunosorbent assays against sera from rabbits infected with F. hepatica. Two dominant epitopes were identified, which were also highly reactive with sera from mice infected with Schistosoma m...

  10. Elevated serum IL-10 levels in diffuse large B-cell lymphoma: a mechanism of aberrant JAK2 activation

    OpenAIRE

    Gupta, Mamta; Han, Jing Jing; Stenson, Mary; Maurer, Matthew; Wellik, Linda; Hu, Guangzhen; Ziesmer, Steve; Dogan, Ahmet; Witzig, Thomas E.

    2012-01-01

    Cytokines are deregulated in cancers and can contribute to tumor growth. In patients with diffuse large-cell lymphoma (DLBCL), we observed higher levels of JAK/STAT pathway-related serum cytokines (ie, IL-6, IL-10, epidermal growth factor, and IL-2) compared with controls. Of these, only IL-10 activated the JAK2 pathway in lymphoma cells in vitro. Patients with high serum IL-10 had shorter event-free survival (EFS) than patients with low levels (P > .01) and high IL-10 was correlated with hig...

  11. Regulation of B Cell Differentiation by Intracellular Membrane-Associated Proteins and microRNAs: Role in the Antibody Response.

    Science.gov (United States)

    Lou, Zheng; Casali, Paolo; Xu, Zhenming

    2015-01-01

    B cells are central to adaptive immunity and their functions in antibody responses are exquisitely regulated. As suggested by recent findings, B cell differentiation is mediated by intracellular membrane structures (including endosomes, lysosomes, and autophagosomes) and protein factors specifically associated with these membranes, including Rab7, Atg5, and Atg7. These factors participate in vesicle formation/trafficking, signal transduction and induction of gene expression to promote antigen presentation, class switch DNA recombination (CSR)/somatic hypermutation (SHM), and generation/maintenance of plasma cells and memory B cells. Their expression is induced in B cells activated to differentiate and further fine-tuned by immune-modulating microRNAs, which coordinates CSR/SHM, plasma cell differentiation, and memory B cell differentiation. These short non-coding RNAs would individually target multiple factors associated with the same intracellular membrane compartments and collaboratively target a single factor in addition to regulating AID and Blimp-1. These, together with regulation of microRNA biogenesis and activities by endosomes and autophagosomes, show that intracellular membranes and microRNAs, two broadly relevant cell constituents, play important roles in balancing gene expression to specify B cell differentiation processes for optimal antibody responses. PMID:26579118

  12. Regulation of B cell differentiation by intracellular membrane associated proteins and microRNAs: role in the antibody response

    Directory of Open Access Journals (Sweden)

    Zheng eLou

    2015-10-01

    Full Text Available B cells are central to adaptive immunity and their functions in antibody responses are exquisitely regulated. As suggested by recent findings, B cell differentiation is mediated by intracellular membrane structures (including endosomes, lysosomes and autophagosomes and protein factors specifically associated with these membranes, including Rab7, Atg5 and Atg7. These factors participate in vesicle formation/trafficking, signal transduction and induction of gene expression to promote antigen presentation, CSR/SHM, and generation/maintenance of plasma cells and memory B cells. Their expression is induced in B cells activated to differentiate and further fine-tuned by immune-modulating microRNAs, which coordinates CSR/SHM, plasma cell differentiation and memory B cell differentiation. These short non-coding RNAs would individually target multiple factors associated with the same intracellular membrane compartments and collaboratively target a single factor in addition to regulate AID and Blimp-1. These, together with regulation of microRNA biogenesis and activities by endosomes and autophagosomes, show that intracellular membranes and microRNAs, two broadly relevant cell constituents, play important roles in balancing gene expression to specify B cell differentiation processes for optimal antibody responses.

  13. Differential and site specific impact of B cells in the protective immune response to Mycobacterium tuberculosis in the mouse.

    Directory of Open Access Journals (Sweden)

    Egídio Torrado

    Full Text Available Cell-mediated immune responses are known to be critical for control of mycobacterial infections whereas the role of B cells and humoral immunity is unclear. B cells can modulate immune responses by secretion of immunoglobulin, production of cytokines and antigen-presentation. To define the impact of B cells in the absence of secreted immunoglobulin, we analyzed the progression of Mycobacterium tuberculosis (Mtb infection in mice that have B cells but which lack secretory immunoglobulin (AID(-/-µS(-/-mice. AID(-/-µS(-/- mice accumulated a population of activated B cells in the lungs when infected and were more susceptible to aerosol Mtb when compared to wild type (C57BL/6 mice or indeed mice that totally lack B cells. The enhanced susceptibility of AID(-/-µS(-/- mice was not associated with defective T cell activation or expression of a type 1 immune response. While delivery of normal serum to AID(-/-µS(-/- mice did not reverse susceptibility, susceptibility in the spleen was dependent upon the presence of B cells and susceptibility in the lungs of AID(-/-µS(-/-mice was associated with elevated expression of the cytokines IL-6, GM-CSF, IL-10 and molecules made by alternatively activated macrophages. Blocking of IL-10 signaling resulted in reversal of susceptibility in the spleens and lungs of AID(-/-µS(-/- mice. These data support the hypothesis that B cells can modulate immunity to Mtb in an organ specific manner via the modulation of cytokine production and macrophage activation.

  14. Differential and site specific impact of B cells in the protective immune response to Mycobacterium tuberculosis in the mouse.

    Science.gov (United States)

    Torrado, Egídio; Fountain, Jeffrey J; Robinson, Richard T; Martino, Cynthia A; Pearl, John E; Rangel-Moreno, Javier; Tighe, Michael; Dunn, Robert; Cooper, Andrea M

    2013-01-01

    Cell-mediated immune responses are known to be critical for control of mycobacterial infections whereas the role of B cells and humoral immunity is unclear. B cells can modulate immune responses by secretion of immunoglobulin, production of cytokines and antigen-presentation. To define the impact of B cells in the absence of secreted immunoglobulin, we analyzed the progression of Mycobacterium tuberculosis (Mtb) infection in mice that have B cells but which lack secretory immunoglobulin (AID(-/-)µS(-/-)mice). AID(-/-)µS(-/-) mice accumulated a population of activated B cells in the lungs when infected and were more susceptible to aerosol Mtb when compared to wild type (C57BL/6) mice or indeed mice that totally lack B cells. The enhanced susceptibility of AID(-/-)µS(-/-) mice was not associated with defective T cell activation or expression of a type 1 immune response. While delivery of normal serum to AID(-/-)µS(-/-) mice did not reverse susceptibility, susceptibility in the spleen was dependent upon the presence of B cells and susceptibility in the lungs of AID(-/-)µS(-/-)mice was associated with elevated expression of the cytokines IL-6, GM-CSF, IL-10 and molecules made by alternatively activated macrophages. Blocking of IL-10 signaling resulted in reversal of susceptibility in the spleens and lungs of AID(-/-)µS(-/-) mice. These data support the hypothesis that B cells can modulate immunity to Mtb in an organ specific manner via the modulation of cytokine production and macrophage activation. PMID:23613902

  15. Epstein-Barr virus nuclear antigen 3C targets p53 and modulates its transcriptional and apoptotic activities

    International Nuclear Information System (INIS)

    The p53 tumor suppressor gene is one of the most commonly mutated genes in human cancers and the corresponding encoded protein induces apoptosis or cell-cycle arrest at the G1/S checkpoint in response to DNA damage. To date, previous studies have shown that antigens encoded by human tumor viruses such as SV40 large T antigen, adenovirus E1A and HPV E6 interact with p53 and disrupt its functional activity. In a similar fashion, we now show that EBNA3C, one of the EBV latent antigens essential for the B-cell immortalization in vitro, interacts directly with p53. Additionally, we mapped the interaction of EBNA3C with p53 to the C-terminal DNA-binding and the tetramerization domain of p53, and the region of EBNA3C responsible for binding to p53 was mapped to the N-terminal domain of EBNA3C (residues 130-190), previously shown to interact with a number of important cell-cycle components, specifically SCFSkp2, cyclin A, and cMyc. Furthermore, we demonstrate that EBNA3C substantially represses the transcriptional activity of p53 in luciferase based reporter assays, and rescues apoptosis induced by ectopic p53 expression in SAOS-2 (p53-/-) cells. Interestingly, we also show that the DNA-binding ability of p53 is diminished in the presence of EBNA3C. Thus, the interaction between the p53 and EBNA3C provides new insights into the mechanism(s) by which the EBNA3C oncoprotein can alter cellular gene expression in EBV associated human cancers.

  16. Increased prevalence of late stage T cell activation antigen (VLA-1) in active juvenile chronic arthritis

    DEFF Research Database (Denmark)

    Ødum, Niels; Morling, Niels; Platz, P; Hofmann, B; Ryder, L P; Heilmann, C; Pedersen, F K; Nielsen, L P; Friis, J; Svejgaard, A

    1987-01-01

    the various HLA class II antigens was observed between the groups. Similarly, no significant differences in stimulatory capability in secondary mixed lymphocyte culture (MLC) were seen. The distribution of T helper/inducer (CD4+), T suppressor/cytotoxic (CD8+), and NK cells was similar in active JCA...

  17. Determination of antigenicity by radioimmunoassay and of trypsin inhibitory activities in heat or enzyme denatured ovomucoid

    International Nuclear Information System (INIS)

    The effects of heat treatment and enzymic digestion on the antigenic reactivity of ovomucoid were studied. This reactivity was mainly examined by a solid phase competitive radioimmunoassay. The trypsin inhibitory activity was also measured to elucidate its relationship to the antigenic reactivity. The antigenic reactivity and trypsin inhibitory activity diminished in parallel with increase of heating time. However, enzymic digestion caused a more rapid decrease in antigenic reactivity than in trypsin inhibitory activity. From these results, it is suggested that the structure of the antigenic determinants is destroyed in a similar manner to that of the trypsin inhibitory active site when ovomucoid is heated, while the former structure is destroyed more easily than the latter by enzymic digestion

  18. Immune activation by casein dietary antigens in bipolar disorder

    NARCIS (Netherlands)

    Severance, E.G.; Dupont, D.; Dickerson, F.B.; Stallings, C.R.; Origoni, A.E.; Krivogorsky, B.; Yang, S.; Haasnoot, W.; Yolken, R.H.

    2010-01-01

    Objectives: Inflammation and other immune processes are increasingly linked to psychiatric diseases. Antigenic triggers specific to bipolar disorder are not yet defined. We tested whether antibodies to bovine milk caseins were associated with bipolar disorder, and whether patients recognized differe

  19. Design of targeted B cell killing agents.

    Directory of Open Access Journals (Sweden)

    Alexey V Stepanov

    Full Text Available B cells play an important role in the pathogenesis of both systemic and organ-specific autoimmune diseases. Autoreactive B cells not only produce autoantibodies, but also are capable to efficiently present specific autoantigens to T cells. Furthermore, B cells can secrete proinflammatory cytokines and amplify the vicious process of self-destruction. B cell-directed therapy is a potentially important approach for treatment of various autoimmune diseases. The depletion of B cells by anti-CD20/19 monoclonal antibody Retuximab® used in autoimmune diseases therapy leads to systemic side effects and should be significantly improved. In this study we designed a repertoire of genetically engineered B cell killers that specifically affected one kind of cells carrying a respective B cell receptor. We constructed immunotoxins (ITs, fused with c-myc epitope as a model targeting sequence, based on barnase, Pseudomonas toxin, Shiga-like toxin E.coli and Fc domain of human antibody IgGγ1. C-MYC hybridoma cell line producing anti-c-myc IgG was chosen as a model for targeted cell depletion. C-myc sequence fused with toxins provided addressed delivery of the toxic agent to the target cells. We demonstrated functional activity of designed ITs in vitro and showed recognition of the fusion molecules by antibodies produced by targeted hybridoma. To study specificity of the proposed B cells killing molecules, we tested a set of created ITs ex vivo, using C-MYC and irrelevant hybridoma cell lines. Pseudomonas-containing IT showed one of the highest cytotoxic effects on the model cells, however, possessed promiscuous specificity. Shiga-like toxin construct demonstrated mild both cytotoxicity and specificity. Barnase and Fc-containing ITs revealed excellent balance between their legibility and toxic properties. Moreover, barnase and Fc molecules fused with c-myc epitope were able to selectively deplete c-myc-specific B cells and decrease production of anti

  20. Murine germinal center B cells require functional Fms-like tyrosine kinase 3 signaling for IgG1 class-switch recombination.

    Science.gov (United States)

    Svensson, Mattias N D; Andersson, Karin M E; Wasén, Caroline; Erlandsson, Malin C; Nurkkala-Karlsson, Merja; Jonsson, Ing-Marie; Brisslert, Mikael; Bemark, Mats; Bokarewa, Maria I

    2015-12-01

    Switched antibody classes are important for efficient immune responses. Aberrant antibody production to otherwise harmless antigens may result in autoimmunity. The protein kinase fms-like tyrosine kinase 3 receptor (Flt3) has an important role during early B-cell development, but the role of Flt3 in peripheral B cells has not been assessed before. Herein we describe a previously unappreciated role for Flt3 in IgG1 class-switch recombination (CSR) and production. We show that Flt3 is reexpressed on B-cell lymphoma 6(+) germinal center B cells in vivo and following LPS activation of peripheral B cells in vitro. Absence of Flt3 signaling in Flt3 ligand-deficient mice results in impaired IgG1 CSR and accumulation of IgM-secreting plasma cells. On activated B cells, Flt3 is coexpressed and functions in synergy with the common-gamma chain receptor family. B cells from Flt3 ligand-deficient mice have impaired IL-4R signaling, with reduced phosphorylation of signal transducer and activator of transcription (Stat) 6, and demonstrate a failure to initiate CSR to IgG1 with low expression of γ1 germ-line transcripts, resulting in impaired IgG1 production. Thus, functional synergy between Flt3 and IL-4R signaling is critical for Stat-mediated regulation of sterile γ1 germ-line transcripts and CSR to IgG1. PMID:26627255

  1. Transcriptome Analysis of B Cell Immune Functions in Periodontitis: Mucosal Tissue Responses to the Oral Microbiome in Aging.

    Science.gov (United States)

    Ebersole, Jeffrey L; Kirakodu, Sreenatha S; Novak, M John; Orraca, Luis; Martinez, Janis Gonzalez; Cunningham, Larry L; Thomas, Mark V; Stromberg, Arnold; Pandruvada, Subramanya N; Gonzalez, Octavio A

    2016-01-01

    Evidence has shown activation of T and B cells in gingival tissues in experimental models and in humans diagnosed with periodontitis. The results of this adaptive immune response are noted both locally and systemically with antigenic specificity for an array of oral bacteria, including periodontopathic species, e.g., Porphyromonas gingivalis and Aggregatibacter actinomycetemcomitans. It has been recognized through epidemiological studies and clinical observations that the prevalence of periodontitis increases with age. This report describes our studies evaluating gingival tissue transcriptomes in humans and specifically exploiting the use of a non-human primate model of naturally occurring periodontitis to delineate gingival mucosal tissue gene expression profiles focusing on cells/genes critical for the development of humoral adaptive immune responses. Patterns of B cell and plasmacyte genes were altered in aging healthy gingival tissues. Substantial increases in a large number of genes reflecting antigen-dependent activation, B cell activation, B cell proliferation, and B cell differentiation/maturation were observed in periodontitis in adults and aged animals. Finally, evaluation of the relationship of these gene expression patterns with those of various tissue destructive molecules (MMP2, MMP9, CTSK, TNFα, and RANKL) showed a greater frequency of positive correlations in healthy tissues versus periodontitis tissues, with only MMP9 correlations similar between the two tissue types. These results are consistent with B cell response activities in healthy tissues potentially contributing to muting the effects of the tissue destructive biomolecules, whereas with periodontitis this relationship is adversely affected and enabling a progression of tissue destructive events. PMID:27486459

  2. Transcriptome Analysis of B Cell Immune Functions in Periodontitis: Mucosal Tissue Responses to the Oral Microbiome in Aging

    Science.gov (United States)

    Ebersole, Jeffrey L.; Kirakodu, Sreenatha S.; Novak, M. John; Orraca, Luis; Martinez, Janis Gonzalez; Cunningham, Larry L.; Thomas, Mark V.; Stromberg, Arnold; Pandruvada, Subramanya N.; Gonzalez, Octavio A.

    2016-01-01

    Evidence has shown activation of T and B cells in gingival tissues in experimental models and in humans diagnosed with periodontitis. The results of this adaptive immune response are noted both locally and systemically with antigenic specificity for an array of oral bacteria, including periodontopathic species, e.g., Porphyromonas gingivalis and Aggregatibacter actinomycetemcomitans. It has been recognized through epidemiological studies and clinical observations that the prevalence of periodontitis increases with age. This report describes our studies evaluating gingival tissue transcriptomes in humans and specifically exploiting the use of a non-human primate model of naturally occurring periodontitis to delineate gingival mucosal tissue gene expression profiles focusing on cells/genes critical for the development of humoral adaptive immune responses. Patterns of B cell and plasmacyte genes were altered in aging healthy gingival tissues. Substantial increases in a large number of genes reflecting antigen-dependent activation, B cell activation, B cell proliferation, and B cell differentiation/maturation were observed in periodontitis in adults and aged animals. Finally, evaluation of the relationship of these gene expression patterns with those of various tissue destructive molecules (MMP2, MMP9, CTSK, TNFα, and RANKL) showed a greater frequency of positive correlations in healthy tissues versus periodontitis tissues, with only MMP9 correlations similar between the two tissue types. These results are consistent with B cell response activities in healthy tissues potentially contributing to muting the effects of the tissue destructive biomolecules, whereas with periodontitis this relationship is adversely affected and enabling a progression of tissue destructive events. PMID:27486459

  3. The Mertk Receptor Tyrosine Kinase Promotes T-B interaction Stimulated by IgD B-cell Receptor Cross-linking

    OpenAIRE

    Shao, Wen-Hai; Zhen, Yuxuan; Finkelman, Fred D.; Cohen, Philip L.

    2014-01-01

    The Mertk receptor tyrosine kinase facilitates macrophage and DC apoptotic-cell clearance and regulates immune tolerance. Mertk may also contribute to B-cell activation, because Mertk-KO mice fail to develop autoantibodies when allo-activated by T cells. We investigated this possibility with a well-characterized model in which injection of mice with goat anti-IgD antibody causes membrane IgD cross-linking that induces T-independent B cell activation and antigen presentation to T cells. Goat a...

  4. Differential programming of B cells in AID deficient mice.

    Directory of Open Access Journals (Sweden)

    Marc A Hogenbirk

    Full Text Available The Aicda locus encodes the activation induced cytidine deaminase (AID and is highly expressed in germinal center (GC B cells to initiate somatic hypermutation (SHM and class switch recombination (CSR of immunoglobulin (Ig genes. Besides these Ig specific activities in B cells, AID has been implicated in active DNA demethylation in non-B cell systems. We here determined a potential role of AID as an epigenetic eraser and transcriptional regulator in B cells. RNA-Seq on different B cell subsets revealed that Aicda(-/- B cells are developmentally affected. However as shown by RNA-Seq, MethylCap-Seq, and SNP analysis these transcriptome alterations may not relate to AID, but alternatively to a CBA mouse strain derived region around the targeted Aicda locus. These unexpected confounding parameters provide alternative, AID-independent interpretations on genotype-phenotype correlations previously reported in numerous studies on AID using the Aicda(-/- mouse strain.

  5. IFN-γ licenses CD11b(+) cells to induce progression of systemic lupus erythematosus.

    Science.gov (United States)

    Shaabani, Namir; Honke, Nadine; Dolff, Sebastian; Görg, Boris; Khairnar, Vishal; Merches, Katja; Duhan, Vikas; Metzger, Sabine; Recher, Mike; Barthuber, Carmen; Hardt, Cornelia; Proksch, Peter; Häussinger, Dieter; Witzke, Oliver; Lang, Philipp A; Lang, Karl S

    2015-08-01

    Autoantibodies are a hallmark of autoimmune diseases, such as rheumatoid arthritis, autoimmune hepatitis, and systemic lupus erythematosus (SLE). High titers of anti-nuclear antibodies are used as surrogate marker for SLE, however their contribution to pathogenesis remains unclear. Using murine model of SLE and human samples, we studied the effect of immune stimulation on relapsing of SLE. Although autoantibodies bound to target cells in vivo, only additional activation of CD8(+) T cells converted this silent autoimmunity into overt disease. In mice as well as in humans CD8(+) T cells derived IFN-γ enhanced expression of Fc-receptors on CD11b(+) cells. High expression of Fc-receptors allowed CD11b(+) cells to bind to antibody covered target cells and to destroy them in vivo. We found that autoantibodies induce clinically relevant disease when adaptive immunity, specific for disease non-related antigen, is activated. PMID:26094774

  6. Role of AKT/ PKB and 14-3-3 in the regulation of B cell receptor signaling and signalosome assembly

    OpenAIRE

    Mohammad, Dara

    2015-01-01

    AKT/PKB is an oncogenic serine/threonine kinase regulated via the PI3K pathways. 14-3-3s represent a large group of adaptor proteins that are known to interact with a plethora of signaling proteins and regulate diverse signal transduction pathways. The B-cell antigen receptor (BCR) activation and signalosome assembly are dynamic processes controlled by protein phosphorylation. The signaling events share their functions in controlling cell proliferation, differentiation, and/or apoptosis. ...

  7. 嵌合型抗原受体T细胞治疗技术在B细胞肿瘤中的研究进展:第56届美国血液学会年会报道%Progress of chimeric antigen receptor T cell immunotherapy in B cell hematologic malignancies:reports from the 56th American Society of Hematology annual meeting

    Institute of Scientific and Technical Information of China (English)

    邓碧萍; 刘红星; 童春容

    2015-01-01

    Chimeric antigen receptor T cell (CAR-T) therapy has shown promising perspective in clinical trails of B cell hematologic malignancies.Meanwhile,this therapy still need to be further improved in the following aspects:design of CAR-T,cancer antigens selection,T cells origin,and clinical application strategy.CAR-T immunotherapy is one of hot topics in the 56th American Society of Hematology (ASH) annual meeting.Some breakthroughs have been reported in both basic research and clinical trails.%嵌合型抗原受体T细胞(chimeric antigen receptorT cell,CAR-T)在B细胞肿瘤临床试验中已经显示了良好的前景,但是CAR-T的设计、肿瘤抗原的选择、T细胞的来源、CAR-T的临床应用策略等还有待进一步开发完善.CAR-T免疫细胞治疗技术是2014年第56届美国血液学会(ASH)年会的热点话题之一,其基础研究及临床试验均有突破性进展.

  8. Immune-inflammatory responses in atherosclerosis: Role of an adaptive immunity mainly driven by T and B cells.

    Science.gov (United States)

    Chistiakov, Dimitry A; Orekhov, Alexander N; Bobryshev, Yuri V

    2016-09-01

    Adaptive immune response plays an important role in atherogenesis. In atherosclerosis, the proinflammatory immune response driven by Th1 is predominant but the anti-inflammatory response mediated mainly by regulatory T cells is also present. The role of Th2 and Th17 cells in atherogenesis is still debated. In the plaque, other T helper cells can be observed such as Th9 and Th22 but is little is known about their impact in atherosclerosis. Heterogeneity of CD4(+) T cell subsets presented in the plaque may suggest for plasticity of T cell that can switch the phenotype dependening on the local microenvironment and activating/blocking stimuli. Effector T cells are able to recognize self-antigens released by necrotic and apoptotic vascular cells and induce a humoral immune reaction. Tth cells resided in the germinal centers help B cells to switch the antibody class to the production of high-affinity antibodies. Humoral immunity is mediated by B cells that release antigen-specific antibodies. A variety of B cell subsets were found in human and murine atherosclerotic plaques. In mice, B1 cells could spontaneously produce atheroprotective natural IgM antibodies. Conventional B2 lymphocytes secrete either proatherogenic IgG, IgA, and IgE or atheroprotective IgG and IgM antibodies reactive with oxidation-specific epitopes on atherosclerosis-associated antigens. A small population of innate response activator (IRA) B cells, which is phenotypically intermediate between B1 and B2 cells, produces IgM but possesses proatherosclerotic properties. Finally, there is a minor subset of splenic regulatory B cells (Bregs) that protect against atherosclerotic inflammation through support of generation of Tregs and production of anti-inflammatory cytokines IL-10 and TGF-β and proapoptotic molecules. PMID:27262513

  9. Monoclonal antibodies putatively recognising activation and differentiation antigens

    Czech Academy of Sciences Publication Activity Database

    Šinkora, Jiří; Řeháková, Zuzana; Haverson, K.; Šinkora, Marek; Dominquez, J.; Huang, CH. A.

    2001-01-01

    Roč. 80, - (2001), s. 143-164. ISSN 0165-2427 R&D Projects: GA ČR GA524/00/1280; GA AV ČR IPP2052002 Institutional research plan: CEZ:AV0Z5020903 Keywords : pig * cluster of differentiation antigens * haematopoisis Subject RIV: EC - Immunology Impact factor: 1.389, year: 2001

  10. Antigen-specific T cell activation independently of the MHC: chimeric antigen receptor (CAR-redirected T cells.

    Directory of Open Access Journals (Sweden)

    Hinrich eAbken

    2013-11-01

    Full Text Available Adoptive T cell therapy has recently shown powerful in initiating a lasting anti-tumor response with spectacular therapeutic success in some cases. Specific T cell therapy, however, is limited since a number of cancer cells are not recognized by T cells due to various mechanisms including the limited availability of tumor-specific T cells and deficiencies in antigen processing or major histocompatibility complex (MHC expression of cancer cells. To make adoptive cell therapy applicable for the broad variety of cancer entities, patient's T cells are engineered ex vivo with pre-defined specificity by a recombinant chimeric antigen receptor (CAR which consists in the extracellular part of an antibody-derived domain for binding with a tumor-associated antigen and in the intracellular part of a TCR-derived signaling moiety for T cell activation. The specificity of CAR mediated T cell recognition is defined by the antibody domain, is independent of MHC presentation and can be extended to any target for which an antibody is available. We discuss the advantages and limitations of MHC-independent T cell targeting by an engineered CAR and review most significant progress recently made in early stage clinical trials to treat cancer.

  11. B7h-expressing dendritic cells and plasma B cells mediate distinct outcomes of ICOS costimulation in T cell-dependent antibody responses

    Directory of Open Access Journals (Sweden)

    Larimore Kevin

    2012-06-01

    Full Text Available Abstract Background The ICOS-B7h costimulatory receptor-ligand pair is required for germinal center formation, the production of isotype-switched antibodies, and antibody affinity maturation in response to T cell-dependent antigens. However, the potentially distinct roles of regulated B7h expression on B cells and dendritic cells in T cell-dependent antibody responses have not been defined. Results We generated transgenic mice with lineage-restricted B7h expression to assess the cell-type specific roles of B7h expression on B cells and dendritic cells in regulating T cell-dependent antibody responses. Our results show that endogenous B7h expression is reduced on B cells after activation in vitro and is also reduced in vivo on antibody-secreting plasma B cells in comparison to both naïve and germinal center B cells from which they are derived. Increasing the level of B7h expression on activated and plasma B cells in B-B7hTg mice led to an increase in the number of antibody-secreting plasma cells generated after immunization and a corresponding increase in the concentration of antigen-specific high affinity serum IgG antibodies of all isotypes, without affecting the number of responding germinal center B cells. In contrast, ICOS costimulation mediated by dendritic cells in DC-B7hTg mice contributed to germinal center formation and selectively increased IgG2a production without affecting the overall magnitude of antibody responses. Conclusions Using transgenic mice with lineage-restricted B7h expression, we have revealed distinct roles of ICOS costimulation mediated by dendritic cells and B cells in the regulation of T cell-dependent antibody responses.

  12. Exploiting human memory B cell heterogeneity for improved vaccine efficacy

    Directory of Open Access Journals (Sweden)

    Noel Thomas Pauli

    2011-12-01

    Full Text Available The major goal in vaccination is establishment of long-term, prophylactic humoral memory to a pathogen. Two major components to long-lived humoral memory are plasma cells for the production of specific immunoglobulin and memory B cells that survey for their specific antigen in the periphery for later affinity maturation, proliferation, and differentiation. The study of human B cell memory has been aided by the discovery of a general marker for B cell memory, expression of CD27; however, new data suggests the existence of CD27- memory B cells as well. These recently described non-canonical memory populations have increasingly pointed to the heterogeneity of the memory compartment. The novel B memory subsets in humans appear to have unique origins, localization, and functions compared to what was considered to be a classical memory B cell. In this article, we review the known B cell memory subsets, the establishment of B cell memory in vaccination and infection, and how understanding these newly described subsets can inform vaccine design and disease treatment.

  13. 猪带绦虫六钩蚴TSO45-4B抗原FnⅢ结构域相应的线性B细胞表位肽免疫原性研究%Research on immunity response about Taenia solium oncosphere TSO45-4B antigens FnⅢ structure domain linear B cells epitope peptides

    Institute of Scientific and Technical Information of China (English)

    王媛媛; 陶志勇; 杨小迪; 王小莉; 常雪莲; 陈勇; 孙新; 夏惠; 方强

    2013-01-01

    Objective: To observe the humoral immune response induced by Taenia solium oncosphere TSO45-4B antigens Fn Ⅲ structure domain linear B cells epitope peptides in mice. Methods: The two predicted B cell epitope peptides of TSO45-4B Fn Ⅲ structure domain conjugated with carrier protein of keyhole limpet hemocyanin were synthesized and used to immunize mice. The mice specific serum antibody titer to the epitope peptides synthesized was tested by ELISA. Results: The specific antibody to one of the predicted epitope peptides synthesized was found in mice serum,and the titer was 1: 1 280. Conclusions: One of the two predicted linear B cell epitope peptides of TSO45-4B FnⅢ structure domain can induce the humoral immune response in mice.%目的:观察载体蛋白偶联的TSO45-4B抗原FnⅢ结构域相应的线性B细胞表位肽诱导的体液免疫反应.方法:人工合成TSO45-4B抗原FnⅢ结构域2条预测表位肽,偶联钥孔血蓝蛋白免疫小鼠,采用ELISA法检测小鼠血清中预测表位肽特异性抗体滴度.结果:免疫小鼠血清中检测到1条预测表位肽特异性抗体,其效价达到1:1 280.结论:设计的1条TSO45-4B抗原FnⅢ结构域线性B细胞表位肽可诱导小鼠产生体液免疫反应.

  14. Novel vaccine strategies to T-independent antigens.

    Science.gov (United States)

    Lesinski, G B; Westerink, M A

    2001-11-01

    T cell independent antigens do not require T cell help to induce an immune response, and are characterized by a lack of immunologic memory. These antigens can be divided into two classes, TI-1 or TI-2. TI-1 antigens, such as bacterial lipopolysaccharide, are potent B-cell mitogens, capable of non-specific, polyclonal activation of B cells. In contrast, TI-2 antigens can only activate mature B cells and consist of highly repetitive structures, such as capsular polysaccharides (CPS) from bacteria. Many vaccines currently in use consist of purified capsular polysaccharides from pathogenic bacteria such as Streptococcus pneumoniae and Neisseria meningitidis. These vaccines are efficacious in immune-competent adults, however, due to their TI-2 nature, are not effective in children <2 years of age. Converting polysaccharides into T cell dependent (TD) antigens, allows children, <2, to produce an effective immune response. This review focuses on various strategies used to convert the immune response to polysaccharide antigens from TI-2 to a TD response. Conjugate vaccines, anti-idiotypic antibodies, phage display library technology and DNA vaccines are discussed. PMID:11576678

  15. An in vitro study of liposomal curcumin: stability, toxicity and biological activity in human lymphocytes and Epstein-Barr virus-transformed human B-cells.

    Science.gov (United States)

    Chen, Changguo; Johnston, Thomas D; Jeon, Hoonbae; Gedaly, Roberto; McHugh, Patrick P; Burke, Thomas G; Ranjan, Dinesh

    2009-01-21

    Curcumin is a multi-functional and pharmacologically safe natural agent. Used as a food additive for centuries, it also has anti-inflammatory, anti-virus and anti-tumor properties. We previously found that it is a potent inhibitor of cyclosporin A (CsA)-resistant T-cell co-stimulation pathway. It inhibits mitogen-stimulated lymphocyte proliferation, NFkappaB activation and IL-2 signaling. In spite of its safety and efficacy, the in vivo bioavailability of curcumin is poor, and this may be a major obstacle to its utility as a therapeutic agent. Liposomes are known to be excellent carriers for drug delivery. In this in vitro study, we report the effects of different liposome formulations on curcumin stability in phosphate buffered saline (PBS), human blood, plasma and culture medium RPMI-1640+10% FBS (pH 7.4, 37 degrees C). Liposomal curcumin had higher stability than free curcumin in PBS. Liposomal and free curcumin had similar stability in human blood, plasma and RPMI-1640+10% FBS. We looked at the toxicity of non-drug-containing liposomes on (3)H-thymidine incorporation by concanavalin A (Con A)-stimulated human lymphocytes, splenocytes and Epstein-Barr virus (EBV)-transformed human B-cell lymphoblastoid cell line (LCL). We found that dimyristoylphosphatidylcholine (DMPC) and dimyristoylphosphatidylglycerol (DMPG) were toxic to the tested cells. However, addition of cholesterol to the lipids at DMPC:DMPG:cholesterol=7:1:8 (molar ratio) almost completely eliminated the lipid toxicity to these cells. Liposomal curcumin had similar or even stronger inhibitory effects on Con A-stimulated human lymphocyte, splenocyte and LCL proliferation. We conclude that liposomal curcumin may be useful for intravenous administration to improve the bioavailability and efficacy, facilitating in vivo studies that could ultimately lead to clinical application of curcumin. PMID:18840516

  16. Maternal immunization with ovalbumin prevents neonatal allergy development and up-regulates inhibitory receptor FcγRIIB expression on B cells

    Directory of Open Access Journals (Sweden)

    Duarte Alberto JS

    2010-03-01

    Full Text Available Abstract Background Preconception allergen immunization prevents neonatal allergen sensitization in mice by a complex interaction between regulatory cells/factors and antibodies. The present study assessed the influence of maternal immunization with ovalbumin (OVA on the immune response of 3 day-old and 3 week-old offspring immunized or non-immunized with OVA and evaluated the effect of IgG treatment during fetal development or neonatal period. Results Maternal immunization with OVA showed increased levels of FcγRIIb expression in splenic B cells of neonates, which were maintained for up to 3 weeks and not affected by additional postnatal OVA immunization. Maternal immunization also exerted a down-modulatory effect on both IL-4 and IFN-γ-secreting T cells and IL-4 and IL-12- secreting B cells. Furthermore, immunized neonates from immunized mothers showed a marked inhibition of antigen-specifc IgE Ab production and lowered Th2/Th1 cytokine levels, whereas displaying enhanced FcγRIIb expression on B cells. These offspring also showed reduced antigen-specific proliferative response and lowered B cell responsiveness. Moreover, in vitro evaluation revealed an impairment of B cell activation upon engagement of B cell antigen receptor by IgG from OVA-immunized mice. Finally, in vivo IgG transference during pregnancy or breastfeeding revealed that maternal Ab transference was able to increase regulatory cytokines, such as IL-10, in the prenatal stage; yet only the postnatal treatment prevented neonatal sensitization. None of the IgG treatments induced immunological changes in the offspring, as it was observed for those from OVA-immunized mothers. Conclusion Maternal immunization upregulates the inhibitory FcγRIIb expression on offspring B cells, avoiding skewed Th2 response and development of allergy. These findings contribute to the advancement of prophylactic strategies to prevent allergic diseases in early life.

  17. Human B cells produce chemokine CXCL10 in the presence of Mycobacterium tuberculosis specific T cells

    DEFF Research Database (Denmark)

    Hoff, Soren T; Salman, Ahmed M; Ruhwald, Morten;

    2015-01-01

    BACKGROUND: The role of B cells in human host response to Mycobacterium tuberculosis (Mtb) infection is still controversial, but recent evidence suggest that B cell follicle like structures within the lung may influence host responses through regulation of the local cytokine environment. A...... candidate for such regulation could be the chemokine CXCL10. CXCL10 is mainly produced by human monocytes, but a few reports have also found CXCL10 production by human B cells. The objective of this study was to investigate CXCL10 production by human B cells in response to in vitro stimulation with Mtb...... antigens. METHODOLOGY/PRINCIPAL FINDINGS: We analyzed human blood samples from 30 volunteer donors using multiparameter flow cytometry, and identified a subgroup of B cells producing CXCL10 in response to in vitro stimulation with antigens. T cells did not produce CXCL10, but CXCL10 production by B cells...

  18. Antigen-activated dendritic cells ameliorate influenza A infections

    OpenAIRE

    Boonnak, Kobporn; Vogel, Leatrice; Orandle, Marlene; Zimmerman, Daniel; Talor, Eyal; Subbarao, Kanta

    2013-01-01

    Influenza A viruses cause significant morbidity and mortality worldwide. There is a need for alternative or adjunct therapies, as resistance to currently used antiviral drugs is emerging rapidly. We tested ligand epitope antigen presentation system (LEAPS) technology as a new immune-based treatment for influenza virus infection in a mouse model. Influenza-J-LEAPS peptides were synthesized by conjugating the binding ligand derived from the β2-microglobulin chain of the human MHC class I molecu...

  19. Early events associated with infection of Epstein-Barr virus infection of primary B-cells.

    Directory of Open Access Journals (Sweden)

    Sabyasachi Halder

    Full Text Available Epstein Barr virus (EBV is closely associated with the development of a vast number of human cancers. To develop a system for monitoring early cellular and viral events associated with EBV infection a self-recombining BAC containing 172-kb of the Epstein Barr virus genome BAC-EBV designated as MD1 BAC (Chen et al., 2005, J.Virology was used to introduce an expression cassette of green fluorescent protein (GFP by homologous recombination, and the resultant BAC clone, BAC-GFP-EBV was transfected into the HEK 293T epithelial cell line. The resulting recombinant GFP EBV was induced to produce progeny virus by chemical inducer from the stable HEK 293T BAC GFP EBV cell line and the virus was used to immortalize human primary B-cell as monitored by green fluorescence and outgrowth of the primary B cells. The infection, B-cell activation and cell proliferation due to GFP EBV was monitored by the expression of the B-cell surface antigens CD5, CD10, CD19, CD23, CD39, CD40 , CD44 and the intercellular proliferation marker Ki-67 using Flow cytometry. The results show a dramatic increase in Ki-67 which continues to increase by 6-7 days post-infection. Likewise, CD40 signals showed a gradual increase, whereas CD23 signals were increased by 6-12 hours, maximally by 3 days and then decreased. Monitoring the viral gene expression pattern showed an early burst of lytic gene expression. This up-regulation of lytic gene expression prior to latent genes during early infection strongly suggests that EBV infects primary B-cell with an initial burst of lytic gene expression and the resulting progeny virus is competent for infecting new primary B-cells. This process may be critical for establishment of latency prior to cellular transformation. The newly infected primary B-cells can be further analyzed for investigating B cell activation due to EBV infection.

  20. Multidimensional single-cell analysis of BCR signaling reveals proximal activation defect as a hallmark of chronic lymphocytic leukemia B cells.

    Directory of Open Access Journals (Sweden)

    M Lia Palomba

    Full Text Available PURPOSE: Chronic Lymphocytic Leukemia (CLL is defined by a perturbed B-cell receptor-mediated signaling machinery. We aimed to model differential signaling behavior between B cells from CLL and healthy individuals to pinpoint modes of dysregulation. EXPERIMENTAL DESIGN: We developed an experimental methodology combining immunophenotyping, multiplexed phosphospecific flow cytometry, and multifactorial statistical modeling. Utilizing patterns of signaling network covariance, we modeled BCR signaling in 67 CLL patients using Partial Least Squares Regression (PLSR. Results from multidimensional modeling were validated using an independent test cohort of 38 patients. RESULTS: We identified a dynamic and variable imbalance between proximal (pSYK, pBTK and distal (pPLCγ2, pBLNK, ppERK phosphoresponses. PLSR identified the relationship between upstream tyrosine kinase SYK and its target, PLCγ2, as maximally predictive and sufficient to distinguish CLL from healthy samples, pointing to this juncture in the signaling pathway as a hallmark of CLL B cells. Specific BCR pathway signaling signatures that correlate with the disease and its degree of aggressiveness were identified. Heterogeneity in the PLSR response variable within the B cell population is both a characteristic mark of healthy samples and predictive of disease aggressiveness. CONCLUSION: Single-cell multidimensional analysis of BCR signaling permitted focused analysis of the variability and heterogeneity of signaling behavior from patient-to-patient, and from cell-to-cell. Disruption of the pSYK/pPLCγ2 relationship is uncovered as a robust hallmark of CLL B cell signaling behavior. Together, these observations implicate novel elements of the BCR signal transduction as potential therapeutic targets.

  1. Overexpression of Activation-Induced Cytidine Deaminase in MTX- and Age-Related Epstein-Barr Virus-Associated B-Cell Lymphoproliferative Disorders of the Head and Neck

    Directory of Open Access Journals (Sweden)

    Kentaro Kikuchi

    2015-01-01

    Full Text Available Recent research has shown that activation-induced cytidine deaminase (AID triggers somatic hypermutation and recombination, in turn contributing to lymphomagenesis. Such aberrant AID expression is seen in B-cell leukemia/lymphomas, including Burkitt lymphoma which is associated with c-myc translocation. Moreover, Epstein-Barr virus (EBV latent membrane protein-1 (LMP-1 increases genomic instability through early growth transcription response-1 (Egr-1 mediated upregulation of AID in B-cell lymphoma. However, few clinicopathological studies have focused on AID expression in lymphoproliferative disorders (LPDs. Therefore, we conducted an immunohistochemical study to investigate the relationship between AID and LMP-1 expression in LPDs (MTX-/Age-related EBV-associated, including diffuse large B-cell lymphomas (DLBCLs. More intense AID expression was detected in LPDs (89.5% than in DLBCLs (20.0%, and the expression of LMP-1 and EBER was more intense in LPDs (68.4% and 94.7% than in DLBCLs (10.0% and 20.0%. Furthermore, stronger Egr-1 expression was found in MTX/Age-EBV-LPDs (83.3% than in DLBCLs (30.0%. AID expression was significantly constitutively overexpressed in LPDs as compared with DLBCLs. These results suggest that increased AID expression in LPDs may be one of the processes involved in lymphomagenesis, thereby further increasing the survival of genetically destabilized B-cells. AID expression may be a useful indicator for differentiation between LPDs and DLBCLs.

  2. Prediction and identification of Taenia solium oncosphere TSO45-4B antigen FnⅢ structure domain linear B cell epitopes%猪带绦虫六钩蚴TSO45-4B抗原FnⅢ结构域线性B细胞表位预测及鉴定

    Institute of Scientific and Technical Information of China (English)

    王媛媛; 王小莉; 常雪莲; 陶志勇; 夏惠; 方强

    2012-01-01

    Objective To predict and identify Taenia solium oncosphere TSO45-4B antigen FnⅢ structure domain linear B cell epitopes. Methods The B cell epitopes of TSO45-4B FnⅢ structure domain were predicted through the sequence analysis by using bioinformatics online tools and the protein space conformation was predicted by SWISS-MODEL. The peptides were synthesized according to the predicted linear epitopes. The immunoreactivity of sera of cysticercosis patients to the peptides synthesized was tested by using ELISA. Results Two linear B cell epitopes of TSO45-4B FnⅢ structure domain were predicted, and one of the predicted epitope peptides synthesized could be recognized by the sera of cysticercosis patients. Conclusion Two linear B cell epitopes of TSO45-4B Fn Ⅲ structure domain are predicted and one of them has been confirmed successfully.%目的 预测并鉴定猪带绦虫六钩蚴TSO45-4B抗原FnⅢ结构域线性B细胞表位.方法 利用生物信息学在线工具分析TSO45-4B抗原FnⅢ结构域序列并预测其线性B细胞表位;采用SWISS-MODEL软件预测TSO45-4B的蛋白空间结构,并合成两条预测表位肽.采用酶联免疫吸附试验(ELISA)检测猪囊尾蚴病人血清与两条预测表位肽的免疫反应性.结果 获得2个TSO45-4B抗原FnⅢ结构域线性B细胞预测表位,其中1条预测表位合成肽可为猪囊尾蚴病患者血清识别.结论 成功预测并鉴定了猪带绦虫六钩蚴TSO45-4B抗原FnⅢ结构域线性B细胞表位.

  3. The Mycobacterium tuberculosis PPE protein Rv1168c induces stronger B cell response than Rv0256c in active TB patients.

    Science.gov (United States)

    Abraham, Philip Raj; Udgata, Atul; Latha, Gaddam Suman; Mukhopadhyay, Sangita

    2016-06-01

    Tuberculosis (TB) caused by Mycobacterium tuberculosis is a serious global health problem and is responsible for millions of deaths every year. For effective control of this dreadful disease, it is necessary to diagnose TB cases at the initial stages of infection. The serodiagnosis of disease represents simple, rapid and inexpensive method that can be used at the primary health care levels. In this study we have compared sensitivity of two PPE proteins of M. tuberculosis, i.e., Rv0256c and Rv1168c for their use as serodiagnostic markers in active tuberculosis patients. Employing a standardized enzyme immunoassay with these PPE proteins as candidate antigens we were able to successfully discriminate the TB patients' sera from the BCG-vaccinated healthy controls. Further, we observed that Rv1168c displayed higher sensitivity in detecting extrapulmonary and smear negative pulmonary TB cases which are difficult to diagnose by available diagnostic methods. Overall the study highlights that Rv1168c can be used as a potential serodiagnostic marker for the diagnosis of active tuberculosis disease. PMID:26364913

  4. 非经典性NF-κB活性在慢性B淋巴细胞白血病中的作用%Function of alternative NF-κB activity in B-cell chronic lymphocytic leukemia cells

    Institute of Scientific and Technical Information of China (English)

    徐晶晶; 周鹏; 国风

    2014-01-01

    Objective To investigate the function of alternative NF-κB activity in B-cell chronic lymphocytic leukemia cells (B-CLL).Methods The mRNA expression of individual NF-κB subunits in CD5+CD19+ cells (CLL B-cells) from bone marrow (BM) of 56 patients with B-CLL was analyzed by quantitative RT-PCR.An ELISA-based NF-κB family transcription factor activity assay was performed to quantify the κB DNA-binding activity in nuclear extracts from CLL B-cells.Cell death of CLL-B cells was determined by PI staining,RelA and RelB expression at protein level of CLL B-cells by Western blot analyses.Results The expression levels of RelA,p50,RelB and p52 mRNA in CLL B-cells were all higher than that of normal B cells with statistical significance (P<0.05).RelA was activated in almost all the patients detected while RelB activity was induced in part of samples.The average RelA activity in CLL B-cells was increased compared to that in normal B cells while the average RelB activity was similar to that of normal B cells.When cultured in vitro for 24,48 and 72 hours,the frequencies of cell death of CLL B-cells from RelA+/RelB-group were (35.54±4.43)%,(50.92±8.44)%,and (49.24±8.16)%,respectively;that of the RelA+/RelB+ group were (20.65±2.37)%,(18.17± 1.36)%,and (26.55±4.08)%,respectively.When the cells from RelA+/RelB-group were co-cultured with bone marrow stromal cells (hBMSCs),the frequencies of cell death of CLL B-cells were decreased compared to that of the cells cultured alone,while the frequencies of cell death of RelA+/RelB-CLL B-cells were higher than that of CLL B-cells from RelA+/ RelB+ group when co-cultured with hBMSCs.RelA and RelB expression in CLL-B cells from the RelA+/RelB-group was induced after co-cultured with hBMSCs for 48h.RelB was reduced in the cytoplasm and increased in the nucleus in CLL-B cells from the RelA+/RelB+ group.Conclusion The alternative NF-κB was indeed activated and presented heterogenous in CLL B-cells from BM.Activation of

  5. Merkel cell polyomavirus large T antigen has growth-promoting and inhibitory activities.

    Science.gov (United States)

    Cheng, Jingwei; Rozenblatt-Rosen, Orit; Paulson, Kelly G; Nghiem, Paul; DeCaprio, James A

    2013-06-01

    Merkel cell carcinoma (MCC) is a rare and aggressive form of skin cancer. In at least 80% of all MCC, Merkel cell polyomavirus (MCPyV) DNA has undergone clonal integration into the host cell genome, and most tumors express the MCPyV large and small T antigens. In all cases of MCC reported to date, the integrated MCPyV genome has undergone mutations in the large T antigen. These mutations result in expression of a truncated large T antigen that retains the Rb binding or LXCXE motif but deletes the DNA binding and helicase domains. However, the transforming functions of full-length and truncated MCPyV large T antigen are unknown. We compared the transforming activities of full-length, truncated, and alternatively spliced 57kT forms of MCPyV large T antigen. MCPyV large T antigen could bind to Rb but was unable to bind to p53. Furthermore, MCPyV-truncated large T antigen was more effective than full-length and 57kT large T antigen in promoting the growth of human and mouse fibroblasts. In contrast, expression of the MCPyV large T antigen C-terminal 100 residues could inhibit the growth of several different cell types. These data imply that the deletion of the C terminus of MCPyV large T antigen found in MCC serves not only to disrupt viral replication but also results in the loss of a distinct growth-inhibitory function intrinsic to this region. PMID:23514892

  6. Suppressive effects of antigens on the activity of specific activated lymphocytes: A test to define the specificity of activated lymphocytes

    Institute of Scientific and Technical Information of China (English)

    HU Jun; PAN Sheng-jun; CAI Zhen-jie; GUAN De-lin; LIU Xiao-cheng

    2006-01-01

    Objective:With the regular mixed lymphocytes culture (MLC) to detect the allograft rejection, the reactivity of the activated lymphocytes (primed lymphocytes) of a recipient shows sometimes increase and sometimes decrease against the antigens from the donor, which is inconsistent with the clinical results. In order to establish a convenient method for testing the specificity of the activated lymphocytes in vitro, so as to know the rejection occurred or not by testing the existence of the specific activated lymphocytes against donor's HLA antigens in the recipient's peripheral blood. Methods: Anti-IL-2 neutralizing monoclonal antibody (anti-IL-2 N-mAb) and immunosuppressors were introduced in this test system in the presence of specific stimulators and activated lymphocytes. Results: When the activated lymphocytes were chosen from the one-way MLC 4 d to undergo re-stimulation by specific stimulators, the activity of activated lymphocytes in the treatment group was suppressed significantly compared with that in the control group. The result of this test method is consistent with the biopsy in the clinical diagnosis of rejection.Conclusion :It suggests that the activated lymphocytes can be inactivated by specific antigens in certain conditions. This can be a useful tool to define the specificity of the activated lymphocytes.

  7. Regulation of B cell differentiation by the ubiquitin-binding protein TAX1BP1

    Science.gov (United States)

    Matsushita, Nobuko; Suzuki, Midori; Ikebe, Emi; Nagashima, Shun; Inatome, Ryoko; Asano, Kenichi; Tanaka, Masato; Matsushita, Masayuki; Kondo, Eisaku; Iha, Hidekatsu; Yanagi, Shigeru

    2016-01-01

    Tax1-binding protein 1 (TAX1BP1) is a ubiquitin-binding protein that restricts nuclear factor-κB (NF-κB) activation and facilitates the termination of aberrant inflammation. However, its roles in B-cell activation and differentiation are poorly understood. To evaluate the function of TAX1BP1 in B cells, we established TAX1BP1-deficient DT40 B cells that are hyper-responsive to CD40-induced extracellular signal-regulated kinase (ERK) activation signaling, exhibit prolonged and exaggerated ERK phosphorylation and show enhanced B lymphocyte-induced maturation protein 1 (Blimp-1; a transcription factor inducing plasma cell differentiation) expression that is ERK-dependent. Furthermore, TAX1BP1-deficient cells exhibit significantly decreased surface IgM expression and increased IgM secretion. Moreover, TAX1BP1-deficient mice display reduced germinal center formation and antigen-specific antibody production. These findings show that TAX1BP1 restricts ERK activation and Blimp-1 expression and regulates germinal center formation. PMID:27515252

  8. Construction and Screening of Antigen Targeted Immune Yeast Surface Display Antibody Libraries

    Energy Technology Data Exchange (ETDEWEB)

    Miller, Keith D.; Pefaur, Noah B.; Baird, Cheryl L.

    2008-07-01

    These protocols describe a yeast surface display-based process for the rapid selection of antibodies from immunized mice, eliminating the need for creating and screening hybridoma fusions. A yeast surface display library of single-chain antibody fragments (scFvs) is created from antigen-binding B cells from the splenocytes of immunized mice. The antigen targeted library is then screened for antigen specific scFv by magnetic activated cell sorting (MACS) and fluorescence activated cell sorting (FACS). Library construction and screening can be accomplished in as little as 2 weeks resulting in a panel of scFvs specific for the target antigen.

  9. CEACAM1 induces B-cell survival and is essential for protective antiviral antibody production

    Science.gov (United States)

    Khairnar, Vishal; Duhan, Vikas; Maney, Sathish Kumar; Honke, Nadine; Shaabani, Namir; Pandyra, Aleksandra A.; Seifert, Marc; Pozdeev, Vitaly; Xu, Haifeng C.; Sharma, Piyush; Baldin, Fabian; Marquardsen, Florian; Merches, Katja; Lang, Elisabeth; Kirschning, Carsten; Westendorf, Astrid M.; Häussinger, Dieter; Lang, Florian; Dittmer, Ulf; Küppers, Ralf; Recher, Mike; Hardt, Cornelia; Scheffrahn, Inka; Beauchemin, Nicole; Göthert, Joachim R.; Singer, Bernhard B.; Lang, Philipp A.; Lang, Karl S.

    2015-01-01

    B cells are essential for antiviral immune defence because they produce neutralizing antibodies, present antigen and maintain the lymphoid architecture. Here we show that intrinsic signalling of CEACAM1 is essential for generating efficient B-cell responses. Although CEACAM1 exerts limited influence on the proliferation of B cells, expression of CEACAM1 induces survival of proliferating B cells via the BTK/Syk/NF-κB-axis. The absence of this signalling cascade in naive Ceacam1−/− mice limits the survival of B cells. During systemic infection with cytopathic vesicular stomatitis virus, Ceacam1−/− mice can barely induce neutralizing antibody responses and die early after infection. We find, therefore, that CEACAM1 is a crucial regulator of B-cell survival, influencing B-cell numbers and protective antiviral antibody responses. PMID:25692415

  10. CEACAM1 induces B-cell survival and is essential for protective antiviral antibody production.

    Science.gov (United States)

    Khairnar, Vishal; Duhan, Vikas; Maney, Sathish Kumar; Honke, Nadine; Shaabani, Namir; Pandyra, Aleksandra A; Seifert, Marc; Pozdeev, Vitaly; Xu, Haifeng C; Sharma, Piyush; Baldin, Fabian; Marquardsen, Florian; Merches, Katja; Lang, Elisabeth; Kirschning, Carsten; Westendorf, Astrid M; Häussinger, Dieter; Lang, Florian; Dittmer, Ulf; Küppers, Ralf; Recher, Mike; Hardt, Cornelia; Scheffrahn, Inka; Beauchemin, Nicole; Göthert, Joachim R; Singer, Bernhard B; Lang, Philipp A; Lang, Karl S

    2015-01-01

    B cells are essential for antiviral immune defence because they produce neutralizing antibodies, present antigen and maintain the lymphoid architecture. Here we show that intrinsic signalling of CEACAM1 is essential for generating efficient B-cell responses. Although CEACAM1 exerts limited influence on the proliferation of B cells, expression of CEACAM1 induces survival of proliferating B cells via the BTK/Syk/NF-κB-axis. The absence of this signalling cascade in naive Ceacam1(-/-) mice limits the survival of B cells. During systemic infection with cytopathic vesicular stomatitis virus, Ceacam1(-/-) mice can barely induce neutralizing antibody responses and die early after infection. We find, therefore, that CEACAM1 is a crucial regulator of B-cell survival, influencing B-cell numbers and protective antiviral antibody responses. PMID:25692415

  11. Regulation of murine macrophage Ia-antigen expression by products of activated spleen cells

    OpenAIRE

    1980-01-01

    This investigation examined the effects of mediators derived form activated spleen cells on macrophage Ia-antigen expression and function. Incubation of adherent thioglycollate-induced murine peritoneal macrophages(> 90% Ia-) with concanavalin A (Con A)- stimulated spleen cell supernate (Con A sup) resulted in a dose- dependent increase in the percentage of Ia-containing (Ia+) phagocytic cells, as detected by antiserum-and-complement-mediated cytotoxicity. The Ia-antigen expression of macroph...

  12. Human herpesvirus-8 infection leads to expansion of the preimmune/natural effector B cell compartment.

    Directory of Open Access Journals (Sweden)

    Silvia Della Bella

    Full Text Available BACKGROUND: Human herpesvirus-8 (HHV-8 is the etiological agent of Kaposi's sarcoma (KS and of some lymphoproliferative disorders of B cells. Most malignancies develop after long-lasting viral dormancy, and a preventing role for both humoral and cellular immune control is suggested by the high frequency of these pathologies in immunosuppressed patients. B cells, macrophages and dendritic cells of peripheral lymphoid organs and blood represent the major reservoir of HHV-8. Due to the dual role of B cells in HHV-8 infection, both as virus reservoir and as agents of humoral immune control, we analyzed the subset distribution and the functional state of peripheral blood B cells in HHV-8-infected individuals with and without cKS. METHODOLOGY/PRINCIPAL FINDINGS: Circulating B cells and their subsets were analyzed by 6-color flow cytometry in the following groups: 1- patients HHV-8 positive with classic KS (cKS (n = 47; 2- subjects HHV-8 positive and cKS negative (HSP (n = 10; 3- healthy controls, HHV-8 negative and cKS negative (HC (n = 43. The number of B cells belonging to the preimmune/natural effector compartment, including transitional, pre-naïve, naïve and MZ-like subsets, was significantly higher among HHV-8 positive subjects, with or without cKS, while was comparable to healthy controls in the antigen-experienced T-cell dependent compartment. The increased number of preimmune/natural effector B cells was associated with increased resistance to spontaneous apoptosis, while it did not correlate with HHV-8 viral load. CONCLUSIONS/SIGNIFICANCE: Our results indicate that long-lasting HHV-8 infection promotes an imbalance in peripheral B cell subsets, perturbing the equilibrium between earlier and later steps of maturation and activation processes. This observation may broaden our understanding of the complex interplay between viral and immune factors leading HHV-8-infected individuals to develop HHV-8-associated malignancies.

  13. Systemic Inflammation in Progressive Multiple Sclerosis Involves Follicular T-Helper, Th17- and Activated B-Cells and Correlates with Progression

    DEFF Research Database (Denmark)

    Romme Christensen, Jeppe; Börnsen, Lars; Ratzer, Rikke;

    2013-01-01

    Pathology studies of progressive multiple sclerosis (MS) indicate a major role of inflammation including Th17-cells and meningeal inflammation with ectopic lymphoid follicles, B-cells and plasma cells, the latter indicating a possible role of the newly identified subset of follicular T-helper (TF...... pathogenic role of systemic inflammation in progressive MS. These observations may have implications for the treatment of progressive MS....

  14. Cadmium induces carcinogenesis in BEAS-2B cells through ROS-dependent activation of PI3K/AKT/GSK-3β/β-catenin signaling

    OpenAIRE

    Son, Young-Ok; Wang, Lei; Poyil, Pratheeshkumar; Budhraja, Amit; Hitron, J. Andrew; Zhang, Zhuo; Lee, Jeong-Chae; Shi, Xianglin

    2012-01-01

    Cadmium has been widely used in industry and is known to be carcinogenic to humans. Although it is widely accepted that chronic exposure to cadmium increases the incidence of cancer, the mechanisms underlying cadmium-induced carcinogenesis are unclear. The main aim of this study was to investigate the role of reactive oxygen species (ROS) in cadmium-induced carcinogenesis and the signal transduction pathways involved. Chronic exposure of human bronchial epithelial BEAS-2B cells to cadmium ind...

  15. Translocations activating IRF4 identify a subtype of germinal center-derived B-cell lymphoma affecting predominantly children and young adults.

    Science.gov (United States)

    Salaverria, Itziar; Philipp, Claudia; Oschlies, Ilske; Kohler, Christian W; Kreuz, Markus; Szczepanowski, Monika; Burkhardt, Birgit; Trautmann, Heiko; Gesk, Stefan; Andrusiewicz, Miroslaw; Berger, Hilmar; Fey, Miriam; Harder, Lana; Hasenclever, Dirk; Hummel, Michael; Loeffler, Markus; Mahn, Friederike; Martin-Guerrero, Idoia; Pellissery, Shoji; Pott, Christiane; Pfreundschuh, Michael; Reiter, Alfred; Richter, Julia; Rosolowski, Maciej; Schwaenen, Carsten; Stein, Harald; Trümper, Lorenz; Wessendorf, Swen; Spang, Rainer; Küppers, Ralf; Klapper, Wolfram; Siebert, Reiner

    2011-07-01

    The prognosis of germinal center-derived B-cell (GCB) lymphomas, including follicular lymphoma and diffuse large-B-cell lymphoma (DLBCL), strongly depends on age. Children have a more favorable outcome than adults. It is not known whether this is because of differences in host characteristics, treatment protocols, or tumor biology, including the presence of chromosomal alterations. By screening for novel IGH translocation partners in pediatric and adult lymphomas, we identified chromosomal translocations juxtaposing the IRF4 oncogene next to one of the immunoglobulin (IG) loci as a novel recurrent aberration in mature B-cell lymphoma. FISH revealed 20 of 427 lymphomas to carry an IG/IRF4-fusion. Those were predominantly GCB-type DLBCL or follicular lymphoma grade 3, shared strong expression of IRF4/MUM1 and BCL6, and lacked PRDM1/BLIMP1 expression and t(14;18)/BCL2 breaks. BCL6 aberrations were common. The gene expression profile of IG/IRF4-positive lymphomas differed from other subtypes of DLBCL. A classifier for IG/IRF4 positivity containing 27 genes allowed accurate prediction. IG/IRF4 positivity was associated with young age and a favorable outcome. Our results suggest IRF4 translocations to be primary alterations in a molecularly defined subset of GCB-derived lymphomas. The probability for this subtype of lymphoma significantly decreases with age, suggesting that diversity in tumor biology might contribute to the age-dependent differences in prognosis of lymphoma. PMID:21487109

  16. Binding of recombinant T cell receptor ligands (RTL) to antigen presenting cells prevents upregulation of CD11b and inhibits T cell activation and transfer of experimental autoimmune encephalomyelitis.

    Science.gov (United States)

    Sinha, Sushmita; Miller, Lisa; Subramanian, Sandhya; McCarty, Owen J T; Proctor, Thomas; Meza-Romero, Roberto; Huan, Jianya; Burrows, Gregory G; Vandenbark, Arthur A; Offner, Halina

    2010-08-25

    Recombinant T cell ligands (RTLs) ameliorate experimental autoimmune encephalomyelitis (EAE) in an antigen-specific manner. We evaluated effects of RTL401 (I-A(s) alpha1beta1+PLP-139-151) on splenocytes from SJL/J mice with EAE to study RTL-T cell tolerance-inducing mechanisms. RTLs bound to B, macrophages and DCs, through RTL-MHC-alpha1beta1 moiety. RTL binding reduced CD11b expression on splenic macrophages/DC, and RTL401-conditioned macrophages/DC, not B cells, inhibited T cell activation. Reduced ability of RTL- incubated splenocytes to transfer EAE was likely mediated through macrophages/DC, since B cells were unnecessary for RTL treatment of EAE. These results demonstrate a novel pathway of T cell regulation by RTL-bound APCs. PMID:20546940

  17. Anti-PEG IgM and complement system are required for the association of second doses of PEGylated liposomes with splenic marginal zone B cells.

    Science.gov (United States)

    Shimizu, Taro; Mima, Yu; Hashimoto, Yosuke; Ukawa, Masami; Ando, Hidenori; Kiwada, Hiroshi; Ishida, Tatsuhiro

    2015-10-01

    The accelerated blood clearance (ABC) phenomenon makes it crucial to use PEGylated liposomes and micelles to deliver drugs. The ABC phenomenon is an immune response against an initial dose of PEGylated liposome, which causes subsequent doses to be rapidly cleared by macrophages in the liver. We recently found that in the early phase of the ABC phenomenon, subsequent doses of PEGylated liposomes were associated with splenic marginal zone (MZ)-B cells and were transported from the MZ to the follicle (FO). In this study, we investigated the underlying mechanisms behind the association of subsequent doses of PEGylated liposomes with MZ-B cells in the spleen. Serum factors, anti-PEG IgM and complement system, were crucial to the association of PEGylated liposomes with MZ-B cells, while the sensitization of MZ-B cells by the first dose of PEGylated liposomes was not significant. It was the complement receptors (CRs) on the MZ-B cells, rather than either the PEG-specific B-cell receptors or the IgM Fc receptors, that were the main contributors to the association between PEGylated liposomes and MZ-B cells. It appeared that anti-PEG IgM would bind to PEGylated liposomes and causes subsequent complement activation, resulting in the formation of immune complexes of PEGylated liposome-anti-PEG IgM-complement. The MZ-B cells then recognized these immune complexes via their CRs. Such an association via CRs might have triggered the transport of the immune complex by MZ-B cells to the FO in the spleen. The information obtained in this study might be useful in the development of an efficient antigen delivery system to usher PEGylated nanoparticles into FO dendritic cells. PMID:26095176

  18. Uptake and presentation of myelin basic protein by normal human B cells.

    Directory of Open Access Journals (Sweden)

    Marie Klinge Brimnes

    Full Text Available B cells may play both pathogenic and protective roles in T-cell mediated autoimmune diseases such as multiple sclerosis (MS. These functions relate to the ability of B cells to bind and present antigens. Under serum-free conditions we observed that 3-4% of circulating B cells from healthy donors were capable of binding the MS-associated self-antigen myelin basic protein (MBP and of presenting the immunodominant peptide MBP85-99, as determined by staining with the mAb MK16 recognising the peptide presented by HLA-DR15-positive cells. In the presence of serum, however, the majority of B cells bound MBP in a complement-dependent manner, and almost half of the B cells became engaged in presentation of MBP85-99. Even though complement receptor 1 (CR1, CD35 and CR2 (CD21 both contributed to binding of MBP to B cells, only CR2 was important for the subsequent presentation of MBP85-99. A high proportion of MBP85-99 presenting B cells expressed CD27, and showed increased expression of CD86 compared to non-presenting B cells. MBP-pulsed B cells induced a low frequency of IL-10-producing CD4+ T cells in 3 out of 6 donors, indicating an immunoregulatory role of B cells presenting MBP-derived peptides. The mechanisms described here refute the general assumption that B-cell presentation of self-antigens requires uptake via specific B-cell receptors, and may be important for maintenance of tolerance as well as for driving T-cell responses in autoimmune diseases.

  19. KSHV-Mediated Regulation of Par3 and SNAIL Contributes to B-Cell Proliferation

    Science.gov (United States)

    Jha, Hem C.; Sun, Zhiguo; Upadhyay, Santosh K.; El-Naccache, Darine W.; Singh, Rajnish K.; Sahu, Sushil K.; Robertson, Erle S.

    2016-01-01

    Studies have suggested that Epithelial–Mesenchymal Transition (EMT) and transformation is an important step in progression to cancer. Par3 (partitioning-defective protein) is a crucial factor in regulating epithelial cell polarity. However, the mechanism by which the latency associated nuclear antigen (LANA) encoded by Kaposi's Sarcoma associated herpesvirus (KSHV) regulates Par3 and EMTs markers (Epithelial-Mesenchymal Transition) during viral-mediated B-cell oncogenesis has not been fully explored. Moreover, several studies have demonstrated a crucial role for EMT markers during B-cell malignancies. In this study, we demonstrate that Par3 is significantly up-regulated in KSHV-infected primary B-cells. Further, Par3 interacted with LANA in KSHV positive and LANA expressing cells which led to translocation of Par3 from the cell periphery to a predominantly nuclear signal. Par3 knockdown led to reduced cell proliferation and increased apoptotic induction. Levels of SNAIL was elevated, and E-cadherin was reduced in the presence of LANA or Par3. Interestingly, KSHV infection in primary B-cells led to enhancement of SNAIL and down-regulation of E-cadherin in a temporal manner. Importantly, knockdown of SNAIL, a major EMT regulator, in KSHV cells resulted in reduced expression of LANA, Par3, and enhanced E-cadherin. Also, SNAIL bound to the promoter region of p21 and can regulate its activity. Further a SNAIL inhibitor diminished NF-kB signaling through upregulation of Caspase3 in KSHV positive cells in vitro. This was also supported by upregulation of SNAIL and Par3 in BC-3 transplanted NOD-SCID mice which has potential as a therapeutic target for KSHV-associated B-cell lymphomas. PMID:27463802

  20. Improved Method for Linear B-Cell Epitope Prediction Using Antigen’s Primary Sequence

    OpenAIRE

    Singh, Harinder; Ansari, Hifzur Rahman; Gajendra P. S. Raghava

    2013-01-01

    One of the major challenges in designing a peptide-based vaccine is the identification of antigenic regions in an antigen that can stimulate B-cell’s response, also called B-cell epitopes. In the past, several methods have been developed for the prediction of conformational and linear (or continuous) B-cell epitopes. However, the existing methods for predicting linear B-cell epitopes are far from perfection. In this study, an attempt has been made to develop an improved method for predicting ...

  1. Tissue-specific B-cell dysfunction and generalized memory B-cell loss during acute SIV infection.

    Directory of Open Access Journals (Sweden)

    Sandrine Peruchon

    organs. Characterization of underlying mechanisms would be helpful in designing new therapeutic strategies to dampen B-cell activation and increases HIV/SIV specific antibody response.

  2. Inhibitory activities of microalgal extracts against Epstein-Barr Virus (EBV antigen expression in lymphoblastoid cells

    Directory of Open Access Journals (Sweden)

    Koh Yih Yih

    2014-01-01

    Full Text Available The inhibitory activities of microalgal extracts against the expression of three EBV antigens, latent membrane protein (LMP1, Epstein-Barr nuclear antigen (EBNA1 and Z Epstein-Barr reactivation activator (ZEBRA were assessed by immunocytochemistry. The observation that the methanol extracts and their fractions from Ankistrodesmus convolutus, Synechococcus elongatus and Spirulina platensis exhibited inhibitory activity against EBV proteins in three Burkitt’s lymphoma cell lines at concentrations as low as 20 μg/ml suggests that microalgae could be a potential source of antiviral compounds against EBV.

  3. Molecular definition of a polymorphic antigen (LA45) of free HLA-A and -B heavy chains found on the surfaces of activated B and T cells.

    Science.gov (United States)

    Madrigal, J A; Belich, M P; Benjamin, R J; Little, A M; Hildebrand, W H; Mann, D L; Parham, P

    1991-11-01

    A monomoprhic monoclonal antibody (LA45 antibody) reactive with "a new activation-induced surface structure on human T lymphocytes" (LA45 antigen) that resembled free class I heavy chains has recently been described (Schnabl, E., H. Stockinger, O. Majdic, H. Gaugitsch, I.J.D. Lindley, D. Maurer, A. Hajek-Rosenmayr, and W. Knapp. 1990. J. Exp. Med. 171:1431). This antibody was used to clone a class I-like heavy chain (LA45 gene) from the HUT 102 tumor cell, which paradoxically did not give rise to the LA45 antigen on transfection into monkey COS cells. We show here that the LA45 gene is HLA-Aw66.2, a previously uncharacterized allele of the HLA-A locus. The previously determined LA45 sequence differs from that of HLA-Aw66.2, from HUT 102, and the CR-B B cell line derived from the same individual as HUT 102 by substitution of tryptophan for serine at position 4 in the alpha 1 domain. Transfection of HLA-Aw66.2, and of a mutant of this gene with serine 4 substituted for tryptophan, into a human B cell line (C1R) both resulted in expression of the LA45 epitope. Furthermore, we find expression of the LA45 epitope on Epstein Barr virus-transformed B cell lines as well as lectin-activated T cells, but not on long-term T cell lines or unstimulated peripheral blood T cells. The specificity of the LA45 antibody is polymorphic and the presence of the LA45 epitope is precisely correlated with the sequence arginine, asparagine (RN) at residues 62 and 63 of the helix of the alpha 1 domain. The LA45 epitope is broadly distributed, being associated with half the alleles of both HLA-A and -B loci but none of the HLA-C locus. All the results are consistent with the presence of pools of free HLA-A and -B heavy chains at the surfaces of certain cell types but not others. Such molecules are probably responsible for the HLA-associated class I alloantigens of lectin-activated T cells. We hypothesize the free heavy chains result from dissociation of beta 2-microglobulin from

  4. Binding of recombinant T cell receptor ligands (RTL) to antigen presenting cells prevents upregulation of CD11b and inhibits T cell activation and transfer of experimental autoimmune encephalomyelitis

    OpenAIRE

    Sinha, Sushmita; Miller, Lisa; Subramanian, Sandhya; McCarty, Owen; Proctor, Thomas; Meza-Romero, Roberto; Burrows, Gregory G.; Vandenbark, Arthur A.; Offner, Halina

    2010-01-01

    Recombinant T cell ligands (RTLs) ameliorate experimental autoimmune encephalomyelitis (EAE) in antigen specific manner. We evaluated effects of RTL401 (I-As α1β1 + PLP-139-151) on splenocytes from mice with EAE to study RTL- T cell-tolerance-inducing mechanisms. RTLs bound to B, macrophages and DCs, through RTL-MHC-α1β1 moiety. RTL binding reduced CD11b expression on splenic macrophages/DC, and RTL401-conditioned macrophages/DC, not B cells, inhibited T cell activation. Reduced ability of RT...

  5. Evolution of B Cell Immunity

    OpenAIRE

    Parra, David; Takizawa, Fumio; Sunyer, J Oriol

    2013-01-01

    Two types of adaptive immune strategies are known to have evolved in vertebrates: the VLR-based system, which is present in jawless organisms and is mediated by VLRA and VLRB lymphocytes, and the BCR/TCR-based system, which is present in jawed species and is provided by B and T cell receptors expressed on B and T cells, respectively. Here we summarize features of B cells and their predecessors in the different animal phyla, focusing the review on B cells from jawed vertebrates. We point out t...

  6. CEACAM1 induces B-cell survival and is essential for protective antiviral antibody production

    OpenAIRE

    Khairnar, Vishal; Duhan, Vikas; Maney, Sathish Kumar; Honke, Nadine; Shaabani, Namir; Pandyra, Aleksandra A; Seifert, Marc; Pozdeev, Vitaly; Xu, Haifeng C.; Sharma, Piyush; Baldin, Fabian; Marquardsen, Florian; Merches, Katja; Lang, Elisabeth; Kirschning, Carsten

    2015-01-01

    B cells are essential for antiviral immune defence because they produce neutralizing antibodies, present antigen and maintain the lymphoid architecture. Here we show that intrinsic signalling of CEACAM1 is essential for generating efficient B-cell responses. Although CEACAM1 exerts limited influence on the proliferation of B cells, expression of CEACAM1 induces survival of proliferating B cells via the BTK/Syk/NF-κB-axis. The absence of this signalling cascade in naive Ceacam1 −/− mice limits...

  7. Primary B-cell deficiencies reveal a link between human IL-17-producing CD4 T-cell homeostasis and B-cell differentiation.

    Directory of Open Access Journals (Sweden)

    Rita R Barbosa

    Full Text Available IL-17 is a pro-inflammatory cytokine implicated in autoimmune and inflammatory conditions. The development/survival of IL-17-producing CD4 T cells (Th17 share critical cues with B-cell differentiation and the circulating follicular T helper subset was recently shown to be enriched in Th17 cells able to help B-cell differentiation. We investigated a putative link between Th17-cell homeostasis and B cells by studying the Th17-cell compartment in primary B-cell immunodeficiencies. Common Variable Immunodeficiency Disorders (CVID, defined by defects in B-cell differentiation into plasma and memory B cells, are frequently associated with autoimmune and inflammatory manifestations but we found no relationship between these and Th17-cell frequency. In fact, CVID patients showed a decrease in Th17-cell frequency in parallel with the expansion of activated non-differentiated B cells (CD21(lowCD38(low. Moreover, Congenital Agammaglobulinemia patients, lacking B cells due to impaired early B-cell development, had a severe reduction of circulating Th17 cells. Finally, we found a direct correlation in healthy individuals between circulating Th17-cell frequency and both switched-memory B cells and serum BAFF levels, a crucial cytokine for B-cell survival. Overall, our data support a relationship between Th17-cell homeostasis and B-cell maturation, with implications for the understanding of the pathogenesis of inflammatory/autoimmune diseases and the physiology of B-cell depleting therapies.

  8. Construction and Screening of Antigen Targeted Immune Yeast Surface Display Antibody Libraries

    Energy Technology Data Exchange (ETDEWEB)

    Miller, Keith D.; Pefaur, Noah B.; Baird, Cheryl L.

    2009-08-02

    These protocols describe a yeast surface display-based process for the rapid selection of antibodies from immunized mice, eliminating the need for creating and screening hybridoma fusions. A yeast surface display library of single-chain antibody fragments (scFvs) is created from antigen-binding B cells from the splenocytes of immunized mice. The antigen targeted library is then screened for antigen specific scFv by magneticactivated cell sorting (MACS) and fluorescence-activated cell sorting (FACS). Library construction and screening can be accomplished in as little as 2 weeks, resulting in a panel of scFvs specific for the target antigen.

  9. iNKT-cell help to B cells: a cooperative job between innate and adaptive immune responses.

    Science.gov (United States)

    Dellabona, Paolo; Abrignani, Sergio; Casorati, Giulia

    2014-08-01

    T-cell help to B lymphocytes is one of the most important events in adaptive immune responses in health and disease. It is generally delivered by cognate CD4(+) T follicular helper (T(FH)) cells via both cell-to-cell contacts and soluble mediators, and it is essential for both the clonal expansion of antibody (Ab)-secreting B cells and memory B-cell formation. CD1d-restricted invariant natural killer T (iNKT) cells are a subset of innate-like T lymphocytes that rapidly respond to stimulation with specific lipid antigens (Ags) that are derived from infectious pathogens or stressed host cells. Activated iNKT cells produce a wide range of cytokines and upregulate costimulatory molecules that can promote activation of dendritic cells (DCs), natural killer (NK) cells, and T cells. A decade ago, we discovered that iNKT cells can help B cells to proliferate and to produce IgG Abs in vitro and in vivo. This adjuvant-like function of Ag-activated iNKT cells provides a flexible set of helper mechanisms that expand the current paradigm of T-cell-B-cell interaction and highlights the potential of iNKT-cell targeting vaccine formulations. PMID:24782127

  10. How to Make a Non-Antigenic Protein (Auto) Antigenic: Molecular Complementarity Alters Antigen Processing and Activates Adaptive-Innate Immunity Synergy.

    Science.gov (United States)

    Root-Bernstein, Robert

    2015-01-01

    Evidence is reviewed that complementary proteins and peptides form complexes with increased antigenicity and/or autoimmunogenicity. Five case studies are highlighted: 1) diphtheria toxin-antitoxin (antibody), which induces immunity to the normally non-antigenic toxin, and autoimmune neuritis; 2) tryptophan peptide of myelin basic protein and muramyl dipeptide ("adjuvant peptide"), which form a complex that induces experimental allergic encephalomyelitis; 3) an insulin and glucagon complex that is far more antigenic than either component individually; 4) various causes of experimental autoimmune myocarditis such as C protein in combination with its antibody, or coxsackie B virus in combination with the coxsackie and adenovirus receptor; 5) influenza A virus haemagglutinin with the outer membrane protein of the Haemophilus influenzae, which increases antigenicity. Several mechanisms cooperate to alter immunogenicity. Complexation alters antigen processing, protecting the components against proteolysis, altering fragmentation and presenting novel antigens to the immune system. Complementary antigens induce complementary adaptive immune responses (complementary antibodies and/or T cell receptors) that produce circulating immune complexes (CIC). CIC stimulate innate immunity. Concurrently, complementary antigens stimulate multiple Toll-like receptors that synergize to over-produce cytokines, which further stimulate adaptive immunity. Thus innate and adaptive immunity form a positive feedback loop. If components of the complex mimic a host protein, then autoimmunity may result. Enhanced antigenicity for production of improved vaccines and/or therapeutic autoimmunity (e.g., against cancer cells) might be achieved by using information from antibody or TCR recognition sites to complement an antigen; by panning for complements in randomized peptide libraries; or using antisense peptide strategies to design complements. PMID:26179268

  11. Microbial Translocation and B Cell Dysfunction in Human Immunodeficiency Virus Disease

    Directory of Open Access Journals (Sweden)

    Wei Jiang

    2012-01-01

    Full Text Available The gut mucosal barrier disrupted in HIV disease, resulting in increased systemic exposure to microbial products such as Lipo Polys Accharide (LPS. The association of enhanced microbial translocation and B cell dysfunction in HIV disease is not fully understood. High dose and short term exposure of microbial Toll-Like Receptor (TLR agonists were used as vaccine adjuvants, however, low dose and long term exposure of TLR agonists could be harmful. The characteristics of B cell dysfunction in HIV disease included B cell, especially memory B cell depletion, enhanced levels of autoimmune antibodies and impaired vaccine or antigen responsiveness. This review discusses and explores the possibility of the effect of microbial translocation on memory B cell depletion and impaired vaccine responses in HIV infection. By determining the mechanisms of B cell depletion and perturbations in HIV disease, it may be possible to design interventions that can improve immune responses to vaccines, reduce selected opportunistic infections and perhaps slow disease progression.

  12. Examination of antigenic structure and some biological activities of hemagglutinin-neuraminidase (hn) and fusion (f) glycoprotein antigens of parainfluenza 3 virus, in vitro

    OpenAIRE

    Milić Nenad S.; Gađanski-Omerović Gordana N.; Ašanin Ružica; Nisavić Jakov; Radojičić Marina

    2003-01-01

    The objective of our study was examination of the antigenic structure fusional and hemolytic activities of the surface glycoprotein HN and F antigens of purified parainfluenza (PI 3) viruses activated with 0.025 g/dl trypsin-versen (molecular weights of 112 kD, 81-82 kD and 30-31 kD), in vitro. The samples of activated PI3 virions with total protein concentrations of 0.55 and 0.27mg/ml and hemagglutinating titre of 256 and 128 HAU/0.1 ml, induced bovine turbinate(BT) cell fusion and hemolysis...

  13. Identification of a B cell-dependent subpopulation of multiple sclerosis by measurements of brain-reactive B cells in the blood.

    Science.gov (United States)

    Kuerten, Stefanie; Pommerschein, Giovanna; Barth, Stefanie K; Hohmann, Christopher; Milles, Bianca; Sammer, Fabian W; Duffy, Cathrina E; Wunsch, Marie; Rovituso, Damiano M; Schroeter, Michael; Addicks, Klaus; Kaiser, Claudia C; Lehmann, Paul V

    2014-01-01

    B cells are increasingly coming into play in the pathogenesis of multiple sclerosis (MS). Here, we screened peripheral blood mononuclear cells (PBMC) from patients with clinically isolated syndrome (CIS), MS, other non-inflammatory neurological, inflammatory neurological or autoimmune diseases, and healthy donors for their B cell reactivity to CNS antigen using the enzyme-linked immunospot technique (ELISPOT) after 96 h of polyclonal stimulation. Our data show that nine of 15 patients with CIS (60.0%) and 53 of 67 patients with definite MS (79.1%) displayed CNS-reactive B cells, compared to none of the control donors. The presence of CNS-reactive B cells in the blood of the majority of patients with MS or at risk to develop MS along with their absence in control subjects suggests that they might be indicative of a B cell-dependent subpopulation of the disease. PMID:24607792

  14. Microsomal triglyceride transfer protein lipidation and control of CD1d on antigen-presenting cells

    OpenAIRE

    Dougan, Stephanie K.; Salas, Azucena; Rava, Paul; Agyemang, Amma; Kaser, Arthur; Morrison, Jamin; Khurana, Archana; Kronenberg, Mitchell; Johnson, Caroline; Exley, Mark; Hussain, M. Mahmood; Blumberg, Richard S.

    2005-01-01

    Microsomal triglyceride transfer protein (MTP), an endoplasmic reticulum (ER) chaperone that loads lipids onto apolipoprotein B, also regulates CD1d presentation of glycolipid antigens in the liver and intestine. We show MTP RNA and protein in antigen-presenting cells (APCs) by reverse transcription–polymerase chain reaction and by immunoblotting of mouse liver mononuclear cells and mouse and human B cell lines. Functional MTP, demonstrated by specific triglyceride transfer activity, is prese...

  15. Antigen receptor signaling: integration of protein tyrosine kinase functions.

    Science.gov (United States)

    Tamir, I; Cambier, J C

    1998-09-17

    Antigen receptors on T and B cells function to transduce signals leading to a variety of biologic responses minimally including antigen receptor editing, apoptotic death, developmental progression, cell activation, proliferation and survival. The response to antigen depends upon antigen affinity and valence, involvement of coreceptors in signaling and differentiative stage of the responding cell. The requirement that these receptors integrate signals that drive an array of responses may explain their evolved structural complexity. Antigen receptors are composed of multiple subunits compartmentalized to provide antigen recognition and signal transduction function. In lieu of on-board enzymatic activity these receptors rely on associated Protein Tyrosine Kinases (PTKs) for their signaling function. By aggregating the receptors, and hence their appended PTKs, antigens induce PTK transphosphorylation, activating them to phosphorylate the receptor within conserved motifs termed Immunoreceptor Tyrosine-based Activation Motifs (ITAMs) found in transducer subunits. The tyrosyl phosphorylated ITAMs then interact with Src Homology 2 (SH2) domains within the PTKs leading to their further activation. As receptor phosphorylation is amplified, other effectors, such as Shc, dock by virtue of SH2 binding, and serve, in-turn, as substrates for these PTKs. This sequence of events not only provides a signal amplification mechanism by combining multiple consecutive steps with positive feedback, but also allows for signal diversification by differential recruitment of effectors that provide access to distinct parallel downstream signaling pathways. The subject of antigen receptor signaling has been recently reviewed in depth (DeFranco, 1997; Kurosaki, 1997). Here we discuss the biochemical basis of antigen receptor signal transduction, using the B cell receptor (BCR) as a paradigm, with specific emphasis on the involved PTKs. We review several specific mechanisms by which responses

  16. Human Memory B Cells in Healthy Gingiva, Gingivitis, and Periodontitis.

    Science.gov (United States)

    Mahanonda, Rangsini; Champaiboon, Chantrakorn; Subbalekha, Keskanya; Sa-Ard-Iam, Noppadol; Rattanathammatada, Warattaya; Thawanaphong, Saranya; Rerkyen, Pimprapa; Yoshimura, Fuminobu; Nagano, Keiji; Lang, Niklaus P; Pichyangkul, Sathit

    2016-08-01

    The presence of inflammatory infiltrates with B cells, specifically plasma cells, is the hallmark of periodontitis lesions. The composition of these infiltrates in various stages of homeostasis and disease development is not well documented. Human tissue biopsies from sites with gingival health (n = 29), gingivitis (n = 8), and periodontitis (n = 21) as well as gingival tissue after treated periodontitis (n = 6) were obtained and analyzed for their composition of B cell subsets. Ag specificity, Ig secretion, and expression of receptor activator of NF-κB ligand and granzyme B were performed. Although most of the B cell subsets in healthy gingiva and gingivitis tissues were CD19(+)CD27(+)CD38(-) memory B cells, the major B cell component in periodontitis was CD19(+)CD27(+)CD38(+)CD138(+)HLA-DR(low) plasma cells, not plasmablasts. Plasma cell aggregates were observed at the base of the periodontal pocket and scattered throughout the gingiva, especially apically toward the advancing front of the lesion. High expression of CXCL12, a proliferation-inducing ligand, B cell-activating factor, IL-10, IL-6, and IL-21 molecules involved in local B cell responses was detected in both gingivitis and periodontitis tissues. Periodontitis tissue plasma cells mainly secreted IgG specific to periodontal pathogens and also expressed receptor activator of NF-κB ligand, a bone resorption cytokine. Memory B cells resided in the connective tissue subjacent to the junctional epithelium in healthy gingiva. This suggested a role of memory B cells in maintaining periodontal homeostasis. PMID:27335500

  17. Intravenously delivered glucocorticoid liposomes inhibit osteoclast activity and bone erosion in murine antigen-induced arthritis

    NARCIS (Netherlands)

    Hofkens, Wouter; Grevers, Lilyanne C.; Walgreen, Birgitte; de Vries, Teun J.; Leenen, Pieter J. M.; Everts, Vincent; Storm, Gert; van den Berg, Wim B.; van Lent, Peter L.

    2011-01-01

    The objective of this study was to determine the effect of systemic delivery of prednisolone phosphate (PLP) encapsulated within long circulating 'stealth' liposomes on bone erosion and osteoclast activity during experimental antigen-induced arthritis (AIA). Liposomal PLP strongly suppressed knee jo

  18. Enhancement of human natural cytotoxicity by Plasmodium falciparum antigen activated lymphocytes

    DEFF Research Database (Denmark)

    Theander, T G; Pedersen, B K; Bygbjerg, I C;

    1987-01-01

    Mononuclear cells (MNC) isolated from malaria immune donors and from donors never exposed to malaria were stimulated in vitro with soluble purified Plasmodium falciparum antigens (SPag) or PPD. After 7 days of culture the proliferative response and the cytotoxic activity against the natural killer...

  19. Inhibition of Ly-6A antigen expression prevents T cell activation

    OpenAIRE

    1990-01-01

    Antisense oligonucleotides complementary to the 5' end of the mRNA encoding the Ly-6A protein were used to block the expression of that protein. Using this approach we could inhibit the expression of Ly-6A by 60-80% in antigen-primed lymph node (LN) T cells as well as in the D10 T cell clone. Inhibition of Ly-6 expression resulted in the inability to restimulate in vitro, antigen-primed T cells. It also blocked the activation of normal spleen cells by Con A, monoclonal antibody (mAb) to CD3, ...

  20. Clear cell variant of diffuse large B-cell lymphoma: a case report

    Directory of Open Access Journals (Sweden)

    Sahatciu-Meka Vjollca

    2011-05-01

    Full Text Available Abstract Introduction Diffuse large B-cell lymphoma is a diffuse proliferation of large neoplastic B lymphoid cells with a nuclear size equal to or exceeding the normal macrophage nuclei. We report a case of a clear cell variant of diffuse large B-cell lymphoma involving a lymph node in the neck, which was clinically suspected of being metastatic carcinoma. Case presentation A 39-year-old Caucasian ethnic Albanian man from Kosovo presented with a rapidly enlarging lymph node in his neck, but he also disclosed B symptoms and fatigue. A cytological aspirate of the lymph node revealed pleomorphic features. Our patient underwent a cervical lymph node biopsy (large excision. The mass was homogeneously fish-flesh, pale white tissue replacing almost the whole structure of the lymph node. The lymph node biopsy showed a partial alveolar growth pattern, which raised clinical suspicion that it was an epithelial neoplasm. With regard to morphological and phenotypic features, we discovered large nodules in diffuse areas, comprising large cells with slightly irregular nuclei and clear cytoplasm admixed with a few mononuclear cells. In these areas, there was high mitotic activity, and in some areas there were macrophages with tangible bodies. Staining for cytokeratins was negative. These areas had the following phenotypes: cluster designation marker 20 (CD20 positive, B-cell lymphoma (Bcl-2-positive, Bcl-6-, CD5-, CD3-, CD21+ (in alveolar patterns, prostate-specific antigen-negative, human melanoma black marker 45-negative, melanoma marker-negative, cytokeratin-7-negative and multiple myeloma marker 1-positive in about 30% of cells, and exhibited a high proliferation index marker (Ki-67, 80%. Conclusion According to the immunohistochemical findings, we concluded that this patient has a clear cell variant of diffuse large B-cell lymphoma of activated cell type, post-germinal center cell origin. Our patient is undergoing R-CHOP chemotherapy treatment.

  1. Modulation of B-cell endoplasmic reticulum calcium homeostasis by Epstein-Barr virus Latent Membrane Protein-1

    Directory of Open Access Journals (Sweden)

    Joab Irène

    2009-08-01

    Full Text Available Abstract Background Calcium signaling plays an important role in B lymphocyte survival and activation, and is critically dependent on the inositol-1,4,5-tris-phosphate-induced release of calcium stored in the endoplasmic reticulum (ER. Calcium is accumulated in the ER by Sarco/Endoplasmic Reticulum Calcium ATPases (SERCA enzymes, and therefore these enzymes play an important role in ER calcium homeostasis and in the control of B of cell activation. Because Epstein-Barr virus (EBV can immortalize B cells and contributes to lymphomagenesis, in this work the effects of the virus on SERCA-type calcium pump expression and calcium accumulation in the endoplasmic reticulum of B cells was investigated. Results Two Sarco-Endoplasmic Reticulum Calcium transport ATPase isoforms, the low Ca2+-affinity SERCA3, and the high Ca2+-affinity SERCA2 enzymes are simultaneously expressed in B cells. Latency type III infection of Burkitt's lymphoma cell lines with immortalization-competent virus expressing the full set of latency genes selectively decreased the expression of SERCA3 protein, whereas infection with immortalization-deficient virus that does not express the EBNA2 or LMP-1 viral genes was without effect. Down-modulation of SERCA3 expression could be observed upon LMP-1, but not EBNA2 expression in cells carrying inducible transgenes, and LMP-1 expression was associated with enhanced resting cytosolic calcium levels and increased calcium storage in the endoplasmic reticulum. Similarly to virus-induced B cell immortalisation, SERCA3 expression was also decreased in normal B cells undergoing activation and blastic transformation in germinal centers of lymph node follicles. Conclusion The data presented in this work indicate that EBV-induced immortalization leads to the remodelling of ER calcium homeostasis of B cells by LMP-1 that copies a previously unknown normal phenomenon taking place during antigen driven B cell activation. The functional remodelling of

  2. Higher numbers of memory B-cells and Th2-cytokine skewing in high responders to hepatitis B vaccination.

    Science.gov (United States)

    Doedée, A M C M; Kannegieter, N; Öztürk, K; van Loveren, H; Janssen, R; Buisman, A M

    2016-04-27

    In the present study, differences in hepatitis B surface antigen (HBsAg)-specific memory B-cell responses between low and high responders to hepatitis B vaccine (HepB), based on levels of antibodies against HBsAg (anti-HBs), were determined. In addition, HBsAg specific T-cell responses between high (anti-HBs level >20,000IU/L) and low (anti-HBs level vaccination. In advance, an Enzyme-linked Immunosorbent Spot (ELISpot) Assay was optimized for the determination of HBsAg-specific B-cell responses. The number of HBsAg-specific B-cells was significantly higher (pvaccination with HepB. In addition, the plasma IgG levels and numbers of HBsAg-specific B-cells were significantly correlated (RS=0.66, pvaccination with HepB. However, a significant correlation (RS=0.6975, p=0.007) between the IL-13 levels and the plasma IgG levels post-booster was found. Subsequently, the IL-13 level in the high-responder group post-booster was significantly higher compared to the low-responder group. Since activation of the B-cell response after vaccination is induced by Th2 cells and IL-13 is produced by these cells, we conclude that the difference in HBsAg-specific Th2 cells is involved in determining the differences in anti-HBs level and memory B-cell numbers between low and high responders. PMID:26721327

  3. Tumorigenic activity of Merkel cell polyomavirus T antigens expressed in the stratified epithelium of mice

    Science.gov (United States)

    Spurgeon, Megan E.; Cheng, Jingwei; Bronson, Roderick T.; Lambert, Paul F.; DeCaprio, James A.

    2015-01-01

    Merkel cell polyomavirus (MCPyV) is frequently associated with Merkel cell carcinoma (MCC), a highly aggressive neuroendocrine skin cancer. Most MCC tumors contain integrated copies of the viral genome with persistent expression of the MCPyV large T (LT) and small T (ST) antigen. MCPyV isolated from MCC typically contain wild type ST but truncated forms of LT that retain the N-terminus but delete the C-terminus and render LT incapable of supporting virus replication. To determine the oncogenic activity of MCC tumor-derived T antigens in vivo, a conditional, tissue-specific mouse model was developed. Keratin 14-mediated Cre recombinase expression induced expression of MCPyV T antigens in stratified squamous epithelial cells and Merkel cells of the skin epidermis. Mice expressing MCPyV T antigens developed hyperplasia, hyperkeratosis, and acanthosis of the skin with additional abnormalities in whisker pads, footpads and eyes. Nearly half of the mice also developed cutaneous papillomas. Evidence for neoplastic progression within stratified epithelia included increased cellular proliferation, unscheduled DNA synthesis, increased E2F-responsive genes levels, disrupted differentiation, and presence of a DNA damage response. These results indicate that MCPyV T antigens are tumorigenic in vivo, consistent with their suspected etiological role in human cancer. PMID:25596282

  4. Tumorigenic activity of merkel cell polyomavirus T antigens expressed in the stratified epithelium of mice.

    Science.gov (United States)

    Spurgeon, Megan E; Cheng, Jingwei; Bronson, Roderick T; Lambert, Paul F; DeCaprio, James A

    2015-03-15

    Merkel cell polyomavirus (MCPyV) is frequently associated with Merkel cell carcinoma (MCC), a highly aggressive neuroendocrine skin cancer. Most MCC tumors contain integrated copies of the viral genome with persistent expression of the MCPyV large T (LT) and small T (ST) antigen. MCPyV isolated from MCC typically contains wild-type ST but truncated forms of LT that retain the N-terminus but delete the C-terminus and render LT incapable of supporting virus replication. To determine the oncogenic activity of MCC tumor-derived T antigens in vivo, a conditional, tissue-specific mouse model was developed. Keratin 14-mediated Cre recombinase expression induced expression of MCPyV T antigens in stratified squamous epithelial cells and Merkel cells of the skin epidermis. Mice expressing MCPyV T antigens developed hyperplasia, hyperkeratosis, and acanthosis of the skin with additional abnormalities in whisker pads, footpads, and eyes. Nearly half of the mice also developed cutaneous papillomas. Evidence for neoplastic progression within stratified epithelia included increased cellular proliferation, unscheduled DNA synthesis, increased E2F-responsive genes levels, disrupted differentiation, and presence of a DNA damage response. These results indicate that MCPyV T antigens are tumorigenic in vivo, consistent with their suspected etiologic role in human cancer. PMID:25596282

  5. Attenuated recombinant vaccinia virus expressing oncofetal antigen (tumor-associated antigen) 5T4 induces active therapy of established tumors.

    Science.gov (United States)

    Mulryan, Kate; Ryan, Matthew G; Myers, Kevin A; Shaw, David; Wang, Who; Kingsman, Susan M; Stern, Peter L; Carroll, Miles W

    2002-10-01

    The human oncofetal antigen 5T4 (h5T4) is a transmembrane glycoprotein overexpressed by a wide spectrum of cancers, including colorectal, ovarian, and gastric, but with a limited normal tissue expression. Such properties make 5T4 an excellent putative target for cancer immunotherapy. The murine homologue of 5T4 (m5T4) has been cloned and characterized, which allows for the evaluation of immune intervention strategies in "self-antigen" in vivo tumor models. We have constructed recombinant vaccinia viruses based on the highly attenuated and modified vaccinia virus ankara (MVA strain), expressing h5T4 (MVA-h5T4), m5T4 (MVA-m5T4), and Escherichia coli LacZ (MVA-LacZ). Immunization of BALB/c and C57BL/6 mice with MVA-h5T4 and MVA-m5T4 constructs induced antibody responses to human and mouse 5T4, respectively. C57BL/6 and BALB/c mice vaccinated with MVA-h5T4 were challenged with syngeneic tumor line transfectants, B16 melanoma, and CT26 colorectal cells that express h5T4. MVA-h5T4-vaccinated mice showed significant tumor retardation compared with mice vaccinated with MVA-LacZ or PBS. In active treatment studies, inoculation with MVA-h5T4 was able to treat established CT26-h5T4 lung tumor and to a lesser extent B16.h5T4 s.c. tumors. Additionally, when C57BL/6 mice vaccinated with MVA-m5T4 were challenged with B16 cells expressing m5T4, resulting growth of the tumors was significantly retarded compared with control animals. Furthermore, mice vaccinated with MVA-m5T4 showed no signs of autoimmune toxicity. These data support the use of MVA-5T4 for tumor immunotherapy. PMID:12481437

  6. Crystal structure of a Staphylococcus aureus protein A domain complexed with the Fab fragment of a human IgM antibody: Structural basis for recognition of B-cell receptors and superantigen activity

    OpenAIRE

    Graille, Marc; Stura, Enrico A; Corper, Adam L.; Sutton, Brian J.; Michael J. Taussig; Charbonnier, Jean-Baptiste; Silverman, Gregg J.

    2000-01-01

    Staphylococcus aureus produces a virulence factor, protein A (SpA), that contains five homologous Ig-binding domains. The interactions of SpA with the Fab region of membrane-anchored Igs can stimulate a large fraction of B cells, contributing to lymphocyte clonal selection. To understand the molecular basis for this activity, we have solved the crystal structure of the complex between domain D of SpA and the Fab fragment of a human IgM antibody to 2.7-Å resolution. In the complex, helices II ...

  7. YY1 Is Required for Germinal Center B Cell Development

    Science.gov (United States)

    Vuyyuru, Raja; Jha, Vibha; Hodewadekar, Suchita; Manser, Tim; Atchison, Michael L.

    2016-01-01

    YY1 has been implicated as a master regulator of germinal center B cell development as YY1 binding sites are frequently present in promoters of germinal center-expressed genes. YY1 is known to be important for other stages of B cell development including the pro-B and pre-B cells stages. To determine if YY1 plays a critical role in germinal center development, we evaluated YY1 expression during B cell development, and used a YY1 conditional knock-out approach for deletion of YY1 in germinal center B cells (CRE driven by the immunoglobulin heavy chain γ1 switch region promoter; γ1-CRE). We found that YY1 is most highly expressed in germinal center B cells and is increased 3 fold in splenic B cells activated by treatment with anti-IgM and anti-CD40. In addition, deletion of the yy1 gene by action of γ1-CRE recombinase resulted in significant loss of GC cells in both un-immunized and immunized contexts with corresponding loss of serum IgG1. Our results show a crucial role for YY1 in the germinal center reaction. PMID:27167731

  8. A multi-scale model for correlation in B cell VDJ usage of zebrafish

    International Nuclear Information System (INIS)

    The zebrafish (Danio rerio) is one of the model animals used for the study of immunology because the dynamics in the adaptive immune system of zebrafish are similar to that in higher animals. In this work, we built a multi-scale model to simulate the dynamics of B cells in the primary and secondary immune responses of zebrafish. We use this model to explain the reported correlation between VDJ usage of B cell repertoires in individual zebrafish. We use a delay ordinary differential equation (ODE) system to model the immune responses in the 6-month lifespan of a zebrafish. This mean field theory gives the number of high-affinity B cells as a function of time during an infection. The sequences of those B cells are then taken from a distribution calculated by a 'microscopic' random energy model. This generalized NK model shows that mature B cells specific to one antigen largely possess a single VDJ recombination. The model allows first-principle calculation of the probability, p, that two zebrafish responding to the same antigen will select the same VDJ recombination. This probability p increases with the B cell population size and the B cell selection intensity. The probability p decreases with the B cell hypermutation rate. The multi-scale model predicts correlations in the immune system of the zebrafish that are highly similar to that from experiment

  9. A multi-scale model for correlation in B cell VDJ usage of zebrafish

    Science.gov (United States)

    Pan, Keyao; Deem, Michael W.

    2011-10-01

    The zebrafish (Danio rerio) is one of the model animals used for the study of immunology because the dynamics in the adaptive immune system of zebrafish are similar to that in higher animals. In this work, we built a multi-scale model to simulate the dynamics of B cells in the primary and secondary immune responses of zebrafish. We use this model to explain the reported correlation between VDJ usage of B cell repertoires in individual zebrafish. We use a delay ordinary differential equation (ODE) system to model the immune responses in the 6-month lifespan of a zebrafish. This mean field theory gives the number of high-affinity B cells as a function of time during an infection. The sequences of those B cells are then taken from a distribution calculated by a 'microscopic' random energy model. This generalized NK model shows that mature B cells specific to one antigen largely possess a single VDJ recombination. The model allows first-principle calculation of the probability, p, that two zebrafish responding to the same antigen will select the same VDJ recombination. This probability p increases with the B cell population size and the B cell selection intensity. The probability p decreases with the B cell hypermutation rate. The multi-scale model predicts correlations in the immune system of the zebrafish that are highly similar to that from experiment.

  10. Identification of Functional Human Splenic Memory B Cells by Expression of CD148 and CD27

    OpenAIRE

    Tangye, Stuart G.; Liu, Yong-Jun; Aversa, Gregorio; Phillips, Joseph H.; Vries, Jan E. de

    1998-01-01

    Memory B cells isolated from human tonsils are characterized by an activated cell surface phenotype, localization to mucosal epithelium, expression of somatically mutated immunoglobulin (Ig) variable (V) region genes, and a preferential differentiation into plasma cells in vitro. In spleens of both humans and rodents, a subset of memory B cells is believed to reside in the marginal zone of the white pulp. Similar to tonsil-derived memory B cells, splenic marginal zone B cells can be distingui...

  11. Immature myeloid Gr-1+ CD11b+ cells from lipopolysaccharide-immunosuppressed mice acquire inhibitory activity in the bone marrow and migrate to lymph nodes to exert their suppressive function.

    Science.gov (United States)

    Landoni, Veronica I; Martire-Greco, Daiana; Rodriguez-Rodrigues, Nahuel; Chiarella, Paula; Schierloh, Pablo; Isturiz, Martin A; Fernández, Gabriela C

    2016-02-01

    Secondary infections due to post-sepsis immunosuppression are a major cause of death in patients with sepsis. Repetitive inoculation of increasing doses of lipopolysaccharide (LPS) into mice mimics the immunosuppression associated with sepsis. Myeloid-derived suppressor cells (MDSCs, Gr-1(+) CD11b(+)) are considered a major component of the immunosuppressive network, interfering with T-cell responses in many pathological conditions. We used LPS-immunosuppressed (IS) mice to address whether MDSCs acquired their suppressive ability in the bone marrow (BM) and whether they could migrate to lymph nodes (LNs) to exert their suppressive function. Our results showed that Gr-1(+) CD11b(+) cells of IS mice already had the potential to inhibit T-cell proliferation in the BM. Moreover, soluble factors present in the BM from IS mice were responsible for inducing this inhibitory ability in control BM cells. In addition, migration of Gr-1(+) CD11b(+) to LNs in vivo was maximal when cells obtained from the BM of IS mice were inoculated into an IS context. In this regard, we found chemoattractant activity in cell-free LN extracts (LNEs) from IS mice and an increased expression of the LN-homing chemokine receptor C-C chemokine receptor type 7 (CCR7) in IS BM Gr-1(+) CD11b(+) cells. These results indicate that Gr-1(+) CD11b(+) cells found in BM from IS mice acquire their suppressive activity in the same niche where they are generated, and migrate to LNs to exert their inhibitory role. A better understanding of MDSC generation and/or regulation of factors able to induce their inhibitory function may provide new and more effective tools for the treatment of sepsis-associated immunosuppression. PMID:26582821

  12. Molecular signals in antigen presentation. II. Activation of cytolytic cells in vitro after ultraviolet radiation or combined gamma and ultraviolet radiation treatment of antigen-presenting cells

    International Nuclear Information System (INIS)

    Murine low-density spleen cells have potent antigen-presenting ability in a hapten-specific cytolytic T lymphocyte (CTL) system using the hapten azobenzenearsonate (ABA). Exposure of these cells to 0.33 KJ/m2 of ultraviolet radiation (UVR) after coupling to hapten results in markedly inhibited antigen-presenting function that can be substantially corrected or bypassed by interleukin 1 (IL 1). These results have been interpreted to reflect an inhibition of Lyt-1+ T cell activation by UVR-treated APC. Treatment of these cells sequentially with 1500 rad of γ-radiation (GR) prior to hapten coupling, followed by 0.33 KJ/m2 of UVR radiation after coupling, results in an antigen-resenting defect only minimally improved by IL 1. However, partially purified interleukin 2 (IL 2) can completely bypass or correct this defect. Thus, combined Cr and UVR induces a different or more profound defect in APC function when compared to UVR alone. However, these cells do provide a signal(s) other than hapten necessary for CTL activation because ABA-coupled high density spleen cells do not activate CTL cells, even with the addition of IL 2. Fluorescence-activated cell sorter analysis demonstrates that exposure of these low density spleen cells to GP or UVR results in decreased I-A antigen expression at 24 hr; exposure to both GR and UVR results in a greater decrease in I-A antigen expression at 24 hr than either alone. The addition of nonhapten-coupled low-density APC partially reconstitutes the ability of combined GR/UVR-treated LD-APC to present antigen, and this effect is enhanced by the administration of exogenous IL 1

  13. Immunoglobulin heavy chain variable region gene utilization by B cell hybridomas derived from rheumatoid synovial tissue.

    Science.gov (United States)

    Brown, C M; Longhurst, C; Haynes, G; Plater-Zyberk, C; Malcolm, A; Maini, R N

    1992-08-01

    Rheumatoid arthritis (RA) is a chronic inflammatory disease that primarily affects synovial joints. Activated B lymphocytes and plasma cells are present in the synovial tissue and are thought to contribute to the immunopathology of the rheumatoid joint. To investigate rheumatoid synovial B lymphocytes, we have generated B cell hybridomas from synovial tissue of an RA patient. Here we describe the immunoglobulin VH gene repertoire of eight IgM- and 10 IgG-secreting synovial-derived hybridomas. The VH4 gene family is highly represented (38.5%) in this panel of hybridomas compared with the frequency of VH4 gene expression in circulating B lymphocytes reported previously (19-22%) and with the VH4 gene frequency we observed in a panel of hybridomas derived in the same manner from the spleen and tonsil of normal individuals (19%). The increased frequency of VH4 gene expression was not due to the expansion of a single B cell clone in vivo as none of these hybridomas was clonally related. Two synovial-derived hybridomas secreted autoantibodies; one (VH3+) secreted an IgM-rheumatoid factor (RF) and the other (VH4+) secreted IgM with polyreactive binding to cytoskeletal proteins and cardiolipin. The antibodies secreted by the remaining synovial-derived hybridomas were not reactive with the autoantigens tested. The VH gene usage in a proportion (5/17) of synovial-derived hybridomas that expressed CD5 antigen provided preliminary evidence that CD5+ B cells in RA synovium have a similar increase of VH4 gene expression reported for CD5+ B cells from normal individuals and patients with chronic lymphocytic leukaemia. PMID:1379132

  14. HIV-dependent depletion of influenza-specific memory B cells impacts B cell responsiveness to seasonal influenza immunisation.

    Science.gov (United States)

    Wheatley, Adam K; Kristensen, Anne B; Lay, William N; Kent, Stephen J

    2016-01-01

    Infection with HIV drives significant alterations in B cell phenotype and function that can markedly influence antibody responses to immunisation. Anti-retroviral therapy (ART) can partially reverse many aspects of B cell dysregulation, however complete normalisation of vaccine responsiveness is not always observed. Here we examine the effects of underlying HIV infection upon humoral immunity to seasonal influenza vaccines. Serological and memory B cell responses were assessed in 26 HIV+ subjects receiving ART and 30 healthy controls immunised with the 2015 Southern Hemisphere trivalent inactivated influenza vaccine (IIV3). Frequencies and phenotypes of influenza hemagglutinin (HA)-specific B cells were assessed by flow cytometry using recombinant HA probes. Serum antibody was measured using hemagglutination inhibition assays. Serological responses to IIV3 were comparable between HIV+ and HIV- subjects. Likewise, the activation and expansion of memory B cell populations specific for vaccine-component influenza strains was observed in both cohorts, however peak frequencies were diminished in HIV+ subjects compared to uninfected controls. Lower circulating frequencies of memory B cells recognising vaccine-component and historical influenza strains were observed in HIV+ subjects at baseline, that were generally restored to levels comparable with HIV- controls post-vaccination. HIV infection is therefore associated with depletion of selected HA-specific memory B cell pools. PMID:27220898

  15. HIV-dependent depletion of influenza-specific memory B cells impacts B cell responsiveness to seasonal influenza immunisation

    Science.gov (United States)

    Wheatley, Adam K.; Kristensen, Anne B.; Lay, William N.; Kent, Stephen J.

    2016-01-01

    Infection with HIV drives significant alterations in B cell phenotype and function that can markedly influence antibody responses to immunisation. Anti-retroviral therapy (ART) can partially reverse many aspects of B cell dysregulation, however complete normalisation of vaccine responsiveness is not always observed. Here we examine the effects of underlying HIV infection upon humoral immunity to seasonal influenza vaccines. Serological and memory B cell responses were assessed in 26 HIV+ subjects receiving ART and 30 healthy controls immunised with the 2015 Southern Hemisphere trivalent inactivated influenza vaccine (IIV3). Frequencies and phenotypes of influenza hemagglutinin (HA)-specific B cells were assessed by flow cytometry using recombinant HA probes. Serum antibody was measured using hemagglutination inhibition assays. Serological responses to IIV3 were comparable between HIV+ and HIV− subjects. Likewise, the activation and expansion of memory B cell populations specific for vaccine-component influenza strains was observed in both cohorts, however peak frequencies were diminished in HIV+ subjects compared to uninfected controls. Lower circulating frequencies of memory B cells recognising vaccine-component and historical influenza strains were observed in HIV+ subjects at baseline, that were generally restored to levels comparable with HIV− controls post-vaccination. HIV infection is therefore associated with depletion of selected HA-specific memory B cell pools. PMID:27220898

  16. Antigen Presentation and T-Cell Activation Are Critical for RBP4-Induced Insulin Resistance.

    Science.gov (United States)

    Moraes-Vieira, Pedro M; Castoldi, Angela; Aryal, Pratik; Wellenstein, Kerry; Peroni, Odile D; Kahn, Barbara B

    2016-05-01

    Adipose tissue (AT) inflammation contributes to impaired insulin action, which is a major cause of type 2 diabetes. RBP4 is an adipocyte- and liver-derived protein with an important role in insulin resistance, metabolic syndrome, and AT inflammation. RBP4 elevation causes AT inflammation by activating innate immunity, which elicits an adaptive immune response. RBP4-overexpressing mice (RBP4-Ox) are insulin resistant and glucose intolerant and have increased AT macrophages and T-helper 1 cells. We show that high-fat diet-fed RBP4(-/-) mice have reduced AT inflammation and improved insulin sensitivity versus wild type. We also elucidate the mechanism for RBP4-induced macrophage antigen presentation and subsequent T-cell activation. In RBP4-Ox, AT macrophages display enhanced c-Jun N-terminal kinase, extracellular signal-related kinase, and p38 phosphorylation. Inhibition of these pathways and of NF-κB reduces activation of macrophages and CD4 T cells. MyD88 is an adaptor protein involved in proinflammatory signaling. In macrophages from MyD88(-/-) mice, RBP4 fails to stimulate secretion of tumor necrosis factor, IL-12, and IL-6 and CD4 T-cell activation. In vivo blockade of antigen presentation by treating RBP4-Ox mice with CTLA4-Ig, which blocks costimulation of T cells, is sufficient to reduce AT inflammation and improve insulin resistance. Thus, MyD88 and downstream mitogen-activated protein kinase and NF-κB pathways are necessary for RBP4-induced macrophage antigen presentation and subsequent T-cell activation. Also, blocking antigen presentation with CTLA4-Ig improves RBP4-induced insulin resistance and macrophage-induced T-cell activation. PMID:26936962

  17. Anti-Ia antibody in the sera of normal subjects after in vivo antigenic stimulation

    OpenAIRE

    1982-01-01

    We showed that sera from normal subjects after antigenic challenge with intradermal PPD or Candida antigens or with subcutaneous tetanus vaccine contain a factor that blocks the binding of mouse monoclonal anti-Ia antibody to Ia-positive T cells or to B35 M cells, an Ia- positive human B cell line. The blocking activity appears 48 to 72 h after antigenic challenge and is gone by day 7. The appearance of the anti-Ia blocking activity coincided with a drop in the percentage of Ia- positive T ce...

  18. Complement-dependent transport of antigen into B cell follicles

    DEFF Research Database (Denmark)

    Gonzalez, Santiago F.; Lukacs-Kornek, Veronika; Kuligowski, Michael P.;

    2010-01-01

    Since the original proposal by Fearon and Locksley (Fearon and Locksley. 1996. Science 272: 50-53) that the complement system linked innate and adaptive immunity, there has been a rapid expansion of studies on this topic. With the advance of intravital imaging, a number of recent papers revealed ...... opsonization of influenza and uptake by macrophages, and the capture of virus by dendritic cells residing in the medullary compartment of peripheral lymph nodes....

  19. New helping friends for B cells.

    OpenAIRE

    Cerutti, Andrea; Puga, Irene; Cols, Montserrat

    2012-01-01

    Over the past decade, a growing recognition of the importance of neutralizing antibodies in host defense combined with the success of B-cell depletion therapies in treating autoimmune disorders has led to an increased focus on better understanding the pathways underpinning B-cell antibody production. In general, B cells require cognate interaction with T helper cells in the germinal center of lymphoid follicles to generate protective antibodies. However, recent evidence shows that B cells rec...

  20. Expansion of IgG+ B-cells during mitogen stimulation for memory B-cell ELISpot analysis is influenced by size and composition of the B-cell pool

    NARCIS (Netherlands)

    Scholzen, A.; Nahrendorf, W.; Langhorne, J.; Sauerwein, R.W.

    2014-01-01

    The memory B-cell (MBC) ELISpot assay is the main technique used to measure antigen-specific MBCs as a readout of humoral immune memory. This assay relies on the ability of MBCs to differentiate into antibody-secreting cells (ASC) upon polyclonal stimulation. The total number of IgG+ ASCs generated

  1. B-cell survival factors in autoimmune rheumatic disorders

    Science.gov (United States)

    Morais, Sandra A.; Vilas-Boas, Andreia

    2015-01-01

    Autoimmune rheumatic disorders have complex etiopathogenetic mechanisms in which B cells play a central role. The importance of factors stimulating B cells, notably the B-cell activating factor (BAFF) and A proliferation inducing ligand (APRIL) axis is now recognized. BAFF and APRIL are cytokines essential for B-cell proliferation and survival from the immature stages to the development of plasma cells. Their levels are increased in some subsets of patients with autoimmune disorders. Several recent biologic drugs have been developed to block this axis, namely belimumab [already licensed for systemic lupus erythematosus (SLE) treatment], tabalumab, atacicept and blisibimod. Many clinical trials to evaluate the safety and efficacy of these drugs in several autoimmune disorders are ongoing, or have been completed recently. This review updates the information on the use of biologic agents blocking BAFF/APRIL for patients with SLE, rheumatoid arthritis, Sjögren’s syndrome and myositis. PMID:26288664

  2. The anti-lymphoma activity of antiviral therapy in HCV-associated B-cell non-Hodgkin lymphomas: a meta-analysis.

    Science.gov (United States)

    Peveling-Oberhag, J; Arcaini, L; Bankov, K; Zeuzem, S; Herrmann, E

    2016-07-01

    Many epidemiological studies provide solid evidence for an association of chronic hepatitis C virus (HCV) infection with B-cell non-Hodgkin's lymphoma (B-NHL). However, the most convincing evidence for a causal relationship between HCV infection and lymphoma development is the observation of B-NHL regression after HCV eradication by antiviral therapy (AVT). We conducted a literature search to identify studies that included patients with HCV-associated B-NHL (HCV-NHL) who received AVT, with the intention to treat lymphoma and viral disease at the same time. The primary end point was the correlation of sustained virological response (SVR) under AVT with lymphoma response. Secondary end points were overall lymphoma response rates and HCV-NHL response in correlation with lymphoma subtypes. We included 20 studies that evaluated the efficacy of AVT in HCV-NHL (n = 254 patients). Overall lymphoma response rate through AVT was 73% [95%>confidence interval, (CI) 67-78%]. Throughout studies there was a strong association between SVR and lymphoma response (83% response rate, 95%>CI, 76-88%) compared to a failure in achieving SVR (53% response rate, 95%>CI, 39-67%, P = 0.0002). There was a trend towards favourable response for AVT in HCV-associated marginal zone lymphomas (response rate 81%, 95%>CI, 74-87%) compared to nonmarginal zone origin (response rate 71%, 95%>CI, 61-79%, P = 0.07). In conclusion, in the current meta-analysis, the overall response rate of HCV-NHL under AVT justifies the recommendation for AVT as first-line treatment in patients who do not need immediate conventional treatment. The strong correlation of SVR and lymphoma regression supports the hypothesis of a causal relationship of HCV and lymphomagenesis. PMID:26924533

  3. Exposure to ozone enhances antigen-presenting activity concentration dependently in rats

    International Nuclear Information System (INIS)

    The effect of ozone (O3) on the symptoms of allergic asthma and the mechanisms underlying have not yet been fully elucidated. Antigen presentation is one of the factors contributing to the allergic reaction. Therefore, we investigated the effects of repeated exposure to O3 on antigen-presenting (AP) activity, on the expression of cell-surface molecules associated with antigen presentation (Ia, B7.1, B7.2 and CD11b/c) in bronchoalveolar lavage cells (BAL cells), and on allergic asthma-like symptoms. Rats were exposed to 0.3, 0.56, 1 ppm O3 or filtered air for a 3-day period every 2 weeks, this was replicated three times. AP activity was assessed by measuring antigen-specific T-cell proliferation; and the expression of cell-surface molecules, by flow cytometry. Rats were also made to inhale aerosolized 1% ovalbumin (OVA) or saline for 10 min post-exposure to O3, and allergic asthma-like symptoms were measured by determining the increase in enhanced pause (Penh), which correlates well with lung resistance. O3 increased both AP activity and expression of Ia and costimulatory molecules in BAL cells concentration dependently. It also increased lung resistance, and the increase in lung resistance after O3 exposure was significantly higher in the OVA-inhaled group than in the saline-inhaled group. The present results show that O3 increased AP activity concentration dependently and suggest that O3 might aggravate allergy symptoms by enhancing AP activity

  4. CEACAM1 mediates B cell aggregation in central nervous system autoimmunity.

    Science.gov (United States)

    Rovituso, Damiano M; Scheffler, Laura; Wunsch, Marie; Kleinschnitz, Christoph; Dörck, Sebastian; Ulzheimer, Jochen; Bayas, Antonios; Steinman, Lawrence; Ergün, Süleyman; Kuerten, Stefanie

    2016-01-01

    B cell aggregates in the central nervous system (CNS) have been associated with rapid disease progression in patients with multiple sclerosis (MS). Here we demonstrate a key role of carcinoembryogenic antigen-related cell adhesion molecule1 (CEACAM1) in B cell aggregate formation in MS patients and a B cell-dependent mouse model of MS. CEACAM1 expression was increased on peripheral blood B cells and CEACAM1(+) B cells were present in brain infiltrates of MS patients. Administration of the anti-CEACAM1 antibody T84.1 was efficient in blocking aggregation of B cells derived from MS patients. Along these lines, application of the monoclonal anti-CEACAM1 antibody mCC1 was able to inhibit CNS B cell aggregate formation and significantly attenuated established MS-like disease in mice in the absence of any adverse effects. CEACAM1 was co-expressed with the regulator molecule T cell immunoglobulin and mucin domain -3 (TIM-3) on B cells, a novel molecule that has recently been described to induce anergy in T cells. Interestingly, elevated coexpression on B cells coincided with an autoreactive T helper cell phenotype in MS patients. Overall, these data identify CEACAM1 as a clinically highly interesting target in MS pathogenesis and open new therapeutic avenues for the treatment of the disease. PMID:27435215

  5. Isolation of human monoclonal antibodies from peripheral blood B cells.

    Science.gov (United States)

    Huang, Jinghe; Doria-Rose, Nicole A; Longo, Nancy S; Laub, Leo; Lin, Chien-Li; Turk, Ellen; Kang, Byong H; Migueles, Stephen A; Bailer, Robert T; Mascola, John R; Connors, Mark

    2013-10-01

    Isolation of monoclonal antibodies is an important technique for understanding the specificities and characteristics of antibodies that underlie the humoral immune response to a given antigen. Here we describe a technique for isolating monoclonal antibodies from human peripheral blood mononuclear cells. The protocol includes strategies for the isolation of switch-memory B cells from peripheral blood, the culture of B cells, the removal of the supernatant for screening and the lysis of B cells in preparation for immunoglobulin heavy-chain and light-chain amplification and cloning. We have observed that the addition of cytokines IL-2, IL-21 and irradiated 3T3-msCD40L feeder cells can successfully stimulate switch-memory B cells to produce high concentrations of IgG in the supernatant. The supernatant may then be screened by appropriate assays for binding or for other functions. This protocol can be completed in 2 weeks. It is adaptable to use in other species and enables the efficient isolation of antibodies with a desired functional characteristic without prior knowledge of specificity. PMID:24030440

  6. Preparation of antisera specific for human B cells by immunization of rabbits with immune complexes

    International Nuclear Information System (INIS)

    Three rabbit antisera are described which are specific without absorption (titer 1:100) for separated human B cells, as measured by complement and non-complement fixing assays. The method of production of these sera involved injections of rabbits with precipitin lines formed between 10μ1 of three separate detergent solubilized membrane preparations and 4μ1 aliquots of rabbit antisera to human B cells. In addition to being B cell specific, the three sera block the MLC reaction, inhibit aggregated IgG binding to B cells, and show differential degrees of B cell lysis when tested on a panel of separated B and T cells. These and other properties suggest that the target specificities of the antibodies are the human equivalent of the murine Ia antigens. (author)

  7. Uptake and presentation of myelin basic protein by normal human B cells

    DEFF Research Database (Denmark)

    Brimnes, Marie Klinge; Hansen, Bjarke Endel; Nielsen, Leif Kofoed;

    2014-01-01

    were capable of binding the MS-associated self-antigen myelin basic protein (MBP) and of presenting the immunodominant peptide MBP85-99, as determined by staining with the mAb MK16 recognising the peptide presented by HLA-DR15-positive cells. In the presence of serum, however, the majority of B cells...... proportion of MBP85-99 presenting B cells expressed CD27, and showed increased expression of CD86 compared to non-presenting B cells. MBP-pulsed B cells induced a low frequency of IL-10-producing CD4+ T cells in 3 out of 6 donors, indicating an immunoregulatory role of B cells presenting MBP-derived peptides...

  8. Soluble Expression and Characterization of Biologically Active Bacillus anthracis Protective Antigen in Escherichia coli

    OpenAIRE

    Nagendra Suryanarayana; Vanlalhmuaka,; Bharti Mankere; Monika Verma; Kulanthaivel Thavachelvam; Urmil Tuteja

    2016-01-01

    Bacillus anthracis secretory protein protective antigen (PA) is primary candidate for subunit vaccine against anthrax. Attempts to obtain large quantity of PA from Escherichia coli expression system often result in the formation of insoluble inclusion bodies. Therefore, it is always better to produce recombinant proteins in a soluble form. In the present study, we have obtained biologically active recombinant PA in small scale E. coli shake culture system using three different expression cons...

  9. Antibody-independent control of gamma-herpesvirus latency via B cell induction of anti-viral T cell responses.

    Directory of Open Access Journals (Sweden)

    Kelly B McClellan

    2006-06-01

    Full Text Available B cells can use antibody-dependent mechanisms to control latent viral infections. It is unknown whether this represents the sole function of B cells during chronic viral infection. We report here that hen egg lysozyme (HEL-specific B cells can contribute to the control of murine gamma-herpesvirus 68 (gammaHV68 latency without producing anti-viral antibody. HEL-specific B cells normalized defects in T cell numbers and proliferation observed in B cell-/- mice during the early phase of gammaHV68 latency. HEL-specific B cells also reversed defects in CD8 and CD4 T cell cytokine production observed in B cell-/- mice, generating CD8 and CD4 T cells necessary for control of latency. Furthermore, HEL-specific B cells were able to present virally encoded antigen to CD8 T cells. Therefore, B cells have antibody independent functions, including antigen presentation, that are important for control of gamma-herpesvirus latency. Exploitation of this property of B cells may allow enhanced vaccine responses to chronic virus infection.

  10. Regulatory B cells and tolerance in transplantation: from animal models to human.

    Directory of Open Access Journals (Sweden)

    Melanie eChesneau

    2013-12-01

    Full Text Available Until recently, the role of B cells in transplantation was thought to be restricted to producing antibodies that have been clearly shown to be deleterious in the long term, but, in fact, B cells are also able to produce cytokine and to present antigen. Their role as regulatory cells in various pathological situations has also been highlighted, and their role in transplantation is beginning to emerge in animal, and also in human, models. This review summarizes the different studies in animals and humans that suggest a B-cell regulatory role in the transplant tolerance mechanisms.

  11. Human Lyb-2 homolog CD72 is a marker for progenitor B-cell leukemias.

    Science.gov (United States)

    Schwarting, R; Castello, R; Moldenhauer, G; Pezzutto, A; von Hoegen, I; Ludwig, W D; Parnes, J R; Dörken, B

    1992-11-01

    S-HCL 2 is the prototype antibody of the recently defined CD72 cluster (human Lyb-2). Under nonreducing conditions, S-HCL 2 monoclonal antibody (mAb) precipitates a glycoprotein of 80-86 kDa. Under reducing conditions, a dimer of 43 and 39 kDa, with core proteins of 40 and 36 kDa, is precipitated. CD72 expression in normal and malignant tissues is different from expression of all other previously described human B-cell antigens. In peripheral blood and bone marrow, the antigen appears to be present on all B lymphocytes, with the exception of plasma cells. In tissue, immunohistochemical staining revealed positivity for all known B-cell compartments; however, pulpa macrophages of the spleen and von Kupffer cells exhibited distinct positivity for CD72 also. Among 83 malignant non-Hodgkin's lymphomas examined by immunohistochemistry (alkaline phosphatase anti-alkaline phosphatase technique), all 54 B-cell lymphomas, including precursor B-cell lymphomas, Burkitt's lymphomas, germinal center lymphomas, chronic lymphocytic leukemias, and hairy cell leukemias, were CD72 positive, but no T-cell lymphomas were. Flow cytometry study of more than 80 mainly acute leukemias (52 B-cell leukemias) showed reactivity with S-HCL 2 mAb over the full range of B-cell differentiation. In particular, very early B cells in cytoplasmic Ig (cIg)-negative, CD19-positive pre-pre-B-cell leukemias and hybrid leukemias (mixed myeloid and B-cell type) were consistently positive for CD72 on the cell surface. Therefore, CD72 may become an important marker for progenitor B-cell leukemias. PMID:1384316

  12. Memory B cell antibodies to HIV-1 gp140 cloned from individuals infected with clade A and B viruses.

    Directory of Open Access Journals (Sweden)

    Hugo Mouquet

    Full Text Available Understanding the antibody response to HIV-1 in humans that show broad neutralizing serologic activity is a crucial step in trying to reproduce such responses by vaccination. Investigating antibodies with cross clade reactivity is particularly important as these antibodies may target conserved epitopes on the HIV envelope gp160 protein. To this end we have used a clade B YU-2 gp140 trimeric antigen and single-cell antibody cloning methods to obtain 189 new anti-gp140 antibodies representing 51 independent B cell clones from the IgG memory B cells of 3 patients infected with HIV-1 clade A or B viruses and exhibiting broad neutralizing serologic activity. Our results support previous findings showing a diverse antibody response to HIV gp140 envelope protein, characterized by differentially expanded B-cell clones producing highly hypermutated antibodies with heterogenous gp140-specificity and neutralizing activity. In addition to their high-affinity binding to the HIV spike, the vast majority of the new anti-gp140 antibodies are also polyreactive. Although none of the new antibodies are as broad or potent as VRC01 or PG9, two clonally-related antibodies isolated from a clade A HIV-1 infected donor, directed against the gp120 variable loop 3, rank in the top 5% of the neutralizers identified in our large collection of 185 unique gp140-specific antibodies in terms of breadth and potency.

  13. Clonality Analysis of Immunoglobulin Gene Rearrangement by Next-Generation Sequencing in Endemic Burkitt Lymphoma Suggests Antigen Drive Activation of BCR as Opposed to Sporadic Burkitt Lymphoma

    Science.gov (United States)

    Amato, Teresa; Abate, Francesco; Piccaluga, Pierpaolo; Iacono, Michele; Fallerini, Chiara; Renieri, Alessandra; De Falco, Giulia; Ambrosio, Maria Raffaella; Mourmouras, Vaselious; Ogwang, Martin; Calbi, Valeria; Rabadan, Roul; Hummel, Michael; Pileri, Stefano; Bellan, Cristiana

    2016-01-01

    Objectives: Recent studies using next-generation sequencing (NGS) analysis disclosed the importance of the intrinsic activation of the B-cell receptor (BCR) pathway in the pathogenesis of sporadic Burkitt lymphoma (sBL) due to mutations of TCF3/ID3 genes. Since no definitive data are available on the genetic landscape of endemic Burkitt (eBL), we first assessed the mutation frequency of TCF3/ID3 in eBL compared with sBL and subsequently the somatic hypermutation status of the BCR to answer whether an extrinsic activation of BCR signaling could also be demonstrated in Burkitt lymphoma. Methods: We assessed the mutations of TCF3/ID3 by RNAseq and the BCR status by NGS analysis of the immunoglobulin genes (IGs). Results: We detected mutations of TCF3/ID3 in about 30% of the eBL cases. This rate is significantly lower than that detected in sBL (64%). The NGS analysis of IGs revealed intraclonal diversity, suggesting an active targeted somatic hypermutation process in eBL compared with sBL. Conclusions: These findings support the view that the antigenic pressure plays a key role in the pathogenetic pathways of eBL, which may be partially distinct from those driving sBL development. PMID:26712879

  14. The Analysis of B-Cell Epitopes of Influenza Virus Hemagglutinin.

    Science.gov (United States)

    Shcherbinin, D N; Alekseeva, S V; Shmarov, M M; Smirnov, Yu A; Naroditskiy, B S; Gintsburg, A L

    2016-01-01

    Vaccination has been successfully used to prevent influenza for a long time. Influenza virus hemagglutinin (HA), which induces a humoral immune response in humans and protection against the flu, is the main antigenic component of modern influenza vaccines. However, new seasonal and pandemic influenza virus variants with altered structures of HA occasionally occur. This allows the pathogen to avoid neutralization with antibodies produced in response to previous vaccination. Development of a vaccine with the new variants of HA acting as antigens takes a long time. Therefore, during an epidemic, it is important to have passive immunization agents to prevent and treat influenza, which can be monoclonal or single-domain antibodies with universal specificity (broad-spectrum agents). We considered antibodies to conserved epitopes of influenza virus antigens as universal ones. In this paper, we tried to characterize the main B-cell epitopes of hemagglutinin and analyze our own and literature data on broadly neutralizing antibodies. We conducted a computer analysis of the best known conformational epitopes of influenza virus HAs using materials of different databases. The analysis showed that the core of the HA molecule, whose antibodies demonstrate pronounced heterosubtypic activity, can be used as a target for the search for and development of broad-spectrum antibodies to the influenza virus. PMID:27099781

  15. CD19 CAR-targeted T cells induce long-term remission and B Cell Aplasia in an immunocompetent mouse model of B cell acute lymphoblastic leukemia.

    Directory of Open Access Journals (Sweden)

    Marco L Davila

    Full Text Available Although many adults with B cell acute lymphoblastic leukemia (B-ALL are induced into remission, most will relapse, underscoring the dire need for novel therapies for this disease. We developed murine CD19-specific chimeric antigen receptors (CARs and an immunocompetent mouse model of B-ALL that recapitulates the disease at genetic, cellular, and pathologic levels. Mouse T cells transduced with an all-murine CD3ζ/CD28-based CAR that is equivalent to the one being used in our clinical trials, eradicate B-ALL in mice and mediate long-term B cell aplasias. In this model, we find that increasing conditioning chemotherapy increases tumor eradication, B cell aplasia, and CAR-modified T cell persistence. Quantification of recipient B lineage cells allowed us to estimate an in vivo effector to endogenous target ratio for B cell aplasia maintenance. In mice exhibiting a dramatic B cell reduction we identified a small population of progenitor B cells in the bone marrow that may serve as a reservoir for long-term CAR-modified T cell stimulation. Lastly, we determine that infusion of CD8+ CAR-modified T cells alone is sufficient to maintain long-term B cell eradication. The mouse model we report here should prove valuable for investigating CAR-based and other therapies for adult B-ALL.

  16. The B-cell receptor orchestrates environment-mediated lymphoma survival and drug resistance in B-cell malignancies.

    Science.gov (United States)

    Shain, K H; Tao, J

    2014-08-01

    Specific niches within the lymphoma tumor microenvironment (TME) provide sanctuary for subpopulations of tumor cells through stromal cell-tumor cell interactions. These interactions notably dictate growth, response to therapy and resistance of residual malignant B cells to therapeutic agents. This minimal residual disease (MRD) remains a major challenge in the treatment of B-cell malignancies and contributes to subsequent disease relapse. B-cell receptor (BCR) signaling has emerged as essential mediator of B-cell homing, survival and environment-mediated drug resistance (EMDR). Central to EMDR are chemokine- and integrin-mediated interactions between lymphoma and the TME. Further, stromal cell-B cell adhesion confers a sustained BCR signaling leading to chemokine and integrin activation. Recently, the inhibitors of BCR signaling have garnered a substantial clinical interest because of their effectiveness in B-cell disorders. The efficacy of these agents is, at least in part, attributed to attenuation of BCR-dependent lymphoma-TME interactions. In this review, we discuss the pivotal role of BCR signaling in the integration of intrinsic and extrinsic determinants of TME-mediated lymphoma survival and drug resistance. PMID:24037527

  17. Activation/Inhibition of mast cells by supra-optimal antigen concentrations

    OpenAIRE

    Huber, Michael

    2013-01-01

    Mast cells (MCs) are tissue resident cells of hemopoietic origin and are critically involved in allergic diseases. MCs bind IgE by means of their high-affinity receptor for IgE (FcεRI). The FcεRI belongs to a family of multi-chain immune recognition receptors and is activated by cross-linking in response to multivalent antigens (Ags)/allergens. Activation of the FcεRI results in immediate release of preformed granular substances (e.g. histamine, heparin, and proteases), generation of arachido...

  18. Primary parotid B-cell lymphoma successfully treated with chemotherapy plus highly active antiretroviral therapy with prolonged survival and immune reconstitution in an acquired immunodeficiency syndrome patient: Case report and review of the literature

    Directory of Open Access Journals (Sweden)

    Marcelo Corti

    2014-01-01

    Full Text Available Non-Hodgkin′s lymphoma (NHL is the second most common acquired immunodeficiency syndrome (AIDS-defining cancer. In this population, up to 70-80% of cases may present as extranodal location as the primary clinical manifestation of the neoplasm disease. Gastrointestinal tract is the most frequent location of AIDS-associated NHL. However, salivary gland involvement, including the parotid gland is a rare complication in human immunodeficiency virus (HIV-patients. Here, we describe a patient seropositive for the HIV, who developed a primary NHL of the parotid gland histologically classified as a high-grade diffuse large B-cell lymphoma. Patient was treated with a combination of chemotherapy plus highly active antiretroviral therapy with a good clinical, virological and immunological response and a prolonged survival, more than 5 years, without evidence of neoplasm relapse.

  19. Constitutive activation of extracellular signal-regulated kinase predisposes diffuse large B-cell lymphoma cell lines to CD40-mediated cell death

    DEFF Research Database (Denmark)

    Hollmann, C Annette; Owens, Trevor; Nalbantoglu, Josephine; Hudson, Thomas J; Sladek, Robert

    2006-01-01

    , including LCK and VAV. In addition, CD40-sensitive DLBCL cell lines also displayed constitutive activation of extracellular signal-regulated kinase (ERK) and failed to undergo apoptosis when ERK phosphorylation was inhibited. In contrast, CD40-resistant lines showed no constitutive activation of ERK and no...

  20. Antibodies recognizing a variety of different structural motifs on meningococcal Lip antigen fail to demonstrate bactericidal activity.

    Science.gov (United States)

    Tinsley, C R; Virji, M; Heckels, J E

    1992-11-01

    The neisserial Lip antigen is a conserved antigen associated with the pathogenic Neisseria species, and is composed of multiple repeats of a consensus pentapeptide. A series of monoclonal antibodies reacting with meningococcal Lip antigen were subjected to epitope mapping, using solid-phase synthetic peptides based on the consensus repeat sequence. The antibodies were found to recognize different continuous epitopes based on the consensus sequence. One monoclonal antibody was utilized in affinity chromatography to obtain purified Lip antigen and the antigen was used for immunization of mice. The resulting antisera did not recognize Lip antigen on Western blots but reacted specifically with Lip antigen in immune precipitation experiments, indicating that the predominant polyclonal immune response was directed against conformational epitopes. Despite the diversity of both continuous and conformational epitopes recognized by the antibodies produced, none of the antibodies demonstrated the ability to promote complement-mediated bactericidal activity. Thus despite its initial apparent promise as a potential vaccine candidate the case for the inclusion of Lip antigen in vaccine formulation cannot be supported at present. PMID:1282535

  1. T and B cell cooperation: a dance of life and death.

    Science.gov (United States)

    Bernard, Alain; Coitot, Sylvie; Brémont, Aurore; Bernard, Ghislaine

    2005-02-15

    In order for the body to develop a good antibody response, B cells need to react intimately with antigen specific T cells. Experimental evidence using hapten-carriers revealed that T and B cells do not recognize the same epitope and this led to the view that the physical contact is mediated by the antigen. Although the modern concept of antigen presentation has changed our perception on how the antigen can bridge both cells, the basic virtues of earlier bridging models remain. Over the past few years, a number of surface ligand-receptor pathways have been described, most of them belonging either to the CD28/B7 Ig or to the TNF/TNFR-like families. These act in concert, whether they are agonist or antagonist, in a timely and spatially organized manner. They form cascades of successive induction and recruitment to ensure that T-B cooperation is closely controlled at all stages of antibody induction. PMID:15699753

  2. Kinetics of T cell-activation molecules in response to Mycobacterium tuberculosis antigens

    Directory of Open Access Journals (Sweden)

    Antas Paulo RZ

    2002-01-01

    Full Text Available The phenotypic features acquired subsequent to antigen-specific stimulation in vitro were evaluated by means of the kinetic expressions of CD69 and CD25 activation molecules on T lymphocytes and assayed by flow cytometry in response to PPD, Ag85B, and ferritin in PPD-positive healthy control individuals. In response to PHA, CD69 staining on both CD4+ and CD8+ T cells became initially marked after 4 h, peaked at 24 h, and quickly decreased after 120 h. For CD25, a latter expression was detected around 8 h, having increased after 96 h. As expected, the response rate to the mycobacterial antigens was much lower than that to the mitogen. Positive staining was high after 96 h for CD25 and after 24 h for CD69. CD69 expression was significantly enhanced (p < 0.05 on CD8+ as compared to CD4+ T cells. High levels were also found between 96-120 h. Regarding Ag85B, CD25+ cells were mostly CD4+ instead of CD8+ T cells. Moreover, in response to ferritin, a lower CD25 expression was noted. The present data will allow further characterization of the immune response to new mycobacterial-specific antigens and their evaluation for possible inclusion in developing new diagnostic techniques for tuberculosis as well in a new vaccine to prevent the disease.

  3. Evaluation of the antigenicity of hydrolyzed cow's milk protein formulas using the mouse basophil activation test.

    Science.gov (United States)

    Iwamoto, Hiroshi; Matsubara, Takeshi; Nakazato, Yuki; Namba, Kazuyoshi; Takeda, Yasuhiro

    2016-02-01

    Hypoallergenic infant formulas are widely used for infants with cow's milk allergy. The aim of this study was to assess the utility of the mouse basophil activation test (BAT) in the evaluation of residual antigenicity in these formulas. Whole blood samples derived from β-lactoglobulin- or casein-immunized mice were incubated with one of the following formulas: conventional, partially hydrolyzed, or extensively hydrolyzed. Basophilic activation was analyzed by flow cytometry using an IgE-dependent activation marker CD200R1 and an IgG-dependent activation marker CD200R3. Systemic anaphylaxis was induced by i.v. injection of milk formula and results were compared. Conventional formula induced pronounced changes in CD200R1 and CD200R3 expression on basophils, whereas extensively hydrolyzed formulas did not elicit any changes in these markers. Similarly, challenge with conventional formula induced anaphylaxis, whereas extensively hydrolyzed formulas did not induce anaphylaxis. Although the partially hydrolyzed formula also induced basophilic activation and systemic anaphylaxis, the magnitude of these effects was smaller than that observed with the conventional formula. Compared to CD200R1, the observed trend in CD200R3 expression resembled the results obtained from systemic anaphylaxis test more closely. These findings show that mouse BAT, in particular using CD200R3, is highly useful for the evaluation of antigenicity of milk formulas. PMID:26626100

  4. Bacteroides gingivalis antigens and bone resorbing activity in root surface fractions of periodontally involved teeth

    Energy Technology Data Exchange (ETDEWEB)

    Patters, M.R.; Landsberg, R.L.; Johansson, L.A.; Trummel, C.L.; Robertson, P.R. (Department of Periodontology, University of Connecticut, School of Dental Medicine, Farmington, Connecticut, U.S.A.)

    1982-01-01

    Bone resorbing activity and the presence of antigens of Bacteroides gingivalis were assessed in plaque, calculus, cementum, and dentin obtained from roots of teeth previously exposed to periodontitis. Each fraction was obtained by scaling the root surface. The fraction were extracted by stirring and sonication, and the soluble centrifuged, sterilized, dialyzed, and adjusted to equivalent protein concentrations. Cementum and dentin extracts from impacted teeth were prepared similarly and served as controls. Stimulation of bone resorption by each extract was assessed in organ cultures of fetal rat bones by measurement of release of previously-incorporated /sup 45/Ca from the bone into the medium. In some groups of teeth, calculus and cementum were treated with acid prior to scaling. Citric acid washes were recovered and dialyzed. An enzyme-linked immunosorbent assay (ELISA) was used to assess the extracts for the presence of antigens reactive with an antiserum to B. gingivalis. Significant stimulation of bone resorption was found in all calculus and periodontally-involved cementum preparations. ELISA showed significant levels of B.gingivalis antigens in plaque, calculus, and cementum of periodontally-involved teeth, but not in involved dentin nor in cementum or dentin of impact teeth. Treatment with citric acid removed essentially all B.gingivalis antigens from cementum but not calculus. The results suggest that substances which stimulate bone resorption and substances which react with B. gingivalis antiserum are present in surface plaque, calculus, and cementum or periodontally-involved teeth. These substances are not present in cementum and dentin of impacted teeth nor in dentin of periodontally-involved teeth. Treatment by both scaling and citric demineralization will remove most of these substances from cementum of teeth previously exposed to periodontitis.

  5. Bacteroides gingivalis antigens and bone resorbing activity in root surface fractions of periodontally involved teeth

    International Nuclear Information System (INIS)

    Bone resorbing activity and the presence of antigens of Bacteroides gingivalis were assessed in plaque, calculus, cementum, and dentin obtained from roots of teeth previously exposed to periodontitis. Each fraction was obtained by scaling the root surface. The fraction were extracted by stirring and sonication, and the soluble centrifuged, sterilized, dialyzed, and adjusted to equivalent protein concentrations. Cementum and dentin extracts from impacted teeth were prepared similarly and served as controls. Stimulation of bone resorption by each extract was assessed in organ cultures of fetal rat bones by measurement of release of previously-incorporated 45Ca from the bone into the medium. In some groups of teeth, calculus and cementum were treated with acid prior to scaling. Citric acid washes were recovered and dialyzed. An enzyme-linked immunosorbent assay (ELISA) was used to assess the extracts for the presence of antigens reactive with an antiserum to B. gingivalis. Significant stimulation of bone resorption was found in all calculus and periodontally-involved cementum preparations. ELISA showed significant levels of B.gingivalis antigens in plaque, calculus, and cementum of periodontally-involved teeth, but not in involved dentin nor in cementum or dentin of impact teeth. Treatment with citric acid removed essentially all B.gingivalis antigens from cementum but not calculus. The results suggest that substances which stimulate bone resorption and substances which react with B. gingivalis antiserum are present in surface plaque, calculus, and cementum or periodontally-involved teeth. These substances are not present in cementum and dentin of impacted teeth nor in dentin of periodontally-involved teeth. Treatment by both scaling and citric demineralization will remove most of these substances from cementum of teeth previously exposed to periodontitis. (author)

  6. B-Cell Hematologic Malignancy Vaccination Registry

    Science.gov (United States)

    2015-09-15

    Monoclonal Gammopathy of Undetermined Significance; Multiple Myeloma; Waldenstrom Macroglobulinemia; Lymphocytosis; Lymphoma, Non-Hodgkin; B-Cell Chronic Lymphocytic Leukemia; Hematological Malignancies

  7. Reconstructing and mining the B cell repertoire with ImmunediveRsity

    OpenAIRE

    Cortina-Ceballos, Bernardo; Godoy-Lozano, Elizabeth Ernestina; Sámano-Sánchez, Hugo; Aguilar-Salgado, Andrés; Velasco-Herrera, Martín Del Castillo; Vargas-Chávez, Carlos; Velázquez-Ramírez, Daniel; Romero, Guillermo; Moreno, José; Téllez-Sosa, Juan; Martínez-Barnetche, Jesús

    2015-01-01

    The B cell antigen receptor repertoire is highly diverse and constantly modified by clonal selection. High-throughput DNA sequencing (HTS) of the lymphocyte repertoire (Rep-Seq) represents a promising technology to explore such diversity ex-vivo and assist in the identification of antigen-specific antibodies based on molecular signatures of clonal selection. Therefore, integrative tools for repertoire reconstruction and analysis from antibody sequences are needed. We developed ImmunediveRity,...

  8. Positive-unlabeled learning for the prediction of conformational B-cell epitopes

    OpenAIRE

    Ren, Jing; Liu, Qian; Ellis, John; Li, Jinyan

    2015-01-01

    Background The incomplete ground truth of training data of B-cell epitopes is a demanding issue in computational epitope prediction. The challenge is that only a small fraction of the surface residues of an antigen are confirmed as antigenic residues (positive training data); the remaining residues are unlabeled. As some of these uncertain residues can possibly be grouped to form novel but currently unknown epitopes, it is misguided to unanimously classify all the unlabeled residues as negati...

  9. Valency and density matter: Deciphering impacts of immunogen structures on immune responses against a tumor associated carbohydrate antigen using synthetic glycopolymers.

    Science.gov (United States)

    Qin, Qian; Yin, Zhaojun; Wu, Xuanjun; Haas, Karen M; Huang, Xuefei

    2016-09-01

    For successful carbohydrate based anti-cancer vaccines, it is critical that B cells are activated to secret antibodies targeting the tumor associated carbohydrate antigens (TACAs). Despite the availability of many TACA based constructs, systematic understanding of the effects of structural features on anti-glycan antibody responses is lacking. In this study, a series of defined synthetic glyco-polymers bearing a representative TACA, i.e., the Thomsen-nouveau (Tn) antigen, have been prepared to probe the induction of early B cell activation and antibody production via a T cell independent mechanism. Valency and density of the antigen in the polymers turned out to be critical. An average of greater than 6 Tn per chain was needed to induce antibody production. Glycopolymers with 40 antigens per chain and backbone molecular weight of 450 kDa gave the strongest stimulation to B cells in vitro, which correlated well with its in vivo activity. Deviations from the desired valency and density led to decreased antibody production or even antigen specific B cell non-responsiveness. These findings provide important insights on how to modulate anti-TACA immune responses facilitating the development of TACA based anti-cancer vaccines using glycopolymers. PMID:27294537

  10. Integrating understanding of epidemiology and genomics in B-cell non-Hodgkin lymphoma as a pathway to novel management strategies.

    Science.gov (United States)

    Glass, Samantha; Phan, Anh; Williams, Jessica N; Flowers, Christopher R; Koff, Jean L

    2016-03-01

    Non-Hodgkin lymphomas include a biologically and clinically heterogeneous group of cancers distinguished by genetics, histology, and treatment outcomes. New discoveries regarding the genomic alterations and epidemiological exposures associated with these lymphomas have enhanced our understanding of factors that contribute to lymphomagenesis for specific subtypes. We explore the impact of normal B-cell biology engineered for recognizing a wide variety of antigens on the development of specific lymphoma subtypes, review lymphoma genetics, and examine the epidemiology of B-cell NHLs including recent investigations of risk factors for particular lymphoma subtypes based on large pooled analyses. Burkitt lymphoma, an aggressive form of B-cell NHL involving translocation of the MYC gene and an immunoglobulin gene has been associated with a history of eczema, hepatitis C, and occupation as a cleaner. Increased risk of diffuse large B-cell lymphoma has been associated with increased young adult body mass index, history of B-cell-activating autoimmune diseases, hepatitis C, and several single nucleotide variants involving the human leukocyte antigen (HLA) region of chromosome 6 and non-HLA loci near EXOC2, PVT1, MYC, and NCOA1. Tumor sequencing studies suggest that multiple pathways are involved in the development of DLBCL. Additional studies of epidemiological exposures, genome wide associations, and tumor sequencing in follicular, lymphoplasmacytic, marginal zone, and mantle cell lymphoma demonstrate overlapping areas of increased risk factors and unique factors for specific subtypes. Integrating these findings is important for constructing comprehensive models of NHL pathogenesis, which could yield novel targets for therapy and strategies for lymphoma prevention in certain populations. PMID:27115168

  11. Evaluation of Intracellular Signaling Downstream Chimeric Antigen Receptors.

    Directory of Open Access Journals (Sweden)

    Hannah Karlsson

    Full Text Available CD19-targeting CAR T cells have shown potency in clinical trials targeting B cell leukemia. Although mainly second generation (2G CARs carrying CD28 or 4-1BB have been investigated in patients, preclinical studies suggest that third generation (3G CARs with both CD28 and 4-1BB have enhanced capacity. However, little is known about the intracellular signaling pathways downstream of CARs. In the present work, we have analyzed the signaling capacity post antigen stimulation in both 2G and 3G CARs. 3G CAR T cells expanded better than 2G CAR T cells upon repeated stimulation with IL-2 and autologous B cells. An antigen-driven accumulation of CAR+ cells was evident post antigen stimulation. The cytotoxicity of both 2G and 3G CAR T cells was maintained by repeated stimulation. The phosphorylation status of intracellular signaling proteins post antigen stimulation showed that 3G CAR T cells had a higher activation status than 2G. Several proteins involved in signaling downstream the TCR were activated, as were proteins involved in the cell cycle, cell adhesion and exocytosis. In conclusion, 3G CAR T cells had a higher degree of intracellular signaling activity than 2G CARs which may explain the increased proliferative capacity seen in 3G CAR T cells. The study also indicates that there may be other signaling pathways to consider when designing or evaluating new generations of CARs.

  12. Evaluation of Intracellular Signaling Downstream Chimeric Antigen Receptors.

    Science.gov (United States)

    Karlsson, Hannah; Svensson, Emma; Gigg, Camilla; Jarvius, Malin; Olsson-Strömberg, Ulla; Savoldo, Barbara; Dotti, Gianpietro; Loskog, Angelica

    2015-01-01

    CD19-targeting CAR T cells have shown potency in clinical trials targeting B cell leukemia. Although mainly second generation (2G) CARs carrying CD28 or 4-1BB have been investigated in patients, preclinical studies suggest that third generation (3G) CARs with both CD28 and 4-1BB have enhanced capacity. However, little is known about the intracellular signaling pathways downstream of CARs. In the present work, we have analyzed the signaling capacity post antigen stimulation in both 2G and 3G CARs. 3G CAR T cells expanded better than 2G CAR T cells upon repeated stimulation with IL-2 and autologous B cells. An antigen-driven accumulation of CAR+ cells was evident post antigen stimulation. The cytotoxicity of both 2G and 3G CAR T cells was maintained by repeated stimulation. The phosphorylation status of intracellular signaling proteins post antigen stimulation showed that 3G CAR T cells had a higher activation status than 2G. Several proteins involved in signaling downstream the TCR were activated, as were proteins involved in the cell cycle, cell adhesion and exocytosis. In conclusion, 3G CAR T cells had a higher degree of intracellular signaling activity than 2G CARs which may explain the increased proliferative capacity seen in 3G CAR T cells. The study also indicates that there may be other signaling pathways to consider when designing or evaluating new generations of CARs. PMID:26700307

  13. Engineered T Cells for the Adoptive Therapy of B-Cell Chronic Lymphocytic Leukaemia

    Directory of Open Access Journals (Sweden)

    Philipp Koehler

    2012-01-01

    Full Text Available B-cell chronic lymphocytic leukaemia (B-CLL remains an incurable disease due to the high risk of relapse, even after complete remission, raising the need to control and eliminate residual tumor cells in long term. Adoptive T cell therapy with genetically engineered specificity is thought to fulfil expectations, and clinical trials for the treatment of CLL are initiated. Cytolytic T cells from patients are redirected towards CLL cells by ex vivo engineering with a chimeric antigen receptor (CAR which binds to CD19 on CLL cells through an antibody-derived domain and triggers T cell activation through CD3ζ upon tumor cell engagement. Redirected T cells thereby target CLL cells in an MHC-unrestricted fashion, secret proinflammatory cytokines, and eliminate CD19+ leukaemia cells with high efficiency. Cytolysis of autologous CLL cells by patient's engineered T cells is effective, however, accompanied by lasting elimination of healthy CD19+ B-cells. In this paper we discuss the potential of the strategy in the treatment of CLL, the currently ongoing trials, and the future challenges in the adoptive therapy with CAR-engineered T cells.

  14. The microRNA miR-148a functions as a critical regulator of B cell tolerance and autoimmunity.

    Science.gov (United States)

    Gonzalez-Martin, Alicia; Adams, Brian D; Lai, Maoyi; Shepherd, Jovan; Salvador-Bernaldez, Maria; Salvador, Jesus M; Lu, Jun; Nemazee, David; Xiao, Changchun

    2016-04-01

    Autoreactive B cells have critical roles in a large diversity of autoimmune diseases, but the molecular pathways that control these cells remain poorly understood. We performed an in vivo functional screen of a lymphocyte-expressed microRNA library and identified miR-148a as a potent regulator of B cell tolerance. Elevated miR-148a expression impaired B cell tolerance by promoting the survival of immature B cells after engagement of the B cell antigen receptor by suppressing the expression of the autoimmune suppressor Gadd45α, the tumor suppressor PTEN and the pro-apoptotic protein Bim. Furthermore, increased expression of miR-148a, which occurs frequently in patients with lupus and lupus-prone mice, facilitated the development of lethal autoimmune disease in a mouse model of lupus. Our studies demonstrate a function for miR-148a as a regulator of B cell tolerance and autoimmunity. PMID:26901150

  15. Targeting CD19 in B-cell lymphoma: emerging role of SAR3419

    International Nuclear Information System (INIS)

    Non-Hodgkin lymphoma symbolizes a heterogeneous group of diseases resulting from malignant transformation of lymphocytes with differing patterns of behavior and responses to treatment. The potential curability of non-Hodgkin lymphoma differs among the various histologic subtypes and is associated in part with the stage at presentation. CD19 antigen is a type I transmembrane glycoprotein belonging to the immunoglobulin Ig superfamily. CD19 is specifically expressed in normal and neoplastic B-cells. Recent study showed that in a mouse model, CD19 and c-Myc synergize functionally to accelerate B-cell lymphomagenesis, which is associated with increased disease severity. Specificity is the most important challenge in cancer therapeutics. Antibody–drug conjugates have the prospect of enhancing the therapeutic efficacy over unconjugated monoclonal antibodies through the selective delivery of cytotoxic agents to cancer cells. The ubiquitous expression of CD19 in these tumors, especially at an earlier stage and the property of efficient internalization, makes CD19 an attractive and affective target for antibody–drug conjugate therapy as compared to CD20. SAR3419 (huB4-DM4) is a novel antibody–drug conjugate that is composed of a humanized monoclonal IgG1 anti-CD19 antibody (huB4) attached to the potent cytotoxic drug, a maytansine derivative (DM4), through a cleavable disulfide cross-linking agent N-Succinimidyl-4-2-pyridyldithio butanoic acid (SPDB). The preclinical efficacy of maytansine derivative–anti-CD19 conjugate was demonstrated in our laboratory, and SAR3419 was found to be more effective than CHOP in a xenograft model. Phase I trials have also been conducted on the basis of preclinical studies that demonstrated promising antitumor activity with acceptable safety results in human B-cell lymphoma models. Additional trials are ongoing and will provide additional insight into the full potential of this novel drug

  16. Intravenous immunoglobulin replacement therapy in common variable immunodeficiency induces B cell depletion through differentiation into apoptosis-prone CD21(low) B cells.

    Science.gov (United States)

    Mitrevski, Milica; Marrapodi, Ramona; Camponeschi, Alessandro; Lazzeri, Cristina; Todi, Laura; Quinti, Isabella; Fiorilli, Massimo; Visentini, Marcella

    2014-12-01

    Intravenous immunoglobulin (IVIG), besides its use as replacement therapy in patients with antibody deficiencies, is broadly used as an immunomodulatory agent for the treatment of autoimmune and inflammatory disorders. The mechanisms of action of IVIG include Fc receptor blockade, inhibition of cytokines and growth factors, modulation of macrophages and dendritic cells, enhancement of regulatory T cells, and modulation of B cells through the FcγRIIB receptor and CD22. Recent studies suggest that in vitro exposure of human B cells to IVIG determines functional changes reminiscent of anergy and that IVIG treatment of patients with common variable immunodeficiency (CVID) induces in B cells ERK activation, a feature of anergy. Here, we show that IVIG therapy drives the B cells of patients with CVID to down-regulate CD21 expression and to assume the peculiar phenotype of the anergic-like, apoptosis-prone CD21(low) B cells that are spontaneously expanded in a subset of CVID and in some other immunological disorders. The CD21(low) B cells newly generated after IVIG infusion undergo spontaneous apoptosis upon in vitro culture. Furthermore, IVIG infusion is rapidly followed by a significant, although discrete, decrease in the number of circulating B cells, but not of T cells or of natural killer cells. These findings suggest that IVIG therapy may constrain antibody responses by inducing B cell depletion through differentiation into CD21(low) B cells that undergo accelerated apoptosis. PMID:25407649

  17. PAX5 promotes pre-B cell proliferation by regulating the expression of pre-B cell receptor and its downstream signaling.

    Science.gov (United States)

    Xue, Kai; Song, Jiazhe; Yang, Yan; Li, Zhi; Wu, Chunhua; Jin, Jinhua; Li, Wenzhe

    2016-05-01

    PAX5 is indispensable for the commitment of early lymphoid progenitors to the B cell lineage as well as for the development of B cells. Although previous studies have indicated that the Pax5-conditional-knockout mouse exhibited dedifferentiation of mature B cell and the development of aggressive lymphomas, the changes of Pax5 gene expressions in pre-B cells have not been analyzed. To understand the functional importance of Pax5 gene in the proliferation and survival of pre-B cells, we established a Pax5-knockdown model using 70Z/3 pre-B cell line. Pax5 knockdown 70Z/3 cells (70Z/3-KD cells) showed down-regulations of pre-BCR compounds such as CD19, BLNK, Id2 and λ5. The signaling via pre-BCRs was significantly diminished in the 70Z/3-KD cells, and this alteration was normalized by restored Pax5 gene expression. Loss of PAX5 reduced the growth rates in the 70Z/3-KD cells, compared to the mock cells. Meanwhile, the proliferation of pre-B cells was reduced by the knockdown of Pax5 gene. Moreover, further examinations showed that PAX5 was also activated in B cell acute lymphoblastic leukemia (B-ALL) as a cell proliferation enhancer. These findings suggested that pax5 is critically important for the proliferation and survival of pre-B cells. PMID:27016671

  18. Fucoidan prevents Cε germline transcription and NFκB p52 translocation for IgE production in B cells

    International Nuclear Information System (INIS)

    Fucoidan, a dietary fiber contained in seaweed, reduces the increase of antigen-specific IgE in mice exposed to ovalbumin. In this study, we investigated the effect of fucoidan on IgE production and intracellular events in B cells in vitro. Fucoidan inhibited the production of IgE and Cε germline transcription in murine B cells induced by IL-4 (100 ng/ml) and anti-CD40 antibodies (10 μg/ml), whereas it stimulated cell proliferation. A significant effect of fucoidan on IgE production was observed when B cells were stimulated with a higher dose (5 μg/ml) of anti-CD40 antibodies, but not when stimulated with lower doses (1.25, 2.5 μg/ml), regardless of the IL-4 concentrations. Moreover, nuclear translocation of NFκB p52, but neither that of NFκB p65, nor the phosphorylation of JAK1 and STAT6 was reduced by fucoidan. These results suggest that fucoidan inhibited IgE production by preventing the NFκB p52-mediated pathways activated by CD40

  19. Role of Phospholipase C-L2, a Novel Phospholipase C-Like Protein That Lacks Lipase Activity, in B-Cell Receptor Signaling

    OpenAIRE

    Takenaka, Kei; Fukami, Kiyoko; Otsuki, Makiko; Nakamura, Yoshikazu; Kataoka, Yuki; Wada, Mika; Tsuji, Kohichiro; Nishikawa, Shin-ichi; Yoshida, Nobuaki; Takenawa, Tadaomi

    2003-01-01

    Phospholipase C (PLC) plays important roles in phosphoinositide turnover by regulating the calcium-protein kinase C signaling pathway. PLC-L2 is a novel PLC-like protein which lacks PLC activity, although it is very homologous with PLC δ. PLC-L2 is expressed in hematopoietic cells, but its physiological roles and intracellular functions in the immune system have not yet been clarified. To elucidate the physiological function of PLC-L2, we generated mice which had a genetic PLC-L2 deficiency. ...

  20. Hepatitis C virus - associated B cell non-Hodgkin's lymphoma

    Science.gov (United States)

    Mihăilă, Romeo-Gabriel

    2016-01-01

    The hepatitis C virus (HCV) infected patients are prone to develop bone marrow or various tissue infiltrates with monoclonal B cells, monoclonal B lymphocytosis or different types of B cell non-Hodgkin’s lymphoma (BCNHL), of which the most common are splenic marginal zone BCNHL, diffuse large BCNHL and follicular lymphoma. The association between chronic HCV infection and non Hodgkin’s lymphoma has been observed especially in areas with high prevalence of this viral infection. Outside the limitations of some studies that have been conducted, there are also geographic, environmental, and genetic factors that contribute to the epidemiological differences. Various microenvironmental signals, such as cytokines, viral antigenic external stimulation of lymphocyte receptors by HCV antigens, and intercellular interactions contribute to B cell proliferation. HCV lymphotropism and chronic antigenic stimulation are involved in B-lymphocyte expansion, as mixted cryoglobulinemia or monoclonal gammopathy of undetermined significance, which can progress to BCNHL. HCV replication in B lymphocytes has oncogenic effect mediated by intracellular HCV proteins. It is also involved in an important induction of reactive oxygen species that can lead to permanent B lymphocyte damage, as DNA mutations, after binding to surface B-cell receptors. Post-transplant lymphoproliferative disorder could appear and it has a multiclonal potentiality that may develop into different types of lymphomas. The hematopoietic stem cell transplant made for lymphoma in HCV-infected patients can increase the risk of earlier progression to liver fibrosis and cirrhosis. HCV infected patients with indolent BCNHL who receive antiviral therapy can be potentially cured. Viral clearance was related to lymphoma response, fact that highlights the probable involvement of HCV in lymphomagenesis. Direct acting antiviral drugs could be a solution for the patients who did not tolerate or respond to interferon, as they

  1. A Viral Vectored Prime-Boost Immunization Regime Targeting the Malaria Pfs25 Antigen Induces Transmission-Blocking Activity

    OpenAIRE

    Goodman, Anna L.; Blagborough, Andrew M.; Sumi Biswas; Yimin Wu; Hill, Adrian V.; Sinden, Robert E.; Draper, Simon J

    2011-01-01

    The ookinete surface protein Pfs25 is a macrogamete-to-ookinete/ookinete stage antigen of Plasmodium falciparum, capable of exerting high-level anti-malarial transmission-blocking activity following immunization with recombinant protein-in-adjuvant formulations. Here, this antigen was expressed in recombinant chimpanzee adenovirus 63 (ChAd63), human adenovirus serotype 5 (AdHu5) and modified vaccinia virus Ankara (MVA) viral vectored vaccines. Two immunizations were administered to mice in a ...

  2. Tritium (3H) radiolabeling of protein A and antibody to high specific activity: Application to cell surface antigen radioimmunoassays

    International Nuclear Information System (INIS)

    Staphylococcal protein A and several different immunoglobulins have been radiolabeled to high specific activities (> 106 cpm/μg) by reductive methylation with tritiated (3H) sodium borohydride. The proteins retain excellent functional and antigenic properties. The utility of these reagents in a variety of assays for cell surface antigens is illustrated. The results indicate that this radiolabeling procedure may become the method of choice for many cell surface and solution immunoassays. (Auth.)

  3. Analysis of immune responses against a wide range of Mycobacterium tuberculosis antigens in patients with active pulmonary tuberculosis

    OpenAIRE

    Kassa, D.; Ran, L.; Geberemeskel, W.; Tebeje, M; Alemu, A.; Selase, A.; Tegbaru, B.; Franken, K. L. M. C.; Friggen, A. H.; van Meijgaarden, K. E.; Ottenhoff, T. H. M.; Wolday, D.; Messele, T.; van Baarle, D.

    2012-01-01

    Characterizing host immune responses to molecular targets of Mycobacterium tuberculosis is essential to develop effective immunodiagnostics and better vaccines. We investigated the immune response against a large series of M. tuberculosis antigens, including 5 classical and 64 nonclassical (39 DosR regulon-encoded, 4 resuscitation-promoting factor [RPF], and 21 reactivation-associated) antigens in active-pulmonary-tuberculosis (TB) patients. Whole blood from TB patients (n = 34) was stimulate...

  4. INDUCEMENT OF ANTITUMOR-IMMUNITY BY DC ACTIVATED BY HSP70-H22 TUMOR ANTIGEN PEPTIDE

    Institute of Scientific and Technical Information of China (English)

    冯作化; 黄波; 张桂梅; 李东; 王洪涛

    2003-01-01

    Objective: To investigate the feasibility of decreasing the dosage of tumor antigen peptides by dendritic cell (DC)-presenting and the characteristics of modification of DC by heat shock protein (Hsp70) and antigen peptides. Methods: Peptides were bound to Hsp70 and used to modify DC in vitro. The metabolism of the modified DC and the cytokines secreted by the modified DC were determined. The activation of lymphocytes by the modified DC and Hsp70-H22 peptides was tested. The cytotoxicity of the activated lymphocytes to H22 tumor cells was analyzed. The inhibitory effect of tumor in mice by the injection of DC and Hsp70-H22 peptides was tested. Results: 0.15μg of H22 peptides bound with Hsp70 could make 2×105 DC mature. 4×103 matured DC could activate 2×106 lymphocytes. The same amount of lymphocytes could be activated to produce similar cytotoxicity to tumor cells by either DC modified by 0.003μg of peptides bound with Hsp70 or by direct stimulation with 0.15μg of peptides bound with Hsp70. The dosage of peptides could be reduced by about 50 folds if the modified DC was used for injection instead of Hsp70-peptides. Peptides from normal hepatocytes, bound with Hsp70, could not make DC mature, nor activate lymphocytes through DC. Conclusion: The dosage of Hsp70-H22 peptides can be reduced significantly by DC-presenting to activate lymphocytes. Peptides from normal cells could not activate lymphocytes by either Hsp70-presenting or DC-presenting and they have little chance to induce autoimmunity.

  5. Structural analysis of B-cell epitopes in antibody:protein complexes

    DEFF Research Database (Denmark)

    Kringelum, Jens Vindahl; Nielsen, Morten; Padkjær, Søren Berg; Lund, Ole

    developed a novel framework for comparing and superimposing B-cell epitopes and applied it on a dataset of 107 non-similar antigen:antibody structures extracted from the PDB database. With the presented framework, we were able to describe the general B-cell epitope as a flat, oblong, oval shaped volume......The binding of antigens to antibodies is one of the key events in an immune response against foreign molecules and is a critical element of several biomedical applications including vaccines and immunotherapeutics. For development of such applications, the identification of antibody binding sites...... (B-cell epitopes) is essential. However experimental epitope mapping is highly cost-intensive and computer-aided methods do in general have moderate performance. One major reason for this moderate performance is an incomplete understanding of what characterizes an epitope. To fill this gap, we here...

  6. B-cell activating CpG ODN 1668 enhance the immune response of Pacific red snapper (Lutjanus peru) exposed to Vibrio parahaemolitycus.

    Science.gov (United States)

    Cárdenas-Reyna, Tomás; Angulo, Carlos; Hori-Oshima, Sawako; Velázquez-Lizárraga, Esteban; Reyes-Becerril, Martha

    2016-09-01

    B-class CpG ODN 1668 is known to possess clear immunostimulatory properties. In this study, we investigated the potential ability of CpG ODN 1668 to enhance the immune response of Pacific red snapper exposed to Vibrio parahaemolyticus. Four different treatments were evaluated in Pacific red snapper: (1) stimulatory CpG ODN 1668, (2) stimulatory CpG ODN 1668 and V. parahaemolyticus, (3) exposure only to V. parahaemolyticus and (4) PBS. Samples were taken at 24, 72, 168 and 240 h of stimulation/infection. The results show that intraperitoneal injection of CpG-ODN 1668 enhanced the anti-protease, superoxide dismutase and catalase activities in serum. CpG ODN 1668 upregulated TLR9 and IgM gene expression in head-kidney, intestine and skin, with higher expression in head-kidney. A higher correlation was observed between TLR9 and IgM in head-kidney and intestine. Finally, no histopathological damages were observed in fish stimulated with CpG ODN 1668. In contrast, melanomacrophages-like structures were present in higher numbers in infected fish. Taken together, these results indicate that CpG ODN 1668 activates innate immune response and upregulate the TLR9 and IgM-mediated immune response. These results may be exploited for the control of Vibriosis in farmed Pacific red snapper. PMID:27143535

  7. Street-Experienced Peripheral B cells Traffic to the CNS in Multiple Sclerosis

    OpenAIRE

    Lu, Daniel R.; Robinson, William H

    2014-01-01

    High-throughput sequencing of immunoglobulin repertoires in patients with multiple sclerosis reveals that antigen-experienced B cells traffic between the peripheral lymph nodes and the central nervous system, suggesting the involvement of peripheral immunity in the autoimmune pathogenesis of multiple sclerosis.

  8. BiovaxID, a personalized therapeutic vaccine against B-cell lymphomas

    Czech Academy of Sciences Publication Activity Database

    Reiniš, Milan

    2008-01-01

    Roč. 10, č. 5 (2008), s. 526-534. ISSN 1464-8431 Institutional research plan: CEZ:AV0Z50520514 Keywords : B-cell lymphomas * tumor antigen * therapeutic vaccine Subject RIV: EB - Genetics ; Molecular Biology Impact factor: 2.913, year: 2008

  9. Neutralizing antibodies derived from the B cells of 1918 influenza pandemic survivors.

    Science.gov (United States)

    Yu, Xiaocong; Tsibane, Tshidi; McGraw, Patricia A; House, Frances S; Keefer, Christopher J; Hicar, Mark D; Tumpey, Terrence M; Pappas, Claudia; Perrone, Lucy A; Martinez, Osvaldo; Stevens, James; Wilson, Ian A; Aguilar, Patricia V; Altschuler, Eric L; Basler, Christopher F; Crowe, James E

    2008-09-25

    Investigation of the human antibody response to influenza virus infection has been largely limited to serology, with relatively little analysis at the molecular level. The 1918 H1N1 influenza virus pandemic was the most severe of the modern era. Recent work has recovered the gene sequences of this unusual strain, so that the 1918 pandemic virus could be reconstituted to display its unique virulence phenotypes. However, little is known about adaptive immunity to this virus. We took advantage of the 1918 virus sequencing and the resultant production of recombinant 1918 haemagglutinin (HA) protein antigen to characterize at the clonal level neutralizing antibodies induced by natural exposure of survivors to the 1918 pandemic virus. Here we show that of the 32 individuals tested that were born in or before 1915, each showed seroreactivity with the 1918 virus, nearly 90 years after the pandemic. Seven of the eight donor samples tested had circulating B cells that secreted antibodies that bound the 1918 HA. We isolated B cells from subjects and generated five monoclonal antibodies that showed potent neutralizing activity against 1918 virus from three separate donors. These antibodies also cross-reacted with the genetically similar HA of a 1930 swine H1N1 influenza strain, but did not cross-react with HAs of more contemporary human influenza viruses. The antibody genes had an unusually high degree of somatic mutation. The antibodies bound to the 1918 HA protein with high affinity, had exceptional virus-neutralizing potency and protected mice from lethal infection. Isolation of viruses that escaped inhibition suggested that the antibodies recognize classical antigenic sites on the HA surface. Thus, these studies demonstrate that survivors of the 1918 influenza pandemic possess highly functional, virus-neutralizing antibodies to this uniquely virulent virus, and that humans can sustain circulating B memory cells to viruses for many decades after exposure-well into the tenth

  10. Self-enforcing Feedback Activation between BCL6 and Pre-B Cell Receptor Signaling Defines a Distinct Subtype of Acute Lymphoblastic Leukemia

    Science.gov (United States)

    Geng, Huimin; Hurtz, Christian; Lenz, Kyle B.; Chen, Zhengshan; Baumjohann, Dirk; Thompson, Sarah; Goloviznina, Natalya; Chen, Wei-Yi; Huan, Jianya; LaTocha, Dorian; Ballabio, Erica; Xiao, Gang; Lee, Jae-Woong; Deucher, Anne; Qi, Zhongxia; Park, Eugene; Huang, Chuanxin; Nahar, Rahul; Kweon, Soo-Mi; Shojaee, Seyedmehdi; Chan, Lai N.; Yu, Jingwei; Kornblau, Steven M.; Bijl, Janetta J.; Ye, B. Hilda; Ansel, Mark; Paietta, Elisabeth; Melnick, Ari; Hunger, Stephen P.; Kurre, Peter; Tyner, Jeffrey W.; Loh, Mignon L.; Roeder, Robert G.; Druker, Brian J.; Burger, Jan. A.; Milne, Thomas A.; Chang, Bill H.; Müschen, Markus

    2015-01-01

    SUMMARY Studying 830 pre-B ALL cases from four clinical trials, we found that human ALL can be divided into two fundamentally distinct subtypes based on pre-BCR function. While absent in the majority of ALL cases, tonic pre-BCR signaling was found in 112 cases (13.5%). In these cases, tonic pre-BCR signaling induced activation of BCL6, which in turn increased pre-BCR signaling output at the transcriptional level. Interestingly, inhibition of pre-BCR-related tyrosine kinases reduced constitutive BCL6 expression and selectively killed patient-derived pre-BCR+ ALL cells. These findings identify a genetically and phenotypically distinct subset of human ALL that critically depends on tonic pre-BCR signaling. In vivo treatment studies suggested that pre-BCR tyrosine kinase inhibitors are useful for the treatment of patients with pre-BCR+ ALL. PMID:25759025

  11. Epigenetic Control of B Cell Development and B-Cell-Related Immune Disorders.

    Science.gov (United States)

    Bao, Yan; Cao, Xuetao

    2016-06-01

    B lymphocytes are generally recognized as the essential component of humoral immunity and also a regulator of innate immunity. The development of B cells is precisely regulated by a variety of factors via different mechanisms, including cytokine/cytokine receptors, signal transduction molecules, and transcription factors. Recent findings suggest that epigenetic factors, such as DNA methylation, histone modification, and non-coding RNA, play critical roles in establishing B cell lineage-specific gene expression profiles to define and sustain B cell identity and function. Epigenetic modifications are also sensitive to external stimuli and might bridge genetic and environmental factors in the pathogenesis or control of B-cell-related immune disorders, such as autoimmune diseases, lymphoma, and leukemia. Better understanding of the epigenetic mechanisms for regulating B cell development and involving B cell abnormal differentiation and function will shed light on the design of new therapeutic approaches to B-cell-related diseases, and potential candidates of epigenetic modulators may be identified to target epigenetic pathways to prevent or treat B cell disorders. We summarize the relevance of epigenetic marks and landscapes in the stages of B cell development, discuss the interaction of the transcriptional networks and epigenetic changes, and review the involvement of epigenetic risk in the pathogenesis of B-cell-related diseases. Understanding how specific epigenetic alterations contribute to the development of B-cell-related autoimmunity and malignancies is instrumental to control B cell disorders. PMID:26066671

  12. B-cell subsets, signaling and their roles in secretion of autoantibodies.

    Science.gov (United States)

    Iwata, S; Tanaka, Y

    2016-07-01

    B cells play a pivotal role in the pathogenesis of autoimmune diseases. In patients with systemic lupus erythematosus (SLE), the percentages of plasmablasts and IgD(-)CD27(-) double-negative memory B cells in peripheral blood are significantly increased, while IgD(+)CD27(+) IgM memory B cells are significantly decreased compared to healthy donors. The phenotypic change is significantly associated with disease activity and concentration of autoantibodies. Treatment of B-cell depletion using rituximab results in the reconstitution of peripheral B cells in SLE patients with subsequent improvement in disease activity. Numerous studies have described abnormalities in B-cell receptor (BCR)-mediated signaling in B cells of SLE patients. Since differences in BCR signaling are considered to dictate the survival or death of naïve and memory B cells, aberrant BCR signal can lead to abnormality of B-cell subsets in SLE patients. Although Syk and Btk function as key molecules in BCR signaling, their pathological role in SLE remains unclear. We found that Syk and Btk do not only transduce activation signal through BCR, but also mediate crosstalk between BCR and Toll-like receptor (TLR) as well as BCR and JAK-STAT pathways in human B cells in vitro. In addition, pronounced Syk and Btk phosphorylation was observed in B cells of patients with active SLE compared to those of healthy individuals. The results suggest the involvement of Syk and Btk activation in abnormalities of BCR-mediated signaling and B-cell phenotypes during the pathological process of SLE and that Syk, Btk and JAK are potential therapeutic targets in SLE. PMID:27252261

  13. B-cell repertoire responses to varicella-zoster vaccination in human identical twins.

    Science.gov (United States)

    Wang, Chen; Liu, Yi; Cavanagh, Mary M; Le Saux, Sabine; Qi, Qian; Roskin, Krishna M; Looney, Timothy J; Lee, Ji-Yeun; Dixit, Vaishali; Dekker, Cornelia L; Swan, Gary E; Goronzy, Jörg J; Boyd, Scott D

    2015-01-13

    Adaptive immune responses in humans rely on somatic genetic rearrangements of Ig and T-cell receptor loci to generate diverse antigen receptors. It is unclear to what extent an individual's genetic background affects the characteristics of the antibody repertoire used in responding to vaccination or infection. We studied the B-cell repertoires and clonal expansions in response to attenuated varicella-zoster vaccination in four pairs of adult identical twins and found that the global antibody repertoires of twin pair members showed high similarity in antibody heavy chain V, D, and J gene segment use, and in the length and features of the complementarity-determining region 3, a major determinant of antigen binding. These twin similarities were most pronounced in the IgM-expressing B-cell pools, but were seen to a lesser extent in IgG-expressing B cells. In addition, the degree of antibody somatic mutation accumulated in the B-cell repertoire was highly correlated within twin pair members. Twin pair members had greater numbers of shared convergent antibody sequences, including mutated sequences, suggesting similarity among memory B-cell clonal lineages. Despite these similarities in the memory repertoire, the B-cell clones used in acute responses to ZOSTAVAX vaccination were largely unique to each individual. Taken together, these results suggest that the overall B-cell repertoire is significantly shaped by the underlying germ-line genome, but that stochastic or individual-specific effects dominate the selection of clones in response to an acute antigenic stimulus. PMID:25535378

  14. Fish oil disrupts MHC class II lateral organization on the B-cell side of the immunological synapse independent of B-T cell adhesion.

    Science.gov (United States)

    Rockett, Benjamin Drew; Melton, Mark; Harris, Mitchel; Bridges, Lance C; Shaikh, Saame Raza

    2013-11-01

    Fish oil-enriched long chain n-3 polyunsaturated fatty acids disrupt the molecular organization of T-cell proteins in the immunological synapse. The impact of fish oil derived n-3 fatty acids on antigen-presenting cells, particularly at the animal level, is unknown. We previously demonstrated B-cells isolated from mice fed with fish oil-suppressed naïve CD4(+) T-cell activation. Therefore, here we determined the mechanistic effects of fish oil on murine B-cell major histocompatibility complex (MHC) class II molecular distribution using a combination of total internal reflection fluorescence, Förster resonance energy transfer and confocal imaging. Fish oil had no impact on presynaptic B-cell MHC II clustering. Upon conjugation with transgenic T-cells, fish-oil suppressed MHC II accumulation at the immunological synapse. As a consequence, T-cell protein kinase C theta (PKCθ) recruitment to the synapse was also diminished. The effects were independent of changes in B-T cell adhesion, as measured with microscopy, flow cytometry and static cell adhesion assays with select immune ligands. Given that fish oil can reorganize the membrane by lowering membrane cholesterol levels, we then compared the results with fish oil to cholesterol depletion using methyl-B-cyclodextrin (MβCD). MβCD treatment of B-cells suppressed MHC II and T-cell PKCθ recruitment to the immunological synapse, similar to fish oil. Overall, the results reveal commonality in the mechanism by which fish oil manipulates protein lateral organization of B-cells compared to T-cells. Furthermore, the data establish MHC class II lateral organization on the B-cell side of the immunological synapse as a novel molecular target of fish oil. PMID:23791516

  15. Expansion of IgG+ B-cells during mitogen stimulation for memory B-cell ELISpot analysis is influenced by size and composition of the B-cell pool.

    Science.gov (United States)

    Scholzen, Anja; Nahrendorf, Wiebke; Langhorne, Jean; Sauerwein, Robert W

    2014-01-01

    The memory B-cell (MBC) ELISpot assay is the main technique used to measure antigen-specific MBCs as a readout of humoral immune memory. This assay relies on the ability of MBCs to differentiate into antibody-secreting cells (ASC) upon polyclonal stimulation. The total number of IgG+ ASCs generated by mitogen-stimulation is often used as a reference point; alternatively antigen-specific MBCs are expressed as a frequency of post-culture peripheral blood mononuclear cells (PBMC) as a surrogate for absolute frequencies. Therefore, it is important to know whether IgG+ B-cells are uniformly expanded during the preceding mitogen-culture as a true reflection of MBC frequencies ex vivo. We systematically compared B-cell phenotype and proportions before and after mitogen stimulation in cultures of 269 peripheral blood mononuclear cell samples from 62 volunteers by flow cytometry and analyzed the number of resulting ASCs. Our data show that the number of total IgG+ ASCs detected by ELISpot after mitogen stimulation correlates with the proportion of IgG+ MBCs ex vivo, highlighting its general robustness for comparisons of study cohorts at group level. The expansion of total and IgG+ B-cells during mitogen-stimulation, however, was not identical in all cultures, but influenced by size and composition of the ex vivo B-cell compartment. The uncorrected readout of antigen-specific MBCs per million post-culture PBMCs therefore only preserves the quality, but not the magnitude of differences in the ex vivo MBC response between groups or time points, particularly when comparing samples where the B-cell compartment substantially differs between cohorts or over time. Therefore, expressing antigen-specific cells per total IgG+ ASCs is currently the best measure to correct for mitogen-culture effects. Additionally, baseline information on the size and composition of the ex vivo B-cell compartment should be supplied to additionally inform about differences or changes in the size and

  16. Expansion of IgG+ B-cells during mitogen stimulation for memory B-cell ELISpot analysis is influenced by size and composition of the B-cell pool.

    Directory of Open Access Journals (Sweden)

    Anja Scholzen

    Full Text Available The memory B-cell (MBC ELISpot assay is the main technique used to measure antigen-specific MBCs as a readout of humoral immune memory. This assay relies on the ability of MBCs to differentiate into antibody-secreting cells (ASC upon polyclonal stimulation. The total number of IgG+ ASCs generated by mitogen-stimulation is often used as a reference point; alternatively antigen-specific MBCs are expressed as a frequency of post-culture peripheral blood mononuclear cells (PBMC as a surrogate for absolute frequencies. Therefore, it is important to know whether IgG+ B-cells are uniformly expanded during the preceding mitogen-culture as a true reflection of MBC frequencies ex vivo. We systematically compared B-cell phenotype and proportions before and after mitogen stimulation in cultures of 269 peripheral blood mononuclear cell samples from 62 volunteers by flow cytometry and analyzed the number of resulting ASCs. Our data show that the number of total IgG+ ASCs detected by ELISpot after mitogen stimulation correlates with the proportion of IgG+ MBCs ex vivo, highlighting its general robustness for comparisons of study cohorts at group level. The expansion of total and IgG+ B-cells during mitogen-stimulation, however, was not identical in all cultures, but influenced by size and composition of the ex vivo B-cell compartment. The uncorrected readout of antigen-specific MBCs per million post-culture PBMCs therefore only preserves the quality, but not the magnitude of differences in the ex vivo MBC response between groups or time points, particularly when comparing samples where the B-cell compartment substantially differs between cohorts or over time. Therefore, expressing antigen-specific cells per total IgG+ ASCs is currently the best measure to correct for mitogen-culture effects. Additionally, baseline information on the size and composition of the ex vivo B-cell compartment should be supplied to additionally inform about differences or changes in

  17. Surface expression of Mo3e antigen by activated human monocytes and U-937 cells

    Energy Technology Data Exchange (ETDEWEB)

    Todd R.F. III; Bury, M.J.; Liu, D.Y.

    1986-03-05

    The surface expression of a protease-sensitive antigen, Mo3e, by activated human monocytes and U-937 cells is a plasma membrane feature of the activated state. Mo3e, which is an 80 kD protein on Western blot analysis, may represent the surface receptor for migration inhibitory factor (MIF), as evidenced by inhibition of MIF responsiveness produced by anti-Mo3e monoclonal antibody. Mo3e is barely detectable (by surface immunofluorescence) on freshly isolated monocytes but becomes expressed in high antigen density during 18-24 hrs culture in medium containing E. coli lipopolysaccharide (> 1 ng/ml), 4..beta..-phorbol 12-myristate 13-acetate (PMA) (5-10 nM), or muramyl dipeptide (0.1-1 ..mu..M). In U-937 cells, Mo3e surface expression is detectable after 24 hrs exposure to PMA and other pharmacological activators of protein kinase C: 4..beta..-phorbol 12, 13 dibutyrate, 4..beta..-phorbol 12, 13 didecanoate, mezerein, or Sn-1,2-dioctanoylglycerol. The biologically-inactivate phorbol compounds, 4..cap alpha..-phorbol 12, 13 didecanoate and 4/sub ..beta../-phorbol do not stimulate Mo3e expression. The calcium ionophore, ionomycin, has a synergistic effect on Mo3e expression stimulated by PMA; conversely, calcium antagonists block PMA-induced Mo3e expression. These results suggest the involvement of protein kinase C activation and intracellular calcium mobilization in the stimulated expression of Mo3e by activated human mononuclear phagocytes.

  18. Naive T lymphocytes traffic to inflamed central nervous system, but require antigen recognition for activation

    DEFF Research Database (Denmark)

    Krakowski, M L; Owens, T

    2000-01-01

    Organ-specific autoimmune diseases may be induced by infiltration of the target tissue by CD4(+) T cells with specificity for self antigen(s). As disease progresses, T cells of other specificities appear in the tissue. Traffic of naive, antigen-inexperienced T cells to target tissues has not been...

  19. Influence of circulating antigen on blood pool activity of a radioiodinated monoclonal antibody

    International Nuclear Information System (INIS)

    Athymic mice with and without circulating CA 125 antigen were injected with 0.1-100 μg of 131I-labeled OC 125 F(ab')2 antibody fragment. Both the blood clearance of 131I activity and the change in serum CA 125 were monitored over 24 h. Influence of CA 125 on blood pool activity could be avoided only at the 100 μg dose. In patient studies, circulating CA 125 levels decreased for the first 2 h after injection of OC 125 F(ab')2 but generally returned to preinjection levels shortly thereafter. In vitro binding studies using the sera from patients injected with 131I-labeled OC 125 F(ab')2 suggest that circulating CA 125 could interfere with the tumor uptake of the labeled antibody. (author)

  20. Vitamin D controls T cell antigen receptor signaling and activation of human T cells

    DEFF Research Database (Denmark)

    von Essen, Marina Rode; Kongsbak, Martin; Schjerling, Peter;

    2010-01-01

    Phospholipase C (PLC) isozymes are key signaling proteins downstream of many extracellular stimuli. Here we show that naive human T cells had very low expression of PLC-gamma1 and that this correlated with low T cell antigen receptor (TCR) responsiveness in naive T cells. However, TCR triggering...... led to an upregulation of approximately 75-fold in PLC-gamma1 expression, which correlated with greater TCR responsiveness. Induction of PLC-gamma1 was dependent on vitamin D and expression of the vitamin D receptor (VDR). Naive T cells did not express VDR, but VDR expression was induced by TCR...... signaling via the alternative mitogen-activated protein kinase p38 pathway. Thus, initial TCR signaling via p38 leads to successive induction of VDR and PLC-gamma1, which are required for subsequent classical TCR signaling and T cell activation....

  1. Systemic Sclerosis Patients Present Alterations in the Expression of Molecules Involved in B-Cell Regulation

    Science.gov (United States)

    Soto, Lilian; Ferrier, Ashley; Aravena, Octavio; Fonseca, Elianet; Berendsen, Jorge; Biere, Andrea; Bueno, Daniel; Ramos, Verónica; Aguillón, Juan Carlos; Catalán, Diego

    2015-01-01

    The activation threshold of B cells is tightly regulated by an array of inhibitory and activator receptors in such a way that disturbances in their expression can lead to the appearance of autoimmunity. The aim of this study was to evaluate the expression of activating and inhibitory molecules involved in the modulation of B cell functions in transitional, naive, and memory B-cell subpopulations from systemic sclerosis patients. To achieve this, blood samples were drawn from 31 systemic sclerosis patients and 53 healthy individuals. Surface expression of CD86, MHC II, CD19, CD21, CD40, CD22, Siglec 10, CD35, and FcγRIIB was determined by flow cytometry. IL-10 production was evaluated by intracellular flow cytometry from isolated B cells. Soluble IL-6 and IL-10 levels were measured by ELISA from supernatants of stimulated B cells. Systemic sclerosis patients exhibit an increased frequency of transitional and naive B cells related to memory B cells compared with healthy controls. Transitional and naive B cells from patients express higher levels of CD86 and FcγRIIB than healthy donors. Also, B cells from patients show high expression of CD19 and CD40, whereas memory cells from systemic sclerosis patients show reduced expression of CD35. CD19 and CD35 expression levels associate with different autoantibody profiles. IL-10+ B cells and secreted levels of IL-10 were markedly reduced in patients. In conclusion, systemic sclerosis patients show alterations in the expression of molecules involved in B-cell regulation. These abnormalities may be determinant in the B-cell hyperactivation observed in systemic sclerosis. PMID:26483788

  2. Adhesion of Human B Cells to Germinal Centers in Vitro Involves VLA-4 and INCAM-110

    Science.gov (United States)

    Freedman, Arnold S.; Munro, J. Michael; Rice, G. Edgar; Bevilacqua, Michael P.; Morimoto, Chikao; McIntyre, Bradley W.; Rhynhart, Kurt; Pober, Jordan S.; Nadler, Lee M.

    1990-08-01

    Human B lymphocytes localize and differentiate within the microenvironment of lymphoid germinal centers. A frozen section binding assay was developed for the identification of those molecules involved in the adhesive interactions between B cells and lymphoid follicles. Activated human B cells and B cell lines were found to selectively adhere to germinal centers. The VLA-4 molecule on the lymphocyte and the adhesion molecule INCAM-110, expressed on follicular dendritic cells, supported this interaction. This cellular interaction model can be used for the study of how B cells differentiate.

  3. A novel approach for reliable detection of cathepsin S activities in mouse antigen presenting cells.

    Science.gov (United States)

    Steimle, Alex; Kalbacher, Hubert; Maurer, Andreas; Beifuss, Brigitte; Bender, Annika; Schäfer, Andrea; Müller, Ricarda; Autenrieth, Ingo B; Frick, Julia-Stefanie

    2016-05-01

    Cathepsin S (CTSS) is a eukaryotic protease mostly expressed in professional antigen presenting cells (APCs). Since CTSS activity regulation plays a role in the pathogenesis of various autoimmune diseases like multiple sclerosis, atherosclerosis, Sjögren's syndrome and psoriasis as well as in cancer progression, there is an ongoing interest in the reliable detection of cathepsin S activity. Various applications have been invented for specific detection of this enzyme. However, most of them have only been shown to be suitable for human samples, do not deliver quantitative results or the experimental procedure requires technical equipment that is not commonly available in a standard laboratory. We have tested a fluorogen substrate, Mca-GRWPPMGLPWE-Lys(Dnp)-DArg-NH2, that has been described to specifically detect CTSS activities in human APCs for its potential use for mouse samples. We have modified the protocol and thereby offer a cheap, easy, reproducible and quick activity assay to detect CTSS activities in mouse APCs. Since most of basic research on CTSS is performed in mice, this method closes a gap and offers a possibility for reliable and quantitative CTSS activity detection that can be performed in almost every laboratory. PMID:26899824

  4. Characterization of four functionally distinct human B-cell subsets that are defined by the expression of CD21 and CD86

    OpenAIRE

    García Márquez, María Alejandra

    2015-01-01

    It is now recognized that B lymphocytes are phenotypically and functionally heterogeneous. In addition to their essential role as antibody producing B cells, they serve as antigen-presenting cells, contribute to immunoregulation and represent an important source of cytokines and chemokines. Although several subpopulations have been identified, a deeper understanding of the role of B cells in the pathophysiology of human diseases has been hampered by the lack of well-defined functional B-cell ...

  5. B-cell waste classification sampling plan

    Energy Technology Data Exchange (ETDEWEB)

    HOBART, R.L.

    1999-11-20

    This report documents the methods used to collect samples and analyze data necessary to verify and/or determine the radionuclide content of the 324 Facility B-Cell decontamination and decommissioning waste stream.

  6. B-cell waste classification sampling plan

    International Nuclear Information System (INIS)

    This report documents the methods used to collect samples and analyze data necessary to verify and/or determine the radionuclide content of the 324 Facility B-Cell decontamination and decommissioning waste stream

  7. MicroRNAs in B-cells

    DEFF Research Database (Denmark)

    Marques, Sara Correia; Laursen, Maria Bach; Bødker, Julie Støve; Kjeldsen, Malene Krag; Larsen, Steffen Falgreen; Schmitz, Alexander; Bøgsted, Martin; Johnsen, Hans Erik; Dybkær, Karen

    2015-01-01

    transformation due to their function as oncogenes or tumor suppressors. We know that miRNAs are involved in the development of normal B-cells and that different B-cell subsets express specific miRNA profiles according to their degree of differentiation. B-cell-derived malignancies contain transcription......MicroRNAs (miRNAs) are small non-coding RNAs that play important post-transcriptional regulatory roles in a wide range of biological processes. They are fundamental to the normal development of cells, and evidence suggests that the deregulation of specific miRNAs is involved in malignant...... signatures reminiscent of their cell of origin. Therefore, we believe that normal and malignant B-cells share features of regulatory networks controlling differentiation and the ability to respond to treatment. The involvement of miRNAs in these processes makes them good biomarker candidates. B...

  8. Farnesyl transferase inhibitors induce extended remissions in transgenic mice with mature B cell lymphomas

    Directory of Open Access Journals (Sweden)

    Refaeli Yosef

    2008-05-01

    Full Text Available Abstract Background We have used a mouse model based on overexpression of c-Myc in B cells genetically engineered to be self-reactive to test the hypothesis that farnesyl transferase inhibitors (FTIs can effectively treat mature B cell lymphomas. FTIs are undergoing clinical trials to treat both lymphoid and non-lymphoid malignancies and we wished to obtain evidence to support the inclusion of B cell lymphomas in future trials. Results We report that two FTIs, L-744,832 and SCH66336, blocked the growth of mature B cell lymphoma cells in vitro and in vivo. The FTI treatment affected the proliferation and survival of the transformed B cells to a greater extent than naïve B cells stimulated with antigen. In syngeneic mice transplanted with the transgenic lymphoma cells, L-744,832 treatment prevented the growth of the tumor cells and the morbidity associated with the resulting lymphoma progression. Tumors that arose from transplantation of the lymphoma cells regressed with as little as three days of treatment with L-744,832 or SCH66336. Treatment of these established lymphomas with L-744,832 for seven days led to long-term remission of the disease in approximately 25% of animals. Conclusion FTI treatment can block the proliferation and survival of self-reactive transformed B cells that overexpress Myc. In mice transplanted with mature B cell lymphomas, we found that FTI treatment led to regression of disease. FTIs warrant further consideration as therapeutic agents for mature B cell lymphomas and other lymphoid tumors.

  9. A small molecule inhibitor for ATPase activity of Hsp70 and Hsc70 enhances the immune response to protein antigens

    Science.gov (United States)

    Baek, Kyung-Hwa; Zhang, Haiying; Lee, Bo Ryeong; Kwon, Young-Guen; Ha, Sang-Jun; Shin, Injae

    2015-12-01

    The ATPase activities of Hsp70 and Hsc70 are known to be responsible for regulation of various biological processes. However, little is known about the roles of Hsp70 and Hsc70 in modulation of immune responses to antigens. In the present study, we investigated the effect of apoptozole (Az), a small molecule inhibitor of Hsp70 and Hsc70, on immune responses to protein antigens. The results show that mice administered with both protein antigen and Az produce more antibodies than those treated with antigen alone, showing that Az enhances immune responses to administered antigens. Treatment of mice with Az elicits production of antibodies with a high IgG2c/IgG1 ratio and stimulates the release of Th1 and Th2-type cytokines, suggesting that Az activates the Th1 and Th2 immune responses. The observations made in the present study suggest that inhibition of Hsp70 and Hsc70 activities could be a novel strategy designing small molecule-based adjuvants in protein vaccines.

  10. Myelin Basic Protein-Induced Production of Tumor Necrosis Factor-α and Interleukin-6, and Presentation of the Immunodominant Peptide MBP85-99 by B Cells from Patients with Relapsing-Remitting Multiple Sclerosis.

    Science.gov (United States)

    Nielsen, Claus H; Börnsen, Lars; Sellebjerg, Finn; Brimnes, Marie K

    2016-01-01

    B cells are involved in driving relapsing-remitting multiple sclerosis (RRMS), as demonstrated by the positive effect of therapeutic B-cell depletion. Aside from producing antibodies, B cells are efficient antigen-presenting and cytokine-secreting cells. Diverse polyclonal stimuli have been used to study cytokine production by B cells, but here we used the physiologically relevant self-antigen myelin basic protein (MBP) to stimulate B cells from untreated patients with RRMS and healthy donors. Moreover, we took advantage of the unique ability of the monoclonal antibody MK16 to recognize the immunodominant peptide MBP85-99 presented on HLA-DR15, and used it as a probe to directly study B-cell presentation of self-antigenic peptide. The proportions of B cells producing TNF-α or IL-6 after stimulation with MBP were higher in RRMS patients than in healthy donors, indicating a pro-inflammatory profile for self-reactive patient B cells. In contrast, polyclonal stimulation with PMA + ionomycin and MBP revealed no difference in cytokine profile between B cells from RRMS patients and healthy donors. Expanded disability status scale (EDSS) as well as multiple sclerosis severity score (MSSS) correlated with reduced ability of B cells to produce IL-10 after stimulation with MBP, indicative of diminished B-cell immune regulatory function in patients with the most severe disease. Moreover, EDSS correlated positively with the frequencies of TNF-α, IL-6 and IL-10 producing B cells after polyclonal stimulation. Patient-derived, IL-10-producing B cells presented MBP85-99 poorly, as did IL-6-producing B cells, particulary in the healthy donor group. B cells from MS patients thus present antigen to T cells in a pro-inflammatory context. These findings contribute to understanding the therapeutic effects of B-cell depletion in human autoimmune diseases, including MS. PMID:26756931

  11. Strategies for B-cell receptor repertoire analysis in Primary Immunodeficiencies:From severe combined immunodeficiency to common variable immunodeficiency

    Directory of Open Access Journals (Sweden)

    Hanna eIJspeert

    2015-04-01

    Full Text Available The antigen receptor repertoires of B and T cells form the basis of the adaptive immune response. The repertoires should be sufficiently diverse to recognize all possible pathogens. However, careful selection is needed to prevent responses to self or harmless antigens. Limited antigen receptor repertoire diversity leads to immunodeficiency, whereas unselected or misdirected repertoires can result in autoimmunity. The antigen receptor repertoire harbors information about abnormalities in many immunological disorders. Recent developments in next generation sequencing allow the analysis of the antigen receptor repertoire in much greater detail than ever before. Analyzing the antigen receptor repertoire in patients with mutations in genes responsible for the generation of the antigen receptor repertoire will give new insights into repertoire formation and selection. In this perspective we describe strategies and considerations for analysis of the naive and antigen selected B-cell repertoires in primary immunodeficiency (PID patients with a focus on severe combined immunodeficiency (SCID and common variable immunodeficiency (CVID.

  12. Role of recombination activating genes in the generation of antigen receptor diversity and beyond.

    Science.gov (United States)

    Nishana, Mayilaadumveettil; Raghavan, Sathees C

    2012-12-01

    V(D)J recombination is the process by which antibody and T-cell receptor diversity is attained. During this process, antigen receptor gene segments are cleaved and rejoined by non-homologous DNA end joining for the generation of combinatorial diversity. The major players of the initial process of cleavage are the proteins known as RAG1 (recombination activating gene 1) and RAG2. In this review, we discuss the physiological function of RAGs as a sequence-specific nuclease and its pathological role as a structure-specific nuclease. The first part of the review discusses the basic mechanism of V(D)J recombination, and the last part focuses on how the RAG complex functions as a sequence-specific and structure-specific nuclease. It also deals with the off-target cleavage of RAGs and its implications in genomic instability. PMID:23039142

  13. Complexes between nuclear factor-κB p65 and signal transducer and activator of transcription 3 are key actors in inducing activation-induced cytidine deaminase expression and immunoglobulin A production in CD40L plus interleukin-10-treated human blood B cells.

    Science.gov (United States)

    Lafarge, S; Hamzeh-Cognasse, H; Richard, Y; Pozzetto, B; Cogné, M; Cognasse, F; Garraud, O

    2011-11-01

    The signal transducer and activator of transcription 3 (STAT3) transcription factor pathway plays an important role in many biological phenomena. STAT3 transcription is triggered by cytokine-associated signals. Here, we use isolated human B cells to analyse the role of STAT3 in interleukin (IL)-10 induced terminal B cell differentiation and in immunoglobulin (Ig)A production as a characteristic readout of IL-10 signalling. We identified optimal conditions for inducing in-vitro IgA production by purified blood naive B cells using IL-10 and soluble CD40L. We show that soluble CD40L consistently induces the phosphorylation of nuclear factor (NF)-κB p65 but not of STAT3, while IL-10 induces the phosphorylation of STAT3 but not of NF-κB p65. Interestingly, while soluble CD40L and IL-10 were synergistic in driving the terminal maturation of B cells into IgA-producing plasma cells, they did not co-operate earlier in the pathway with regard to the transcription factors NF-κB p65 or STAT3. Blocking either NF-κB p65 or STAT3 profoundly altered the production of IgA and mRNA for activation-induced cytidine deaminase (AID), an enzyme strictly necessary for Ig heavy chain recombination. Finally, the STAT3 pathway was directly activated by IL-10, while IL-6, the main cytokine otherwise known for activating the STAT3 pathway, did not appear to be involved in IL-10-induced-STAT3 activation. Our results suggest that STAT3 and NF-κB pathways co-operate in IgA production, with soluble CD40L rapidly activating the NF-κB pathway, probably rendering STAT3 probably more reactive to IL-10 signalling. This novel role for STAT3 in B cell development reveals a potential therapeutic or vaccine target for eliciting IgA humoral responses at mucosal interfaces. PMID:21985363

  14. Transformation and radiosensitivity of human diploid skin fibroblasts transfected with activated RAS oncogene and SV40 T-antigen

    Energy Technology Data Exchange (ETDEWEB)

    Su, L.-N.; Little, J.B. (Harvard School of Public Health, Boston, MA (United States))

    1992-08-01

    Three normal human diploid cell strains were transfected with an activated Ha-ras oncogene (EJ ras) or SV40 T-antigen. Multiple clones were examined for morphological alterations, growth requirements, ability to grow under anchorage independent conditions, immortality and tumorigenicity in nude mice. Clones expressing SV40 T-antigen alone or in combination with ras protein p21 were significantly radioresistant as compared with their parent cells or clones transfected with the neo gene only. This radioresistant phenotype persisted in post-crisis, immortalized cell lines. These data suggest that expression of the SV40 T-antigen but not activated Ha-ras plays an important role in the radiosensitivity of human diploid cells. The radioresistant phenotype in SV40 T transfected cells was not related to the enhanced level of genetic instability seen in pre-crisis and newly immortalized cells, nor to the process of immortalization itself. (author).

  15. Identification of uniquely expressed transcription factors in highly purified B-cell lymphoma samples

    DEFF Research Database (Denmark)

    Andréasson, Ulrika; Edén, Patrik; Peterson, Carsten; Högerkorp, Carl-Magnus; Jerkeman, Mats; Andersen, Niels Smedegaard; Berglund, Mattias; Sundström, Christer; Rosenquist, Richard; Borrebaeck, Carl A K; Ek, Sara

    2010-01-01

    Transcription factors (TFs) are critical for B-cell differentiation, affecting gene expression both by repression and transcriptional activation. Still, this information is not used for classification of B-cell lymphomas (BCLs). Traditionally, BCLs are diagnosed based on a phenotypic resemblance ...

  16. A new role for complement C3: regulation of antigen processing through an inhibitory activity.

    OpenAIRE

    Villiers, Christian; Cretin, François,; Lefebvre, Nicole; Marche, Patrice; Villiers, Marie-Bernadette

    2008-01-01

    International audience Increasing evidence underlines the involvement of complement component C3 in the establishment of acquired immunity which appears to play a complex role and to act at different levels. As antigen proteolysis by antigen presenting cells is a key event in the control of antigen presentation efficiency, and consequently in the quality of the immune response, we investigated whether C3 could modulate this step. Our results demonstrate for the first time that C3 can inter...

  17. Individualized leukemia cell-population profiles in common B-cell acute lymphoblastic leukemia patients

    OpenAIRE

    Xian-Ming Mo; Hong Xu; Ting-Ting Zeng; Neng-Gang Jiang; Yong-Qian Jia; Jing-Tao Dong; Jian-Hua Yu; Wen-Tong Meng

    2013-01-01

    Immunophenotype is critical for diagnosing common B-cell acute lymphoblastic leukemia (common ALL) and detecting minimal residual disease. We developed a protocol to explore the immunophenotypic profiles of common ALL based on the expression levels of the antigens associated with B lymphoid development, including IL-7R alpha (CD127), cytoplasmic CD79a (cCD79a), CD19, VpreB (CD179a), and sIgM, which are successive and essential for progression of B cells along their developmental pathway. Anal...

  18. T cell activation. II. Activation of human T lymphoma cells by cross-linking of their MHC class I antigens

    DEFF Research Database (Denmark)

    Dissing, S; Geisler, C; Rubin, B; Plesner, T; Claesson, M H

    1990-01-01

    The present work demonstrates that antibody-induced cross-linking of MHC class I antigens on Jurkat T lymphoma cells leads to a rise in intracellular calcium (Cai2+) and, in the presence of phorbol ester (PMA), to IL-2 production and IL-2 receptor expression. The rise in Cai2+ exhibited a profile...... very different from that obtained after anti-CD3 antibody-induced activation suggesting that activation signals are transduced differently after binding of anti-CD3 antibody and class I cross-linking, respectively. However, when Cai2+ was examined in individual Jurkat cells by means of a digital image...... the T cell receptor complex and MHC class I molecules....

  19. CD85/LIR-1/ILT2 and CD152 (cytotoxic T lymphocyte antigen 4) inhibitory molecules down-regulate the cytolytic activity of human CD4+ T-cell clones specific for Mycobacterium tuberculosis.

    Science.gov (United States)

    Merlo, A; Saverino, D; Tenca, C; Grossi, C E; Bruno, S; Ciccone, E

    2001-10-01

    Antigen-specific cytolytic CD4+ T lymphocytes control Mycobacterium tuberculosis infection by secreting cytokines and by killing macrophages that have phagocytosed the pathogen. However, lysis of the latter cells promotes microbial dissemination, and other macrophages engulf the released bacteria. Subsequently, CD4+ T-cell-mediated killing of macrophages goes on, and this persistent process may hamper control of infection, unless regulatory mechanisms maintain a subtle balance between lysis of macrophages by cytolytic CD4+ cells and activation of cytolytic CD4+ cells by infected macrophages. We asked whether inhibitory molecules expressed by CD4+ cytolytic T lymphocytes could play a role in such a balance. To this end, human CD4+ T-cell clones specific for M. tuberculosis were produced that displayed an autologous major histocompatibility complex class II-restricted lytic ability against purified protein derivative (PPD)-pulsed antigen-presenting cells. All T-cell clones expressed CD152 (cytotoxic T-lymphocyte antigen 4 [CTLA-4]) and CD85/leukocyte immunoglobulin-like receptor 1 (LIR-1)/immunoglobulin-like transcript 2 (ILT2) inhibitory receptors, but not CD94 and the killer inhibitory receptor (or killer immunoglobulin-like receptor [KIR]) p58.2. CD3-mediated activation of the clones was inhibited in a redirected killing assay in which CD152 and CD85/LIR-1/ILT2 were cross-linked. Specific antigen-mediated proliferation of the clones was also sharply reduced when CD152 and CD85/LIR-1/ILT2 were cross-linked by specific monoclonal antibody (MAb) followed by goat anti-mouse antiserum. In contrast, blockade of the receptors by specific MAb only increased their proliferation. Production of interleukin 2 (IL-2) and gamma interferon (IFN-gamma) by the T-cell clones was also strongly reduced when CD152 and CD85/LIR-1/ILT2 were cross-linked. The lytic activity of the T-cell clones against PPD-pulsed autologous monocytes or Epstein-Barr virus-activated B cells was increased

  20. Proteolytic activity of prostate-specific antigen (PSA towards protein substrates and effect of peptides stimulating PSA activity.

    Directory of Open Access Journals (Sweden)

    Johanna M Mattsson

    Full Text Available Prostate-specific antigen (PSA or kallikrein-related peptidase-3, KLK3 exerts chymotrypsin-like proteolytic activity. The main biological function of PSA is the liquefaction of the clot formed after ejaculation by cleavage of semenogelins I and II in seminal fluid. PSA also cleaves several other substrates, which may explain its putative functions in prostate cancer and its antiangiogenic activity. We compared the proteolytic efficiency of PSA towards several protein and peptide substrates and studied the effect of peptides stimulating the activity of PSA with these substrates. An endothelial cell tube formation model was used to analyze the effect of PSA-degraded protein fragments on angiogenesis. We showed that PSA degrades semenogelins I and II much more efficiently than other previously identified protein substrates, e.g., fibronectin, galectin-3 and IGFBP-3. We identified nidogen-1 as a new substrate for PSA. Peptides B2 and C4 that stimulate the activity of PSA towards small peptide substrates also enhanced the proteolytic activity of PSA towards protein substrates. Nidogen-1, galectin-3 or their fragments produced by PSA did not have any effect on endothelial cell tube formation. Although PSA cleaves several other protein substrates, in addition to semenogelins, the physiological importance of this activity remains speculative. The PSA levels in prostate are very high, but several other highly active proteases, such as hK2 and trypsin, are also expressed in the prostate and may cleave protein substrates that are weakly cleaved by PSA.

  1. Clinical and pharmacologic aspects of blinatumomab in the treatment of B-cell acute lymphoblastic leukemia

    Directory of Open Access Journals (Sweden)

    Portell CA

    2013-04-01

    Full Text Available Craig A Portell, Candice M Wenzell, Anjali S Advani Leukemia Program, Taussig Cancer Institute, Cleveland Clinic, Cleveland, OH, USA Abstract: Acute lymphoblastic leukemia (ALL in adults remains a challenging disease to treat, and novel therapies are needed. Precursor-B ALL comprises 80% of cases, and the CD19 antigen is expressed in nearly all precursor-B ALL patients. Bispecific T-cell-engaging antibodies are novel bioengineered proteins. The bispecific T-cell-engaging antibody blinatumomab engages polyclonal T cells to CD19-expressing B cells. By binding to both CD3 and CD19, blinatumomab physically brings these T cells in close proximity to malignant B cells and potentiates T-cell-induced cytotoxic cell kill. Blinatumomab requires continuous intravenous infusion due to its short half-life, the need for continuous exposure for the drug to exert sufficient efficacy, and lessened toxicity. A phase II trial of B-cell ALL patients with persistent or relapsed minimal residual disease demonstrated an 80% rate of complete molecular remission. Cytokine-release syndrome and central nervous system events, such as seizures and encephalopathy, are reversible toxicities. Promising results in B-cell ALL with minimal residual disease have led to further evaluation of this drug in newly diagnosed and relapsed B-cell ALL. Keywords: blinatumomab, B-cell acute lymphoblastic leukemia, CD19, BiTE antibodies

  2. Engineered nanomaterials cause cytotoxicity and activation on mouse antigen presenting cells

    International Nuclear Information System (INIS)

    Nanomaterials improve everyday products but their safety for human health is poorly known. In this study we explored immunological effects of five different nanomaterials on antigen presenting cells (APC) in vitro. Nanomaterials studied were rutile titanium dioxide (TiO2), amorphous silica-coated rutile titanium dioxide (TiO2-silica), zinc oxide (ZnO), single-walled carbon nanotubes (SWCNT) and multi-walled carbon nanotubes (MWCNT). APCs included mouse macrophages (RAW 264.7 cell line) and murine bone marrow-derived dendritic cells (bmDC). All studied particles were cytotoxic to bmDCs, and ZnO, TiO2 and TiO2-silica-induced dose-dependently cell death also in macrophages. ZnO had the most drastic immunological effects leading to high expression of proinflammatory cytokine, IL-1β, and enhanced production of neutrophil chemoattractant CXCL-9 on both cell types. TiO2 and TiO2-silica stimulated the expression of IL-6, MIP-1α and TNF-α in macrophages, and increased their maturation, antigen presentation and co-stimulation activity. In contrast, SWCNT or MWCNT did not seem to have any significant immunological effects on the cell types studied suggesting that APCs might not be the target cells for carbon nanotubes. Due to diverse effects on different nanomaterials on immune cells we suggest that each new nanomaterial should be extensively studied in vitro and in vivo for risk assessment before their use in final products.

  3. Enforced expression of the transcriptional coactivator OBF1 impairs B cell differentiation at the earliest stage of development.

    Directory of Open Access Journals (Sweden)

    Alain Bordon

    Full Text Available OBF1, also known as Bob.1 or OCA-B, is a B lymphocyte-specific transcription factor which coactivates Oct1 and Oct2 on B cell specific promoters. So far, the function of OBF1 has been mainly identified in late stage B cell populations. The central defect of OBF1 deficient mice is a severely reduced immune response to T cell-dependent antigens and a lack of germinal center formation in the spleen. Relatively little is known about a potential function of OBF1 in developing B cells. Here we have generated transgenic mice overexpressing OBF1 in B cells under the control of the immunoglobulin heavy chain promoter and enhancer. Surprisingly, these mice have greatly reduced numbers of follicular B cells in the periphery and have a compromised immune response. Furthermore, B cell differentiation is impaired at an early stage in the bone marrow: a first block is observed during B cell commitment and a second differentiation block is seen at the large preB2 cell stage. The cells that succeed to escape the block and to differentiate into mature B cells have post-translationally downregulated the expression of transgene, indicating that expression of OBF1 beyond the normal level early in B cell development is deleterious. Transcriptome analysis identified genes deregulated in these mice and Id2 and Id3, two known negative regulators of B cell differentiation, were found to be upregulated in the EPLM and preB cells of the transgenic mice. Furthermore, the Id2 and Id3 promoters contain octamer-like sites, to which OBF1 can bind. These results provide evidence that tight regulation of OBF1 expression in early B cells is essential to allow efficient B lymphocyte differentiation.

  4. Immunotherapy for B-cell lymphoma: current status and prospective advances.

    Science.gov (United States)

    Hollander, Nurit

    2012-01-01

    Therapy for non-Hodgkin's lymphoma has progressed significantly over the last decades. However, the majority of patients remain incurable, and novel therapies are needed. Because immunotherapy ideally offers target selectivity, an ever increasing number of immunotherapies, both passive and active, are undergoing development. The champion of passive immunotherapy to date is the anti-CD20 monoclonal antibody rituximab that revolutionized the standard of care for lymphoma. The great success of rituximab catalyzed the development of new passive immunotherapy strategies that are currently undergoing clinical evaluation. These include improvement of rituximab efficacy, newer generation anti-CD20 antibodies, drug-conjugated and radio labeled anti-CD20 antibodies, monoclonal antibodies targeting non-CD20 lymphoma antigens, and bispecific antibodies. Active immunotherapy aims at inducing long-lasting antitumor immunity, thereby limiting the likelihood of relapse. Current clinical studies of active immunotherapy for lymphoma consist largely of vaccination and immune checkpoint blockade. A variety of protein- and cell-based vaccines are being tested in ongoing clinical studies. Recently completed phase III clinical trials of an idiotype protein vaccine suggest that the vaccine may have clinical activity in a subset of patients. Efforts to enhance the efficacy of active immunotherapy are ongoing with an emphasis on optimization of antigen delivery and presentation of vaccines and modulation of the immune system toward counteracting immunosuppression, using antibodies against immune regulatory checkpoints. This article discusses results of the various immunotherapy approaches applied to date for B-cell lymphoma and the ongoing trials to improve their effect. PMID:22566889

  5. Immunotherapy for B-cell lymphoma: current status and prospective advances

    Directory of Open Access Journals (Sweden)

    Nurit eHollander

    2012-01-01

    Full Text Available Therapy for non-Hodgkin's lymphoma has progressed significantly over the last decades. However, the majority of patients remain incurable, and novel therapies are needed. Because immunotherapy ideally offers target selectivity, an ever increasing number of immunotherapies, both passive and active, are undergoing development. The champion of passive immunotherapy to date is the anti-CD20 monoclonal antibody rituximab that revolutionized the standard of care for lymphoma. The great success of rituximab catalyzed the development of new passive immunotherapy strategies that are currently undergoing clinical evaluation. These include improvement of rituximab efficacy, newer generation anti-CD20 antibodies, drug-conjugated and radiolabelled anti-CD20 antibodies, monoclonal antibodies targeting non-CD20 lymphoma antigens, and bispecific antibodies. Active immunotherapy aims at inducing long-lasting antitumor immunity, thereby limiting the likelihood of relapse. Current clinical studies of active immunotherapy for lymphoma consist largely of vaccination and immune checkpoint blockade. A variety of protein- and cell-based vaccines are being tested in ongoing clinical studies. Recently completed phase III clinical trials of an idiotype protein vaccine suggest that the vaccine may have clinical activity in a subset of patients. Efforts to enhance the efficacy of active immunotherapy are ongoing with an emphasis on optimization of antigen delivery and presentation of vaccines and modulation of the immune system toward counteracting immunosuppression, using antibodies against immune regulatory checkpoints. This article discusses results of the various immunotherapy approaches applied to date for B-cell lymphoma and the ongoing trials to improve their effect.

  6. Longitudinal microarray analysis of cell surface antigens on peripheral blood mononuclear cells from HIV+ individuals on highly active antiretroviral therapy

    Directory of Open Access Journals (Sweden)

    Wang Bin

    2008-03-01

    Full Text Available Abstract Background The efficacy of highly active antiretroviral therapy (HAART determined by simultaneous monitoring over 100 cell-surface antigens overtime has not been attempted. We used an antibody microarray to analyze changes in the expression of 135 different cell-surface antigens overtime on PBMC from HIV+ patients on HAART. Two groups were chosen, one (n = 6 achieved sustainable response by maintaining below detectable plasma viremia and the other (n = 6 responded intermittently. Blood samples were collected over an average of 3 years and 5–8 time points were selected for microarray assay and statistical analysis. Results Significant trends over time were observed for the expression of 7 cell surface antigens (CD2, CD3epsilon, CD5, CD95, CD36, CD27 and CD28 for combined patient groups. Between groups, expression levels of 10 cell surface antigens (CD11a, CD29, CD38, CD45RO, CD52, CD56, CD57, CD62E, CD64 and CD33 were found to be differential. Expression levels of CD9, CD11a, CD27, CD28 and CD52, CD44, CD49d, CD49e, CD11c strongly correlated with CD4+ and CD8+ T cell counts, respectively. Conclusion Our findings not only detected markers that may have potential prognostic/diagnostic values in evaluating HAART efficacy, but also showed how density of cell surface antigens could be efficiently exploited in an array-like manner in relation to HAART and HIV-infection. The antigens identified in this study should be further investigated by other methods such as flow cytometry for confirmation as biological analysis of these antigens may help further clarify their role during HAART and HIV infection.

  7. Soluble Expression and Characterization of Biologically Active Bacillus anthracis Protective Antigen in Escherichia coli

    Directory of Open Access Journals (Sweden)

    Nagendra Suryanarayana

    2016-01-01

    Full Text Available Bacillus anthracis secretory protein protective antigen (PA is primary candidate for subunit vaccine against anthrax. Attempts to obtain large quantity of PA from Escherichia coli expression system often result in the formation of insoluble inclusion bodies. Therefore, it is always better to produce recombinant proteins in a soluble form. In the present study, we have obtained biologically active recombinant PA in small scale E. coli shake culture system using three different expression constructs. The PA gene was cloned in expression vectors bearing trc, T5, and T7 promoters and transformed into their respective E. coli hosts. The growth conditions were optimized to obtain maximum expression of PA in soluble form. The expression construct PA-pET32c in DE3-pLysS E. coli host resulted in a maximum production of soluble PA (15 mg L−1 compared to other combinations. Purified PA was subjected to trypsin digestion and binding assay with lethal factor to confirm the protein’s functionality. Biological activity was confirmed by cytotoxicity assay on J774.1 cells. Balb/c mice were immunized with PA and the immunogenicity was tested by ELISA and toxin neutralization assay. This study highlights the expression of soluble and biologically active recombinant PA in larger quantity using simpler E. coli production platform.

  8. Soluble Expression and Characterization of Biologically Active Bacillus anthracis Protective Antigen in Escherichia coli.

    Science.gov (United States)

    Suryanarayana, Nagendra; Vanlalhmuaka; Mankere, Bharti; Verma, Monika; Thavachelvam, Kulanthaivel; Tuteja, Urmil

    2016-01-01

    Bacillus anthracis secretory protein protective antigen (PA) is primary candidate for subunit vaccine against anthrax. Attempts to obtain large quantity of PA from Escherichia coli expression system often result in the formation of insoluble inclusion bodies. Therefore, it is always better to produce recombinant proteins in a soluble form. In the present study, we have obtained biologically active recombinant PA in small scale E. coli shake culture system using three different expression constructs. The PA gene was cloned in expression vectors bearing trc, T5, and T7 promoters and transformed into their respective E. coli hosts. The growth conditions were optimized to obtain maximum expression of PA in soluble form. The expression construct PA-pET32c in DE3-pLysS E. coli host resulted in a maximum production of soluble PA (15 mg L(-1)) compared to other combinations. Purified PA was subjected to trypsin digestion and binding assay with lethal factor to confirm the protein's functionality. Biological activity was confirmed by cytotoxicity assay on J774.1 cells. Balb/c mice were immunized with PA and the immunogenicity was tested by ELISA and toxin neutralization assay. This study highlights the expression of soluble and biologically active recombinant PA in larger quantity using simpler E. coli production platform. PMID:26966576

  9. Functional activation of the T-cell antigen receptor induces tyrosine phosphorylation of phospholipase C-gamma 1.

    OpenAIRE

    Weiss, A; Koretzky, G; Schatzman, R C; Kadlecek, T

    1991-01-01

    Stimulation of the T-cell antigen receptor (TCR), which itself is not a protein-tyrosine kinase (PTK), activates a PTK and phospholipase C (PLC). Using the human T-cell leukemic line Jurkat and normal peripheral blood lymphocytes, we demonstrate that stimulation of the TCR specifically induces the recovery of PLC activity in eluates from anti-phosphotyrosine immunoprecipitates. Stimulation of the human muscarinic receptor, subtype 1, when expressed in Jurkat activates PLC through a guanine nu...

  10. Humans with chronic granulomatous disease maintain humoral immunologic memory despite low frequencies of circulating memory B cells

    OpenAIRE

    Moir, Susan; De Ravin, Suk See; Santich, Brian H.; Kim, Jin Young; Posada, Jacqueline G.; Ho, Jason; Buckner, Clarisa M.; Wang, Wei; Kardava, Lela; Garofalo, Mary; Marciano, Beatriz E.; Manischewitz, Jody; King, Lisa R.; Khurana, Surender; Chun, Tae-Wook

    2012-01-01

    CD27+ memory B cells are reduced in the blood of patients with chronic granulomatous disease (CGD) for reasons and consequences that remain unclear. Here we confirm not only decreased CD27+ but also IgG+ B cells in the blood of CGD patients compared with healthy donors (HDs). However, among IgG+ B cells, the ratio of CD27− to CD27+ was significantly higher in CGD patients compared with HDs. Similar to conventional memory B cells, CD27−IgG+ B cells of CGD patients expressed activation markers ...

  11. Hepatitis B virus e antigen induces activation of rat hepatic stellate cells

    Energy Technology Data Exchange (ETDEWEB)

    Zan, Yanlu [Center for Molecular Virology, CAS Key Laboratory of Pathogenic Microbiology and Immunology, Institute of Microbiology, Chinese Academy of Sciences, Beijing 100101 (China); University of Chinese Academy of Sciences, Beijing 100049 (China); Zhang, Yuxia, E-mail: yzhang@wehi.edu.au [Center for Molecular Virology, CAS Key Laboratory of Pathogenic Microbiology and Immunology, Institute of Microbiology, Chinese Academy of Sciences, Beijing 100101 (China); Tien, Po, E-mail: tienpo@sun.im.ac.cn [Center for Molecular Virology, CAS Key Laboratory of Pathogenic Microbiology and Immunology, Institute of Microbiology, Chinese Academy of Sciences, Beijing 100101 (China)

    2013-06-07

    Highlights: •HBeAg expression in HSCs induced production of ECM protein and liver fibrotic markers. •The activation and proliferation of HSCs were mediated by TGF-β. •HBeAg protein purified from cell medium directly activated HSCs. -- Abstract: Chronic hepatitis B virus infection is a major cause of hepatic fibrosis, leading to liver cirrhosis and hepatocellular carcinoma. Hepatitis B virus e antigen (HBeAg) is an accessory protein of HBV, not required for viral replication but important for natural infection in vivo. Hepatic stellate cells (HSCs) are the major producers of excessive extracellular matrix during liver fibrogenesis. Therefore, we examined the influence of HBeAg on HSCs. The rat HSC line HSC-T6 was transfected with HBeAg plasmids, and expression of α-smooth muscle actin, collagen I, transforming growth factor-β1 (TGF-β), and tissue inhibitors of metalloproteinase 1 (TIMP-1) was investigated by quantitative real-time PCR. The proliferation of HSCs was determined by MTS analysis. HBeAg transduction induced up-regulation of these fibrogenic genes and proliferation of HSCs. We found that HBeAg induced TGF-β secretion in HSCs, and the activation of HSCs was prevented by a neutralizing anti-TGF-β antibody. Depletion and addition of HBeAg protein in conditioned medium from HSC-T6 cells transduced with HBeAg indicated that HBeAg directly induced the activation and proliferation of rat primary HSCs. Taken together, HBeAg induces the activation and proliferation of HSCs, mainly mediated by TGF-β, and HBeAg protein purified from cell medium can directly activate HSCs.

  12. Hepatitis B virus e antigen induces activation of rat hepatic stellate cells

    International Nuclear Information System (INIS)

    Highlights: •HBeAg expression in HSCs induced production of ECM protein and liver fibrotic markers. •The activation and proliferation of HSCs were mediated by TGF-β. •HBeAg protein purified from cell medium directly activated HSCs. -- Abstract: Chronic hepatitis B virus infection is a major cause of hepatic fibrosis, leading to liver cirrhosis and hepatocellular carcinoma. Hepatitis B virus e antigen (HBeAg) is an accessory protein of HBV, not required for viral replication but important for natural infection in vivo. Hepatic stellate cells (HSCs) are the major producers of excessive extracellular matrix during liver fibrogenesis. Therefore, we examined the influence of HBeAg on HSCs. The rat HSC line HSC-T6 was transfected with HBeAg plasmids, and expression of α-smooth muscle actin, collagen I, transforming growth factor-β1 (TGF-β), and tissue inhibitors of metalloproteinase 1 (TIMP-1) was investigated by quantitative real-time PCR. The proliferation of HSCs was determined by MTS analysis. HBeAg transduction induced up-regulation of these fibrogenic genes and proliferation of HSCs. We found that HBeAg induced TGF-β secretion in HSCs, and the activation of HSCs was prevented by a neutralizing anti-TGF-β antibody. Depletion and addition of HBeAg protein in conditioned medium from HSC-T6 cells transduced with HBeAg indicated that HBeAg directly induced the activation and proliferation of rat primary HSCs. Taken together, HBeAg induces the activation and proliferation of HSCs, mainly mediated by TGF-β, and HBeAg protein purified from cell medium can directly activate HSCs

  13. Levels of expression of CD19 and CD20 in chronic B cell leukaemias.

    Science.gov (United States)

    Ginaldi, L; De Martinis, M; Matutes, E; Farahat, N; Morilla, R; Catovsky, D

    1998-01-01

    AIMS: To investigate whether the antigen levels of the B cell lineage markers CD19 and CD20 can distinguish between normal and neoplastic B cells or characterise distinct expression patterns among the chronic B cell leukaemias. METHODS: Peripheral blood cells from 70 patients with B cell disorders and 17 healthy donors were analysed by quantitative flow cytometry. Direct immunofluorescence staining was performed with phycoerythrin conjugated CD19 and CD20 monoclonal antibodies. Standard microbeads with different capacities to bind mouse immunoglobulins were used to convert the mean fluorescence intensity (MFI) values into number of antigen molecules/cell, expressed as antibody binding capacity (ABC). RESULTS: CD19 and CD20 ABC values in leukaemic B cells differed from those of normal blood B lymphocytes. The results identified distinct profiles of CD19 and CD20 expression in the various types of B cell leukaemias. In all leukaemias studied except hairy cell leukaemia (HCL), CD19 expression was significantly lower than the mean (SD) value in normal B cells (22 (7) x 10(3) molecules/cell), as follows: chronic lymphocytic leukaemia (CLL), 13 (7) x 10(3); B prolymphocytic leukaemia (B-PLL), 16 (9) x 10(3); splenic lymphoma with villous lymphocytes (SLVL), 15 (11) x 10(3); mantle cell lymphoma (MCL), 10 (7) x 10(3). In HCL there was strong CD19 expression (38 (16) x 10(3)). In contrast, the level of expression of membrane CD20 was higher than the mean (SD) value in normal B cells (94 (16) x 10(3) molecules/cell) in MCL (123 (51) x 10(3)); B-PLL (129 (47) x 10(3)); SLVL (167 (72) x 10(3)); and HCL (312 (110) x 10(3)); while it was significantly lower (65 (11) x 10(3)) in CLL compared with normal B cells and the other B cell leukaemias. CONCLUSIONS: Quantitative determination of CD19 and CD20 may provide useful diagnostic information for the study of B lymphoproliferative disorders. PMID:9708202

  14. Transcription Activity of Ectogenic Human Carcinoembryonic Antigen Promoter in Lung Adenocarcinoma Cells A549

    Institute of Scientific and Technical Information of China (English)

    XIONG Weining; FANG Huijuan; XU Yongjian; XIONG Shendao; CAO Yong; SONG Qingfeng; ZENG Daxiong; ZHANG Huilan

    2006-01-01

    The transcription activity of ectogenic human carcinoembryonic antigen (CEA) promoter in lung adenocarcinoma cells A549 was investigated for the further gene-targeting therapy. The reporter gene green fluorescent protein (GFP) driven by CEA promoter and human cytomegalovirus (CMV) promoter were relatively constructed and named plasmid pCEA-EGFP and pCMV-GFP respectively. The intensity of fluorescence was detected by fluorescence microscope and flow cytometry analysis after the pCEA-GFP and pSNAV-GFP plasmids were transfected into A549 cells through liposome respectively. The results showed (4.08±0.63) % of the A549 cells transfected with pCEA-AFP plasmid expressed, significantly lower than that of the A549 cells transfected with pCMV-GFP [(43.27±3.54) %]. It was suggested that ectogenic human CEA promoter in lung adenocarcinoma cells A549 was weakly expressed. The distinct specificity of CEA promoter in CEA high expression cells was regarded as a tool in selective gene therapy, but the transcription activity of ectogenic human CEA promoter was needed to increase in the future.

  15. HIV-Exposed Uninfected Infants Show Robust Memory B-Cell Responses in Spite of a Delayed Accumulation of Memory B Cells: an Observational Study in the First 2 Years of Life.

    Science.gov (United States)

    Nduati, Eunice W; Nkumama, Irene N; Gambo, Faith K; Muema, Daniel M; Knight, Miguel G; Hassan, Amin S; Jahangir, Margaret N; Etyang, Timothy J; Berkley, James A; Urban, Britta C

    2016-07-01

    Improved HIV care has led to an increase in the number of HIV-exposed uninfected (HEU) infants born to HIV-infected women. Although they are uninfected, these infants experience increased morbidity and mortality. One explanation may be that their developing immune system is altered by HIV exposure, predisposing them to increased postnatal infections. We explored the impact of HIV exposure on the B-cell compartment by determining the B-cell subset distribution, the frequency of common vaccine antigen-specific memory B cells (MBCs), and the levels of antibodies to the respective antigens in HEU and HIV-unexposed uninfected (HUU) infants born to uninfected mothers, using flow cytometry, a B-cell enzyme-linked immunosorbent spot assay, and an enzyme-linked immunosorbent assay, respectively, during the first 2 years of life. For the majority of the B-cell subsets, there were no differences between HEU and HUU infants. However, HIV exposure was associated with a lower proportion of B cells in general and MBCs in particular, largely due to a lower proportion of unswitched memory B cells. This reduction was maintained even after correcting for age. These phenotypic differences in the MBC compartment did not affect the ability of HEU infants to generate recall responses to previously encountered antigens or reduce the antigen-specific antibody levels at 18 months of life. Although HIV exposure was associated with a transient reduction in the proportion of MBCs, we found that the ability of HEU infants to mount robust MBC and serological responses was unaffected. PMID:27170641

  16. Regulation of B cell function by plasmacytoid dendritic cells

    OpenAIRE

    Gujer, Cornelia

    2011-01-01

    Dendritic cells (DCs) are early sentinels of pathogen exposure and central in the initiation and orchestration of adaptive immune responses. Apart from the prominent role of DCs in the activation of T cells, DCs have been shown to influence humoral B cell mediated responses. DCs are therefore important cells for regulating immune responses to vaccines. Many of the vaccines under development today are against pathogens such as Mycobacterium tuberculosis and HIV-1 that likely r...

  17. Anatomy of a new B-cell-specific enhancer.

    OpenAIRE

    Koch, W; Benoist, C.; Mathis, D

    1989-01-01

    The major histocompatibility complex class II molecules, like the immunoglobulins, are prominent B-lymphocyte markers. Herein, we describe a B-cell-specific enhancer associated with the murine class II gene, Ek alpha. This enhancer has a complex anatomy that suggests interactions between remotely spaced elements. Of particular interest is the finding that two CCAAT boxes spaced one kilobase apart are important for enhancer activity. Somewhat surprisingly, the E alpha and immunoglobulin enhanc...

  18. Merkel Cell Polyomavirus Large T Antigen Has Growth-Promoting and Inhibitory Activities

    OpenAIRE

    Cheng, Jingwei; Rozenblatt-Rosen, Orit; Paulson, Kelly G.; Nghiem, Paul; DeCaprio, James A.

    2013-01-01

    Merkel cell carcinoma (MCC) is a rare and aggressive form of skin cancer. In at least 80% of all MCC, Merkel cell polyomavirus (MCPyV) DNA has undergone clonal integration into the host cell genome, and most tumors express the MCPyV large and small T antigens. In all cases of MCC reported to date, the integrated MCPyV genome has undergone mutations in the large T antigen. These mutations result in expression of a truncated large T antigen that retains the Rb binding or LXCXE motif but deletes...

  19. The relationship between colonization and haemagglutination inhibiting and B cell epitopes of Porphyromonas gingivalis

    Science.gov (United States)

    KELLY, C G; BOOTH, V; KENDAL, H; SLANEY, J M; CURTIS, M A; LEHNER, T

    1997-01-01

    Passive immunization with the monoclonal antibody 61BG1.3 selectively prevents colonization by Porphyromonas gingivalis in humans (Booth V, Ashley FP, Lehner T. Infect Immun 1996; 64:422-7). The protective MoAb recognizes the j3 component of the RI protease of P. gingivalis which is formed by proteolytic processing of a polyprotein precursor termed PrpRl. This subunit is both a haemagglutinin and an antigen which is recognized by sera from patients with periodontitis. In this study the relationship was investigated between a colonization epitope which is recognized by the MoAb 61BG1.3, a haemagglutinating and B cell epitope which are recognized by sera from patients with periodontitis. B cell epitopes were mapped by Western blotting with a series of truncated recombinant polypeptides spanning the adhesion domain within residues 784–1130 of PrpRl and by ELISA using a panel of synthetic peptides spanning the same sequence. The epitope which is recognized by the protective MoAb was mapped within residues 907–931 of PrpRl, while serum responses of patients were directed predominantly to the adjacent carboxy-terminal sequence within residues 934–1042. The haemagglutinating epitope was mapped to residues 1073–1112. In view of our previous findings that the MoAb 61BG1.3 prevents colonization of P. gingivalis in vivo and inhibits haemagglutination, these two epitopes may be in proximity in the native protein. Active or passive immunization strategies which target the protective or haemagglutinating epitopes of the adhesion domain of PrpRl may provide a means of preventing infection with P. gingivalis. PMID:9367414

  20. Preparation, Characterization, and Determination of Immunological Activities of Transfer Factor Specific to Human Sperm Antigen

    Directory of Open Access Journals (Sweden)

    Jianwei Zhou

    2013-01-01

    Full Text Available Objective. The objective of this study was to prepare, characterize, and determine immunological activities of specific transfer factor (STF specific to human sperm antigen (HSA for the preparation of antisperm contraceptive vaccine that can be used as an immunocontraceptive. Methods. HSA-STF was prepared using the spleens of rabbits vaccinated with HSA. The specific immunological activities were examined by lymphocyte proliferation test (LPT, leukocyte adhesion inhibition test (LAIT, and by determining the concentrations of IL-4, γ-IFN, and IL-21. HSA-STF was a helveolous substance, having a pH value of 7.0±0.4 and UV absorption maxima at 258 ± 6 nm. It contained seventeen amino acids; glycine and glutamic acids were the highest in terms of concentrations (38.8 μg/mL and 36.3 μg/mL, resp.. Results. The concentration of polypeptide was 2.34±0.31 mg/mL, and ribose was 0.717±0.043 mg/mL. The stimulation index for lymphocyte proliferation test was 1.84, and the leukocyte adhesion inhibition rate was 37.7%. There was a statistically significant difference between the cultural lymphocytes with HSA-STF and non-HSA-STF for γ-IFN and IL-21 (P0.05. Conclusion. HSA-STF was prepared and characterized successfully. It had immunological activity which could transfer the immune response specific to HSA and prove to be a potential candidate for the development of male immunocontraceptive agents.