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Sample records for human b-cell lines

  1. Interleukin-2 production by human leukemia cell lines of pre-B cell origin

    International Nuclear Information System (INIS)

    Holan, V.; Minowada, J.

    1993-01-01

    Cells of 7 tested human leukemia cell lines of pre-B cell origin (as characterized by immunophenotyping and by the expression of cytoplasmic micro chains, but not by surface immunoglobulins) produced after stimulation with bacterial lipopolysaccharide (LPS) or phorbol myristate acetate (PMA) a lymphokine activity which supported the growth of the interleukin-2 (IL-2)-dependent CTLL-2 cell line. Three pieces of evidence indicate that the secreted lymphokine was functionally and antigenically very similar, if not identical, to human IL-2: (1) The lymphokine supported the growth of murine IL-2-dependent CTLL-2 cells, which did not respond to human lymphokines other than IL-2, but it did not stimulate the growth of murine IL-3-dependent FDC-P2 cells, (2) the biological activity of the lymphokine was was inhibited by monoclonal antibody (mAb) anti-human-IL-2, and (3) the proliferation of IL-2-dependent cells in the presence of the active materials was completely inhibited by the inclusion of the anti-mouse-IL-2 receptor (IL-2R) mAb. Since leukemia cells of immature B-cell origin also synthesize IL-2R, the human pre-B cell leukemias could represent another type of hematological malignancy where the autocrine processes of IL-2 production and utilization are involved in the expansion of the disease. (author)

  2. Heterogeneity in both cytokine production and responsiveness of a panel of monoclonal human Epstein-Barr virus-transformed B-cell lines

    NARCIS (Netherlands)

    Jochems, G. J.; Klein, M. R.; Jordens, R.; Pascual-Salcedo, D.; van Boxtel-Oosterhof, F.; van Lier, R. A.; Zeijlemaker, W. P.

    1991-01-01

    To optimize growth and Ig production of in vitro-cultured Epstein-Barr virus (EBV)-transformed B cells, a panel of six monoclonal EBV B-cell lines was analyzed for autocrine growth factor production and responsiveness to various cytokines. Three cell lines produced Il-I and four produced Il-6,

  3. Generation of a human immunodeficiency virus type 1 chronically infected monkey B cell line expressing low levels of endogenous TRIM5alpha.

    Science.gov (United States)

    Ridolfi, Barbara; Catone, Stefania; Sgarbanti, Marco; Sernicola, Leonardo; Battistini, Angela; Parolin, Cristina; Titti, Fausto; Borsetti, Alessandra

    2009-12-01

    Several innate cellular antiviral factors exist in mammalian cells that prevent the replication of retroviruses. Among them, the tripartite motif protein (TRIM)5alpha has been shown to block human immunodeficiency virus type 1 (HIV-1) infection in several types of Old World monkey cells. Here we report a novel HIV-1 chronically infected monkey B cell line, F6/HIV-1, characterized by very low levels of TRIM5alpha expression that allows HIV-1 to overcome the restriction. Virus produced by F6/HIV-1 cells fails to infect monkey cells but retains the ability to infect human peripheral blood mononuclear cells (PBMCs) and T cell lines, although with a reduced infectivity compared to the input virus. Ultrastructural analyses revealed the presence of budding virions at the F6/HIV-1 cells plasma membrane characterized by a typical conical core shell. To our knowledge F6/HIV-1 is the first monkey cell line chronically infected by HIV-1 and able to release infectious particles thus representing a useful tool to gain further insights into the molecular mechanisms of HIV-1 pathogenesis.

  4. Interaction of Staphylococci with Human B cells.

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    Tyler K Nygaard

    Full Text Available Staphylococcus aureus is a leading cause of human infections worldwide. The pathogen produces numerous molecules that can interfere with recognition and binding by host innate immune cells, an initial step required for the ingestion and subsequent destruction of microbes by phagocytes. To better understand the interaction of this pathogen with human immune cells, we compared the association of S. aureus and S. epidermidis with leukocytes in human blood. We found that a significantly greater proportion of B cells associated with S. epidermidis relative to S. aureus. Complement components and complement receptors were important for the binding of B cells with S. epidermidis. Experiments using staphylococci inactivated by ultraviolet radiation and S. aureus isogenic deletion mutants indicated that S. aureus secretes molecules regulated by the SaeR/S two-component system that interfere with the ability of human B cells to bind this bacterium. We hypothesize that the relative inability of B cells to bind S. aureus contributes to the microbe's success as a human pathogen.

  5. Continuous Lymphoid Cell Lines with Characteristics of B Cells (Bone-Marrow-Derived), Lacking the Epstein-Barr Virus Genome and Derived from Three Human Lymphomas

    Science.gov (United States)

    Klein, George; Lindahl, Tomas; Jondal, Mikael; Leibold, Wolfgang; Menézes, José; Nilsson, Kenneth; Sundström, Christer

    1974-01-01

    Three exceptional cell lines have been tested for the presence of the Epstein-Barr virus genome by nucleic acid hybridization (complementary RNA·DNA) and Epstein-Barr virus-determined nuclear antigen tests. Two lines were derived from Swedish lymphoma cases and one from an African Burkitt-like lymphoma biopsy that was negative for Epstein-Barr virus DNA and the virus-determined nuclear antigen. All three lines apparently lacked the viral genome. Two of the three lines clearly had characteristics of B-cells (bone-marrow-derived). PMID:4369887

  6. JSI-124 inhibits IgE production in an IgE B cell line

    International Nuclear Information System (INIS)

    Cui, Lulu; Bi, Jiacheng; Yan, Dehong; Ye, Xiufeng; Zheng, Mingxing; Yu, Guang; Wan, Xiaochun

    2017-01-01

    IgE is a key effector molecule in atopic diseases; however, the regulation mechanisms of IgE production in IgE B cells remain poorly understood. In the present study, we demonstrate that JSI-124 (cucurbitacin I), a selective STAT3 inhibitor, selectively inhibits production of IgE by a human IgE B cell line, CRL-8033 cells, while does not affect the IgG production by IgG B cell lines. In the aspect of molecular mechanism, we found that Igλ, but not Ighe, gene expression was suppressed by JSI-124. The above effects of JSI-124 were not mediated by affecting cellular proliferation or apoptosis. Furthermore, multiple B cell differentiation-related genes expression was not significantly affected by JSI-124. Taken together, we demonstrate a potential strategy of therapeutically suppressing IgE production without affecting IgG production in atopic patients. - Highlights: • JSI-124 inhibits IgE production in an IgE B cell line, CRL-8033 cells. • JSI-124 does not affect IgG production by IgG B cell lines. • JSI-124 inhibits IgE production mainly by suppressing transcription of Igλ.

  7. The small FOXP1 isoform predominantly expressed in activated B cell-like diffuse large B-cell lymphoma and full-length FOXP1 exert similar oncogenic and transcriptional activity in human B cells.

    Science.gov (United States)

    van Keimpema, Martine; Grüneberg, Leonie J; Schilder-Tol, Esther J M; Oud, Monique E C M; Beuling, Esther A; Hensbergen, Paul J; de Jong, Johann; Pals, Steven T; Spaargaren, Marcel

    2017-03-01

    The forkhead transcription factor FOXP1 is generally regarded as an oncogene in activated B cell-like diffuse large B-cell lymphoma. Previous studies have suggested that a small isoform of FOXP1 rather than full-length FOXP1, may possess this oncogenic activity. Corroborating those studies, we herein show that activated B cell-like diffuse large B-cell lymphoma cell lines and primary activated B cell-like diffuse large B-cell lymphoma cells predominantly express a small FOXP1 isoform, and that the 5'-end of the Foxp1 gene is a common insertion site in murine lymphomas in leukemia virus- and transposon-mediated insertional mutagenesis screens. By combined mass spectrometry, (quantative) reverse transcription polymerase chain reaction/sequencing, and small interfering ribonucleic acid-mediated gene silencing, we determined that the small FOXP1 isoform predominantly expressed in activated B cell-like diffuse large B-cell lymphoma lacks the N-terminal 100 amino acids of full-length FOXP1. Aberrant overexpression of this FOXP1 isoform (ΔN100) in primary human B cells revealed its oncogenic capacity; it repressed apoptosis and plasma cell differentiation. However, no difference in potency was found between this small FOXP1 isoform and full-length FOXP1. Furthermore, overexpression of full-length FOXP1 or this small FOXP1 isoform in primary B cells and diffuse large B-cell lymphoma cell lines resulted in similar gene regulation. Taken together, our data indicate that this small FOXP1 isoform and full-length FOXP1 have comparable oncogenic and transcriptional activity in human B cells, suggesting that aberrant expression or overexpression of FOXP1, irrespective of the specific isoform, contributes to lymphomagenesis. These novel insights further enhance the value of FOXP1 for the diagnostics, prognostics, and treatment of diffuse large B-cell lymphoma patients. Copyright© Ferrata Storti Foundation.

  8. Preparation of antisera specific for human B cells by immunization of rabbits with immune complexes

    International Nuclear Information System (INIS)

    Welsh, K.I.; Turner, M.J.

    1976-01-01

    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)

  9. 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. Copyright © 2016 by The American Association of Immunologists, Inc.

  10. Human regulatory B cells control the TFH cell response.

    Science.gov (United States)

    Achour, Achouak; Simon, Quentin; Mohr, Audrey; Séité, Jean-François; Youinou, Pierre; Bendaoud, Boutahar; Ghedira, Ibtissem; Pers, Jacques-Olivier; Jamin, Christophe

    2017-07-01

    Follicular helper T (T FH ) cells support terminal B-cell differentiation. Human regulatory B (Breg) cells modulate cellular responses, but their control of T FH cell-dependent humoral immune responses is unknown. We sought to assess the role of Breg cells on T FH cell development and function. Human T cells were polyclonally stimulated in the presence of IL-12 and IL-21 to generate T FH cells. They were cocultured with B cells to induce their terminal differentiation. Breg cells were included in these cultures, and their effects were evaluated by using flow cytometry and ELISA. B-cell lymphoma 6, IL-21, inducible costimulator, CXCR5, and programmed cell death protein 1 (PD-1) expressions increased on stimulated human T cells, characterizing T FH cell maturation. In cocultures they differentiated B cells into CD138 + plasma and IgD - CD27 + memory cells and triggered immunoglobulin secretions. Breg cells obtained by Toll-like receptor 9 and CD40 activation of B cells prevented T FH cell development. Added to T FH cell and B-cell cocultures, they inhibited B-cell differentiation, impeded immunoglobulin secretions, and expanded Foxp3 + CXCR5 + PD-1 + follicular regulatory T cells. Breg cells modulated IL-21 receptor expressions on T FH cells and B cells, and their suppressive activities involved CD40, CD80, CD86, and intercellular adhesion molecule interactions and required production of IL-10 and TGF-β. Human Breg cells control T FH cell maturation, expand follicular regulatory T cells, and inhibit the T FH cell-mediated antibody secretion. These novel observations demonstrate a role for the Breg cell in germinal center reactions and suggest that deficient activities might impair the T FH cell-dependent control of humoral immunity and might lead to the development of aberrant autoimmune responses. Copyright © 2016 American Academy of Allergy, Asthma & Immunology. Published by Elsevier Inc. All rights reserved.

  11. Immunomodulating effects of heparin on human B cell proliferation

    International Nuclear Information System (INIS)

    Wasik, Maria; Stepien-Sopniewska, Barbara; Gorski, Andrzej

    1993-01-01

    Recent data indicate that heparin may act as an immunomodulator. In this paper we have analyzed the effect of this agent on human B cell proliferation ''in vitro'' induced by ''S. aureus'' Cowan. The action of heparin is complex, but there was a trend for inhibition of B cell responses obtained from defibrinated but not heparinized blood samples. This suggest that heparin interacts with platelet products (growth factors, cytokines) and the results of such interactions determine the final effect. (author). 6 refs, 4 figs

  12. Human B cells fail to secrete type I interferons upon cytoplasmic DNA exposure.

    Science.gov (United States)

    Gram, Anna M; Sun, Chenglong; Landman, Sanne L; Oosenbrug, Timo; Koppejan, Hester J; Kwakkenbos, Mark J; Hoeben, Rob C; Paludan, Søren R; Ressing, Maaike E

    2017-11-01

    Most cells are believed to be capable of producing type I interferons (IFN I) as part of an innate immune response against, for instance, viral infections. In macrophages, IFN I is potently induced upon cytoplasmic exposure to foreign nucleic acids. Infection of these cells with herpesviruses leads to triggering of the DNA sensors interferon-inducible protein 16 (IFI16) and cyclic GMP-AMP (cGAMP) synthase (cGAS). Thereby, the stimulator of interferon genes (STING) and the downstream molecules TANK-binding kinase 1 (TBK1) and interferon regulatory factor 3 (IRF3) are sequentially activated culminating in IFN I secretion. Human gamma-herpesviruses, such as Epstein-Barr virus (EBV), exploit B cells as a reservoir for persistent infection. In this study, we investigated whether human B cells, similar to macrophages, engage the cytoplasmic DNA sensing pathway to induce an innate immune response. We found that the B cells fail to secrete IFN I upon cytoplasmic DNA exposure, although they express the DNA sensors cGAS and IFI16 and the signaling components TBK1 and IRF3. In primary human B lymphocytes and EBV-negative B cell lines, this deficiency is explained by a lack of detectable levels of the central adaptor protein STING. In contrast, EBV-transformed B cell lines did express STING, yet both these lines as well as STING-reconstituted EBV-negative B cells did not produce IFN I upon dsDNA or cGAMP stimulation. Our combined data show that the cytoplasmic DNA sensing pathway is dysfunctional in human B cells. This exemplifies that certain cell types cannot induce IFN I in response to cytoplasmic DNA exposure providing a potential niche for viral persistence. Copyright © 2017 The Authors. Published by Elsevier Ltd.. All rights reserved.

  13. Molecular characterization of neoplastic and normal "sister" lymphoblastoid B-cell lines from chronic lymphocytic leukemia

    DEFF Research Database (Denmark)

    Lanemo Myhrinder, Anna; Hellqvist, Eva; Bergh, Ann-Charlotte

    2013-01-01

    Chronic lymphocytic leukemia (CLL) B-cells resemble self-renewing CD5 + B-cells carrying auto/xeno-antigen-reactive B-cell receptors (BCRs) and multiple innate pattern-recognition receptors, such as Toll-like receptors and scavenger receptors. Integration of signals from BCRs with multiple surface...... a comprehensive genotypic and phenotypic characterization of available CLL and normal B-cell-derived lymphoblastoid cell lines (LCLs) from the same individuals (n = 17). Authenticity and verification studies of CLL-patient origin were done by IGHV sequencing, fluorescence in situ hybridization (FISH) and DNA...

  14. Dissection of pathways leading to antigen receptor-induced and Fas/CD95-induced apoptosis in human B cells

    NARCIS (Netherlands)

    Lens, S. M.; den Drijver, B. F.; Pötgens, A. J.; Tesselaar, K.; van Oers, M. H.; van Lier, R. A.

    1998-01-01

    To dissect intracellular pathways involved in B cell Ag receptor (BCR)-mediated and Fas-induced human B cell death, we isolated clones of the Burkitt lymphoma cell line Ramos with different apoptosis sensitivities. Selection for sensitivity to Fas-induced apoptosis also selected for clones with

  15. Absence of annexin I expression in B-cell non-Hodgkin's lymphomas and cell lines

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    Gopalakrishnan Velliyur K

    2004-03-01

    Full Text Available Abstract Background Annexin I, one of the 20 members of the annexin family of calcium and phospholipid-binding proteins, has been implicated in diverse biological processes including signal transduction, mediation of apoptosis and immunosuppression. Previous studies have shown increased annexin I expression in pancreatic and breast cancers, while it is absent in prostate and esophageal cancers. Results Data presented here show that annexin I mRNA and protein are undetectable in 10 out of 12 B-cell lymphoma cell lines examined. Southern blot analysis indicates that the annexin I gene is intact in B-cell lymphoma cell lines. Aberrant methylation was examined as a cause for lack of annexin I expression by treating cells 5-Aza-2-deoxycytidine. Reexpression of annexin I was observed after prolonged treatment with the demethylating agent indicating methylation may be one of the mechanisms of annexin I silencing. Treatment of Raji and OMA-BL-1 cells with lipopolysaccharide, an inflammation inducer, and with hydrogen peroxide, a promoter of oxidative stress, also failed to induce annexin I expression. Annexin I expression was examined in primary lymphoma tissues by immunohistochemistry and presence of annexin I in a subset of normal B-cells and absence of annexin I expression in the lymphoma tissues were observed. These results show that annexin I is expressed in normal B-cells, and its expression is lost in all primary B-cell lymphomas and 10 of 12 B-cell lymphoma cell lines. Conclusions Our results suggest that, similar to prostate and esophageal cancers, annexin I may be an endogenous suppressor of cancer development, and loss of annexin I may contribute to B-cell lymphoma development.

  16. The nanoscale spatial organization of B-cell receptors on immunoglobulin M- and G-expressing human B-cells.

    Science.gov (United States)

    Lee, Jinmin; Sengupta, Prabuddha; Brzostowski, Joseph; Lippincott-Schwartz, Jennifer; Pierce, Susan K

    2017-02-15

    B-cell activation is initiated by the binding of antigen to the B-cell receptor (BCR). Here we used dSTORM superresolution imaging to characterize the nanoscale spatial organization of immunoglobulin M (IgM) and IgG BCRs on the surfaces of resting and antigen--activated human peripheral blood B-cells. We provide insights into both the fundamental process of antigen-driven BCR clustering and differences in the spatial organization of IgM and IgG BCRs that may contribute to the characteristic differences in the responses of naive and memory B-cells to antigen. We provide evidence that although both IgM and IgG BCRs reside in highly heterogeneous protein islands that vary in size and number of BCR single-molecule localizations, both resting and activated B-cells intrinsically maintain a high -frequency of single isolated BCR localizations, which likely represent BCR monomers. IgG BCRs are more clustered than IgM BCRs on resting cells and form larger protein islands after antigen activation. Small, dense BCR clusters likely formed via protein-protein interactions are present on the surface of resting cells, and antigen activation induces these to come together to form less dense, larger islands, a process likely governed, at least in part, by protein-lipid interactions. © 2017 Lee, Sengupta, et al. This article is distributed by The American Society for Cell Biology under license from the author(s). Two months after publication it is available to the public under an Attribution–Noncommercial–Share Alike 3.0 Unported Creative Commons License (http://creativecommons.org/licenses/by-nc-sa/3.0).

  17. Invasin of Yersinia pseudotuberculosis activates human peripheral B cells.

    OpenAIRE

    Lundgren, E; Carballeira, N; Vazquez, R; Dubinina, E; Bränden, H; Persson, H; Wolf-Watz, H

    1996-01-01

    The Yersinia pseudotuberculosis cell surface-located protein invasin was found to promote binding between the pathogen and resting peripheral B cells via beta 1 integrin receptors (CD29). B cells responded by expressing several activation markers and by growing, In contrast, T cells did not react, although these cells express CD29. An isogenic invA mutant failed to activate B cells. The mutation could be complemented by providing the invA+ gene in trans. Purified invasin alone did not activat...

  18. A role for gut-associated lymphoid tissue in shaping the human B cell repertoire.

    Science.gov (United States)

    Vossenkämper, Anna; Blair, Paul A; Safinia, Niloufar; Fraser, Louise D; Das, Lisa; Sanders, Theodore J; Stagg, Andrew J; Sanderson, Jeremy D; Taylor, Kirstin; Chang, Fuju; Choong, Lee M; D'Cruz, David P; Macdonald, Thomas T; Lombardi, Giovanna; Spencer, Jo

    2013-08-26

    We have tracked the fate of immature human B cells at a critical stage in their development when the mature B cell repertoire is shaped. We show that a major subset of bone marrow emigrant immature human B cells, the transitional 2 (T2) B cells, homes to gut-associated lymphoid tissue (GALT) and that most T2 B cells isolated from human GALT are activated. Activation in GALT is a previously unknown potential fate for immature human B cells. The process of maturation from immature transitional B cell through to mature naive B cell includes the removal of autoreactive cells from the developing repertoire, a process which is known to fail in systemic lupus erythematosus (SLE). We observe that immature B cells in SLE are poorly equipped to access the gut and that gut immune compartments are depleted in SLE. Thus, activation of immature B cells in GALT may function as a checkpoint that protects against autoimmunity. In healthy individuals, this pathway may be involved in generating the vast population of IgA plasma cells and also the enigmatic marginal zone B cell subset that is poorly understood in humans.

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

    OpenAIRE

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

    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.

  20. 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 candid......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...

  1. Epigenetic inactivation of Notch-Hes pathway in human B-cell acute lymphoblastic leukemia.

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    Shao-Qing Kuang

    Full Text Available The Notch pathway can have both oncogenic and tumor suppressor roles, depending on cell context. For example, Notch signaling promotes T cell differentiation and is leukemogenic in T cells, whereas it inhibits early B cell differentiation and acts as a tumor suppressor in B cell leukemia where it induces growth arrest and apoptosis. The regulatory mechanisms that contribute to these opposing roles are not understood. Aberrant promoter DNA methylation and histone modifications are associated with silencing of tumor suppressor genes and have been implicated in leukemogenesis. Using methylated CpG island amplification (MCA/DNA promoter microarray, we identified Notch3 and Hes5 as hypermethylated in human B cell acute lymphoblastic leukemia (ALL. We investigated the methylation status of other Notch pathway genes by bisulfite pyrosequencing. Notch3, JAG1, Hes2, Hes4 and Hes5 were frequently hypermethylated in B leukemia cell lines and primary B-ALL, in contrast to T-ALL cell lines and patient samples. Aberrant methylation of Notch3 and Hes5 in B-ALL was associated with gene silencing and was accompanied by decrease of H3K4 trimethylation and H3K9 acetylation and gain of H3K9 trimethylation and H3K27 trimethylation. 5-aza-2'-deoxycytidine treatment restored Hes5 expression and decreased promoter hypermethylation in most leukemia cell lines and primary B-ALL samples. Restoration of Hes5 expression by lentiviral transduction resulted in growth arrest and apoptosis in Hes5 negative B-ALL cells but not in Hes5 expressing T-ALL cells. These data suggest that epigenetic modifications are implicated in silencing of tumor suppressor of Notch/Hes pathway in B-ALL.

  2. Epigenetic inactivation of Notch-Hes pathway in human B-cell acute lymphoblastic leukemia.

    Science.gov (United States)

    Kuang, Shao-Qing; Fang, Zhihong; Zweidler-McKay, Patrick A; Yang, Hui; Wei, Yue; Gonzalez-Cervantes, Emilio A; Boumber, Yanis; Garcia-Manero, Guillermo

    2013-01-01

    The Notch pathway can have both oncogenic and tumor suppressor roles, depending on cell context. For example, Notch signaling promotes T cell differentiation and is leukemogenic in T cells, whereas it inhibits early B cell differentiation and acts as a tumor suppressor in B cell leukemia where it induces growth arrest and apoptosis. The regulatory mechanisms that contribute to these opposing roles are not understood. Aberrant promoter DNA methylation and histone modifications are associated with silencing of tumor suppressor genes and have been implicated in leukemogenesis. Using methylated CpG island amplification (MCA)/DNA promoter microarray, we identified Notch3 and Hes5 as hypermethylated in human B cell acute lymphoblastic leukemia (ALL). We investigated the methylation status of other Notch pathway genes by bisulfite pyrosequencing. Notch3, JAG1, Hes2, Hes4 and Hes5 were frequently hypermethylated in B leukemia cell lines and primary B-ALL, in contrast to T-ALL cell lines and patient samples. Aberrant methylation of Notch3 and Hes5 in B-ALL was associated with gene silencing and was accompanied by decrease of H3K4 trimethylation and H3K9 acetylation and gain of H3K9 trimethylation and H3K27 trimethylation. 5-aza-2'-deoxycytidine treatment restored Hes5 expression and decreased promoter hypermethylation in most leukemia cell lines and primary B-ALL samples. Restoration of Hes5 expression by lentiviral transduction resulted in growth arrest and apoptosis in Hes5 negative B-ALL cells but not in Hes5 expressing T-ALL cells. These data suggest that epigenetic modifications are implicated in silencing of tumor suppressor of Notch/Hes pathway in B-ALL.

  3. Impact of tofacitinib treatment on human B-cells in vitro and in vivo.

    Science.gov (United States)

    Rizzi, Marta; Lorenzetti, Raquel; Fischer, Kathleen; Staniek, Julian; Janowska, Iga; Troilo, Arianna; Strohmeier, Valentina; Erlacher, Miriam; Kunze, Mirjam; Bannert, Bettina; Kyburz, Diego; Voll, Reinhard E; Venhoff, Nils; Thiel, Jens

    2017-02-01

    B-cells are pivotal to the pathogenesis of rheumatoid arthritis and tofacitinib, a JAK inhibitor, is effective and safe in its treatment. Tofacitinib interferes with signal transduction via cytokine receptors using the common γ-chain. Despite extensive data on T-lymphocytes, the impact of tofacitinib on B-lymphocytes is poorly understood. In this study we assessed the effect of tofacitinib on B-lymphocyte differentiation and function. Tofacitinib treatment strongly impaired in vitro plasmablast development, immunoglobulin secretion and induction of B-cell fate determining transcription factors, Blimp-1, Xbp-1, and IRF-4, in naïve B-cells. Interestingly, class switch and activation-induced cytidine deaminase (AICDA) induction was only slightly reduced in activated naïve B-cells. The effect of tofacitinib on plasmablast formation, immunoglobulin secretion and proliferation was less profound, when peripheral blood B-cells, including not only naïve but also memory B-cells, were stimulated. In line with these in vitro results, the relative distribution of B-cell populations remained stable in tofacitinib treated patients. Nevertheless, a temporary increase in absolute B-cell numbers was observed 6-8 weeks after start of treatment. In addition, B-cells isolated from tofacitinib treated patients responded rapidly to in vitro activation. We demonstrate that tofacitinib has a direct impact on human naïve B-lymphocytes, independently from its effect on T-lymphocytes, by impairing their development into plasmablasts and immunoglobulin secretion. The major effect of tofacitinib on naïve B-lymphocyte development points to the potential inability of tofacitinib-treated patients to respond to novel antigens, and suggests planning vaccination strategies prior to tofacitinib treatment. Copyright © 2016 Elsevier Ltd. All rights reserved.

  4. Selective induction of DNA repair pathways in human B cells activated by CD4+ T cells.

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    Xiaosheng Wu

    Full Text Available Greater than 75% of all hematologic malignancies derive from germinal center (GC or post-GC B cells, suggesting that the GC reaction predisposes B cells to tumorigenesis. Because GC B cells acquire expression of the highly mutagenic enzyme activation-induced cytidine deaminase (AID, GC B cells may require additional DNA repair capacity. The goal of this study was to investigate whether normal human B cells acquire enhanced expression of DNA repair factors upon AID induction. We first demonstrated that several DNA mismatch repair, homologous recombination, base excision repair, and ATR signaling genes were overexpressed in GC B cells relative to naïve and memory B cells, reflecting activation of a process we have termed somatic hyperrepair (SHR. Using an in vitro system, we next characterized activation signals required to induce AID expression and SHR. Although AID expression was induced by a variety of polyclonal activators, SHR induction strictly required signals provided by contact with activated CD4+ T cells, and B cells activated in this manner displayed reduced levels of DNA damage-induced apoptosis. We further show the induction of SHR is independent of AID expression, as GC B cells from AID-/-mice retained heightened expression of SHR proteins. In consideration of the critical role that CD4+ T cells play in inducing the SHR process, our data suggest a novel role for CD4+ T cells in the tumor suppression of GC/post-GC B cells.

  5. Establishment of a novel feline leukemia virus (FeLV)-negative B-cell cell line from a cat with B-cell lymphoma.

    Science.gov (United States)

    Mochizuki, Hiroyuki; Takahashi, Masashi; Nishigaki, Kazuo; Ide, Tetsuya; Goto-Koshino, Yuko; Watanabe, Shinya; Sato, Hirofumi; Sato, Masahiko; Kotera, Yukiko; Fujino, Yasuhito; Ohno, Koichi; Uchida, Kazuyuki; Tsujimoto, Hajime

    2011-04-15

    We established a novel feline B-cell line, MS4, from the neoplastic pleural effusion of a cat with cutaneous B-cell lymphoma. Immunophenotype staining of the MS4 cells was positive for CD20, CD79α, and IgA and negative for CD3, CD4, CD5, CD8α, CD18, CD21, CD22, IgM, IgG, Ig light chain, and MHC class II. PCR analysis for immunoglobulin heavy chain gene rearrangements revealed a monoclonal rearrangement, whereas no clonal rearrangement of the T-cell receptor γ gene was detected. Southern blotting with an exogenous feline leukemia virus (FeLV) U3 probe revealed no integration of exogenous FeLV provirus. The MS4 cell line is the first FeLV-negative feline B-cell lymphoma cell line, and may be used to investigate the pathogenesis of spontaneously occurring feline lymphoma and the development of new therapies. Copyright © 2011 Elsevier B.V. All rights reserved.

  6. The chicken B-cell line DT40 proteome, beadome and interactomes

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    Johanna S. Rees

    2015-06-01

    Full Text Available In developing a new quantitative AP-MS method for exploring interactomes in the chicken B-cell line DT40, we also surveyed the most abundant proteins in this organism and explored the likely contaminants that bind to a variety of affinity resins that would later be confirmed quantitatively [1]. We present the ‘Top 150 abundant DT40 proteins list’, the DT40 beadomes as well as protein interaction lists for the Phosphatidyl inositol 5-phosphate 4-kinase 2β and Fanconi anaemia protein complexes.

  7. Genetic modification of human B-cell development: B-cell development is inhibited by the dominant negative helix loop helix factor Id3

    NARCIS (Netherlands)

    Jaleco, A. C.; Stegmann, A. P.; Heemskerk, M. H.; Couwenberg, F.; Bakker, A. Q.; Weijer, K.; Spits, H.

    1999-01-01

    Transgenic and gene targeted mice have contributed greatly to our understanding of the mechanisms underlying B-cell development. We describe here a model system that allows us to apply molecular genetic techniques to the analysis of human B-cell development. We constructed a retroviral vector with a

  8. Maternal and fetal mechanisms of B cell regulation during pregnancy: human Chorionic Gonadotropin stimulates B cells to produce IL-10 while alpha-fetoprotein drives them into apoptosis

    Directory of Open Access Journals (Sweden)

    Franziska Fettke

    2016-12-01

    Full Text Available Maternal immune tolerance towards the fetus is an essential requisite for pregnancy. While T cell functions are well documented, little is known about the participation of B cells. We have previously suggested that IL-10 producing B cells are involved in pregnancy tolerance in mice and humans. By employing murine and human systems, we report now that fetal trophoblasts positively regulate the generation of IL-10 producing B cells. We next studied the participation of hormones produced by the placenta as well as the fetal protein alpha-fetoprotein (AFP in B cell modulation. Human Chorionic Gonadotropin (hCG, but not progesterone, estrogen or a combination of both, was able to promote changes in B cell phenotype and boost their IL-10 production, which was abolished after blocking hCG. The hCG-induced B cell phenotype was not associated with augmented galactosylation, sialylation or fucosylation of IgG subclasses in their Fc. In vitro, hCG induced the synthesis of asymmetrically glycosylated antibodies in their Fab region. Interestingly, AFP had dual effects depending on the concentration. At concentrations corresponding to maternal serum levels, it did not modify the phenotype or IL-10 secretion of B cells. At fetal concentrations, however, AFP was able to drive B cells into apoptosis, which may indicate a protective mechanism to avoid maternal B cells to reach the fetus.Our data suggests that the fetus secrete factors that promote a pregnancy-friendly B cell phenotype, unraveling interesting aspects of B cell function and modulation by pregnancy hormones and fetal proteins.

  9. The synthetic epoxyquinoids jesterone dimer and epoxyquinone A monomer induce apoptosis and inhibit REL (human c-Rel) DNA binding in an IkappaBalpha-deficient diffuse large B-cell lymphoma cell line.

    Science.gov (United States)

    Liang, Mei-Chih; Bardhan, Sujata; Porco, John A; Gilmore, Thomas D

    2006-09-08

    The NF-kappaB transcription factor signaling pathway is constitutively active in many human cancers, and inhibition of this pathway can often kill cancer cells by inducing apoptosis. In this study, we show that two synthetic epoxyquinoids, jesterone dimer (JD) and epoxyquinone A monomer (EqM), are equally effective at inhibiting the growth of two human lymphoma cell lines that have constitutively nuclear REL (human c-Rel) DNA-binding complexes, but either express (SUDHL-4 cells) or do not express (RC-K8 cells) the NF-kappaB inhibitor IkappaBalpha. Furthermore, in these cells, both JD and EqM dose-dependently induced apoptosis, inhibited REL DNA-binding activity, and converted REL to a high molecular weight form. In A293 cells, JD and EqM inhibited the DNA-binding activity of overexpressed REL, but not p50. Replacement of Cys-27 with Ser in REL reduced JD- and EqM-mediated inhibition of REL DNA-binding activity. These results suggest that JD and EqM can induce apoptosis in IkappaBalpha-deficient lymphoma cells through a mechanism involving direct inhibition of transcription factor REL.

  10. Authentication of Primordial Characteristics of the CLBL-1 Cell Line Prove the Integrity of a Canine B-Cell Lymphoma in a Murine In Vivo Model

    OpenAIRE

    Rütgen, Barbara C.; Willenbrock, Saskia; Reimann-Berg, Nicola; Walter, Ingrid; Fuchs-Baumgartinger, Andrea; Wagner, Siegfried; Kovacic, Boris; Essler, Sabine E.; Schwendenwein, Ilse; Nolte, Ingo; Saalmüller, Armin; Escobar, Hugo Murua

    2012-01-01

    Cell lines are key tools in cancer research allowing the generation of neoplasias in animal models resembling the initial tumours able to mimic the original neoplasias closely in vivo. Canine lymphoma is the major hematopoietic malignancy in dogs and considered as a valuable spontaneous large animal model for human Non-Hodgkin's Lymphoma (NHL). Herein we describe the establishment and characterisation of an in vivo model using the canine B-cell lymphoma cell line CLBL-1 analysing the stabilit...

  11. Signaling through intercellular adhesion molecule 1 (ICAM-1) in a B cell lymphoma line

    DEFF Research Database (Denmark)

    Holland, J; Owens, T

    1997-01-01

    Intercellular adhesion molecule 1 (ICAM-1) (CD54) is an adhesion molecule of the immunoglobulin superfamily. The interaction between ICAM-1 on B lymphocytes and leukocyte function-associated antigen 1 on T cells plays a major role in several aspects of the immune response, including T-dependent B...... cell activation. While it was originally believed that ICAM-1 played a purely adhesive role, recent evidence suggests that it can itself transduce biochemical signals. We demonstrate that cross-linking of ICAM-1 results in the up-regulation of class II major histocompatibility complex, and we...... investigate the biochemical mechanism for the signaling role of ICAM-1. We show that cross-linking of ICAM-1 on the B lymphoma line A20 induces an increase in tyrosine phosphorylation of several cellular proteins, including the Src family kinase p53/p56(lyn). In vitro kinase assays showed that Lyn kinase...

  12. Evidence for idiotypic- and antiidiotypic B-B cellular interaction with the use of cloned antiidiotypic B cell line.

    Science.gov (United States)

    Bitoh, S; Fujimoto, S; Yamamoto, H

    1990-03-15

    Immunization of BALB/c mice with MOPC104E myeloma protein induces antiidiotypic B lymphocytes that have Id-specific enhancing activity on antibody production. The B-B cell interaction was restricted to both Igh and class II MHC. However, anti-Thy-1 and C-treated splenic B cells were maintained for more than 1 y in a mixture of Con A-stimulated splenocyte culture supernatant and synthetic medium. In applying the long term culture method, we have established a cloned B cell line named B19-1d, B19-1d cells are specific to MOPC104E or J558 cross-reactive Id and they express surface mu, lambda but no Ly-1. B19-1d do not spontaneously secrete Ig but produce them upon stimulation with bacterial LPS. The effect of B19-1d cell line on idiotypic antibody production was tested. Addition of only 10 to 100 B19-1d cells into dextran-immune B cell culture greatly enhanced the Id+ antidextran antibody responses. On the contrary, the antidextran antibody production was suppressed by the higher doses of B19-1d cells. The effective cooperation between dextran-immune B cells and B19-1d cloned B cells was restricted to class II MHC. The role of idiotypic- and antiidiotypic B-B cell interaction in immune regulation and repertoire generation was suggested.

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

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    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.

  14. TGFβ activated kinase 1 (TAK1 at the crossroad of B cell receptor and Toll-like receptor 9 signaling pathways in human B cells.

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    Dániel Szili

    Full Text Available B cell development and activation are regulated by combined signals mediated by the B cell receptor (BCR, receptors for the B-cell activating factor of the tumor necrosis factor family (BAFF-R and the innate receptor, Toll-like receptor 9 (TLR9. However, the underlying mechanisms by which these signals cooperate in human B cells remain unclear. Our aim was to elucidate the key signaling molecules at the crossroads of BCR, BAFF-R and TLR9 mediated pathways and to follow the functional consequences of costimulation.Therefore we stimulated purified human B cells by combinations of anti-Ig, B-cell activating factor of the tumor necrosis factor family (BAFF and the TLR9 agonist, CpG oligodeoxynucleotide. Phosphorylation status of various signaling molecules, B cell proliferation, cytokine secretion, plasma blast generation and the frequency of IgG producing cells were investigated. We have found that BCR induced signals cooperate with BAFF-R- and TLR9-mediated signals at different levels of cell activation. BCR and BAFF- as well as TLR9 and BAFF-mediated signals cooperate at NFκB activation, while BCR and TLR9 synergistically costimulate mitogen activated protein kinases (MAPKs, ERK, JNK and p38. We show here for the first time that the MAP3K7 (TGF beta activated kinase, TAK1 is responsible for the synergistic costimulation of B cells by BCR and TLR9, resulting in an enhanced cell proliferation, plasma blast generation, cytokine and antibody production. Specific inhibitor of TAK1 as well as knocking down TAK1 by siRNA abrogates the synergistic signals. We conclude that TAK1 is a key regulator of receptor crosstalk between BCR and TLR9, thus plays a critical role in B cell development and activation.

  15. 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 proliferation. Thus, B...

  16. Anti-leukemic activity of bortezomib and carfilzomib on B-cell precursor ALL cell lines.

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    Kazuya Takahashi

    Full Text Available Prognosis of childhood acute lymphoblastic leukemia (ALL has been dramatically improved. However, prognosis of the cases refractory to primary therapy is still poor. Recent phase 2 study on the efficacy of combination chemotherapy with bortezomib (BTZ, a proteasome inhibitor, for refractory childhood ALL demonstrated favorable clinical outcomes. However, septic death was observed in over 10% of patients, indicating the necessity of biomarkers that could predict BTZ sensitivity. We investigated in vitro BTZ sensitivity in a large panel of ALL cell lines that acted as a model system for refractory ALL, and found that Philadelphia chromosome-positive (Ph+ ALL, IKZF1 deletion, and biallelic loss of CDKN2A were associated with favorable response. Even in Ph-negative ALL cell lines, IKZF1 deletion and bilallelic loss of CDKN2A were independently associated with higher BTZ sensitivity. BTZ showed only marginal cross-resistance to four representative chemotherapeutic agents (vincristine, dexamethasone, l-asparaginase, and daunorubicin in B-cell precursor-ALL cell lines. To improve the efficacy and safety of proteasome inhibitor combination chemotherapy, we also analyzed the anti-leukemic activity of carfilzomib (CFZ, a second-generation proteasome inhibitor, as a substitute for BTZ. CFZ showed significantly higher activity than BTZ in the majority of ALL cell lines except for the P-glycoprotein-positive t(17;19 ALL cell lines, and IKZF1 deletion was also associated with a favorable response to CFZ treatment. P-glycoprotein inhibitors effectively restored the sensitivity to CFZ, but not BTZ, in P-glycoprotein-positive t(17;19 ALL cell lines. P-glycoprotein overexpressing ALL cell line showed a CFZ-specific resistance, while knockout of P-glycoprotein by genome editing with a CRISPR/Cas9 system sensitized P-glycoprotein-positive t(17;19 ALL cell line to CFZ. These observations suggested that IKZF1 deletion could be a useful biomarker to predict good

  17. B-cell lymphoma 6 protein stimulates oncogenicity of human breast cancer cells

    International Nuclear Information System (INIS)

    Wu, Qiang; Kong, Xiang-jun; Xu, Xiao-chun; Lobie, Peter E; Zhu, Tao; Wu, Zheng-sheng; Liu, Xue; Yan, Hong; He, Yin-huan; Ye, Shan; Cheng, Xing-wang; Zhu, Gui-lu; Wu, Wen-yong; Wang, Xiao-nan

    2014-01-01

    B-cell lymphoma 6 (BCL6) protein, an evolutionarily conserved zinc finger transcription factor, showed to be highly expressed in various human cancers in addition to malignancies in the lymphoid system. This study investigated the role of BCL6 expression in breast cancer and its clinical significance in breast cancer patients. Expression of BCL6 protein was assessed using in situ hybridization and immunohistochemistry in 127 breast cancer patients and 50 patients with breast benign disease as well as in breast cell lines. Expression of BCL6 was restored or knocked down in two breast cancer cell lines (MCF-7 and T47D) using BCL6 cDNA and siRNA, respectively. The phenotypic change of these breast cancer cell lines was assessed using cell viability MTT, Transwell invasion, colony formation, and flow cytometry assays and in a xenograft mice model. Luciferase reporter gene, immunoblot, and qRT-PCR were used to investigate the molecular events after manipulated BCL6 expression in breast cancer cells. BCL6 protein was highly expressed in breast cancer cell lines and tissue specimens and expression of BCL6 protein was associated with disease progression and poor survival of breast cancer patients. In vitro, the forced expression of BCL6 results in increased proliferation, anchorage-independent growth, migration, invasion and survival of breast cancer cell lines, whereas knockdown of BCL6 expression reduced these oncogenic properties of breast cancer cells. Moreover, forced expression of BCL6 increased tumor growth and invasiveness in a nude mouse xenograft model. At the gene level, BCL6 was a target gene of miR-339-5p. Expression of BCL6 induced expression of CXCR4 and cyclinD1 proteins. The current study demonstrated the oncogenic property of BCL6 in breast cancer and further study could target BCL6 as a novel potential therapeutic strategy for breast cancer

  18. A novel cell growth-promoting factor identified in a B cell leukemia cell line, BALL-1

    International Nuclear Information System (INIS)

    Dao, T.; Holan, V.; Minowada, J.

    1993-01-01

    A novel leukemia cell growth-promoting activity has been identified in the culture supernatant from a human B cell leukemia cell line, BALL-1. The supernatant from unstimulated cultures of the BALL-1 cells significantly promoted the growth of 16 out of 24 leukemia/lymphoma cell lines of different lineages (T, B and non-lymphoid) in a minimal concentration of fetal bovine serum (FBS), and 5 out of 12 cases of fresh leukemia cells in FBS-free medium. The growth-promoting sieve filtration and dialysis. The MW of the factor was less than 10 kDa. The growth-promoting activity was heat and acid stable and resistant to trypsin treatment. The factor isolated from the BALL-1 supernatant was distinct from known polypeptide growth factors with MW below 10 kDa, such as epidermal growth factor, transforming growth factor α, insulin-like growth factor I (IGF-I), IGF-II and insulin, as determine by specific antibodies and by cell-growth-promoting tests. The factor is the BALL-1 supernatant did not promote the proliferation of normal human fresh peripheral blood lymphocytes or mouse fibroblast cell line, BALB/C 3T3. In addition to the BALL-1 supernatant, a similar growth-promoting activity was found in the culture supernatant from 13 of 17 leukemia/lymphoma cell lines tested. The activity in these culture supernatant promoted the growth of leukemia/lymphoma cell lines in autocrine and/or paracrine fashions. These observations suggest that the low MW cell growth-promoting activity found in the BALL-1 culture supernatant is mediated by a novel factor which may be responsible for the clonal expansion of particular leukemic clones. (author)

  19. Overexpression of the human BCL-2 gene product results in growth enhancement of Epstein-Barr virus-immortalized B cells

    International Nuclear Information System (INIS)

    Tsujimoto, Yoshihide

    1989-01-01

    The biological activity of the human BCL-2 gene product was analyzed in an Epstein-Barr virus (EBV)-infected human lymphoblastoid B-cell line transfected with BCL-2 sequences driven by the simian virus 40 promoter and enhancer. Overproduction of the BCL-2 protein conferred a selective growth advantage to the EBV-infected B cells as compared with control transfectants in low-serum medium and also after seeding at limiting dilution but did not render the cells tumorigenic in athymic nude mice. This growth enhancement was also seen in cells transfected with the BCL-2 gene with its own promoter juxtaposed to the immunoglobulin heavy chain gene enhancer, which represents the translocated form of the BCL-2 gene observed in follicular lymphomas with the t(14;18) translocation. The growth advantage of EBV-infected B cells overproducing the BCL-2 protein is neither due to the enhanced growth factor production nor due to an enhanced sensitivity of the BCL-2 transfectants to interleukins 1 or 6, although both lymphokines are known to stimulate proliferation of EBV-infected B-cell lines. The growth advantage of EBV-infected B-cell lines. The growth advantage of EBV-infected B cells by overproduction of the BCL-2 protein suggests the direct involvement of the BCL-2 gene product in the pathogenesis of follicular lymphoma

  20. Molecular Cytogenetic Characterization Identified the Murine B-Cell Lymphoma Cell Line A-20 as a Model for Sporadic Burkitt's Lymphoma.

    Science.gov (United States)

    Guja, Karolina; Liehr, Thomas; Rincic, Martina; Kosyakova, Nadezda; Hussein Azawi, Shaymaa S

    2017-11-01

    Here, we report the first molecular cytogenetic characterization of the BALB/cAnN mouse derived B-cell non-Hodgkin lymphoma (B-cell NHL) cell lines A-20. Even though previously used as a model for testing of, for example, dexametason, up to present, no data in the genetic properties of A-20 were available. The present study closed this gap and provides evidence that A-20 is a model for B-cell NHL subgroup sporadic Burkitt's lymphoma. C-myc oncogene is involved in a translocation and copy number alterations as gain of murine 14q material could be observed. Interestingly, the cell line showed the karyotype 39,X,-X or -Y,t(2;15)(qE5;qD2),del(6)(qB3qC3),del(9)(qA3qA4),dup(14)(qE1qE4) in ~95% of the cells, being exceptionally stable for cell lines being established 38 years ago. Still, ~5% of the cells showed polyploidization followed by chromothripsis. It remains to be determined if this can be observed also in other cell lines, just has not been reported yet, and/or if it is a unique feature of A-20. Overall, finally here, the necessary genetic data to identify A-20 as a model for human sporadic Burkitt's lymphoma are provided.

  1. Effect of TH-lines and clones on the growth and differentiation of B cell clones in microculture.

    Science.gov (United States)

    Kotloff, D B; Cebra, J J

    1988-02-01

    Antibody isotype expression by B cell clones was analyzed using in vitro microcultures containing low numbers of hapten-gelatin-enriched B cells and higher numbers of hemocyanin-specific helper T cell lines or clones. Twenty-eight to sixty-three percent of clones grown in microculture with haptenated hemocyanin and T cells from established lines expressed IgG and/or IgA isotypes in random mixtures, almost always accompanied by IgM. Helper T cells from hemocyanin-specific clones also supported the expression of non-IgM isotypes by the B cell clones, suggesting that a single specificity of T cell can provide sufficient growth and differentiation factors for the display of isotype switching. A positive correlation between the antibody output of clones and the expression of non-IgM isotypes indicated that the switching process may be associated with cell division. Although memory B cells that give clones expressing IgG and/or IgA in the absence of IgM are also enriched on haptenated gelatin, they are not stimulable under conditions of this microculture assay.

  2. 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.

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

    Science.gov (United States)

    Carraz, Maëlle; Lavergne, Cédric; Jullian, Valérie; Wright, Michel; Gairin, Jean Edouard; Gonzales de la Cruz, Mercedes; Bourdy, Geneviève

    2015-05-26

    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. 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: Chiclayo (Lambayeque department, Chiclayo province) and Huaraz (Ancash department, Huaraz province). Altogether 51 species were collected and 61 ethanol extracts were prepared to be tested. Biological assessment: All extracts were first assessed against the HCC cell line Hep3B according a 3-step multi-parametric phenotypic assay. It included 1) the evaluation of phenotypic changes on cells by light microscopy, 2) the measurement of the antiproliferative activity and 3) the analysis of the cytoskeleton and mitosis by immunofluorescence. Best extracts were further assessed against other HCC cell lines HepG2, PLC/PRF/5 and SNU-182 and their toxicity measured in vitro on primary human hepatocytes. Ethnopharmacological survey: Some of the species collected had a high reputation spreading over the surveyed locations for treating liver problems, i.e. Baccharis genistelloides, Bejaria aestuans, Centaurium pulchellum, Desmodium molliculum, Dipsacus fullonum, Equisetum bogotense, Gentianella spp., Krameria lapacea, Otholobium spp., Schkuhria pinnata, Taraxacum officinale. Hep3B evaluation: Fourteen extracts from 13 species (Achyrocline alata, Ambrosia arborescens, Baccharis latifolia, Hypericum laricifolium, Krameria lappacea, Niphidium crassifolium, Ophryosporus chilca, Orthrosanthus chimboracensis, Otholobium pubescens, Passiflora ligularis, Perezia coerulescens, Perezia multiflora and Schkuhria pinnata) showed a significant antiproliferative activity against Hep3B cells (IC50≤ 50µg/mL). This was associated with a lack of toxicity on primary

  4. Expression of Immunoglobulin Receptors with Distinctive Features Indicating Antigen Selection by Marginal Zone B Cells from Human Spleen

    Science.gov (United States)

    Colombo, Monica; Cutrona, Giovanna; Reverberi, Daniele; Bruno, Silvia; Ghiotto, Fabio; Tenca, Claudya; Stamatopoulos, Kostas; Hadzidimitriou, Anastasia; Ceccarelli, Jenny; Salvi, Sandra; Boccardo, Simona; Calevo, Maria Grazia; De Santanna, Amleto; Truini, Mauro; Fais, Franco; Ferrarini, Manlio

    2013-01-01

    Marginal zone (MZ) B cells, identified as surface (s)IgMhighsIgDlowCD23low/−CD21+CD38− B cells, were purified from human spleens, and the features of their V(D)J gene rearrangements were investigated and compared with those of germinal center (GC), follicular mantle (FM) and switched memory (SM) B cells. Most MZ B cells were CD27+ and exhibited somatic hypermutations (SHM), although to a lower extent than SM B cells. Moreover, among MZ B-cell rearrangements, recurrent sequences were observed, some of which displayed intraclonal diversification. The same diversifying sequences were detected in very low numbers in GC and FM B cells and only when a highly sensitive, gene-specific polymerase chain reaction was used. This result indicates that MZ B cells could expand and diversify in situ and also suggested the presence of a number of activation-induced cytidine deaminase (AID)-expressing B cells in the MZ. The notion of antigen-driven expansion/selection in situ is further supported by the VH CDR3 features of MZ B cells with highly conserved amino acids at specific positions and by the finding of shared (“stereotyped”) sequences in two different spleens. Collectively, the data are consistent with the notion that MZ B cells are a special subset selected by in situ antigenic stimuli. PMID:23877718

  5. B cell signatures of BCWD-resistant and susceptible lines of rainbow trout: a shift towards more EBF-expressing progenitors and fewer mature B cells in resistant animals.

    Science.gov (United States)

    Zwollo, Patty; Ray, Jocelyn C; Sestito, Michael; Kiernan, Elizabeth; Wiens, Gregory D; Kaattari, Steve; StJacques, Brittany; Epp, Lidia

    2015-01-01

    Bacterial cold water disease (BCWD) is a chronic disease of rainbow trout, and is caused by the Gram-negative bacterium Flavobacterium psychrophilum (Fp), a common aquaculture pathogen. The National Center for Cool and Cold Water Aquaculture has bred two genetic lines of rainbow trout: a line of Fp-resistant trout (ARS-Fp-R or R-line trout) and a line of susceptible trout (ARS-Fp-S, or S-line). Little is known about how phenotypic selection alters immune response parameters or how such changes relate to genetic disease resistance. Herein, we quantify interindividual variation in the distribution and abundance of B cell populations (B cell signatures) and examine differences between genetic lines of naive animals. There are limited trout-specific cell surface markers currently available to resolve B cell subpopulations and thus we developed an alternative approach based on detection of differentially expressed transcription factors and intracellular cytokines. B cell signatures were compared between R-line and S-line trout by flow cytometry using antibodies against transcription factors early B cell factor-1 (EBF1) and paired domain box protein Pax5, the pro-inflammatory cytokine IL-1β, and the immunoglobulin heavy chain mu. R-line trout had higher percentages of EBF(+) B myeloid/ progenitor and pre-B cells in PBL, anterior and posterior kidney tissues compared to S-line trout. The opposite pattern was detected in more mature B cell populations: R-line trout had lower percentages of both IgM(+) mature B cells and IgM-secreting cells in anterior kidney and PBL compared to S-line trout. In vitro LPS-activation studies of PBL and spleen cell cultures revealed no significant induction differences between R-line and S-line trout. Together, our findings suggest that selective resistance to BCWD may be associated with shifts in naive animal developmental lineage commitment that result in decreased B lymphopoiesis and increased myelopoiesis in BCWD resistant trout relative

  6. Authentication of primordial characteristics of the CLBL-1 cell line prove the integrity of a canine B-cell lymphoma in a murine in vivo model.

    Directory of Open Access Journals (Sweden)

    Barbara C Rütgen

    Full Text Available Cell lines are key tools in cancer research allowing the generation of neoplasias in animal models resembling the initial tumours able to mimic the original neoplasias closely in vivo. Canine lymphoma is the major hematopoietic malignancy in dogs and considered as a valuable spontaneous large animal model for human Non-Hodgkin's Lymphoma (NHL. Herein we describe the establishment and characterisation of an in vivo model using the canine B-cell lymphoma cell line CLBL-1 analysing the stability of the induced tumours and the ability to resemble the original material. CLBL-1 was injected into Rag2(-/-γ(c (-/- mice. The generated tumor material was analysed by immunophenotyping and histopathology and used to establish the cell line CLBL-1M. Both cell lines were karyotyped for detection of chromosomal aberrations. Additionally, CLBL-1 was stimulated with IL-2 and DSP30 as described for primary canine B-cell lymphomas and NHL to examine the stimulatory effect on cell proliferation. CLBL-1 in vivo application resulted in lymphoma-like disease and tumor formation. Immunophenotypic analysis of tumorous material showed expression of CD45(+, MHCII(+, CD11a(+ and CD79αcy(+. PARR analysis showed positivity for IgH indicating a monoclonal character. These cytogenetic, molecular, immunophenotypical and histological characterisations of the in vivo model reveal that the induced tumours and thereof generated cell line resemble closely the original material. After DSP30 and IL-2 stimulation, CLBL-1 showed to respond in the same way as primary material. The herein described CLBL-1 in vivo model provides a highly stable tool for B-cell lymphoma research in veterinary and human medicine allowing various further in vivo studies.

  7. Authentication of primordial characteristics of the CLBL-1 cell line prove the integrity of a canine B-cell lymphoma in a murine in vivo model.

    Science.gov (United States)

    Rütgen, Barbara C; Willenbrock, Saskia; Reimann-Berg, Nicola; Walter, Ingrid; Fuchs-Baumgartinger, Andrea; Wagner, Siegfried; Kovacic, Boris; Essler, Sabine E; Schwendenwein, Ilse; Nolte, Ingo; Saalmüller, Armin; Murua Escobar, Hugo

    2012-01-01

    Cell lines are key tools in cancer research allowing the generation of neoplasias in animal models resembling the initial tumours able to mimic the original neoplasias closely in vivo. Canine lymphoma is the major hematopoietic malignancy in dogs and considered as a valuable spontaneous large animal model for human Non-Hodgkin's Lymphoma (NHL). Herein we describe the establishment and characterisation of an in vivo model using the canine B-cell lymphoma cell line CLBL-1 analysing the stability of the induced tumours and the ability to resemble the original material. CLBL-1 was injected into Rag2(-/-)γ(c) (-/-) mice. The generated tumor material was analysed by immunophenotyping and histopathology and used to establish the cell line CLBL-1M. Both cell lines were karyotyped for detection of chromosomal aberrations. Additionally, CLBL-1 was stimulated with IL-2 and DSP30 as described for primary canine B-cell lymphomas and NHL to examine the stimulatory effect on cell proliferation. CLBL-1 in vivo application resulted in lymphoma-like disease and tumor formation. Immunophenotypic analysis of tumorous material showed expression of CD45(+), MHCII(+), CD11a(+) and CD79αcy(+). PARR analysis showed positivity for IgH indicating a monoclonal character. These cytogenetic, molecular, immunophenotypical and histological characterisations of the in vivo model reveal that the induced tumours and thereof generated cell line resemble closely the original material. After DSP30 and IL-2 stimulation, CLBL-1 showed to respond in the same way as primary material. The herein described CLBL-1 in vivo model provides a highly stable tool for B-cell lymphoma research in veterinary and human medicine allowing various further in vivo studies.

  8. Atypical memory B cells in human chronic infectious diseases: An interim report.

    Science.gov (United States)

    Portugal, Silvia; Obeng-Adjei, Nyamekye; Moir, Susan; Crompton, Peter D; Pierce, Susan K

    2017-11-01

    Immunological memory is a remarkable phenomenon in which survival of an initial infection by a pathogen leads to life-long protection from disease upon subsequent exposure to that same pathogen. For many infectious diseases, long-lived protective humoral immunity is induced after only a single infection in a process that depends on the generation of memory B cells (MBCs) and long-lived plasma cells. However, over the past decade it has become increasingly evident that many chronic human infectious diseases to which immunity is not readily established, including HIV-AIDS, malaria and TB, are associated with fundamental alterations in the composition and functionality of MBC compartments. A common feature of these diseases appears to be a large expansion of what have been termed exhausted B cells, tissue-like memory B cells or atypical memory B cells (aMBCs) that, for simplicity's sake, we refer to here as aMBCs. It has been suggested that chronic immune activation and inflammation drive the expansion of aMBCs and that in some way aMBCs contribute to deficiencies in the acquisition of immunity in chronic infectious diseases. Although aMBCs are heterogeneous both within individuals and between diseases, they have several features in common including low expression of the cell surface markers that define classical MBCs in humans including CD21 and CD27 and high expression of genes not usually expressed by classical MBCs including T-bet, CD11c and a variety of inhibitory receptors, notably members of the FcRL family. Another distinguishing feature is their greatly diminished ability to be stimulated through their B cell receptors to proliferate, secrete cytokines or produce antibodies. In this review, we describe our current understanding of the phenotypic markers of aMBCs, their specificity in relation to the disease-causing pathogen, their functionality, the drivers of their expansion in chronic infections and their life span. We briefly summarize the features of a

  9. Ontogeny of human IgE?expressing B cells and plasma cells

    OpenAIRE

    Ramadani, F.; Bowen, H.; Upton, N.; Hobson, P. S.; Chan, Y.?C.; Chen, J.?B.; Chang, T. W.; McDonnell, J. M.; Sutton, B. J.; Fear, D. J.; Gould, H. J.

    2016-01-01

    BACKGROUND: IgE-expressing (IgE+) plasma cells (PCs) provide a continuous source of allergen specific IgE that is central to allergic responses. The extreme sparsity of IgE+ cells in vivo has confined their study almost entirely to mouse models.OBJECTIVE: To characterise the development pathway of human IgE+ PCs and to determine the ontogeny of human IgE+ PCs.METHODS: To generate human IgE+ cells we cultured tonsil B cells with IL-4 and anti-CD40. Using FACS and RT-PCR we examined the phenoty...

  10. Altered distribution of peripheral blood memory B cells in humans chronically infected with Trypanosoma cruzi.

    Science.gov (United States)

    Fernández, Esteban R; Olivera, Gabriela C; Quebrada Palacio, Luz P; González, Mariela N; Hernandez-Vasquez, Yolanda; Sirena, Natalia María; Morán, María L; Ledesma Patiño, Oscar S; Postan, Miriam

    2014-01-01

    Numerous abnormalities of the peripheral blood T cell compartment have been reported in human chronic Trypanosoma cruzi infection and related to prolonged antigenic stimulation by persisting parasites. Herein, we measured circulating lymphocytes of various phenotypes based on the differential expression of CD19, CD4, CD27, CD10, IgD, IgM, IgG and CD138 in a total of 48 T. cruzi-infected individuals and 24 healthy controls. Infected individuals had decreased frequencies of CD19+CD27+ cells, which positively correlated with the frequencies of CD4+CD27+ cells. The contraction of CD19+CD27+ cells was comprised of IgG+IgD-, IgM+IgD- and isotype switched IgM-IgD- memory B cells, CD19+CD10+CD27+ B cell precursors and terminally differentiated CD19+CD27+CD138+ plasma cells. Conversely, infected individuals had increased proportions of CD19+IgG+CD27-IgD- memory and CD19+IgM+CD27-IgD+ transitional/naïve B cells. These observations prompted us to assess soluble CD27, a molecule generated by the cleavage of membrane-bound CD27 and used to monitor systemic immune activation. Elevated levels of serum soluble CD27 were observed in infected individuals with Chagas cardiomyopathy, indicating its potentiality as an immunological marker for disease progression in endemic areas. In conclusion, our results demonstrate that chronic T. cruzi infection alters the distribution of various peripheral blood B cell subsets, probably related to the CD4+ T cell deregulation process provoked by the parasite in humans.

  11. Altered distribution of peripheral blood memory B cells in humans chronically infected with Trypanosoma cruzi.

    Directory of Open Access Journals (Sweden)

    Esteban R Fernández

    Full Text Available Numerous abnormalities of the peripheral blood T cell compartment have been reported in human chronic Trypanosoma cruzi infection and related to prolonged antigenic stimulation by persisting parasites. Herein, we measured circulating lymphocytes of various phenotypes based on the differential expression of CD19, CD4, CD27, CD10, IgD, IgM, IgG and CD138 in a total of 48 T. cruzi-infected individuals and 24 healthy controls. Infected individuals had decreased frequencies of CD19+CD27+ cells, which positively correlated with the frequencies of CD4+CD27+ cells. The contraction of CD19+CD27+ cells was comprised of IgG+IgD-, IgM+IgD- and isotype switched IgM-IgD- memory B cells, CD19+CD10+CD27+ B cell precursors and terminally differentiated CD19+CD27+CD138+ plasma cells. Conversely, infected individuals had increased proportions of CD19+IgG+CD27-IgD- memory and CD19+IgM+CD27-IgD+ transitional/naïve B cells. These observations prompted us to assess soluble CD27, a molecule generated by the cleavage of membrane-bound CD27 and used to monitor systemic immune activation. Elevated levels of serum soluble CD27 were observed in infected individuals with Chagas cardiomyopathy, indicating its potentiality as an immunological marker for disease progression in endemic areas. In conclusion, our results demonstrate that chronic T. cruzi infection alters the distribution of various peripheral blood B cell subsets, probably related to the CD4+ T cell deregulation process provoked by the parasite in humans.

  12. Human B cells induce dendritic cell maturation and favour Th2 polarization by inducing OX-40 ligand

    Science.gov (United States)

    Maddur, Mohan S.; Sharma, Meenu; Hegde, Pushpa; Stephen-Victor, Emmanuel; Pulendran, Bali; Kaveri, Srini V.; Bayry, Jagadeesh

    2015-01-01

    Dendritic cells (DCs) play a critical role in immune homeostasis by regulating the functions of various immune cells, including T and B cells. Notably, DCs also undergo education on reciprocal signalling by these immune cells and environmental factors. Various reports demonstrated that B cells have profound regulatory functions, although only few reports have explored the regulation of human DCs by B cells. Here we demonstrate that activated but not resting B cells induce maturation of DCs with distinct features to polarize Th2 cells that secrete interleukin (IL)-5, IL-4 and IL-13. B-cell-induced maturation of DCs is contact dependent and implicates signalling of B-cell activation molecules CD69, B-cell-activating factor receptor, and transmembrane activator and calcium-modulating cyclophilin ligand interactor. Mechanistically, differentiation of Th2 cells by B-cell-matured DCs is dependent on OX-40 ligand. Collectively, our results suggest that B cells have the ability to control their own effector functions by enhancing the ability of human DCs to mediate Th2 differentiation. PMID:24910129

  13. The role of the interleukin-10 subfamily members in immunoglobulin production by human B cells

    DEFF Research Database (Denmark)

    Hummelshoj, L; Ryder, L P; Poulsen, Lars K.

    2006-01-01

    Interleukin (IL)-10 has been shown to have various effects on B cells, including positively affecting the production of immunoglobulin A (IgA) and IgG. Several human IL-10-related molecules have been identified. These include IL-19, IL-20, IL-22, IL-24, IL-26, IL-28 and IL-29. To determine...... the effects of the IL-10 analogues on the class switch recombination in B cells, we analysed Ig production from naïve B cells stimulated with these cytokines in the presence of anti-CD40. None of the cytokines were found to induce Ig production by themselves in the presence of anti-CD40 Ab. However, all...... cytokines inhibited the production of IgA and IgG induced by anti-CD40 Ab alone. In combination with anti-CD40 Ab and IL-4, IgG4 were inhibited in cultures stimulated with IL-20, IL-22, IL-26, IL-28 and IL-29 compared with IL-4 and anti-CD40 Ab alone, whereas all IL-10 analogues increased the production...

  14. Neurotoxic, cytotoxic, apoptotic and antiproliferative effects of some marine algae extracts on the NA2B cell line.

    Science.gov (United States)

    Kurt, O; Özdal-Kurt, F; Akçora, C M; Özkut, M; Tuğlu, M I

    2018-02-01

    Oxidative stress contributes to cancer pathologies and to apoptosis. Marine algae exhibit cytotoxic, antiproliferative and apoptotic effects; their metabolites have been used to treat many types of cancer. We investigated in culture extracts of Petalonia fascia, Jania longifurca and Halimeda tuna to determine their effects on mouse neuroblastoma cell line, NA2B. NA2B cells were treated with algae extracts, and the survival and proliferation of NA2B cells were assessed using the 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay. The effects of algae extracts on oxidative stress in NA2B cells also were investigated using nitric oxide synthase (NOS) immunocytochemistry and apoptosis was assessed using terminal deoxynucleotidyl transferase dUTP nick end labeling. We observed significant neurite inhibition with moderate damage by the neurotoxicity-screening test (NST) at IC 50 dilutions of the extracts. MTT demonstrated that J. longifurca extracts were more toxic than P. fascia and H. tuna extracts. We found an increase of endothelial and inducible NOS immunostaining for oxidative stress and TUNEL analysis revealed increased apoptosis after application of extract. Our findings suggest that the algae we tested may have potential use for treatment of cancer.

  15. Extinct type of human parvovirus B19 persists in tonsillar B cells

    OpenAIRE

    Pyöriä, Lari; Toppinen, Mari; Mantyla, Elina; Hedman, Lea; Aaltonen, Leena-Maija; Vihinen-Ranta, Maija; Ilmarinen, Taru; Soderlund-Venermo, Maria; Hedman, Klaus; Perdomo, Maria

    2017-01-01

    Parvovirus B19 (B19V) DNA persists lifelong in human tissues, but the cell type harbouring it remains unclear. We here explore B19V DNA distribution in B, T and monocyte cell lineages of recently excised tonsillar tissues from 77 individuals with an age range of 2?69 years. We show that B19V DNA is most frequent and abundant among B cells, and within them we find a B19V genotype that vanished from circulation >40 years ago. Since re-infection or re-activation are unlikely with this virus type...

  16. High Inter-Individual Diversity of Point Mutations, Insertions, and Deletions in Human Influenza Virus Nucleoprotein-Specific Memory B Cells.

    Directory of Open Access Journals (Sweden)

    Sven Reiche

    Full Text Available The diversity of virus-specific antibodies and of B cells among different individuals is unknown. Using single-cell cloning of antibody genes, we generated recombinant human monoclonal antibodies from influenza nucleoprotein-specific memory B cells in four adult humans with and without preceding influenza vaccination. We examined the diversity of the antibody repertoires and found that NP-specific B cells used numerous immunoglobulin genes. The heavy chains (HCs originated from 26 and the kappa light chains (LCs from 19 different germ line genes. Matching HC and LC chains gave rise to 43 genetically distinct antibodies that bound influenza NP. The median lengths of the CDR3 of the HC, kappa and lambda LC were 14, 9 and 11 amino acids, respectively. We identified changes at 13.6% of the amino acid positions in the V gene of the antibody heavy chain, at 8.4% in the kappa and at 10.6 % in the lambda V gene. We identified somatic insertions or deletions in 8.1% of the variable genes. We also found several small groups of clonal relatives that were highly diversified. Our findings demonstrate broadly diverse memory B cell repertoires for the influenza nucleoprotein. We found extensive variation within individuals with a high number of point mutations, insertions, and deletions, and extensive clonal diversification. Thus, structurally conserved proteins can elicit broadly diverse and highly mutated B-cell responses.

  17. Development Refractoriness of MLL-Rearranged Human B Cell Acute Leukemias to Reprogramming into Pluripotency

    Directory of Open Access Journals (Sweden)

    Alvaro Muñoz-López

    2016-10-01

    Full Text Available Induced pluripotent stem cells (iPSCs are a powerful tool for disease modeling. They are routinely generated from healthy donors and patients from multiple cell types at different developmental stages. However, reprogramming leukemias is an extremely inefficient process. Few studies generated iPSCs from primary chronic myeloid leukemias, but iPSC generation from acute myeloid or lymphoid leukemias (ALL has not been achieved. We attempted to generate iPSCs from different subtypes of B-ALL to address the developmental impact of leukemic fusion genes. OKSM(L-expressing mono/polycistronic-, retroviral/lentiviral/episomal-, and Sendai virus vector-based reprogramming strategies failed to render iPSCs in vitro and in vivo. Addition of transcriptomic-epigenetic reprogramming “boosters” also failed to generate iPSCs from B cell blasts and B-ALL lines, and when iPSCs emerged they lacked leukemic fusion genes, demonstrating non-leukemic myeloid origin. Conversely, MLL-AF4-overexpressing hematopoietic stem cells/B progenitors were successfully reprogrammed, indicating that B cell origin and leukemic fusion gene were not reprogramming barriers. Global transcriptome/DNA methylome profiling suggested a developmental/differentiation refractoriness of MLL-rearranged B-ALL to reprogramming into pluripotency.

  18. BMP-6 inhibits growth of mature human B cells; induction of Smad phosphorylation and upregulation of Id1

    Directory of Open Access Journals (Sweden)

    Kersten Christian

    2005-05-01

    Full Text Available Abstract Background Bone morphogenetic proteins (BMPs belong to the TGF-β superfamily and are secreted proteins with pleiotropic roles in many different cell types. A potential role of BMP-6 in the immune system has been implied by various studies of malignant and rheumatoid diseases. In the present study, we explored the role of BMP-6 in normal human peripheral blood B cells. Results The B cells were found to express BMP type I and type II receptors and BMP-6 rapidly induced phosphorylation of Smad1/5/8. Furthermore, Smad-phosphorylation was followed by upregulation of Id1 mRNA and Id1 protein, whereas Id2 and Id3 expression was not affected. Furthermore, we found that BMP-6 had an antiproliferative effect both in naïve (CD19+CD27- and memory B cells (CD19+CD27+ stimulated with anti-IgM alone or the combined action of anti-IgM and CD40L. Additionally, BMP-6 induced cell death in activated memory B cells. Importantly, the antiproliferative effect of BMP-6 in B-cells was completely neutralized by the natural antagonist, noggin. Furthermore, B cells were demonstrated to upregulate BMP-6 mRNA upon stimulation with anti-IgM. Conclusion In mature human B cells, BMP-6 inhibited cell growth, and rapidly induced phosphorylation of Smad1/5/8 followed by an upregulation of Id1.

  19. Piperlongumine inhibits the proliferation and survival of B-cell acute lymphoblastic leukemia cell lines irrespective of glucocorticoid resistance

    Energy Technology Data Exchange (ETDEWEB)

    Han, Seong-Su, E-mail: seong-su-han@uiowa.edu [Division of Pediatric Hematology-Oncology, University of Iowa Carver College of Medicine, Iowa City, IA (United States); Han, Sangwoo [Health and Human Physiology, University of Iowa Carver College of Medicine, Iowa City, IA (United States); Kamberos, Natalie L. [Division of Pediatric Hematology-Oncology, University of Iowa Carver College of Medicine, Iowa City, IA (United States)

    2014-09-26

    Highlights: • PL inhibits the proliferation of B-ALL cell lines irrespective of GC-resistance. • PL selectively kills B-ALL cells by increasing ROS, but not normal counterpart. • PL does not sensitize majority of B-ALL cells to DEX. • PL represses the network of constitutively activated TFs and modulates their target genes. • PL may serve as a new therapeutic molecule for GC-resistant B-ALL. - Abstract: Piperlongumine (PL), a pepper plant alkaloid from Piper longum, has anti-inflammatory and anti-cancer properties. PL selectively kills both solid and hematologic cancer cells, but not normal counterparts. Here we evaluated the effect of PL on the proliferation and survival of B-cell acute lymphoblastic leukemia (B-ALL), including glucocorticoid (GC)-resistant B-ALL. Regardless of GC-resistance, PL inhibited the proliferation of all B-ALL cell lines, but not normal B cells, in a dose- and time-dependent manner and induced apoptosis via elevation of ROS. Interestingly, PL did not sensitize most of B-ALL cell lines to dexamethasone (DEX). Only UoC-B1 exhibited a weak synergistic effect between PL and DEX. All B-ALL cell lines tested exhibited constitutive activation of multiple transcription factors (TFs), including AP-1, MYC, NF-κB, SP1, STAT1, STAT3, STAT6 and YY1. Treatment of the B-ALL cells with PL significantly downregulated these TFs and modulated their target genes. While activation of AURKB, BIRC5, E2F1, and MYB mRNA levels were significantly downregulated by PL, but SOX4 and XBP levels were increased by PL. Intriguingly, PL also increased the expression of p21 in B-ALL cells through a p53-independent mechanism. Given that these TFs and their target genes play critical roles in a variety of hematological malignancies, our findings provide a strong preclinical rationale for considering PL as a new therapeutic agent for the treatment of B-cell malignancies, including B-ALL and GC-resistant B-ALL.

  20. Piperlongumine inhibits the proliferation and survival of B-cell acute lymphoblastic leukemia cell lines irrespective of glucocorticoid resistance

    International Nuclear Information System (INIS)

    Han, Seong-Su; Han, Sangwoo; Kamberos, Natalie L.

    2014-01-01

    Highlights: • PL inhibits the proliferation of B-ALL cell lines irrespective of GC-resistance. • PL selectively kills B-ALL cells by increasing ROS, but not normal counterpart. • PL does not sensitize majority of B-ALL cells to DEX. • PL represses the network of constitutively activated TFs and modulates their target genes. • PL may serve as a new therapeutic molecule for GC-resistant B-ALL. - Abstract: Piperlongumine (PL), a pepper plant alkaloid from Piper longum, has anti-inflammatory and anti-cancer properties. PL selectively kills both solid and hematologic cancer cells, but not normal counterparts. Here we evaluated the effect of PL on the proliferation and survival of B-cell acute lymphoblastic leukemia (B-ALL), including glucocorticoid (GC)-resistant B-ALL. Regardless of GC-resistance, PL inhibited the proliferation of all B-ALL cell lines, but not normal B cells, in a dose- and time-dependent manner and induced apoptosis via elevation of ROS. Interestingly, PL did not sensitize most of B-ALL cell lines to dexamethasone (DEX). Only UoC-B1 exhibited a weak synergistic effect between PL and DEX. All B-ALL cell lines tested exhibited constitutive activation of multiple transcription factors (TFs), including AP-1, MYC, NF-κB, SP1, STAT1, STAT3, STAT6 and YY1. Treatment of the B-ALL cells with PL significantly downregulated these TFs and modulated their target genes. While activation of AURKB, BIRC5, E2F1, and MYB mRNA levels were significantly downregulated by PL, but SOX4 and XBP levels were increased by PL. Intriguingly, PL also increased the expression of p21 in B-ALL cells through a p53-independent mechanism. Given that these TFs and their target genes play critical roles in a variety of hematological malignancies, our findings provide a strong preclinical rationale for considering PL as a new therapeutic agent for the treatment of B-cell malignancies, including B-ALL and GC-resistant B-ALL

  1. HSC extrinsic sex-related and intrinsic autoimmune disease-related human B-cell variation is recapitulated in humanized mice.

    Science.gov (United States)

    Borsotti, Chiara; Danzl, Nichole M; Nauman, Grace; Hölzl, Markus A; French, Clare; Chavez, Estefania; Khosravi-Maharlooei, Mohsen; Glauzy, Salome; Delmotte, Fabien R; Meffre, Eric; Savage, David G; Campbell, Sean R; Goland, Robin; Greenberg, Ellen; Bi, Jing; Satwani, Prakash; Yang, Suxiao; Bathon, Joan; Winchester, Robert; Sykes, Megan

    2017-10-24

    B cells play a major role in antigen presentation and antibody production in the development of autoimmune diseases, and some of these diseases disproportionally occur in females. Moreover, immune responses tend to be stronger in female vs male humans and mice. Because it is challenging to distinguish intrinsic from extrinsic influences on human immune responses, we used a personalized immune (PI) humanized mouse model, in which immune systems were generated de novo from adult human hematopoietic stem cells (HSCs) in immunodeficient mice. We assessed the effect of recipient sex and of donor autoimmune diseases (type 1 diabetes [T1D] and rheumatoid arthritis [RA]) on human B-cell development in PI mice. We observed that human B-cell levels were increased in female recipients regardless of the source of human HSCs or the strain of immunodeficient recipient mice. Moreover, mice injected with T1D- or RA-derived HSCs displayed B-cell abnormalities compared with healthy control HSC-derived mice, including altered B-cell levels, increased proportions of mature B cells and reduced CD19 expression. Our study revealed an HSC-extrinsic effect of recipient sex on human B-cell reconstitution. Moreover, the PI humanized mouse model revealed HSC-intrinsic defects in central B-cell tolerance that recapitulated those in patients with autoimmune diseases. These results demonstrate the utility of humanized mouse models as a tool to better understand human immune cell development and regulation.

  2. Human invariant NKT cell subsets differentially promote differentiation, antibody production, and T cell stimulation by B cells in vitro.

    OpenAIRE

    O'REILLY, VINCENT

    2013-01-01

    PUBLISHED Invariant NK T (iNKT) cells can provide help for B cell activation and Ab production. Because B cells are also capable of cytokine production, Ag presentation, and T cell activation, we hypothesized that iNKT cells will also influence these activities. Furthermore, subsets of iNKT cells based on CD4 and CD8 expression that have distinct functional activities may differentially affect B cell functions. We investigated the effects of coculturing expanded human CD4(+), CD8α(+), and ...

  3. Comprehensive microRNA profiling in B-cells of human centenarians by massively parallel sequencing

    Directory of Open Access Journals (Sweden)

    Gombar Saurabh

    2012-07-01

    Full Text Available Abstract Background MicroRNAs (miRNAs are small, non-coding RNAs that regulate gene expression and play a critical role in development, homeostasis, and disease. Despite their demonstrated roles in age-associated pathologies, little is known about the role of miRNAs in human aging and longevity. Results We employed massively parallel sequencing technology to identify miRNAs expressed in B-cells from Ashkenazi Jewish centenarians, i.e., those living to a hundred and a human model of exceptional longevity, and younger controls without a family history of longevity. With data from 26.7 million reads comprising 9.4 × 108 bp from 3 centenarian and 3 control individuals, we discovered a total of 276 known miRNAs and 8 unknown miRNAs ranging several orders of magnitude in expression levels, a typical characteristics of saturated miRNA-sequencing. A total of 22 miRNAs were found to be significantly upregulated, with only 2 miRNAs downregulated, in centenarians as compared to controls. Gene Ontology analysis of the predicted and validated targets of the 24 differentially expressed miRNAs indicated enrichment of functional pathways involved in cell metabolism, cell cycle, cell signaling, and cell differentiation. A cross sectional expression analysis of the differentially expressed miRNAs in B-cells from Ashkenazi Jewish individuals between the 50th and 100th years of age indicated that expression levels of miR-363* declined significantly with age. Centenarians, however, maintained the youthful expression level. This result suggests that miR-363* may be a candidate longevity-associated miRNA. Conclusion Our comprehensive miRNA data provide a resource for further studies to identify genetic pathways associated with aging and longevity in humans.

  4. Prevalence of polyreactive innate clones among graft--infiltrating B cells in human cardiac allograft vasculopathy.

    Science.gov (United States)

    Chatterjee, Debanjana; Moore, Carolina; Gao, Baoshan; Clerkin, Kevin J; See, Sarah B; Shaked, David; Rogers, Kortney; Nunez, Sarah; Veras, Yokarla; Addonizio, Linda; Givertz, Michael M; Naka, Yoshifumi; Mancini, Donna; Vasilescu, Rodica; Marboe, Charles; Restaino, Susan; Madsen, Joren C; Zorn, Emmanuel

    2018-03-01

    Cardiac allograft vasculopathy (CAV) has been associated with graft-infiltrating B cells, although their characteristics are still unclear. In this study we examined the frequency, localization and reactivity profile of graft-infiltrating B cells to determine their contribution to the pathophysiology of CAV. B cells, plasma cells and macrophages were examined by immunohistochemistry in 56 allografts with CAV, 49 native failed hearts and 25 autopsy specimens. A total of 102 B-cell clones were immortalized directly from the infiltrates of 3 fresh cardiac samples with CAV. Their secreted antibodies were assessed using enzyme-linked immunoassay and flow cytometry. B-cell infiltration was observed around coronary arteries in 93% of allograft explants with CAV. Comparatively, intragraft B cells were less frequent and less dense in the intraventricular myocardium from where routine biopsies are obtained. Plasma cells and macrophages were also detected in 85% and 95% of explants, respectively. Remarkably, B-cell infiltrates were not associated with circulating donor-specific antibodies (DSA) or prior episodes of antibody-mediated rejection (AMR). Among all B-cell clones generated from 3 explants with CAV, a majority secreted natural antibodies reactive to multiple autoantigens and apoptotic cells, a characteristic of innate B cells. Our study reveals a high frequency of infiltrating B cells around the coronary arteries of allografts with CAV, independent of DSA or AMR. These cells are enriched for innate B cells with a polyreactive profile. The findings shift the focus from conventional DSA-producing B cells to the potentially pathogenic polyreactive B cells in the development of clinical CAV. Copyright © 2018 International Society for the Heart and Lung Transplantation. Published by Elsevier Inc. All rights reserved.

  5. Radiation-induced apoptosis of chicken lymphocyte B-cell line DT40

    International Nuclear Information System (INIS)

    Furusawa, Y.; Aoki, M.; Takakura, K.

    2003-01-01

    Full text: Ionizing radiation causes lesions of DNA, cell cycle arrest, induced cell death, and apoptosis in the irradiated cells. Then it is easy to expect that those events would be increased in a cell line which is defective in DNA repair system. However, induction of apoptosis by irradiation takes so complicated process when the cells are defective of DNA repair system. Indeed by many recent studies it has been clarified that DNA repair gene is also concerned with apoptotic event and some study shows the contrary data. Thus, the relationship between the genetics of apoptosis and that of DNA repair is still unclear. In this study two kinds of DNA repair proteins, Rad54 and Ku70, were focused. Proteins of Rad54 and Ku70 have important role at two type of DNA repair systems called homologous recombination repair and non-homologous end joining repair, respectively. 4 phenotypes of DT40, parent type, ku70-/-, rad54-/- and ku70-/-/rad54-/- were used to study the radiation-induced apoptosis (Previous study shows that survival fraction of 4 phenotypes of DT40 is decreased in the cell line, in which DNA repair gene is defective). From the results in this study, two things are clarifies. One is that the dependence of apoptotic index on phenotypes is so different between at low dose and at high dose irradiation. The other is that Ku70 has effective role to induce apoptosis in DT40 irradiated with high dose X-rays

  6. 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.

  7. 1,25-dihydroxyvitamin D3 impairs NF-κB activation in human naive B cells

    International Nuclear Information System (INIS)

    Geldmeyer-Hilt, Kerstin; Heine, Guido; Hartmann, Bjoern; Baumgrass, Ria; Radbruch, Andreas; Worm, Margitta

    2011-01-01

    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 D 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-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.

  8. Coordinate viral induction of tumor necrosis factor α and interferon β in human B cells and monocytes

    International Nuclear Information System (INIS)

    Goldfeld, A.E.; Maniatis, T.

    1989-01-01

    Human tumor necrosis factor α (TNF-α) gene expression can be induced primarily in cells of the monocyte/macrophage lineage by a variety of inducers, including lipopolysaccharide, phorbol esters such as phorbol 12-myristate 13-acetate, and virus or synthetic double-stranded RNA [poly(I)·poly(C)]. In this paper the authors show that the TNF-α gene also responds to virus and phorbol 12-myristate 13-acetate in B lymphocytes and that virus is the most potent inducer of TNF-α mRNA in both monocyte and B-cell lines. In addition, they show that viral infection coinduces the expression of TNF-α and interferon β mRNA and that viral induction of both genes is blocked by the kinase inhibitor 2-aminopurine. Inhibition of protein synthesis with cycloheximide had no effect on mRNA expression of the genes in one of three cell lines tested (U937) but blocked the viral induction of both genes in another (Namalwa). Thus, the regulatory factors required for mRNA induction of both genes are present prior to the addition of virus in U937 but not in Namalwa cells. However, in a third cell line (JY), cycloheximide blocked viral induction of the interferon β gene but not the TNF-α gene. Taken together, these observations suggest that viral induction of TNF-α and interferon β gene expression may involve overlapping pathways with both common and distinct regulatory factors

  9. IL-10-produced by human transitional B-cells down-regulates CD86 expression on B-cells leading to inhibition of CD4+T-cell responses.

    Science.gov (United States)

    Nova-Lamperti, Estefania; Fanelli, Giorgia; Becker, Pablo D; Chana, Prabhjoat; Elgueta, Raul; Dodd, Philippa C; Lord, Graham M; Lombardi, Giovanna; Hernandez-Fuentes, Maria P

    2016-01-22

    A novel subset of human regulatory B-cells has recently been described. They arise from within the transitional B-cell subpopulation and are characterised by the production of IL-10. They appear to be of significant importance in regulating T-cell immunity in vivo. Despite this important function, the molecular mechanisms by which they control T-cell activation are incompletely defined. Here we show that transitional B-cells produced more IL-10 and expressed higher levels of IL-10 receptor after CD40 engagement compared to other B-cell subsets. Furthermore, under this stimulatory condition, CD86 expressed by transitional B-cells was down regulated and T-cell proliferation was reduced. We provide evidence to demonstrate that the down-regulation of CD86 expression by transitional B-cells was due to the autocrine effect of IL-10, which in turn leads to decreased T-cell proliferation and TNF-α production. This analysis was further extended to peripheral B-cells in kidney transplant recipients. We observed that B-cells from patients tolerant to the graft maintained higher IL-10 production after CD40 ligation, which correlates with lower CD86 expression compared to patients with chronic rejection. Hence, the results obtained in this study shed light on a new alternative mechanism by which transitional B-cells inhibit T-cell proliferation and cytokine production.

  10. IL-10-produced by human transitional B-cells down-regulates CD86 expression on B-cells leading to inhibition of CD4+T-cell responses

    Science.gov (United States)

    Nova-Lamperti, Estefania; Fanelli, Giorgia; Becker, Pablo D.; Chana, Prabhjoat; Elgueta, Raul; Dodd, Philippa C.; Lord, Graham M.; Lombardi, Giovanna; Hernandez-Fuentes, Maria P.

    2016-01-01

    A novel subset of human regulatory B-cells has recently been described. They arise from within the transitional B-cell subpopulation and are characterised by the production of IL-10. They appear to be of significant importance in regulating T-cell immunity in vivo. Despite this important function, the molecular mechanisms by which they control T-cell activation are incompletely defined. Here we show that transitional B-cells produced more IL-10 and expressed higher levels of IL-10 receptor after CD40 engagement compared to other B-cell subsets. Furthermore, under this stimulatory condition, CD86 expressed by transitional B-cells was down regulated and T-cell proliferation was reduced. We provide evidence to demonstrate that the down-regulation of CD86 expression by transitional B-cells was due to the autocrine effect of IL-10, which in turn leads to decreased T-cell proliferation and TNF-α production. This analysis was further extended to peripheral B-cells in kidney transplant recipients. We observed that B-cells from patients tolerant to the graft maintained higher IL-10 production after CD40 ligation, which correlates with lower CD86 expression compared to patients with chronic rejection. Hence, the results obtained in this study shed light on a new alternative mechanism by which transitional B-cells inhibit T-cell proliferation and cytokine production. PMID:26795594

  11. Proton pump inhibitors induce apoptosis of human B-cell tumors through a caspase-independent mechanism involving reactive oxygen species.

    Science.gov (United States)

    De Milito, Angelo; Iessi, Elisabetta; Logozzi, Mariantonia; Lozupone, Francesco; Spada, Massimo; Marino, Maria Lucia; Federici, Cristina; Perdicchio, Maurizio; Matarrese, Paola; Lugini, Luana; Nilsson, Anna; Fais, Stefano

    2007-06-01

    Proton pumps like the vacuolar-type H+ ATPase (V-ATPase) are involved in the control of cellular pH in normal and tumor cells. Treatment with proton pump inhibitors (PPI) induces sensitization of cancer cells to chemotherapeutics via modifications of cellular pH gradients. It is also known that low pH is the most suitable condition for a full PPI activation. Here, we tested whether PPI treatment in unbuffered culture conditions could affect survival and proliferation of human B-cell tumors. First, we showed that PPI treatment increased the sensitivity to vinblastine of a pre-B acute lymphoblastic leukemia (ALL) cell line. PPI, per se, induced a dose-dependent inhibition of proliferation of tumor B cells, which was associated with a dose- and time-dependent apoptotic-like cytotoxicity in B-cell lines and leukemic cells from patients with pre-B ALL. The effect of PPI was mediated by a very early production of reactive oxygen species (ROS), that preceded alkalinization of lysosomal pH, lysosomal membrane permeabilization, and cytosol acidification, suggesting an early destabilization of the acidic vesicular compartment. Lysosomal alterations were followed by mitochondrial membrane depolarization, release of cytochrome c, chromatin condensation, and caspase activation. However, inhibition of caspase activity did not affect PPI-induced cell death, whereas specific inhibition of ROS by an antioxidant (N-acetylcysteine) significantly delayed cell death and protected both lysosomal and mitochondrial membranes. The proapoptotic activity of PPI was consistent with a clear inhibition of tumor growth following PPI treatment of B-cell lymphoma in severe combined immunodeficient mice. This study further supports the importance of acidity and pH gradients in tumor cell homeostasis and suggests new therapeutic approaches for human B-cell tumors based on PPI.

  12. Human CD40 ligand-expressing type 3 innate lymphoid cells induce IL-10-producing immature transitional regulatory B cells.

    Science.gov (United States)

    Komlósi, Zsolt I; Kovács, Nóra; van de Veen, Willem; Kirsch, Anna Isabella; Fahrner, Heinz Benedikt; Wawrzyniak, Marcin; Rebane, Ana; Stanic, Barbara; Palomares, Oscar; Rückert, Beate; Menz, Günter; Akdis, Mübeccel; Losonczy, György; Akdis, Cezmi A

    2017-09-20

    Type 3 innate lymphoid cells (ILC3s) are involved in maintenance of mucosal homeostasis; however, their role in immunoregulation has been unknown. Immature transitional regulatory B (itBreg) cells are innate-like B cells with immunosuppressive properties, and the in vivo mechanisms by which they are induced have not been fully clarified. We aimed to investigate the ILC3-B-cell interaction that probably takes place in human tonsils. ILC3s were isolated from peripheral blood and palatine tonsils, expanded, and cocultured with naive B cells. Tonsillar ILC3s and regulatory B cells were visualized with immunofluorescence histology. ILC3 frequencies were measured in tonsil tissue of allergic and nonallergic patients and in peripheral blood of allergic asthmatic patients and healthy control subjects. A mutually beneficial relationship was revealed between ILC3s and B cells: ILC3s induced IL-15 production in B cells through B cell-activating factor receptor, whereas IL-15, a potent growth factor for ILC3s, induced CD40 ligand (CD40L) expression on circulating and tonsillar ILC3s. IL-15-activated CD40L + ILC3s helped B-cell survival, proliferation, and differentiation of IL-10-secreting, PD-L1-expressing functional itBreg cells in a CD40L- and B cell-activating factor receptor-dependent manner. ILC3s and regulatory B cells were in close connection with each other in palatine tonsils. ILC3 frequency was reduced in tonsil tissue of allergic patients and in peripheral blood of allergic asthmatic patients. Human CD40L + ILC3s provide innate B-cell help and are involved in an innate immunoregulatory mechanism through induction of itBreg cell differentiation, which takes place in palatine tonsils in vivo. This mechanism, which can contribute to maintenance of immune tolerance, becomes insufficient in allergic diseases. Copyright © 2017 American Academy of Allergy, Asthma & Immunology. Published by Elsevier Inc. All rights reserved.

  13. Antibody repertoires in humanized NOD-scid-IL2Rγ(null mice and human B cells reveals human-like diversification and tolerance checkpoints in the mouse.

    Directory of Open Access Journals (Sweden)

    Gregory C Ippolito

    Full Text Available Immunodeficient mice reconstituted with human hematopoietic stem cells enable the in vivo study of human hematopoiesis. In particular, NOD-scid-IL2Rγ(null engrafted mice have been shown to have reasonable levels of T and B cell repopulation and can mount T-cell dependent responses; however, antigen-specific B-cell responses in this model are generally poor. We explored whether developmental defects in the immunoglobulin gene repertoire might be partly responsible for the low level of antibody responses in this model. Roche 454 sequencing was used to obtain over 685,000 reads from cDNA encoding immunoglobulin heavy (IGH and light (IGK and IGL genes isolated from immature, naïve, or total splenic B cells in engrafted NOD-scid-IL2Rγ(null mice, and compared with over 940,000 reads from peripheral B cells of two healthy volunteers. We find that while naïve B-cell repertoires in humanized mice are chiefly indistinguishable from those in human blood B cells, and display highly correlated patterns of immunoglobulin gene segment use, the complementarity-determining region H3 (CDR-H3 repertoires are nevertheless extremely diverse and are specific for each individual. Despite this diversity, preferential D(H-J(H pairings repeatedly occur within the CDR-H3 interval that are strikingly similar across all repertoires examined, implying a genetic constraint imposed on repertoire generation. Moreover, CDR-H3 length, charged amino-acid content, and hydropathy are indistinguishable between humans and humanized mice, with no evidence of global autoimmune signatures. Importantly, however, a statistically greater usage of the inherently autoreactive IGHV4-34 and IGKV4-1 genes was observed in the newly formed immature B cells relative to naïve B or total splenic B cells in the humanized mice, a finding consistent with the deletion of autoreactive B cells in humans. Overall, our results provide evidence that key features of the primary repertoire are shaped by

  14. The progeny of a single virgin B cell predominates the human recall B cell response to the capsular polysaccharide of Haemophilus influenzae type b

    DEFF Research Database (Denmark)

    Barington, T; Hougs, L; Juul, L

    1996-01-01

    of the cells originated from a common virgin B cell. Kinetic considerations implied that an extremely selected population of hypermutated memory B cells must have existed in these individuals before the first systemic immunization with the Ag. A possible role for the mucosal immune system in the priming...

  15. Toll-like receptors in the pathogenesis of human B cell malignancies

    NARCIS (Netherlands)

    Isaza-Correa, Johana M.; Liang, Zheng; van den Berg, Anke; Diepstra, Arjan; Visser, Lydia

    2014-01-01

    Toll-like receptors (TLRs) are important players in B-cell activation, maturation and memory and may be involved in the pathogenesis of B-cell lymphomas. Accumulating studies show differential expression in this heterogeneous group of cancers. Stimulation with TLR specific ligands, or agonists of

  16. 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)

    Lu Haitian; Crawford, Robert B.; Kaplan, Barbara L.F.; Kaminski, Norbert E.

    2011-01-01

    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.

  17. Complex forms of mitochondrial DNA in human B cells transformed by Epstein-Barr virus (EBV)

    DEFF Research Database (Denmark)

    Christiansen, Gunna; Christiansen, C; Zeuthen, J

    1983-01-01

    Human lymphocytes and lymphoid cell lines were analyzed for the presence of complex forms of mitochondrial DNA (mtDNA) by electron microscopy. A high frequency (9%-14.5%) of catenated dimers, circular dimers, or oligomers were found in samples from Epstein-Barr-virus-(EBV) transformed lymphoblast......Human lymphocytes and lymphoid cell lines were analyzed for the presence of complex forms of mitochondrial DNA (mtDNA) by electron microscopy. A high frequency (9%-14.5%) of catenated dimers, circular dimers, or oligomers were found in samples from Epstein-Barr-virus-(EBV) transformed...

  18. Comparative analysis of TCDD-induced AhR-mediated gene expression in human, mouse and rat primary B cells

    Energy Technology Data Exchange (ETDEWEB)

    Kovalova, Natalia, E-mail: kovalova@msu.edu [Department of Pharmacology and Toxicology, Michigan State University, Lansing, MI 48824 (United States); Institute for Integrative Toxicology, Michigan State University, Lansing, MI 48824 (United States); Nault, Rance, E-mail: naultran@msu.edu [Institute for Integrative Toxicology, Michigan State University, Lansing, MI 48824 (United States); Department of Biochemistry & Molecular Biology, Michigan State University, East Lansing, MI 48824 (United States); Crawford, Robert, E-mail: crawfo28@msu.edu [Institute for Integrative Toxicology, Michigan State University, Lansing, MI 48824 (United States); Zacharewski, Timothy R., E-mail: tzachare@msu.edu [Institute for Integrative Toxicology, Michigan State University, Lansing, MI 48824 (United States); Department of Biochemistry & Molecular Biology, Michigan State University, East Lansing, MI 48824 (United States); Kaminski, Norbert E., E-mail: kamins11@msu.edu [Department of Pharmacology and Toxicology, Michigan State University, Lansing, MI 48824 (United States); Institute for Integrative Toxicology, Michigan State University, Lansing, MI 48824 (United States)

    2017-02-01

    2,3,7,8-Tetrachlorodibenzo-p-dioxin (TCDD) is a persistent environmental pollutant that activates the aryl hydrocarbon receptor (AhR) resulting in altered gene expression. In vivo, in vitro, and ex vivo studies have demonstrated that B cells are directly impaired by TCDD, and are a sensitive target as evidenced by suppression of antibody responses. The window of sensitivity to TCDD-induced suppression of IgM secretion among mouse, rat and human B cells is similar. Specifically, TCDD must be present within the initial 12 h post B cell stimulation, indicating that TCDD disrupts early signaling network(s) necessary for B lymphocyte activation and differentiation. Therefore, we hypothesized that TCDD treatment across three different species (mouse, rat and human) triggers a conserved, B cell-specific mechanism that is involved in TCDD-induced immunosuppression. RNA sequencing (RNA-Seq) was used to identify B cell-specific orthologous genes that are differentially expressed in response to TCDD in primary mouse, rat and human B cells. Time course studies identified TCDD-elicited differential expression of 515 human, 2371 mouse and 712 rat orthologous genes over the 24-h period. 28 orthologs were differentially expressed in response to TCDD in all three species. Overrepresented pathways enriched in all three species included cytokine-cytokine receptor interaction, ECM-receptor interaction, focal adhesion, regulation of actin cytoskeleton and pathways in cancer. Differentially expressed genes functionally associated with cell-cell signaling in humans, immune response in mice, and oxidation reduction in rats. Overall, these results suggest that despite the conservation of the AhR and its signaling mechanism, TCDD elicits species-specific gene expression changes. - Highlights: • Kovalova TAAP Highlights Nov. 2016 • RNA-Seq identified TCDD-induced gene expression in PWM-activated primary B cells. • TCDD elicited differential expression of 515 human, 2371 mouse and 712

  19. Enhancement of hypermutation frequency in the chicken B cell line DT40 for efficient diversification of the antibody repertoire

    Energy Technology Data Exchange (ETDEWEB)

    Magari, Masaki; Kanehiro, Yuichi; Todo, Kagefumi; Ikeda, Mika; Kanayama, Naoki, E-mail: nkanayama@cc.okayama-u.ac.jp; Ohmori, Hitoshi

    2010-05-28

    Chicken B cell line DT40 continuously accumulates mutations in the immunoglobulin variable region (IgV) gene by gene conversion and point mutation, both of which are mediated by activation-induced cytidine deaminase (AID), thereby producing an antibody (Ab) library that is useful for screening monoclonal Abs (mAbs) in vitro. We previously generated an engineered DT40 line named DT40-SW, whose AID expression can be reversibly switched on or off, and developed an in vitro Ab generation system using DT40-SW cells. To efficiently create an Ab library with sufficient diversity, higher hypermutation frequency is advantageous. To this end, we generated a novel cell line DT40-SW{Delta}C, which conditionally expresses a C-terminus-truncated AID mutant lacking the nuclear export signal. The transcription level of the mutant AID gene in DT40-SW{Delta}C cells was similar to that of the wild-type gene in DT40-SW cells. However, the protein level of the truncated AID mutant was less than that of the wild type. The mutant protein was enriched in the nuclei of DT40-SW{Delta}C cells, although the protein might be highly susceptible to degradation. In DT40-SW{Delta}C cells, both gene conversion and point mutation occurred in the IgV gene with over threefold higher frequency than in DT40-SW cells, suggesting that a lower level of the mutant AID protein was sufficient to increase mutation frequency. Thus, DT40-SW{Delta}C cells may be useful for constructing Ab libraries for efficient screening of mAbs in vitro.

  20. The sensitivity of diffuse large B-cell lymphoma cell lines to histone deacetylase inhibitor-induced apoptosis is modulated by BCL-2 family protein activity.

    Directory of Open Access Journals (Sweden)

    Ryan C Thompson

    Full Text Available BACKGROUND: Diffuse large B-cell lymphoma (DLBCL is a genetically heterogeneous disease and this variation can often be used to explain the response of individual patients to chemotherapy. One cancer therapeutic approach currently in clinical trials uses histone deacetylase inhibitors (HDACi's as monotherapy or in combination with other agents. METHODOLOGY/PRINCIPAL FINDINGS: We have used a variety of cell-based and molecular/biochemical assays to show that two pan-HDAC inhibitors, trichostatin A and vorinostat, induce apoptosis in seven of eight human DLBCL cell lines. Consistent with previous reports implicating the BCL-2 family in regulating HDACi-induced apoptosis, ectopic over-expression of anti-apoptotic proteins BCL-2 and BCL-XL or pro-apoptotic protein BIM in these cell lines conferred further resistance or sensitivity, respectively, to HDACi treatment. Additionally, BCL-2 family antgonist ABT-737 increased the sensitivity of several DLBCL cell lines to vorinostat-induced apoptosis, including one cell line (SUDHL6 that is resistant to vorinostat alone. Moreover, two variants of the HDACi-sensitive SUDHL4 cell line that have decreased sensitivity to vorinostat showed up-regulation of BCL-2 family anti-apoptotic proteins such as BCL-XL and MCL-1, as well as decreased sensitivity to ABT-737. These results suggest that the regulation and overall balance of anti- to pro-apoptotic BCL-2 family protein expression is important in defining the sensitivity of DLBCL to HDACi-induced apoptosis. However, the sensitivity of DLBCL cell lines to HDACi treatment does not correlate with expression of any individual BCL-2 family member. CONCLUSIONS/SIGNIFICANCE: These studies indicate that the sensitivity of DLBCL to treatment with HDACi's is dependent on the complex regulation of BCL-2 family members and that BCL-2 antagonists may enhance the response of a subset of DLBCL patients to HDACi treatment.

  1. Mapping of Minimal Motifs of B-Cell Epitopes on Human Zona Pellucida Glycoprotein-3

    Directory of Open Access Journals (Sweden)

    Wan-Xiang Xu

    2012-01-01

    Full Text Available The human zona pellucida glycoprotein-3 (hZP3 by virtue of its critical role during fertilization has been proposed as a promising candidate antigen to develop a contraceptive vaccine. In this direction, it is imperative to map minimal motifs of the B cell epitopes (BCEs so as to avoid ZP-specific oophoritogenic T cell epitopes (TCEs in the ZP3-based immunogens. In this study, based on known results of mapping marmoset and bonnet monkey ZP3 (mstZP3 and bmZP3, two predictable epitopes23–30  and  301–320 on hZP3 were first confirmed and five minimal motifs within four epitopes on hZP3 were defined using serum to recombinant hZP3a22–176 or hZP3b177–348 as well as a biosynthetic peptide strategy. These defined minimal motifs were QPLWLL23–28 for hZP323–30, MQVTDD103–108 for hZP393–110, EENW178–181 for hZP3172–190, as well as SNSWF306–310 and EGP313–315 for hZP3301–320, respectively. Furthermore, the antigenicity of two peptides for hZP3172–187 and hZP3301–315 and specificity of the antibody response to these peptides were also evaluated, which produced high-titer antibodies in immunized animals that were capable of reacting to ZP on human oocytes, r-hZP3b177–348 protein, as well as r-hZP3172–190, r-hZP3303–310, and r-hZP3313–320 epitope peptides fused with truncated GST188 protein.

  2. Src inhibitor herbimycin A prevents 132.7 kDa tyrosine phosphatase activity in Ramos Burkitt's lymphoma B cell line

    International Nuclear Information System (INIS)

    Hristov, K.; Mitev, V.; Knox, K.

    2006-01-01

    Reversible tyrosine phosphorylation, regulation of expression and proteolytic cleavage control tyrosine phosphatase contribution for the signalling pathways of B-cell antigen receptor (BCR), and CD40 during B cell selection. We used Ramos-BL B cell line to determine whether BCR and CD40 stimulation, or inhibition of the Src - tyrosine kinase, tyrosine phosphatase and caspase activity have an effect on the tyrosine phosphatase activities determined on in-gel phosphatase assay. The tyrosine phosphatase activities present in whole cell lysates of Ramos-BL B cells following treatment with 20 μg/ml anti-IgM, 1 μg/ml anti-CD40, 10 μM herbimycin A, 178 μM vanadate,100 μM phenylarsine oxide and 10 μM zVAD-fmk were detected with an in-gel phosphatase assay. Seven major tyrosine phosphatase activities with approximate molecular weight of 132.7, 63.9, 60.3, 54.2, 49.7, 44.6, and 39 kDa are present in whole cell lysates of Ramos-BL B cells. Treatment with Src-PTK inhibitor herbimycin A prevents 132.7 kDa tyrosine phosphatase activity. We conclude that the catalytic activity of Src-PTK in Ramos-BL B cells is critical for the presence of this 132.7 kDa tyrosine phosphatase activity. (authors)

  3. Conserved B-cell epitopes among human bocavirus species indicate potential diagnostic targets.

    Directory of Open Access Journals (Sweden)

    Zhuo Zhou

    Full Text Available BACKGROUND: Human bocavirus species 1-4 (HBoV1-4 have been associated with respiratory and enteric infections in children. However, the immunological mechanisms in response to HBoV infections are not fully understood. Though previous studies have shown cross-reactivities between HBoV species, the epitopes responsible for this phenomenon remain unknown. In this study, we used genomic and immunologic approaches to identify the reactive epitopes conserved across multiple HBoV species and explored their potential as the basis of a novel diagnostic test for HBoVs. METHODOLOGY/PRINCIPAL FINDINGS: We generated HBoV1-3 VP2 gene fragment phage display libraries (GFPDLs and used these libraries to analyze mouse antisera against VP2 protein of HBoV1, 2, and 3, and human sera positive for HBoVs. Using this approach, we mapped four epitope clusters of HBoVs and identified two immunodominant peptides--P1 (¹MSDTDIQDQQPDTVDAPQNT²⁰, and P2 (¹⁶²EHAYPNASHPWDEDVMPDL¹⁸⁰--that are conserved among HBoV1-4. To confirm epitope immunogenicity, we immunized mice with the immunodominant P1 and P2 peptides identified in our screen and found that they elicited high titer antibodies in mice. These two antibodies could only recognize the VP2 of HBoV 1-4 in Western blot assays, rather than those of the two other parvoviruses human parvovirus B19 and human parvovirus 4 (PARV4. Based on our findings, we evaluated epitope-based peptide-IgM ELISAs as potential diagnostic tools for HBoVs IgM antibodies. We found that the P1+P2-IgM ELISA showed a higher sensitivity and specificity in HBoVs IgM detection than the assays using a single peptide. CONCLUSIONS/SIGNIFICANCE: The identification of the conserved B-cell epitopes among human bocavirus species contributes to our understanding of immunological cross-reactivities of HBoVs, and provides important insights for the development of HBoV diagnostic tools.

  4. 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

    2006-01-01

    CD40 promotes survival, proliferation, and differentiation of normal B cells but can cause activation-induced cell death in malignant B lymphocytes. CD40 ligand and anti-CD40 antibodies have been used successfully to induce apoptosis in lymphoma lines both in vitro and in xenograft tumor models. ...

  5. Human Memory B Cells Targeting Staphylococcus aureus Exotoxins Are Prevalent with Skin and Soft Tissue Infection

    Directory of Open Access Journals (Sweden)

    Adam J. Pelzek

    2018-03-01

    Full Text Available Staphylococcus aureus is a Gram-positive opportunistic pathogen that causes superficial and invasive infections in the hospital and community. High mortality from infection emphasizes the need for improved methods for prevention and treatment. Although S. aureus possesses an arsenal of virulence factors that contribute to evasion of host defenses, few studies have examined long-term humoral and B-cell responses. Adults with acute-phase skin and soft tissue infections were recruited; blood samples were obtained; and S. aureus isolates, including methicillin-resistant strains, were subjected to genomic sequence analysis. In comparisons of acute-phase sera with convalescent-phase sera, a minority (37.5% of patients displayed 2-fold or greater increases in antibody titers against three or more S. aureus antigens, whereas nearly half exhibited no changes, despite the presence of toxin genes in most infecting strains. Moreover, enhanced antibody responses waned over time, which could reflect a defect in B-cell memory or long-lived plasma cells. However, memory B cells reactive with a range of S. aureus antigens were prevalent at both acute-phase and convalescent-phase time points. While some memory B cells exhibited toxin-specific binding, those cross-reactive with structurally related leucocidin subunits were dominant across patients, suggesting the targeting of conserved epitopes. Memory B-cell reactivity correlated with serum antibody levels for selected S. aureus exotoxins, suggesting a relationship between the cellular and humoral compartments. Overall, although there was no global defect in the representation of anti-S. aureus memory B cells, there was evidence of restrictions in the range of epitopes recognized, which may suggest potential therapeutic approaches for augmenting host defenses.

  6. Mitogen-stimulated phospholipid synthesis in normal and immune-deficient human B cells

    International Nuclear Information System (INIS)

    Chien, M.M.; Yokoyama, W.M.; Ashman, R.F.

    1986-01-01

    Eight patients with common variable panhypogammaglobulinemia were shown in the in vitro Ig biosynthesis assay to have defective B cell responses to pokeweed mitogen (PWM). Phospholipid synthesis was assessed in the B cell plus monocyte fraction (MB) and irradiated T cells (T*) of patients and paired normal controls. Cell populations were studied separately and in the four possible combinations (1:1), with and without PWM, to reveal the effect of cell interactions. At 16 to 20 hr the mean stimulation index (SI) +/- standard error for MB cells alone was 1.01 +/- 0.02 for eight patients and 0.99 +/- 0.02 for the paired normals; the T* cell SI was 1.25 +/- 0.04 for patients and 1.28 +/- 0.05 for normals. Combinations of normal MB cells with normal T* cells showed significantly higher SI when compared with the combinations of normal MB cells with patient T* cells (p less than 0.005). However, the combination of patient MB cells with patient T* cells and the combination of patient MB cells with normal T* cells were not significantly different in SI (0.05 less than p less than 0.1). Isolation of patient and normal B cells, T* cells, and monocytes after the choline pulse showed that patient B cells gave a higher SI with normal T* help than with patient T* help. Of greatest interest is the finding that patient B cells that were defective in PWM-stimulated Ig production nevertheless showed a phospholipid synthesis response to PWM in the normal range, suggesting that the maturation defect in these B cells occurs later than the phospholipid synthesis acceleration step, or on a different pathway

  7. Toxic effects of various pollutants in 11B7501 lymphoma B cell line from harbour seal (Phoca vitulina)

    International Nuclear Information System (INIS)

    Frouin, Heloise; Fortier, Marlene; Fournier, Michel

    2010-01-01

    Although, heavy metals and polycyclic aromatic hydrocarbons (PAHs) have been reported at high levels in marine mammals, little is known about the toxic effects of some of these contaminants. In this study, we assessed the immunotoxic and genotoxic effects of seven heavy metals (arsenic, vanadium, selenium, iron, zinc, silver and chromium) and one PAH (benzo[a]pyrene or B[a]P) on a lymphoma B cell line from harbour seal (Phoca vitulina). A significant reduction in lymphocyte proliferation was registered following an exposure to 0.05 μM of B[a]P, 5 μM of arsenic or selenium, 50 μM of vanadium, 100 μM of silver and 200 μM of iron. On the contrary, zinc increased the lymphoproliferative response at 200 μM. Decreased phagocytosis was observed at 20 μM of arsenic, 50 μM of B[a]P or selenium, 200 μM of zinc and 500 μM of vanadium. Micronuclei induction occurred with 0.2 μM of B[a]P, 100 μM of vanadium and with 200 μM of arsenic or selenium. Exposure to 50 μM of arsenic decreased G 2 /M phase of the cell cycle. Chromium did not induce any effects at the concentrations tested. Concentrations of heavy metals (except silver and vanadium) and B[a]P inducing an toxic effect are within the environmental ranges reported in the blood tissue of pinnipeds. The reduction of some functional activities of the harbour seal immune system may cause a significant weakness capable of altering host resistance to disease in free-ranging pinnipeds.

  8. The occurrence and significance of V gene mutations in B cell-derived human malignancy.

    Science.gov (United States)

    Stevenson, F K; Sahota, S S; Ottensmeier, C H; Zhu, D; Forconi, F; Hamblin, T J

    2001-01-01

    The classification of B cell tumors has relevance for refining and improving clinical strategies. However, consensus has been difficult to establish, and although a scheme is now available, objective criteria are desirable. Genetic technology will underpin and extend current knowledge, and it is certain to reveal further subdivisions of current tumor categories. The Ig variable region genes of B cell tumors present a considerable asset for this area of investigation. The unique sequences carried in neoplastic B cells are easily isolated and sequenced. In addition to acting as clone-specific markers of each tumor, they indicate where the cell has come from and track its history following transformation. There is emerging clinical value in knowing whether the cell of origin has encountered antigen and has moved from the naive compartment to the germinal center, where somatic mutation is activated. This is amply illustrated by the subdivision of chronic lymphocytic leukemia into two subsets, unmutated or mutated, each with very different prognosis. Other tumors may be subdivided in a similar way. Microarray technology is developing rapidly to probe gene expression and to further divide tumor categories. All these genetic analyses will provide objective data to enhance both our understanding of B cell tumors and our ability to treat them.

  9. Signals sustaining human immunoglobulin V gene hypermutation in isolated germinal centre B cells

    NARCIS (Netherlands)

    K. Dahlenborg; J.D. Pound (J.); J. Gordon (Jocelynne); C.A.K. Borrebaeck (C. A K); R. Carlsson (R.)

    2000-01-01

    textabstractAffinity maturation of antibody responses depends on somatic hypermutation of the immunoglobulin V genes. Hypermutation is initiated specifically in proliferating B cells in lymphoid germinal centres but the signals driving this process remain unknown. This study identifies signals that

  10. Corruption of human follicular B-lymphocyte trafficking by a B-cell superantigen.

    Science.gov (United States)

    Borhis, Gwenoline; Viau, Muriel; Badr, Gamal; Richard, Yolande; Zouali, Moncef

    2012-05-09

    Protein A (SpA) of Staphylococcus aureus is known to target the paratope of immunoglobulins expressing V(H)3 genes, and to delete marginal zone B cells and B-1a in vivo. We have discovered that SpA endows S. aureus with the potential to subvert B-cell trafficking in the host. We found that SpA, whose Fc-binding site has been inactivated, binds essentially to naïve B cells and induces a long-lasting decrease in CXCR4 expression and in B-cell chemotaxis to CXCL12. Competition experiments indicated that SpA does not interfere with binding of CXCR4 ligands and does not directly bind to CXCR4. This conclusion is strongly supported by the inability of SpA to modulate clathrin-mediated CXCR4 internalization, which contrasts with the potent effect of anti-immunoglobin M (IgM) antibodies. Microscopy and biochemical experiments confirmed that SpA binds to the surface IgM/IgD complex and induces its clathrin-dependent internalization. Concomitantly, the SpA-induced signaling leads to protein kinase C-dependent CXCR4 downmodulation, suggesting that SpA impairs the recycling of CXCR4, a postclathrin process that leads to either degradation into lysozomes or de novo expression at the cell surface. In addition to providing novel insight into disruption of B-cell trafficking by an infectious agent, our findings may have therapeutic implications. Because CXCR4 has been associated with cancer metastasis and with certain autoimmune diseases, SpA behaves as an evolutionary tailored highly specific, chemokine receptor inhibitor that may have value in addition to conventional cytotoxic therapy in patients with various malignancies and immune-mediated diseases.

  11. Monitoring the systemic human memory B cell compartment of melanoma patients for anti-tumor IgG antibodies.

    Directory of Open Access Journals (Sweden)

    Amy E Gilbert

    Full Text Available Melanoma, a potentially lethal skin cancer, is widely thought to be immunogenic in nature. While there has been much focus on T cell-mediated immune responses, limited knowledge exists on the role of mature B cells. We describe an approach, including a cell-based ELISA, to evaluate mature IgG antibody responses to melanoma from human peripheral blood B cells. We observed a significant increase in antibody responses from melanoma patients (n = 10 to primary and metastatic melanoma cells compared to healthy volunteers (n = 10 (P<0.0001. Interestingly, we detected a significant reduction in antibody responses to melanoma with advancing disease stage in our patient cohort (n = 21 (P<0.0001. Overall, 28% of melanoma patient-derived B cell cultures (n = 1,800 compared to 2% of cultures from healthy controls (n = 600 produced antibodies that recognized melanoma cells. Lastly, a patient-derived melanoma-specific monoclonal antibody was selected for further study. This antibody effectively killed melanoma cells in vitro via antibody-mediated cellular cytotoxicity. These data demonstrate the presence of a mature systemic B cell response in melanoma patients, which is reduced with disease progression, adding to previous reports of tumor-reactive antibodies in patient sera, and suggesting the merit of future work to elucidate the clinical relevance of activating humoral immune responses to cancer.

  12. Monitoring the Systemic Human Memory B Cell Compartment of Melanoma Patients for Anti-Tumor IgG Antibodies

    Science.gov (United States)

    Gilbert, Amy E.; Karagiannis, Panagiotis; Dodev, Tihomir; Koers, Alexander; Lacy, Katie; Josephs, Debra H.; Takhar, Pooja; Geh, Jenny L. C.; Healy, Ciaran; Harries, Mark; Acland, Katharine M.; Rudman, Sarah M.; Beavil, Rebecca L.; Blower, Philip J.; Beavil, Andrew J.; Gould, Hannah J.; Spicer, James; Nestle, Frank O.; Karagiannis, Sophia N.

    2011-01-01

    Melanoma, a potentially lethal skin cancer, is widely thought to be immunogenic in nature. While there has been much focus on T cell-mediated immune responses, limited knowledge exists on the role of mature B cells. We describe an approach, including a cell-based ELISA, to evaluate mature IgG antibody responses to melanoma from human peripheral blood B cells. We observed a significant increase in antibody responses from melanoma patients (n = 10) to primary and metastatic melanoma cells compared to healthy volunteers (n = 10) (P<0.0001). Interestingly, we detected a significant reduction in antibody responses to melanoma with advancing disease stage in our patient cohort (n = 21) (P<0.0001). Overall, 28% of melanoma patient-derived B cell cultures (n = 1,800) compared to 2% of cultures from healthy controls (n = 600) produced antibodies that recognized melanoma cells. Lastly, a patient-derived melanoma-specific monoclonal antibody was selected for further study. This antibody effectively killed melanoma cells in vitro via antibody-mediated cellular cytotoxicity. These data demonstrate the presence of a mature systemic B cell response in melanoma patients, which is reduced with disease progression, adding to previous reports of tumor-reactive antibodies in patient sera, and suggesting the merit of future work to elucidate the clinical relevance of activating humoral immune responses to cancer. PMID:21559411

  13. Circulating CXCR5+CD4+ T Follicular-Like Helper Cell and Memory B Cell Responses to Human Papillomavirus Vaccines

    Science.gov (United States)

    Matsui, Ken; Adelsberger, Joseph W.; Kemp, Troy J.; Baseler, Michael W.; Ledgerwood, Julie E.; Pinto, Ligia A.

    2015-01-01

    Through the interaction of T follicular helper (Tfh) cells and B cells, efficacious vaccines can generate high-affinity, pathogen-neutralizing antibodies, and memory B cells. Using CXCR5, CXCR3, CCR6, CCR7, PD1, and ICOS as markers, Tfh-like cells can be identified in the circulation and be classified into three functionally distinct subsets that are PD1+ICOS+, PD1+ ICOS-, or PD1-ICOS-. We used these markers to identify different subsets of CXCR5+CD4+ Tfh-like cells in response to highly immunogenic and efficacious vaccines for human papillomaviruses (HPV): Cervarix and Gardasil. In this small study, we used PBMC samples from 11 Gardasil recipients, and 8 Cervarix recipients from the Vaccine Research Center 902 Study to examine the induction of circulating Tfh-like cells and IgD-CD38HiCD27+ memory B cells by flow cytometry. PD1+ICOS+ CXCR3+CCR6-CXCR5+CD4+ (Tfh1-like) cells were induced and peaked on Day (D) 7 post-first vaccination, but not as much on D7 post-third vaccination. We also observed a trend toward increase in PD1+ICOS+ CXCR3-CCR6-CXCR5+CD4+ (Tfh2-like) cells for both vaccines, and PD1+ICOS+ CXCR3-CCR6+CXCR5+CD4+ (Tfh17-like) subset was induced by Cervarix post-first vaccination. There were also minimal changes in the other cellular subsets. In addition, Cervarix recipients had more memory B cells post-first vaccination than did Gardasil recipients at D14 and D30. We found frequencies of memory B cells at D30 correlated with anti-HPV16 and 18 antibody titers from D30, and the induction levels of memory B cells at D30 and PD1+ICOS+Tfh1-like cells at D7 post-first vaccination correlated for Cervarix. Our study showed that induction of circulating CXCR5+CD4+ Tfh-like subsets can be detected following immunization with HPV vaccines, and potentially be useful as a marker of immunogenicity of vaccines. However, further investigations should be extended to different cohorts with larger sample size to better understand the functions of these T cells, as well as

  14. Protection against Pertussis in Humans Correlates to Elevated Serum Antibodies and Memory B Cells

    Directory of Open Access Journals (Sweden)

    Valentina Marcellini

    2017-09-01

    Full Text Available Pertussis is a respiratory infection caused by Bordetella pertussis that may be particularly severe and even lethal in the first months of life when infants are still too young to be vaccinated. Adults and adolescents experience mild symptoms and are the source of infection for neonates. Adoptive maternal immunity does not prevent pertussis in the neonate. We compared the specific immune response of mothers of neonates diagnosed with pertussis and mothers of control children. We show that women have pre-existing pertussis-specific antibodies and memory B cells and react against the infection with a recall response increasing the levels specific serum IgG, milk IgA, and the frequency of memory B cells of all isotypes. Thus, the maternal immune system is activated in response to pertussis and effectively prevents the disease indicating that the low levels of pre-formed serum antibodies are insufficient for protection. For this reason, memory B cells play a major role in the adult defense. The results of this study suggest that new strategies for vaccine design should aim at increasing long-lived plasma cells and their antibodies.

  15. Responses to recipient and donor B cells by genetically donor T cells from human haploidentical chimeras

    International Nuclear Information System (INIS)

    Schiff, S.; Sampson, H.; Buckley, R.

    1986-01-01

    Following administration of haploidentical stem cells to infants with severe combined immunodeficiency (SCID), mature T cells of donor karyotype appear later in the recipient without causing graft-versus-host disease. To investigate the effect of the host environment on the responsiveness of these genetically donor T cells, blood B and T lymphocytes from 6 SCID recipients, their parental donors and unrelated controls were purified by double SRBC rosetting. T cells were stimulated by irradiated B cells at a 1:1 ratio in 6 day cultures. Engrafted T cells of donor karyotype gave much smaller responses to irradiated genetically recipient B cells than did fresh donor T cells. Moreover, engrafted T cells of donor karyotype from two of the three SCIDs who are longest post-transplantation responded more vigorously (14,685 and 31,623 cpm) than fresh donor T cells (5141 and 22,709 cpm) to donor B cells. These data indicate that T lymphocytes which have matured from donor stem cells in the recipient microenvironment behave differently from those that have matured in the donor

  16. Induction of apoptosis by epigallocatechin-3-gallate in human lymphoblastoid B cells

    International Nuclear Information System (INIS)

    Noda, Chiseko; He, Jinsong; Takano, Tomoko; Tanaka, Chisato; Kondo, Toshinori; Tohyama, Kaoru; Yamamura, Hirohei; Tohyama, Yumi

    2007-01-01

    (-)-Epigallocatechin-3-gallate (EGCG), a major constituent of green tea polyphenols, has been shown to suppress cancer cell proliferation and induce apoptosis. In this study we investigated its efficacy and the mechanism underlying its effect using human B lymphoblastoid cell line Ramos, and effect of co-treatment with EGCG and a chemotherapeutic agent on apoptotic cell death. EGCG induced dose- and time-dependent apoptotic cell death accompanied by loss of mitochondrial transmembrane potential, release of cytochrome c into the cytosol, and cleavage of pro-caspase-9 to its active form. EGCG also enhanced production of intracellular reactive oxygen species (ROS). Pretreatment with diphenylene iodonium chloride, an inhibitor of NAD(P)H oxidase and an antioxidant, partially suppressed both EGCG-induced apoptosis and production of ROS, implying that oxidative stress is involved in the apoptotic response. Furthermore, we showed that combined-treatment with EGCG and a chemotherapeutic agent, etoposide, synergistically induced apoptosis in Ramos cells

  17. Vav-1 expression correlates with NFkappaB activation and CD40-mediated cell death in diffuse large B-cell lymphoma cell lines

    DEFF Research Database (Denmark)

    Hollmann, Annette; Aloyz, Raquel; Baker, Kristi

    2010-01-01

    Diffuse large B-cell lymphoma (DLBCL) is an aggressive malignancy with a variable response to therapy. We have previously shown that DLBCL cell lines differ in their susceptibility to CD40-mediated cell death, and that resistance to CD40-targeted antibodies correlated with increased expression...... as a potential marker to identify tumours likely to respond to CD40-targeted therapies. Copyright (c) 2010 John Wiley & Sons, Ltd....

  18. 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)

    Løbner, Morten; 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 u...

  19. Role of aryl hydrocarbon receptor polymorphisms on TCDD-mediated CYP1B1 induction and IgM suppression by human B cells

    Energy Technology Data Exchange (ETDEWEB)

    Kovalova, Natalia, E-mail: kovalova@msu.edu [Department of Pharmacology and Toxicology, Michigan State University, Lansing, MI 48824 (United States); Institute for Integrative Toxicology, Michigan State University, Lansing, MI 48824 (United States); Manzan, Maria, E-mail: ale.manzan@gmail.com [Institute for Integrative Toxicology, Michigan State University, Lansing, MI 48824 (United States); Crawford, Robert, E-mail: crawfo28@msu.edu [Institute for Integrative Toxicology, Michigan State University, Lansing, MI 48824 (United States); Kaminski, Norbert, E-mail: kamins11@msu.edu [Department of Pharmacology and Toxicology, Michigan State University, Lansing, MI 48824 (United States); Institute for Integrative Toxicology, Michigan State University, Lansing, MI 48824 (United States)

    2016-10-15

    Previous studies have demonstrated that most of the intraspecies variation in sensitivity to the toxic effects of 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD), including suppression of antibody responses, in murine models is due to single nucleotide polymorphisms (SNPs) within the aryl hydrocarbon receptor (AhR) gene. The underlying reason for variation in sensitivity to TCDD-induced suppression of IgM responses among humans is not well understood, but is thought, in part, to be a result of different polymorphic forms of the AhR expressed by different individuals. In this study, the functional properties of six (P517S, R554K, V570I, V570I + P517S, R554K + V570I and P517S + R554K + V570I) human AhR variants were examined in the human B cell line, SKW 6.4. TCDD-induced Cyp1B1 and Cyp1A2 mRNA expression levels and Cyp1B1-regulated reporter gene activity, used for comparative purposes, were markedly lower in SKW cells containing the R554K SNP than in SKW-AHR{sup +} (control AhR) cells. Furthermore, all AhR variants were able to mediate TCDD-induced suppression of the IgM response; however, a combined P517S + R554K + V570I variant partially reduced sensitivity to TCDD-mediated suppression of IgM secretion. Collectively, our findings show that the R554K human AhR SNP alone altered sensitivity of human B cells to TCDD-mediated induction of Cyp1B1 and Cyp1A2. By contrast, attenuation of TCDD-induced IgM suppression required a combination of all three SNPs P517S, R554K, and V570I. - Highlights: • Mouse, rat and SKW-AHR{sup +} B cells have a similar window of sensitivity to TCDD. • R554K AhR SNP alters B cell sensitivity to TCDD-mediated Cyp1B1 and Cyp1A2 induction. • Combination of P517S, R554K, and V570I SNPs attenuates TCDD-induced IgM suppression.

  20. Human Cytotoxic T Lymphocytes Form Dysfunctional Immune Synapses with B Cells Characterized by Non-Polarized Lytic Granule Release

    Directory of Open Access Journals (Sweden)

    Anna Kabanova

    2016-04-01

    Full Text Available Suppression of the cytotoxic T cell (CTL immune response has been proposed as one mechanism for immune evasion in cancer. In this study, we have explored the underlying basis for CTL suppression in the context of B cell malignancies. We document that human B cells have an intrinsic ability to resist killing by freshly isolated cytotoxic T cells (CTLs, but are susceptible to lysis by IL-2 activated CTL blasts and CTLs isolated from immunotherapy-treated patients with chronic lymphocytic leukemia (CLL. Impaired killing was associated with the formation of dysfunctional non-lytic immune synapses characterized by the presence of defective linker for activation of T cells (LAT signaling and non-polarized release of the lytic granules transported by ADP-ribosylation factor-like protein 8 (Arl8. We propose that non-lytic degranulation of CTLs are a key regulatory mechanism of evasion through which B cells may interfere with the formation of functional immune synapses by CTLs.

  1. Endogenous Nur77 Is a Specific Indicator of Antigen Receptor Signaling in Human T and B Cells.

    Science.gov (United States)

    Ashouri, Judith F; Weiss, Arthur

    2017-01-15

    Distinguishing true Ag-stimulated lymphocytes from bystanders activated by the inflammatory milieu has been difficult. Nur77 is an immediate early gene whose expression is rapidly upregulated by TCR signaling in murine T cells and human thymocytes. Nur77-GFP transgenes serve as specific TCR and BCR signaling reporters in murine transgenic models. In this study, we demonstrate that endogenous Nur77 protein expression can serve as a reporter of TCR and BCR specific signaling in human PBMCs. Nur77 protein amounts were assessed by immunofluorescence and flow cytometry in T and B cells isolated from human PBMCs obtained from healthy donors that had been stimulated by their respective Ag receptors. We demonstrate that endogenous Nur77 is a more specific reporter of Ag-specific signaling events than the commonly used CD69 activation marker in both human T and B cells. This is reflective of the disparity in signaling pathways that regulate the expression of Nur77 and CD69. Assessing endogenous Nur77 protein expression has great potential to identify Ag-activated lymphocytes in human disease. Copyright © 2017 by The American Association of Immunologists, Inc.

  2. CD24(hi)CD27⁺ and plasmablast-like regulatory B cells in human chronic graft-versus-host disease.

    Science.gov (United States)

    de Masson, Adèle; Bouaziz, Jean-David; Le Buanec, Hélène; Robin, Marie; O'Meara, Alix; Parquet, Nathalie; Rybojad, Michel; Hau, Estelle; Monfort, Jean-Benoît; Branchtein, Mylène; Michonneau, David; Dessirier, Valérie; Sicre de Fontbrune, Flore; Bergeron, Anne; Itzykson, Raphaël; Dhédin, Nathalie; Bengoufa, Djaouida; Peffault de Latour, Régis; Xhaard, Aliénor; Bagot, Martine; Bensussan, Armand; Socié, Gérard

    2015-03-12

    Interleukin 10 (IL-10)-producing B cells (regulatory B cells [Bregs]) regulate autoimmunity in mice and humans, and a regulatory role of IL-10-producing plasma cells has been described in mice. Dysfunction of B cells that maintain homeostasis may play a role in the pathogenesis of chronic graft-versus-host disease (cGVHD) after allogeneic stem cell transplantation. Here, we found a relation between decreased Breg frequencies and cGVHD severity. An impaired ability of B cells to produce IL-10, possibly linked to poor signal transducer and activator of transcription 3 and extracellular signal-regulated kinase phosphorylation, was found in patients with active cGVHD. IL-10 production was not confined to a single B-cell subset, but enriched in both the CD24(hi)CD27(+) and CD27(hi)CD38(hi) plasmablast B-cell compartments. In vitro plasmablast differentiation increased the frequency of IL-10-producing B cells. We confirmed that allogeneic transplant recipients had an impaired reconstitution of the memory B-cell pool. cGVHD patients had less CD24(hi)CD27(+) B cells and IL-10-producing CD24(hi)CD27(+) B cells. Patients with cGVHD had increased plasmablast frequencies but decreased IL-10-producing plasmablasts. These results suggest a role of CD24(hi)CD27(+) B-cell and plasmablast-derived IL-10 in the regulation of human cGVHD. © 2015 by The American Society of Hematology.

  3. Combined immunodeficiency and Epstein-Barr virus-induced B cell malignancy in humans with inherited CD70 deficiency

    DEFF Research Database (Denmark)

    Abolhassani, Hassan; Edwards, Emily S. J.; Ikinciogullari, Aydan

    2017-01-01

    In this study, we describe four patients from two unrelated families of different ethnicities with a primary immunodeficiency, predominantly manifesting as susceptibility to Epstein-Barr virus (EBV)-related diseases. Three patients presented with EBV-associated Hodgkin's lymphoma...... is a novel cause of combined immunodeficiency and EBV-associated diseases, reminiscent of inherited CD27 deficiency. Overall, human CD70-CD27 interactions therefore play a nonredundant role in T and B cell-mediated immunity, especially for protection against EBV and humoral immunity....

  4. In Vitro Measles Virus Infection of Human Lymphocyte Subsets Demonstrates High Susceptibility and Permissiveness of both Naive and Memory B Cells.

    Science.gov (United States)

    Laksono, Brigitta M; Grosserichter-Wagener, Christina; de Vries, Rory D; Langeveld, Simone A G; Brem, Maarten D; van Dongen, Jacques J M; Katsikis, Peter D; Koopmans, Marion P G; van Zelm, Menno C; de Swart, Rik L

    2018-04-15

    Measles is characterized by a transient immune suppression, leading to an increased risk of opportunistic infections. Measles virus (MV) infection of immune cells is mediated by the cellular receptor CD150, expressed by subsets of lymphocytes, dendritic cells, macrophages, and thymocytes. Previous studies showed that human and nonhuman primate memory T cells express higher levels of CD150 than naive cells and are more susceptible to MV infection. However, limited information is available about the CD150 expression and relative susceptibility to MV infection of B-cell subsets. In this study, we assessed the susceptibility and permissiveness of naive and memory T- and B-cell subsets from human peripheral blood or tonsils to in vitro MV infection. Our study demonstrates that naive and memory B cells express CD150, but at lower frequencies than memory T cells. Nevertheless, both naive and memory B cells proved to be highly permissive to MV infection. Furthermore, we assessed the susceptibility and permissiveness of various functionally distinct T and B cells, such as helper T (T H ) cell subsets and IgG- and IgA-positive memory B cells, in peripheral blood and tonsils. We demonstrated that T H 1T H 17 cells and plasma and germinal center B cells were the subsets most susceptible and permissive to MV infection. Our study suggests that both naive and memory B cells, along with several other antigen-experienced lymphocytes, are important target cells of MV infection. Depletion of these cells potentially contributes to the pathogenesis of measles immune suppression. IMPORTANCE Measles is associated with immune suppression and is often complicated by bacterial pneumonia, otitis media, or gastroenteritis. Measles virus infects antigen-presenting cells and T and B cells, and depletion of these cells may contribute to lymphopenia and immune suppression. Measles has been associated with follicular exhaustion in lymphoid tissues in humans and nonhuman primates, emphasizing the

  5. Chemo-sensitivity in a panel of B-cell precursor acute lymphoblastic leukemia cell lines, YCUB series, derived from children.

    Science.gov (United States)

    Goto, Hiroaki; Naruto, Takuya; Tanoshima, Reo; Kato, Hiromi; Yokosuka, Tomoko; Yanagimachi, Masakatsu; Fujii, Hisaki; Yokota, Shumpei; Komine, Hiromi

    2009-10-01

    Sensitivity to 10 anticancer drugs was evaluated in 6 childhood B-cell precursor acute lymphoblastic leukemia (BCP-ALL) cell lines. Authenticity of newly established cell lines was confirmed by genomic fingerprinting. The line YCUB-5R established at relapse was more resistant to 4-hydroperoxy-cyclophosphamide, cytarabine, L-asparaginase, topotecan, fludarabine, and etoposide than YCUB-5 from the same patient at diagnosis. Of the drugs tested, etoposide and SN-38 (irinotecan) showed highest efficacy in the panel, with 50% growth inhibition at 0.22-1.8 microg/ml and 0.57-3.6 ng/ml, respectively. This cell line panel offers an in vitro model for the development of new therapies for childhood BCP-ALL.

  6. CD83 Antibody Inhibits Human B Cell Responses to Antigen as well as Dendritic Cell-Mediated CD4 T Cell Responses.

    Science.gov (United States)

    Wong, Kuan Y; Baron, Rebecca; Seldon, Therese A; Jones, Martina L; Rice, Alison M; Munster, David J

    2018-05-15

    Anti-CD83 Ab capable of Ab-dependent cellular cytotoxicity can deplete activated CD83 + human dendritic cells, thereby inhibiting CD4 T cell-mediated acute graft-versus-host disease. As CD83 is also expressed on the surface of activated B lymphocytes, we hypothesized that anti-CD83 would also inhibit B cell responses to stimulation. We found that anti-CD83 inhibited total IgM and IgG production in vitro by allostimulated human PBMC. Also, Ag-specific Ab responses to immunization of SCID mice xenografted with human PBMC were inhibited by anti-CD83 treatment. This inhibition occurred without depletion of all human B cells because anti-CD83 lysed activated CD83 + B cells by Ab-dependent cellular cytotoxicity and spared resting (CD83 - ) B cells. In cultured human PBMC, anti-CD83 inhibited tetanus toxoid-stimulated B cell proliferation and concomitant dendritic cell-mediated CD4 T cell proliferation and expression of IFN-γ and IL-17A, with minimal losses of B cells (80% of B cells but had no effect on CD4 T cell proliferation and cytokine expression. By virtue of the ability of anti-CD83 to selectively deplete activated, but not resting, B cells and dendritic cells, with the latter reducing CD4 T cell responses, anti-CD83 may be clinically useful in autoimmunity and transplantation. Advantages might include inhibited expansion of autoantigen- or alloantigen-specific B cells and CD4 T cells, thus preventing further production of pathogenic Abs and inflammatory cytokines while preserving protective memory and regulatory cells. Copyright © 2018 by The American Association of Immunologists, Inc.

  7. Analysis of B Cell Repertoire Dynamics Following Hepatitis B Vaccination in Humans, and Enrichment of Vaccine-specific Antibody Sequences

    Directory of Open Access Journals (Sweden)

    Jacob D. Galson

    2015-12-01

    Full Text Available Generating a diverse B cell immunoglobulin repertoire is essential for protection against infection. The repertoire in humans can now be comprehensively measured by high-throughput sequencing. Using hepatitis B vaccination as a model, we determined how the total immunoglobulin sequence repertoire changes following antigen exposure in humans, and compared this to sequences from vaccine-specific sorted cells. Clonal sequence expansions were seen 7 days after vaccination, which correlated with vaccine-specific plasma cell numbers. These expansions caused an increase in mutation, and a decrease in diversity and complementarity-determining region 3 sequence length in the repertoire. We also saw an increase in sequence convergence between participants 14 and 21 days after vaccination, coinciding with an increase of vaccine-specific memory cells. These features allowed development of a model for in silico enrichment of vaccine-specific sequences from the total repertoire. Identifying antigen-specific sequences from total repertoire data could aid our understanding B cell driven immunity, and be used for disease diagnostics and vaccine evaluation.

  8. Analysis of B Cell Repertoire Dynamics Following Hepatitis B Vaccination in Humans, and Enrichment of Vaccine-specific Antibody Sequences.

    Science.gov (United States)

    Galson, Jacob D; Trück, Johannes; Fowler, Anna; Clutterbuck, Elizabeth A; Münz, Márton; Cerundolo, Vincenzo; Reinhard, Claudia; van der Most, Robbert; Pollard, Andrew J; Lunter, Gerton; Kelly, Dominic F

    2015-12-01

    Generating a diverse B cell immunoglobulin repertoire is essential for protection against infection. The repertoire in humans can now be comprehensively measured by high-throughput sequencing. Using hepatitis B vaccination as a model, we determined how the total immunoglobulin sequence repertoire changes following antigen exposure in humans, and compared this to sequences from vaccine-specific sorted cells. Clonal sequence expansions were seen 7 days after vaccination, which correlated with vaccine-specific plasma cell numbers. These expansions caused an increase in mutation, and a decrease in diversity and complementarity-determining region 3 sequence length in the repertoire. We also saw an increase in sequence convergence between participants 14 and 21 days after vaccination, coinciding with an increase of vaccine-specific memory cells. These features allowed development of a model for in silico enrichment of vaccine-specific sequences from the total repertoire. Identifying antigen-specific sequences from total repertoire data could aid our understanding B cell driven immunity, and be used for disease diagnostics and vaccine evaluation.

  9. Association of CLEC16A with human common variable immunodeficiency disorder and role in murine B cells.

    Science.gov (United States)

    Li, Jin; Jørgensen, Silje F; Maggadottir, S Melkorka; Bakay, Marina; Warnatz, Klaus; Glessner, Joseph; Pandey, Rahul; Salzer, Ulrich; Schmidt, Reinhold E; Perez, Elena; Resnick, Elena; Goldacker, Sigune; Buchta, Mary; Witte, Torsten; Padyukov, Leonid; Videm, Vibeke; Folseraas, Trine; Atschekzei, Faranaz; Elder, James T; Nair, Rajan P; Winkelmann, Juliane; Gieger, Christian; Nöthen, Markus M; Büning, Carsten; Brand, Stephan; Sullivan, Kathleen E; Orange, Jordan S; Fevang, Børre; Schreiber, Stefan; Lieb, Wolfgang; Aukrust, Pål; Chapel, Helen; Cunningham-Rundles, Charlotte; Franke, Andre; Karlsen, Tom H; Grimbacher, Bodo; Hakonarson, Hakon; Hammarström, Lennart; Ellinghaus, Eva

    2015-04-20

    Common variable immunodeficiency disorder (CVID) is the most common symptomatic primary immunodeficiency in adults, characterized by B-cell abnormalities and inadequate antibody response. CVID patients have considerable autoimmune comorbidity and we therefore hypothesized that genetic susceptibility to CVID may overlap with autoimmune disorders. Here, in the largest genetic study performed in CVID to date, we compare 778 CVID cases with 10,999 controls across 123,127 single-nucleotide polymorphisms (SNPs) on the Immunochip. We identify the first non-HLA genome-wide significant risk locus at CLEC16A (rs17806056, P=2.0 × 10(-9)) and confirm the previously reported human leukocyte antigen (HLA) associations on chromosome 6p21 (rs1049225, P=4.8 × 10(-16)). Clec16a knockdown (KD) mice showed reduced number of B cells and elevated IgM levels compared with controls, suggesting that CLEC16A may be involved in immune regulatory pathways of relevance to CVID. In conclusion, the CLEC16A associations in CVID represent the first robust evidence of non-HLA associations in this immunodeficiency condition.

  10. Immunoglobulin lambda light chain gene rearrangements in human B-cell malignancies

    NARCIS (Netherlands)

    T. Tümkaya (Talip)

    1997-01-01

    textabstractLymphocytes form the specific immune system, capable of recognizing and responding to any foreign antigen, while remaining indifferent to self components. Throughout human life, lymphocytes are continuously generated from pluripotent hematopoietic stem cells. These hematopoietic stem

  11. The FOX and the mutants in mature human B cells and DLBCL: The role of FOXP1 in mature human B cell biology and lymphomagenesis & prevalence of oncogenic MyD88 and CD79B mutations in diffuse large B cell lymphoma

    NARCIS (Netherlands)

    van Keimpema, M.

    2015-01-01

    The transcription factor FOXP1 is prominently expressed in mature B cells and is a potential oncogene in B cell non-Hodgkin lymphomas; however, the functions of FOXP1 in mature B cells and B cell lymphomagenesis have not yet been fully explored. In the first part of this thesis, the roles of FOXP1

  12. Restoration of human B-cell differentiation into NOD-SCID mice engrafted with gene-corrected CD34+ cells isolated from Artemis or RAG1-deficient patients.

    Science.gov (United States)

    Lagresle-Peyrou, Chantal; Benjelloun, Fatine; Hue, Christophe; Andre-Schmutz, Isabelle; Bonhomme, Delphine; Forveille, Monique; Beldjord, Kheira; Hacein-Bey-Abina, Salima; De Villartay, Jean-Pierre; Charneau, Pierre; Durandy, Anne; Fischer, Alain; Cavazzana-Calvo, Marina

    2008-02-01

    Severe combined immunodeficiency (SCID) caused by mutation of the recombination-activating gene 1 (RAG1) or Artemis gene lead to the absence of B- and T-cell differentiation. The only curative treatment is allogeneic bone marrow (BM) transplantation, which displays a high survival rate when an HLA compatible donor is available but has a poorer prognosis when the donor is partially compatible. Consequently, gene therapy may be a promising alternative strategy for these diseases. Here, we report that lentiviral gene-corrected BM CD34(+) cells (isolated from Artemis- or RAG1-deficient patients) sustain human B-cell differentiation following injection into non-obese diabetic/SCID (NOD-SCID) mice previously infused with anti-interleukin-2 receptor beta chain monoclonal antibody. In most of the mice BM, engrafted with Artemis-transduced cells, human B-cell differentiation occurred until the mature stage. The B cells were functional as human immunoglobulin M (IgM) was present in the serum. Following injection with RAG1-transduced cells, human engraftment occurred in vivo but B-cell differentiation until the mature stage was less frequent. However, when it occurred, it was always associated with human IgM production. This overall approach represents a useful tool for evaluating gene transfer efficiency in human SCID forms affecting B-cell development (such as Artemis deficiency) and for testing new vectors for improving in vivo RAG1 complementation.

  13. Epstein–Barr Virus-Induced Metabolic Rearrangements in Human B-Cell Lymphomas

    Directory of Open Access Journals (Sweden)

    Pier P. Piccaluga

    2018-06-01

    Full Text Available Tumor metabolism has been the object of several studies in the past, leading to the pivotal observation of a consistent shift toward aerobic glycolysis (so-called Warburg effect. More recently, several additional investigations proved that tumor metabolism is profoundly affected during tumorigenesis, including glucose, lipid and amino-acid metabolism. It is noticeable that metabolic reprogramming can represent a suitable therapeutic target in many cancer types. Epstein–Barr virus (EBV was the first virus linked with cancer in humans when Burkitt lymphoma (BL was described. Besides other well-known effects, it was recently demonstrated that EBV can induce significant modification in cell metabolism, which may lead or contribute to neoplastic transformation of human cells. Similarly, virus-induced tumorigenesis is characterized by relevant metabolic abnormalities directly induced by the oncoviruses. In this article, the authors critically review the most recent literature concerning EBV-induced metabolism alterations in lymphomas.

  14. The role of T and B cells in human atherosclerosis and atherothrombosis.

    Science.gov (United States)

    Ammirati, E; Moroni, F; Magnoni, M; Camici, P G

    2015-02-01

    Far from being merely a passive cholesterol accumulation within the arterial wall, the development of atherosclerosis is currently known to imply both inflammation and immune effector mechanisms. Adaptive immunity has been implicated in the process of disease initiation and progression interwined with traditional cardiovascular risk factors. Although the body of knowledge regarding the correlation between atherosclerosis and immunity in humans is growing rapidly, a relevant proportion of it derives from studies carried out in animal models of cardiovascular disease (CVD). However, while the mouse is a well-suited model, the results obtained therein are not fully transferrable to the human setting due to intrinsic genomic and environmental differences. In the present review, we will discuss mainly human findings, obtained either by examination of post-mortem and surgical atherosclerotic material or through the analysis of the immunological profile of peripheral blood cells. In particular, we will discuss the findings supporting a pro-atherogenic role of T cell subsets, such as effector memory T cells or the potential protective function of regulatory T cells. Recent studies suggest that traditional T cell-driven B2 cell responses appear to be atherogenic, while innate B1 cells appear to exert a protective action through the secretion of naturally occurring antibodies. The insights into the immune pathogenesis of atherosclerosis can provide new targets in the quest for novel therapeutic targets to abate CVD morbidity and mortality. © 2014 British Society for Immunology.

  15. Generation and characterization of tabalumab, a human monoclonal antibody that neutralizes both soluble and membrane-bound B-cell activating factor

    Directory of Open Access Journals (Sweden)

    Manetta J

    2014-08-01

    Full Text Available Joseph Manetta, Holly Bina, Paul Ryan, Niles Fox, Derrick R Witcher, Kristine Kikly Biotechnology Discovery Research, Lilly Research Laboratories, Eli Lilly and Company, Indianapolis, IN, USA Abstract: B-cell activating factor (BAFF is a B-cell survival factor with a key role in B-cell homeostasis and tolerance. Dysregulated BAFF expression may contribute to autoimmune diseases or B-cell malignancies via effects on abnormal B-lymphocyte activation, proliferation, survival, and immunoglobulin secretion. Monoclonal antibodies were generated against human BAFF, characterized for species specificity and affinity, and screened for the ability to neutralize both membrane-bound and soluble BAFF. In addition, studies were undertaken to determine the relative potency of membrane-bound and soluble BAFF. Tabalumab has a high affinity for human, cynomolgus monkey, and rabbit BAFF. No binding to mouse BAFF was detected. Tabalumab was able to neutralize soluble human, cynomolgus monkey, or rabbit BAFF with equal potency. Our data demonstrate that membrane-bound BAFF can be a more potent stimulus for B-cells than soluble BAFF, and tabalumab also neutralized membrane-bound BAFF. Tabalumab prevented BAFF from binding to BAFF receptors and demonstrated pharmacodynamic effects in human BAFF transgenic mice. Tabalumab is a high-affinity human antibody with neutralizing activity against membrane-bound and soluble BAFF. Given our findings that membrane-bound BAFF can have greater in vitro potency than soluble BAFF, neutralization of both forms of BAFF is likely to be important for optimal therapeutic effect. Keywords: autoimmunity, B-cell malignancies, B-cell survival factor, BAFF

  16. Role of reactive oxygen species in arsenic-induced transformation of human lung bronchial epithelial (BEAS-2B) cells

    Energy Technology Data Exchange (ETDEWEB)

    Zhang, Zhuo, E-mail: zhuo.zhang@uky.edu [Graduate Center for Toxicology, University of Kentucky, Lexington, KY 40536 (United States); Pratheeshkumar, Poyil; Budhraja, Amit; Son, Young-Ok [Center for Research on Environmental Diseases, University of Kentucky, Lexington, KY 40536 (United States); Kim, Donghern [Graduate Center for Toxicology, University of Kentucky, Lexington, KY 40536 (United States); Shi, Xianglin [Center for Research on Environmental Diseases, University of Kentucky, Lexington, KY 40536 (United States)

    2015-01-09

    Highlights: • Short term exposure of cells to arsenic causes ROS generation. • Chronical exposure of cells to arsenic causes malignant cell transformation. • Inhibition of ROS generation reduces cell transformation by arsenic. • Arsenic-transformed cells exhibit reduced capacity of generating ROS. • Arsenic-transformed cells exhibit increased levels of antioxidants. - Abstract: Arsenic is an environmental carcinogen, its mechanisms of carcinogenesis remain to be investigated. Reactive oxygen species (ROS) are considered to be important. A previous study (Carpenter et al., 2011) has measured ROS level in human lung bronchial epithelial (BEAS-2B) cells and arsenic-transformed BEAS-2B cells and found that ROS levels were higher in transformed cells than that in parent normal cells. Based on these observations, the authors concluded that cell transformation induced by arsenic is mediated by increased cellular levels of ROS. This conclusion is problematic because this study only measured the basal ROS levels in transformed and parent cells and did not investigate the role of ROS in the process of arsenic-induced cell transformation. The levels of ROS in arsenic-transformed cells represent the result and not the cause of cell transformation. Thus question concerning whether ROS are important in arsenic-induced cell transformation remains to be answered. In the present study, we used expressions of catalase (antioxidant against H{sub 2}O{sub 2}) and superoxide dismutase 2 (SOD2, antioxidant against O{sub 2}{sup ·−}) to decrease ROS level and investigated their role in the process of arsenic-induced cell transformation. Our results show that inhibition of ROS by antioxidant enzymes decreased arsenic-induced cell transformation, demonstrating that ROS are important in this process. We have also shown that in arsenic-transformed cells, ROS generation was lower and levels of antioxidants are higher than those in parent cells, in a disagreement with the previous

  17. Role of reactive oxygen species in arsenic-induced transformation of human lung bronchial epithelial (BEAS-2B) cells

    International Nuclear Information System (INIS)

    Zhang, Zhuo; Pratheeshkumar, Poyil; Budhraja, Amit; Son, Young-Ok; Kim, Donghern; Shi, Xianglin

    2015-01-01

    Highlights: • Short term exposure of cells to arsenic causes ROS generation. • Chronical exposure of cells to arsenic causes malignant cell transformation. • Inhibition of ROS generation reduces cell transformation by arsenic. • Arsenic-transformed cells exhibit reduced capacity of generating ROS. • Arsenic-transformed cells exhibit increased levels of antioxidants. - Abstract: Arsenic is an environmental carcinogen, its mechanisms of carcinogenesis remain to be investigated. Reactive oxygen species (ROS) are considered to be important. A previous study (Carpenter et al., 2011) has measured ROS level in human lung bronchial epithelial (BEAS-2B) cells and arsenic-transformed BEAS-2B cells and found that ROS levels were higher in transformed cells than that in parent normal cells. Based on these observations, the authors concluded that cell transformation induced by arsenic is mediated by increased cellular levels of ROS. This conclusion is problematic because this study only measured the basal ROS levels in transformed and parent cells and did not investigate the role of ROS in the process of arsenic-induced cell transformation. The levels of ROS in arsenic-transformed cells represent the result and not the cause of cell transformation. Thus question concerning whether ROS are important in arsenic-induced cell transformation remains to be answered. In the present study, we used expressions of catalase (antioxidant against H 2 O 2 ) and superoxide dismutase 2 (SOD2, antioxidant against O 2 ·− ) to decrease ROS level and investigated their role in the process of arsenic-induced cell transformation. Our results show that inhibition of ROS by antioxidant enzymes decreased arsenic-induced cell transformation, demonstrating that ROS are important in this process. We have also shown that in arsenic-transformed cells, ROS generation was lower and levels of antioxidants are higher than those in parent cells, in a disagreement with the previous report. The

  18. Identification and fine mapping of a linear B cell epitope of human vimentin

    DEFF Research Database (Denmark)

    Dam, Catharina Essendrup; Houen, Gunnar; Hansen, Paul R.

    2014-01-01

    Knowledge about antibody-antigen interactions is important for the understanding of the immune system mechanisms and for supporting development of drugs and biomarkers. A tool for identification of these antigenic epitopes of specific antibodies is epitope mapping. In this study, a modified enzyme......-linked immunosorbent assay was applied for epitope mapping of a mouse monoclonal vimentin antibody using overlapping resin-bound peptides covering the entire vimentin protein. The minimal epitope required for binding was identified as the LDSLPLVD sequence using N- and C-terminally truncated peptides. The peptide...... sequence LDSLPLVDTH was identified as the complete epitope, corresponding to amino acids 428-437 in the C-terminal end of the human vimentin protein. Alanine scanning and functionality scanning applying substituted peptides were used to identify amino acids essential for antibody reactivity. In particular...

  19. Resveratrol suppresses constitutive activation of AKT via generation of ROS and induces apoptosis in diffuse large B cell lymphoma cell lines.

    Directory of Open Access Journals (Sweden)

    Azhar R Hussain

    Full Text Available BACKGROUND: We have recently shown that deregulation PI3-kinase/AKT survival pathway plays an important role in pathogenesis of diffuse large B cell lymphoma (DLBCL. In an attempt to identify newer therapeutic agents, we investigated the role of Resveratrol (trans-3,4', 5-trihydroxystilbene, a naturally occurring polyphenolic compound on a panel of diffuse large B-cell lymphoma (DLBCL cells in causing inhibition of cell viability and inducing apoptosis. METHODOLOGY/PRINCIPAL FINDINGS: We investigated the action of Resveratrol on DLBCL cells and found that Resveratrol inhibited cell viability and induced apoptosis by inhibition of constitutively activated AKT and its downstream targets via generation of reactive oxygen species (ROS. Simultaneously, Resveratrol treatment of DLBCL cell lines also caused ROS dependent upregulation of DR5; and interestingly, co-treatment of DLBCL with sub-toxic doses of TRAIL and Resveratrol synergistically induced apoptosis via utilizing DR5, on the other hand, gene silencing of DR5 abolished this effect. CONCLUSION/SIGNIFICANCE: Altogether, these data suggest that Resveratrol acts as a suppressor of AKT/PKB pathway leading to apoptosis via generation of ROS and at the same time primes DLBCL cells via up-regulation of DR5 to TRAIL-mediated apoptosis. These data raise the possibility that Resveratrol may have a future therapeutic role in DLBCL and possibly other malignancies with constitutive activation of the AKT/PKB pathway.

  20. Identification of continuous human B-cell epitopes in the VP35, VP40, nucleoprotein and glycoprotein of Ebola virus.

    Directory of Open Access Journals (Sweden)

    Pierre Becquart

    Full Text Available Ebola virus (EBOV is a highly virulent human pathogen. Recovery of infected patients is associated with efficient EBOV-specific immunoglobulin G (IgG responses, whereas fatal outcome is associated with defective humoral immunity. As B-cell epitopes on EBOV are poorly defined, we sought to identify specific epitopes in four EBOV proteins (Glycoprotein (GP, Nucleoprotein (NP, and matrix Viral Protein (VP40 and VP35. For the first time, we tested EBOV IgG+ sera from asymptomatic individuals and symptomatic Gabonese survivors, collected during the early humoral response (seven days after the end of symptoms and the late memory phase (7-12 years post-infection. We also tested sera from EBOV-seropositive patients who had never had clinical signs of hemorrhagic fever or who lived in non-epidemic areas (asymptomatic subjects. We found that serum from asymptomatic individuals was more strongly reactive to VP40 peptides than to GP, NP or VP35. Interestingly, anti-EBOV IgG from asymptomatic patients targeted three immunodominant regions of VP40 reported to play a crucial role in virus assembly and budding. In contrast, serum from most survivors of the three outbreaks, collected a few days after the end of symptoms, reacted mainly with GP peptides. However, in asymptomatic subjects the longest immunodominant domains were identified in GP, and analysis of the GP crystal structure revealed that these domains covered a larger surface area of the chalice bowl formed by three GP1 subunits. The B-cell epitopes we identified in the EBOV VP35, VP40, NP and GP proteins may represent important tools for understanding the humoral response to this virus and for developing new antibody-based therapeutics or detection methods.

  1. Identification of continuous human B-cell epitopes in the VP35, VP40, nucleoprotein and glycoprotein of Ebola virus.

    Science.gov (United States)

    Becquart, Pierre; Mahlakõiv, Tanel; Nkoghe, Dieudonné; Leroy, Eric M

    2014-01-01

    Ebola virus (EBOV) is a highly virulent human pathogen. Recovery of infected patients is associated with efficient EBOV-specific immunoglobulin G (IgG) responses, whereas fatal outcome is associated with defective humoral immunity. As B-cell epitopes on EBOV are poorly defined, we sought to identify specific epitopes in four EBOV proteins (Glycoprotein (GP), Nucleoprotein (NP), and matrix Viral Protein (VP)40 and VP35). For the first time, we tested EBOV IgG+ sera from asymptomatic individuals and symptomatic Gabonese survivors, collected during the early humoral response (seven days after the end of symptoms) and the late memory phase (7-12 years post-infection). We also tested sera from EBOV-seropositive patients who had never had clinical signs of hemorrhagic fever or who lived in non-epidemic areas (asymptomatic subjects). We found that serum from asymptomatic individuals was more strongly reactive to VP40 peptides than to GP, NP or VP35. Interestingly, anti-EBOV IgG from asymptomatic patients targeted three immunodominant regions of VP40 reported to play a crucial role in virus assembly and budding. In contrast, serum from most survivors of the three outbreaks, collected a few days after the end of symptoms, reacted mainly with GP peptides. However, in asymptomatic subjects the longest immunodominant domains were identified in GP, and analysis of the GP crystal structure revealed that these domains covered a larger surface area of the chalice bowl formed by three GP1 subunits. The B-cell epitopes we identified in the EBOV VP35, VP40, NP and GP proteins may represent important tools for understanding the humoral response to this virus and for developing new antibody-based therapeutics or detection methods.

  2. IgE production by normal human B cells induced by alloreactive T cell clones is mediated by IL-4 and suppressed by IFN-gamma

    NARCIS (Netherlands)

    Pène, J.; Rousset, F.; Brière, F.; Chrétien, I.; Paliard, X.; Banchereau, J.; Spits, H.; de Vries, J. E.

    1988-01-01

    Seven T cell clones were established from mixed leukocyte cultures in which PBMC from two healthy donors and from one patient suffering from the hyper-IgE syndrome were stimulated by the irradiated EBV-transformed B cell lines JY or UD53. Five of seven T cell clones, after activation by

  3. Second-line therapy in diffuse large B-cell lymphoma (DLBCL): treatment patterns and outcomes in older patients receiving outpatient chemotherapy.

    Science.gov (United States)

    Danese, Mark D; Griffiths, Robert I; Gleeson, Michelle L; Dalvi, Tapashi; Li, Jingyi; Mikhael, Joseph R; Deeter, Robert; Dreyling, Martin

    2017-05-01

    Using SEER-Medicare linked data we identified elderly patients diagnosed with diffuse large B-cell lymphoma (DLBCL) between January 2000 and December 2007 who received second-line outpatient chemotherapy for relapsed or refractory disease. Second-line regimens were classified into three mutually exclusive groups: aggressive, conventional, and palliative. Of the 632 (426 relapsed, 206 refractory) patients in the cohort, 27.8% received aggressive second-line therapy, 39.1% received conventional therapy, and 33.1% received palliative therapy. There were no differences in survival by type of therapy received, either for relapsed or refractory patients, although the patient risk profile differed significantly. However, duration of remission, male gender, and anemia at diagnosis were important predictors in relapsed patients, and male gender, B-symptoms, comorbidity burden, and poverty status were important predictors in refractory patients. Survival in elderly patients receiving second-line therapy remains poor, and the 24-month cost of all care exceeds $97,000. Patients would benefit from improved treatment options.

  4. B-Cell and T-Cell Immune Responses to Experimental Helicobacter pylori Infection in Humans

    Science.gov (United States)

    Nurgalieva, Zhannat Z.; Conner, Margaret E.; Opekun, Antone R.; Zheng, Carl Q.; Elliott, Susan N.; Ernst, Peter B.; Osato, Michael; Estes, Mary K.; Graham, David Y.

    2005-01-01

    The acute antibody and T-cell immune response to Helicobacter pylori infection in humans has not been studied systematically. Serum from H. pylori-naive volunteers challenged with H. pylori and cured after 4 or 12 weeks was tested by enzyme-linked immunosorbent assays for anti-H. pylori-specific immunoglobulin M (IgM) and IgA established using bacterial lysates from homologous (the infecting strain) and heterologous H. pylori. Proteins recognized by IgM antibody were identified by mass spectrometry of immunoreactive bands separated by two-dimensional gel electrophoresis. Mucosal T-cell subsets (CD4, CD8, CD3, and CD30 cells) were assessed by immunohistochemistry. All 18 infected volunteers developed H. pylori-specific IgM responses to both homologous or heterologous H. pylori antigens. H. pylori antigens reacted with IgM antibody at 4 weeks postinfection. IgM Western blotting showed immunoreactivity of postinfection serum samples to multiple H. pylori proteins with molecular weights ranging between 9,000 (9K) to 150K with homologous strains but only a 70K band using heterologous antigens. Two-dimensional electrophoresis demonstrated that production of H. pylori-specific IgM antibodies was elicited by H. pylori flagellins A and B, urease B, ABC transporter binding protein, heat shock protein 70 (DnaK), and alkyl hydroperoxide reductase. Mucosal CD3, CD4, and CD8 T-cell numbers increased following infection. IgM antibody responses were detected to a range of homologous H. pylori antigens 2 to 4 weeks postchallenge. The majority of H. pylori proteins were those involved in motility and colonization and may represent targets for vaccine development. PMID:15845507

  5. Diverse phosphorylation patterns of B cell receptor-associated signaling in naïve and memory human B cells revealed by phosphoflow, a powerful technique to study signaling at the single cell level

    Directory of Open Access Journals (Sweden)

    Franklin R Toapanta

    2012-10-01

    Full Text Available Following interaction with cognate antigens, B cells undergo cell activation, proliferation and differentiation. Ligation of the B cell receptor (BCR leads to the phosphorylation of BCR-associated signaling proteins within minutes of antigen binding, a process with profound consequences for the fate of the cells and development of effector immunity. Phosphoflow allows a rapid evaluation of various signaling pathways in complex heterogenous cell subsets. This novel technique was used in combination with multi-chromatic flow cytometry and fluorescent-cell barcoding to study phosphorylation of BCR-associated signaling pathways in naïve and memory human B cell subsets. Proteins of the initiation (Syk, propagation (Btk, Akt and integration (p38MAPK and Erk1/2 signaling units were studied. Switched memory (Sm CD27+ and Sm CD27- phosphorylation patterns were similar when stimulated with anti-IgA or -IgG. In contrast, naïve and unswitched memory (Um cells showed significant differences following IgM stimulation. Enhanced phosphorylation of Syk was observed in Um cells, suggesting a lower activation threshold. This is likely the result of higher amounts of IgM on the cell surface, higher pan-Syk levels and enhanced susceptibility to phosphatase inhibition. All other signaling proteins evaluated also showed some degree of enhanced phosphorylation in Um cells. Furthermore, both the PLC-γ2 and PI3K pathways were activated in Um cells, while only the PI3K pathway was activated on naïve cells. Um cells were the only ones that activated signaling pathways when stimulated with fluorescently-labeled S. Typhi and S. pneumoniae. Finally, simultaneous evaluation of signaling proteins at the single cell level (multi-phosphorylated cells revealed that interaction with gram positive and negative bacteria resulted in complex and diverse signaling patterns. Phosphoflow holds great potential to accelerate vaccine development by identifying signaling profiles in good

  6. Expression of the Grb2-related protein of the lymphoid system in B cell subsets enhances B cell antigen receptor signaling through mitogen-activated protein kinase pathways.

    Science.gov (United States)

    Yankee, Thomas M; Solow, Sasha A; Draves, Kevin D; Clark, Edward A

    2003-01-01

    Adapter proteins play a critical role in regulating signals triggered by Ag receptor cross-linking. These small molecules link receptor proximal events with downstream signaling pathways. In this study, we explore the expression and function of the Grb2-related protein of the lymphoid system (GrpL)/Grb2-related adaptor downstream of Shc adapter protein in human B cells. GrpL is expressed in naive B cells and is down-regulated following B cell Ag receptor ligation. By contrast, germinal center and memory B cells express little or no GrpL. Using human B cell lines, we detected constitutive interactions between GrpL and B cell linker protein, Src homology (SH)2 domain-containing leukocyte protein of 76 kDa, hemopoietic progenitor kinase 1, and c-Cbl. The N-terminal SH3 domain of GrpL binds c-Cbl while the C-terminal SH3 domain binds B cell linker protein and SH2 domain-containing leukocyte protein of 76 kDa. Exogenous expression of GrpL in a GrpL-negative B cell line leads to enhanced Ag receptor-induced extracellular signal-related kinase and p38 mitogen-activated protein kinase phosphorylation. Thus, GrpL expression in human B cell subsets appears to regulate Ag receptor-mediated signaling events.

  7. B-cell infiltration and frequency of cytokine producing cells differ between localized and disseminated human cutaneous leishmaniases

    Directory of Open Access Journals (Sweden)

    MGS Vieira

    2002-10-01

    Full Text Available Biopsies from human localized cutaneous lesions (LCL n = 7 or disseminated lesions (DL n = 8 cases were characterized according to cellular infiltration,frequency of cytokine (IFN-g, TNF-alpha or iNOS enzyme producing cells. LCL, the most usual form of the disease with usually one or two lesions, exhibits extensive tissue damage. DL is a rare form with widespread lesions throughout the body; exhibiting poor parasite containment but less tissue damage. We demonstrated that LCL lesions exhibit higher frequency of B lymphocytes and a higher intensity of IFN-gamma expression. In both forms of the disease CD8+ were found in higher frequency than CD4+ T cells. Frequency of TNF-alpha and iNOS producing cells, as well as the frequency of CD68+ macrophages, did not differ between LCL and DL. Our findings reinforce the link between an efficient control of parasite and tissue damage, implicating higher frequency of IFN-gamma producing cells, as well as its possible counteraction by infiltrated B cells and hence possible humoral immune response in situ.

  8. Radioimmunotherapy for first-line and relapse treatment of aggressive B-cell non-Hodgkin lymphoma: an analysis of 215 patients registered in the international RIT-Network

    Energy Technology Data Exchange (ETDEWEB)

    Hohloch, Karin; Lankeit, H.K.; Truemper, L. [Georg August University, Hematology and Oncology, Goettingen (Germany); Zinzani, P.L. [University of Bologna, Institute of Hematology and Medical Oncology ' ' L. e A. Seragnoli' ' , Bologna (Italy); Scholz, C.W. [Charite, University Berlin, Hematology, Oncology and Tumor Immunology, Berlin (Germany); Lorsbach, M.; Windemuth-Kieselbach, C. [Alcedis GmbH, Giessen (Germany)

    2014-08-15

    Very few reliable clinical data about the use of radioimmunotherapy in aggressive B-cell lymphoma exist. Patients with aggressive B-cell lymphoma registered in the international RIT-Network were analysed with regard to prior treatment, response and side effects. The RIT-Network is a web-based registry that collects observational data from radioimmunotherapy-treated patients with malignant lymphoma across 13 countries. This analysis included 215 with aggressive B-cell lymphoma out of 232 patients registered in the RIT-Network. Histological subtypes were as follows: 190 diffuse large B-cell, 15 primary mediastinal, 9 anaplastic large cell, and 1 intravascular lymphoma. The median age of the patients was 62 years (range 17 - 88), with 27 % above the age of 70 years. Radioimmunotherapy was mainly used as consolidation after first-line or second-line chemotherapy (56.1 %), as part of third-line to eighth-line therapy for relapse (16.4 %), and in refractory disease (12.2 %). Grade IV neutropenia and thrombopenia and grade III anaemia were observed. The median time to recovery of blood count was 81 days (range 0 - 600 days). The overall response rate was 63.3 %. The complete response rate was 76.4 % in patients treated as part of first-line therapy, and 44.3 % in patients with relapse. Mean overall survival in first-line therapy patients was 32.7 months and 14.0 months in patients with relapse or refractory disease, respectively. Most patients with aggressive B-cell lymphoma in the RIT-Network received radioimmunotherapy as consolidation after first-line therapy with excellent complete remission and overall survival rates compared to published data. In relapsed aggressive B-cell lymphoma, radioimmunotherapy is a safe and feasible treatment leading to satisfactory response rates with acceptable toxicity. (orig.)

  9. Human lymphoma mutations reveal CARD11 as the switch between self-antigen–induced B cell death or proliferation and autoantibody production

    Science.gov (United States)

    Jeelall, Yogesh S.; Wang, James Q.; Law, Hsei-Di; Domaschenz, Heather; Fung, Herman K.H.; Kallies, Axel; Nutt, Stephen L.

    2012-01-01

    Self-tolerance and immunity are actively acquired in parallel through a poorly understood ability of antigen receptors to switch between signaling death or proliferation of antigen-binding lymphocytes in different contexts. It is not known whether this tolerance-immunity switch requires global rewiring of the signaling apparatus or if it can arise from a single molecular change. By introducing individual CARD11 mutations found in human lymphomas into antigen-activated mature B lymphocytes in mice, we find here that lymphoma-derived CARD11 mutations switch the effect of self-antigen from inducing B cell death into T cell–independent proliferation, Blimp1-mediated plasmablast differentiation, and autoantibody secretion. Our findings demonstrate that regulation of CARD11 signaling is a critical switch governing the decision between death and proliferation in antigen-stimulated mature B cells and that mutations in this switch represent a powerful initiator for aberrant B cell responses in vivo. PMID:23027925

  10. Effects of JS-K, a novel anti-cancer nitric oxide prodrug, on gene expression in human hepatoma Hep3B cells.

    Science.gov (United States)

    Dong, Ray; Wang, Xueqian; Wang, Huan; Liu, Zhengyun; Liu, Jie; Saavedra, Joseph E

    2017-04-01

    JS-K is a novel anticancer nitric oxide (NO) prodrug effective against a variety of cancer cells, including the inhibition of AM-1 hepatoma cell growth in rats. To further evaluate anticancer effects of JS-K, human hepatoma Hep3B cells were treated with JS-K and the compound control JS-43-126 at various concentrations (0-100μM) for 24h, and cytotoxicity was determined by the MTS assay. The compound control JS-43-126 was not cytotoxic to Hep3B cells at concentrations up to 100μM, while the LC 50 for JS-K was about 10μM. To examine the molecular mechanisms of antitumor effects of JS-K, Hep3B cells were treated with 1-10μM of JS-K for 24h, and then subjected to gene expression analysis via real time RT-PCR and protein immunostain via confocal images. JS-K is a GST-α targeting NO prodrug, and decreased immunostaining for GST-α was associated with JS-K treatment. JS-K activated apoptosis pathways in Hep3B cells, including induction of caspase-3, caspase-9, Bax, TNF-α, and IL-1β, and immunostaining for caspase-3 was intensified. The expressions of thrombospondin-1 (TSP-1) and the tissue inhibitors of metalloproteinase-1 (TIMP-1) were increased by JS-K at both transcript and protein levels. JS-K treatment also increased the expression of differentiation-related genes CD14 and CD11b, and depressed the expression of c-myc in Hep3B cells. Thus, multiple molecular events appear to be associated with anticancer effects of JS-K in human hepatoma Hep3B cells, including activation of genes related to apoptosis and induction of genes involved in antiangiogenesis and tumor cell migration. Copyright © 2017 Elsevier Masson SAS. All rights reserved.

  11. B-cell depletion is protective against anti-AAV capsid immune response: a human subject case study

    Directory of Open Access Journals (Sweden)

    M Corti

    2014-01-01

    Full Text Available Gene therapy strategies for congenital myopathies may require repeat administration of adeno-associated viral (AAV vectors due to aspects of the clinical application, such as: (i administration of doses below therapeutic efficacy in patients enrolled in early phase clinical trials; (ii progressive reduction of the therapeutic gene expression over time as a result of increasing muscle mass in patients treated at a young age; and (iii a possibly faster depletion of pathogenic myofibers in this patient population. Immune response triggered by the first vector administration, and to subsequent doses, represents a major obstacle for successful gene transfer in young patients. Anti-capsid and anti-transgene product related humoral and cell-mediated responses have been previously observed in all preclinical models and human subjects who received gene therapy or enzyme replacement therapy (ERT for congenital myopathies. Immune responses may result in reduced efficacy of the gene transfer over time and/or may preclude for the possibility of re-administration of the same vector. In this study, we evaluated the immune response of a Pompe patient dosed with an AAV1-GAA vector after receiving Rituximab and Sirolimus to modulate reactions against ERT. A key finding of this single subject case report is the observation that B-cell ablation with rituximab prior to AAV vector exposure results in non-responsiveness to both capsid and transgene, therefore allowing the possibility of repeat administration in the future. This observation is significant for future gene therapy studies and establishes a clinically relevant approach to blocking immune responses to AAV vectors.

  12. 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...... peripheral blood lymphocytes caused by the low abundance of antigen-specific B cells in the circulation. The preselection of B cells is based on the specificity of the surface Ig receptor and is accomplished using the antigen of interest conjugated to magnetic beads. This method should significantly increase...... the frequency of antibody phage particles of interest in the library and allow for efficient isolation monoclonal antibodies with the predefined specificity....

  13. Proof of the concept to use a malignant B cell line drug screen strategy for identification and weight of melphalan resistance genes in multiple myeloma.

    Directory of Open Access Journals (Sweden)

    Martin Bøgsted

    Full Text Available In a conceptual study of drug resistance we have used a preclinical model of malignant B-cell lines by combining drug induced growth inhibition and gene expression profiling. In the current report a melphalan resistance profile of 19 genes were weighted by microarray data from the MRC Myeloma IX trial and time to progression following high dose melphalan, to generate an individual melphalan resistance index. The resistance index was subsequently validated in the HOVON65/GMMG-HD4 trial data set to prove the concept. Biologically, the assigned resistance indices were differentially distributed among translocations and cyclin D expression classes. Clinically, the 25% most melphalan resistant, the intermediate 50% and the 25% most sensitive patients had a median progression free survival of 18, 32 and 28 months, respectively (log-rank P-value  = 0.05. Furthermore, the median overall survival was 45 months for the resistant group and not reached for the intermediate and sensitive groups (log-rank P-value  = 0.003 following 38 months median observation. In a multivariate analysis, correcting for age, sex and ISS-staging, we found a high resistance index to be an independent variable associated with inferior progression free survival and overall survival. This study provides clinical proof of concept to use in vitro drug screen for identification of melphalan resistance gene signatures for future functional analysis.

  14. Homeostatic 'bystander' proliferation of human peripheral blood B cells in response to polyclonal T-cell stimulation in vitro.

    Science.gov (United States)

    Jasiulewicz, Aleksandra; Lisowska, Katarzyna A; Pietruczuk, Krzysztof; Frąckowiak, Joanna; Fulop, Tamas; Witkowski, Jacek M

    2015-11-01

    The mechanisms of maintenance of adequate numbers of B lymphocytes and of protective levels of immunoglobulins in the absence of antigenic (re)stimulation remain not fully understood. Meanwhile, our results presented here show that both peripheral blood naive and memory B cells can be activated strongly and non-specifically (in a mitogen-like fashion) in 5-day in vitro cultures of anti-CD3- or concanavalin A (Con A)-stimulated peripheral blood mononuclear cells of healthy people. This polyclonal, bystander activation of the B cells includes multiple divisions of most of them (assessed here by the flow cytometric technique of dividing cell tracking) and significant antibody [immunoglobulin M (IgM) and IgG] secretion. Observed proliferation of the CD19(+) B cells depends on contact with stimulated T helper (Th) cells (via CD40-CD40L interaction) and on the response of B cells to secreted interleukins IL-5, IL-10 and IL-4, and is correlated with the levels of these Th-derived molecules, while it does not involve the ligation of the BCR/CD19 complex. We suggest that the effect might reflect the situation occurring in vivo as the homeostatic proliferation of otherwise non-stimulated, peripheral B lymphocytes, providing an always ready pool for efficient antibody production to any new (or cognate) antigen challenge. © The Japanese Society for Immunology. 2015. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.

  15. Activation of cross-reactive mucosal T and B cell responses in human nasopharynx-associated lymphoid tissue in vitro by Modified Vaccinia Ankara-vectored influenza vaccines.

    Science.gov (United States)

    Mullin, Jennifer; Ahmed, Muhammed S; Sharma, Ravi; Upile, Navdeep; Beer, Helen; Achar, Priya; Puksuriwong, Suttida; Ferrara, Francesca; Temperton, Nigel; McNamara, Paul; Lambe, Teresa; Gilbert, Sarah C; Zhang, Qibo

    2016-03-29

    Recent efforts have been focused on the development of vaccines that could induce broad immunity against influenza virus, either through T cell responses to conserved internal antigens or B cell response to cross-reactive haemagglutinin (HA). We studied the capacity of Modified Vaccinia Ankara (MVA)-vectored influenza vaccines to induce cross-reactive immunity to influenza virus in human nasopharynx-associated lymphoid tissue (NALT) in vitro. Adenotonsillar cells were isolated and stimulated with MVA vaccines expressing either conserved nucleoprotein (NP) and matrix protein 1 (M1) (MVA-NP-M1) or pandemic H1N1 HA (MVA-pdmH1HA). The MVA vaccine uptake and expression, and T and B cell responses were analyzed. MVA-vectored vaccines were highly efficient infecting NALT and vaccine antigens were highly expressed by B cells. MVA-NP-M1 elicited T cell response with greater numbers of IFNγ-producing CD4+ T cells and tissue-resident memory T cells than controls. MVA-pdmH1HA induced cross-reactive anti-HA antibodies to a number of influenza subtypes, in an age-dependent manner. The cross-reactive antibodies include anti-avian H5N1 and mainly target HA2 domain. MVA vaccines are efficient in infecting NALT and the vaccine antigen is highly expressed by B cells. MVA vaccines expressing conserved influenza antigens induce cross-reactive T and B cell responses in human NALT in vitro, suggesting the potential as mucosal vaccines for broader immunity against influenza. Copyright © 2016 Elsevier Ltd. All rights reserved.

  16. Human recombinant Fab fragment from combinatorial libraries of a B-cell lymphoma patient recognizes core protein of chondroitin sulphate proteoglycan 4.

    Science.gov (United States)

    Egami, Yoko; Narushima, Yuta; Ohshima, Motohiro; Yoshida, Akira; Yoneta, Naruki; Masaki, Yasufumi; Itoh, Kunihiko

    2018-01-01

    CD antigens are well known as therapeutic targets of B-cell lymphoma. To isolate therapeutic antibodies that recognize novel targets other than CD antigens, we constructed a phage display combinatorial antibody Fab library from bone marrow lymphocytes of B-cell lymphoma patient. To eliminate antibodies reactive with known B-cell lymphoma antigen, non-hematopoietic and patient's sera reactive HeLaS3 cells was selected as a target of whole cell panning. Five rounds of panning against live HeLaS3 cells retrieved single Fab clone, termed AHSA (Antibody to HeLa Surface Antigen). Using phage display random peptide library, LSYLEP was identified as an epitope sequence of AHSA. LC-MS/MS analysis of AHSA-precipitated HeLaS3 cell lysates detected several fragments corresponding to the sequence of chondroitin sulphate proteoglycan 4 (CSPG4) core protein. Since LSYLEP sequence was at the position of 313-318 of CSPG4, we considered that CSPG4 was AHSA-associated antigen. Double staining of CSPG4-postive MDA-MB-435S cells with AHSA and anti-CSPG4 rabbit antibody showed identical staining position, and reduced AHSA reactivity was observed in CSPG4-siRNA treated MDA-MB-435S cells. In conclusion, we retrieved a human Fab from antibody library of B-cell lymphoma patient, and identified CSPG4 as a recognizing antigen. AHSA may have potential benefits for development of CSPG4-targeting theranostics for B-cell lymphoma. © The Authors 2017. Published by Oxford University Press on behalf of the Japanese Biochemical Society. All rights reserved.

  17. Circulating CXCR5⁺CD4⁺ T Follicular-Like Helper Cell and Memory B Cell Responses to Human Papillomavirus Vaccines.

    Directory of Open Access Journals (Sweden)

    Ken Matsui

    Full Text Available Through the interaction of T follicular helper (Tfh cells and B cells, efficacious vaccines can generate high-affinity, pathogen-neutralizing antibodies, and memory B cells. Using CXCR5, CXCR3, CCR6, CCR7, PD1, and ICOS as markers, Tfh-like cells can be identified in the circulation and be classified into three functionally distinct subsets that are PD1+ICOS+, PD1+ ICOS-, or PD1-ICOS-. We used these markers to identify different subsets of CXCR5+CD4+ Tfh-like cells in response to highly immunogenic and efficacious vaccines for human papillomaviruses (HPV: Cervarix and Gardasil. In this small study, we used PBMC samples from 11 Gardasil recipients, and 8 Cervarix recipients from the Vaccine Research Center 902 Study to examine the induction of circulating Tfh-like cells and IgD-CD38HiCD27+ memory B cells by flow cytometry. PD1+ICOS+ CXCR3+CCR6-CXCR5+CD4+ (Tfh1-like cells were induced and peaked on Day (D 7 post-first vaccination, but not as much on D7 post-third vaccination. We also observed a trend toward increase in PD1+ICOS+ CXCR3-CCR6-CXCR5+CD4+ (Tfh2-like cells for both vaccines, and PD1+ICOS+ CXCR3-CCR6+CXCR5+CD4+ (Tfh17-like subset was induced by Cervarix post-first vaccination. There were also minimal changes in the other cellular subsets. In addition, Cervarix recipients had more memory B cells post-first vaccination than did Gardasil recipients at D14 and D30. We found frequencies of memory B cells at D30 correlated with anti-HPV16 and 18 antibody titers from D30, and the induction levels of memory B cells at D30 and PD1+ICOS+Tfh1-like cells at D7 post-first vaccination correlated for Cervarix. Our study showed that induction of circulating CXCR5+CD4+ Tfh-like subsets can be detected following immunization with HPV vaccines, and potentially be useful as a marker of immunogenicity of vaccines. However, further investigations should be extended to different cohorts with larger sample size to better understand the functions of these T

  18. Phosphodiesterase profile of human B lymphocytes from normal and atopic donors and the effects of PDE inhibition on B cell proliferation

    Science.gov (United States)

    Gantner, Florian; Götz, Christine; Gekeler, Volker; Schudt, Christian; Wendel, Albrecht; Hatzelmann, Armin

    1998-01-01

    CD19+ B lymphocytes were purified from the peripheral blood of normal and atopic subjects to analyse and compare the phosphodiesterase (PDE) activity profile, PDE mRNA expression and the importance of PDE activity for the regulation of B cell function.The majority of cyclic AMP hydrolyzing activity of human B cells was cytosolic PDE4, followed by cytosolic PDE7-like activity; marginal PDE3 activity was found only in the particulate B cell fraction. PDE1, PDE2 and PDE5 activities were not detected.By cDNA-PCR analysis mRNA of the PDE4 subtypes A, B (splice variant PDE4B2) and D were detected. In addition, a weak signal for PDE3A was found.No differences in PDE activities or mRNA expression of PDE subtypes were found in B cells from either normal or atopic subjects.Stimulation of B lymphocytes with the polyclonal stimulus lipopolysaccharide (LPS) induced a proliferative response in a time- and concentration-dependent manner, which was increased in the presence of interleukin-4 (IL-4). PDE4 inhibitors (rolipram, piclamilast) led to an increase in the cellular cyclic AMP concentration and to an augmentation of proliferation, whereas a PDE3 inhibitor (motapizone) was ineffective, which is in accordance with the PDE profile found. The proliferation enhancing effect of the PDE4 inhibitors was partly mimicked by the cyclic AMP analogues dibutyryl (db) cyclic AMP and 5,6-dichloro-1-β-D-ribofuranosylbenzimidazole-3′,5′-cyclic monophosphorothioate, Sp-isomer (dcl-cBIMPS), respectively. However, at concentrations exceeding 100 μM db-cyclic AMP suppressed B lymphocyte proliferation, probably as a result of cytotoxicity. Prostaglandin E2 (PGE2, 1 μM) and forskolin (10 μM) did not affect B cell proliferation, even when given in combination with rolipram.Inhibition of protein kinase A (PKA) by differentially acting selective inhibitors (KT 5720, Rp-8-Br-cyclic AMPS) decreased the proliferative response of control cells and reversed the proliferation enhancing effects

  19. 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)

    Fleisher, T.A.; Greene, W.C.; Blaese, R.M.; Waldmann, T.A.

    1981-01-01

    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

  20. Modulation of the human equilibrative nucleoside transporter1 (hENT1) activity by IL-4 and PMA in B cells from chronic lymphocytic leukemia.

    Science.gov (United States)

    Fernández Calotti, Paula; Galmarini, Carlos María; Cañones, Cristian; Gamberale, Romina; Saénz, Daniel; Avalos, Julio Sánchez; Chianelli, Mónica; Rosenstein, Ruth; Giordano, Mirta

    2008-02-15

    Nucleoside transporters (NTs) are essential for the uptake of therapeutic nucleoside analogs, broadly used in cancer treatment. The mechanisms responsible for NT regulation are largely unknown. IL-4 is a pro-survival signal for chronic lymphocytic leukemia (CLL) cells and has been shown to confer resistance to nucleoside analogs. The aim of this study was to investigate whether IL-4 is able to modulate the expression and function of the human equilibrative NT1 (hENT1) in primary cultures of CLL cells and, consequently, to affect cytotoxicity induced by therapeutic nucleosides analogs. We found that treatment with IL-4 (20 ng/ml for 24 h) increased mRNA hENT1 expression in CLL cells without affecting that of normal B cells. Given that the enhanced mRNA levels of hENT1 in CLL cells did not result in increased transport activity, we examined the possibility that hENT1 induced by IL-4 may require post-translational modifications to become active. We found that the acute stimulation of PKC in IL-4-treated CLL cells by short-term incubation with PMA significantly increased hENT1 transport activity and favoured fludarabine-induced apoptosis. By contrast, and in line with previous reports, IL-4 plus PMA protected CLL cells from a variety of cytotoxic agents. Our findings indicate that the combined treatment with IL-4 and PMA enhances hENT1 activity and specifically sensitizes CLL cells to undergo apoptosis induced by fludarabine.

  1. Polymorphism of 41 kD Flagellin Gene and Its Human B-Cell Epitope in Borrelia burgdorferi Strains of China

    Directory of Open Access Journals (Sweden)

    Huixin Liu

    2016-01-01

    Full Text Available The 41 kD flagellin of Borrelia burgdorferi (B. burgdorferi is a major component of periplasmic flagellar filament core and a good candidate for serodiagnosis in early stage of Lyme disease. Here, we chose 89 B. burgdorferi strains in China, amplified the gene encoding the 41 kD flagellin, and compared the sequences. The results showed that genetic diversity presented in the 41 kD flagellin genes of all 89 strains among the four genotypes of B. burgdorferi, especially in the genotype of B. garinii. Some specific mutation sites for each genotype of the 41 kD flagellin genes were found, which could be used for genotyping B. burgdorferi strains in China. Human B-cell epitope analysis showed that thirteen of 15 nonsynonymous mutations occurred in the epitope region of 41 kD flagellin and thirty of 42 B-cell epitopes were altered due to all 13 nonsynonymous mutations in the epitope region, which may affect the function of the antigen. Nonsynonymous mutations and changed human B-cell epitopes exist in 41 kD flagellin of B. burgdorferi sensu lato strains; these changes should be considered in serodiagnosis of Lyme disease.

  2. Rituximab plus chemotherapy as first-line treatment in Chinese patients with diffuse large B-cell lymphoma in routine practice: a prospective, multicentre, non-interventional study

    International Nuclear Information System (INIS)

    Wu, Jianqiu; Song, Yongping; Su, Liping; Xu, Li; Chen, Tingchao

    2016-01-01

    The efficacy and safety of rituximab-based chemotherapy (R-chemo), the standard regimen for patients with diffuse large B-cell lymphoma (DLBCL), which is more common in Asia than in Western countries, are well confirmed in randomized controlled trials (RCTs). However, the safety and effectiveness of R-chemo in patients who are largely excluded from RCTs have not been well characterized. This real-world study investigated the safety and effectiveness of R-chemo as first-line treatment in Chinese patients with DLBCL. Treatment-naive DLBCL patients who were CD20 positive and eligible to receive R-chemo were enrolled with no specific exclusion criteria. Data collected at baseline included age, gender, disease stage, international prognostic index (IPI), B symptoms, extranodal involvement, performance status, and medical history. In the present study, data on safety, treatment effectiveness, and HBV infection management were collected 120 days after the last R-chemo administration. Overall, R-chemo was well tolerated. The safety profile of R-chemo in patients with a history of heart or liver disease was well described without any additional unexpected safety concerns. The overall response rate (ORR) in the Chinese patients from this study was 94.2 % (complete response [CR], 55.0 %; CR unconfirmed [CRu] 18.2 %; and partial response [PR], 20.9 %). Compared to patients with no history of disease, the CR and PR rates of patients with a history of heart or liver disease were lower and higher, respectively; this tendency could be in part explained by treatment interruptions in patients with heart or liver diseases. HBsAg positivity and a maximum tumor diameter of ≥7.5 cm negatively correlated with CR + CRu, whereas age and HBsAg positivity negatively correlated with CR. This study further validated the safety and effectiveness of R-chemo in Chinese patients with DLBCL. Patients with a history of heart or liver disease may further benefit from R-chemo if preventive measures

  3. A human monoclonal antibody drug and target discovery platform for B-cell chronic lymphocytic leukemia based on allogeneic hematopoietic stem cell transplantation and phage display

    OpenAIRE

    Baskar, Sivasubramanian; Suschak, Jessica M.; Samija, Ivan; Srinivasan, Ramaprasad; Childs, Richard W.; Pavletic, Steven Z.; Bishop, Michael R.; Rader, Christoph

    2009-01-01

    Allogeneic hematopoietic stem cell transplantation (alloHSCT) is the only potentially curative treatment available for patients with B-cell chronic lymphocytic leukemia (B-CLL). Here, we show that post-alloHSCT antibody repertoires can be mined for the discovery of fully human monoclonal antibodies to B-CLL cell-surface antigens. Sera collected from B-CLL patients at defined times after alloHSCT showed selective binding to primary B-CLL cells. Pre-alloHSCT sera, donor sera, and control sera w...

  4. NKT Cell Responses to B Cell Lymphoma.

    Science.gov (United States)

    Li, Junxin; Sun, Wenji; Subrahmanyam, Priyanka B; Page, Carly; Younger, Kenisha M; Tiper, Irina V; Frieman, Matthew; Kimball, Amy S; Webb, Tonya J

    2014-06-01

    Natural killer T (NKT) cells are a unique subset of CD1d-restricted T lymphocytes that express characteristics of both T cells and natural killer cells. NKT cells mediate tumor immune-surveillance; however, NKT cells are numerically reduced and functionally impaired in lymphoma patients. Many hematologic malignancies express CD1d molecules and co-stimulatory proteins needed to induce anti-tumor immunity by NKT cells, yet most tumors are poorly immunogenic. In this study, we sought to investigate NKT cell responses to B cell lymphoma. In the presence of exogenous antigen, both mouse and human NKT cell lines produce cytokines following stimulation by B cell lymphoma lines. NKT cell populations were examined ex vivo in mouse models of spontaneous B cell lymphoma, and it was found that during early stages, NKT cell responses were enhanced in lymphoma-bearing animals compared to disease-free animals. In contrast, in lymphoma-bearing animals with splenomegaly and lymphadenopathy, NKT cells were functionally impaired. In a mouse model of blastoid variant mantle cell lymphoma, treatment of tumor-bearing mice with a potent NKT cell agonist, α-galactosylceramide (α-GalCer), resulted in a significant decrease in disease pathology. Ex vivo studies demonstrated that NKT cells from α-GalCer treated mice produced IFN-γ following α-GalCer restimulation, unlike NKT cells from vehicle-control treated mice. These data demonstrate an important role for NKT cells in the immune response to an aggressive hematologic malignancy like mantle cell lymphoma.

  5. Enhancement of B-cell receptor signaling by a point mutation of adaptor protein 3BP2 identified in human inherited disease cherubism.

    Science.gov (United States)

    Ogi, Kazuhiro; Nakashima, Kenji; Chihara, Kazuyasu; Takeuchi, Kenji; Horiguchi, Tomoko; Fujieda, Shigeharu; Sada, Kiyonao

    2011-09-01

    Tyrosine phosphorylation of adaptor protein c-Abl-Src homology 3 (SH3) domain-binding protein-2 (3BP2, also referred to SH3BP2) positively regulates the B-cell antigen receptor (BCR)-mediated signal transduction, leading to the activation of nuclear factor of activated T cells (NFAT). Here we showed the effect of the proline to arginine substitution of 3BP2 in which is the most common mutation in patients with cherubism (P418R) on B-cell receptor signaling. Comparing to the wild type, overexpression of the mutant form of 3BP2 (3BP2-P416R, corresponding to P418R in human protein) enhanced BCR-mediated activation of NFAT. 3BP2-P416R increased the signaling complex formation with Syk, phospholipase C-γ2 (PLC-γ2), and Vav1. In contrast, 3BP2-P416R could not change the association with the negative regulator 14-3-3. Loss of the association mutant that was incapable to associate with 14-3-3 could not mimic BCR-mediated NFAT activation in Syk-deficient cells. Moreover, BCR-mediated phosphorylation of extracellular signal regulated kinase (ERK) and c-Jun N-terminal kinase (JNK) was not affected by P416R mutation. These results showed that P416R mutation of 3BP2 causes the gain of function in B cells by increasing the interaction with specific signaling molecules. © 2011 The Authors. Journal compilation © 2011 by the Molecular Biology Society of Japan/Blackwell Publishing Ltd.

  6. Lymphoid-Like Structures with Distinct B Cell Areas in Kidney Allografts are not Predictive for Graft Rejection. A Non-human Primate Study

    NARCIS (Netherlands)

    Jonker, Margreet; Wubben, Jacqueline A. M.; 't Hart, Bert A.; Haanstra, Krista G.

    2015-01-01

    Kidney allograft biopsies were analyzed for the presence of B cell clusters/aggregates using CD20 staining. Few B cells were found in the diffuse interstitial infiltrates, but clusters of B cells were found in nodular infiltrates. These nodular infiltrates were smaller shortly after transplantation,

  7. TLR5 signaling enhances the proliferation of human allogeneic CD40-activated B cell induced CD4hiCD25+ regulatory T cells.

    Directory of Open Access Journals (Sweden)

    Ping-Lung Chan

    Full Text Available Although diverse functions of different toll-like receptors (TLR on human natural regulatory T cells have been demonstrated recently, the role of TLR-related signals on human induced regulatory T cells remain elusive. Previously our group developed an ex vivo high-efficient system in generating human alloantigen-specific CD4(hiCD25(+ regulatory T cells from naïve CD4(+CD25(- T cells using allogeneic CD40-activated B cells as stimulators. In this study, we investigated the role of TLR5-related signals on the generation and function of these novel CD4(hiCD25(+ regulatory T cells. It was found that induced CD4(hiCD25(+ regulatory T cells expressed an up-regulated level of TLR5 compared to their precursors. The blockade of TLR5 using anti-TLR5 antibodies during the co-culture decreased CD4(hiCD25(+ regulatory T cells proliferation by induction of S phase arrest. The S phase arrest was associated with reduced ERK1/2 phosphorylation. However, TLR5 blockade did not decrease the CTLA-4, GITR and FOXP3 expressions, and the suppressive function of CD4(hiCD25(+ regulatory T cells. In conclusion, we discovered a novel function of TLR5-related signaling in enhancing the proliferation of CD4(hiCD25(+ regulatory T cells by promoting S phase progress but not involved in the suppressive function of human CD40-activated B cell-induced CD4(hiCD25(+ regulatory T cells, suggesting a novel role of TLR5-related signals in the generation of induced regulatory T cells.

  8. Immunoglobulin production induced in vitro by glucocorticoid hormones: T cell-dependent stimulation of immunoglobulin production without B cell proliferation in cultures of human peripheral blood lymphocytes

    International Nuclear Information System (INIS)

    Grayson, J.; Dooley, N.J.; Koski, I.R.; Blaese, R.M.

    1981-01-01

    The direct effects of steroid hormones on the production of immunoglobulins and DNA synthesis by human T and B lymphocytes was evaluated in cultures of peripheral blood mononuclear cells. As detected by a reverse hemolytic plaque assay, the addition of 0.1 mM to 10 nM hydrocortisone to lymphocytes in culture in the absence of other stimulants or mitogens, resulted in the dramatic induction of immunoglobulin production with responses comparable to those seen in similar cultures stimulated with pokeweed mitogen. Steroid-stimulated immunoglobulin production was first seen after 48 h and peaked at 8-10 d of culture. The production of IgG, IgA, and IgM was induced following incubation with steroid. Glucocorticoids, but not estrogens or androgens, were capable of mediating this effect, and only compounds with affinity for the glucocorticoid receptor were active. The induction of immunoglobulin production was dependent on both T cells and monocytes; cultures depleted of either cell type did not produce immunoglobulin when stimulated with glucocorticoid hormones. Proliferation of B cells or T cells could not be detected by [/sup 3/H]thymidine incorporation or total cell recovery from steroid-stimulated cultures, even though such cultures demonstrated marked increases in immunoglobulin production. The mechanism responsible for this functional maturation of B cells to become high rate immunoglobulin producing cells is as yet undefined, although it appears to involve more than merely steroid mediated inactivation of suppressor T cells

  9. Nicotine Component of Cigarette Smoke Extract (CSE) Decreases the Cytotoxicity of CSE in BEAS-2B Cells Stably Expressing Human Cytochrome P450 2A13.

    Science.gov (United States)

    Ji, Minghui; Zhang, Yudong; Li, Na; Wang, Chao; Xia, Rong; Zhang, Zhan; Wang, Shou-Lin

    2017-10-13

    Cytochrome P450 2A13 (CYP2A13), an extrahepatic enzyme mainly expressed in the human respiratory system, has been reported to mediate the metabolism and toxicity of cigarette smoke. We previously found that nicotine inhibited 4-(methylnitrosamino)-1-(3-pyridyl)-1-butanone (NNK) metabolism by CYP2A13, but its influence on other components of cigarette smoke remains unclear. The nicotine component of cigarette smoke extract (CSE) was separated, purified, and identified using high-performance liquid chromatography (HPLC) and ultra-performance liquid chromatography tandem mass spectrometry (UPLC-MS/MS), splitting CSE into a nicotine section (CSE-N) and nicotine-free section (CSE-O). Cell viability and apoptosis by Cell Counting Kit-8 (CCK-8) and flow cytometry assays were conducted on immortalized human bronchial epithelial (BEAS-2B) cells stably expressing CYP2A13 (B-2A13) or vector (B-V), respectively. Interestingly, CSE and CSE-O were toxic to BEAS-2B cells whereas CSE-N showed less cytotoxicity. CSE-O was more toxic to B-2A13 cells than to B-V cells (IC 50 of 2.49% vs. 7.06%), which was flatted by 8-methoxypsoralen (8-MOP), a CYP inhibitor. CSE-O rather than CSE or CSE-N increased apoptosis of B-2A13 cells rather than B-V cells. Accordingly, compared to CSE-N and CSE, CSE-O significantly changed the expression of three pairs of pro- and anti-apoptotic proteins, Bcl-2 Associated X Protein/B cell lymphoma-2 (Bax/Bcl-2), Cleaved Poly (Adenosine Diphosphate-Ribose) Polymerase/Poly (Adenosine Diphosphate-Ribose) Polymerase (C-PARP/PARP), and C-caspase-3/caspase-3, in B-2A13 cells. In addition, recombination of CSE-N and CSE-O (CSE-O/N) showed similar cytotoxicity and apoptosis to the original CSE. These results demonstrate that the nicotine component decreases the metabolic activation of CYP2A13 to CSE and aids in understanding the critical role of CYP2A13 in human respiratory diseases caused by cigarette smoking.

  10. A human monoclonal antibody drug and target discovery platform for B-cell chronic lymphocytic leukemia based on allogeneic hematopoietic stem cell transplantation and phage display.

    Science.gov (United States)

    Baskar, Sivasubramanian; Suschak, Jessica M; Samija, Ivan; Srinivasan, Ramaprasad; Childs, Richard W; Pavletic, Steven Z; Bishop, Michael R; Rader, Christoph

    2009-11-12

    Allogeneic hematopoietic stem cell transplantation (alloHSCT) is the only potentially curative treatment available for patients with B-cell chronic lymphocytic leukemia (B-CLL). Here, we show that post-alloHSCT antibody repertoires can be mined for the discovery of fully human monoclonal antibodies to B-CLL cell-surface antigens. Sera collected from B-CLL patients at defined times after alloHSCT showed selective binding to primary B-CLL cells. Pre-alloHSCT sera, donor sera, and control sera were negative. To identify post-alloHSCT serum antibodies and subsequently B-CLL cell-surface antigens they recognize, we generated a human antibody-binding fragment (Fab) library from post-alloHSCT peripheral blood mononuclear cells and selected it on primary B-CLL cells by phage display. A panel of Fab with B-CLL cell-surface reactivity was strongly enriched. Selection was dominated by highly homologous Fab predicted to bind the same antigen. One Fab was converted to immunoglobulin G1 and analyzed for reactivity with peripheral blood mononuclear cells from B-CLL patients and healthy volunteers. Cell-surface antigen expression was restricted to primary B cells and up-regulated in primary B-CLL cells. Mining post-alloHSCT antibody repertoires offers a novel route to discover fully human monoclonal antibodies and identify antigens of potential therapeutic relevance to B-CLL and possibly other cancers. Trials described herein were registered at www.clinicaltrials.gov as nos. NCT00055744 and NCT00003838.

  11. Improving serodiagnosis of human and canine leishmaniasis with recombinant Leishmania braziliensis cathepsin l-like protein and a synthetic peptide containing its linear B-cell epitope.

    Directory of Open Access Journals (Sweden)

    Daniel Menezes-Souza

    2015-01-01

    Full Text Available The early and correct diagnosis of human leishmaniasis is essential for disease treatment. Another important step in the control of visceral leishmaniasis is the identification of infected dogs, which are the main domestic reservoir of L. infantum. Recombinant proteins and synthetic peptides based on Leishmania genes have emerged as valuable targets for serodiagnosis due to their increased sensitivity, specificity and potential for standardization. Cathepsin L-like genes are surface antigens that are secreted by amastigotes and have little similarity to host proteins, factors that enable this protein as a good target for serodiagnosis of the leishmaniasis.We mapped a linear B-cell epitope within the Cathepsin L-like protein from L. braziliensis. A synthetic peptide containing the epitope and the recombinant protein was evaluated for serodiagnosis of human tegumentary and visceral leishmaniasis, as well as canine visceral leishmaniasis.The recombinant protein performed best for human tegumentary and canine visceral leishmaniasis, with 96.30% and 89.33% accuracy, respectively. The synthetic peptide was the best to discriminate human visceral leishmaniasis, with 97.14% specificity, 94.55% sensitivity and 96.00% accuracy. Comparison with T. cruzi-infected humans and dogs suggests that the identified epitope is specific to Leishmania parasites, which minimizes the likelihood of cross-reactions.

  12. Immunization of rabbits with highly purified, soluble, trimeric human immunodeficiency virus type 1 envelope glycoprotein induces a vigorous B cell response and broadly cross-reactive neutralization.

    Directory of Open Access Journals (Sweden)

    Gerald V Quinnan

    Full Text Available Previously we described induction of cross-reactive HIV-1 neutralizing antibody responses in rabbits using a soluble HIV-1 gp140 envelope glycoprotein (Env in an adjuvant containing monophosphoryl lipid A (MPL and QS21 (AS02A. Here, we compared different forms of the same HIV-1 strain R2 Env for antigenic and biophysical characteristics, and in rabbits characterized the extent of B cell induction for specific antibody expression and secretion and neutralizing responses. The forms of this Env that were produced in and purified from stably transformed 293T cells included a primarily dimeric gp140, a trimeric gp140 appended to a GCN4 trimerization domain (gp140-GCN4, gp140-GCN4 with a 15 amino acid flexible linker between the gp120 and gp41 ectodomain (gp140-GCN4-L, also trimeric, and a gp140 with the flexible linker purified from cell culture supernatants as either dimer (gp140-L(D or monomer (gp140-L(M. Multimeric states of the Env proteins were assessed by native gel electrophoresis and analytical ultracentrifugation. The different forms of gp140 bound broadly cross-reactive neutralizing (BCN human monoclonal antibodies (mAbs similarly in ELISA and immunoprecipitation assays. All Envs bound CD4i mAbs in the presence and absence of sCD4, as reported for the R2 Env. Weak neutralization of some strains of HIV-1 was seen after two additional doses in AS02A. Rabbits that were given a seventh dose of gp140-GCN4-L developed BCN responses that were weak to moderate, similar to our previous report. The specificity of these responses did not appear similar to that of any of the known BCN human mAbs. Induction of spleen B cell and plasma cells producing immunoglobulins that bound trimeric gp140-GCN4-L was vigorous, based on ELISpot and flow cytometry analyses. The results demonstrate that highly purified gp140-GCN4-L trimer in adjuvant elicits BCN responses in rabbits accompanied by vigorous B cell induction.

  13. B cells promote obesity-associated periodontitis and oral pathogen-associated inflammation.

    Science.gov (United States)

    Zhu, Min; Belkina, Anna C; DeFuria, Jason; Carr, Jordan D; Van Dyke, Thomas E; Gyurko, Robert; Nikolajczyk, Barbara S

    2014-08-01

    Individuals with T2D and PD suffer significantly from the ability of one disease to intensify the other. Disease-associated inflammation is one mechanism thought to fuel this pathogenic feed-forward loop. Several lines of evidence indicate that proinflammatory B cells promote T2D and PD; thus, B cells are top candidates for a cell type that predisposes PD in T2D. To test directly the role of B cells in T2D-associated PD, we compared outcomes from oral Porphyromonas gingivalis challenge of lean WT or B cell-null mice with outcomes from mice that were obese and insulin-resistant before challenge. Obese WT mice responded to oral P. gingivalis challenge with significant periodontal bone loss, whereas obese B cell-null mice were protected completely from PD. By contrast, lean WT and B cell-null mice suffer similar periodontal bone loss in response to oral pathogen. B cells from obese/insulin-resistant hosts also support oral osteoclastogenesis and both oral and systemic production of inflammatory cytokines, including pro-osteoclastogenic TNF-α and MIP-2, an ortholog of human IL-8. B cells furthermore impact AT inflammation in obese, P. gingivalis-infected hosts. Taken together, these data show that fundamentally different mechanisms regulate PD in lean and obese hosts, with B cells able to promote PD only if the hosts are "primed" by obesity. These results justify more intense analysis of obesity-associated changes in B cells that predispose PD in human T2D. © 2014 Society for Leukocyte Biology.

  14. Immunoreactivity, stability, pharmacokinetics and biodistribution of a monoclonal antibody to human leukemic B cells after three different methods of radioiodination

    International Nuclear Information System (INIS)

    Zhenping Zhu; Ghose, T.; Kralovec, Y.; Chunzheng Yang

    1994-01-01

    Dal B02, a murine monoclonal antibody against human chronic lymphocytic leukemia (CLL) was radioiodinated using chloramine T (Chl.T), Bolton-Hunter (B-H) or N-succinimidyl-p-iodobenzoate (PIB). The preparations had comparable radiochemical purity (>97%) and immunoreactive fraction (65-80%) but the Chl.T-based product was most susceptible to deiodination and loss of immunoreactivity. After i.v. injection into CLL-xenografted nude mice, the preparations had identical patterns of clearance from the blood but the PIB-based product led to more radioactivity in liver and spleen and less in the thyroid compared to the other preparations. The Chl.T-based product showed loss of immunoreactivity in circulation and less tumor-localized radioactivity 168 h after administration. The differences between the B-H-based and PIB-based products were less impressive than between PIB-based and Chl.T-based products. (author)

  15. Antigen-specific human NKT cells from tuberculosis patients produce IL-21 to help B cells for the production of immunoglobulins.

    Science.gov (United States)

    Wu, Changyou; Li, Zitao; Fu, Xiaoying; Yu, Sifei; Lao, Suihua; Yang, Binyan

    2015-10-06

    Natural killer T (NKT) cells from mouse and human play an important role in the immune responses against Mycobacterium tuberculosis. However, the function of CD3(+)TCRvβ11(+) NKT cells at the local site of M. tuberculosis infection remains poorly defined. In the present study, we found that after stimulation with M. tuberculosis antigens, NKT cells isolated from tuberculosis (TB) pleural fluid mononuclear cells (PFMCs) produced IL-21 and other cytokines including IFN-γ, TNF-α, IL-2 and IL-17. IL-21-expressing NKT cells in PFMCs displayed effector memory phenotype, expressing CD45RO(high)CD62L(low)CCR7(low). Moreover, NKT cells expressed high levels of CXCR5 and all of IL-21-expressing NKT cells co-expressed CXCR5. The frequency of BCL-6-expression was higher in IL-21-expressing but not in non-IL-21-expressing CD3(+)TCRvβ11(+) NKT cells. Sorted CD3(+)TCRvβ11(+) NKT cells from PFMCs produced IFN-γ and IL-21 after stimulation, which expressed CD40L. Importantly, CD3(+)TCRvβ11(+) NKT cells provided help to B cells for the production of IgG and IgA. Taken together, our data demonstrate that CD3(+)TCRvβ11(+) NKT cells from a local site of M. tuberculosis infection produce IL-21, express CXCR5 and CD40L, help B cells to secrete IgG and IgA, and may participate in local immune responses against M. tuberculosis infection.

  16. Synthesis of interleukin 6 (interferon-β2/B cell stimulatory factor 2) in human fibroblasts is triggered by an increase in intracellular cyclic AMP

    International Nuclear Information System (INIS)

    Zhange, Y.; Lin, J.X.; Vilcek, J.

    1988-01-01

    Interleukin 6 (IL-6; also referred to as interferon-β 2 , 26-kDa protein, and B cell stimulatory factor 2) is a cytokine whose actions include a stimulation of immunoglobulin synthesis, enhancement of B cell growth, and modulation of acute phase protein synthesis by hepatocytes. Synthesis of IL-6 is stimulated by interleukin 1 (IL-1), tumor necrosis factor (TNF), or platelet-derived growth factor. The authors examined the role of the cyclic AMP (cAMP)-dependent signal transduction pathway in IL-6 gene expression. Several activators of adenylate cyclase, including prostaglandin E1, forskolin, and cholera toxin, as well as the phosphodiesterase inhibitor isobutylmethylxanthine and the cAMP analog dibutyryl cAMP, shared the ability to cause a dramatic and sustained increase in IL-6 mRNA levels in human FS-4 fibroblasts. Actinomycin D treatment abolished this enhancement. Treatments that increased intracellular cAMP also stimulated the secretion of the IL-6 protein in a biologically active form. Increased intracellular cAMP appears to enhance IL-6 gene expression by a protein kinase C-independent mechanism because down-regulation of protein kinase C by a chronic exposure of cells to a high dose of 12-O-tetradecanoylphorbol 13-acetate did not abolish the enhancement of IL-6 expression by treatments that increase cAMP. IL-1 and TNF too increased IL-6 mRNA levels by a protein kinase C-independent mechanism. The results suggest a role for the cAMP-dependent pathway(s) in IL-6 gene activation by TNF and IL-1

  17. Bruton's tyrosine kinase mediates the synergistic signalling between TLR9 and the B cell receptor by regulating calcium and calmodulin.

    Directory of Open Access Journals (Sweden)

    Elaine F Kenny

    Full Text Available B cells signal through both the B cell receptor (BCR which binds antigens and Toll-like receptors (TLRs including TLR9 which recognises CpG DNA. Activation of TLR9 synergises with BCR signalling when the BCR and TLR9 co-localise within an auto-phagosome-like compartment. Here we report that Bruton's tyrosine kinase (BTK is required for synergistic IL6 production and up-regulation of surface expression of MHC-class-II, CD69 and CD86 in primary murine and human B cells. We show that BTK is essential for co-localisation of the BCR and TLR9 within a potential auto-phagosome-like compartment in the Namalwa human B cell line. Downstream of BTK we find that calcium acting via calmodulin is required for this process. These data provide new insights into the role of BTK, an important target for autoimmune diseases, in B cell activation.

  18. Ectopic expression of AID in a non-B cell line triggers A:T and G:C point mutations in non-replicating episomal vectors.

    Directory of Open Access Journals (Sweden)

    Tihana Jovanic

    Full Text Available Somatic hypermutation (SHM of immunoglobulin genes is currently viewed as a two step process initiated by the deamination of deoxycytidine (C to deoxyuridine (U, catalysed by the activation induced deaminase (AID. Phase 1 mutations arise from DNA replication across the uracil residue or the abasic site, generated by the uracil-DNA glycosylase, yielding transitions or transversions at G:C pairs. Phase 2 mutations result from the recognition of the U:G mismatch by the Msh2/Msh6 complex (MutS Homologue, followed by the excision of the mismatched nucleotide and the repair, by the low fidelity DNA polymerase eta, of the gap generated by the exonuclease I. These mutations are mainly focused at A:T pairs. Whereas in activated B cells both G:C and A:T pairs are equally targeted, ectopic expression of AID was shown to trigger only G:C mutations on a stably integrated reporter gene. Here we show that when using non-replicative episomal vectors containing a GFP gene, inactivated by the introduction of stop codons at various positions, a high level of EGFP positive cells was obtained after transient expression in Jurkat cells constitutively expressing AID. We show that mutations at G:C and A:T pairs are produced. EGFP positive cells are obtained in the absence of vector replication demonstrating that the mutations are dependent only on the mismatch repair (MMR pathway. This implies that the generation of phase 1 mutations is not a prerequisite for the expression of phase 2 mutations.

  19. B Cell Receptor-Mediated Internalization of Salmonella: A Novel Pathway for Autonomous B Cell Activation and Antibody Production

    NARCIS (Netherlands)

    Souwer, Yuri; Griekspoor, Alexander; Jorritsma, Tineke; de Wit, Jelle; Janssen, Hans; Neefjes, Jacques; van Ham, S. Marieke

    2009-01-01

    The present paradigm is that primary B cells are nonphagocytosing cells. In this study, we demonstrate that human primary B cells are able to internalize bacteria when the bacteria are recognized by the BCR. BCR-mediated internalization of Salmonella typhimurium results in B cell differentiation and

  20. Expression and function of β-adrenergic receptors in human hematopoietic cell lines

    International Nuclear Information System (INIS)

    Maeki, T.; Andersson, L.C.; Kontula, K.K.

    1992-01-01

    We investigated the expression and functional characteristics of β-adrenoceptors in a panel of 10 phenotypically different human hematopoietic cell lines. A binding assay with [ 125 I]iodocyanopindolol as the ligand revealed that cell lines of myelomonocytic or histiocytic derivation (HL-60, ML-2, RC-2A, U-937) expressed high numbers of β-adrenoceptors. An intermediate density of receptors was found in a non-T, non-B cell leukemia line (Nall-1), whereas T-cell (JM, CCRF-CEM), B-cell (Raji) or erythroleukemic cell lines (K-562, HEL) displayed minimala or undetectable binding of the radioligand. Isoprenaline-stimulated cAMP production by the cells correlated to their extent of β-adrenoceptor expression. Southern blot hybridization analysis of genomic DNA from the cell lines with a 32 P-labelled β 2 -adrenoceptor cDNA probe revealed no evidence for major rearrangement or amplification of the receptor gene. Incubation with isoprenaline in vitro suppressed the proliferation of the receptor-rich RC-2A cells but did not affect the growth rate of the receptor-deficient K-562 cells. Treatment with propranolol slightly enhanced the proliferation of the RC-2A cells but did not markedly alter the growth rate of two other cell lines, regardless of their β-adrenoceptor status. These findings indicate a regulatory influence by the sympathoadrenergic system on selected cells of the myelomonocytic lineage. (au)

  1. Epstein-Barr virus (EBV) provides survival factors to EBV+ diffuse large B-cell lymphoma (DLBCL) lines and modulates cytokine induced specific chemotaxis in EBV+  DLBCL.

    Science.gov (United States)

    Wu, Liang; Ehlin-Henriksson, Barbro; Zhou, Xiaoying; Zhu, Hong; Ernberg, Ingemar; Kis, Lorand L; Klein, George

    2017-12-01

    Diffuse large B-cell lymphoma (DLBCL), the most common type of malignant lymphoma, accounts for 30% of adult non-Hodgkin lymphomas. Epstein-Barr virus (EBV) -positive DLBCL of the elderly is a newly recognized subtype that accounts for 8-10% of DLBCLs in Asian countries, but is less common in Western populations. Five DLBCL-derived cell lines were employed to characterize patterns of EBV latent gene expression, as well as response to cytokines and chemotaxis. Interleukin-4 and interleukin-21 modified LMP1, EBNA1 and EBNA2 expression depending on cell phenotype and type of EBV latent programme (type I, II or III). These cytokines also affected CXCR4- or CCR7-mediated chemotaxis in two of the cell lines, Farage (type III) and Val (type II). Further, we investigated the effect of EBV by using dominant-negative EBV nuclear antigen 1(dnEBNA1) to eliminate EBV genomes. This resulted in decreased chemotaxis. By employing an alternative way to eliminate EBV genomes, Roscovitine, we show an increase of apoptosis in the EBV-positive lines. These results show that EBV plays an important role in EBV-positive DLBCL lines with regard to survival and chemotactic response. Our findings provide evidence for the impact of microenvironment on EBV-carrying DLBCL cells and might have therapeutic implications. © 2017 John Wiley & Sons Ltd.

  2. Piper betle leaf extracts induced human hepatocellular carcinoma Hep3B cell death via MAPKs regulating the p73 pathway in vitro and in vivo.

    Science.gov (United States)

    Wu, Pei-Fang; Tseng, Hsien-Chun; Chyau, Charng-Cherng; Chen, Jing-Hsien; Chou, Fen-Pi

    2014-12-01

    Extracts of Piper betle leaf (PBLs) are rich in bioactive compounds with potential chemopreventive ability. In this study, Hep3B cells which are p53 null were used to investigate the anti-tumor effect of PBLs in the cell and in the xenograft model. The results revealed that PBLs (0.1 to 1 mg mL(-1)) induced a dose- and time-dependent increase of cell toxicity. The underlying mechanisms as evidenced by flow cytometry and western blot analysis showed that PBLs triggered ATM, cAbl, and p73 expressions and activated JNK and p38 pathways that subsequently led to cell cycle arrest and mitochondria-dependent apoptosis. PBLs also inhibited tumor growth in Hep3B-bearing mice via inducing the MAPK-p73 pathway. Our results demonstrated the in vitro and in vivo anti-tumor potential of PBLs, supporting their application as a novel chemopreventive agent for the treatment of human hepatocellular carcinoma (HCC) in the future via targeting the p73 pathway.

  3. Fully human broadly neutralizing monoclonal antibodies against influenza A viruses generated from the memory B cells of a 2009 pandemic H1N1 influenza vaccine recipient

    Energy Technology Data Exchange (ETDEWEB)

    Hu, Weibin [Molecular Virus Unit, Key Laboratory of Molecular Virology and Immunology, Institut Pasteur of Shanghai, Chinese Academy of Sciences, Shanghai 200025 (China); Chen, Aizhong [Key Laboratory of Molecular Cell Biology, Institute of Biochemistry and Cell Biology, Shanghai Institutes for Biological Sciences, Chinese Academy of Sciences, Shanghai 200031 (China); Miao, Yi [Shanghai Xuhui Central Hospital, Shanghai 200031 (China); Xia, Shengli [Center for Disease Control and Prevention of Henan Province, Zhengzhou 450016 (China); Ling, Zhiyang; Xu, Ke; Wang, Tongyan [Molecular Virus Unit, Key Laboratory of Molecular Virology and Immunology, Institut Pasteur of Shanghai, Chinese Academy of Sciences, Shanghai 200025 (China); Xu, Ying; Cui, Jun; Wu, Hongqiang; Hu, Guiyu; Tian, Lin; Wang, Lingling [Key Laboratory of Molecular Cell Biology, Institute of Biochemistry and Cell Biology, Shanghai Institutes for Biological Sciences, Chinese Academy of Sciences, Shanghai 200031 (China); Shu, Yuelong [Chinese Center for Disease Control and Prevention, Beijing 102206 (China); Ma, Xiaowei [Hualan Biological Bacterin Company, Xinxiang 453003 (China); Xu, Bianli; Zhang, Jin [Center for Disease Control and Prevention of Henan Province, Zhengzhou 450016 (China); Lin, Xiaojun, E-mail: linxiaojun@hualan.com [Hualan Biological Bacterin Company, Xinxiang 453003 (China); Bian, Chao, E-mail: cbian@sibs.ac.cn [Key Laboratory of Molecular Cell Biology, Institute of Biochemistry and Cell Biology, Shanghai Institutes for Biological Sciences, Chinese Academy of Sciences, Shanghai 200031 (China); Sun, Bing, E-mail: bsun@sibs.ac.cn [Molecular Virus Unit, Key Laboratory of Molecular Virology and Immunology, Institut Pasteur of Shanghai, Chinese Academy of Sciences, Shanghai 200025 (China); Key Laboratory of Molecular Cell Biology, Institute of Biochemistry and Cell Biology, Shanghai Institutes for Biological Sciences, Chinese Academy of Sciences, Shanghai 200031 (China)

    2013-01-20

    Whether the 2009 pandemic H1N1 influenza vaccine can induce heterosubtypic cross-protective anti-hemagglutinin (HA) neutralizing antibodies is an important issue. We obtained a panel of fully human monoclonal antibodies from the memory B cells of a 2009 pandemic H1N1 influenza vaccine recipient. Most of the monoclonal antibodies targeted the HA protein but not the HA1 fragment. Among the analyzed antibodies, seven mAbs exhibited neutralizing activity against several influenza A viruses of different subtypes. The conserved linear epitope targeted by the neutralizing mAbs (FIEGGWTGMVDGWYGYHH) is part of the fusion peptide on HA2. Our work suggests that a heterosubtypic neutralizing antibody response primarily targeting the HA stem region exists in recipients of the 2009 pandemic H1N1 influenza vaccine. The HA stem region contains various conserved neutralizing epitopes with the fusion peptide as an important one. This work may aid in the design of a universal influenza A virus vaccine.

  4. Fully human broadly neutralizing monoclonal antibodies against influenza A viruses generated from the memory B cells of a 2009 pandemic H1N1 influenza vaccine recipient

    International Nuclear Information System (INIS)

    Hu, Weibin; Chen, Aizhong; Miao, Yi; Xia, Shengli; Ling, Zhiyang; Xu, Ke; Wang, Tongyan; Xu, Ying; Cui, Jun; Wu, Hongqiang; Hu, Guiyu; Tian, Lin; Wang, Lingling; Shu, Yuelong; Ma, Xiaowei; Xu, Bianli; Zhang, Jin; Lin, Xiaojun; Bian, Chao; Sun, Bing

    2013-01-01

    Whether the 2009 pandemic H1N1 influenza vaccine can induce heterosubtypic cross-protective anti-hemagglutinin (HA) neutralizing antibodies is an important issue. We obtained a panel of fully human monoclonal antibodies from the memory B cells of a 2009 pandemic H1N1 influenza vaccine recipient. Most of the monoclonal antibodies targeted the HA protein but not the HA1 fragment. Among the analyzed antibodies, seven mAbs exhibited neutralizing activity against several influenza A viruses of different subtypes. The conserved linear epitope targeted by the neutralizing mAbs (FIEGGWTGMVDGWYGYHH) is part of the fusion peptide on HA2. Our work suggests that a heterosubtypic neutralizing antibody response primarily targeting the HA stem region exists in recipients of the 2009 pandemic H1N1 influenza vaccine. The HA stem region contains various conserved neutralizing epitopes with the fusion peptide as an important one. This work may aid in the design of a universal influenza A virus vaccine.

  5. The experimental study on biodistribution and radioimmunoimaging of 131I labeled anti-lymphoma Fab antibody in nude mice bearing human B cell lymphoma

    International Nuclear Information System (INIS)

    Yang Xiaochun; Zhang Meihua; Shen Junkang; Shen Yongmei; Shi Yizhen; Liu Zengli

    2008-01-01

    Objective: Radioimmunoimaging is still an interesting study in the domain of nuclear medicine. The aim of this study was to evaluate the biodistribution and radioimmunoimaging of 131 I-Fab anti- body in nude mice beating human B cell lymphoma. Methods: The immunoreactivity of Fab antibody to Raji cells was analyzed by immunohistochemistry and flow cytometry. Fab antibody and CD20 monoclonal antibody (as control) were labeled with 131 I using Iodogen method. 131 I-Fab antibody or 131 I-CD20 was injected into nude mice bearing B cell lymphoma via tail veins. The biodistribution and radioimmunoimaging results were obtained at 2, 4, 8 and 24 h postinjection, respectively. Results: The results of immunohistochemistry and flow cytometry indicated that both Fab antibody and 131 I-Fab antibody could bind strongly with membrane antigens on Raji cells, and the binding rate reached above 87%. Clear tumor image was obtained at 8 h after injection with 131 I-Fab and elimination was observed at 24 h postinjection. The clear tumor image for 131 I-CD20 antibody was obtained at 24 h post injection. The biodistribution in vivo showed that the percentage activities of injection dose per gram of tumor (% ID/g) of 131 I-Fab group at 2, 4, 8 h postinjection were higher than that of 131 I-CD20 antibody [(1.37±0.28), (1.84±0.13), (2.21±0.15)% ID/g vs (0.33±0.06), (0.62±0.08), (1.46±0.24)% ID/g, respectively; F=52.22, 278.42 and 29.00, all P 131 I-Fad and 131 I-CD20 groups at 2, 4, 8 and 24 h were [(0.22±0.03)-(5.44± 0.31)] vs[(0.04±0.01)-(3.10±0.29)], [(0.43±0.11) - (21.01±3.97)] vs [(0.11±0.05) - (7.99±1.81)], [(1.09±0.07) -(20.28±2.77)] vs [(0.48±0.06) - (23.55±1.69)], [(1.12± 0.02) - (10.29±1.78)] vs [(2.32 ± 0.34) - (33.23±6.83)], respectively. Conclusion: 131 I-Fab anti- body has advantages of small molecular weight, excellent targeting characteristics, early imaging and fast elimination, which indicates the potential application value in diagnosing B cell

  6. Changes in Tonsil B Cell Phenotypes and EBV Receptor Expression in Children Under 5-Years-Old.

    Science.gov (United States)

    Wohlford, Eric M; Baresel, Paul C; Wilmore, Joel R; Mortelliti, Anthony J; Coleman, Carrie B; Rochford, Rosemary

    2018-03-01

    Palatine tonsils are principally B cell organs that are the initial line of defense against many oral pathogens, as well as the site of infection for others. While the size of palatine tonsils changes greatly in the first five years of life, the cellular changes during this period are not well studied. Epstein Barr virus (EBV) is a common orally transmitted virus that infects tonsillar B cells. Naïve B cells are thought to be the target of primary infection with EBV in vivo, suggesting that they are targeted by the virus. EBV enters B cells through CD21, but studies of older children and adults have not shown differences in surface CD21 between naïve B cells and other tonsil B cell populations. In this study, we used an 11-color flow cytometry panel to detail the changes in B cell subpopulations in human tonsils over the first five years of life from 33 healthy US children. We provide reference ranges for tonsil B cell subpopulations over this age range. We show that the frequency of naïve tonsil B cells decreases over the early years of life, and that naïve B cells expressed higher surface levels of CD21 relative to other tonsil B cell populations. We show that young children have a higher frequency of naïve tonsil B cells, and importantly that these cells express increased surface EBV receptor, suggesting that young children have a larger pool of cells that can be infected by the virus. © 2017 International Clinical Cytometry Society. © 2017 International Clinical Cytometry Society.

  7. Subclass of individual IgA-secreting human lymphocytes. Investigation of in vivo pneumococcal polysaccharide-induced and in vitro mitogen-induced blood B cells by monolayer plaque-forming cell assays

    DEFF Research Database (Denmark)

    Heilmann, C; Barington, T; Sigsgaard, T

    1988-01-01

    The subclass of individual human IgA B cells was investigated by means of monolayer plaque-forming cell assays permitting analysis of all IgA-secreting cells as well as of cells secreting IgA anti-pneumococcal polysaccharide antibody. Center cells were examined by indirect immunofluorescence...

  8. Membranes of activated CD4+ T cells expressing T cell receptor (TcR) alpha beta or TcR gamma delta induce IgE synthesis by human B cells in the presence of interleukin-4

    NARCIS (Netherlands)

    Gascan, H.; Aversa, G. G.; Gauchat, J. F.; van Vlasselaer, P.; Roncarolo, M. G.; Yssel, H.; Kehry, M.; Spits, H.; de Vries, J. E.

    1992-01-01

    In the present study it is demonstrated that human B cells can be induced to switch to IgE production following a contact-mediated signal provided by activated T cell receptor (TcR) gamma delta+, CD4+ and TcR alpha beta+, CD4+ T cell clones and interleukin (IL)-4. The signal provided by these T cell

  9. CHLORAMBUCIL PLUS RITUXIMAB AS FRONT-LINE THERAPY IN ELDERLY/UNFIT PATIENTS AFFECTED BY B-CELL CHRONIC LYMPHOCYTIC LEUKEMIA: RESULTS OF A SINGLE-CENTRE EXPERIENCE.

    Directory of Open Access Journals (Sweden)

    Luca Laurenti

    2013-05-01

    Full Text Available Currently standard first line therapy for fit patients with B-CLL/SLL are fludarabine-based regimens. Elderly patients or patients with comorbidities poorly tolerate purine analogue-based chemotherapy and they are often treated with Chlorambucil (Chl. However, complete response (CR and overall response (OR rates with Chl are relatively low. We now investigated whether the addition of Rituximab to Chl will improve the efficacy without impairing the tolerability in elderly and unfit patients. We included in our study 27 elderly or unfit patients that had not received prior therapy. All patients were treated with Chl (1mg/Kg per 28-day cycle for 8 cycles plus Rituximab (375 mg/m2 for the first course and 500 mg/m2 for subsequent cycles until the 6th cycle. We obtained an OR rate of 74%. The most frequent adverse effect was grade 3-4 neutropenia, which occurred in 18.5% of the patients. Infections or grade 3-4 extra-hematological side effects were not recorded. None of the patients required reduction of dose, delay of therapy or hospitalization. Overall, these data suggest that Chl-R is an effective and well tolerated regimen in elderly/unfit patients with CLL.

  10. Cell of origin associated classification of B-cell malignancies by gene signatures of the normal B-cell hierarchy.

    Science.gov (United States)

    Johnsen, Hans Erik; Bergkvist, Kim Steve; Schmitz, Alexander; Kjeldsen, Malene Krag; Hansen, Steen Møller; Gaihede, Michael; Nørgaard, Martin Agge; Bæch, John; Grønholdt, Marie-Louise; Jensen, Frank Svendsen; Johansen, Preben; Bødker, Julie Støve; Bøgsted, Martin; Dybkær, Karen

    2014-06-01

    Recent findings have suggested biological classification of B-cell malignancies as exemplified by the "activated B-cell-like" (ABC), the "germinal-center B-cell-like" (GCB) and primary mediastinal B-cell lymphoma (PMBL) subtypes of diffuse large B-cell lymphoma and "recurrent translocation and cyclin D" (TC) classification of multiple myeloma. Biological classification of B-cell derived cancers may be refined by a direct and systematic strategy where identification and characterization of normal B-cell differentiation subsets are used to define the cancer cell of origin phenotype. Here we propose a strategy combining multiparametric flow cytometry, global gene expression profiling and biostatistical modeling to generate B-cell subset specific gene signatures from sorted normal human immature, naive, germinal centrocytes and centroblasts, post-germinal memory B-cells, plasmablasts and plasma cells from available lymphoid tissues including lymph nodes, tonsils, thymus, peripheral blood and bone marrow. This strategy will provide an accurate image of the stage of differentiation, which prospectively can be used to classify any B-cell malignancy and eventually purify tumor cells. This report briefly describes the current models of the normal B-cell subset differentiation in multiple tissues and the pathogenesis of malignancies originating from the normal germinal B-cell hierarchy.

  11. 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)

    Brines, R.D.; Sia, D.Y.; Lehner, T.

    1987-01-01

    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

  12. MUTATION ON WD DIPEPTIDE MOTIFS OF THE p48 SUBUNIT OF CHROMATIN ASSEMBLY FACTOR-1 CAUSING VIABILITY AND GROWTH OF DT40 CHICKEN B CELL LINE

    Directory of Open Access Journals (Sweden)

    Ahyar Ahmad

    2010-07-01

    Full Text Available Chromatin assembly factor-1 (CAF-1, a protein complex consisting of three subunits, p150, p60, and p48, is highly conserved from yeast to humans and facilitated nucleosome assembly of newly replicated DNA. The p48 subunit, CAF-1p48 (p48, with seven WD (Trp-Asp repeat motifs, is a member of the WD protein family. The immunoprecipitation experiment revealed that ß-propeller structure of p48 was less stringent for it's binding to HDAC-1, but more stringent for its binding to both histones H4 and CAF-1p60 but not to ASF-1, indicating that the proper ß-propeller structure of p48 is essential for the binding to these two proteins histone H4 and CAF-1p60. Complementation experiments, involving missense and truncated mutants of FLAG-tagged p48, revealed that mutations of every of seven WD dipeptide motifs, like both the N-terminal and C-terminal truncated mutations, could not rescue for the tet-induced lethality. These results indicate not only that p48 is essential for the viability of vertebrate cells, although the yeast p48 homolog is nonessential, but also that all the seven WD dipeptide motifs are necessary for the maintenance of the proper structure of p48 that is fundamentally important for cell viability.   Keywords: Chromatin assembly factor-1, complementation experiments, viability

  13. Cotton Leaf Curl Multan Betasatellite DNA as a Tool to Deliver and Express the Human B-Cell Lymphoma 2 (Bcl-2) Gene in Plants.

    Science.gov (United States)

    Kharazmi, Sara; Ataie Kachoie, Elham; Behjatnia, Seyed Ali Akbar

    2016-05-01

    The betasatellite DNA associated with Cotton leaf curl Multan virus (CLCuMB) contains a single complementary-sense ORF, βC1, which is a pathogenicity determinant. CLCuMB was able to replicate in plants in the presence of diverse helper geminiviruses, including Tomato leaf curl virus-Australia (TLCV-Au), Iranian isolate of Tomato yellow leaf curl virus (TYLCV-[Ab]), and Beet curly top virus (BCTV-Svr), and can be used as a plant gene delivery vector. To test the hypothesis that CLCuMB has the potential to act as an animal gene delivery vector, a specific insertion construct was produced by the introduction of a human B-cell lymphoma 2 (Bcl-2) cDNA into a mutant DNA of CLCuMB in which the βC1 was deleted (β∆C1). The recombinant βΔC1-Bcl-2 construct was successfully replicated in tomato and tobacco plants in the presence of TLCV-Au, BCTV-Svr and TYLCV-[Ab]. Real-time PCR and Western blot analyses of plants containing the replicative forms of recombinant βΔC1-Bcl-2 DNA showed that Bcl-2 gene was expressed in an acceptable level in these plants, indicating that β∆C1 can be used as a tool to deliver and express animal genes in plants. This CLCuMB-based system, having its own promoter activity, offers the possibility of production of animal recombinant proteins in plants.

  14. DNA damage and DNA damage response in human bronchial epithelial BEAS-2B cells following exposure to 2-nitrobenzanthrone and 3-nitrobenzanthrone: role in apoptosis.

    Science.gov (United States)

    Oya, Elisabeth; Ovrevik, Johan; Arlt, Volker M; Nagy, Eszter; Phillips, David H; Holme, Jørn A

    2011-11-01

    Nitro-polycyclic aromatic hydrocarbons (nitro-PAHs) are mutagenic and carcinogenic environmental pollutants found in diesel exhaust and on urban air pollution particles. In the present study, human bronchial epithelial BEAS-2B cells were exposed to 2-nitrobenzanthrone (2-NBA) and 3-nitrobenzanthrone (3-NBA). DNA damage responses were compared to those observed after exposure to 1-nitropyrene (1-NP) and benzo[a]pyrene (B[a]P). Examination by microscopy revealed that 3-NBA was the most potent toxic compound while weaker responses were observed with 1-NP and B[a]P. Most interestingly, 2-NBA did not induce cell death or any other stress-related responses. 3-NBA induced a typical apoptotic cell death judged by nuclear condensation and little plasma membrane damage as well as cleavage of caspase 3 and poly-(ADP-ribose) polymerase (PARP). Exposure to 3-NBA resulted in an accumulation of cells in S-phase, and further analysis by Western blotting, immunocytochemistry and flow cytometry revealed that 3-NBA induced a DNA damage response characterized by phosphorylation of ATM (ataxia-telangiectasia mutated), checkpoint kinase (Chk) 2/Chk1, H2AX and p53. The p53 inhibitor pifithrin-α inhibited 3-NBA-induced apoptosis while small effects were seen using pifithrin-μ, suggesting that 3-NBA-induced cell death is a result of transcriptional activation of p53. In conclusion, 3-NBA is a potent inducer of apoptosis, which seemed to be triggered by the DNA damage response. Furthermore, a change of the nitro-group to the second position (i.e. 2-NBA) dramatically changed the cellular reactivity of the compound.

  15. 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

    The FOXP1 (forkhead box P1) transcription factor is a marker of poor prognosis in diffuse large B-cell lymphoma (DLBCL). Here microarray analysis of FOXP1-silenced DLBCL cell lines identified differential regulation of immune response signatures and major histocompatibility complex class II (MHC II......) genes as some of the most significant differences between germinal center B-cell (GCB)-like DLBCL with full-length FOXP1 protein expression versus activated B-cell (ABC)-like DLBCL expressing predominantly short FOXP1 isoforms. In an independent primary DLBCL microarray data set, multiple MHC II genes......, including human leukocyte antigen DR alpha chain (HLA-DRA), were inversely correlated with FOXP1 transcript expression (PABC-DLBCL cells led to increased cell-surface expression of HLA-DRA and CD74. In R-CHOP (rituximab, cyclophosphamide, doxorubicin, vincristine and prednisone...

  16. Human Infant Memory B Cell and CD4+ T Cell Responses to HibMenCY-TT Glyco-Conjugate Vaccine.

    Directory of Open Access Journals (Sweden)

    Angela Fuery

    Full Text Available Carrier-specific T cell and polysaccharide-specific B cell memory responses are not well characterised in infants following glyco-conjugate vaccination. We aimed to determine if the number of Meningococcal (Men C- and Y- specific memory B cells and; number and quality of Tetanus Toxoid (TT carrier-specific memory CD4+ T cells are associated with polysaccharide-specific IgG post HibMenCY-TT vaccination. Healthy infants received HibMenCY-TT vaccine at 2, 4 and 6 months with a booster at 12 months. Peripheral blood mononuclear cells were isolated and polysaccharide-specific memory B cells enumerated using ELISpot. TT-specific memory CD4+ T cells were detected and phenotyped based on CD154 expression and intracellular TNF-α, IL-2 and IFN-γ expression following stimulation. Functional polysaccharide-specific IgG titres were measured using the serum bactericidal activity (SBA assay. Polysaccharide-specific Men C- but not Men Y- specific memory B cell frequencies pre-boost (12 months were significantly associated with post-boost (13 months SBA titres. Regression analysis showed no association between memory B cell frequencies post-priming (at 6 or 7 months and SBA at 12 months or 13 months. TT-specific CD4+ T cells were detected at frequencies between 0.001 and 0.112 as a percentage of CD3+ T cells, but their numbers were not associated with SBA titres. There were significant negative associations between SBA titres at M13 and cytokine expression at M7 and M12.Induction of persistent polysaccharide-specific memory B cells prior to boosting is an important determinant of secondary IgG responses in infants. However, polysaccharide-specific functional IgG responses appear to be independent of the number and quality of circulating carrier-specific CD4+ T cells after priming.

  17. B cell biology: implications for treatment of systemic lupus erythematosus.

    Science.gov (United States)

    Anolik, J H

    2013-04-01

    utilization of B cell depletion therapy in autoimmune diseases and the need for new therapeutic approaches in SLE, a better understanding of human B cell subsets and the balance of pathogenic and regulatory functions is of the essence.

  18. Cellular cooperation in lymphocyte activation. III. B-cell helper effect in the enhancement of T-cell response.

    Science.gov (United States)

    Kasahara, T; Kin, K; Itoh, Y; Kawai, T; Kano, Y; Shioiri-Nakano, K

    1979-01-01

    T and B cells were purified from human tonsil and peripheral blood by the removal of phagocytic cells, followed by filtration through a nylon fiber column (NC) and E-rosette formation. Purified T and B cells contained less than 1% of other cell types. The responses of T cells to concanavalin A (Con A) and soluble protein A were greatly enhanced in the presence of autologous B cells. Participation of B cells in T-cell enhancement was confirmed by the following observations: (a) purified B copulation, which was separated further from adherent B cells, retained its enhancing activity. (b) Another adherent cell-free B-cell preparation, which was purified from the NC-passed fraction, and (c) no T lymphoid but some B lymphoid cell lines, elicited strong T-cell enhancement. It was also found that the enhancing capacity of B cells required no metabolic activity, but rather an intact cell form and direct cell-to-cell contact with responding cells. The stimulatory determinants on B cells were resistant to trypsin and neuraminidase treatment. In this paper a hypothesis will be presented that at least two signals are prerequisite for the effective activation of T cells.

  19. Continuous signaling of CD79b and CD19 is required for the fitness of Burkitt lymphoma B cells.

    Science.gov (United States)

    He, Xiaocui; Kläsener, Kathrin; Iype, Joseena M; Becker, Martin; Maity, Palash C; Cavallari, Marco; Nielsen, Peter J; Yang, Jianying; Reth, Michael

    2018-04-18

    Expression of the B-cell antigen receptor (BCR) is essential not only for the development but also for the maintenance of mature B cells. Similarly, many B-cell lymphomas, including Burkitt lymphoma (BL), require continuous BCR signaling for their tumor growth. This growth is driven by immunoreceptor tyrosine-based activation motif (ITAM) and PI3 kinase (PI3K) signaling. Here, we employ CRISPR/Cas9 to delete BCR and B-cell co-receptor genes in the human BL cell line Ramos. We find that Ramos B cells require the expression of the BCR signaling component Igβ (CD79b), and the co-receptor CD19, for their fitness and competitive growth in culture. Furthermore, we show that in the absence of any other BCR component, Igβ can be expressed on the B-cell surface, where it is found in close proximity to CD19 and signals in an ITAM-dependent manner. These data suggest that Igβ and CD19 are part of an alternative B-cell signaling module that use continuous ITAM/PI3K signaling to promote the survival of B lymphoma and normal B cells. © 2018 The Authors. Published under the terms of the CC BY 4.0 license.

  20. Generation of B-cell chronic lymphocytic leukemia (B-CLL)-reactive T-cell lines and clones from HLA class I-matched donors using modified B-CLL cells as stimulators: implications for adoptive immunotherapy.

    Science.gov (United States)

    Hoogendoorn, M; Wolbers, J Olde; Smit, W M; Schaafsma, M R; Barge, R M Y; Willemze, R; Falkenburg, J H F

    2004-07-01

    Allogeneic stem cell transplantation following reduced-intensity conditioning is being evaluated in patients with advanced B-cell chronic lymphocytic leukemia (B-CLL). The curative potential of this procedure is mediated by donor-derived alloreactive T cells, resulting in a graft-versus-leukemia effect. However, B-CLL may escape T-cell-mediated immune reactivity since these cells lack expression of costimulatory molecules. We examined the most optimal method to transform B-CLL cells into efficient antigen-presenting cells (APC) using activating cytokines, by triggering toll-like receptors (TLRs) using microbial pathogens and by CD40 stimulation with CD40L-transfected fibroblasts. CD40 activation in the presence of IL-4 induced strongest upregulation of costimulatory and adhesion molecules on B-CLL cells and induced the production of high amounts of IL-12 by the leukemic cells. In contrast to primary B-CLL cells as stimulator cells, these malignant APCs were capable of inducing the generation of B-CLL-reactive CD8(+) CTL lines and clones from HLA class I-matched donors. These CTL lines and clones recognized and killed primary B-CLL as well as patient-derived lymphoblasts, but not donor cells. These results show the feasibility of ex vivo generation of B-CLL-reactive CD8(+) CTLs. This opens new perspectives for adoptive immunotherapy, following allogeneic stem cell transplantation in patients with advanced B-CLL.

  1. Human Adipose Tissue-Derived Mesenchymal Stem Cells Abrogate Plasmablast Formation and Induce Regulatory B Cells Independently of T Helper Cells

    NARCIS (Netherlands)

    Franquesa, M.; Mensah, F. K.; Huizinga, R.; Strini, T.; Boon, L.; Lombardo, E.; DelaRosa, O.; Laman, J. D.; Grinyo, J. M.; Weimar, W.; Betjes, M. G. H.; Baan, C. C.; Hoogduijn, M. J.

    Mesenchymal or stromal stem cells (MSC) interact with cells of the immune system in multiple ways. Modulation of the immune system by MSC is believed to be a therapeutic option for autoimmune disease and transplant rejection. In recent years, B cells have moved into the focus of the attention as

  2. IL-6 Triggers IL-21 production by human CD4(+) T cells to drive STAT3-dependent plasma cell differentiation in B cells

    NARCIS (Netherlands)

    Diehl, Sean A.; Schmidlin, Heike; Nagasawa, Maho; Blom, Bianca; Spits, Hergen

    2012-01-01

    Interleukin (IL)-21-producing CD4(+) T cells are central to humoral immunity. Deciphering the signals that induce IL-21 production in CD4(+) T cells and those triggered by IL-21 in B cells are, therefore, of importance for understanding the generation of antibody (Ab) responses. Here, we show that

  3. Establishment, characterization, chemosensitivity, and radiosensitivity of two different cell lines derived from a human breast cancer biopsy

    International Nuclear Information System (INIS)

    Gioanni, J.; Courdi, A.; Lalanne, C.M.; Fischel, J.L.; Zanghellini, E.; Lambert, J.C.; Ettore, F.; Namer, M.

    1985-01-01

    In vitro culture of a human breast cancer biopsy fragment gave rise to two permanent cell lines, CAL 18 A and CAL 18 B, which were differentiated by both morphological and ultrastructural analysis. The karyotypic and growth properties of these two cell lines also differed, providing further evidence of cell heterogeneity within a given tumor. Both cell lines lost their hormone receptors in vitro. CAL 18 A cells grew in agar and were tumorigenic after inoculation into nude mice; neither of these properties was observed in CAL 18 B cells. The chemosensitivity of 12 antineoplastic drugs was assessed by a short-term assay, using inhibition of tritiated thymidine incorporation by the cells after contact with the drugs as the end point. Only a few drugs were active at moderate concentrations. The overall responses of both cell lines were similar. The cell survival curves, established by the colony method following a single dose of radiation, were also very similar, despite the greater heterogeneity of CAL 18 B cells. The two cell lines appear to be interrelated, since CAL 18 B cells were occasionally observed to emerge from CAL 18 A clones, suggesting that malignant cell redifferentiation may occur spontaneously in vitro

  4. The expression analysis of ICOS-L on activated T cells and immature dendritic cells as well as malignant B cells and Grave's-disease-derived thyroid tissues by two novel mAbs against human ICOS-L.

    Science.gov (United States)

    Wang, F; Zhu, W; Liu, T; Sun, Z; Ju, S; Ju, S; Yu, G; Xie, W; Deng, Z; Lu, B; Zhang, X

    2007-01-01

    ICOS-L, a newly identified member of B7 superfamily, plays an important role in immune responses. In this article, we report on two novel mouse anti-human ICOS-L monoclonal antibodies (mAbs) named as 11C4 and 12B11, whose specificities were verified by methods of flow cytometry, western blotting, and epitope competition assay. The two mAbs bound to distinct ICOS-L epitopes on B cells. Interestingly, mAb 11C4 could well recognize ICOS-L molecule on activated T cells and Jurkat cell lines, which is different from commercial anti-ICOS-L mAb (clone number MIH12) and the other mAb 12B11. In addition, we found that the expression of ICOS-L molecule was only detected on the surface of immature monocyte-derived dendritic cells (Mo-DCs) and was sharply decreased after induction of mature Mo-DCs activated by tumor necrosis factor-alpha or CD40. Furthermore, we showed that 11C4 could effectively suppress the maturation of Mo-DCs in vitro as evidenced by the low expression of CD80, CD86, CD83, and human leukocyte antigen-DR, which suggested that ICOS-L may be involved in the maturation of Mo-DCs. Using immunohistochemistry staining with mAb 11C4, the expression of ICOS-L was found in B lymphoma tissues and thyroid tissues from the Grave's disease but not in thyroid adenoma and normal thyroid tissues.

  5. Methodology of clinical trials evaluating the incorporation of new drugs in the first-line treatment of patients with diffuse large B-cell lymphoma (DLBCL): a critical review.

    Science.gov (United States)

    Iacoboni, G; Zucca, E; Ghielmini, M; Stathis, A

    2018-05-01

    The first-line treatment of diffuse large B-cell lymphoma (DLBCL) is the combination of rituximab with CHOP (cyclophosphamide, doxorubicin, vincristine, prednisone) chemotherapy, curing approximately 60% of patients. Many clinical trials have been carried out over the last 10 years trying to improve the results of this treatment, but the appropriateness of their planning strategies could be rediscussed. Reports of phase III trials evaluating the addition of molecularly targeted agents or new monoclonal antibodies to the classic R-CHOP backbone in first-line induction or maintenance treatment were reviewed. The trial design, primary end point, number of patients enrolled, patient selection criteria, treatment schedule and results were registered for each one. In addition, the phases I and II trials which preceded these phase III trials were also reviewed. Among six phase III trials with results, only one trial evaluating lenalidomide maintenance after response to R-CHOP induction was positive and reached its primary end point. The other five trials did not show an improved outcome with the addition of the new agent. The preceding phases I and II trials were very heterogeneous in their end points and design. Even though most of these trials were considered positive, thus encouraging further investigation, so far they failed to predict the results of the subsequent phase III trials. The standard of care for DLBCL is still R-CHOP. Phase I/II trials failed to predict the results of subsequent phase III trials evaluating non-chemotherapeutic agents added to R-CHOP. The methodology of phase II trials evaluating new agents in DLBCL needs to be better defined in the future.

  6. Brucella abortus-infected B cells induce osteoclastogenesis.

    Science.gov (United States)

    Pesce Viglietti, Ayelén Ivana; Arriola Benitez, Paula Constanza; Giambartolomei, Guillermo Hernán; Delpino, María Victoria

    2016-09-01

    Brucella abortus is an intracellular bacterium that establishes lifelong infections in livestock and humans although the mechanisms of its chronicity are poorly understood. Activated B cells have long lifespan and B. abortus infection activates B cells. Our results indicate that the direct infection of B cells with B. abortus induced matrix metalloproteinase-9 (MMP-9), receptor activator for NF κB ligand (RANKL), tumor necrosis factor (TNF)-α and interleukin (IL)-6 secretion. In addition, supernatants from B. abortus-infected B cells induced bone marrow-derived monocytes to undergo osteoclastogenesis. Using osteoprotegerin, RANKL's decoy receptor, we determined that RANKL is involved in osteoclastogenesis induced by supernatants from B. abortus-infected B cells. The results presented here shed light on how the interactions of B. abortus with B cells may have a role in the pathogenesis of brucellar osteoarticular disease. Copyright © 2016 Institut Pasteur. Published by Elsevier Masson SAS. All rights reserved.

  7. LINE-1 couples EMT programming with acquisition of oncogenic phenotypes in human bronchial epithelial cells.

    Science.gov (United States)

    Reyes-Reyes, Elsa M; Aispuro, Ivan; Tavera-Garcia, Marco A; Field, Matthew; Moore, Sara; Ramos, Irma; Ramos, Kenneth S

    2017-11-28

    Although several lines of evidence have established the central role of epithelial-to-mesenchymal-transition (EMT) in malignant progression of non-small cell lung cancers (NSCLCs), the molecular events connecting EMT to malignancy remain poorly understood. This study presents evidence that Long Interspersed Nuclear Element-1 (LINE-1) retrotransposon couples EMT programming with malignancy in human bronchial epithelial cells (BEAS-2B). This conclusion is supported by studies showing that: 1) activation of EMT programming by TGF-β1 increases LINE-1 mRNAs and protein; 2) the lung carcinogen benzo(a)pyrene coregulates TGF-β1 and LINE-1 mRNAs, with LINE-1 positioned downstream of TGF-β1 signaling; and, 3) forced expression of LINE-1 in BEAS-2B cells recapitulates EMT programming and induces malignant phenotypes and tumorigenesis in vivo . These findings identify a TGFβ1-LINE-1 axis as a critical effector pathway that can be targeted for the development of precision therapies during malignant progression of intractable NSCLCs.

  8. Impact of high-dose chemotherapy and autologous transplantation as first-line therapy on the survival of high-risk diffuse large B cell lymphoma patients: a single-center study in Japan.

    Science.gov (United States)

    Inano, Shojiro; Iwasaki, Makoto; Iwamoto, Yoshihiro; Sueki, Yuki; Fukunaga, Akiko; Yanagita, Soshi; Arima, Nobuyoshi

    2014-02-01

    High-dose chemotherapy (HDT), together with autologous stem cell transplantation (ASCT), plays an important role in the treatment of diffuse large B cell lymphoma (DLBCL), especially as second-line therapy. However, its significance in up-front settings remains to be elucidated. In our institute, patients with DLBCL in both the high-intermediate and high international prognostic index (IPI) groups initially underwent CHOP/R-CHOP treatment followed by HDT/ASCT at upfront settings between 2002 and 2011. We retrospectively analyzed 25 patients who were all treated with upfront HDT/ASCT. We excluded one patient who failed to undergo transplantation because of primary refractory disease from the analysis. The median follow-up was 77 months (range 17-110 months). Five-year overall survival (OS) and progression-free survival (PFS) were 91.7 and 79.2 %, respectively, which were higher than the equivalents in previous studies. The OS and PFS in the high-risk group were lower than those in the high-intermediate group. Treatment-related mortalities or fatal complication were not observed. Our results confirm that HDT/ASCT for high-risk aggressive lymphoma is a feasible and promising therapy, but patients with high IPI continued to have poor prognoses; improvements in treatment strategy are clearly needed. Since HDT/ASCT is an aggressive treatment option associated with long-term complications, we need to identify patient groups that will gain the maximum benefit from HDT/ASCT in the upfront setting.

  9. Combining EL4-B5-based B-cell stimulation and phage display technology for the successful isolation of human anti-Scl-70 autoantibody fragments.

    Science.gov (United States)

    Weber, Malte; Weiss, Etienne; Engel, Alfred M

    2003-07-01

    Scl-70 is the major antigen recognised by autoantibodies in the sera of patients with systemic sclerosis (SSc). The autoantibodies that specifically react with Scl-70 are highly characteristic of the disease and represent valuable markers for the diagnosis of SSc. We describe a novel strategy for cloning autoantibody fragments starting with a small blood sample from an SSc patient. B cells isolated from the collected peripheral blood mononuclear cells (PBMCs) were cultured in vitro using the EL4-B5 system. Anti-Scl-70 IgG-producing cells were pooled for RNA preparation followed by the generation of phagemid libraries of approximately 10(7) independent single-chain Fvs (scFvs). The screening of these libraries by phage display allowed us to isolate four anti-Scl-70 scFvs following three rounds of biopanning. About 10 times more starting blood material was needed to generate scFv libraries of similar size from PBMCs of an SSc patient and only two anti-Scl-70 scFvs were isolated after three rounds of phage selection. Together, this work shows that functional autoantibody fragments can be advantageously cloned after in vitro expansion of B cells. The isolated anti-Scl-70 autoantibody fragments represent useful tools for calibrating SSc diagnostic assays.

  10. Trichloroethylene toxicity in a human hepatoma cell line

    Energy Technology Data Exchange (ETDEWEB)

    Thevenin, E.; McMillian, J. [Medical Univ. of Charleston South Carolina, SC (United States)

    1994-12-31

    The experiments conducted in this study were designed to determine the usefullness of hepatocyte cultures and a human hepatoma cell line as model systems for assessing human susceptibility to hepatocellular carcinoma due to exposure to trichloroethylene. The results from these studies will then be analyzed to determine if human cell lines can be used to conduct future experiments of this nature.

  11. Active evolution of memory B-cells specific to viral gH/gL/pUL128/130/131 pentameric complex in healthy subjects with silent human cytomegalovirus infection.

    Science.gov (United States)

    Xia, Lin; Tang, Aimin; Meng, Weixu; Freed, Daniel C; He, Linling; Wang, Dai; Li, Fengsheng; Li, Leike; Xiong, Wei; Gui, Xun; Schultz, Robbie D; Chen, Haotai; He, Xi; Swoyer, Ryan; Ha, Sha; Liu, Yaping; Morris, Charles D; Zhou, Yu; Wang, I-Ming; Zhao, Qinjian; Luo, Wenxin; Xia, Ningshao; Espeseth, Amy S; Hazuda, Daria J; Rupp, Richard E; Barrett, Alan D; Zhang, Ningyan; Zhu, Jiang; Fu, Tong-Ming; An, Zhiqiang

    2017-09-26

    Human cytomegalovirus (HCMV) can cause life-threatening infection in immunosuppressed patients, and in utero infection that may lead to birth defects. No vaccine is currently available. HCMV infection in healthy subjects is generally asymptomatic, and virus persists as latent infection for life. Host immunity is effective against reactivation and super-infection with another strain. Thus, vaccine candidates able to elicit immune responses similar to those of natural infection may confer protection. Since neutralization is essential for prophylactic vaccines, it is important to understand how antiviral antibodies are developed in natural infection. We hypothesized that the developmental path of antibodies in seropositive subjects could be unveiled by interrogating host B-cell repertoires using unique genetic signature sequences of mAbs. Towards this goal, we isolated 56 mAbs from three healthy donors with different neutralizing titers. Antibodies specific to the gH/gL/pUL128/130/131 pentameric complex were more potent in neutralization than those to gB. Using these mAbs as probes, patterns of extended lineage development for B-cells and evidence of active antibody maturation were revealed in two donors with higher neutralizing titers. Importantly, such patterns were limited to mAbs specific to the pentamer, but none to gB. Thus, memory B-cells with antiviral function such as neutralization were active during latent infection in the two donors, and this activity was responsible for their higher neutralizing titers. Our results indicated that memory B-cells of neutralizing capacity could be frequently mobilized in host, probably responding to silent viral episodes, further suggesting that neutralizing antibodies could play a role in control of recurrent infection.

  12. Quantitation of multiple myeloma oncogene 1/interferon-regulatory factor 4 gene expression in malignant B-cell proliferations and normal leukocytes.

    Science.gov (United States)

    Yamada, M; Asanuma, K; Kobayashi, D; Moriai, R; Yajima, T; Yagihashi, A; Yamamori, S; Watanabe, N

    2001-01-01

    We studied multiple myeloma oncogene 1/interferon-regulatory factor 4 (MUM1/IRF4) mRNA expression in various malignant human hematopoietic cell lines and normal leukocyte fractions. A quantitative reverse transcription-polymerase chain reaction was used to assess expression and chromosomes were examined for anomalies by fluorescent in situ hybridization. Among 12 cell lines examined, mRNA transcripts were expressed only in B-lymphoblastic and myeloma cell lines. Myeloma cells and malignant cell lines derived from mature B cells expressed more transcript than cell lines derived from immature B cells. Transcript levels, however, showed no association with chromosomal translocations. Expression in B-cell fractions from healthy donors was much less than in the malignant cells. In addition, MUM1/IRF4 mRNA expressed in samples from patients with acute lymphoblastic leukemia derived from B cells but not T cells. Our results suggested that MUM1/IRF4 gene expression is related to stage of differentiation of malignant B cells and they indicated the possibility that the quantitative analysis of MUM1/IRF4 gene is a useful tool for detection of malignant B-cell proliferations in clinical laboratory tests.

  13. LINE FUSION GENES: a database of LINE expression in human genes

    Directory of Open Access Journals (Sweden)

    Park Hong-Seog

    2006-06-01

    Full Text Available Abstract Background Long Interspersed Nuclear Elements (LINEs are the most abundant retrotransposons in humans. About 79% of human genes are estimated to contain at least one segment of LINE per transcription unit. Recent studies have shown that LINE elements can affect protein sequences, splicing patterns and expression of human genes. Description We have developed a database, LINE FUSION GENES, for elucidating LINE expression throughout the human gene database. We searched the 28,171 genes listed in the NCBI database for LINE elements and analyzed their structures and expression patterns. The results show that the mRNA sequences of 1,329 genes were affected by LINE expression. The LINE expression types were classified on the basis of LINEs in the 5' UTR, exon or 3' UTR sequences of the mRNAs. Our database provides further information, such as the tissue distribution and chromosomal location of the genes, and the domain structure that is changed by LINE integration. We have linked all the accession numbers to the NCBI data bank to provide mRNA sequences for subsequent users. Conclusion We believe that our work will interest genome scientists and might help them to gain insight into the implications of LINE expression for human evolution and disease. Availability http://www.primate.or.kr/line

  14. Drug Modulators of B Cell Signaling Pathways and Epstein-Barr Virus Lytic Activation.

    Science.gov (United States)

    Kosowicz, John G; Lee, Jaeyeun; Peiffer, Brandon; Guo, Zufeng; Chen, Jianmeng; Liao, Gangling; Hayward, S Diane; Liu, Jun O; Ambinder, Richard F

    2017-08-15

    Epstein-Barr virus (EBV) is a ubiquitous human gammaherpesvirus that establishes a latency reservoir in B cells. In this work, we show that ibrutinib, idelalisib, and dasatinib, drugs that block B cell receptor (BCR) signaling and are used in the treatment of hematologic malignancies, block BCR-mediated lytic induction at clinically relevant doses. We confirm that the immunosuppressive drugs cyclosporine and tacrolimus also inhibit BCR-mediated lytic induction but find that rapamycin does not inhibit BCR-mediated lytic induction. Further investigation shows that mammalian target of rapamycin complex 2 (mTORC2) contributes to BCR-mediated lytic induction and that FK506-binding protein 12 (FKBP12) binding alone is not adequate to block activation. Finally, we show that BCR signaling can activate EBV lytic induction in freshly isolated B cells from peripheral blood mononuclear cells (PBMCs) and that activation can be inhibited by ibrutinib or idelalisib. IMPORTANCE EBV establishes viral latency in B cells. Activation of the B cell receptor pathway activates lytic viral expression in cell lines. Here we show that drugs that inhibit important kinases in the BCR signaling pathway inhibit activation of lytic viral expression but do not inhibit several other lytic activation pathways. Immunosuppressant drugs such as cyclosporine and tacrolimus but not rapamycin also inhibit BCR-mediated EBV activation. Finally, we show that BCR activation of lytic infection occurs not only in tumor cell lines but also in freshly isolated B cells from patients and that this activation can be blocked by BCR inhibitors. Copyright © 2017 American Society for Microbiology.

  15. Feedback regulation of mitochondria by caspase-9 in the B cell receptor-mediated apoptosis.

    Science.gov (United States)

    Eeva, J; Nuutinen, U; Ropponen, A; Mättö, M; Eray, M; Pellinen, R; Wahlfors, J; Pelkonen, J

    2009-12-01

    During the germinal centre reaction (GC), B cells with non-functional or self-reactive antigen receptors are negatively selected by apoptosis to generate B cell repertoire with appropriate antigen specificities. We studied the molecular mechanism of Fas/CD95- and B cell receptor (BCR)-induced apoptosis to shed light on the signalling events involved in the negative selection of GC B cells. As an experimental model, we used human follicular lymphoma (FL) cell line HF1A3, which originates from a GC B cell, and transfected HF1A3 cell lines overexpressing Bcl-x(L), c-FLIP(long) or dominant negative (DN) caspase-9. Fas-induced apoptosis was dependent on the caspase-8 activation, since the overexpression of c-FLIP(long), a natural inhibitor of caspase-8 activation, blocked apoptosis induced by Fas. In contrast, caspase-9 activation was not involved in Fas-induced apoptosis. BCR-induced apoptosis showed the typical characteristics of mitochondria-dependent (intrinsic) apoptosis. Firstly, the activation of caspase-9 was involved in BCR-induced DNA fragmentation, while caspase-8 showed only marginal role. Secondly, overexpression of Bcl-x(L) could block all apoptotic changes induced by BCR. As a novel finding, we demonstrate that caspase-9 can enhance the cytochrome-c release and collapse of mitochondrial membrane potential (DeltaPsi(m)) during BCR-induced apoptosis. The requirement of different signalling pathways in apoptosis induced by BCR and Fas may be relevant, since Fas- and BCR-induced apoptosis can thus be regulated independently, and targeted to different subsets of GC B cells.

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

    Directory of Open Access Journals (Sweden)

    Sabyasachi Halder

    2009-09-01

    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.

  17. Cloning of B cell-specific membrane tetraspanning molecule BTS possessing B cell proliferation-inhibitory function.

    Science.gov (United States)

    Suenaga, Tadahiro; Arase, Hisashi; Yamasaki, Sho; Kohno, Masayuki; Yokosuka, Tadashi; Takeuchi, Arata; Hattori, Takamichi; Saito, Takashi

    2007-11-01

    Lymphocyte proliferation is regulated by signals through antigen receptors, co-stimulatory receptors, and other positive and negative modulators. Several membrane tetraspanning molecules are also involved in the regulation of lymphocyte growth and death. We cloned a new B cell-specific tetraspanning (BTS) membrane molecule, which is similar to CD20 in terms of expression, structure and function. BTS is specifically expressed in the B cell line and its expression is increased after the pre-B cell stage. BTS is expressed in intracellular granules and on the cell surface. Overexpression of BTS in immature B cell lines induces growth retardation through inhibition of cell cycle progression and cell size increase without inducing apoptosis. This inhibitory function is mediated predominantly by the N terminus of BTS. The development of mature B cells is inhibited in transgenic mice expressing BTS, suggesting that BTS is involved in the in vivo regulation of B cells. These results indicate that BTS plays a role in the regulation of cell division and B cell growth.

  18. Liraglutide, a once-daily human GLP-1 analogue, improves pancreatic B-cell function and arginine-stimulated insulin secretion during hyperglycaemia in patients with Type 2 diabetes mellitus

    DEFF Research Database (Denmark)

    Vilsbøll, Tina; Brock, Birgitte; Perrild, Hans

    2008-01-01

    To assess the effect of liraglutide, a once-daily human glucagon-like peptide-1 analogue on pancreatic B-cell function. methods: Patients with Type 2 diabetes (n = 39) were randomized to treatment with 0.65, 1.25 or 1.9 mg/day liraglutide or placebo for 14 weeks. First- and second-phase insulin...... release were measured by means of the insulin-modified frequently sampled intravenous glucose tolerance test. Arginine-stimulated insulin secretion was measured during a hyperglycaemic clamp (20 mmol/l). Glucose effectiveness and insulin sensitivity were estimated by means of the insulin...

  19. 3-Bromopyruvate induces endoplasmic reticulum stress, overcomes autophagy and causes apoptosis in human HCC cell lines.

    Science.gov (United States)

    Ganapathy-Kanniappan, Shanmugasundaram; Geschwind, Jean-Francois H; Kunjithapatham, Rani; Buijs, Manon; Syed, Labiq H; Rao, Pramod P; Ota, Shinichi; Kwak, Byung Kook; Loffroy, Romaric; Vali, Mustafa

    2010-03-01

    Autophagy, a cellular response to stress, plays a role in resistance to chemotherapy in cancer cells. Resistance renders systemic chemotherapy generally ineffective against human hepatocellular carcinoma (HCC). Recently, we reported that the pyruvate analog 3-bromopyruvate (3-BrPA) promoted tumor cell death by targeting GAPDH. In continuance, we investigated the intracellular response of two human HCC cell lines (Hep3B and SK-Hep1) that differ in their status of key apoptotic regulators, p53 and Fas. 3-BrPA treatment induced endoplasmic reticulum (ER) stress, translation inhibition and apoptosis based on Western blot and qPCR, pulse labeling, Terminal deoxynucleotidyl transferase dUTP nick end labeling (TUNEL) assay and active caspase-3 in both the cell lines. However, electron microscopy revealed that 3-BrPA treated SK-Hep1 cells underwent classical apoptotic cell death while Hep3B cells initially responded with the protective autophagy that failed to prevent eventual apoptosis. 3-BrPA treatment promotes apoptosis in human HCC cell lines, irrespective of the intracellular response.

  20. Tonsillar CD56brightNKG2A+ NK cells restrict primary Epstein-Barr virus infection in B cells via IFN-γ.

    Science.gov (United States)

    Jud, Aurelia; Kotur, Monika; Berger, Christoph; Gysin, Claudine; Nadal, David; Lünemann, Anna

    2017-01-24

    Natural killer (NK) cells constitute the first line of defense against viruses and cancers cells. Epstein-Barr virus (EBV) was the first human virus to be directly implicated in carcinogenesis, and EBV infection is associated with a broad spectrum of B cell lymphomas. How NK cells restrict EBV-associated oncogenesis is not understood. Here, we investigated the efficacies and mechanisms of distinct NK cell subsets from tonsils, the portal of entry of EBV, in limiting EBV infection in naïve, germinal center-associated and memory B cells. We found that CD56bright and NKG2A expression sufficiently characterizes the potent anti-EBV capacity of tonsillar NK cells. We observed restriction of EBV infection in B cells as early as 18 hours after infection. The restriction was most efficient in naïve B cells and germinal center-associated B cells, the B cell subsets that exhibited highest susceptibility to EBV infection in vitro. IFN-γ release by and partially NKp44 engagement of CD56bright and NKG2A positive NK cells mediated the restriction that eventually inhibited B-cell transformation. Thus, harnessing CD56brightNKG2A+ NK cell function might be promising to improve treatment strategies that target EBV-associated B cell lymphomas.

  1. MODERATE CYTOTOXICITY OF PROANTHOCYANIDINS TO HUMAN TUMOR-CELL LINES

    NARCIS (Netherlands)

    KOLODZIEJ, H; HABERLAND, C; WOERDENBAG, HJ; KONINGS, AWT

    In the present study the cytotoxicity of 16 proanthocyanidins was evaluated in GLC(4), a human small cell lung carcinoma cell line, and in COLO 320, a human colorectal cancer cell line, using the microculture tetrazolium (MTT) assay. With IC50 values ranging from 18 to >200 mu m following continuous

  2. Tuft (caveolated) cells in two human colon carcinoma cell lines.

    OpenAIRE

    Barkla, D. H.; Whitehead, R. H.; Foster, H.; Tutton, P. J.

    1988-01-01

    The presence of an unusual cell type in two human colon carcinoma cell lines is reported. The cells show the same morphology as "tuft" (caveolated) cells present in normal gastrointestinal epithelium. Tuft cells were seen in cell line LIM 1863 growing in vitro and in human colon carcinoma cell line LIM 2210 growing as subcutaneous solid tumour xenografts in nude mice. Characteristic morphologic features of tuft cells included a wide base, narrow apex and a tuft of long microvilli projecting f...

  3. 5′ Rapid Amplification of cDNA Ends and Illumina MiSeq Reveals B Cell Receptor Features in Healthy Adults, Adults With Chronic HIV-1 Infection, Cord Blood, and Humanized Mice

    Directory of Open Access Journals (Sweden)

    Eric Waltari

    2018-03-01

    Full Text Available Using 5′ rapid amplification of cDNA ends, Illumina MiSeq, and basic flow cytometry, we systematically analyzed the expressed B cell receptor (BCR repertoire in 14 healthy adult PBMCs, 5 HIV-1+ adult PBMCs, 5 cord blood samples, and 3 HIS-CD4/B mice, examining the full-length variable region of μ, γ, α, κ, and λ chains for V-gene usage, somatic hypermutation (SHM, and CDR3 length. Adding to the known repertoire of healthy adults, Illumina MiSeq consistently detected small fractions of reads with high mutation frequencies including hypermutated μ reads, and reads with long CDR3s. Additionally, the less studied IgA repertoire displayed similar characteristics to that of IgG. Compared to healthy adults, the five HIV-1 chronically infected adults displayed elevated mutation frequencies for all μ, γ, α, κ, and λ chains examined and slightly longer CDR3 lengths for γ, α, and λ. To evaluate the reconstituted human BCR sequences in a humanized mouse model, we analyzed cord blood and HIS-CD4/B mice, which all lacked the typical SHM seen in the adult reference. Furthermore, MiSeq revealed identical unmutated IgM sequences derived from separate cell aliquots, thus for the first time demonstrating rare clonal members of unmutated IgM B cells by sequencing.

  4. 5' Rapid Amplification of cDNA Ends and Illumina MiSeq Reveals B Cell Receptor Features in Healthy Adults, Adults With Chronic HIV-1 Infection, Cord Blood, and Humanized Mice.

    Science.gov (United States)

    Waltari, Eric; Jia, Manxue; Jiang, Caroline S; Lu, Hong; Huang, Jing; Fernandez, Cristina; Finzi, Andrés; Kaufmann, Daniel E; Markowitz, Martin; Tsuji, Moriya; Wu, Xueling

    2018-01-01

    Using 5' rapid amplification of cDNA ends, Illumina MiSeq, and basic flow cytometry, we systematically analyzed the expressed B cell receptor (BCR) repertoire in 14 healthy adult PBMCs, 5 HIV-1+ adult PBMCs, 5 cord blood samples, and 3 HIS-CD4/B mice, examining the full-length variable region of μ, γ, α, κ, and λ chains for V-gene usage, somatic hypermutation (SHM), and CDR3 length. Adding to the known repertoire of healthy adults, Illumina MiSeq consistently detected small fractions of reads with high mutation frequencies including hypermutated μ reads, and reads with long CDR3s. Additionally, the less studied IgA repertoire displayed similar characteristics to that of IgG. Compared to healthy adults, the five HIV-1 chronically infected adults displayed elevated mutation frequencies for all μ, γ, α, κ, and λ chains examined and slightly longer CDR3 lengths for γ, α, and λ. To evaluate the reconstituted human BCR sequences in a humanized mouse model, we analyzed cord blood and HIS-CD4/B mice, which all lacked the typical SHM seen in the adult reference. Furthermore, MiSeq revealed identical unmutated IgM sequences derived from separate cell aliquots, thus for the first time demonstrating rare clonal members of unmutated IgM B cells by sequencing.

  5. B cells in operational tolerance.

    Science.gov (United States)

    Chesneau, M; Danger, R; Soulillou, J-P; Brouard, S

    2018-02-16

    Transplantation is currently the therapy of choice for endstage organ failure even though it requires long-term immunosuppresive therapy, with its numerous side effects, for acceptance of the transplanted organ. In rare cases however, patients develop operational tolerance, that is, graft survival without immunosuppression. Studies conducted on these patients reveal genetic, phenotypic, and functional signatures. They provide a better understanding of the immunological mechanisms involved in operational tolerance and define biomarkers that could be used to adapt immunosuppressive treatment to the individual, safely reduce immunosuppression doses, and ideally and safely guide immunosuppression withdrawal. This review summarizes studies that suggest a role for B cells as biomarkers of operational tolerance and discusses the use of B cells as a predictive tool for immunologic risk. Copyright © 2018. Published by Elsevier Inc.

  6. Regulation of normal B-cell differentiation and malignant B-cell survival by OCT2.

    Science.gov (United States)

    Hodson, Daniel J; Shaffer, Arthur L; Xiao, Wenming; Wright, George W; Schmitz, Roland; Phelan, James D; Yang, Yandan; Webster, Daniel E; Rui, Lixin; Kohlhammer, Holger; Nakagawa, Masao; Waldmann, Thomas A; Staudt, Louis M

    2016-04-05

    The requirement for the B-cell transcription factor OCT2 (octamer-binding protein 2, encoded by Pou2f2) in germinal center B cells has proved controversial. Here, we report that germinal center B cells are formed normally after depletion of OCT2 in a conditional knockout mouse, but their proliferation is reduced and in vivo differentiation to antibody-secreting plasma cells is blocked. This finding led us to examine the role of OCT2 in germinal center-derived lymphomas. shRNA knockdown showed that almost all diffuse large B-cell lymphoma (DLBCL) cell lines are addicted to the expression of OCT2 and its coactivator OCA-B. Genome-wide chromatin immunoprecipitation (ChIP) analysis and gene-expression profiling revealed the broad transcriptional program regulated by OCT2 that includes the expression of STAT3, IL-10, ELL2, XBP1, MYC, TERT, and ADA. Importantly, genetic alteration of OCT2 is not a requirement for cellular addiction in DLBCL. However, we detected amplifications of the POU2F2 locus in DLBCL tumor biopsies and a recurrent mutation of threonine 223 in the DNA-binding domain of OCT2. This neomorphic mutation subtly alters the DNA-binding preference of OCT2, leading to the transactivation of noncanonical target genes including HIF1a and FCRL3 Finally, by introducing mutations designed to disrupt the OCT2-OCA-B interface, we reveal a requirement for this protein-protein interface that ultimately might be exploited therapeutically. Our findings, combined with the predominantly B-cell-restricted expression of OCT2 and the absence of a systemic phenotype in our knockout mice, suggest that an OCT2-targeted therapeutic strategy would be efficacious in both major subtypes of DLBCL while avoiding systemic toxicity.

  7. Fraction against Human Cancer Cell Lines

    African Journals Online (AJOL)

    fraction of A. sieberi against seven cancer cell lines (Colo20, HCT116, DLD, MCF7, Jurkat, HepG2 and ... The morphology of the HepG2 cell nucleus was investigated by Hoechst 33342, ..... Gong F, Liang Y, Xie P, Chau F. Information theory.

  8. (Asteraceae) Fraction against Human Cancer Cell Lines

    African Journals Online (AJOL)

    Purpose: To investigate the anti-proliferative and apoptotic activity of crude and dichloromethane fraction of A. sieberi against seven cancer cell lines (Colo20, HCT116, DLD, MCF7, Jurkat, HepG2 and L929). Methods: A. sieberi was extracted with methanol and further purification was carried out using liquidliquid extraction ...

  9. Membrane-associated signaling in human B-lymphoma lines

    Energy Technology Data Exchange (ETDEWEB)

    Tauzin, Sebastien; Ding, Heidrun; Burdevet, Dimitri [Department of Pathology and Immunology, Centre medical universitaire, 1, rue Michel-Servet, 1211 Geneva 11 (Switzerland); Borisch, Bettina [Department of Social and Preventive Medicine, Centre medical universitaire, 1, rue Michel-Servet, 1211 Geneva 11 (Switzerland); Hoessli, Daniel C., E-mail: danielhoessli@gmail.com [Department of Pathology and Immunology, Centre medical universitaire, 1, rue Michel-Servet, 1211 Geneva 11 (Switzerland)

    2011-01-15

    In B-non-Hodgkin lymphomas, Lyn and Cbp/PAG constitute the core of an oncogenic signalosome that captures the Phosphatidylinositol-3-kinase, the Spleen tyrosine kinase and the Signal transducer and activator of transcription-3 to generate pro-survival and proliferative signals. Lymphoma lines corresponding to follicular, mantle-cell and Burkitt-derived lymphomas display type-specific signalosome organizations that differentially activate PI3K, Syk and STAT3. In the follicular lymphoma line, PI3K, Syk and STAT3 were optimally activated upon association with the Lyn-Cbp/PAG signalosome, while in the Burkitt lymphoma-derived line, the association with Cbp/PAG and activation of PI3K were interfered with by the latent membrane proteins encoded by the Epstein-Barr virus. In the Jeko-1 mantle-cell line, a weak association of Syk with the Lyn-Cbp/PAG signalosome resulted in poor activation of Syk, but in those cells, as in the follicular and Burkitt-derived lines, efficient apoptosis induction by the Syk inhibitor R406 indicated that Syk is nonetheless an important prosurvival element and therefore a valuable therapeutic target. In all configurations described herein is the Lyn-Cbp/PAG signalosome independent of external signals and provides efficient means of activation for its associated lipid and protein kinases. In follicular and Burkitt-derived lines, Syk appears to be activated following binding to Cbp/PAG and no longer requires B-cell receptor-associated activation motifs for activation. Assessment of the different modalities of Lyn-Cbp/PAG signalosome organization could help in selecting the appropriate combination of kinase inhibitors to eliminate a particular type of lymphoma cells.

  10. Evaluation of Antigen-Conjugated Fluorescent Beads to Identify Antigen-Specific B Cells.

    Science.gov (United States)

    Correa, Isabel; Ilieva, Kristina M; Crescioli, Silvia; Lombardi, Sara; Figini, Mariangela; Cheung, Anthony; Spicer, James F; Tutt, Andrew N J; Nestle, Frank O; Karagiannis, Panagiotis; Lacy, Katie E; Karagiannis, Sophia N

    2018-01-01

    Selection of single antigen-specific B cells to identify their expressed antibodies is of considerable interest for evaluating human immune responses. Here, we present a method to identify single antibody-expressing cells using antigen-conjugated fluorescent beads. To establish this, we selected Folate Receptor alpha (FRα) as a model antigen and a mouse B cell line, expressing both the soluble and the membrane-bound forms of a human/mouse chimeric antibody (MOv18 IgG1) specific for FRα, as test antibody-expressing cells. Beads were conjugated to FRα using streptavidin/avidin-biotin bridges and used to select single cells expressing the membrane-bound form of anti-FRα. Bead-bound cells were single cell-sorted and processed for single cell RNA retrotranscription and PCR to isolate antibody heavy and light chain variable regions. Variable regions were then cloned and expressed as human IgG1/k antibodies. Like the original clone, engineered antibodies from single cells recognized native FRα. To evaluate whether antigen-coated beads could identify specific antibody-expressing cells in mixed immune cell populations, human peripheral blood mononuclear cells (PBMCs) were spiked with test antibody-expressing cells. Antigen-specific cells could comprise up to 75% of cells selected with antigen-conjugated beads when the frequency of the antigen-positive cells was 1:100 or higher. In PBMC pools, beads conjugated to recombinant antigens FRα and HER2 bound antigen-specific anti-FRα MOv18 and anti-HER2 Trastuzumab antibody-expressing cells, respectively. From melanoma patient-derived B cells selected with melanoma cell line-derived protein-coated fluorescent beads, we generated a monoclonal antibody that recognized melanoma antigen-coated beads. This approach may be further developed to facilitate analysis of B cells and their antibody profiles at the single cell level and to help unravel humoral immune repertoires.

  11. Evaluation of Antigen-Conjugated Fluorescent Beads to Identify Antigen-Specific B Cells

    Directory of Open Access Journals (Sweden)

    Isabel Correa

    2018-03-01

    Full Text Available Selection of single antigen-specific B cells to identify their expressed antibodies is of considerable interest for evaluating human immune responses. Here, we present a method to identify single antibody-expressing cells using antigen-conjugated fluorescent beads. To establish this, we selected Folate Receptor alpha (FRα as a model antigen and a mouse B cell line, expressing both the soluble and the membrane-bound forms of a human/mouse chimeric antibody (MOv18 IgG1 specific for FRα, as test antibody-expressing cells. Beads were conjugated to FRα using streptavidin/avidin-biotin bridges and used to select single cells expressing the membrane-bound form of anti-FRα. Bead-bound cells were single cell-sorted and processed for single cell RNA retrotranscription and PCR to isolate antibody heavy and light chain variable regions. Variable regions were then cloned and expressed as human IgG1/k antibodies. Like the original clone, engineered antibodies from single cells recognized native FRα. To evaluate whether antigen-coated beads could identify specific antibody-expressing cells in mixed immune cell populations, human peripheral blood mononuclear cells (PBMCs were spiked with test antibody-expressing cells. Antigen-specific cells could comprise up to 75% of cells selected with antigen-conjugated beads when the frequency of the antigen-positive cells was 1:100 or higher. In PBMC pools, beads conjugated to recombinant antigens FRα and HER2 bound antigen-specific anti-FRα MOv18 and anti-HER2 Trastuzumab antibody-expressing cells, respectively. From melanoma patient-derived B cells selected with melanoma cell line-derived protein-coated fluorescent beads, we generated a monoclonal antibody that recognized melanoma antigen-coated beads. This approach may be further developed to facilitate analysis of B cells and their antibody profiles at the single cell level and to help unravel humoral immune repertoires.

  12. Evaluation of Antigen-Conjugated Fluorescent Beads to Identify Antigen-Specific B Cells

    Science.gov (United States)

    Correa, Isabel; Ilieva, Kristina M.; Crescioli, Silvia; Lombardi, Sara; Figini, Mariangela; Cheung, Anthony; Spicer, James F.; Tutt, Andrew N. J.; Nestle, Frank O.; Karagiannis, Panagiotis; Lacy, Katie E.; Karagiannis, Sophia N.

    2018-01-01

    Selection of single antigen-specific B cells to identify their expressed antibodies is of considerable interest for evaluating human immune responses. Here, we present a method to identify single antibody-expressing cells using antigen-conjugated fluorescent beads. To establish this, we selected Folate Receptor alpha (FRα) as a model antigen and a mouse B cell line, expressing both the soluble and the membrane-bound forms of a human/mouse chimeric antibody (MOv18 IgG1) specific for FRα, as test antibody-expressing cells. Beads were conjugated to FRα using streptavidin/avidin-biotin bridges and used to select single cells expressing the membrane-bound form of anti-FRα. Bead-bound cells were single cell-sorted and processed for single cell RNA retrotranscription and PCR to isolate antibody heavy and light chain variable regions. Variable regions were then cloned and expressed as human IgG1/k antibodies. Like the original clone, engineered antibodies from single cells recognized native FRα. To evaluate whether antigen-coated beads could identify specific antibody-expressing cells in mixed immune cell populations, human peripheral blood mononuclear cells (PBMCs) were spiked with test antibody-expressing cells. Antigen-specific cells could comprise up to 75% of cells selected with antigen-conjugated beads when the frequency of the antigen-positive cells was 1:100 or higher. In PBMC pools, beads conjugated to recombinant antigens FRα and HER2 bound antigen-specific anti-FRα MOv18 and anti-HER2 Trastuzumab antibody-expressing cells, respectively. From melanoma patient-derived B cells selected with melanoma cell line-derived protein-coated fluorescent beads, we generated a monoclonal antibody that recognized melanoma antigen-coated beads. This approach may be further developed to facilitate analysis of B cells and their antibody profiles at the single cell level and to help unravel humoral immune repertoires. PMID:29628923

  13. Peptidomic analysis of human cell lines

    Science.gov (United States)

    Gelman, Julia S.; Sironi, Juan; Castro, Leandro M.; Ferro, Emer S.; Fricker, Lloyd D.

    2011-01-01

    Peptides have been proposed to function in intracellular signaling within the cytosol. Although cytosolic peptides are considered to be highly unstable, a large number of peptides have been detected in mouse brain and other biological samples. In the present study, we evaluated the peptidome of three diverse cell lines: SH-SY5Y, MCF7, and HEK293 cells. A comparison of the peptidomes revealed considerable overlap in the identity of the peptides found in each cell line. The majority of the observed peptides are not derived from the most abundant or least stable proteins in the cell, and approximately half of the cellular peptides correspond to the N- or C- termini of the precursor proteins. Cleavage site analysis revealed a preference for hydrophobic residues in the P1 position. Quantitative peptidomic analysis indicated that the levels of most cellular peptides are not altered in response to elevated intracellular calcium, suggesting that calpain is not responsible for their production. The similarity of the peptidomes of the three cell lines and the lack of correlation with the predicted cellular degradome implies the selective formation or retention of these peptides, consistent with the hypothesis that they are functional in the cells. PMID:21204522

  14. Derivation of the human embryonic stem cell line RCM1

    Directory of Open Access Journals (Sweden)

    P.A. De Sousa

    2016-03-01

    Full Text Available The human embryonic stem cell line RCM-1 was derived from a failed to fertilise egg undergoing parthenogenetic stimulation. The cell line shows normal pluripotency marker expression and differentiation to three germ layers in vitro and in vivo. It has a normal 46XX female karyotype and microsatellite PCR identity, HLA and blood group typing data is available.

  15. Effects of anti-CD40 mAb on inducing malignant B cells proliferation arrest and apoptosis and its mechanism

    International Nuclear Information System (INIS)

    Tang Lin; Zhuang Yumei; Zhou Zhaohua; Yu Gehua; Pan Jianzhong; Zhang Xueguang

    2002-01-01

    Objective: To study the expression of CD 40 molecule and the biological effects mediated by CD 40 molecules on malignant B cells. Methods: Agonistic anti-human CD 40 monoclonal antibody (clone 5C11) was added to cell culture system. Cell counting, PI staining, Annexin-V staining and flow cytometric analysis were used to study the behavior of malignant B cell lines after treatment with mAb clone 5C11. Results: 5C11 induced homotypic aggregation and proliferation arrest and mediated apoptosis in multiple myeloma cell line XG2 that expressed CD 40 strongly; 5C11 induced B lymphoma cell line Daudi homotypic aggregation and proliferation arrest and apoptosis, the apoptosis of XG2 and Daudi by CD40 activation was not mediated by TNF. Conclusion: Agonistic anti-CD 40 mAb 5C11 can inhibit the proliferation of malignant B cells by inducing them to die apoplectically

  16. IL-21: an executor of B cell fate.

    Science.gov (United States)

    Konforte, Danijela; Simard, Nathalie; Paige, Christopher J

    2009-02-15

    IL-21 is a type I cytokine that shares the common receptor gamma-chain with IL-2, IL-4, IL-7, IL-9, and IL-15. B cells are one of the lymphoid cell types whose development and function are regulated by IL-21. Depending on the interplay with costimulatory signals and on the developmental stage of a B cell, IL-21 can induce proliferation, differentiation into Ig-producing plasma cells, or apoptosis in both mice and humans. Alone and in combination with Th cell-derived cytokines IL-21 can regulate class switch recombination to IgG, IgA, or IgE isotypes, indicating its important role in shaping the effector function of B cells. This review highlights the role of IL-21 in B cell development, function, and disease and provides some perspectives on the future studies in this area.

  17. Safety and efficacy of ofatumumab, a fully human monoclonal anti-CD20 antibody, in patients with relapsed or refractory B-cell chronic lymphocytic leukemia

    DEFF Research Database (Denmark)

    Coiffier, Bertrand; Lepretre, Stéphane; Pedersen, Lars Møller

    2008-01-01

    Safety and efficacy of the fully human anti-CD20 monoclonal antibody, ofatumumab, was analyzed in a multicenter dose-escalating study including 33 patients with relapsed or refractory chronic lymphocytic leukemia. Three cohorts of 3 (A), 3 (B), and 27 (C) patients received 4, once weekly, infusio...

  18. Adipose Tissue Inflammation Induces B Cell Inflammation and Decreases B Cell Function in Aging

    Directory of Open Access Journals (Sweden)

    Daniela Frasca

    2017-08-01

    Full Text Available Aging is the greatest risk factor for developing chronic diseases. Inflamm-aging, the age-related increase in low-grade chronic inflammation, may be a common link in age-related diseases. This review summarizes recent published data on potential cellular and molecular mechanisms of the age-related increase in inflammation, and how these contribute to decreased humoral immune responses in aged mice and humans. Briefly, we cover how aging and related inflammation decrease antibody responses in mice and humans, and how obesity contributes to the mechanisms for aging through increased inflammation. We also report data in the literature showing adipose tissue infiltration with immune cells and how these cells are recruited and contribute to local and systemic inflammation. We show that several types of immune cells infiltrate the adipose tissue and these include macrophages, neutrophils, NK cells, innate lymphoid cells, eosinophils, T cells, B1, and B2 cells. Our main focus is how the adipose tissue affects immune responses, in particular B cell responses and antibody production. The role of leptin in generating inflammation and decreased B cell responses is also discussed. We report data published by us and by other groups showing that the adipose tissue generates pro-inflammatory B cell subsets which induce pro-inflammatory T cells, promote insulin resistance, and secrete pathogenic autoimmune antibodies.

  19. Novel human multiple myeloma cell line UHKT-893

    Czech Academy of Sciences Publication Activity Database

    Uherková, L.; Vančurová, I.; Vyhlídalová, I.; Pleschnerová, M.; Špička, I.; Mihalová, R.; Březinová, J.; Hodný, Zdeněk; Čermáková, K.; Polanská, V.; Marinov, I.; Jedelský, P.L.; Kuželová, K.; Stöckbauer, P.

    2013-01-01

    Roč. 37, č. 3 (2013), s. 320-326 ISSN 0145-2126 Institutional support: RVO:68378050 Keywords : human myeloma cell line * human multiple myeloma * plasma cell * IL-6 dependence * immunoglobulin * free light chain Subject RIV: EB - Genetics ; Molecular Biology Impact factor: 2.692, year: 2013

  20. Cell Line Data Base: structure and recent improvements towards molecular authentication of human cell lines.

    Science.gov (United States)

    Romano, Paolo; Manniello, Assunta; Aresu, Ottavia; Armento, Massimiliano; Cesaro, Michela; Parodi, Barbara

    2009-01-01

    The Cell Line Data Base (CLDB) is a well-known reference information source on human and animal cell lines including information on more than 6000 cell lines. Main biological features are coded according to controlled vocabularies derived from international lists and taxonomies. HyperCLDB (http://bioinformatics.istge.it/hypercldb/) is a hypertext version of CLDB that improves data accessibility by also allowing information retrieval through web spiders. Access to HyperCLDB is provided through indexes of biological characteristics and navigation in the hypertext is granted by many internal links. HyperCLDB also includes links to external resources. Recently, an interest was raised for a reference nomenclature for cell lines and CLDB was seen as an authoritative system. Furthermore, to overcome the cell line misidentification problem, molecular authentication methods, such as fingerprinting, single-locus short tandem repeat (STR) profile and single nucleotide polymorphisms validation, were proposed. Since this data is distributed, a reference portal on authentication of human cell lines is needed. We present here the architecture and contents of CLDB, its recent enhancements and perspectives. We also present a new related database, the Cell Line Integrated Molecular Authentication (CLIMA) database (http://bioinformatics.istge.it/clima/), that allows to link authentication data to actual cell lines.

  1. Lym-1 Chimeric Antigen Receptor T Cells Exhibit Potent Anti-Tumor Effects against B-Cell Lymphoma

    Directory of Open Access Journals (Sweden)

    Long Zheng

    2017-12-01

    Full Text Available T cells expressing chimeric antigen receptors (CARs recognizing CD19 epitopes have produced remarkable anti-tumor effects in patients with B-cell malignancies. However, cancer cells lacking recognized epitopes can emerge, leading to relapse and death. Thus, CAR T cells targeting different epitopes on different antigens could improve immunotherapy. The Lym-1 antibody targets a conformational epitope of Human Leukocyte Antigen-antigen D Related (HLA-DR on the surface of human B-cell lymphomas. Lym-1 CAR T cells were thus generated for evaluation of cytotoxic activity towards lymphoma cells in vitro and in vivo. Human T cells from healthy donors were transduced to express a Lym-1 CAR, and assessed for epitope-driven function in culture and towards Raji xenografts in NOD-scidIL2Rgammanull (NSG mice. Lym-1 CAR T cells exhibited epitope-driven activation and lytic function against human B-cell lymphoma cell lines in culture and mediated complete regression of Raji/Luciferase-Green fluorescent protein (Raji/Luc-GFP in NSG mice with similar or better reactivity than CD19 CAR T cells. Lym-1 CAR transduction of T cells is a promising immunotherapy for patients with Lym-1 epitope positive B-cell malignancies.

  2. Effect of irradiation on T-cell suppression of ELISA-determined Ig production by human blood B-cells in vitro

    Energy Technology Data Exchange (ETDEWEB)

    Wasserman, J; Stedingk, L.V. von; Biberfeld, G; Petrini, B; Blomgren, H; Baral, E [Central Microbiologcal Lab. of Stockholm County Council (Sweden)

    1979-11-01

    Human blood B-lymphocytes were co-cultured with in vitro irradiated allogeneic or autologous T-lymphocytes in the presence of pokeweed mitogen (PWM). The production of IgG, IgM and IgA, as assessed by the enzyme-linked immunosorbent assay (ELISA) was increased 2-7 times, as compared to values obtained with non-irradiated T-lymphocytes. It was suggested that the increase of Ig production was due to the selective radiosensitivity of T-lymphocytes with suppressor function. (author).

  3. HCV Infection and B-Cell Lymphomagenesis

    Directory of Open Access Journals (Sweden)

    Masahiko Ito

    2011-01-01

    Full Text Available Hepatitis C virus (HCV has been recognized as a major cause of chronic liver diseases worldwide. It has been suggested that HCV infects not only hepatocytes but also mononuclear lymphocytes including B cells that express the CD81 molecule, a putative HCV receptor. HCV infection of B cells is the likely cause of B-cell dysregulation disorders such as mixed cryoglobulinemia, rheumatoid factor production, and B-cell lymphoproliferative disorders that may evolve into non-Hodgkin's lymphoma (NHL. Epidemiological data indicate an association between HCV chronic infection and the occurrence of B-cell NHL, suggesting that chronic HCV infection is associated at least in part with B-cell lymphomagenesis. In this paper, we aim to provide an overview of recent literature, including our own, to elucidate a possible role of HCV chronic infection in B-cell lymphomagenesis.

  4. Cell- and stage-specific chromatin structure across the Complement receptor 2 (CR2/CD21) promoter coincide with CBF1 and C/EBP-beta binding in B cells.

    Science.gov (United States)

    Cruickshank, Mark N; Fenwick, Emily; Karimi, Mahdad; Abraham, Lawrence J; Ulgiati, Daniela

    2009-08-01

    Stringent developmental transcription requires multiple transcription factor (TF) binding sites, cell-specific expression of signaling molecules, TFs and co-regulators and appropriate chromatin structure. During B-lymphopoiesis, human Complement receptor 2 (CR2/CD21) is detected on immature and mature B cells but not on B cell precursors and plasma cells. We examined cell- and stage-specific human CR2 gene regulation using cell lines modeling B-lymphopoiesis. Chromatin accessibility assays revealed a region between -409 and -262 with enhanced accessibility in mature B cells and pre-B cells, compared to either non-lymphoid or plasma cell-types, however, accessibility near the transcription start site (TSS) was elevated only in CR2-expressing B cells. A correlation between histone acetylation and CR2 expression was observed, while histone H3K4 dimethylation was enriched near the TSS in both CR2-expressing B cells and non-expressing pre-B cells. Candidate sites within the CR2 promoter were identified which could regulate chromatin, including a matrix attachment region associated with CDP, SATB1/BRIGHT and CEBP-beta sites as well as two CBF1 sites. ChIP assays verified that both CBF1 and C/EBP-beta bind the CR2 promoter in B cells raising the possibility that these factors facilitate or respond to alterations in chromatin structure to control the timing and/or level of CR2 transcription.

  5. Reprogramming of B cells into macrophages: mechanistic insights

    OpenAIRE

    Di Tullio, Alessandro, 1982-

    2012-01-01

    Our earlier work has shown that pre-B cells can be converted into macrophages by the transcription factor C/EBPα at very high frequencies and also that a clonal pre-B cell line with an inducible form of C/EBPα can be converted into macrophage-like cells. Using these systems we have performed a systematic analysis of the questions whether during transdifferentiation the cells retrodifferentiate to a precursor cell state and whether cell cycle is required for reprogramming. As for the first ...

  6. Transitional-2 B cells acquire regulatory function during tolerance induction and contribute to allograft survival.

    Science.gov (United States)

    Moreau, Aurélie; Blair, Paul A; Chai, Jian-Guo; Ratnasothy, Kulachelvy; Stolarczyk, Emilie; Alhabbab, Rowa; Rackham, Chloe L; Jones, Peter M; Smyth, Lesley; Elgueta, Raul; Howard, Jane K; Lechler, Robert I; Lombardi, Giovanna

    2015-03-01

    In humans, tolerance to renal transplants has been associated with alterations in B-cell gene transcription and maintenance of the numbers of circulating transitional B cells. Here, we use a mouse model of transplantation tolerance to investigate the contribution of B cells to allograft survival. We demonstrate that transfer of B cells from mice rendered tolerant to MHC class I mismatched skin grafts can prolong graft survival in a dose-dependent and antigen-specific manner to a degree similar to that afforded by graft-specific regulatory T (Treg) cells. Tolerance in this model was associated with an increase in transitional-2 (T2) B cells. Only T2 B cells from tolerized mice, not naïve T2 nor alloantigen experienced T2, were capable of prolonging skin allograft survival, and suppressing T-cell activation. Tolerized T2 B cells expressed lower levels of CD86, increased TIM-1, and demonstrated a preferential survival in vivo. Furthermore, we demonstrate a synergistic effect between tolerized B cells and graft-specific Treg cells. IL-10 production by T2 B cells did not contribute to tolerance, as shown by transfer of B cells from IL-10(-/-) mice. These results suggest that T2 B cells in tolerant patients may include a population of regulatory B cells that directly inhibit graft rejection. © 2014 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  7. Differential radiosensitivity among B cell subpopulations

    International Nuclear Information System (INIS)

    Riggs, J.E.

    1988-01-01

    The selective radiosensitivity of sIgM >> sIgD marginal zone B cells is associated with the selective loss of B cell function. The simultaneous restoration of impaired function and recovery of these cells with time supports this premise. B cell recovery, delayed one week after irradiation, is in progress at two weeks, and virtually complete by three weeks. XID mice reveal similar recovery kinetics although there are fewer recovering cells and these bear reduced levels of Ia. This observation represents additional evidence that xid B cells are distinct from those of normal mice. The simultaneous loss, and concurrent recovery, of sIgM >> sIgD B cells and TI-2 responsiveness in irradiated mice suggests the existence of a unique B cell subpopulation possessing both phenotypes. Additional support for this hypothesis is provided by demonstrating that splenocytes, depleted of IgD + cells adoptively reconstitute this response in XID mice. The peritoneal B cell pool, which, compared to the spleen, consist of increased numbers of sIgM >> sIgD B cells, is shown to be a source of radiosensitive B cells that are TI-2 responsive. These observations represent additional evidence for an association between sIgM >> sIgD B cells and TI-2 responsiveness

  8. Src Family Kinases Regulate Interferon Regulatory Factor 1 K63 Ubiquitination following Activation by TLR7/8 Vaccine Adjuvant in Human Monocytes and B Cells

    Directory of Open Access Journals (Sweden)

    Lorenza Tulli

    2018-03-01

    Full Text Available Toll-like receptors (TLRs play a key role in the activation of innate immune cells, in which their engagement leads to production of cytokines and co-stimulatory molecules. TLRs signaling requires recruitment of toll/IL-1R (TIR domain-containing adaptors, such as MyD88 and/or TRIF, and leads to activation of several transcription factors, such as NF-κB, the AP1 complex, and various members of the interferon regulatory factor (IRF family, which in turn results in triggering of several cellular functions associated with these receptors. A role for Src family kinases (SFKs in this signaling pathway has also been established. Our work and that of others have shown that this type of kinases is activated following engagement of several TLRs, and that this event is essential for the initiation of specific downstream cellular response. In particular, we have previously demonstrated that activation of SFKs is required for balanced production of pro-inflammatory cytokines by monocyte-derived dendritic cells after stimulation with R848, an agonist of human TLRs 7/8. We also showed that TLR7/8 triggering leads to an increase in interferon regulatory factor 1 (IRF-1 protein levels and that this effect is abolished by inhibition of SFKs, suggesting a critical role of these kinases in IRF-1 regulation. In this study, we first confirmed the key role of SFKs in TLR7/8 signaling for cytokine production and accumulation of IRF-1 protein in monocytes and in B lymphocytes, two other type of antigen-presenting cells. Then, we demonstrate that TLR7 triggering leads to an increase of K63-linked ubiquitination of IRF-1, which is prevented by SFKs inhibition, suggesting a key role of these kinases in posttranslational regulation of IRF-1 in the immune cells. In order to understand the mechanism that links SFKs activation to IRF-1 K63-linked ubiquitination, we examined SFKs and IRF-1 possible interactors and proved that activation of SFKs is necessary for their

  9. CP-25, a Novel Anti-inflammatory and Immunomodulatory Drug, Inhibits the Functions of Activated Human B Cells through Regulating BAFF and TNF-alpha Signaling and Comparative Efficacy with Biological Agents

    Directory of Open Access Journals (Sweden)

    Feng Zhang

    2017-12-01

    Full Text Available Paeoniflorin-6′-O-benzene sulfonate (code: CP-25 was the chemistry structural modifications of Paeoniflorin (Pae. CP-25 inhibited B cells proliferation stimulated by B cell activating factor belonging to the TNF family (BAFF or Tumor necrosis factor alpha (TNF-alpha. CP-25, Rituximab and Etanercept reduced the percentage and numbers of CD19+ B cells, CD19+CD20+ B cells, CD19+CD27+ B cells and CD19+CD20+CD27+ B cells induced by BAFF or TNF-alpha. There was significant difference between CP-25 and Rituximab or CP-25 and Etanercept. CP-25 down-regulated the high expression of BAFFR, BCMA, and TACI stimulated by BAFF or TNF-alpha. The effects of Rituximab and Etanercept on BAFFR or BCMA were stronger than that of CP-25. CP-25, Rituximab and Etanercept down-regulated significantly the expression of TNFR1 and TNFR2 on B cell stimulated by BAFF or TNF-alpha. CP-25, Rituximab and Etanercept down-regulated the expression of MKK3, P-p38, P-p65, TRAF2, and p52 in B cells stimulated by BAFF and the expression of TRAF2 and P-p65 in B cells stimulated by TNF-alpha. These results suggest that CP-25 regulated moderately activated B cells function by regulating the classical and alternative NF-κB signaling pathway mediated by BAFF and TNF-alpha-TRAF2-NF-κB signaling pathway. This study suggests that CP-25 may be a promising anti-inflammatory immune and soft regulation drug.

  10. CP-25, a Novel Anti-inflammatory and Immunomodulatory Drug, Inhibits the Functions of Activated Human B Cells through Regulating BAFF and TNF-alpha Signaling and Comparative Efficacy with Biological Agents.

    Science.gov (United States)

    Zhang, Feng; Shu, Jin-Ling; Li, Ying; Wu, Yu-Jing; Zhang, Xian-Zheng; Han, Le; Tang, Xiao-Yu; Wang, Chen; Wang, Qing-Tong; Chen, Jing-Yu; Chang, Yan; Wu, Hua-Xun; Zhang, Ling-Ling; Wei, Wei

    2017-01-01

    Paeoniflorin-6'- O -benzene sulfonate (code: CP-25) was the chemistry structural modifications of Paeoniflorin (Pae). CP-25 inhibited B cells proliferation stimulated by B cell activating factor belonging to the TNF family (BAFF) or Tumor necrosis factor alpha (TNF-alpha). CP-25, Rituximab and Etanercept reduced the percentage and numbers of CD19 + B cells, CD19 + CD20 + B cells, CD19 + CD27 + B cells and CD19 + CD20 + CD27 + B cells induced by BAFF or TNF-alpha. There was significant difference between CP-25 and Rituximab or CP-25 and Etanercept. CP-25 down-regulated the high expression of BAFFR, BCMA, and TACI stimulated by BAFF or TNF-alpha. The effects of Rituximab and Etanercept on BAFFR or BCMA were stronger than that of CP-25. CP-25, Rituximab and Etanercept down-regulated significantly the expression of TNFR1 and TNFR2 on B cell stimulated by BAFF or TNF-alpha. CP-25, Rituximab and Etanercept down-regulated the expression of MKK3, P-p38, P-p65, TRAF2, and p52 in B cells stimulated by BAFF and the expression of TRAF2 and P-p65 in B cells stimulated by TNF-alpha. These results suggest that CP-25 regulated moderately activated B cells function by regulating the classical and alternative NF-κB signaling pathway mediated by BAFF and TNF-alpha-TRAF2-NF-κB signaling pathway. This study suggests that CP-25 may be a promising anti-inflammatory immune and soft regulation drug.

  11. Human rhabdomyosarcoma cell lines for rhabdomyosarcoma research: Utility and pitfalls

    Directory of Open Access Journals (Sweden)

    Ashley R.P. Hinson

    2013-07-01

    Full Text Available Rhabdomyosarcoma (RMS is the most common soft tissue sarcoma of childhood and adolescence. Despite intergroup clinical trials conducted in Europe and North America, outcomes for high risk patients with this disease have not significantly improved in the last several decades, and survival of metastatic or relapsed disease remains extremely poor. Accrual into new clinical trials is slow and difficult, so in vitro cell line research and in vivo xenograft models present an attractive alternative for preclinical research for this cancer type. Currently, 30 commonly used human RMS cell lines exist, with differing origins, karyotypes, histologies, and methods of validation. Selecting an appropriate cell line for RMS research has important implications for outcomes. There are also potential pitfalls in using certain cell lines including contamination with murine stromal cells, cross-contamination between cell lines, discordance between the cell line and its associated original tumor, imposter cell lines, and nomenclature errors that result in the circulation of two or more presumed unique cell lines that are actually from the same origin. These pitfalls can be avoided by testing for species-specific isoenzymes, microarray analysis, assays for subtype-specific fusion products, and short tandem repeat analysis.

  12. Human Rhabdomyosarcoma Cell Lines for Rhabdomyosarcoma Research: Utility and Pitfalls

    Science.gov (United States)

    Hinson, Ashley R. P.; Jones, Rosanne; Crose, Lisa E. S.; Belyea, Brian C.; Barr, Frederic G.; Linardic, Corinne M.

    2013-01-01

    Rhabdomyosarcoma (RMS) is the most common soft tissue sarcoma of childhood and adolescence. Despite intergroup clinical trials conducted in Europe and North America, outcomes for high risk patients with this disease have not significantly improved in the last several decades, and survival of metastatic or relapsed disease remains extremely poor. Accrual into new clinical trials is slow and difficult, so in vitro cell-line research and in vivo xenograft models present an attractive alternative for preclinical research for this cancer type. Currently, 30 commonly used human RMS cell lines exist, with differing origins, karyotypes, histologies, and methods of validation. Selecting an appropriate cell line for RMS research has important implications for outcomes. There are also potential pitfalls in using certain cell lines including contamination with murine stromal cells, cross-contamination between cell lines, discordance between the cell line and its associated original tumor, imposter cell lines, and nomenclature errors that result in the circulation of two or more presumed unique cell lines that are actually from the same origin. These pitfalls can be avoided by testing for species-specific isoenzymes, microarray analysis, assays for subtype-specific fusion products, and short tandem repeat analysis. PMID:23882450

  13. B-Cell Hematologic Malignancy Vaccination Registry

    Science.gov (United States)

    2017-12-29

    Monoclonal Gammopathy of Undetermined Significance; Multiple Myeloma; Waldenstrom Macroglobulinemia; Lymphocytosis; Lymphoma, Non-Hodgkin; B-Cell Chronic Lymphocytic Leukemia; Hematological Malignancies

  14. Genetic errors of the human caspase recruitment domain-B-cell lymphoma 10-mucosa-associated lymphoid tissue lymphoma-translocation gene 1 (CBM) complex: Molecular, immunologic, and clinical heterogeneity.

    Science.gov (United States)

    Pérez de Diego, Rebeca; Sánchez-Ramón, Silvia; López-Collazo, Eduardo; Martínez-Barricarte, Rubén; Cubillos-Zapata, Carolina; Ferreira Cerdán, Antonio; Casanova, Jean-Laurent; Puel, Anne

    2015-11-01

    Three members of the caspase recruitment domain (CARD) family of adaptors (CARD9, CARD10, and CARD11) are known to form heterotrimers with B-cell lymphoma 10 (BCL10) and mucosa-associated lymphoid tissue lymphoma-translocation gene 1 (MALT1). These 3 CARD-BCL10-MALT1 (CBM) complexes activate nuclear factor κB in both the innate and adaptive arms of immunity. Human inherited defects of the 3 components of the CBM complex, including the 2 adaptors CARD9 and CARD11 and the 2 core components BCL10 and MALT1, have recently been reported. Biallelic loss-of-function mutant alleles underlie several different immunologic and clinical phenotypes, which can be assigned to 2 distinct categories. Isolated invasive fungal infections of unclear cellular basis are associated with CARD9 deficiency, whereas a broad range of clinical manifestations, including those characteristic of T- and B-lymphocyte defects, are associated with CARD11, MALT1, and BCL10 deficiencies. Interestingly, human subjects with these mutations have some features in common with the corresponding knockout mice, but other features are different between human subjects and mice. Moreover, germline and somatic gain-of-function mutations of MALT1, BCL10, and CARD11 have also been found in patients with other lymphoproliferative disorders. This broad range of germline and somatic CBM lesions, including loss-of-function and gain-of-function mutations, highlights the contribution of each of the components of the CBM complex to human immunity. Copyright © 2015 American Academy of Allergy, Asthma & Immunology. Published by Elsevier Inc. All rights reserved.

  15. Chronic Lymphocytic Leukemia B-Cell Normal Cellular Counterpart: Clues From a Functional Perspective.

    Science.gov (United States)

    Darwiche, Walaa; Gubler, Brigitte; Marolleau, Jean-Pierre; Ghamlouch, Hussein

    2018-01-01

    Chronic lymphocytic leukemia (CLL) is characterized by the clonal expansion of small mature-looking CD19+ CD23+ CD5+ B-cells that accumulate in the blood, bone marrow, and lymphoid organs. To date, no consensus has been reached concerning the normal cellular counterpart of CLL B-cells and several B-cell types have been proposed. CLL B-cells have remarkable phenotypic and gene expression profile homogeneity. In recent years, the molecular and cellular biology of CLL has been enriched by seminal insights that are leading to a better understanding of the natural history of the disease. Immunophenotypic and molecular approaches (including immunoglobulin heavy-chain variable gene mutational status, transcriptional and epigenetic profiling) comparing the normal B-cell subset and CLL B-cells provide some new insights into the normal cellular counterpart. Functional characteristics (including activation requirements and propensity for plasma cell differentiation) of CLL B-cells have now been investigated for 50 years. B-cell subsets differ substantially in terms of their functional features. Analysis of shared functional characteristics may reveal similarities between normal B-cell subsets and CLL B-cells, allowing speculative assignment of a normal cellular counterpart for CLL B-cells. In this review, we summarize current data regarding peripheral B-cell differentiation and human B-cell subsets and suggest possibilities for a normal cellular counterpart based on the functional characteristics of CLL B-cells. However, a definitive normal cellular counterpart cannot be attributed on the basis of the available data. We discuss the functional characteristics required for a cell to be logically considered to be the normal counterpart of CLL B-cells.

  16. Apoptosis induction of epifriedelinol on human cervical cancer cell line

    African Journals Online (AJOL)

    Background: Present investigation evaluates the antitumor activity of epifriedelinol for the management of cervical cancer by inducing process of apoptosis. Methods: Human Cervical Cancer Cell Line, C33A and HeLa were selected for study and treated with epifriedelinol at a concentration of (50-1000 μg/ml). Cytotoxicity of ...

  17. 77 FR 5489 - Identification of Human Cell Lines Project

    Science.gov (United States)

    2012-02-03

    ... individual or species. With the advent of standardized, simple, and rapid methods for human cell line... project will undergo STR profiling, a DNA profiling method that examines/screens for STRs (DNA elements 2... distinct DNA profile and when the STR DNA fragment sizes are converted to numeric values, the DNA profiles...

  18. Radiation sensitivity of human lung cancer cell lines

    International Nuclear Information System (INIS)

    Carmichael, J.; Degraff, W.G.; Gamson, J.; Russo, G.; Mitchell, J.B.; Gazdar, A.F.; Minna, J.D.; Levitt, M.L.

    1989-01-01

    X-Ray survival curves were determined using a panel of 17 human lung cancer cell lines, with emphasis on non-small cell lung cancer (NSCLC). In contrast to classic small cell lung cancer (SCLC) cell lines, NSCLC cell lines were generally less sensitive to radiation as evidenced by higher radiation survival curve extrapolation numbers, surviving fraction values following a 2Gy dose (SF2) and the mean inactivation dose values (D) values. The spectrum of in vitro radiation responses observed was similar to that expected in clinical practice, although mesothelioma was unexpectedly sensitive in vitro. Differences in radiosensitivity were best distinguished by comparison of SF2 values. Some NSCLC lines were relatively sensitive, and in view of this demonstrable variability in radiation sensitivity, the SF2 value may be useful for in vitro predictive assay testing of clinical specimens. (author)

  19. Peroxisomal abnormalities in the immortalized human hepatocyte (IHH) cell line.

    Science.gov (United States)

    Klouwer, Femke C C; Koster, Janet; Ferdinandusse, Sacha; Waterham, Hans R

    2017-04-01

    The immortalized human hepatocyte (IHH) cell line is increasingly used for studies related to liver metabolism, including hepatic glucose, lipid, lipoprotein and triglyceride metabolism, and the effect of therapeutic interventions. To determine whether the IHH cell line is a good model to investigate hepatic peroxisomal metabolism, we measured several peroxisomal parameters in IHH cells and, for comparison, HepG2 cells and primary skin fibroblasts. This revealed a marked plasmalogen deficiency and a deficient fatty acid α-oxidation in the IHH cells, due to a defect of PEX7, a cytosolic receptor protein required for peroxisomal import of a subset of peroxisomal proteins. These abnormalities have consequences for the lipid homeostasis of these cells and thus should be taken into account for the interpretation of data previously generated by using this cell line and when considering using this cell line for future research.

  20. Radiation response of haematopoietic cell lines of human origin

    International Nuclear Information System (INIS)

    Lehnert, S.; Rybka, W.B.; Suissa, S.; Giambattisto, D.

    1986-01-01

    Six human haematopoietic cell lines, five of leukaemic origin, including cells with myeloid, lymphoid and undifferentiated phenotype have been studied with respect to radiation response. The intrinsic radio-sensitivity of the cells varied widely, the D 0 s ranging from 0.53 to 1.39 Gy. Five of the cell lines showed some capacity to accumulate sublethal damage; in three of these, enhanced survival was demonstrated in split-dose experiments. One cell line (HL-60) was anomalous in that although little accumulation of sublethal damage was demonstrable, survival was enhanced by fractionation of the dose. Five of the six cell lines studied were of leukaemic origin. The results support the belief that, in contrast to the almost constant radiosensitivity of normal haematopoietic cell progenitors, leukaemic cell progenitors may show a wide range of radiosensitivities. (author)

  1. Preclinical Evaluation of the Novel BTK Inhibitor Acalabrutinib in Canine Models of B-Cell Non-Hodgkin Lymphoma.

    Directory of Open Access Journals (Sweden)

    Bonnie K Harrington

    Full Text Available Acalabrutinib (ACP-196 is a second-generation inhibitor of Bruton agammaglobulinemia tyrosine kinase (BTK with increased target selectivity and potency compared to ibrutinib. In this study, we evaluated acalabrutinib in spontaneously occurring canine lymphoma, a model of B-cell malignancy similar to human diffuse large B-cell lymphoma (DLBCL. First, we demonstrated that acalabrutinib potently inhibited BTK activity and downstream effectors in CLBL1, a canine B-cell lymphoma cell line, and primary canine lymphoma cells. Acalabrutinib also inhibited proliferation in CLBL1 cells. Twenty dogs were enrolled in the clinical trial and treated with acalabrutinib at dosages of 2.5 to 20mg/kg every 12 or 24 hours. Acalabrutinib was generally well tolerated, with adverse events consisting primarily of grade 1 or 2 anorexia, weight loss, vomiting, diarrhea and lethargy. Overall response rate (ORR was 25% (5/20 with a median progression free survival (PFS of 22.5 days. Clinical benefit was observed in 30% (6/20 of dogs. These findings suggest that acalabrutinib is safe and exhibits activity in canine B-cell lymphoma patients and support the use of canine lymphoma as a relevant model for human non-Hodgkin lymphoma (NHL.

  2. A salvage chemotherapy of R-P-IMVP16/CBDCA consisting of rituximab, methylprednisolone, ifosfamide, methotrexate, etoposide, and carboplatin for patients with diffuse large B cell lymphoma who had previously received R-CHOP therapy as first-line chemotherapy.

    Science.gov (United States)

    Matsumoto, Takuro; Hara, Takeshi; Shibata, Yuhei; Nakamura, Nobuhiko; Nakamura, Hiroshi; Ninomiya, Soranobu; Kitagawa, Junichi; Kanemura, Nobuhiro; Goto, Naoe; Kito, Yusuke; Kasahara, Senji; Yamada, Toshiki; Sawada, Michio; Miyazaki, Tatsuhiko; Takami, Tsuyoshi; Takeuchi, Tamotsu; Moriwaki, Hisataka; Tsurumi, Hisashi

    2017-09-01

    We have reported the efficacy of the salvage chemotherapy P-IMVP16/CBDCA for patients with diffuse large B cell lymphoma (DLBCL) who had previously received CHOP before the availability of rituximab (R). Here, we confirmed the efficacy of R combined with P-IMVP16/CBDCA as a salvage chemotherapy for patients with DLBCL, who had previously received R-CHOP. We retrospectively analysed 59 patients with relapse or refractory DLBCL (38 male patients and 21 female patients) presenting between June 2004 and June 2013. The patients received R 375 mg/m 2 on day 1, methylprednisolone 1000 mg/body for 3 days (from day 3 to day 5), ifosfamide 1000 mg/m 2 for 5 days (from day 3 to day 7), methotrexate 30 mg/m 2 on day 5 and day 12, etoposide 80 mg/m 2 for 3 days (from day 3 to day 5), and carboplatin 300 mg/m 2 on day 3 every 21 days. Patients aged 70 years or older were given 75% of the standard dose. The overall response rate (complete response + partial response) was 64.4%. The 2-year overall survival rate was 55.3%. The 2-year progression free survival rate was 34.7%. The 2-year overall survival rate was 61.5% for the relapse patients, and 15.6% for the refractory patients (p effects were mild and tolerable. The R-P-IMVP-16/CBDCA regimen displayed a significant activity in relapsed DLBCL, with acceptable toxicity, and should be considered a candidate for salvage chemotherapy. Copyright © 2016 John Wiley & Sons, Ltd. Copyright © 2016 John Wiley & Sons, Ltd.

  3. Communication between B-Cells and Microbiota for the Maintenance of Intestinal Homeostasis

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    Yuying Liu

    2013-10-01

    Full Text Available The human intestine is populated with an extremely dense and diverse bacterial community. Commensal bacteria act as an important antigenic stimulus producing the maturation of gut-associated lymphoid tissue (GALT. The production of immunoglobulin (Ig A by B-cells in the GALT is one of the immune responses following intestinal colonization of bacteria. The switch of B-cells from IgM to IgA-producing cells in the Peyer’s patches and neighboring lamina propria proceeds by T-cell-dependent and T-cell-independent mechanisms. Several grams of secretory IgA (SIgA are released into the intestine each day. SIgA serves as a first-line of defense in protecting the intestinal epithelium from enteric toxins and pathogenic microorganisms. SIgA has a capacity to directly quench bacterial virulence factors, influence the composition of the intestinal microbiota, and promote the transportation of antigens across the intestinal epithelium to GALT and down-regulate proinflammatory responses associated with the uptake of highly pathogenic bacteria and potentially allergenic antigens. This review summarizes the reciprocal interactions between intestinal B cells and bacteria, specifically, the formation of IgA in the gut, the role of intestinal IgA in the regulation of bacterial communities and the maintenance of intestinal homeostasis, and the effects of probiotics on IgA levels in the gastrointestinal tract.

  4. Human embryonic stem cell lines model experimental human cytomegalovirus latency.

    Science.gov (United States)

    Penkert, Rhiannon R; Kalejta, Robert F

    2013-05-28

    Herpesviruses are highly successful pathogens that persist for the lifetime of their hosts primarily because of their ability to establish and maintain latent infections from which the virus is capable of productively reactivating. Human cytomegalovirus (HCMV), a betaherpesvirus, establishes latency in CD34(+) hematopoietic progenitor cells during natural infections in the body. Experimental infection of CD34(+) cells ex vivo has demonstrated that expression of the viral gene products that drive productive infection is silenced by an intrinsic immune defense mediated by Daxx and histone deacetylases through heterochromatinization of the viral genome during the establishment of latency. Additional mechanistic details about the establishment, let alone maintenance and reactivation, of HCMV latency remain scarce. This is partly due to the technical challenges of CD34(+) cell culture, most notably, the difficulty in preventing spontaneous differentiation that drives reactivation and renders them permissive for productive infection. Here we demonstrate that HCMV can establish, maintain, and reactivate in vitro from experimental latency in cultures of human embryonic stem cells (ESCs), for which spurious differentiation can be prevented or controlled. Furthermore, we show that known molecular aspects of HCMV latency are faithfully recapitulated in these cells. In total, we present ESCs as a novel, tractable model for studies of HCMV latency.

  5. Radioimmunotherapy of B-cell non-Hodgkin’s lymphoma

    Directory of Open Access Journals (Sweden)

    Caroline eBodet-Milin

    2013-07-01

    Full Text Available This manuscript reviews current advances in the use of radioimmunotherapy (RIT for the treatment of B-cell non-Hodgkin’s lymphoma (NHL. RIT has been in use for more than 20 years and has progressed significantly with the discovery of new molecular targets, the development of new stable chelates, the humanization of monoclonal antibodies (MAbs, and the use of pretargeting techniques. Today, two products targeting the CD20 antigen are approved: 131I-tositumomab, (Bexxar® and 90Y-ibritumomab tiuxetan, (Zevalin®. 131I-tositumomab is available in the United States, and 90Y-ibritumumab tiuxetan in Europe, the United States, Asia, and Africa. RIT can be integrated in clinical practice using non-ablative activities for treatment of patients with relapsed or refractory follicular lymphoma (FL or as consolidation after induction chemotherapy in front-line treatment in FL patients. Despite the lack of phase III studies to clearly define the efficacy of RIT in the management of B lymphoma in the era of rituximab-based therapy, RIT efficacy in NHL has been demonstrated. In relapsing refractory FL and transformed NHL, RIT as a monotherapy induces around 30% complete response with a possibility of durable remissions. RIT consolidation after induction therapy significantly improves the quality of the response. Dose-limiting toxicity of RIT is hematological, depending on bone marrow involvement and prior treatment. Non-hematological toxicity is generally low. Different studies have been published assessing innovative protocols of RIT or new indications, in particular treatment in patients with aggressive lymphomas. High-dose treatment, RIT as consolidation after different therapeutic induction modalities, RIT in first-line treatment or fractionated RIT showed promising results. New MAbs, in particular humanized MAbs, or combinations of naked and radiolabeled MAbs, also appear promising. Personalized dosimetry protocols should be developed to determine

  6. Generation and Characterization of an Immortalized Human Esophageal Myofibroblast Line.

    Directory of Open Access Journals (Sweden)

    Chao Niu

    Full Text Available Stromal cells with a myofibroblast phenotype present in the normal human esophagus are increased in individuals with gastro-esophageal reflux disease (GERD. We have previously demonstrated that myofibroblasts stimulated with acid and TLR4 agonists increase IL-6 and IL-8 secretion using primary cultures of myofibroblasts established from normal human esophagus. While primary cultures have the advantage of reflecting the in vivo environment, a short life span and unavoidable heterogeneity limits the usefulness of this model in larger scale in vitro cellular signaling studies. The major aim of this paper therefore was to generate a human esophageal myofibroblast line with an extended lifespan. In the work presented here we have generated and characterized an immortalized human esophageal myofibroblast line by transfection with a commercially available GFP-hTERT lentivirus. Immortalized human esophageal myofibroblasts demonstrate phenotypic, genotypic and functional similarity to primary cultures of esophageal myofibroblasts we have previously described. We found that immortalized esophageal myofibroblasts retain myofibroblast spindle-shaped morphology at low and high confluence beyond passage 80, and express α-SMA, vimentin, and CD90 myofibroblast markers. Immortalized human esophageal myofibroblasts also express the putative acid receptor TRPV1 and TLR4 and retain the functional capacity to respond to stimuli encountered in GERD with secretion of IL-6. Finally, immortalized human esophageal myofibroblasts also support the stratified growth of squamous esophageal epithelial cells in 3D organotypic cultures. This newly characterized immortalized human esophageal myofibroblast cell line can be used in future cellular signaling and co-culture studies.

  7. Type I CD20 Antibodies Recruit the B Cell Receptor for Complement-Dependent Lysis of Malignant B Cells

    NARCIS (Netherlands)

    Engelberts, Patrick J.; Voorhorst, Marleen; Schuurman, Janine; van Meerten, Tom; Bakker, Joost M.; Vink, Tom; Mackus, Wendy J. M.; Breij, Esther C. W.; Derer, Stefanie; Valerius, Thomas; van de Winkel, Jan G. J.; Parren, Paul W. H. I.; Beurskens, Frank J.

    2016-01-01

    Human IgG1 type I CD20 Abs, such as rituximab and ofatumumab (OFA), efficiently induce complement-dependent cytotoxicity (CDC) of CD20(+) B cells by binding of C1 to hexamerized Fc domains. Unexpectedly, we found that type I CD20 Ab F(ab ')2 fragments, as well as C1q-binding-deficient IgG mutants,

  8. B cells and B cell products-helping to restore cellular immunity?

    OpenAIRE

    Cascalho, Marilia; Platt, Jeffrey L.

    2006-01-01

    T cells that provide vital protection against tumors, viruses and intracellular bacteria are thought to develop independently of B cells. However, recent discoveries suggest that development of T cells depends on B cells. One way B cells promote T cell development is by providing diverse peptides that may promote positive selection of thymocytes. Diverse peptides and B cells help in diversification of the T cell receptor repertoire and may decrease cross-reactivity in the mature T cell compar...

  9. Understanding B-cell activation and autoantibody repertoire selection in systemic lupus erythematosus: A B-cell immunomics approach.

    Science.gov (United States)

    Tipton, Christopher M; Hom, Jennifer R; Fucile, Christopher F; Rosenberg, Alexander F; Sanz, Inaki

    2018-07-01

    Understanding antibody repertoires and in particular, the properties and fates of B cells expressing potentially pathogenic antibodies is critical to define the mechanisms underlying multiple immunological diseases including autoimmune and allergic conditions as well as transplant rejection. Moreover, an integrated knowledge of the antibody repertoires expressed by B cells and plasma cells (PC) of different functional properties and longevity is essential to develop new therapeutic strategies, better biomarkers for disease segmentation, and new assays to measure restoration of B-cell tolerance or, at least, of normal B-cell homeostasis. Reaching these goals, however, will require a more precise phenotypic, functional and molecular definition of B-cell and PC populations, and a comprehensive analysis of the antigenic reactivity of the antibodies they express. While traditionally hampered by technical and ethical limitations in human experimentation, new technological advances currently enable investigators to address these questions in a comprehensive fashion. In this review, we shall discuss these concepts as they apply to the study of Systemic Lupus Erythematosus. © 2018 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.

  10. Excessive amounts of mu heavy chain block B-cell development.

    Science.gov (United States)

    Zhu, Lingqiao; Chang, Cheong-Hee; Dunnick, Wesley

    2011-09-01

    Antigen-independent B-cell development occurs in several stages that depend on the expression of Ig heavy and light chain. We identified a line of mice that lacked mature B cells in the spleen. This mouse line carried approximately 11 copies of a transgene of the murine heavy chain constant region locus, and B-lineage cells expressed excessive amounts of the intracellular μ heavy chain. B-cell development failed in the bone marrow at the pro/pre B-cell transition, and examination of other lines with various copy numbers of the same transgene suggested that deficiencies in B-cell development increased with increased transgene copy number. Expression of a transgenic (Tg) light chain along with the Tg μ heavy chain led to minimal rescue of B-cell development in the bone marrow and B cells in the spleen. There are several potential mechanisms for the death of pro/pre B cells as a consequence of excess heavy chain expression.

  11. Epigenetic Impact on EBV Associated B-Cell Lymphomagenesis

    Directory of Open Access Journals (Sweden)

    Shatadru Ghosh Roy

    2016-11-01

    Full Text Available Epigenetic modifications leading to either transcriptional repression or activation, play an indispensable role in the development of human cancers. Epidemiological study revealed that approximately 20% of all human cancers are associated with tumor viruses. Epstein-Barr virus (EBV, the first human tumor virus, demonstrates frequent epigenetic alterations on both viral and host genomes in associated cancers—both of epithelial and lymphoid origin. The cell type-dependent different EBV latent gene expression patterns appear to be determined by the cellular epigenetic machinery and similarly viral oncoproteins recruit epigenetic regulators in order to deregulate the cellular gene expression profile resulting in several human cancers. This review elucidates the epigenetic consequences of EBV–host interactions during development of multiple EBV-induced B-cell lymphomas, which may lead to the discovery of novel therapeutic interventions against EBV-associated B-cell lymphomas by alteration of reversible patho-epigenetic markings.

  12. Bifenthrin activates homotypic aggregation in human T-cell lines.

    Science.gov (United States)

    Hoffman, Nataly; Tran, Van; Daniyan, Anthony; Ojugbele, Olutosin; Pryor, Stephen C; Bonventre, Josephine A; Flynn, Katherine; Weeks, Benjamin S

    2006-03-01

    Here, we addressed the concern that, despite the lack of overt toxicity, exposure to low levels of the common household pyrethroid pesticide, bifenthrin, could cause harm to the immune system. To do this, we measure the effect of bifenthrin on phytohemagglutinin (PHA) activation of homotypic aggregation in human T-cell lines. The human CD4+ H9, and Jurkat cell lines and the human promonocyte U937 cell line, were exposed to varying concentrations of bifenthrin. Cell viability was determined using the AlmarBlue Toxicity Assay. Concentrations of bifenthrin which did not reduce cell viability were determined and these concentrations were tested for the effect of bifenthrin on PHA-mediated homotypic aggregation. Blocking antibodies to ICAM and LFA-1 were used to disrupt aggregation and a nonspecific IgG was used as a control. Bifenthrin was found to be nontoxic at concentrations ranging from 10(-4) to 10(-13) M. Bifenthrin did not inhibit PHA induced cell aggregation in all cell lines tested. However, at 10(-4) M, bifenthrin to form aggregates stimulated homotypic aggregation in the H9 and Jurkat T-cell lines. The bifenthrin-induced aggregate formation, like that seen with PHA, was blocked by treating the cells with antibodies to either LFA-1 or ICAM. The results here show that bifenthrin activates T-cell function by stimulating ICAM/LFA-1 mediated homotypic aggregation. This data suggests that exposure to bifenthrin, even at "acceptable" limits, can increase the risk for and frequency of inflammatory responses and diseases such as asthma.

  13. EBI2 overexpression in mice leads to B1 B-cell expansion and chronic lymphocytic leukemia-like B-cell malignancies.

    Science.gov (United States)

    Niss Arfelt, Kristine; Barington, Line; Benned-Jensen, Tau; Kubale, Valentina; Kovalchuk, Alexander L; Daugvilaite, Viktorija; Christensen, Jan Pravsgaard; Thomsen, Allan Randrup; Egerod, Kristoffer L; Bassi, Maria R; Spiess, Katja; Schwartz, Thue W; Wang, Hongsheng; Morse, Herbert C; Holst, Peter J; Rosenkilde, Mette M

    2017-02-16

    Human and mouse chronic lymphocytic leukemia (CLL) develops from CD5 + B cells that in mice and macaques are known to define the distinct B1a B-cell lineage. B1a cells are characterized by lack of germinal center (GC) development, and the B1a cell population is increased in mice with reduced GC formation. As a major mediator of follicular B-cell migration, the G protein-coupled receptor Epstein-Barr virus-induced gene 2 ( EBI2 or GPR183 ) directs B-cell migration in the lymphoid follicles in response to its endogenous ligands, oxysterols. Thus, upregulation of EBI2 drives the B cells toward the extrafollicular area, whereas downregulation is essential for GC formation. We therefore speculated whether increased expression of EBI2 would lead to an expanded B1 cell subset and, ultimately, progression to CLL. Here, we demonstrate that B-cell-targeted expression of human EBI2 (hEBI2) in mice reduces GC-dependent immune responses, reduces total immunoglobulin M (IgM) and IgG levels, and leads to increased proliferation and upregulation of cellular oncogenes. Furthermore, hEBI2 overexpression leads to an abnormally expanded CD5 + B1a B-cell subset (present as early as 4 days after birth), late-onset lymphoid cancer development, and premature death. These findings are highly similar to those observed in CLL patients and identify EBI2 as a promoter of B-cell malignancies.

  14. B Cell Tolerance in Health and Disease

    Directory of Open Access Journals (Sweden)

    Murali Gururajan

    2014-02-01

    Full Text Available B lymphocyte receptors are generated randomly during the bone marrow developmental phase of B cells. Hence, the B cell repertoire consists of both self and foreign antigen specificities necessitating specific tolerance mechanisms to eliminate self-reactive B cells. This review summarizes the major mechanisms of B cell tolerance, which include clonal deletion, anergy and receptor editing. In the bone marrow presentation of antigen in membrane bound form is more effective than soluble form and the role of dendritic cells in this process is discussed. Toll like receptor derived signals affect activation of B cells by certain ligands such as nucleic acids and have been shown to play crucial roles in the development of autoimmunity in several animal models. In the periphery availability of BAFF, a B cell survival factor plays a critical role in the survival of self-reactive B cells. Antibodies against BAFF have been found to be effective therapeutic agents in lupus like autoimmune diseases. Recent developments are targeting anergy to control the growth of chronic lymphocytic leukemia cells.

  15. 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.

  16. Generation of memory B cells and their reactivation.

    Science.gov (United States)

    Inoue, Takeshi; Moran, Imogen; Shinnakasu, Ryo; Phan, Tri Giang; Kurosaki, Tomohiro

    2018-05-01

    The successful establishment of humoral memory response depends on at least two layers of defense. Pre-existing protective antibodies secreted by long-lived plasma cells act as a first line of defense against reinfection ("constitutive humoral memory"). Previously, a second line of defense in which pathogen-experienced memory B cells are rapidly reactivated to produce antibodies ("reactive humoral memory"), was considered as simply a back-up system for the first line (particularly for re-infection with homologous viruses). However, in the case of re-infection with similar but different strains of viruses, or in response to viral escape mutants, the reactive humoral memory plays a crucial role. Here, we review recent progress in our understanding of how memory B cells are generated in the pre-GC stage and during the GC reaction, and how these memory B cells are robustly reactivated with the help of memory Tfh cells to generate the secondary antibody response. In addition, we discuss how these advances may be relevant to the quest for a vaccine that can induce broadly reactive antibodies against influenza and HIV. © 2018 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.

  17. 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)

    Onnis, A; Navari, M; Antonicelli, G; Morettini, F; Mannucci, S; De Falco, G; Vigorito, E; Leoncini, L

    2012-01-01

    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

  18. Dipeptidyl peptidase IV in two human glioma cell lines

    Directory of Open Access Journals (Sweden)

    A Sedo

    2009-12-01

    Full Text Available There is growing evidence that dipeptidyl peptidase IV [DPP-IV, EC 3.4.14.5] takes part in the metabolism of biologically active peptides participating in the regulation of growth and transformation of glial cells. However, the knowledge on the DPP-IV expression in human glial and glioma cells is still very limited. In this study, using histochemical and biochemical techniques, the DPP-IV activity was demonstrated in two commercially available human glioma cell lines of different transformation degree, as represented by U373 astrocytoma (Grade III and U87 glioblastoma multiforme (Grade IV lines. Higher total activity of the enzyme, as well as its preferential localisation in the plasma membrane, was observed in U87 cells. Compared to U373 population, U87 cells were morphologically more pleiomorphic, they were cycling at lower rate and expressing less Glial Fibrillary Acidic Protein. The data revealed positive correlation between the degree of transformation of cells and activity of DPP-IV. Great difference in expression of this enzyme, together with the phenotypic differences of cells, makes these lines a suitable standard model for further 57 studies of function of this enzyme in human glioma cells.

  19. Ikaros limits follicular B cell activation by regulating B cell receptor signaling pathways

    International Nuclear Information System (INIS)

    Heizmann, Beate; Sellars, MacLean; Macias-Garcia, Alejandra; Chan, Susan; Kastner, Philippe

    2016-01-01

    The Ikaros transcription factor is essential for early B cell development, but its effect on mature B cells is debated. We show that Ikaros is required to limit the response of naive splenic B cells to B cell receptor signals. Ikaros deficient follicular B cells grow larger and enter cell cycle faster after anti-IgM stimulation. Unstimulated mutant B cells show deregulation of positive and negative regulators of signal transduction at the mRNA level, and constitutive phosphorylation of ERK, p38, SYK, BTK, AKT and LYN. Stimulation results in enhanced and prolonged ERK and p38 phosphorylation, followed by hyper-proliferation. Pharmacological inhibition of ERK and p38 abrogates the increased proliferative response of Ikaros deficient cells. These results suggest that Ikaros functions as a negative regulator of follicular B cell activation.

  20. Ikaros limits follicular B cell activation by regulating B cell receptor signaling pathways

    Energy Technology Data Exchange (ETDEWEB)

    Heizmann, Beate [Institut de Génétique et de Biologie Moléculaire et Cellulaire (IGBMC), INSERM U964, CNRS UMR 7104, Université de Strasbourg, 67404 Illkirch (France); Sellars, MacLean [Institut de Génétique et de Biologie Moléculaire et Cellulaire (IGBMC), INSERM U964, CNRS UMR 7104, Université de Strasbourg, 67404 Illkirch (France); David Geffen School of Medicine at UCLA, Los Angeles, CA 90095 (United States); Macias-Garcia, Alejandra [Institut de Génétique et de Biologie Moléculaire et Cellulaire (IGBMC), INSERM U964, CNRS UMR 7104, Université de Strasbourg, 67404 Illkirch (France); Institute for Medical Engineering and Science at MIT, Cambridge, MA 02139 (United States); Chan, Susan, E-mail: scpk@igbmc.fr [Institut de Génétique et de Biologie Moléculaire et Cellulaire (IGBMC), INSERM U964, CNRS UMR 7104, Université de Strasbourg, 67404 Illkirch (France); Kastner, Philippe, E-mail: scpk@igbmc.fr [Institut de Génétique et de Biologie Moléculaire et Cellulaire (IGBMC), INSERM U964, CNRS UMR 7104, Université de Strasbourg, 67404 Illkirch (France); Faculté de Médecine, Université de Strasbourg, Strasbourg (France)

    2016-02-12

    The Ikaros transcription factor is essential for early B cell development, but its effect on mature B cells is debated. We show that Ikaros is required to limit the response of naive splenic B cells to B cell receptor signals. Ikaros deficient follicular B cells grow larger and enter cell cycle faster after anti-IgM stimulation. Unstimulated mutant B cells show deregulation of positive and negative regulators of signal transduction at the mRNA level, and constitutive phosphorylation of ERK, p38, SYK, BTK, AKT and LYN. Stimulation results in enhanced and prolonged ERK and p38 phosphorylation, followed by hyper-proliferation. Pharmacological inhibition of ERK and p38 abrogates the increased proliferative response of Ikaros deficient cells. These results suggest that Ikaros functions as a negative regulator of follicular B cell activation.

  1. Sourcing human embryos for embryonic stem cell lines: Problems & perspectives

    Directory of Open Access Journals (Sweden)

    Rajvi H Mehta

    2014-01-01

    Full Text Available The ability to successfully derive human embryonic stem cells (hESC lines from human embryos following in vitro fertilization (IVF opened up a plethora of potential applications of this technique. These cell lines could have been successfully used to increase our understanding of human developmental biology, transplantation medicine and the emerging science of regenerative medicine. The main source for human embryos has been ′discarded′ or ′spare′ fresh or frozen human embryos following IVF. It is a common practice to stimulate the ovaries of women undergoing any of the assisted reproductive technologies (ART and retrieve multiple oocytes which subsequently lead to multiple embryos. Of these, only two or maximum of three embryos are transferred while the rest are cryopreserved as per the decision of the couple. In case a couple does not desire to ′cryopreserve′ their embryos then all the embryos remaining following embryo transfer can be considered ′spare′ or if a couple is no longer in need of the ′cryopreserved′ embryos then these also can be considered as ′spare′. But, the question raised by the ethicists is, "what about ′slightly′ over-stimulating a woman to get a few extra eggs and embryos? The decision becomes more difficult when it comes to ′discarded′ embryos. As of today, the quality of the embryos is primarily assessed based on morphology and the rate of development mainly judged by single point assessment. Despite many criteria described in the literature, the quality assessment is purely subjective. The question that arises is on the decision of ′discarding′ embryos. What would be the criteria for discarding embryos and the potential ′use′ of ESC derived from the ′abnormal appearing′ embryos? This paper discusses some of the newer methods to procure embryos for the derivation of embryonic stem cell lines which will respect the ethical concerns but still provide the source material.

  2. Type i CD20 antibodies recruit the B cell receptor for complement-dependent lysis of malignant B cells

    DEFF Research Database (Denmark)

    Engelberts, P. J.; Voorhorst, M.; Schuurman, J.

    2016-01-01

    . We hypothesized that CD20 Ab-induced clustering of the IgM or IgG BCR was involved in accessory CDC. Indeed, accessory CDC was consistently observed in B cell lines expressing an IgM BCR and in some cell lines expressing an IgG BCR, but it was absent in BCR- B cell lines. A direct relationship...... between BCR expression and accessory CDC was established by transfecting the BCR into CD20+ cells: OFA-F(ab')2 fragments were able to induce CDC in the CD20+BCR+ cell population, but not in the CD20+BCR- population. Importantly, OFA-F(ab')2 fragments were able to induce CDC ex vivo in malignant B cells...... isolated from patients with mantle cell lymphoma and Waldenström macroglobulinemia. In summary, accessory CDC represents a novel effector mechanism that is dependent on type I CD20 Ab-induced BCR clustering. Accessory CDC may contribute to the excellent capacity of type I CD20 Abs to induce CDC...

  3. Anti-HIV-1 B cell responses are dependent on B cell precursor frequency and antigen-binding affinity.

    Science.gov (United States)

    Dosenovic, Pia; Kara, Ervin E; Pettersson, Anna-Klara; McGuire, Andrew T; Gray, Matthew; Hartweger, Harald; Thientosapol, Eddy S; Stamatatos, Leonidas; Nussenzweig, Michel C

    2018-04-16

    The discovery that humans can produce potent broadly neutralizing antibodies (bNAbs) to several different epitopes on the HIV-1 spike has reinvigorated efforts to develop an antibody-based HIV-1 vaccine. Antibody cloning from single cells revealed that nearly all bNAbs show unusual features that could help explain why it has not been possible to elicit them by traditional vaccination and instead would require a sequence of different immunogens. This idea is supported by experiments with genetically modified immunoglobulin (Ig) knock-in mice. Sequential immunization with a series of specifically designed immunogens was required to shepherd the development of bNAbs. However, knock-in mice contain superphysiologic numbers of bNAb precursor-expressing B cells, and therefore how these results can be translated to a more physiologic setting remains to be determined. Here we make use of adoptive transfer experiments using knock-in B cells that carry a synthetic intermediate in the pathway to anti-HIV-1 bNAb development to examine how the relationship between B cell receptor affinity and precursor frequency affects germinal center (GC) B cell recruitment and clonal expansion. Immunization with soluble HIV-1 antigens can recruit bNAb precursor B cells to the GC when there are as few as 10 such cells per mouse. However, at low precursor frequencies, the extent of clonal expansion is directly proportional to the affinity of the antigen for the B cell receptor, and recruitment to GCs is variable and dependent on recirculation.

  4. Homotypic aggregation of human cell lines by HLA class II-, class Ia- and HLA-G-specific monoclonal antibodies

    DEFF Research Database (Denmark)

    Odum, Niels; Ledbetter, J A; Martin, P

    1991-01-01

    Major histocompatibility complex (MHC) class II molecules have been implicated in cell adhesion in two ways. In addition to the well-established role of class II antigens in low-affinity adhesion provided by interactions between class II and CD4, recent data indicated that class II may also induce...... adhesion between T and B cells by activating the CD18/CD11a (LFA-1) adhesion pathway. Here we report that monoclonal antibodies (mAb) against HLA-DR (L243, p4.1, HB10a, VI15) and certain broad class II reacting mAb (TU35, TU39), but not anti-DQ (TU22, Leu-10) mAb, induced homotypic aggregation of human...... class II-positive monocytic (I937) and T leukemic (HUT78) tumor cell lines and Epstein-Barr virus (EBV) transformed B-lymphoid cell lines (EBV-LCL). Class II-negative cell lines (U-937 and the EBV-LCL mutant line 616) were not induced to aggregate. An HLA-G-transfected EBV-LCL, 221-AGN...

  5. Lipid rafts and B cell signaling.

    Science.gov (United States)

    Gupta, Neetu; DeFranco, Anthony L

    2007-10-01

    B cells comprise an essential component of the humoral immune system. They are equipped with the unique ability to synthesize and secrete pathogen-neutralizing antibodies, and share with professional antigen presenting cells the ability to internalize foreign antigens, and process them for presentation to helper T cells. Recent evidence indicates that specialized cholesterol- and glycosphingolipid-rich microdomains in the plasma membrane commonly referred to as lipid rafts, serve to compartmentalize key signaling molecules during the different stages of B cell activation including B cell antigen receptor (BCR)-initiated signal transduction, endocytosis of BCR-antigen complexes, loading of antigenic peptides onto MHC class II molecules, MHC-II associated antigen presentation to helper T cells, and receipt of helper signals via the CD40 receptor. Here we review the recent literature arguing for a role of lipid rafts in the spatial organization of B cell function.

  6. B-cell waste classification sampling plan

    International Nuclear Information System (INIS)

    HOBART, R.L.

    1999-01-01

    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. B-cell stimulating factor

    Energy Technology Data Exchange (ETDEWEB)

    Clevenger, W R; Conlon, P J; Eisenman, J R; Gillis, S; Grabstein, K H; Hopp, T P; March, C J; Mochizuki, D Y; Price, V L; Shanebeck, K D

    1987-10-05

    BSF-1 was derived from malignant cells and purified by use of various techniques, including adsorption, ion exchange chromatography and reverse phase high performance liquid chromatography. By these techniques, the BSF-1 was purified to homogenity. The high purification of the BSF-1 has made possible the sequencing of the amino acid residues at the N-terminal portion of its protein molecules. From the amino acid sequencing information, a radiolabelled oligonucleotide probe corresponding to portion of the amino acid sequence of the BSF-1 molecule was synthesized and then used to probe a cDNA library prepared from polyadenylated mRNA extracted from cell lines known to produce BSF-1. Through this procedure, a cDNA clone containing the BSF-1 gene was isolated, sequenced and mature BSF-1 expressed. The isolated cDNA clone was then radiolabelled and used as a large probe for screening cDNA libraries of other species of animals for homologous BSF-1 clones.

  8. Derivation of novel genetically diverse human embryonic stem cell lines.

    Science.gov (United States)

    Stefanova, Valentina T; Grifo, James A; Hansis, Christoph

    2012-06-10

    Human embryonic stem cells (hESCs) have the potential to revolutionize many biomedical fields ranging from basic research to disease modeling, regenerative medicine, drug discovery, and toxicity testing. A multitude of hESC lines have been derived worldwide since the first 5 lines by Thomson et al. 13 years ago, but many of these are poorly characterized, unavailable, or do not represent desired traits, thus making them unsuitable for application purposes. In order to provide the scientific community with better options, we have derived 12 new hESC lines at New York University from discarded genetically normal and abnormal embryos using the latest techniques. We examined the genetic status of the NYUES lines in detail as well as their molecular and cellular features and DNA fingerprinting profile. Furthermore, we differentiated our hESCs into the tissues most affected by a specific condition or into clinically desired cell types. To our knowledge, a number of characteristics of our hESCs have not been previously reported, for example, mutation for alpha thalassemia X-linked mental retardation syndrome, linkage to conditions with a genetic component such as asthma or poor sperm morphology, and novel combinations of ethnic backgrounds. Importantly, all of our undifferentiated euploid female lines tested to date did not show X chromosome inactivation, believed to result in superior potency. We continue to derive new hESC lines and add them to the NIH registry and other registries. This should facilitate the use of our hESCs and lead to advancements for patient-benefitting applications.

  9. Tuft (caveolated) cells in two human colon carcinoma cell lines.

    Science.gov (United States)

    Barkla, D H; Whitehead, R H; Foster, H; Tutton, P J

    1988-09-01

    The presence of an unusual cell type in two human colon carcinoma cell lines is reported. The cells show the same morphology as "tuft" (caveolated) cells present in normal gastrointestinal epithelium. Tuft cells were seen in cell line LIM 1863 growing in vitro and in human colon carcinoma cell line LIM 2210 growing as subcutaneous solid tumour xenografts in nude mice. Characteristic morphologic features of tuft cells included a wide base, narrow apex and a tuft of long microvilli projecting from the apical surface. The microvilli are attached by a core of long microfilaments passing deep into the apical cytoplasm. Between the microvilli are parallel arrays of vesicles (caveoli) containing flocculent material. Two different but not mutually exclusive explanations for the presence of tuft cells are proposed. The first explanation is that tuft cells came from the resected tumour and have survived by mitotic division during subsequent passages. The second explanation suggests that tuft cells are the progeny of undifferentiated tumour cells. Descriptions of tuft cells in colon carcinomas are uncommon and possible reasons for this are presented. The morphology of tuft cells is consistent with that of a highly differentiated cell specialised for absorption, and these new models provide an opportunity to further investigate the structure and function of tuft cells.

  10. B Cells and B Cell Blasts Withstand Cryopreservation While Retaining Their Functionality for Producing Antibody

    Directory of Open Access Journals (Sweden)

    Philipp Fecher

    2018-05-01

    Full Text Available In individuals who have once developed humoral immunity to an infectious/foreign antigen, the antibodies present in their body can mediate instant protection when the antigen re-enters. Such antigen-specific antibodies can be readily detected in the serum. Long term humoral immunity is, however, also critically dependent on the ability of memory B cells to engage in a secondary antibody response upon re-exposure to the antigen. Antibody molecules in the body are short lived, having a half-life of weeks, while memory B cells have a life span of decades. Therefore, the presence of serum antibodies is not always a reliable indicator of B cell memory and comprehensive monitoring of humoral immunity requires that both serum antibodies and memory B cells be assessed. The prevailing view is that resting memory B cells and B cell blasts in peripheral blood mononuclear cells (PBMC cannot be cryopreserved without losing their antibody secreting function, and regulated high throughput immune monitoring of B cell immunity is therefore confined to—and largely limited by—the need to test freshly isolated PBMC. Using optimized protocols for freezing and thawing of PBMC, and four color ImmunoSpot® analysis for the simultaneous detection of all immunoglobulin classes/subclasses we show here that both resting memory B cells and B cell blasts retain their ability to secrete antibody after thawing, and thus demonstrate the feasibility of B cell immune monitoring using cryopreserved PBMC.

  11. B Cells and B Cell Blasts Withstand Cryopreservation While Retaining Their Functionality for Producing Antibody.

    Science.gov (United States)

    Fecher, Philipp; Caspell, Richard; Naeem, Villian; Karulin, Alexey Y; Kuerten, Stefanie; Lehmann, Paul V

    2018-05-31

    In individuals who have once developed humoral immunity to an infectious/foreign antigen, the antibodies present in their body can mediate instant protection when the antigen re-enters. Such antigen-specific antibodies can be readily detected in the serum. Long term humoral immunity is, however, also critically dependent on the ability of memory B cells to engage in a secondary antibody response upon re-exposure to the antigen. Antibody molecules in the body are short lived, having a half-life of weeks, while memory B cells have a life span of decades. Therefore, the presence of serum antibodies is not always a reliable indicator of B cell memory and comprehensive monitoring of humoral immunity requires that both serum antibodies and memory B cells be assessed. The prevailing view is that resting memory B cells and B cell blasts in peripheral blood mononuclear cells (PBMC) cannot be cryopreserved without losing their antibody secreting function, and regulated high throughput immune monitoring of B cell immunity is therefore confined to-and largely limited by-the need to test freshly isolated PBMC. Using optimized protocols for freezing and thawing of PBMC, and four color ImmunoSpot ® analysis for the simultaneous detection of all immunoglobulin classes/subclasses we show here that both resting memory B cells and B cell blasts retain their ability to secrete antibody after thawing, and thus demonstrate the feasibility of B cell immune monitoring using cryopreserved PBMC.

  12. B cells are multifunctional players in multiple sclerosis pathogenesis: insights from therapeutic interventions

    Directory of Open Access Journals (Sweden)

    Nele eClaes

    2015-12-01

    Full Text Available Multiple sclerosis (MS is a severe disease of the central nervous system (CNS characterized by autoimmune inflammation and neurodegeneration. Historically, damage to the CNS was thought to be mediated predominantly by activated pro-inflammatory T cells. B cell involvement in the pathogenesis of MS was solely attributed to autoantibody production. The first clues for the involvement of antibody-independent B cell functions in MS pathology came from positive results in clinical trials of the B cell depleting treatment rituximab in patients with relapsing-remitting (RR MS. The survival of antibody-secreting plasma cells and decrease in T cell numbers indicated the importance of other B cell functions in MS such as antigen presentation, costimulation and cytokine production. Rituximab provided us with an example of how clinical trials can lead to new research opportunities concerning B cell biology. Moreover, analysis of the antibody-independent B cell functions in MS has gained interest since these trials. Limited information is present on the effects of current immunomodulatory therapies on B cell functions, although effects of both first-line (interferon, glatiramer acetate, dimethyl fumarate and teriflunomide, second-line (fingolimod, natalizumab and even third-line (monoclonal antibody therapies treatments on B cell subtype distribution, expression of functional surface markers and secretion of different cytokines by B cells have been studied to some extent. In this review, we summarize the effects of different MS related treatments on B cell functions that have been described up to now in order to find new research opportunities and contribute to the understanding of the pathogenesis of MS.

  13. Yield of two mutant lines of soybean for human consumption

    International Nuclear Information System (INIS)

    Salmeron E, J.; Mastache L, A. A.; Diaz V, G. E.; Valencia E, F.; Ranfla C, R.; Melendez P, M.; Cervantes S, T.; De la Cruz T, E.; Garcia A, J. M.; Falcon B, T.

    2009-01-01

    The present work has the objective of to evaluate the yield and the agronomic behavior of 2 mutant lines of soybean for human consumption, obtained by means of a process of recurrent irradiation of soybean seed ISAAEG-BM 2 with gammas of Co 60 and selection in the generation R 4 M 18 . For the variable yield significant statistical differences were not observed, but considering the rest of the evaluated agronomic characteristics the mutant lines L 6 and Bombona they were excellent with values of 3,934.6 and 3,806.8 Kg ha- 1 to 15% of grain humidity, they also possess excellent genetic characteristics result of the irradiations and selections of these new genetic materials. (Author)

  14. Immunomodulatory effect of Mesenchymal Stem Cells on B cells

    Directory of Open Access Journals (Sweden)

    Marcella eFranquesa

    2012-07-01

    Full Text Available The research on T cell immunosuppression therapies has attracted most of the attention in clinical transplantation. However, B cells and humoral immune responses are increasingly acknowledged as crucial mediators of chronic allograft rejection. Indeed, humoral immune responses can lead to renal allograft rejection even in patients whose cell-mediated immune responses are well controlled. On the other hand, newly studied B cell subsets with regulatory effects have been linked to tolerance achievement in transplantation. Better understanding of the regulatory and effector B cell responses may therefore lead to new therapeutic approaches.Mesenchymal Stem Cells (MSC are arising as a potent therapeutic tool in transplantation due to their regenerative and immunomodulatory properties. The research on MSCs has mainly focused on their effects on T cells and although data regarding the modulatory effects of MSCs on alloantigen-specific humoral response in humans is scarce, it has been demonstrated that MSCs significantly affect B cell functioning. In the present review we will analyze and discuss the results in this field.

  15. Likelihood-Based Inference of B Cell Clonal Families.

    Directory of Open Access Journals (Sweden)

    Duncan K Ralph

    2016-10-01

    Full Text Available The human immune system depends on a highly diverse collection of antibody-making B cells. B cell receptor sequence diversity is generated by a random recombination process called "rearrangement" forming progenitor B cells, then a Darwinian process of lineage diversification and selection called "affinity maturation." The resulting receptors can be sequenced in high throughput for research and diagnostics. Such a collection of sequences contains a mixture of various lineages, each of which may be quite numerous, or may consist of only a single member. As a step to understanding the process and result of this diversification, one may wish to reconstruct lineage membership, i.e. to cluster sampled sequences according to which came from the same rearrangement events. We call this clustering problem "clonal family inference." In this paper we describe and validate a likelihood-based framework for clonal family inference based on a multi-hidden Markov Model (multi-HMM framework for B cell receptor sequences. We describe an agglomerative algorithm to find a maximum likelihood clustering, two approximate algorithms with various trade-offs of speed versus accuracy, and a third, fast algorithm for finding specific lineages. We show that under simulation these algorithms greatly improve upon existing clonal family inference methods, and that they also give significantly different clusters than previous methods when applied to two real data sets.

  16. Variations in Humanized and Defined Culture Conditions Supporting Derivation of New Human Embryonic Stem Cell Lines

    DEFF Research Database (Denmark)

    Fletcher, Judy M; Ferrier, Patricia M; Gardner, John O

    2006-01-01

    matrix substrate of purified human laminin (Ln) with transitional reliance on mitotically inactivated human fibroblast (HDF) feeder cells. With this integrated system hESC lines were isolated using either HDF conditioned medium supplemented with a bovine-sourced serum replacement (bSRM), or a defined...

  17. Functional analysis of the human calcyclin gene promoter in a panel of human melanoma cell lines

    NARCIS (Netherlands)

    van Groningen, J. J.; Weterman, M. A.; Swart, G. W.; Bloemers, H. P.

    1995-01-01

    By comparing two subsequent human tumor stages we previously described calcyclin as a new potential melanoma associated neoplastic progression marker positively linked with metastasis. In this study the calcyclin expression levels in a representative panel of human melanoma cell lines were

  18. Fibronectin synthesized by a human hepatoma cell line

    International Nuclear Information System (INIS)

    Glasgow, J.E.; Colman, R.W.

    1984-01-01

    Fibronectin is a family of immunologically similar glycoproteins which mediate a variety of cell-cell and cell-substratum interactions. It is a constituent of the extracellular matrix of connective tissue and circulates in plasma. When suspension and adherent cultures of a human hepatoma cell line (SK-HEP-1) were incubated in serum-free medium, the resulting conditioned medium contained material which was specifically immunoprecipitated by antisera to human plasma fibronectin. By double immunodiffusion, a component in the conditioned culture medium was shown to form a line of identity with fibronectin in human plasma and to migrate as an alpha 2- to beta-globulin during immunoelectrophoresis. Human fibronectin was quantified in conditioned medium by electroimmunodiffusion, and was found to increase for at least three days at about 0.1 micrograms/10(6) cells/day. Adherent cultures of SK-HEP-1 cells were incubated with L-[ 35 S]methionine to label newly synthesized proteins. Labeled fibronectin in conditioned medium or in cell extracts comigrated with fibronectin in human plasma as shown by autoradiography following crossed-immunoelectrophoresis. Fibronectin was demonstrated in the extra-cellular matrix of adherent SK-HEP-1 cultures by immunofluorescence. It was shown previously that SK-HEP-1 cells synthesize alpha 1-protease inhibitor, one of the products of normal hepatocytes. The finding that these hepatoma cells also synthesize fibronectin supports the concept that the hepatocyte may be one source of circulating fibronectin, a possibility consistent with the established role of this cell type in blood plasma protein synthesis

  19. Effect of IL-6 on proliferation and IG production of human EBV-transformed cell lines in serum free culture media

    NARCIS (Netherlands)

    Jochems, G. J.; Jordens, R.; van Lier, R. A.; Zeijlemaker, W. P.

    1990-01-01

    To optimalize growth and Ig production of EBV transformed B cells for large scale tissue culture, we analyzed five stable monoclonal EBV-B cell lines for their responsiveness to interleukin (IL)-6 in standard medium with 5% FCS and in several serum-free media. As we previously demonstrated these

  20. Two distinct affinity binding sites for IL-1 on human cell lines

    International Nuclear Information System (INIS)

    Bensimon, C.; Wakasugi, N.; Tagaya, Y.; Takakura, K.; Yodoi, J.; Tursz, T.; Wakasugi, H.

    1989-01-01

    We used two human cell lines, NK-like YT-C3 and an EBV-containing B cell line, 3B6, as models to study the receptor(s) for IL-1. Two distinct types of saturable binding sites were found on both cell lines at 37 degrees C. Between 1 pM and 100 pM of 125I-IL-1-alpha concentration, saturable binding sites were detected on the YT-C3 cells with a K of 4 x 10(-11) M. The K found for the IL-1-alpha binding sites on 3B6 cells was 7.5 x 10(-11) M. An additional binding curve was detected above 100 pM on YT-C3 cells with a K of 7 x 10(-9) M and on 3B6 cells with a K of 5 x 10(-9) M. Scatchard plot analysis revealed 600 sites/cell with high affinity binding and 7000 sites/cell with low affinity for YT-C3 cells and 300 sites/cell with high affinity binding and 6000 sites/cell with low affinity for 3B6 cells. At 37 degrees C, the internalization of 125I-labeled IL-1 occurred via both high and low affinity IL-1R on both YT-C3 and 3B6 cells, whereas the rates of internalization for high affinity binding sites on YT-C3 cells were predominant in comparison to that of low affinity binding sites. In chemical cross-linking studies of 125 I-IL-1-alpha to 3B6 and YT-C3 cells, two protein bands were immunoprecipitated with Mr around 85 to 90 kDa leading to an estimation of the Mr of the IL-1R around 68 to 72 kDa. In similar experiments, the Mr found for the IL-1R expressed on the murine T cell line EL4 was slightly higher (around 80 kDa). Whether these distinct affinity binding sites are shared by a single molecule or by various chains remains to be elucidated

  1. End-of-Treatment Positron Emission Tomography After Uniform First-Line Therapy of B-Cell Posttransplant Lymphoproliferative Disorder Identifies Patients at Low Risk of Relapse in the Prospective German PTLD Registry.

    Science.gov (United States)

    Zimmermann, Heiner; Denecke, Timm; Dreyling, Martin H; Franzius, Christiane; Reinke, Petra; Subklewe, Marion; Amthauer, Holger; Kneba, Michael; Riess, Hanno; Trappe, Ralf U

    2018-05-01

    Fluorine-18 fluorodeoxyglucose (18F-FDG)-positron emission tomography (PET) is a recommended standard in the staging and response assessment of 18F-FDG-avid lymphoma. Posttransplant lymphoproliferative disorder (PTLD) can be detected by 18F-FDG-PET at diagnosis with high sensitivity and specificity. However, the role of response assessment by end-of-treatment (EOT) PET has only been addressed in small case series. We performed a retrospective, multicenter study of 37 patients with CD20-positive PTLD after solid organ transplantation treated with uniform, up-to-date, first-line protocols in the prospective German PTLD registry who had received EOT 18F-FDG-PET between 2006 and 2014. Median follow-up was 5.0 years. Any nonphysiological 18F-FDG uptake (Deauville score greater 2) was interpreted as PET-positive. By computed tomography (CT) final staging, 18 of 37 patients had a complete response, 18 had a partial response and 1 patient had stable disease. EOT PET was negative in 24 of 37 patients and positive in 13 of 37 patients. The positive predictive value of EOT PET for PTLD relapse was 38%, and the negative predictive value was 92%. Time to progression (TTP) and progression-free-survival were significantly longer in the PET negative group (P = 0.019 and P = 0.013). In the 18 patients in a partial response by CT staging, we noted highly significant differences in overall survival (P = 0.001), time to progression (P = 0.007), and progression-free survival (P < 0.001) by EOT PET. Even without baseline imaging, EOT PET in PTLD identifies patients at low risk of relapse and offers clinically relevant information, particularly in patients in a partial remission by CT staging.

  2. Changes in Circulating B Cell Subsets Associated with Aging and Acute SIV Infection in Rhesus Macaques.

    Science.gov (United States)

    Chang, W L William; Gonzalez, Denise F; Kieu, Hung T; Castillo, Luis D; Messaoudi, Ilhem; Shen, Xiaoying; Tomaras, Georgia D; Shacklett, Barbara L; Barry, Peter A; Sparger, Ellen E

    2017-01-01

    Aging and certain viral infections can negatively impact humoral responses in humans. To further develop the nonhuman primate (NHP) model for investigating B cell dynamics in human aging and infectious disease, a flow cytometric panel was developed to characterize circulating rhesus B cell subsets. Significant differences between human and macaque B cells included the proportions of cells within IgD+ and switched memory populations and a prominent CD21-CD27+ unswitched memory population detected only in macaques. We then utilized the expanded panel to analyze B cell alterations associated with aging and acute simian immunodeficiency virus (SIV) infection in the NHP model. In the aging study, distinct patterns of B cell subset frequencies were observed for macaques aged one to five years compared to those between ages 5 and 30 years. In the SIV infection study, B cell frequencies and absolute number were dramatically reduced following acute infection, but recovered within four weeks of infection. Thereafter, the frequencies of activated memory B cells progressively increased; these were significantly correlated with the magnitude of SIV-specific IgG responses, and coincided with impaired maturation of anti-SIV antibody avidity, as previously reported for HIV-1 infection. These observations further validate the NHP model for investigation of mechanisms responsible for B cells alterations associated with immunosenescence and infectious disease.

  3. Culture of human cell lines by a pathogen-inactivated human platelet lysate.

    Science.gov (United States)

    Fazzina, R; Iudicone, P; Mariotti, A; Fioravanti, D; Procoli, A; Cicchetti, E; Scambia, G; Bonanno, G; Pierelli, L

    2016-08-01

    Alternatives to the use of fetal bovine serum (FBS) have been investigated to ensure xeno-free growth condition. In this study we evaluated the efficacy of human platelet lysate (PL) as a substitute of FBS for the in vitro culture of some human cell lines. PL was obtained by pools of pathogen inactivated human donor platelet (PLT) concentrates. Human leukemia cell lines (KG-1, K562, JURKAT, HL-60) and epithelial tumor cell lines (HeLa and MCF-7) were cultured with either FBS or PL. Changes in cell proliferation, viability, morphology, surface markers and cell cycle were evaluated for each cell line. Functional characteristics were analysed by drug sensitivity test and cytotoxicity assay. Our results demonstrated that PL can support growth and expansion of all cell lines, although the cells cultured in presence of PL experienced a less massive proliferation compared to those grown with FBS. We found a comparable percentage of viable specific marker-expressing cells in both conditions, confirming lineage fidelity in all cultures. Functionality assays showed that cells in both FBS- and PL-supported cultures maintained their normal responsiveness to adriamycin and NK cell-mediated lysis. Our findings indicate that PL is a feasible serum substitute for supporting growth and propagation of haematopoietic and epithelial cell lines with many advantages from a perspective of process standardization, ethicality and product safety.

  4. B cell depletion reduces T cell activation in pancreatic islets in a murine autoimmune diabetes model.

    Science.gov (United States)

    Da Rosa, Larissa C; Boldison, Joanne; De Leenheer, Evy; Davies, Joanne; Wen, Li; Wong, F Susan

    2018-06-01

    Type 1 diabetes is a T cell-mediated autoimmune disease characterised by the destruction of beta cells in the islets of Langerhans, resulting in deficient insulin production. B cell depletion therapy has proved successful in preventing diabetes and restoring euglycaemia in animal models of diabetes, as well as in preserving beta cell function in clinical trials in the short term. We aimed to report a full characterisation of B cell kinetics post B cell depletion, with a focus on pancreatic islets. Transgenic NOD mice with a human CD20 transgene expressed on B cells were injected with an anti-CD20 depleting antibody. B cells were analysed using multivariable flow cytometry. There was a 10 week delay in the onset of diabetes when comparing control and experimental groups, although the final difference in the diabetes incidence, following prolonged observation, was not statistically significant (p = 0.07). The co-stimulatory molecules CD80 and CD86 were reduced on stimulation of B cells during B cell depletion and repopulation. IL-10-producing regulatory B cells were not induced in repopulated B cells in the periphery, post anti-CD20 depletion. However, the early depletion of B cells had a marked effect on T cells in the local islet infiltrate. We demonstrated a lack of T cell activation, specifically with reduced CD44 expression and effector function, including IFN-γ production from both CD4 + and CD8 + T cells. These CD8 + T cells remained altered in the pancreatic islets long after B cell depletion and repopulation. Our findings suggest that B cell depletion can have an impact on T cell regulation, inducing a durable effect that is present long after repopulation. We suggest that this local effect of reducing autoimmune T cell activity contributes to delay in the onset of autoimmune diabetes.

  5. Human comment dynamics in on-line social systems

    Science.gov (United States)

    Wu, Ye; Zhou, Changsong; Chen, Maoying; Xiao, Jinghua; Kurths, Jürgen

    2010-12-01

    Human comment is studied using data from ‘tianya’ which is one of the most popular on-line social systems in China. We found that the time interval between two consecutive comments on the same topic, called inter-event time, follows a power-law distribution. This result shows that there is no characteristic decay time on a topic. It allows for very long periods without comments that separate bursts of intensive comments. Furthermore, the frequency of a different ID commenting on a topic also follows a power-law distribution. It indicates that there are some “hubs” in the topic who lead the direction of the public opinion. Based on the personal comments habit, a model is introduced to explain these phenomena. The numerical simulations of the model fit well with the empirical results. Our findings are helpful for discovering regular patterns of human behavior in on-line society and the evolution of the public opinion on the virtual as well as real society.

  6. Effects of hypoxia on human cancer cell line chemosensitivity

    Science.gov (United States)

    2013-01-01

    Background Environment inside even a small tumor is characterized by total (anoxia) or partial oxygen deprivation, (hypoxia). It has been shown that radiotherapy and some conventional chemotherapies may be less effective in hypoxia, and therefore it is important to investigate how different drugs act in different microenvironments. In this study we perform a large screening of the effects of 19 clinically used or experimental chemotherapeutic drugs on five different cell lines in conditions of normoxia, hypoxia and anoxia. Methods A panel of 19 commercially available drugs: 5-fluorouracil, acriflavine, bortezomib, cisplatin, digitoxin, digoxin, docetaxel, doxorubicin, etoposide, gemcitabine, irinotecan, melphalan, mitomycin c, rapamycin, sorafenib, thalidomide, tirapazamine, topotecan and vincristine were tested for cytotoxic activity on the cancer cell lines A2780 (ovarian), ACHN (renal), MCF-7 (breast), H69 (SCLC) and U-937 (lymphoma). Parallel aliquots of the cells were grown at different oxygen pressures and after 72 hours of drug exposure viability was measured with the fluorometric microculture cytotoxicity assay (FMCA). Results Sorafenib, irinotecan and docetaxel were in general more effective in an oxygenated environment, while cisplatin, mitomycin c and tirapazamine were more effective in a low oxygen environment. Surprisingly, hypoxia in H69 and MCF-7 cells mostly rendered higher drug sensitivity. In contrast ACHN appeared more sensitive to hypoxia, giving slower proliferating cells, and consequently, was more resistant to most drugs. Conclusions A panel of standard cytotoxic agents was tested against five different human cancer cell lines cultivated at normoxic, hypoxic and anoxic conditions. Results show that impaired chemosensitivity is not universal, in contrast different cell lines behave different and some drugs appear even less effective in normoxia than hypoxia. PMID:23829203

  7. EBI2 overexpression in mice leads to B1 B cell expansion and chronic lymphocytic leukemia-(CLL)-like B cell malignancies

    DEFF Research Database (Denmark)

    Niss Arfelt, Kristine; Barington, Line; Benned-Jensen, Tau

    2017-01-01

    -targeted expression of human EBI2 in mice reduces germinal center-dependent immune responses, reduces total IgM and IgG levels, and leads to increased proliferation and upregulation of cellular oncogenes. Furthermore, hEBI2 overexpression leads to an abnormally expanded CD5+ B1a B cell subset present as early as 4......Human and mouse chronic lymphocytic leukemia (CLL) develop from CD5+ B cells that in mice and macaques are known to define the distinct B1a B cell lineage. B1a cells are characterized by lack of germinal center development and the B1a cell population is increased in mice with reduced germinal...... cells towards the extrafollicular area, whereas downregulation is essential for germinal center formation. We therefore speculated whether increased expression of EBI2 would lead to an expanded B1 cell subset and, ultimately, progression to chronic lymphocytic leukemia. Here we demonstrate that B cell...

  8. The targeted inhibition of mitochondrial Hsp90 overcomes the apoptosis resistance conferred by Bcl-2 in Hep3B cells via necroptosis

    Energy Technology Data Exchange (ETDEWEB)

    Yan, Chunlan [Department of Anatomy and Cell Biology, Dong-A University College of Medicine and Mitochondria Hub Regulation Center, Busan, 602-714 (Korea, Republic of); Department of Physiology, Zhejiang University School of Medicine, Hangzhou, Zhejiang 310058 (China); Oh, Joon Seok; Yoo, Seung Hee; Lee, Jee Suk [Department of Anatomy and Cell Biology, Dong-A University College of Medicine and Mitochondria Hub Regulation Center, Busan, 602-714 (Korea, Republic of); Yoon, Young Geol [Department of Anatomy and Cell Biology, Dong-A University College of Medicine and Mitochondria Hub Regulation Center, Busan, 602-714 (Korea, Republic of); Department of Biomedical Science, Institute for Biomedical and Health Sciences, Jungwon University, Chungbuk, 367-805 (Korea, Republic of); Oh, Yoo Jin; Jang, Min Seok [Department of Anatomy and Cell Biology, Dong-A University College of Medicine and Mitochondria Hub Regulation Center, Busan, 602-714 (Korea, Republic of); Lee, Sang Yeob [Department of Rheumatology, Dong-A University College of Medicine, Busan, 602-714 (Korea, Republic of); Yang, Jun [Department of Toxicology, Hangzhou Normal University School of Public Health, Hangzhou, Zhejiang, 310036 China (China); Lee, Sang Hwa [Department of Microbiology and, Dong-A University College of Medicine, Busan, 602-714 (Korea, Republic of); Kim, Hye Young [Department of Anatomy and Cell Biology, Dong-A University College of Medicine and Mitochondria Hub Regulation Center, Busan, 602-714 (Korea, Republic of); Yoo, Young Hyun, E-mail: yhyoo@dau.ac.kr [Department of Anatomy and Cell Biology, Dong-A University College of Medicine and Mitochondria Hub Regulation Center, Busan, 602-714 (Korea, Republic of)

    2013-01-01

    Previous studies have reported that a Gamitrinib variant containing triphenylphosphonium (G-TPP) binds to mitochondrial Hsp90 and rapidly inhibits its activity, thus inducing the apoptotic pathway in the cells. Accordingly, G-TPP shows a potential as a promising drug for the treatment of cancer. A cell can die from different types of cell death such as apoptosis, necrosis, necroptosis, and autophagic cell death. In this study, we further investigated the mechanisms and modes of cell death in the G-TPP-treated Hep3B and U937 cell lines. We discovered that G-TPP kills the U937 cells through the apoptotic pathway and the overexpression of Bcl-2 significantly inhibits U937 cell death to G-TPP. We further discovered that G-TPP kills the Hep3B cells by activating necroptosis in combination with the partial activation of caspase-dependent apoptosis. Importantly, G-TPP overcomes the apoptosis resistance conferred by Bcl-2 in Hep3B cells via necroptosis. We also observed that G-TPP induces compensatory autophagy in the Hep3B cell line. We further found that whereas there is a Bcl-2-Beclin 1 interaction in response to G-TPP, silencing the beclin 1 gene failed to block LC3-II accumulation in the Hep3B cells, indicating that G-TPP triggers Beclin 1-independent protective autophagy in Hep3B cells. Taken together, these data reveal that G-TPP induces cell death through a combination of death pathways, including necroptosis and apoptosis, and overcomes the apoptosis resistance conferred by Bcl-2 in Hep3B cells via necroptosis. These findings are important for the therapeutic exploitation of necroptosis as an alternative cell death program to bypass the resistance to apoptosis. Highlights: ► G-TPP binds to mitochondrial Hsp90. ► G-TPP induces apoptosis in U937 human leukemia cancer cells. ► G-TPP induces combination of death pathways in Hep3B cell. ► G-TPP overcomes the resistance conferred by Bcl-2 in Hep3B cells via necroptosis. ► G-TPP triggers Beclin 1-independent

  9. CD3-positive B cells: a storage-dependent phenomenon.

    Directory of Open Access Journals (Sweden)

    Angela Nagel

    Full Text Available The majority of clinical studies requires extensive management of human specimen including e.g. overnight shipping of blood samples in order to convey the samples in a central laboratory or to simultaneously analyze large numbers of patients. Storage of blood samples for periods of time before in vitro/ex vivo testing is known to influence the antigen expression on the surface of lymphocytes. In this context, the present results show for the first time that the T cell antigen CD3 can be substantially detected on the surface of human B cells after ex vivo storage and that the degree of this phenomenon critically depends on temperature and duration after blood withdrawal. The appearance of CD3 on the B cell surface seems to be a result of contact-dependent antigen exchange between T and B lymphocytes and is not attributed to endogenous production by B cells. Since cellular subsets are often classified by phenotypic analyses, our results indicate that ex vivo cellular classification in peripheral blood might result in misleading interpretations. Therefore, in order to obtain results reflecting the in vivo situation, it is suggested to minimize times of ex vivo blood storage after isolation of PBMC. Moreover, to enable reproducibility of results between different research groups and multicenter studies, we would emphasize the necessity to specify and standardize the storage conditions, which might be the basis of particular findings.

  10. Effects of cholera toxin on human colon carcinoma cell lines.

    Science.gov (United States)

    Barkla, D H; Whitehead, R H; Hayward, I P

    1992-10-01

    This study reports on changes in morphology and membrane transport in 5 human colon carcinoma cell lines treated with cholera toxin (CT). Three of the cell lines that grew as monolayers (LIM 1215, LIM 1899, LIM 2099) and 1 that grew as floating clumps (LIM 2408) did not show morphological changes after CT treatment. However, cell line LIM 1863 that grows as floating "crypt-like" organoids showed rapid and distinctive changes in morphology and membrane transport after CT treatment. At 1 and 6 hrs after CT treatment, light and transmission electron microscopy revealed rapid dilatation of the central lumen of organoids and the appearance of 2 populations of apical vesicular inclusions. The first population was unusual in being non-membrane bound and limited by fuzzy filamentous material. The second population was membrane bound. Scanning electron microscopy at 1-6 hr after CT treatment showed swelling and loss of surface microvilli on some, but not all, cells. At 24 hr after CT treatment the majority of organoids showed evidence of fluid accumulation and small apical vesicles coalesced to form large single vacuoles that obliterated normal cell morphology. By 48 hr, continued swelling produced extreme attenuation of the plasma membrane with cells taking on an "endothelial cell-like" appearance. The response to CT was dose-dependent. Uptake studies using 86Rubidium and blocking studies using ouabain and amiloride indicated that CT is acting on the Na+/K+ ATPase membrane pump to cause the increased fluid uptake by LIM 1863 cells. This study is the first to report specific morphological changes in intestine-derived cells in response to CT.(ABSTRACT TRUNCATED AT 250 WORDS)

  11. B cell subset distribution is altered in patients with severe periodontitis.

    Science.gov (United States)

    Demoersman, Julien; Pochard, Pierre; Framery, Camille; Simon, Quentin; Boisramé, Sylvie; Soueidan, Assem; Pers, Jacques-Olivier

    2018-01-01

    Several studies have recently highlighted the implication of B cells in physiopathogenesis of periodontal disease by showing that a B cell deficiency leads to improved periodontal parameters. However, the detailed profiles of circulating B cell subsets have not yet been investigated in patients with severe periodontitis (SP). We hypothesised that an abnormal distribution of B cell subsets could be detected in the blood of patients with severe periodontal lesions, as already reported for patients with chronic inflammatory diseases as systemic autoimmune diseases. Fifteen subjects with SP and 13 subjects without periodontitis, according to the definition proposed by the CDC periodontal disease surveillance work group, were enrolled in this pilot observational study. Two flow cytometry panels were designed to analyse the circulating B and B1 cell subset distribution in association with the RANKL expression. A significantly higher percentage of CD27+ memory B cells was observed in patients with SP. Among these CD27+ B cells, the proportion of the switched memory subset was significantly higher. At the same time, human B1 cells, which were previously associated with a regulatory function (CD20+CD69-CD43+CD27+CD11b+), decreased in SP patients. The RANKL expression increased in every B cell subset from the SP patients and was significantly greater in activated B cells than in the subjects without periodontitis. These preliminary results demonstrate the altered distribution of B cells in the context of severe periodontitis. Further investigations with a larger cohort of patients can elucidate if the analysis of the B cell compartment distribution can reflect the periodontal disease activity and be a reliable marker for its prognosis (clinical trial registration number: NCT02833285, B cell functions in periodontitis).

  12. B cell subset distribution is altered in patients with severe periodontitis

    Science.gov (United States)

    Demoersman, Julien; Pochard, Pierre; Framery, Camille; Simon, Quentin; Boisramé, Sylvie; Soueidan, Assem

    2018-01-01

    Several studies have recently highlighted the implication of B cells in physiopathogenesis of periodontal disease by showing that a B cell deficiency leads to improved periodontal parameters. However, the detailed profiles of circulating B cell subsets have not yet been investigated in patients with severe periodontitis (SP). We hypothesised that an abnormal distribution of B cell subsets could be detected in the blood of patients with severe periodontal lesions, as already reported for patients with chronic inflammatory diseases as systemic autoimmune diseases. Fifteen subjects with SP and 13 subjects without periodontitis, according to the definition proposed by the CDC periodontal disease surveillance work group, were enrolled in this pilot observational study. Two flow cytometry panels were designed to analyse the circulating B and B1 cell subset distribution in association with the RANKL expression. A significantly higher percentage of CD27+ memory B cells was observed in patients with SP. Among these CD27+ B cells, the proportion of the switched memory subset was significantly higher. At the same time, human B1 cells, which were previously associated with a regulatory function (CD20+CD69-CD43+CD27+CD11b+), decreased in SP patients. The RANKL expression increased in every B cell subset from the SP patients and was significantly greater in activated B cells than in the subjects without periodontitis. These preliminary results demonstrate the altered distribution of B cells in the context of severe periodontitis. Further investigations with a larger cohort of patients can elucidate if the analysis of the B cell compartment distribution can reflect the periodontal disease activity and be a reliable marker for its prognosis (clinical trial registration number: NCT02833285, B cell functions in periodontitis). PMID:29447240

  13. Expression of HLA Class II Molecules in Humanized NOD.Rag1KO.IL2RgcKO Mice is Critical for Development and Function of Human T and B Cells

    Science.gov (United States)

    2011-05-17

    class switch was preserved in DRAG mice. The human identity of Ig heavy and Figure 4. Human T cells developed by DRAG mice are functional. At 6 months...HSC-infused DRAG and control mice were immunized with 1 flocculation unit of TT vaccine (Sanofi Pasteur ) by the intramuscular route, and the titers...and CD8 T cells with preserved integrity of both TCR and non-TCR signaling pathways. It has been suggested that low frequency and functional

  14. Non-invasive bioluminescence imaging to monitor the immunological control of a plasmablastic lymphoma-like B cell neoplasia after hematopoietic cell transplantation.

    Directory of Open Access Journals (Sweden)

    Martin Chopra

    Full Text Available To promote cancer research and to develop innovative therapies, refined pre-clinical mouse tumor models that mimic the actual disease in humans are of dire need. A number of neoplasms along the B cell lineage are commonly initiated by a translocation recombining c-myc with the immunoglobulin heavy-chain gene locus. The translocation is modeled in the C.129S1-Igha(tm1(MycJanz/J mouse which has been previously engineered to express c-myc under the control of the endogenous IgH promoter. This transgenic mouse exhibits B cell hyperplasia and develops diverse B cell tumors. We have isolated tumor cells from the spleen of a C.129S1-Igha(tm1(MycJanz/J mouse that spontaneously developed a plasmablastic lymphoma-like disease. These cells were cultured, transduced to express eGFP and firefly luciferase, and gave rise to a highly aggressive, transplantable B cell lymphoma cell line, termed IM380. This model bears several advantages over other models as it is genetically induced and mimics the translocation that is detectable in a number of human B cell lymphomas. The growth of the tumor cells, their dissemination, and response to treatment within immunocompetent hosts can be imaged non-invasively in vivo due to their expression of firefly luciferase. IM380 cells are radioresistant in vivo and mice with established tumors can be allogeneically transplanted to analyze graft-versus-tumor effects of transplanted T cells. Allogeneic hematopoietic stem cell transplantation of tumor-bearing mice results in prolonged survival. These traits make the IM380 model very valuable for the study of B cell lymphoma pathophysiology and for the development of innovative cancer therapies.

  15. B cell markers in Ph1-positive acute lymphoblastic leukemia.

    Science.gov (United States)

    Alimena, G; De Rossi, G; Gastaldi, R; Guglielmi, C; Mandelli, F

    1980-01-01

    A case of acute lymphoblastic leukemia (ALL) where the blast cells had B cell markers and displayed the presence of a typical Ph1 chromosome, originated by a standard t (9;22) translocation, is reported. Cytological and clinical aspects during the entire course of the disease were consistent with the diagnosis of ALL. Evidence of differentiation along a well-defined lymphoid cell line in a Ph1-positive cell confirms the presence of the Ph1 chromosome in conditions other than chronic granulocytic leukemia and shows that it possibly does not occur in an exclusively undifferentiated totipotent stem cell.

  16. Double-hit B-cell lymphomas

    NARCIS (Netherlands)

    Aukema, Sietse M.; Siebert, Reiner; Schuuring, Ed; van Imhoff, Gustaaf W.; Kluin-Nelemans, Hanneke C.; Boerma, Evert-Jan; Kluin, Philip M.

    2011-01-01

    In many B-cell lymphomas, chromosomal translocations are biologic and diagnostic hallmarks of disease. An intriguing subset is formed by the so-called double-hit (DH) lymphomas that are defined by a chromosomal breakpoint affecting the MYC/8q24 locus in combination with another recurrent breakpoint,

  17. Rituximab in the treatment of primary cutaneous B-cell lymphoma: a review.

    Science.gov (United States)

    Fernández-Guarino, M; Ortiz-Romero, P L; Fernández-Misa, R; Montalbán, C

    2014-06-01

    Rituximab is a chimeric mouse-human antibody that targets the CD20 antigen, which is found in both normal and neoplastic B cells. In recent years, it has been increasingly used to treat cutaneous B-cell lymphoma and is now considered an alternative to classic treatment (radiotherapy and surgery) of 2 types of indolent lymphoma, namely, primary cutaneous follicle center lymphoma and primary cutaneous marginal zone B-cell lymphoma. Rituximab is also administered as an alternative to polychemotherapy in the treatment of primary cutaneous large B-cell lymphoma, leg type. Its use as an alternative drug led to it being administered intralesionally, with beneficial effects. In the present article, we review the literature published on the use of rituximab to treat primary cutaneous B-cell lymphoma. Copyright © 2012 Elsevier España, S.L. and AEDV. All rights reserved.

  18. Translocation of the B cell receptor to lipid rafts is inhibited in B cells from BLV-infected, persistent lymphocytosis cattle

    International Nuclear Information System (INIS)

    Hamilton, Valerie T.; Stone, Diana M.; Cantor, Glenn H.

    2003-01-01

    Bovine leukemia virus (BLV) infection causes a significant polyclonal expansion of CD5 + , IgM+ B lymphocytes known as persistent lymphocytosis (PL) in approximately 30% of infected cattle. There is evidence that this expanded B cell population has altered signaling, and resistance to apoptosis has been proposed as one mechanism of B cell expansion. In human and murine B cells, antigen binding initiates movement of the B cell receptor (BCR) into membrane microdomains enriched in sphingolipids and cholesterol, termed lipid rafts. Lipid rafts include members of the Src-family kinases and exclude certain phosphatases. Inclusion of the BCR into lipid rafts plays an important role in regulation of early signaling events and subsequent antigen internalization. Viral proteins may also influence signaling events in lipid rafts. Here we demonstrate that the largely CD5 + B cell population in PL cattle has different mobilization and internalization of the BCR when compared to the largely CD5-negative B cells in BLV-negative cattle. Unlike B cells from BLV-negative cattle, the BCR in B cells of BLV-infected, PL cattle resists movement into lipid rafts upon stimulation and is only weakly internalized. Expression of viral proteins as determined by detection of the BLV transmembrane (TM) envelope glycoprotein gp30 did not alter these events in cells from PL cattle. This exclusion of the BCR from lipid rafts may, in part, explain signaling differences seen between B cells of BLV-infected, PL, and BLV-negative cattle and the resistance to apoptosis speculated to contribute to persistent lymphocytosis

  19. TRPM4 expression is associated with activated B cell subtype and poor survival in diffuse large B cell lymphoma

    DEFF Research Database (Denmark)

    Loo, Suet K; Ch'ng, Ewe S; Md Salleh, Md Salzihan

    2017-01-01

    to investigate TRPM4 protein expression pattern in non-malignant tissues and DLBCL cases, and its association with clinico-demographic parameters and survival in DLBCL. METHODS AND RESULTS: Analysis of publicly available DLBCL microarray data sets showed that TRPM4 transcripts were up-regulated in DLBCL compared...... to normal germinal centre B (GCB) cells, were expressed more highly in the activated B cell-like DLBCL (ABC-DLBCL) subtype and higher TRPM4 transcripts conferred worse overall survival (OS) in R-CHOP (rituximab, cyclophosphamide, doxorubicin, vincristine and prednisone)-treated DLBCL cases (P ... immunohistochemical analysis showed that TRPM4 was expressed in various human tissues but not in normal B cells within lymphoid tissues (reactive tonsil, lymph node and appendix). TRPM4 protein was present in 26% (n = 49 of 189) of our cohort of R-CHOP-treated DLBCL cases and this was associated significantly...

  20. Characterization of Antigen-Specific B Cells Using Nominal Antigen-Coated Flow-Beads

    Science.gov (United States)

    Akl, Ahmed; Lepetit, Maud; Crochette, Romain; Giral, Magali; Lepourry, Julie; Pallier, Annaick; Castagnet, Stéphanie; Dugast, Emilie; Guillot-Gueguen, Cécile; Jacq-Foucher, Marylène; Saulquin, Xavier; Cesbron, Anne; Laplaud, David; Nicot, Arnaud; Brouard, Sophie; Soulillou, Jean-Paul

    2013-01-01

    In order to characterize the reactivity of B cells against nominal antigens, a method based on the coupling of antigens onto the surface of fluorescent core polystyrene beads was developed. We first demonstrate that murine B cells with a human MOG-specific BCR are able to interact with MOG-coated beads and do not recognize beads coated with human albumin or pp65. B cells purified from human healthy volunteer blood or immunized individuals were tested for their ability to interact with various nominal antigens, including viral, vaccine, self and alloantigens, chosen for their usefulness in studying a variety of pathological processes. A substantial amount of B cells binding self-antigen MOG-coated beads can be detected in normal blood. Furthermore, greater frequencies of B cell against anti-Tetanic Toxin or anti-EBNA1 were observed in primed individuals. This method can reveal increased frequencies of anti-HLA committed B cells in patients with circulating anti-HLA antibodies compared to unsensitized patients and normal individuals. Of interest, those specific CD19 cells were preferentially identified within CD27−IgD+ (i-e naïve) subset. These observations suggest that a broad range of medical situations could benefit from a tool that allows the detection, the quantification and the characterization of antigen-specific blood B cells. PMID:24386360

  1. Inactivated Tianjin strain, a novel genotype of Sendai virus, induces apoptosis in HeLa, NCI-H446 and Hep3B cells.

    Science.gov (United States)

    Chen, Jun; Han, Han; Wang, Bin; Shi, Liying

    2016-07-01

    The Sendai virus strain Tianjin is a novel genotype of the Sendai virus. In previous studies, ultraviolet-inactivated Sendai virus strain Tianjin (UV-Tianjin) demonstrated antitumor effects on human breast cancer cells. The aim of the present study was to investigate the in vitro antitumor effects of UV-Tianjin on the human cervical carcinoma HeLa, human small cell lung cancer NCI-H446 and human hepatocellular carcinoma Hep 3B cell lines, and the possible underlying mechanisms of these antitumor effects. A 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl tetrazolium bromide assay revealed that UV-Tianjin treatment inhibited the proliferation of HeLa, NCI-H446 and Hep 3B cells in a dose- and time-dependent manner. Hoechst and Annexin V-fluorescein isothiocyanate/propidium iodide double staining indicated that UV-Tianjin induced dose-dependent apoptosis in all three cell lines with the most significant effect observed in the HeLa cell line. In the HeLa cell line, UV-Tianjin-induced apoptosis was further confirmed by the disruption of the mitochondria membrane potential and the activation of caspases, as demonstrated by fluorescent cationic dye and colorimetric assays, respectively. In addition, western blot analysis revealed that UV-Tianjin treatment resulted in significant upregulation of cytochrome c , apoptosis protease activating factor-1, Fas, Fas ligand and Fas-associated protein with death domain, and activated caspase-9, -8 and -3 in HeLa cells. Based on these results, it is hypothesized that UV-Tianjin exhibits anticancer activity in HeLa, NCI-H446 and Hep 3B cell lines via the induction of apoptosis. In conclusion, the results of the present study indicate that in the HeLa cell line, intrinsic and extrinsic apoptotic pathways may be involved in UV-Tianjin-induced apoptosis.

  2. Chronic Exposure to Malaria Is Associated with Inhibitory and Activation Markers on Atypical Memory B Cells and Marginal Zone-Like B Cells

    Directory of Open Access Journals (Sweden)

    Itziar Ubillos

    2017-08-01

    Full Text Available In persistent infections that are accompanied by chronic immune activation, such as human immunodeficiency virus, hepatitis C virus, and malaria, there is an increased frequency of a phenotypically distinct subset of memory B cells lacking the classic memory marker CD27 and showing a reduced capacity to produce antibodies. However, critical knowledge gaps remain on specific B cell changes and immune adaptation in chronic infections. We hypothesized that expansion of atypical memory B cells (aMBCs and reduction of activated peripheral marginal zone (MZ-like B cells in constantly exposed individuals might be accompanied by phenotypic changes that would confer a tolerogenic profile, helping to establish tolerance to infections. To better understand malaria-associated phenotypic abnormalities on B cells, we analyzed peripheral blood mononuclear cells from 55 pregnant women living in a malaria-endemic area of Papua Nueva Guinea and 9 Spanish malaria-naïve individuals using four 11-color flow cytometry panels. We assessed the expression of markers of B cell specificity (IgG and IgM, activation (CD40, CD80, CD86, b220, TACI, and CD150, inhibition (PD1, CD95, and CD71, and migration (CCR3, CXCR3, and CD62l. We found higher frequencies of active and resting aMBC and marked reduction of MZ-like B cells, although changes in absolute cell counts could not be assessed. Highly exposed women had higher PD1+-, CD95+-, CD40+-, CD71+-, and CD80+-activated aMBC frequencies than non-exposed subjects. Malaria exposure increased frequencies of b220 and proapoptotic markers PD1 and CD95, and decreased expression of the activation marker TACI on MZ-like B cells. The increased frequencies of inhibitory and apoptotic markers on activated aMBCs and MZ-like B cells in malaria-exposed adults suggest an immune-homeostatic mechanism for maintaining B cell development and function while simultaneously downregulating hyperreactive B cells. This mechanism would keep the B cell

  3. Selective Depletion of αβ T Cells and B Cells for Human Leukocyte Antigen-Haploidentical Hematopoietic Stem Cell Transplantation. A Three-Year Follow-Up of Procedure Efficiency.

    Science.gov (United States)

    Li Pira, Giuseppina; Malaspina, David; Girolami, Elia; Biagini, Simone; Cicchetti, Elisabetta; Conflitti, Gianpiero; Broglia, Manuel; Ceccarelli, Stefano; Lazzaro, Stefania; Pagliara, Daria; Meschini, Antonella; Bertaina, Alice; Montanari, Mauro; Locatelli, Franco

    2016-11-01

    HLA-haploidentical family donors represent a valuable option for children requiring allogeneic hematopoietic stem cell transplantation (HSCT). Because graft-versus-host diseases (GVHD) is a major complication of HLA-haploidentical HSCT because of alloreactive T cells in the graft, different methods have been used for ex vivo T cell depletion. Removal of donor αβ T cells, the subset responsible for GVHD, and of B cells, responsible for post-transplantation lymphoproliferative disorders, have been recently developed for HLA-haploidentical HSCT. This manipulation preserves, in addition to CD34 +  progenitors, natural killer, γδ T, and monocytes/dendritic cells, contributing to anti-leukemia activity and protection against infections. We analyzed depletion efficiency and cell yield in 200 procedures performed in the last 3 years at our center. Donors underwent CD34 +   hematopoietic stem cell (HSC) peripheral blood mobilization with granulocyte colony-stimulating factor (G-CSF). Poor CD34 +  cell mobilizers (48 of 189, 25%) received plerixafor in addition to G-CSF. Aphereses containing a median of 52.5 × 10 9 nucleated cells and 494 × 10 6 CD34 +  HSC were manipulated using the CliniMACS device. In comparison to the initial product, αβ T cell depletion produced a median 4.1-log reduction (range, 3.1 to 5.5) and B cell depletion led to a median 3.4-log reduction (range, 2.0 to 4.7). Graft products contained a median of 18.5 × 10 6 CD34 +  HSC/kg recipient body weight, with median values of residual αβ T cells and B cells of 29 × 10 3 /kg and 33 × 10 3 /kg, respectively. Depletion efficiency monitored at 6-month intervals demonstrated steady performance, while improved recovery of CD34 +  cells was observed after the first year (P = .0005). These data indicate that αβ T cell and B cell depletion of HSC grafts from HLA-haploidentical donors was efficient and reproducible. Copyright © 2016 The American Society for Blood and Marrow

  4. Transplantability of human lymphoid cell line, lymphoma, and leukemia in splenectomized and/or irradiated nude mice

    International Nuclear Information System (INIS)

    Watanabe, S.; Shimosato, Y.; Kuroki, M.; Sato, Y.; Nakajima, T.

    1980-01-01

    The effects of splenectomy and/or whole-body irradiation of nude mice before xenotransplantation of lymphoid cell lines, lymphoma, and leukemia were studied. Transplantation after whole-body irradiation resulted in the increased ''take'' rate of three cultured cell lines (two of T-cell-derived acute lymphocytic leukemia and one of B-cell derived acute lymphocytic leukemia) and in the tumorous growth of Burkitt-derived Raji and spontaneously transformed lymphoblastoid cell lines. With splenectomy plus irradiation as a pretreatment, tumorous growth occurred in four other cell lines which were not transplantable after irradiation only (two cell lines of Epstein-Barr virus-transformed cord blood cells and one each of null acute lymphocytic leukemia and nodular lymphoma-derived cell lines). Direct transplantation of leukemia and lymphoma cells into the pretreated mice was successful in 7 of 24 cases (29%). B-cell-derived diffuse large lymphoid lymphoma was transplantable in three of seven cases (43%). However, lymphoma and leukemia of peripheral T-cell origin was difficult to transplant even with pretreatment, and only one pleomorphic T-cell lymphoma grew to a significant size (2 cm). One tumor each of B-cell-derived diffuse large lymphoid and T-cell diffuse lymphoblastic lymphoma became transplantable

  5. Rituximab does not reset defective early B cell tolerance checkpoints

    Science.gov (United States)

    Chamberlain, Nicolas; Massad, Christopher; Oe, Tyler; Cantaert, Tineke; Herold, Kevan C.; Meffre, Eric

    2015-01-01

    Type 1 diabetes (T1D) patients show abnormalities in early B cell tolerance checkpoints, resulting in the accumulation of large numbers of autoreactive B cells in their blood. Treatment with rituximab, an anti-CD20 mAb that depletes B cells, has been shown to preserve β cell function in T1D patients and improve other autoimmune diseases, including rheumatoid arthritis and multiple sclerosis. However, it remains largely unknown how anti–B cell therapy thwarts autoimmunity in these pathologies. Here, we analyzed the reactivity of Abs expressed by single, mature naive B cells from 4 patients with T1D before and 52 weeks after treatment to determine whether rituximab resets early B cell tolerance checkpoints. We found that anti–B cell therapy did not alter the frequencies of autoreactive and polyreactive B cells, which remained elevated in the blood of all patients after rituximab treatment. Moreover, the limited proliferative history of autoreactive B cells after treatment revealed that these clones were newly generated B cells and not self-reactive B cells that had escaped depletion and repopulated the periphery through homeostatic expansion. We conclude that anti–B cell therapy may provide a temporary dampening of autoimmune processes through B cell depletion. However, repletion with autoreactive B cells may explain the relapse that occurs in many autoimmune patients after anti–B cell therapy. PMID:26642366

  6. Inhibition of Oncogenic Transcription Factor REL by the Natural Product Derivative Calafianin Monomer 101 Induces Proliferation Arrest and Apoptosis in Human B-Lymphoma Cell Lines.

    Science.gov (United States)

    Yeo, Alan T; Chennamadhavuni, Spandan; Whitty, Adrian; Porco, John A; Gilmore, Thomas D

    2015-04-23

    Increased activity of transcription factor NF-κB has been implicated in many B-cell lymphomas. We investigated effects of synthetic compound calafianin monomer (CM101) on biochemical and biological properties of NF-κB. In human 293 cells, CM101 selectively inhibited DNA binding by overexpressed NF-κB subunits REL (human c-Rel) and p65 as compared to NF-κB p50, and inhibition of REL and p65 DNA binding by CM101 required a conserved cysteine residue. CM101 also inhibited DNA binding by REL in human B-lymphoma cell lines, and the sensitivity of several B-lymphoma cell lines to CM101-induced proliferation arrest and apoptosis correlated with levels of cellular and nuclear REL. CM101 treatment induced both phosphorylation and decreased expression of anti-apoptotic protein Bcl-XL, a REL target gene product, in sensitive B-lymphoma cell lines. Ectopic expression of Bcl-XL protected SUDHL-2 B-lymphoma cells against CM101-induced apoptosis, and overexpression of a transforming mutant of REL decreased the sensitivity of BJAB B-lymphoma cells to CM101-induced apoptosis. Lipopolysaccharide-induced activation of NF-κB signaling upstream components occurred in RAW264.7 macrophages at CM101 concentrations that blocked NF-κB DNA binding. Direct inhibitors of REL may be useful for treating B-cell lymphomas in which REL is active, and may inhibit B-lymphoma cell growth at doses that do not affect some immune-related responses in normal cells.

  7. B cell activation by outer membrane vesicles--a novel virulence mechanism.

    Directory of Open Access Journals (Sweden)

    Maria Laura A Perez Vidakovics

    2010-01-01

    Full Text Available Secretion of outer membrane vesicles (OMV is an intriguing phenomenon of Gram-negative bacteria and has been suggested to play a role as virulence factors. The respiratory pathogens Moraxella catarrhalis reside in tonsils adjacent to B cells, and we have previously shown that M. catarrhalis induce a T cell independent B cell response by the immunoglobulin (Ig D-binding superantigen MID. Here we demonstrate that Moraxella are endocytosed and killed by human tonsillar B cells, whereas OMV have the potential to interact and activate B cells leading to bacterial rescue. The B cell response induced by OMV begins with IgD B cell receptor (BCR clustering and Ca(2+ mobilization followed by BCR internalization. In addition to IgD BCR, TLR9 and TLR2 were found to colocalize in lipid raft motifs after exposure to OMV. Two components of the OMV, i.e., MID and unmethylated CpG-DNA motifs, were found to be critical for B cell activation. OMV containing MID bound to and activated tonsillar CD19(+ IgD(+ lymphocytes resulting in IL-6 and IgM production in addition to increased surface marker density (HLA-DR, CD45, CD64, and CD86, whereas MID-deficient OMV failed to induce B cell activation. DNA associated with OMV induced full B cell activation by signaling through TLR9. Importantly, this concept was verified in vivo, as OMV equipped with MID and DNA were found in a 9-year old patient suffering from Moraxella sinusitis. In conclusion, Moraxella avoid direct interaction with host B cells by redirecting the adaptive humoral immune response using its superantigen-bearing OMV as decoys.

  8. Primary Hepatosplenic Large B-Cell Lymphoma

    Directory of Open Access Journals (Sweden)

    M.R. Morales-Polanco

    2008-03-01

    Full Text Available Diffuse large B-cell lymphoma is the most common form of lymphoma. It usually begins in the lymph nodes; up to 40% may have an extranodal presentation. According to a definition of primary extranodal lymphoma with presentation only in extranodal sites, there are reports of large B-cell lymphomas limited to liver or spleen as separate entities, and to date there have been only three documented cases of primary hepatosplenic presentation. This paper reports a fourth case. Due to a review of the literature and the clinical course of the case reported, we conclude that primary hepatosplenic large B-cell lymphoma has been found predominantly in females older than 60 years. The patients reported had <2 months of evolution prior to diagnosis, prominent B symptoms, splenomegaly in three and hepatomegaly in two, none with lymph node involvement. All had thrombocytopenia and abnormal liver function tests; three had anemia and elevated serum lactic dehydrogenase levels, two with hemophagocytosis in bone marrow. Because of the previously mentioned data, it can be stated that primary hepatosplenic lymphoma is an uncommon and aggressive form of disease that requires immediate recognition and treatment.

  9. Modulation of B-cell receptor and microenvironment signaling by a guanine exchange factor in B-cell malignancies

    International Nuclear Information System (INIS)

    Liao, Wei; Sharma, Sanjai

    2016-01-01

    Objective: Chronic lymphocytic leukemia (CLL) and mantle cell lymphoma (MCL) cells over-express a guanine exchange factor (GEF), Rasgrf-1. This GEF increases active Ras as it catalyzes the removal of GDP from Ras so that GTP can bind and activate Ras. This study aims to study the mechanism of action of Rasgrf-1 in B-cell malignancies. Methods: N-terminus truncated Rasgrf-1 variants have a higher GEF activity as compared to the full-length transcript therefore a MCL cell line with stable over-expression of truncated Rasgrf-1 was established. The B-cell receptor (BCR) and chemokine signaling pathways were compared in the Rasgrf-1 over-expressing and a control transfected cell line. Results: Cells over-expressing truncated form of Rasgrf-1 have a higher proliferative rate as compared to control transfected cells. BCR was activated by lower concentrations of anti-IgM antibody in Rasgrf-1 over-expressing cells as compared to control cells indicating that these cells are more sensitive to BCR signaling. BCR signaling also phosphorylates Rasgrf-1 that further increases its GEF function and amplifies BCR signaling. This activation of Rasgrf-1 in over-expressing cells resulted in a higher expression of phospho-ERK, AKT, BTK and PKC-alpha as compared to control cells. Besides BCR, Rasgrf-1 over-expressing cells were also more sensitive to microenvironment stimuli as determined by resistance to apoptosis, chemotaxis and ERK pathway activation. Conclusions: This GEF protein sensitizes B-cells to BCR and chemokine mediated signaling and also upregulates a number of other signaling pathways which promotes growth and survival of these cells

  10. 76 FR 16609 - Proposed Information Collection; Comment Request; Identification of Human Cell Lines Project

    Science.gov (United States)

    2011-03-24

    ... differentiate among cell lines, as described in Designation: ASN-0002 Authentication of Human Cell Lines... NIST (contact information above). III. Data OMB Control Number: None. Form Number: None. Type of Review...

  11. Characterization of a novel telomerase-immortalized human endometrial stromal cell line, St-T1b

    Directory of Open Access Journals (Sweden)

    Brosens Jan J

    2009-07-01

    Full Text Available Abstract Background Coordinated differentiation of the endometrial compartments in the second half of the menstrual cycle is a prerequisite for the establishment of pregnancy. Endometrial stromal cells (ESC decidualize under the influence of ovarian progesterone to accommodate implantation of the blastocyst and support establishment of the placenta. Studies into the mechanisms of decidualization are often hampered by the lack of primary ESC. Here we describe a novel immortalized human ESC line. Methods Primary ESC were immortalized by the transduction of telomerase. The resultant cell line, termed St-T1b, was characterized for its morphological and biochemical properties by immunocytochemistry, RT-PCR and immunoblotting. Its progestational response was tested using progesterone and medroxyprogesterone acetate with and without 8-Br-cAMP, an established inducer of decidualization in vitro. Results St-T1b were positive for the fibroblast markers vimentin and CD90 and negative for the epithelial marker cytokeratin-7. They acquired a decidual phenotype indistinguishable from primary ESC in response to cAMP stimulation. The decidual response was characterized by transcriptional activation of marker genes, such as PRL, IGFBP1, and FOXO1, and enhanced protein levels of the tumor suppressor p53 and the metastasis suppressor KAI1 (CD82. Progestins alone had no effect on St-T1b cells, but medroxyprogesterone acetate greatly enhanced the cAMP-stimulated expression of IGFBP-1 after 3 and 7 days. Progesterone, albeit more weakly, also augmented the cAMP-induced IGFBP-1 production but only after 7 days of treatment. The cell line remained stable in continuous culture for more than 150 passages. Conclusion St-T1b express the appropriate phenotypic ESC markers and their decidual response closely mimics that of primary cultures. Decidualization is efficiently induced by cAMP analog and enhanced by medroxyprogesterone acetate, and, to a lesser extent, by natural

  12. Aging-dependent decline of IL-10 producing B cells coincides with production of antinuclear antibodies but not rheumatoid factors.

    Science.gov (United States)

    van der Geest, Kornelis S M; Lorencetti, Pedro G; Abdulahad, Wayel H; Horst, Gerda; Huitema, Minke; Roozendaal, Caroline; Kroesen, Bart-Jan; Brouwer, Elisabeth; Boots, Annemieke M H

    2016-03-01

    Aging is associated with development of autoimmunity. Loss of B cell tolerance in the elderly is suggested by an increased prevalence of anti-nuclear antibodies (ANAs) and rheumatoid factors (RFs). Accumulating evidence indicates that B cells also impact autoimmunity via secretion of cytokines. So far, few studies have directly assessed the effect of aging on the latter B cell function. Here, we determined if and how human aging influences the production of cytokines by B cells. In a cross-sectional study, we found that absolute numbers of circulating B cells were similar in 31 young (ages 19-39) and 73 old (age ≥ 60) individuals. Numbers of transitional B cells (CD19(+)CD27(-)CD38(High)CD24(High)) were decreased in old individuals, whereas numbers of naive and memory B cell subsets were comparable in young and old individuals. Short-term in vitro stimulation of whole blood samples revealed that numbers of B cells capable of producing TNF-α were similar in young and old individuals. In contrast, B cells capable of IL-10 production were decreased in old subjects. This decline of IL-10(+) B cells was observed in old individuals that were ANA positive, and in those that were negative for both ANAs and RFs. However, IL-10(+) B cells were remarkably well retained in the circulation of old subjects that were RF positive. Thus, pro-inflammatory TNF-α(+) B cells are retained in the elderly, whereas IL-10(+) B cells generally decline. In addition, our findings indicate that IL-10(+) B cells may differentially impact the development of ANAs and RFs in the elderly. Copyright © 2015 Elsevier Inc. All rights reserved.

  13. Derivation and characterization of the NIH registry human stem cell line NYSCF100 line under defined feeder-free conditions

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    Ana Sevilla

    2018-05-01

    Full Text Available The human embryonic stem cell line NYSCFe001-A was derived from a day 6 blastocyst in feeder-free and antibiotic free conditions. The blastocyst was voluntarily donated for research as surplus after in vitro fertilization treatment following informed consent. The NYSCFe001-A line, registered as NYSCF100 on the NIH registry, presents normal karyotype, is mycoplasma free, expresses all the pluripotency markers and has the potential to differentiate into all three germ layers in vitro.

  14. Natural autoantibodies and complement promote the uptake of a self antigen, human thyroglobulin, by B cells and the proliferation of thyroglobulin-reactive CD4(+) T cells in healthy individuals

    DEFF Research Database (Denmark)

    Nielsen, C H; Leslie, R G; Jepsen, B S

    2001-01-01

    of complement receptor types 1 (CR1, CD35) and 2 (CR2, CD21). T cell responsiveness to Tg was examined in a preparation of peripheral blood mononuclear cells (PBMC) cultured in the presence of autologous serum. A subset of CD4(+) T cells exhibited a dose-dependent proliferative response to Tg, which...... cells are prerequisites for the proliferation of Tg-reactive CD4(+) T cells, suggesting a novel role for natural autoantibodies and complement in the regulation of autoreactivity under physiological conditions....... was strongly inhibited by complement inactivation and by immunoabsorption of Tg-reactive antibodies. Furthermore, this T cell response was abrogated by depletion of B cells from the PBMC culture. These data imply that uptake of complement-opsonized Tg / anti-Tg complexes and subsequent presentation of Tg by B...

  15. Retinoic acid induction of CD1d expression primes chronic lymphocytic leukemia B cells for killing by CD8+ invariant natural killer T cells.

    Science.gov (United States)

    Ghnewa, Yasmeen G; O'Reilly, Vincent P; Vandenberghe, Elisabeth; Browne, Paul V; McElligott, Anthony M; Doherty, Derek G

    2017-10-01

    Invariant natural killer T (iNKT) cells are cytotoxic T cells that respond to glycolipid antigens presented by CD1d. Therapeutic activation of iNKT cells with α-galactosylceramide (α-GalCer) can prevent and reverse tumor growth in mice and clinical trials involving α-GalCer-stimulated iNKT cells are ongoing in humans. B cells express CD1d, however, we show that CD1d expression is reduced on B cells from patients with chronic lymphocytic leukemia (CLL). B cells from CLL patients pulsed with α-GalCer failed to stimulate cytolytic degranulation by iNKT cell lines, but could present the more potent glycolipid analogue, 7DW8-5. Retinoic acid receptor-α (RAR-α) agonists induced CD1d expression by CLL B cells, restoring their ability to present α-GalCer to CD8α + iNKT cells, resulting in cytolytic degranulation. Thus, RAR-α agonists can augment the anti-tumor activities of iNKT cells against CLL cells in vitro. Their inclusion in iNKT cell-based therapies may benefit patients with CLL. Copyright © 2017 Elsevier Inc. All rights reserved.

  16. Anti-CD20 B-cell depletion enhances monocyte reactivity in neuroimmunological disorders

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    Hohlfeld Reinhard

    2011-10-01

    Full Text Available Abstract Background Clinical trials evaluating anti-CD20-mediated B-cell depletion in multiple sclerosis (MS and neuromyelitis optica (NMO generated encouraging results. Our recent studies in the MS model experimental autoimmune encephalomyelitis (EAE attributed clinical benefit to extinction of activated B-cells, but cautioned that depletion of naïve B-cells may be undesirable. We elucidated the regulatory role of un-activated B-cells in EAE and investigated whether anti-CD20 may collaterally diminish regulatory B-cell properties in treatment of neuroimmunological disorders. Methods Myelin oligodendrocyte glycoprotein (MOG peptide-immunized C57Bl/6 mice were depleted of B-cells. Functional consequences for regulatory T-cells (Treg and cytokine production of CD11b+ antigen presenting cells (APC were assessed. Peripheral blood mononuclear cells from 22 patients receiving anti-CD20 and 23 untreated neuroimmunological patients were evaluated for frequencies of B-cells, T-cells and monocytes; monocytic reactivity was determined by TNF-production and expression of signalling lymphocytic activation molecule (SLAM. Results We observed that EAE-exacerbation upon depletion of un-activated B-cells closely correlated with an enhanced production of pro-inflammatory TNF by CD11b+ APC. Paralleling this pre-clinical finding, anti-CD20 treatment of human neuroimmunological disorders increased the relative frequency of monocytes and accentuated pro-inflammatory monocyte function; when reactivated ex vivo, a higher frequency of monocytes from B-cell depleted patients produced TNF and expressed the activation marker SLAM. Conclusions These data suggest that in neuroimmunological disorders, pro-inflammatory APC activity is controlled by a subset of B-cells which is eliminated concomitantly upon anti-CD20 treatment. While this observation does not conflict with the general concept of B-cell depletion in human autoimmunity, it implies that its safety and

  17. The Tol2 transposon system mediates the genetic engineering of T-cells with CD19-specific chimeric antigen receptors for B-cell malignancies.

    Science.gov (United States)

    Tsukahara, T; Iwase, N; Kawakami, K; Iwasaki, M; Yamamoto, C; Ohmine, K; Uchibori, R; Teruya, T; Ido, H; Saga, Y; Urabe, M; Mizukami, H; Kume, A; Nakamura, M; Brentjens, R; Ozawa, K

    2015-02-01

    Engineered T-cell therapy using a CD19-specific chimeric antigen receptor (CD19-CAR) is a promising strategy for the treatment of advanced B-cell malignancies. Gene transfer of CARs to T-cells has widely relied on retroviral vectors, but transposon-based gene transfer has recently emerged as a suitable nonviral method to mediate stable transgene expression. The advantages of transposon vectors compared with viral vectors include their simplicity and cost-effectiveness. We used the Tol2 transposon system to stably transfer CD19-CAR into human T-cells. Normal human peripheral blood lymphocytes were co-nucleofected with the Tol2 transposon donor plasmid carrying CD19-CAR and the transposase expression plasmid and were selectively propagated on NIH3T3 cells expressing human CD19. Expanded CD3(+) T-cells with stable and high-level transgene expression (~95%) produced interferon-γ upon stimulation with CD19 and specifically lysed Raji cells, a CD19(+) human B-cell lymphoma cell line. Adoptive transfer of these T-cells suppressed tumor progression in Raji tumor-bearing Rag2(-/-)γc(-/-) immunodeficient mice compared with control mice. These results demonstrate that the Tol2 transposon system could be used to express CD19-CAR in genetically engineered T-cells for the treatment of refractory B-cell malignancies.

  18. To B or not to B cells-mediate a healthy start to life.

    Science.gov (United States)

    Nguyen, T G; Ward, C M; Morris, J M

    2013-02-01

    Maternal immune responses during pregnancy are critical in programming the future health of a newborn. The maternal immune system is required to accommodate fetal immune tolerance as well as to provide a protective defence against infections for the immunocompromised mother and her baby during gestation and lactation. Natural immunity and antibody production by maternal B cells play a significant role in providing such immunoprotection. However, aberrations in the B cell compartment as a consequence of maternal autoimmunity can pose serious risks to both the mother and her baby. Despite their potential implication in shaping pregnancy outcomes, the role of B cells in human pregnancy has been poorly studied. This review focuses on the role of B cells and the implications of B cell depletion therapy in pregnancy. It highlights the evidence of an association between aberrant B cell compartment and obstetric conditions. It also alludes to the potential mechanisms that amplify these B cell aberrances and thereby contribute to exacerbation of some maternal autoimmune conditions and poor neonatal outcomes. Clinical and experimental evidence suggests strongly that maternal autoantibodies contribute directly to the pathologies of obstetric and neonatal conditions that have significant implications for the lifelong health of a newborn. The evidence for clinical benefit and safety of B cell depletion therapies in pregnancy is reviewed, and an argument is mounted for further clinical evaluation of B cell-targeted therapies in high-risk pregnancy, with an emphasis on improving neonatal outcomes and prevention of neonatal conditions such as congenital heart block and fetal/neonatal alloimmune thrombocytopenia. © 2012 British Society for Immunology.

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

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    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.

  20. Towards the generation of B-cell receptor retrogenic mice.

    Directory of Open Access Journals (Sweden)

    Jenny Freitag

    Full Text Available Transgenic expression of B- and T-cell receptors (BCRs and TCRs, respectively has been a standard tool to study lymphocyte development and function in vivo. The generation of transgenic mice is time-consuming and, therefore, a faster method to study the biology of defined lymphocyte receptors in vivo would be highly welcome. Using 2A peptide-linked multicistronic retroviral vectors to transduce stem cells, TCRs can be expressed rapidly in mice of any background. We aimed at adopting this retrogenic technology to the in vivo expression of BCRs. Using a well characterised BCR specific for hen egg lysozyme (HEL, we achieved surface expression of the retrogenically encoded BCR in a Rag-deficient pro B-cell line in vitro. In vivo, retrogenic BCRs were detectable only intracellularly but not on the surface of B cells from wild type or Rag2-deficient mice. This data, together with the fact that no BCR retrogenic mouse model has been published in the 7 years since the method was originally published for TCRs, strongly suggests that achieving BCR-expression in vivo with retrogenic technology is highly challenging if not impossible.

  1. Generation of human induced pluripotent stem cell lines from human dermal fibroblasts using a non-integration system

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    Kyung-Ok Uhm

    2017-05-01

    Full Text Available We generated human induced pluripotent stem cells (hiPSCs from dermal fibroblasts using a Sendai virus (SeV-based gene delivery method. The generated hiPSC line, KSCBi002-A, has a normal karyotype (46,XY. The pluripotency and differentiation capacity were characterized by comparison with those of a human embryonic stem cell line. This cell line is registered and available from the National Stem Cell Bank, Korea National Institute of Health.

  2. DNA cytosine methylation in the bovine leukemia virus promoter is associated with latency in a lymphoma-derived B-cell line: potential involvement of direct inhibition of cAMP-responsive element (CRE)-binding protein/CRE modulator/activation transcription factor binding.

    Science.gov (United States)

    Pierard, Valérie; Guiguen, Allan; Colin, Laurence; Wijmeersch, Gaëlle; Vanhulle, Caroline; Van Driessche, Benoît; Dekoninck, Ann; Blazkova, Jana; Cardona, Christelle; Merimi, Makram; Vierendeel, Valérie; Calomme, Claire; Nguyên, Thi Liên-Anh; Nuttinck, Michèle; Twizere, Jean-Claude; Kettmann, Richard; Portetelle, Daniel; Burny, Arsène; Hirsch, Ivan; Rohr, Olivier; Van Lint, Carine

    2010-06-18

    Bovine leukemia virus (BLV) proviral latency represents a viral strategy to escape the host immune system and allow tumor development. Besides the previously demonstrated role of histone deacetylation in the epigenetic repression of BLV expression, we showed here that BLV promoter activity was induced by several DNA methylation inhibitors (such as 5-aza-2'-deoxycytidine) and that overexpressed DNMT1 and DNMT3A, but not DNMT3B, down-regulated BLV promoter activity. Importantly, cytosine hypermethylation in the 5'-long terminal repeat (LTR) U3 and R regions was associated with true latency in the lymphoma-derived B-cell line L267 but not with defective latency in YR2 cells. Moreover, the virus-encoded transactivator Tax(BLV) decreased DNA methyltransferase expression levels, which could explain the lower level of cytosine methylation observed in the L267(LTaxSN) 5'-LTR compared with the L267 5'-LTR. Interestingly, DNA methylation inhibitors and Tax(BLV) synergistically activated BLV promoter transcriptional activity in a cAMP-responsive element (CRE)-dependent manner. Mechanistically, methylation at the -154 or -129 CpG position (relative to the transcription start site) impaired in vitro binding of CRE-binding protein (CREB) transcription factors to their respective CRE sites. Methylation at -129 CpG alone was sufficient to decrease BLV promoter-driven reporter gene expression by 2-fold. We demonstrated in vivo the recruitment of CREB/CRE modulator (CREM) and to a lesser extent activating transcription factor-1 (ATF-1) to the hypomethylated CRE region of the YR2 5'-LTR, whereas we detected no CREB/CREM/ATF recruitment to the hypermethylated corresponding region in the L267 cells. Altogether, these findings suggest that site-specific DNA methylation of the BLV promoter represses viral transcription by directly inhibiting transcription factor binding, thereby contributing to true proviral latency.

  3. Clonogenic cell line survival of a human liver cancer cell line SMMC-7721 after carbon ion irradiation with different LET

    International Nuclear Information System (INIS)

    Lei Suwen; Su Xu; Wang Jifang; Li Wenjian

    2003-01-01

    Objective: To investigate the survival fraction of a human liver cancer cell line SMMC-7721 following irradiation with carbon ions with different LET. Methods: cells of the human liver cancer cell line SMMC-7721 were irradiated with carbon ions (LET=30 and 70 keV/μm). The survival fraction was determined with clonogenic assay after 9 days incubation in a 5% CO 2 incubator at 37 degree C. Results: When the survival fractions of 70 keV/μm were D s = 0.1 and D s=0.01 absorption dose were 2.94 and 5.88 Gy respectively, and those of 30 keV/μm were 4.00 and 8.00 Gy respectively. Conclusion: For the SMMC-7721 cell line, 70 keV/μm is more effective for cell killing than 30 keV/μm

  4. Network signatures of cellular immortalization in human lymphoblastoid cell lines

    Energy Technology Data Exchange (ETDEWEB)

    Shim, Sung-Mi; Jung, So-Young; Nam, Hye-Young; Kim, Hye-Ryun; Lee, Mee-Hee; Kim, Jun-Woo; Han, Bok-Ghee [National Biobank of Korea, Center for Genome Science, Korea National Institute of Health, Osong 363-951 (Korea, Republic of); Jeon, Jae-Pil, E-mail: jaepiljeon@hanmail.net [Division of Brain Diseases, Center for Biomedical Science, Korea National Institute of Health, Osong 363-951 (Korea, Republic of)

    2013-11-15

    Highlights: •We identified network signatures of LCL immortalization from transcriptomic profiles. •More than 41% of DEGs are possibly regulated by miRNAs in LCLs. •MicroRNA target genes in LCLs are involved in apoptosis and immune-related functions. •This approach is useful to find functional miRNA targets in specific cell conditions. -- Abstract: Human lymphoblastoid cell line (LCL) has been used as an in vitro cell model in genetic and pharmacogenomic studies, as well as a good model for studying gene expression regulatory machinery using integrated genomic analyses. In this study, we aimed to identify biological networks of LCL immortalization from transcriptomic profiles of microRNAs and their target genes in LCLs. We first selected differentially expressed genes (DEGs) and microRNAs (DEmiRs) between early passage LCLs (eLCLs) and terminally differentiated late passage LCLs (tLCLs). The in silico and correlation analysis of these DEGs and DEmiRs revealed that 1098 DEG–DEmiR pairs were found to be positively (n = 591 pairs) or negatively (n = 507 pairs) correlated with each other. More than 41% of DEGs are possibly regulated by miRNAs in LCL immortalizations. The target DEGs of DEmiRs were enriched for cellular functions associated with apoptosis, immune response, cell death, JAK–STAT cascade and lymphocyte activation while non-miRNA target DEGs were over-represented for basic cell metabolisms. The target DEGs correlated negatively with miR-548a-3p and miR-219-5p were significantly associated with protein kinase cascade, and the lymphocyte proliferation and apoptosis, respectively. In addition, the miR-106a and miR-424 clusters located in the X chromosome were enriched in DEmiR–mRNA pairs for LCL immortalization. In this study, the integrated transcriptomic analysis of LCLs could identify functional networks of biologically active microRNAs and their target genes involved in LCL immortalization.

  5. B cell lymphoma and myeloma in murine Gaucher's disease

    NARCIS (Netherlands)

    Pavlova, E. V.; Wang, S. Z.; Archer, J.; Dekker, N. [=Nick; Aerts, J. M. F. G.; Karlsson, S.; Cox, T. M.

    2013-01-01

    Multiple myeloma and B cell lymphoma are leading causes of death in Gaucher's disease but the nature of the stimulus driving the often noted clonal expansion of immunoglobulin-secreting B cells and cognate lymphoid malignancy is unknown. We investigated the long-term development of B cell

  6. Unique B cell differentiation profile in tolerant kidney transplant patients.

    Science.gov (United States)

    Chesneau, M; Pallier, A; Braza, F; Lacombe, G; Le Gallou, S; Baron, D; Giral, M; Danger, R; Guerif, P; Aubert-Wastiaux, H; Néel, A; Michel, L; Laplaud, D-A; Degauque, N; Soulillou, J-P; Tarte, K; Brouard, S

    2014-01-01

    Operationally tolerant patients (TOL) display a higher number of blood B cells and transcriptional B cell signature. As they rarely develop an allo-immune response, they could display an abnormal B cell differentiation. We used an in vitro culture system to explore T-dependent differentiation of B cells into plasma cells. B cell phenotype, apoptosis, proliferation, cytokine, immunoglobulin production and markers of differentiation were followed in blood of these patients. Tolerant recipients show a higher frequency of CD20(+) CD24(hi) CD38(hi) transitional and CD20(+) CD38(lo) CD24(lo) naïve B cells compared to patients with stable graft function, correlating with a decreased frequency of CD20(-) CD38(+) CD138(+) differentiated plasma cells, suggestive of abnormal B cell differentiation. B cells from TOL proliferate normally but produce more IL-10. In addition, B cells from tolerant recipients exhibit a defective expression of factors of the end step of differentiation into plasma cells and show a higher propensity for cell death apoptosis compared to patients with stable graft function. This in vitro profile is consistent with down-regulation of B cell differentiation genes and anti-apoptotic B cell genes in these patients in vivo. These data suggest that a balance between B cells producing IL-10 and a deficiency in plasma cells may encourage an environment favorable to the tolerance maintenance. © Copyright 2013 The American Society of Transplantation and the American Society of Transplant Surgeons.

  7. δ-Aminolevulinic acid cytotoxic effects on human hepatocarcinoma cell lines

    Energy Technology Data Exchange (ETDEWEB)

    De Siervi, Adriana; Vazquez, Elba S; Rezaval, Carolina; Rossetti, María V; Batlle, Alcira M del [Centro de Investigaciones sobre Porfirinas y Porfirias (CIPYP), Argentine National Research Council (CONICET), Department of Biological Chemistry, FCEN, University of Buenos Aires (Argentina)

    2002-01-01

    Acute Intermittent Porphyria is a genetic disorder of heme metabolism, characterized by increased levels of porphyrin precursors, δ-aminolevulinic acid (ALA) and porphobilinogen (PBG). ALA has been reported to generate reactive oxygen species and to cause oxidative damage to proteins, subcellular structures and DNA. It is known that oxidative stress can induce apoptosis. The aim of this work was to study the cytotoxic effect of ALA on two hepatocarcinoma cell lines. We have determined the impact of ALA on HEP G2 and HEP 3B hepatocarcinoma cell lines survival as measured by the MTT assay. ALA proved to be cytotoxic in both cell lines however; HEP G2 was more sensitive to ALA than HEP 3B. Addition of hemin or glucose diminished ALA cytotoxicity in HEP G2 cells; instead it was enhanced in HEP 3B cells. Because apoptosis is usually associated with DNA fragmentation, the DNA of ALA treated and untreated cells were analyzed. The characteristic pattern of DNA fragmentation ladders was observed in ALA treated cells. To elucidate the mechanisms of ALA induced apoptosis, we examined its effect on p53 expression. No changes in p53 mRNA levels were observed after exposure of both cell lines to ALA for 24 h. CDK2 and CDK4 protein levels were reduced after ALA treatment at physiological concentrations.

  8. δ-Aminolevulinic acid cytotoxic effects on human hepatocarcinoma cell lines

    Directory of Open Access Journals (Sweden)

    del Batlle Alcira M

    2002-03-01

    Full Text Available Abstract Background Acute Intermittent Porphyria is a genetic disorder of heme metabolism, characterized by increased levels of porphyrin precursors, δ-aminolevulinic acid (ALA and porphobilinogen (PBG. ALA has been reported to generate reactive oxygen species and to cause oxidative damage to proteins, subcellular structures and DNA. It is known that oxidative stress can induce apoptosis. The aim of this work was to study the cytotoxic effect of ALA on two hepatocarcinoma cell lines. Results We have determined the impact of ALA on HEP G2 and HEP 3B hepatocarcinoma cell lines survival as measured by the MTT assay. ALA proved to be cytotoxic in both cell lines however; HEP G2 was more sensitive to ALA than HEP 3B. Addition of hemin or glucose diminished ALA cytotoxicity in HEP G2 cells; instead it was enhanced in HEP 3B cells. Because apoptosis is usually associated with DNA fragmentation, the DNA of ALA treated and untreated cells were analyzed. The characteristic pattern of DNA fragmentation ladders was observed in ALA treated cells. To elucidate the mechanisms of ALA induced apoptosis, we examined its effect on p53 expression. No changes in p53 mRNA levels were observed after exposure of both cell lines to ALA for 24 h. CDK2 and CDK4 protein levels were reduced after ALA treatment at physiological concentrations.

  9. δ-Aminolevulinic acid cytotoxic effects on human hepatocarcinoma cell lines

    International Nuclear Information System (INIS)

    De Siervi, Adriana; Vazquez, Elba S; Rezaval, Carolina; Rossetti, María V; Batlle, Alcira M del

    2002-01-01

    Acute Intermittent Porphyria is a genetic disorder of heme metabolism, characterized by increased levels of porphyrin precursors, δ-aminolevulinic acid (ALA) and porphobilinogen (PBG). ALA has been reported to generate reactive oxygen species and to cause oxidative damage to proteins, subcellular structures and DNA. It is known that oxidative stress can induce apoptosis. The aim of this work was to study the cytotoxic effect of ALA on two hepatocarcinoma cell lines. We have determined the impact of ALA on HEP G2 and HEP 3B hepatocarcinoma cell lines survival as measured by the MTT assay. ALA proved to be cytotoxic in both cell lines however; HEP G2 was more sensitive to ALA than HEP 3B. Addition of hemin or glucose diminished ALA cytotoxicity in HEP G2 cells; instead it was enhanced in HEP 3B cells. Because apoptosis is usually associated with DNA fragmentation, the DNA of ALA treated and untreated cells were analyzed. The characteristic pattern of DNA fragmentation ladders was observed in ALA treated cells. To elucidate the mechanisms of ALA induced apoptosis, we examined its effect on p53 expression. No changes in p53 mRNA levels were observed after exposure of both cell lines to ALA for 24 h. CDK2 and CDK4 protein levels were reduced after ALA treatment at physiological concentrations

  10. Intravascular Large B-Cell Lymphoma

    Directory of Open Access Journals (Sweden)

    Maria S. Khan MD, FACP

    2014-03-01

    Full Text Available Case Presentation . A 69-year-old Hispanic male, with a past history of diabetes and coronary disease, was admitted for fever, diarrhea, and confusion of 4 weeks duration. Physical examination showed a disoriented patient with multiple ecchymoses, possible ascites, and bilateral scrotal swelling. Hemoglobin was 6.7, prothrombin time (PT 21.4 seconds with international normalized ratio 2.1, partial thromboplastin time (PTT 55.6 seconds, fibrin split 10 µg/L, and lactate dehydrogenase (LDH 1231 IU/L. Except for a positive DNA test for Epstein–Barr virus (EBV infection, extensive diagnostic workup for infections, malignancy, or a neurological cause was negative. Mixing studies revealed a nonspecific inhibitor of PT and PTT but Factor VIII levels were normal. The patient was empirically treated with antibiotics but developed hypotension and died on day 27 of admission. At autopsy, patient was found to have intravascular diffuse large B-cell lymphoma involving skin, testes, lung, and muscles. The malignant cells were positive for CD20, CD791, Mum-1, and Pax-5 and negative for CD3, CD5, CD10, CD30, and Bcl-6. The malignant cells were 100% positive for Ki-67. Discussion . Intravascular large cell B-cell lymphoma (IVLBCL is rare form of diffuse large B-cell lymphoma and tends to proliferate within small blood vessels, particularly capillaries and postcapillary venules. The cause of its affinity for vascular bed remains unknown. In many reports, IVLBCL was associated with HIV, HHV8, and EBV infections. The fact that our case showed evidence of EBV infection lends support to the association of this diagnosis to viral illness. The available literature on this subject is scant, and in many cases, the diagnosis was made only at autopsy. The typical presentation of this disorder is with B symptoms, progressive neurologic deficits, and skin findings. Bone marrow, spleen, and liver are involved in a minority of patients. Nearly all patients have elevated LDH

  11. Trypanosoma brucei Co-opts NK Cells to Kill Splenic B2 B Cells.

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    Deborah Frenkel

    2016-07-01

    Full Text Available After infection with T. brucei AnTat 1.1, C57BL/6 mice lost splenic B2 B cells and lymphoid follicles, developed poor parasite-specific antibody responses, lost weight, became anemic and died with fulminating parasitemia within 35 days. In contrast, infected C57BL/6 mice lacking the cytotoxic granule pore-forming protein perforin (Prf1-/- retained splenic B2 B cells and lymphoid follicles, developed high-titer antibody responses against many trypanosome polypeptides, rapidly suppressed parasitemia and did not develop anemia or lose weight for at least 60 days. Several lines of evidence show that T. brucei infection-induced splenic B cell depletion results from natural killer (NK cell-mediated cytotoxicity: i B2 B cells were depleted from the spleens of infected intact, T cell deficient (TCR-/- and FcγRIIIa deficient (CD16-/- C57BL/6 mice excluding a requirement for T cells, NKT cell, or antibody-dependent cell-mediated cytotoxicity; ii administration of NK1.1 specific IgG2a (mAb PK136 but not irrelevant IgG2a (myeloma M9144 prevented infection-induced B cell depletion consistent with a requirement for NK cells; iii splenic NK cells but not T cells or NKT cells degranulated in infected C57BL/6 mice co-incident with B cell depletion evidenced by increased surface expression of CD107a; iv purified NK cells from naïve C57BL/6 mice killed purified splenic B cells from T. brucei infected but not uninfected mice in vitro indicating acquisition of an NK cell activating phenotype by the post-infection B cells; v adoptively transferred C57BL/6 NK cells prevented infection-induced B cell population growth in infected Prf1-/- mice consistent with in vivo B cell killing; vi degranulated NK cells in infected mice had altered gene and differentiation antigen expression and lost cytotoxic activity consistent with functional exhaustion, but increased in number as infection progressed indicating continued generation. We conclude that NK cells in T. brucei

  12. Targeting Bruton Tyrosine Kinase: A novel strategy in the treatment of B-cell lymphomas

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    Sklavenitis-Pistofidis R.

    2017-06-01

    Full Text Available In normal B-cells, Bruton tyrosine kinase (Btk, a non-receptor tyrosine kinase involved in B-cell receptor (BCR signalling, is essential for cell survival and maturation. Not surprisingly, Btk is also implicated in the pathogenesis of B-cell lymphomas, like Chronic Lymphocytic Leukaemia/Small Lymphocytic Lymphoma (CLL/SLL, Mantle Cell Lymphoma (MCL and Waldenström’s Macroglobulinemia (WM, which are driven by aberrant BCR signalling. Thus, targeting Btk represents a promising therapeutic strategy in the treatment of B-cell lymphoma patients. Ibrutinib, a selective Btk inhibitor, has already been approved as second-line treatment of CLL/SLL, MCL and WM patients, while more clinical studies of ibrutinib and novel Btk inhibitors are currently under way. In light of results of the RESONATE-2 trial, the approval of ibrutinib as a first-line treatment of CLL/SLL may well be approaching. Herein, we review Btk’s role in normal and malignant BCR signalling, as well as ibrutinib’s performance in B-cell lymphoma treatment and prognosis.

  13. Generation and characterization of a human iPSC cell line expressing inducible Cas9 in the “safe harbor” AAVS1 locus

    Directory of Open Access Journals (Sweden)

    Julio Castaño

    2017-05-01

    Full Text Available We report the generation-characterization of a fetal liver (FL B-cell progenitor (BCP-derived human induced pluripotent stem cell (hiPSC line CRISPR/Cas9-edited to carry/express a single copy of doxycycline-inducible Cas9 gene in the “safe locus” AAVS1 (iCas9-FL-BCP-hiPSC. Gene-edited iPSCs remained pluripotent after CRISPR/Cas9 genome-edition. Correct genomic integration of a unique copy of Cas9 was confirmed by PCR and Southern blot. Cas9 was robustly and specifically expressed on doxycycline exposure. T7-endonuclease assay demonstrated that iCas9 induces robust gene-edition when gRNAs against hematopoietic transcription factors were tested. This iCas9-FL-BCP-hiPSC will facilitate gene-editing approaches for studies on developmental biology, drug screening and disease modeling.

  14. Derivation of Two New Human Embryonic Stem Cell Lines from Nonviable Human Embryos

    Directory of Open Access Journals (Sweden)

    Svetlana Gavrilov

    2011-01-01

    Full Text Available We report the derivation and characterization of two new human embryonic stem cells (hESC lines (CU1 and CU2 from embryos with an irreversible loss of integrated organismic function. In addition, we analyzed retrospective data of morphological progression from embryonic day (ED 5 to ED6 for 2480 embryos not suitable for clinical use to assess grading criteria indicative of loss of viability on ED5. Our analysis indicated that a large proportion of in vitro fertilization (IVF embryos not suitable for clinical use could be used for hESC derivation. Based on these combined findings, we propose that criteria commonly used in IVF clinics to determine optimal embryos for uterine transfer can be employed to predict the potential for hESC derivation from poor quality embryos without the destruction of vital human embryos.

  15. B cell lymphomas express CX3CR1 a non-B cell lineage adhesion molecule

    DEFF Research Database (Denmark)

    Andreasson, U.; Ek, S.; Merz, H.

    2008-01-01

    normally is not expressed on B cells, is expressed both at the mRNA and protein level in several subtypes of lymphoma. CX3CR1 has also shown to be involved in the homing to specific tissues that express the ligand, CX3CL1, in breast and prostate cancer and may thus be involved in dissemination of lymphoma......To study the differential expression of cell membrane-bound receptors and their potential role in growth and/or survival of the tumor cells, highly purified follicular lymphoma cells were analyzed, using gene expression analysis, and compared to non-malignant B cell populations. Filtering...... the genome for overexpressed genes coding for cell membrane-bound proteins/receptors resulted in a hit list of 27 identified genes. Among these, we have focused on the aberrant over expression of CX3CR1, in different types of B cell lymphoma, as compared to non-malignant B cells. We show that CX3CR1, which...

  16. Lining cells on normal human vertebral bone surfaces

    International Nuclear Information System (INIS)

    Henning, C.B.; Lloyd, E.L.

    1982-01-01

    Thoracic vertebrae from two individuals with no bone disease were studied with the electron microscope to determine cell morphology in relation to bone mineral. The work was undertaken to determine if cell morphology or spatial relationships between the bone lining cells and bone mineral could account for the relative infrequency of bone tumors which arise at this site following radium intake, when compared with other sites, such as the head of the femur. Cells lining the vertebral mineral were found to be generally rounded in appearance with varied numbers of cytoplasmic granules, and they appeared to have a high density per unit of surface area. These features contrasted with the single layer of flattened cells characteristic of the bone lining cells of the femur. A tentative discussion of the reasons for the relative infrequency of tumors in the vertebrae following radium acquisition is presented

  17. Endometrial Polyps and Benign Endometrial Hyperplasia Present Increased Prevalence of DNA Fragmentation Factors 40 and 45 (DFF40 and DFF45) Together With the Antiapoptotic B-Cell Lymphoma (Bcl-2) Protein Compared With Normal Human Endometria.

    Science.gov (United States)

    Banas, Tomasz; Pitynski, Kazimierz; Mikos, Marcin; Cielecka-Kuszyk, Joanna

    2017-09-13

    DNA fragmentation factor 40 (DFF40) is a key executor of apoptosis. It localizes to the nucleus together with DNA fragmentation factor 45 (DFF45), which acts as a DFF40 inhibitor and chaperone. B-cell lymphoma (Bcl-2) protein is a proven antiapoptotic factor present in the cytoplasm. In this study, we aimed to investigate DFF40, DFF45, and Bcl-2 immunoexpression in endometrial polyps (EPs) and benign endometrial hyperplasia (BEH) tissue compared with that in normal proliferative endometrium (NPE) and normal secretory endometrium (NSE) as well as normal post menopausal endometrium (NAE). This study used archived samples from 65 and 62 cases of EPs and BEH, respectively. The control group consisted of 52 NPE, 54 NSE, and 54 NAE specimens. Immunohistochemistry was used to detect DFF40, DFF45, and Bcl-2. DFF40, DFF45, and Bcl-2 were more highly expressed in the glandular layer of EPs and BEH compared with the stroma, and this was not influenced by menopausal status. Both glandular and stromal expression of DFF40, DFF45, and Bcl-2 were significantly higher in EPs compared with NPE, NSE, and NAE. Glandular BEH tissue showed significantly higher DFF40, DFF45, and Bcl-2 expression than in NPE, NSE, and NAE. No differences in the glandular expression of DFF40, DFF45, and Bcl-2 were observed between EP and BEH tissues, while Bcl-2 stromal expression in BEH was significantly lower than in EPs. Glandular, menopause-independent DFF40, DFF45, and Bcl-2 overexpression may play an important role in the pathogenesis of EPs and BEH.

  18. Discovery and validation of the tumor-suppressive function of long noncoding RNA PANDA in human diffuse large B-cell lymphoma through the inactivation of MAPK/ERK signaling pathway.

    Science.gov (United States)

    Wang, Yingjun; Zhang, Mingzhi; Xu, Huanan; Wang, Yifei; Li, Zhaoming; Chang, Yu; Wang, Xinhuan; Fu, Xiaorui; Zhou, Zhiyuan; Yang, Siyuan; Wang, Bei; Shang, Yufeng

    2017-09-22

    Diffuse large B-cell lymphoma (DLBCL) is one of the leading causes of cancer-related mortality, and responds badly to existing treatment. Thus, it is of urgent need to identify novel prognostic markers and therapeutic targets of DLBCL. Recent studies have shown that long non-coding RNAs (lncRNAs) play an important role in the development of cancer. By using the next generation HiSeq sequencing assay, we determined lncRNAs exhibiting differential expression between DLBCL patients and healthy controls. Then, RT-qPCR was performed for identification in clinical samples and cell materials, and lncRNA PANDA was verified to be down-regulated in DLBCL patients and have considerable diagnostic potential. In addition, decreased serum PANDA level was correlated to poorer clinical outcome and lower overall survival in DLBCL patients. Subsequently, we determined the experimental role of lncRNA PANDA in DLBCL progression. Luciferase reporter assay and chromatin immunoprecipitation assay suggested that lncRNA PANDA was induced by p53 and p53 interacts with the promoter region of PANDA. Cell functional assay further indicated that PANDA functioned as a tumor suppressor gene through the suppression of cell growth by a G0/G1 cell cycle arrest in DLBCL. More importantly, Cignal Signal Transduction Reporter Array and western blot assay showed that lncRNA PANDA inactivated the MAPK/ERK signaling pathway. In conclusion, our integrated approach demonstrates that PANDA in DLBCL confers a tumor suppressive function through inhibiting cell proliferation and silencing MAPK/ERK signaling pathway. Thus, PANDA may be a promising therapeutic target for patients with DLBCL.

  19. Citric acid induces cell-cycle arrest and apoptosis of human immortalized keratinocyte cell line (HaCaT) via caspase- and mitochondrial-dependent signaling pathways.

    Science.gov (United States)

    Ying, Tsung-Ho; Chen, Chia-Wei; Hsiao, Yu-Ping; Hung, Sung-Jen; Chung, Jing-Gung; Yang, Jen-Hung

    2013-10-01

    Citric acid is an alpha-hydroxyacid (AHA) widely used in cosmetic dermatology and skincare products. However, there is concern regarding its safety for the skin. In this study, we investigated the cytotoxic effects of citric acid on the human keratinocyte cell line HaCaT. HaCaT cells were treated with citric acid at 2.5-12.5 mM for different time periods. Cell-cycle arrest and apoptosis were investigated by 4,6-diamidino-2-phenylindole dihydrochloride (DAPI) staining, flow cytometry, western blot and confocal microscopy. Citric acid not only inhibited proliferation of HaCaT cells in a dose-dependent manner, but also induced apoptosis and cell cycle-arrest at the G2/M phase (before 24 h) and S phase (after 24 h). Citric acid increased the level of Bcl-2-associated X protein (BAX) and reduced the levels of B-cell lymphoma-2 (BCL-2), B-cell lymphoma-extra large (BCL-XL) and activated caspase-9 and caspase-3, which subsequently induced apoptosis via caspase-dependent and caspase-independent pathways. Citric acid also activated death receptors and increased the levels of caspase-8, activated BH3 interacting-domain death agonist (BID) protein, Apoptosis-inducing factor (AIF), and Endonuclease G (EndoG). Therefore, citric acid induces apoptosis through the mitochondrial pathway in the human keratinocyte cell line HaCaT. The study results suggest that citric acid is cytotoxic to HaCaT cells via induction of apoptosis and cell-cycle arrest in vitro.

  20. The regulatory roles of B cell subsets in transplantation.

    Science.gov (United States)

    Chu, Zhulang; Zou, Weilong; Xu, Yanan; Sun, Qiquan; Zhao, Yong

    2018-02-01

    B cells mediate allograft rejection through antigen presentation, and production of cytokines and antibodies. More and more immunosuppressive agents specifically targeting B cells and plasma cells have been applied in clinical transplantation. However, recent studies have indicated the regulatory roles of B cells. Therefore, it is vital to clarify the different effects of B cell subsets in organ transplantation so that we can completely understand the diverse functions of B cells in transplantation. Areas covered: This review focuses on the regulatory roles of B cells in transplantation. B cell subsets with immune modulation and factors mediating immunosuppressive functions of regulatory B (Breg) cells were analyzed. Therapies targeting B cells and the application of B cells for transplant tolerance induction were discussed. Expert commentary: Besides involving rejection, B cells could also play regulatory roles in transplantation. Breg cells and the related markers may be used to predict the immune tolerant state in transplant recipients. New therapeutic strategies targeting B cells should be explored to promote tolerance induction with less impact on the host's protective immunity in organ transplanted patients.

  1. Dependence of Immunoglobulin Class Switch Recombination in B Cells on Vesicular Release of ATP and CD73 Ectonucleotidase Activity

    Directory of Open Access Journals (Sweden)

    Francesca Schena

    2013-06-01

    Full Text Available Immunoglobulin (Ig isotype diversification by class switch recombination (CSR is an essential process for mounting a protective humoral immune response. Ig CSR deficiencies in humans can result from an intrinsic B cell defect; however, most of these deficiencies are still molecularly undefined and diagnosed as common variable immunodeficiency (CVID. Here, we show that extracellular adenosine critically contributes to CSR in human naive and IgM memory B cells. In these cells, coordinate stimulation of B cell receptor and toll-like receptors results in the release of ATP stored in Ca2+-sensitive secretory vesicles. Plasma membrane ectonucleoside triphosphate diphosphohydrolase 1 CD39 and ecto-5′-nucleotidase CD73 hydrolyze ATP to adenosine, which induces CSR in B cells in an autonomous fashion. Notably, CVID patients with impaired class-switched antibody responses are selectively deficient in CD73 expression in B cells, suggesting that CD73-dependent adenosine generation contributes to the pathogenesis of this disease.

  2. Lines of evidence for environmentally driven human migration

    Science.gov (United States)

    Davis, K. F.; D'Odorico, P.

    2012-12-01

    International human migration is an important mechanism that affects, and is affected by, various human and natural systems. With the number of people living outside their countries of origin currently estimated at 214 million people and projected to potentially reach more than 400 million people by mid-century, the topic of international human movements presents possible advantages and pitfalls for both sending and receiving countries on multiple fronts (e.g. economic, environmental, political and cultural). Understanding how human migration interacts with human and natural systems is therefore essential in realizing a sustainable and balanced future. While the study of international migration has historically been motivated largely by economic and political interests, the issue of environmentally induced migration has become increasingly important in light of a rapidly changing climate in conjunction with increasing population pressure on many important resources. Particularly in terms of theoretical and conceptual discussions, environmentally induced human migration has been receiving increased attention in the literature. To date, few studies - many of which focus on internal (intra-national) or regional migration - have attempted to quantify the interactions of human migration and the environment, with little attention paid to the global scale as a result of varying regional factors and lack of sufficient data. Recently available global bilateral migration datasets have been developed that allow for a more comprehensive understanding of human movements between all countries. With these datasets, we seek to elucidate environmental drivers of human migration over the past half-century using a multi-pronged approach. First, using a recently developed universal radiation model, we examine human movements based solely on global population distribution. Next, by comparison of migration movements with selected economic, environmental and human welfare indicators, we

  3. Human pathogenic Mycoplasma species induced cytokine gene expression in Epstein-Barr virus (EBV)-positive lymphoblastoid cell lines.

    Science.gov (United States)

    Schäffner, E; Opitz, O; Pietsch, K; Bauer, G; Ehlers, S; Jacobs, E

    1998-04-01

    We addressed the question whether the in vitro interaction of two Epstein-Barr virus (EBV)-genome-positive B cell lines (EB-3 and HilB-gamma) with either Mycoplasma pneumoniae or M. hominis, with the mycoplasma species (M. fermentans, M. fermentans subsp. incognitus, M. penetrans, M. genitalium) or with mycoplasma species known to be mere commensals of the respiratory tract (M. orale and M. salivarium) would result in expression of mRNAs for IL-2, IL-2R, IL-4 and IL-6 as determined by reverse transcriptase (RT)-PCR after 4 and 24 h of cocultivation. The pattern of cytokine gene expression observed depended on (i) the origin of the transformed cell line, (ii) the pathogenicity of the Mycoplasma species, and (iii) the length of cocultivation. The EBV-immortalized lymphoblastoid cell line HilB-gamma showed mRNA expression for IL-2, IL-2-receptor, IL-4 and IL-6 peaking 24 h after stimulation with M. pneumoniae and all AIDS-related mycoplasma species tested. The Burkitt lymphoma cell line EB-3 showed a distinct and isolated strong II-2/IL-2 R-mRNA expression within 4 h after contact with the pathogenic and all of the AIDS related mycoplasma species. In neither EBV-containing cell line cytokine was gene expression detectable after stimulation with the commensal mycoplasma species, M. orale and M. salivarium, indicating species differences in the ability of mycoplasmas to interact with and stimulate B-cell lines. Our data suggest that some mcyoplasma species may act as immunomodulatory cofactors by eliciting inappropriate cytokine gene expression in B cells latently infected with EBV. Therefore, this cultivation model may prove useful in evaluating the pathogenetic potential of novel isolated mycoplasma species. Copyright 1998 Academic Press Limited.

  4. Foxp1 controls mature B cell survival and the development of follicular and B-1 B cells

    Science.gov (United States)

    Patzelt, Thomas; Keppler, Selina J.; Gorka, Oliver; Thoene, Silvia; Wartewig, Tim; Reth, Michael; Förster, Irmgard; Lang, Roland; Buchner, Maike; Ruland, Jürgen

    2018-01-01

    The transcription factor Foxp1 is critical for early B cell development. Despite frequent deregulation of Foxp1 in B cell lymphoma, the physiological functions of Foxp1 in mature B cells remain unknown. Here, we used conditional gene targeting in the B cell lineage and report that Foxp1 disruption in developing and mature B cells results in reduced numbers and frequencies of follicular and B-1 B cells and in impaired antibody production upon T cell-independent immunization in vivo. Moreover, Foxp1-deficient B cells are impaired in survival even though they exhibit an increased capacity to proliferate. Transcriptional analysis identified defective expression of the prosurvival Bcl-2 family gene Bcl2l1 encoding Bcl-xl in Foxp1-deficient B cells, and we identified Foxp1 binding in the regulatory region of Bcl2l1. Transgenic overexpression of Bcl2 rescued the survival defect in Foxp1-deficient mature B cells in vivo and restored peripheral B cell numbers. Thus, our results identify Foxp1 as a physiological regulator of mature B cell survival mediated in part via the control of Bcl-xl expression and imply that this pathway might contribute to the pathogenic function of aberrant Foxp1 expression in lymphoma. PMID:29507226

  5. Effect of 5-azacytidine and galectin-1 on growth and differentiation of the human b lymphoma cell line bl36

    Directory of Open Access Journals (Sweden)

    Joubert-Caron Raymonde

    2001-12-01

    Full Text Available Abstract Background 5-AzaCytidine (AzaC is a DNA demethylating drugs that has been shown to inhibit cell growth and to induce apoptosis in certain cancer cells. Induced expression of the galectin1 (Gal1 protein, a galactoside-binding protein distributed widely in immune cells, has been described in cultured hepatoma-derived cells treated with AzaC and this event may have a role in the effect of the drug. According to this hypothesis, we investigated the effect of AzaC and Gal1 on human lymphoid B cells phenotype. Methods The effect of AzaC and Gal1 on cell growth and phenotype was determined on the Burkitt lymphoma cell line BL36. An immunocytochemical analysis for detection of Gal1 protein expression was performed in AzaC-treated cells. To investigate the direct effects of Gal1, recombinant Gal1 was added to cells. Results Treatment of lymphoid B cells with AzaC results in: i a decrease in cell growth with an arrest of the cell cycle at G0/G1 phase, ii phenotypic changes consistent with a differentiated phenotype, and iii the expression of p16, a tumor-suppressor gene whose expression was dependent of its promoter demethylation, and of Gal1. A targeting of Gal 1 to the plasma membrane follows its cytosolic expression. To determine which of the effects of AzaC might be secondary to the induction of Gal1, recombinant Gal1 was added to BL36 cells. Treated cells displayed growth inhibition and phenotypic changes consistent with a commitment toward differentiation. Conclusions Altered cell growth and expression of the cell surface plasma cell antigen, CD138 are detectable in BL36 cells treated by AzaC as well as by Gal1. It seems that AzaC-induced Gal1 expression and consequent binding of Gal1 on its cell membrane receptor may be, in part, involved in AzaC-induced plasmacytic differentiation.

  6. Glycoproteins and sialyl transferase of human B lymphoblastoid cell lines

    International Nuclear Information System (INIS)

    Lui, S.W.L.; Ng, M.H.

    1980-01-01

    We used two radiolabeling methods to study glycoproteins on the surface of lymphoblastoid cells. One of the methods affects tritiation of residues which are oxidized with galactose oxidase and the other causes tritiation of neuraminic acid residues. This approach was shown to allow a better resolution of cell surface glycoproteins than if either method were used alone. Glycoproteins of B 1 - 19 cells which harbor the Epstein-Barr virus genomes were compared with those of its parental cell line, BJAB, which does not harbor the viral genomes. These studies did not reveal a unique viral protein. A 28,000 mol. wt. glycoprotein was found to be the most prominent neuraminic acidlabeled product of B 1 - 19 cells and also of the two other cell lines, Raji and Ly38, which harbor the EBV genomes. A similar molecular weight species from BJAB cells identified by galactose oxidase labeling might be deficient in neuraminic acid residues as it was poorly labeled by the periodate oxidation method. The neuraminic acid content and level of sialyl transferase of BJAB cells were found to be lower than those of the other cell lines studied. (auth.)

  7. Method and cell lines for the production of monoclonal antibodies to human glycophorin A

    Science.gov (United States)

    Bigbee, W.L.; Fong, S.S.N.; Jensen, R.H.; Vanderlaan, M.

    Cloned mouse hybridoma cell lines have been established which continuously produce antibodies that differentiate between the M and N forms of human glycophorin A. These antibodies have potential application as human blood group reagents, as markers for terminally differentiated erythroid cells and as immunofluorescent labels of somatically variant human erythrocytes.

  8. LINES

    Directory of Open Access Journals (Sweden)

    Minas Bakalchev

    2015-10-01

    Full Text Available The perception of elements in a system often creates their interdependence, interconditionality, and suppression. The lines from a basic geometrical element have become the model of a reductive world based on isolation according to certain criteria such as function, structure, and social organization. Their traces are experienced in the contemporary world as fragments or ruins of a system of domination of an assumed hierarchical unity. How can one release oneself from such dependence or determinism? How can the lines become less “systematic” and forms more autonomous, and less reductive? How is a form released from modernistic determinism on the new controversial ground? How can these elements or forms of representation become forms of action in the present complex world? In this paper, the meaning of lines through the ideas of Le Corbusier, Leonidov, Picasso, and Hitchcock is presented. Spatial research was made through a series of examples arising from the projects of the architectural studio “Residential Transformations”, which was a backbone for mapping the possibilities ranging from playfulness to exactness, as tactics of transformation in the different contexts of the contemporary world.

  9. CYP1A1 and CYP1A2 expression: Comparing 'humanized' mouse lines and wild-type mice; comparing human and mouse hepatoma-derived cell lines

    International Nuclear Information System (INIS)

    Uno, Shigeyuki; Endo, Kaori; Ishida, Yuji; Tateno, Chise; Makishima, Makoto; Yoshizato, Katsutoshi; Nebert, Daniel W.

    2009-01-01

    Human and rodent cytochrome P450 (CYP) enzymes sometimes exhibit striking species-specific differences in substrate preference and rate of metabolism. Human risk assessment of CYP substrates might therefore best be evaluated in the intact mouse by replacing mouse Cyp genes with human CYP orthologs; however, how 'human-like' can human gene expression be expected in mouse tissues? Previously a bacterial-artificial-chromosome-transgenic mouse, carrying the human CYP1A1 C YP1A2 locus and lacking the mouse Cyp1a1 and Cyp1a2 orthologs, was shown to express robustly human dioxin-inducible CYP1A1 and basal versus inducible CYP1A2 (mRNAs, proteins, enzyme activities) in each of nine mouse tissues examined. Chimeric mice carrying humanized liver have also been generated, by transplanting human hepatocytes into a urokinase-type plasminogen activator(+/+) s evere-combined-immunodeficiency (uPA/SCID) line with most of its mouse hepatocytes ablated. Herein we compare basal and dioxin-induced CYP1A mRNA copy numbers, protein levels, and four enzymes (benzo[a]pyrene hydroxylase, ethoxyresorufin O-deethylase, acetanilide 4-hydroxylase, methoxyresorufin O-demethylase) in liver of these two humanized mouse lines versus wild-type mice; we also compare these same parameters in mouse Hepa-1c1c7 and human HepG2 hepatoma-derived established cell lines. Most strikingly, mouse liver CYP1A1-specific enzyme activities are between 38- and 170-fold higher than human CYP1A1-specific enzyme activities (per unit of mRNA), whereas mouse versus human CYP1A2 enzyme activities (per unit of mRNA) are within 2.5-fold of one another. Moreover, both the mouse and human hepatoma cell lines exhibit striking differences in CYP1A mRNA levels and enzyme activities. These findings are relevant to risk assessment involving human CYP1A1 and CYP1A2 substrates, when administered to mice as environmental toxicants or drugs.

  10. Overexpression of a transcription factor LYL1 induces T- and B-cell lymphoma in mice.

    Science.gov (United States)

    Zhong, Y; Jiang, L; Hiai, H; Toyokuni, S; Yamada, Y

    2007-10-18

    LYL1, a member of the class II basic helix-loop-helix transcription factors, is aberrantly expressed in a fraction of human T-cell acute lymphoblastic leukemia. Here, we generated transgenic mice ubiquitously overexpressing LYL1 using a construct expressing full-length cDNA driven by a human elongation factor 1alpha promoter. Four independent lines exhibiting high LYL1 expression were established. Of these transgenic mice, 96% displayed loss of hair with a short kinked tail. Furthermore, 30% of them developed malignant lymphoma, with an average latent period of 352 days. In these mice, histological examination revealed tumor cell infiltration in multiple organs and immunohistochemical analysis showed that the infiltrated tumor cells were either CD3 or CD45R/B220-positive; fluorescence-activated cell sorter analysis indicated that each tumor consisted either of mainly CD4, CD8 double-positive T cells or mature B cells; the clonality of LYL1-induced lymphoma was confirmed by T-cell receptor rearrangement and immunoglobulin heavy-chain gene rearrangement analyses. Mammalian two-hybrid analysis and luciferase assay suggested that excess LYL1 blocked the dimerization of E2A and thus inhibited the regulatory activity of E2A on the CD4 promoter. Reverse transcription-polymerase chain reaction results showed that the expression of certain E2A/HEB target genes was downregulated. Taken together, our results provide direct evidence that aberrant expression of LYL1 plays a role in lymphomagenesis.

  11. Derivation and characterization of the NYSCFe003-A human embryonic stem cell line

    Directory of Open Access Journals (Sweden)

    Ana Sevilla

    2017-12-01

    Full Text Available The human embryonic stem cell line NYSCFe003-A was derived from a day 5 to day 6 blastocyst in feeder-free and antibiotic free conditions. The blastocyst was voluntarily donated for research as surplus after in vitro fertilization treatment following informed consent. The NYSCFe003-A line expresses all the pluripotency markers and has the potential to differentiate into all three germ layers in vitro. The line presents normal karyotype and is mycoplasma free.

  12. Comparison of immunological characteristics of peripheral, splenic and tonsilar naïve B cells by differential gene expression meta-analyses.

    Science.gov (United States)

    Chokeshai-u-saha, Kaj; Lepoivre, Cyrille; Grieco, Luca; Nguyen, Catherine; Ruxrungtham, Kiat

    2012-12-01

    Naïve B cells isolated from peripheral blood, spleen and tonsil are commonly used in human B cell studies. However, little has been written about their possible variations in immunological properties. This study compared differential gene expression in human naive B subsets by meta-analysis using expression data available in Gene Expression Onimbus (GEO). Gene expression files of the Affymetrix Human Genome U133A Array (Affymetrix) were downloaded to collect 21 total array data samples of peripheral naïve B cells (n=10), splenic naïve B cells (n=2), tonsilar naïve B cells (n=3), peripheral memory B cells (n=4) and splenic memory B cells (n=2). Prior to differential gene expression analyses, data were normalized in order to reduce non-biological variation among the datasets. Comparisons of peripheral naive B cells with their splenic and tonsilar counterparts showed remarkable differences in terms of gene expression (29 and 202 genes, respectively). However, only minor differences were detected between splenic and tonsilar naive B cells (10 genes), consistent with the clustering results classifying both of them as lymphoid naive B cells. Differential gene expression results also implied higher stimulating states of lymphoid naive B cells when compared with peripheral blood naive B cells. These included enhanced expressions of CD27, CR2, EGR1, GADD45B, ICAM1, ICOSLG, IGHA, IL6, MMP9, SAMSN1, SMAD7, TNFAIP3, but reduced HLA-DOB expression. Our findings suggest that results generated from peripheral naive B cells may not always be applicable to the biological activities of other lymphoid naïve B cells. Nonetheless, further biological study is warranted.

  13. Molecular cytogenetic characterization of a human thyroid cancercell line

    Energy Technology Data Exchange (ETDEWEB)

    Weier, Heinz-Ulrich G.; Tuton, Tiffany B.; Ito, Yuko; Chu, LisaW.; Lu, Chung-Mei; Baumgartner, Adolf; Zitzelsberger, Horst F.; Weier,Jingly F.

    2006-01-04

    The incidence of papillary thyroid carcinoma (PTC) increases significantly after exposure of the head and neck region to ionizing radiation, yet we know neither the steps involved in malignant transformation of thyroid epithelium nor the specific carcinogenic mode of action of radiation. Such increased tumor frequency became most evident in children after the 1986 nuclear accident in Chernobyl, Ukraine. In the twelve years following the accident, the average incidence of childhood PTCs (chPTC) increased over one hundred-fold compared to the rate of about 1 tumor incidence per 10{sup 6} children per year prior to 1986. To study the etiology of radiation-induced thyroid cancer, we formed an international consortium to investigate chromosomal changes and altered gene expression in cases of post-Chernobyl chPTC. Our approach is based on karyotyping of primary cultures established from chPTC specimens, establishment of cell lines and studies of genotype-phenotype relationships through high resolution chromosome analysis, DNA/cDNA micro-array studies, and mouse xenografts that test for tumorigenicity. Here, we report the application of fluorescence in situ hybridization (FISH)-based techniques for the molecular cytogenetic characterization of a highly tumorigenic chPTC cell line, S48TK, and its subclones. Using chromosome 9 rearrangements as an example, we describe a new approach termed ''BAC-FISH'' to rapidly delineate chromosomal breakpoints, an important step towards a better understanding of the formation of translocations and their functional consequences.

  14. Caffeine markedly sensitizes human mesothelioma cell lines to pemetrexed

    Science.gov (United States)

    Min, Sang Hee; Goldman, I. David; Zhao, Rongbao

    2013-01-01

    Pemetrexed is a new generation antifolate approved for the treatment of mesothelioma and non-small cell lung cancer. Caffeine is known to augment radiation or chemotherapeutic drug-induced cell killing. The current study addresses the impact of caffeine on the activity of pemetrexed in mesothelioma cell lines. Caffeine enhanced pemetrexed activity in all four mesothelioma cell lines tested (H2052, H2373, H28 and MSTO-211H). Caffeine sensitized H2052 cells in a dose- and schedule-dependent manner, and was associated with a markedly decreased clonogenic survival. Caffeine sensitization occurred only in cells subjected to pulse, but not continuous, exposure to pemetrexed. Similar pemetrexed sensitization was also observed with the clinically better tolerated caffeine analog, theobromine. Pemetrexed sensitization by caffeine was associated with an increase in pemetrexed-induced phosphorylation of ataxia-telangiectasia-mutated (ATM) and Chk1. These data indicate that caffeine and its analog, theobromine, may be a useful approach to enhance pemetrexed-based chemotherapy. PMID:17594092

  15. In vitro cytotoxicity of Indonesian stingless bee products against human cancer cell lines

    Directory of Open Access Journals (Sweden)

    Paula M. Kustiawan

    2014-07-01

    Conclusions: Propolis from T. incisa and Trigona fusco-balteata contain an in vitro cytotoxic activity against human cancer cell lines. Further study is required, including the isolation and characterization of the active antiproliferative agent(s.

  16. 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.

  17. Restoring balance to B cells in ADA deficiency.

    Science.gov (United States)

    Luning Prak, Eline T

    2012-06-01

    It is paradoxical that immunodeficiency disorders are associated with autoimmunity. Adenosine deaminase (ADA) deficiency, a cause of X-linked severe combined immunodeficiency (SCID), is a case in point. In this issue of the JCI, Sauer and colleagues investigate the B cell defects in ADA-deficient patients. They demonstrate that ADA patients receiving enzyme replacement therapy had B cell tolerance checkpoint defects. Remarkably, gene therapy with a retrovirus that expresses ADA resulted in the apparent correction of these defects, with normalization of peripheral B cell autoantibody frequencies. In vitro, agents that either block ADA or overexpress adenosine resulted in altered B cell receptor and TLR signaling. Collectively, these data implicate a B cell-intrinsic mechanism for alterations in B cell tolerance in the setting of partial ADA deficiency that is corrected by gene therapy.

  18. Radiobiological studies with a series of human cell lines of varying glutathione content

    International Nuclear Information System (INIS)

    Astor, M.B.

    1984-01-01

    Radiation responses of a series of four human fibroblast lines obtained from a family affected with 5-oxoprolinuria were determined. Cell suspensions were irradiated under hypoxic conditions and the oxygen enhancement ratio was determined for each cell line. Results are compared with previous studies

  19. Entry of Francisella tularensis into Murine B Cells: The Role of B Cell Receptors and Complement Receptors.

    Directory of Open Access Journals (Sweden)

    Lenka Plzakova

    Full Text Available Francisella tularensis, the etiological agent of tularemia, is an intracellular pathogen that dominantly infects and proliferates inside phagocytic cells but can be seen also in non-phagocytic cells, including B cells. Although protective immunity is known to be almost exclusively associated with the type 1 pathway of cellular immunity, a significant role of B cells in immune responses already has been demonstrated. Whether their role is associated with antibody-dependent or antibody-independent B cell functions is not yet fully understood. The character of early events during B cell-pathogen interaction may determine the type of B cell response regulating the induction of adaptive immunity. We used fluorescence microscopy and flow cytometry to identify the basic requirements for the entry of F. tularensis into B cells within in vivo and in vitro infection models. Here, we present data showing that Francisella tularensis subsp. holarctica strain LVS significantly infects individual subsets of murine peritoneal B cells early after infection. Depending on a given B cell subset, uptake of Francisella into B cells is mediated by B cell receptors (BCRs with or without complement receptor CR1/2. However, F. tularensis strain FSC200 ΔiglC and ΔftdsbA deletion mutants are defective in the ability to enter B cells. Once internalized into B cells, F. tularensis LVS intracellular trafficking occurs along the endosomal pathway, albeit without significant multiplication. The results strongly suggest that BCRs alone within the B-1a subset can ensure the internalization process while the BCRs on B-1b and B-2 cells need co-signaling from the co receptor containing CR1/2 to initiate F. tularensis engulfment. In this case, fluidity of the surface cell membrane is a prerequisite for the bacteria's internalization. The results substantially underline the functional heterogeneity of B cell subsets in relation to F. tularensis.

  20. Generation of hiPSTZ16 (ISMMSi003-A cell line from normal human foreskin fibroblasts

    Directory of Open Access Journals (Sweden)

    Marion Dejosez

    2018-01-01

    Full Text Available Human foreskin fibroblasts from a commercial source were reprogrammed into induced pluripotent stem cells to establish a clonal stem cell line, hiPSTZ16 (ISMMSi003-A. These cells show a normal karyotype and full differentiation potential in teratoma assays. The described cells provide a useful resource in combination with other iPS cell lines generated from normal human foreskin fibroblasts to study source- and reprogramming method-independent effects in downstream applications.

  1. Generation of iPSC line epiHUVEC from human umbilical vein endothelial cells

    Directory of Open Access Journals (Sweden)

    Peggy Matz

    2015-11-01

    Full Text Available Human umbilical vein endothelial cells (HUVECs were used to generate the iPSC line epiHUVEC employing a combination of three episomal-based plasmids expressing OCT4, SOX2, NANOG, LIN28, c-MYC and KLF4. Pluripotency was confirmed both in vivo and in vitro. The transcriptome profile of epiHUVEC and the human embryonic stem cell line — H1 have a Pearson correlation of 0.899.

  2. Generation of iPSC lines from primary human chorionic villi cells

    Directory of Open Access Journals (Sweden)

    Björn Lichtner

    2015-11-01

    Full Text Available Primary human chorionic villi (CV cells were used to generate the iPSC line by retroviral transduction of the four Yamanaka-factors OCT4, SOX2, KLF4 and c-MYC. Pluripotency was confirmed both in vivo and in vitro. The transcriptomes of the CV-derived iPSC lines and the human embryonic stem cell lines—H1 and H9 have a Pearson correlation of 0.929 and 0.943 respectively.

  3. Anti-B cell antibody therapies for inflammatory rheumatic diseases

    DEFF Research Database (Denmark)

    Faurschou, Mikkel; Jayne, David R W

    2014-01-01

    Several monoclonal antibodies targeting B cells have been tested as therapeutics for inflammatory rheumatic diseases. We review important observations from randomized clinical trials regarding the efficacy and safety of anti-B cell antibody-based therapies for rheumatoid arthritis, systemic lupus...... and functions in rheumatic disorders. Future studies should also evaluate how to maintain disease control by means of conventional and/or biologic immunosuppressants after remission-induction with anti-B cell antibodies....

  4. UV light blocks EGFR signalling in human cancer cell lines

    DEFF Research Database (Denmark)

    Olsen, BB; Neves-Petersen, M T; Klitgaard, S

    2007-01-01

    UV light excites aromatic residues, causing these to disrupt nearby disulphide bridges. The EGF receptor is rich in aromatic residues near the disulphide bridges. Herein we show that laser-pulsed UV illumination of two different skin-derived cancer cell lines i.e. Cal-39 and A431, which both...... antibodies. There was a threshold level, below which the receptor could not be blocked. In addition, illumination caused the cells to upregulate the cyclin-dependent kinase inhibitor p21WAF1, irrespective of the p53 status. Since the EGF receptor is often overexpressed in cancers and other proliferative skin...... disorders, it might be possible to significantly reduce the proliferative potential of these cells making them good targets for laser-pulsed UV light treatment....

  5. Metabolic Abnormalities Detected in Phase II Evaluation of Doxycycline in Dogs with Multicentric B-Cell Lymphoma.

    Science.gov (United States)

    Hume, Kelly R; Sylvester, Skylar R; Borlle, Lucia; Balkman, Cheryl E; McCleary-Wheeler, Angela L; Pulvino, Mary; Casulo, Carla; Zhao, Jiyong

    2018-01-01

    Doxycycline has antiproliferative effects in human lymphoma cells and in murine xenografts. We hypothesized that doxycycline would decrease canine lymphoma cell viability and prospectively evaluated its clinical tolerability in client-owned dogs with spontaneous, nodal, multicentric, substage a, B-cell lymphoma, not previously treated with chemotherapy. Treatment duration ranged from 1 to 8 weeks (median and mean, 3 weeks). Dogs were treated with either 10 ( n  = 6) or 7.5 ( n  = 7) mg/kg by mouth twice daily. One dog had a stable disease for 6 weeks. No complete or partial tumor responses were observed. Five dogs developed grade 3 and/or 4 metabolic abnormalities suggestive of hepatopathy with elevations in bilirubin, ALT, ALP, and/or AST. To evaluate the absorption of oral doxycycline in our study population, serum concentrations in 10 treated dogs were determined using liquid chromatography tandem mass spectrometry. Serum levels were variable and ranged from 3.6 to 16.6 µg/ml (median, 7.6 µg/ml; mean, 8.8 µg/ml). To evaluate the effect of doxycycline on canine lymphoma cell viability in vitro , trypan blue exclusion assay was performed on canine B-cell lymphoma cell lines (17-71 and CLBL) and primary B-cell lymphoma cells from the nodal tissue of four dogs. A doxycycline concentration of 6 µg/ml decreased canine lymphoma cell viability by 80%, compared to matched, untreated, control cells (mixed model analysis, p  canine lymphoma, combination therapy may be worthwhile if future research determines that doxycycline can alter cell survival pathways in canine lymphoma cells. Due to the potential for metabolic abnormalities, close monitoring is recommended with the use of this drug in tumor-bearing dogs. Additional research is needed to assess the tolerability of chronic doxycycline therapy.

  6. B cell repertoires in HLA-sensitized kidney transplant candidates undergoing desensitization therapy.

    Science.gov (United States)

    Beausang, John F; Fan, H Christina; Sit, Rene; Hutchins, Maria U; Jirage, Kshama; Curtis, Rachael; Hutchins, Edward; Quake, Stephen R; Yabu, Julie M

    2017-01-13

    Kidney transplantation is the most effective treatment for end-stage renal disease. Sensitization refers to pre-existing antibodies against human leukocyte antigen (HLA) protein and remains a major barrier to successful transplantation. Despite implementation of desensitization strategies, many candidates fail to respond. Our objective was to determine whether measuring B cell repertoires could differentiate candidates that respond to desensitization therapy. We developed an assay based on high-throughput DNA sequencing of the variable domain of the heavy chain of immunoglobulin genes to measure changes in B cell repertoires in 19 highly HLA-sensitized kidney transplant candidates undergoing desensitization and 7 controls with low to moderate HLA sensitization levels. Responders to desensitization had a decrease of 5% points or greater in cumulated calculated panel reactive antibody (cPRA) levels, and non-responders had no decrease in cPRA. Dominant B cell clones were not observed in highly sensitized candidates, suggesting that the B cells responsible for sensitization are either not present in peripheral blood or present at comparable levels to other circulating B cells. Candidates that responded to desensitization therapy had pre-treatment repertoires composed of a larger fraction of class-switched (IgG and IgA) isotypes compared to non-responding candidates. After B cell depleting therapy, the proportion of switched isotypes increased and the mutation frequencies of the remaining non-switched isotypes (IgM and IgD) increased in both responders and non-responders, perhaps representing a shift in the repertoire towards memory B cells or plasmablasts. Conversely, after transplantation, non-switched isotypes with fewer mutations increased, suggesting a shift in the repertoire towards naïve B cells. Relative abundance of different B cell isotypes is strongly perturbed by desensitization therapy and transplantation, potentially reflecting changes in the relative

  7. Leukemia - B-Cell Prolymphocytic Leukemia and Hairy Cell Leukemia

    Science.gov (United States)

    ... Leukemia - B-cell Prolymphocytic Leukemia and Hairy Cell Leukemia Introduction Statistics Risk Factors Symptoms and Signs Diagnosis Stages Treatment Options About Clinical Trials Latest Research ...

  8. 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.

  9. Regulatory T cells and B cells: implication on autoimmune diseases

    OpenAIRE

    Wang, Ping; Zheng, Song Guo

    2013-01-01

    The regulatory T (Treg) cells play an important role in the maintenance of homeostasis and the prevention of autoimmune diseases. Although most studies are focusing on the role of Treg cells in T cells and T cells-mediated diseases, these cells also directly affect B cells and other non-T cells. This manuscript updates the role of Treg cells on the B cells and B cell-mediated diseases. In addition, the mechanisms whereby Treg cells suppress B cell responses have been discussed.

  10. Antigen presentation by resting B cells. Radiosensitivity of the antigen-presentation function and two distinct pathways of T cell activation

    International Nuclear Information System (INIS)

    Ashwell, J.D.; DeFranco, A.L.; Paul, W.E.; Schwartz, R.H.

    1984-01-01

    In this report we have examined the ability of small resting B cells to act as antigen-presenting cells (APC) to antigen-specific MHC-restricted T cells as assessed by either T cell proliferation or T cell-dependent B cell stimulation. We found that 10 of 14 in vitro antigen-specific MHC-restricted T cell clones and lines and three of four T cell hybridomas could be induced to either proliferate or secrete IL-2 in the presence of lightly irradiated (1,000 rads) purified B cells and the appropriate foreign antigen. All T cell lines and hybridomas were stimulated to proliferate or make IL-2 by macrophage- and dendritic cell-enriched populations and all T cells tested except one hybridoma caused B cell activation when stimulated with B cells as APC. Furthermore, lightly irradiated, highly purified syngeneic B cells were as potent a source of APC for inducing B cell activation as were low density dendritic and macrophage-enriched cells. Lymph node T cells freshly taken from antigen-primed animals were also found to proliferate when cultured with purified B cells and the appropriate antigen. This APC function was easily measured when the cells were irradiated with 1,000 rads, but was greatly diminished or absent when they were irradiated with 3,300 rads. In addition, this radiosensitivity allowed us to easily distinguish B cell antigen presentation from presentation by the dendritic cell and macrophage, as the latter was resistant to 3,300 rads. Finally, one T cell clone that failed to proliferate when B cells were used as APC was able to recruit allogeneic B cells to proliferate in the presence of syngeneic B cells and the appropriate antigen. This result suggests that there are at least two distinct pathways of activation in T cells, one that leads to T cell proliferation and one that leads to the secretion of B cell recruitment factor(s)

  11. Clonal heterogeneity of thymic B cells from early-onset myasthenia gravis patients with antibodies against the acetylcholine receptor.

    Science.gov (United States)

    Vrolix, Kathleen; Fraussen, Judith; Losen, Mario; Stevens, Jo; Lazaridis, Konstantinos; Molenaar, Peter C; Somers, Veerle; Bracho, Maria Alma; Le Panse, Rozen; Stinissen, Piet; Berrih-Aknin, Sonia; Maessen, Jos G; Van Garsse, Leen; Buurman, Wim A; Tzartos, Socrates J; De Baets, Marc H; Martinez-Martinez, Pilar

    2014-08-01

    Myasthenia gravis (MG) with antibodies against the acetylcholine receptor (AChR-MG) is considered as a prototypic autoimmune disease. The thymus is important in the pathophysiology of the disease since thymus hyperplasia is a characteristic of early-onset AChR-MG and patients often improve after thymectomy. We hypothesized that thymic B cell and antibody repertoires of AChR-MG patients differ intrinsically from those of control individuals. Using immortalization with Epstein-Barr Virus and Toll-like receptor 9 activation, we isolated and characterized monoclonal B cell lines from 5 MG patients and 8 controls. Only 2 of 570 immortalized B cell clones from MG patients produced antibodies against the AChR (both clones were from the same patient), suggesting that AChR-specific B cells are not enriched in the thymus. Surprisingly, many B cell lines from both AChR-MG and control thymus samples displayed reactivity against striated muscle proteins. Striational antibodies were produced by 15% of B cell clones from AChR-MG versus 6% in control thymus. The IgVH gene sequence analysis showed remarkable similarities, concerning VH family gene distribution, mutation frequency and CDR3 composition, between B cells of AChR-MG patients and controls. MG patients showed clear evidence of clonal B cell expansion in contrast to controls. In this latter aspect, MG resembles multiple sclerosis and clinically isolated syndrome, but differs from systemic lupus erythematosus. Our results support an antigen driven immune response in the MG thymus, but the paucity of AChR-specific B cells, in combination with the observed polyclonal expansions suggest a more diverse immune response than expected. Copyright © 2013 Elsevier Ltd. All rights reserved.

  12. In Silico Prediction Analysis of Idiotope-Driven T–B Cell Collaboration in Multiple Sclerosis

    Directory of Open Access Journals (Sweden)

    Rune A. Høglund

    2017-10-01

    Full Text Available Memory B cells acting as antigen-presenting cells are believed to be important in multiple sclerosis (MS, but the antigen they present remains unknown. We hypothesized that B cells may activate CD4+ T cells in the central nervous system of MS patients by presenting idiotopes from their own immunoglobulin variable regions on human leukocyte antigen (HLA class II molecules. Here, we use bioinformatics prediction analysis of B cell immunoglobulin variable regions from 11 MS patients and 6 controls with other inflammatory neurological disorders (OINDs, to assess whether the prerequisites for such idiotope-driven T–B cell collaboration are present. Our findings indicate that idiotopes from the complementarity determining region (CDR 3 of MS patients on average have high predicted affinities for disease associated HLA-DRB1*15:01 molecules and are predicted to be endosomally processed by cathepsin S and L in positions that allows such HLA binding to occur. Additionally, complementarity determining region 3 sequences from cerebrospinal fluid (CSF B cells from MS patients contain on average more rare T cell-exposed motifs that could potentially escape tolerance and stimulate CD4+ T cells than CSF B cells from OIND patients. Many of these features were associated with preferential use of the IGHV4 gene family by CSF B cells from MS patients. This is the first study to combine high-throughput sequencing of patient immune repertoires with large-scale prediction analysis and provides key indicators for future in vitro and in vivo analyses.

  13. Interferon-alpha triggers B cell effector 1 (Be1 commitment.

    Directory of Open Access Journals (Sweden)

    Marie-Ghislaine de Goër de Herve

    Full Text Available B-cells can contribute to the pathogenesis of autoimmune diseases not only through auto-antibody secretion but also via cytokine production. Therapeutic depletion of B-cells influences the functions and maintenance of various T-cell subsets. The mechanisms governing the functional heterogeneity of B-cell subsets as cytokine-producing cells are poorly understood. B-cells can differentiate into two functionally polarized effectors, one (B-effector-1-cells producing a Th-1-like cytokine pattern and the other (Be2 producing a Th-2-like pattern. IL-12 and IFN-γ play a key role in Be1 polarization, but the initial trigger of Be1 commitment is unclear. Type-I-interferons are produced early in the immune response and prime several processes involved in innate and adaptive responses. Here, we report that IFN-α triggers a signaling cascade in resting human naive B-cells, involving STAT4 and T-bet, two key IFN-γ gene imprinting factors. IFN-α primed naive B-cells for IFN-γ production and increased IFN-γ gene responsiveness to IL-12. IFN-γ continues this polarization by re-inducing T-bet and up-regulating IL-12Rβ2 expression. IFN-α and IFN-γ therefore pave the way for the action of IL-12. These results point to a coordinated action of IFN-α, IFN-γ and IL-12 in Be1 polarization of naive B-cells, and may provide new insights into the mechanisms by which type-I-interferons favor autoimmunity.

  14. Derivation of Huntington Disease affected Genea046 human embryonic stem cell line

    Directory of Open Access Journals (Sweden)

    Biljana Dumevska

    2016-03-01

    Full Text Available The Genea046 human embryonic stem cell line was derived from a donated, fully commercially consented ART blastocyst, carrying HTT gene CAG expansion of 45 repeats, indicative of Huntington Disease. Following ICM outgrowth on inactivated human feeders, karyotype was confirmed as 46, XX by CGH and STR analysis demonstrated a female Allele pattern. The hESC line had pluripotent cell morphology, 85% of cells expressed Nanog, 92% Oct4, 75% Tra1–60 and 99% SSEA4 and demonstrated Alkaline Phosphatase activity. The cell line was negative for Mycoplasma and visible contamination.

  15. Effect of recombinant adenovirus encoding human p53 tumor suppressor gene combined with radiation therapy on human lymphoma cells lines

    International Nuclear Information System (INIS)

    Yu Zeyang; Fan Wo; Li Dongqing; Zhu Ran; Wan Jianmei; Wang Yongqing; Wu Jinchang

    2008-01-01

    This paper analyzes the inhibitory effect and radiation sensitization of recombinant adenovirus encoding human p53 tumor suppressor gene (rAd-p53) on human lymphoma cell lines. Human lymphoma cell lines were treated with rAd-p53, radiation therapy and combined treatment, respectively. The cell growth inhibition was assessed by MTF. The cell cycle and apoptosis were detected by flow cytometry, and the p53 protein expression was detected by Western blotting. The results showed that extrinsic p53 gene have expressed to some degree, but not at high level. The role of inhibition and radiation sensitivity of rAd-p53 was not significant to human lymphoma cell lines. (authors)

  16. Identifying anti-growth factors for human cancer cell lines through genome-scale metabolic modeling

    DEFF Research Database (Denmark)

    Ghaffari, Pouyan; Mardinoglu, Adil; Asplund, Anna

    2015-01-01

    Human cancer cell lines are used as important model systems to study molecular mechanisms associated with tumor growth, hereunder how genomic and biological heterogeneity found in primary tumors affect cellular phenotypes. We reconstructed Genome scale metabolic models (GEMs) for eleven cell lines...... based on RNA-Seq data and validated the functionality of these models with data from metabolite profiling. We used cell line-specific GEMs to analyze the differences in the metabolism of cancer cell lines, and to explore the heterogeneous expression of the metabolic subsystems. Furthermore, we predicted...... for inhibition of cell growth may provide leads for the development of efficient cancer treatment strategies....

  17. The role of complement receptors type 1 (CR1, CD35) and 2 (CR2, CD21) in promoting C3 fragment deposition and membrane attack complex formation on normal peripheral human B cells

    DEFF Research Database (Denmark)

    Nielsen, Claus Henrik; Pedersen, Morten Løbner; Marquart, Hanne Vibeke Hansen

    2002-01-01

    Normal human B lymphocytes are known to activate the alternative pathway (AP) of complement, leading to C3-fragment deposition and membrane attack complex (MAC) formation. The process is mediated via complement receptor type 2 (CR2, CD21), with complement receptor type 1 (CR1, CD35) playing...... a subsidiary role. In this study, we examine the relative contributions of CR1 and CR2 to the deposition of C3 fragments and MAC on B lymphocytes under circumstances where all complement pathways are operational. C3-fragment deposition and MAC formation were assessed on human peripheral B lymphocytes...... in the presence of 30% autologous serum. Blocking the CR2 ligand-binding site with monoclonal antibody (mAb) FE8 resulted in significant reduction (37.9+/-11.9%) in C3-fragment deposition, whereas MAC formation was only marginally affected (12.1+/-22.2% reduction). Blocking the CR1 binding-site resulted...

  18. Natural autoantibodies and complement promote the uptake of a self antigen, human thyroglobulin, by B cells and the proliferation of thyroglobulin-reactive CD4(+) T cells in healthy individuals

    DEFF Research Database (Denmark)

    Nielsen, C H; Leslie, R G; Jepsen, B S

    2001-01-01

    Serum from normal individuals contains substantial amounts of natural antibodies (NA) capable of recognizing self antigens. However, the physiological implications of this autoreactivity remain unclear. We have examined the role of self-reactive NA and complement in mediating the uptake of human...... cells are prerequisites for the proliferation of Tg-reactive CD4(+) T cells, suggesting a novel role for natural autoantibodies and complement in the regulation of autoreactivity under physiological conditions....

  19. B-cell-mediated strategies to fight chronic allograft rejection

    Directory of Open Access Journals (Sweden)

    Ali H Dalloul

    2013-12-01

    Full Text Available Solid organs have been transplanted for decades. Since the improvement in graft selection and in medical and surgical procedures, the likelihood of graft function after one year is now close to 90%. Nonetheless even well-matched recipients continue to need medications for the rest of their lives hence adverse side effects and enhanced morbidity. Understanding Immune rejection mechanisms, is of increasing importance since the greater use of living-unrelated donors and genetically unmatched individuals. Chronic rejection is devoted to T-cells, however the role of B-cells in rejection has been appreciated recently by the observation that B-cell depletion improve graft survival. By contrast however, B-cells can be beneficial to the grafted tissue. This protective effect is secondary to either the secretion of protective antibodies or the induction of B-cells that restrain excessive inflammatory responses, chiefly by local provision of IL-10, or inhibit effector T-cells by direct cellular interactions. As a proof of concept B-cell-mediated infectious transplantation tolerance could be achieved in animal models, and evidence emerged that the presence of such B-cells in transplanted patients correlate with a favorable outcome. Among these populations, regulatory B-cells constitute a recently described population. These cells may develop as a feedback mechanism to prevent uncontrolled reactivity to antigens and inflammatory stimuli. The difficult task for the clinician, is to quantify the respective ratios and functions of tolerant vs effector B-cells within a transplanted organ, at a given time point in order to modulate B-cell-directed therapy. Several receptors at the B-cell membrane as well as signaling molecules, can now be targeted for this purpose. Understanding the temporal expansion of regulatory B-cells in grafted patients and the stimuli that activate them will help in the future to implement specific strategies aimed at fighting chronic

  20. How B cells influence bone biology in health and disease.

    Science.gov (United States)

    Horowitz, Mark C; Fretz, Jackie A; Lorenzo, Joseph A

    2010-09-01

    It is now well established that important regulatory interactions occur between the cells in the hematopoietic, immune and skeletal systems (osteoimmunology). B lymphocytes (B cells) are responsible for the generation and production of antibodies or immunoglobulins in the body. Together with T cells these lymphocytes comprise the adaptive immune system, which allows an individual to develop specific responses to an infection and retain memory of that infection, allowing for a faster and more robust response if that same infection occurs again. In addition to this immune function, B cells have a close and multifaceted relationship with bone cells. B cells differentiate from hematopoietic stem cells (HSCs) in supportive niches found on endosteal bone surfaces. Cells in the osteoblast lineage support HSC and B cell differentiation in these niches. B cell differentiation is regulated, at least in part, by a series of transcription factors that function in a temporal manner. While these transcription factors are required for B cell differentiation, their loss causes profound changes in the bone phenotype. This is due, in part, to the close relationship between macrophage/osteoclast and B cell differentiation. Cross talk between B cells and bone cells is reciprocal with defects in the RANKL-RANK, OPG signaling axis resulting in altered bone phenotypes. While the role of B cells during normal bone remodeling appears minimal, activated B cells play an important role in many inflammatory diseases with associated bony changes. This review examines the relationship between B cells and bone cells and how that relationship affects the skeleton and hematopoiesis during health and disease. Copyright 2010 Elsevier Inc. All rights reserved.

  1. Tissue-specific B-cell dysfunction and generalized memory B-cell loss during acute SIV infection.

    Directory of Open Access Journals (Sweden)

    Sandrine Peruchon

    Full Text Available BACKGROUND: Primary HIV-infected patients display severe and irreversible damage to different blood B-cell subsets which is not restored by highly efficient anti-retroviral therapy (HAART. Because longitudinal investigations of primary HIV-infection is limited by the availability of lymphoid organs, we studied the tissue-specific B-cell dysfunctions in acutely simian immunodeficiency virus (SIV mac251-infected Cynomolgus macaques. METHODS AND FINDINGS: Experiments were performed on three groups of macaques infected for 14, 21 or 28 days and on three groups of animals treated with HAART for two-weeks either initiated at 4 h, 7 or 14 days post-infection (p.i.. We have simultaneously compared changes in B-cell phenotypes and functions and tissue organization of B-cell areas in various lymphoid organs. We showed that SIV induced a steady decline in SIgG-expressing memory (SIgD(-CD27(+ B-cells in spleen and lymph nodes during the first 4 weeks of infection, concomitant to selective homing/sequestration of B-cells to the small intestine and spleen. SIV non-specific Ig production was transiently increased before D14p.i., whereas SIV-specific Ig production was only detectable after D14p.i., coinciding with the presence of CD8(+ T-cells and IgG-expressing plasma cells within germinal centres. Transient B-cell apoptosis on D14p.i. and commitment to terminal differentiation contributed to memory B-cell loss. HAART abrogated B-cell apoptosis, homing to the small intestine and SIV-specific Ig production but had minimal effect on early Ig production, increased B-cell proportions in spleen and loss of memory B-cells. Therefore, virus-B-cell interactions and SIV-induced inflammatory cytokines may differently contribute to early B-cell dysfunction and impaired SIV/HIV-specific antibody response. CONCLUSIONS: These data establish tissue-specific impairments in B-cell trafficking and functions and a generalized and steady memory B-cell loss in secondary lymphoid

  2. [The characters and specific features of new human embryonic stem cells lines].

    Science.gov (United States)

    Krylova, T A; Kol'tsova, A M; Zenin, V V; Gordeeva, O F; Musorina, A S; Goriachaia, T S; Shlykova, S A; Kamenetskaia, Iu K; Pinaev, G P; Polianskaia, G G

    2009-01-01

    Four continuous human embryonic stem cell lines (SC1, SC2, SC3 and SC4), derived from the blastocysts has been described. The cell lines were cultivated on mitotically inactivated human feeder cells. The cell lines SC1 and SC2 have passed through 150 population doublings and the cell lines SC3 and SC4 -- near 120 populations doublings, which exceeds Hayflick limit sufficiently. These cell lines maintain high activity of alkaline phosphatase, expression of transcription factor OCT-4 and cell surface antigens (SSEA-4, TRA-1-60 and TRA-1-81), confirming their ESC status and human specificity. Immunofluorescent detection of antigens, characteristic of ectoderm, endoderm and mesoderm confirms the ability of these cells to retain their pluripotency under in vitro condition. PCR analysis revealed expression of six genes specific for pluripotent cells (OCT-4, NANOG, DPPA3/STELLA, TDGF/CRIPTO and LEFTYA). Correlation between the level of proliferative activity and the character of DNA-bound fluorescent staining was found. Fluorescent dyes, Hoechst 33342 and PI, produced diffuse staining of the nuclei in slowly proliferating cells of the SC1 and SC2 lines. In contrast, in actively proliferating cells of the SC3 and SC4 lines, the clear staining of the nuclei was observed. Upon changing the cultivation condition, proliferative activity of SC3 and SC4 lines decreased and became similar to that of SC1 and SC2 lines. The character of the fluorescent staining of all these lines was also shown to be similar. These results show that quality of the fluorescent staining with Hoechst 33342 and PI reflects the level of proliferation. Possible causes and mechanisms of this feature of human ESC are discussed.

  3. Epigenetic alterations differ in phenotypically distinct human neuroblastoma cell lines

    International Nuclear Information System (INIS)

    Yang, Qiwei; Tian, Yufeng; Ostler, Kelly R; Chlenski, Alexandre; Guerrero, Lisa J; Salwen, Helen R; Godley, Lucy A; Cohn, Susan L

    2010-01-01

    Epigenetic aberrations and a CpG island methylator phenotype have been shown to be associated with poor outcomes in children with neuroblastoma (NB). Seven cancer related genes (THBS-1, CASP8, HIN-1, TIG-1, BLU, SPARC, and HIC-1) that have been shown to have epigenetic changes in adult cancers and play important roles in the regulation of angiogenesis, tumor growth, and apoptosis were analyzed to investigate the role epigenetic alterations play in determining NB phenotype. Two NB cell lines (tumorigenic LA1-55n and non-tumorigenic LA1-5s) that differ in their ability to form colonies in soft agar and tumors in nude mice were used. Quantitative RNA expression analyses were performed on seven genes in LA1-5s, LA1-55n and 5-Aza-dC treated LA1-55n NB cell lines. The methylation status around THBS-1, HIN-1, TIG-1 and CASP8 promoters was examined using methylation specific PCR. Chromatin immunoprecipitation assay was used to examine histone modifications along the THBS-1 promoter. Luciferase assay was used to determine THBS-1 promoter activity. Cell proliferation assay was used to examine the effect of 5-Aza-dC on NB cell growth. The soft agar assay was used to determine the tumorigenicity. Promoter methylation values for THBS-1, HIN-1, TIG-1, and CASP8 were higher in LA1-55n cells compared to LA1-5s cells. Consistent with the promoter methylation status, lower levels of gene expression were detected in the LA1-55n cells. Histone marks associated with repressive chromatin states (H3K9Me3, H3K27Me3, and H3K4Me3) were identified in the THBS-1 promoter region in the LA1-55n cells, but not the LA1-5s cells. In contrast, the three histone codes associated with an active chromatin state (acetyl H3, acetyl H4, and H3K4Me3) were present in the THBS-1 promoter region in LA1-5s cells, but not the LA1-55n cells, suggesting that an accessible chromatin structure is important for THBS-1 expression. We also show that 5-Aza-dC treatment of LA1-55n cells alters the DNA methylation

  4. Biobanking human embryonic stem cell lines: policy, ethics and efficiency.

    Science.gov (United States)

    Holm, Søren

    2015-12-01

    Stem cell banks curating and distributing human embryonic stem cells have been established in a number of countries and by a number of private institutions. This paper identifies and critically discusses a number of arguments that are used to justify the importance of such banks in policy discussions relating to their establishment or maintenance. It is argued (1) that 'ethical arguments' are often more important in the establishment phase and 'efficiency arguments' more important in the maintenance phase, and (2) that arguments relating to the interests of embryo and gamete donors are curiously absent from the particular stem cell banking policy discourse. This to some extent artificially isolates this discourse from the broader discussions about the flows of reproductive materials and tissues in modern society, and such isolation may lead to the interests of important actors being ignored in the policy making process.

  5. An Exponential Regression Model Reveals the Continuous Development of B Cell Subpopulations Used as Reference Values in Children

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    Christoph Königs

    2018-05-01

    Full Text Available B lymphocytes are key players in humoral immunity, expressing diverse surface immunoglobulin receptors directed against specific antigenic epitopes. The development and profile of distinct subpopulations have gained awareness in the setting of primary immunodeficiency disorders, primary or secondary autoimmunity and as therapeutic targets of specific antibodies in various diseases. The major B cell subpopulations in peripheral blood include naïve (CD19+ or CD20+IgD+CD27−, non-switched memory (CD19+ or CD20+IgD+CD27+ and switched memory B cells (CD19+ or CD20+IgD−CD27+. Furthermore, less common B cell subpopulations have also been described as having a role in the suppressive capacity of B cells to maintain self-tolerance. Data on reference values for B cell subpopulations are limited and only available for older age groups, neglecting the continuous process of human B cell development in children and adolescents. This study was designed to establish an exponential regression model to produce continuous reference values for main B cell subpopulations to reflect the dynamic maturation of the human immune system in healthy children.

  6. Radiosensitization of human prostate cell line LNCAP by [6]- gingerol

    Energy Technology Data Exchange (ETDEWEB)

    Silva, Josias Paulino Leal; Bellini, Maria Helena [Instituto de Pesquisas Energéticas e Nucleares (IPEN/CNEN-SP), São Paulo, SP (Brazil)

    2017-07-01

    Full text: Introduction: Prostate cancer is the second most prevalent malignancy and second leading cause of cancer-related deaths among men in the world. Several different diagnostic and therapeutic approaches have been developed in order to decrease the death rates. A number of experimental and clinical studies have showed antiproliferative, pro-apoptotic, anti-metastatic and anti-angiogenic effects of several phytochemicals. [6]-Gingerol (1-[4'-hydroxy-3'-methoxyphenyl]-5-hydroxy-3- decanone), the major pungent principle of ginger, has anti-oxidant, anti-inflammation and antitumor promoting activities. Aim: The purpose of this study was to evaluate the radiosensitizing activity of [6]-Gingerol in the human prostate cancer cells. Methods: The viability was assessed (3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl tetrazolium bromide) tetrazolium (MTS) assay. The prostate human cells (LNCAP) (2,5×103 cells/well) were seeded into 96-well plates, after 24 hr they were treated with 150 and 300μg/mL of [6]-Gingerol or vehicle alone (0.1% DMSO) in serum containing media. After incubation, MTS solution was added to the plate at a final concentration of 0.5 mg/mL. The cells were incubated for 2 hr in dark at 37. The resulting MTS-products were determined by measuring the absorbance at 490 nm with ELISA reader. In the clonogenic cell survival assay, the cells were divided into two groups: A) control, B) treated with [6]-Gingerol, C) irradiated control and D) treated with [6]-Gingerol and irradiated. The cells were irradiated by 60Co source in the range from 0 to 15 Gy, using the GammaCell 220 - Irradiation Unit of Canadian-Atomic Energy Commision Ltd. (CTR-IPEN). After 10-14 days of culture in normoxia conditions, cell colonies were fixed and stained with methanol 20% and crystal violet 0.5% and counted. Multiple comparisons were assessed by One-way ANOVA followed by Bonferroni´s tests with GraphPad Prism version 6.0 software. p< 0.05 was considered statistically

  7. Radiosensitization of human prostate cell line LNCAP by [6]- gingerol

    International Nuclear Information System (INIS)

    Silva, Josias Paulino Leal; Bellini, Maria Helena

    2017-01-01

    Full text: Introduction: Prostate cancer is the second most prevalent malignancy and second leading cause of cancer-related deaths among men in the world. Several different diagnostic and therapeutic approaches have been developed in order to decrease the death rates. A number of experimental and clinical studies have showed antiproliferative, pro-apoptotic, anti-metastatic and anti-angiogenic effects of several phytochemicals. [6]-Gingerol (1-[4'-hydroxy-3'-methoxyphenyl]-5-hydroxy-3- decanone), the major pungent principle of ginger, has anti-oxidant, anti-inflammation and antitumor promoting activities. Aim: The purpose of this study was to evaluate the radiosensitizing activity of [6]-Gingerol in the human prostate cancer cells. Methods: The viability was assessed (3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl tetrazolium bromide) tetrazolium (MTS) assay. The prostate human cells (LNCAP) (2,5×103 cells/well) were seeded into 96-well plates, after 24 hr they were treated with 150 and 300μg/mL of [6]-Gingerol or vehicle alone (0.1% DMSO) in serum containing media. After incubation, MTS solution was added to the plate at a final concentration of 0.5 mg/mL. The cells were incubated for 2 hr in dark at 37. The resulting MTS-products were determined by measuring the absorbance at 490 nm with ELISA reader. In the clonogenic cell survival assay, the cells were divided into two groups: A) control, B) treated with [6]-Gingerol, C) irradiated control and D) treated with [6]-Gingerol and irradiated. The cells were irradiated by 60Co source in the range from 0 to 15 Gy, using the GammaCell 220 - Irradiation Unit of Canadian-Atomic Energy Commision Ltd. (CTR-IPEN). After 10-14 days of culture in normoxia conditions, cell colonies were fixed and stained with methanol 20% and crystal violet 0.5% and counted. Multiple comparisons were assessed by One-way ANOVA followed by Bonferroni´s tests with GraphPad Prism version 6.0 software. p< 0.05 was considered statistically

  8. DNA breaks early in replication in B cell cancers

    Science.gov (United States)

    Research by scientists at the NCI has identified a new class of DNA sites in cells that break early in the replication process. They found that these break sites correlate with damage often seen in B cell cancers, such as diffuse large B cell lymphoma.

  9. Identification of cDNA encoding an additional α subunit of a human GTP-binding protein: Expression of three αi subtypes in human tissues and cell lines

    International Nuclear Information System (INIS)

    Kim, S.; Ang, S.L.; Bloch, D.B.; Bloch, K.D.; Kawahara, Y.; Tolman, C.; Lee, R.; Seidman, J.G.; Neer, E.J.

    1988-01-01

    The guanine nucleotide-binding proteins (G proteins), which mediate hormonal regulation of many membrane functions, are composed of α, β, and γ subunits. The authors have cloned and characterized cDNA from a human T-cell library encoding a form of α i that is different from the human α i subtypes previously reported. α i is the α subunit of a class of G proteins that inhibits adenylate cyclase and regulates other enzymes and ion channels. This cDNA encodes a polypeptide of 354 amino acids and is assigned to encode the α i-3 subtype of G proteins on the basis of its similarity to other α i -like cDNAs and the presence of a predicted site for ADP ribosylation by pertussis toxin. They have determined the expression of mRNA for this and two other subtypes of human α i (α i-1 and α i-2 ) in a variety of human fetal tissues and in human cell lines. All three α i subtypes were present in the tissues tested. However, analysis of individual cell types reveals specificity of α i-1 expression. mRNA for α i-1 is absent in T cells, B cells, and monocytes but is present in other cell lines. The finding of differential expression of α i-1 genes may permit characterization of distinct physiological roles for this α i subunit. mRNA for α i-2 and α i-3 was found in all the primary and transformed cell lines tested. Thus, some cells contain all three α i subtypes. This observation raises the question of how cells prevent cross talk among receptors that are coupled to effectors through such similar α proteins

  10. B cell signature during inactive systemic lupus is heterogeneous: toward a biological dissection of lupus.

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    Jean-Claude Garaud

    Full Text Available Systemic lupus erythematosous (SLE is an autoimmune disease with an important clinical and biological heterogeneity. B lymphocytes appear central to the development of SLE which is characterized by the production of a large variety of autoantibodies and hypergammaglobulinemia. In mice, immature B cells from spontaneous lupus prone animals are able to produce autoantibodies when transferred into immunodeficient mice, strongly suggesting the existence of intrinsic B cell defects during lupus. In order to approach these defects in humans, we compared the peripheral B cell transcriptomas of quiescent lupus patients to normal B cell transcriptomas. When the statistical analysis is performed on the entire group of patients, the differences between patients and controls appear quite weak with only 14 mRNA genes having a false discovery rate ranging between 11 and 17%, with 6 underexpressed genes (PMEPA1, TLR10, TRAF3IP2, LDOC1L, CD1C and EGR1. However, unforced hierarchical clustering of the microarrays reveals a subgroup of lupus patients distinct from both the controls and the other lupus patients. This subgroup has no detectable clinical or immunological phenotypic peculiarity compared to the other patients, but is characterized by 1/an IL-4 signature and 2/the abnormal expression of a large set of genes with an extremely low false discovery rate, mainly pointing to the biological function of the endoplasmic reticulum, and more precisely to genes implicated in the Unfolded Protein Response, suggesting that B cells entered an incomplete BLIMP1 dependent plasmacytic differentiation which was undetectable by immunophenotyping. Thus, this microarray analysis of B cells during quiescent lupus suggests that, despite a similar lupus phenotype, different biological roads can lead to human lupus.

  11. Murid herpesvirus-4 exploits dendritic cells to infect B cells.

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    Miguel Gaspar

    2011-11-01

    Full Text Available Dendritic cells (DCs play a central role in initiating immune responses. Some persistent viruses infect DCs and can disrupt their functions in vitro. However, these viruses remain strongly immunogenic in vivo. Thus what role DC infection plays in the pathogenesis of persistent infections is unclear. Here we show that a persistent, B cell-tropic gamma-herpesvirus, Murid Herpesvirus-4 (MuHV-4, infects DCs early after host entry, before it establishes a substantial infection of B cells. DC-specific virus marking by cre-lox recombination revealed that a significant fraction of the virus latent in B cells had passed through a DC, and a virus attenuated for replication in DCs was impaired in B cell colonization. In vitro MuHV-4 dramatically altered the DC cytoskeleton, suggesting that it manipulates DC migration and shape in order to spread. MuHV-4 therefore uses DCs to colonize B cells.

  12. Response of the MG-63 human osteosarcoma cell line grown as multicellular spheroids to neutron irradiation

    International Nuclear Information System (INIS)

    Kubota, Nobuo; Kakehi, Masae; Matsubara, Shou; Koike, Sachiko; Ando, Koichi.

    1993-01-01

    Multicellular tumor spheroids are composed of the mixed populations of cells with regard to cell proliferation, nutrition, oxygenation and radiosensitivity. Human osteogenic sarcoma is generally considered clinically radioresistant. However, the in vitro cell survival curves for human osteogenic sarcoma cell lines do not differ from those of other tumor cell lines. In this study, the responses of human osteogenic sarcoma cell line to gamma ray and neutrons were investigated by using spheroid system. The spheroids of the osteogenic sarcoma cell line are considered to be a good in vitro model of radioresistant tumors. The purpose of this study is to measure the response of the spheroids to fast neutron irradiation. MG-63 human osteogenic sarcoma cell line was used for this study. The cell line was cultured in alpha-MEM with supplement. Cell survival was estimated after the trypsinization of spheroids 24 hours after irradiation. The method of measuring spheroid cure is explained. The mean number of surviving cells per spheroid can be obtained from the mean clonogenic number and cell survival curve. The cell survival of MG-63 spheroids exposed to gamma ray and neutrons and the dose effect curves for spheroid cure after irradiation are shown. (K.I.)

  13. Dihydrochalcone Compounds Isolated from Crabapple Leaves Showed Anticancer Effects on Human Cancer Cell Lines

    Directory of Open Access Journals (Sweden)

    Xiaoxiao Qin

    2015-11-01

    Full Text Available Seven dihydrochalcone compounds were isolated from the leaves of Malus crabapples, cv. “Radiant”, and their chemical structures were elucidated by UV, IR, ESI-MS, 1H-NMR and 13C-NMR analyses. These compounds, which include trilobatin (A1, phloretin (A2, 3-hydroxyphloretin (A3, phloretin rutinoside (A4, phlorizin (A5, 6′′-O-coumaroyl-4′-O-glucopyranosylphloretin (A6, and 3′′′-methoxy-6′′-O-feruloy-4′-O-glucopyranosyl-phloretin (A7, all belong to the phloretin class and its derivatives. Compounds A6 and A7 are two new rare dihydrochalcone compounds. The results of a MTT cancer cell growth inhibition assay demonstrated that phloretin and these derivatives showed significant positive anticancer activities against several human cancer cell lines, including the A549 human lung cancer cell line, Bel 7402 liver cancer cell line, HepG2 human ileocecal cancer cell line, and HT-29 human colon cancer cell line. A7 had significant effects on all cancer cell lines, suggesting potential applications for phloretin and its derivatives. Adding a methoxyl group to phloretin dramatically increases phloretin’s anticancer activity.

  14. Feasibility of drug screening with panels of human tumor cell lines using a microculture tetrazolium assay.

    Science.gov (United States)

    Alley, M C; Scudiero, D A; Monks, A; Hursey, M L; Czerwinski, M J; Fine, D L; Abbott, B J; Mayo, J G; Shoemaker, R H; Boyd, M R

    1988-02-01

    For the past 30 years strategies for the preclinical discovery and development of potential anticancer agents have been based largely upon the testing of agents in mice bearing transplantable leukemias and solid tumors derived from a limited number of murine as well as human sources. The feasibility of implementing an alternate approach, namely combined in vitro/in vivo screening for selective cytotoxicity among panels of human tumor cell lines derived from a broad spectrum of human solid tumors is under investigation. A group of 30 cell lines acquired from a variety of sources and representing 8 lung cancer pathologies as well as 76 cell lines representing 10 other categories of human cancer (carcinomas of colon, breast, kidney, prostate, ovary, head and neck; glioma; leukemia; melanoma; and sarcoma) have exhibited acceptable growth characteristics and suitable colorimetric profiles in a single, standard culture medium. Measurements of in vitro growth in microculture wells by cell-mediated reduction of tetrazolium showed excellent correlation (0.89 less than r2 less than 0.98) with measurements of cellular protein in adherent cell line cultures as well as viable cell count in suspension cell line cultures (0.94 less than r2 less than 0.99). Since the microculture tetrazolium assay provides sensitive and reproducible indices of growth as well as drug sensitivity in individual cell lines over the course of multiple passages and several months' cultivation, it appears suitable for initial-stage in vitro drug screening.

  15. Can resting B cells present antigen to T cells

    International Nuclear Information System (INIS)

    Ashwell, J.D.; DeFranco, A.L.; Paul, W.E.; Schwartz, R.H.

    1985-01-01

    Antigen stimulation of T lymphocytes can occur only in the presence of an antigen-presenting cell (APC). An ever-increasing number of cell types have been found to act as APCs; these include macrophages, splenic and lymph node dendritic cells, and Langerhans cells of the skin. Although activated B lymphocytes and B cell lymphomas are known to serve as APCs, it has been generally believed that resting B cells cannot perform this function. However, in recent studies the authors have found that resting B cells can indeed present soluble antigen to T cell clones as well as to antigen-primed T cells. The previous difficulty in demonstrating this activity can be explained by the finding that, in contrast to macrophages and dendritic cells, the antigen-presenting ability of resting B cells is very radiosensitive. Macrophages are usually irradiated with 2000-3300 rads to prevent them from incorporating [ 3 H]thymidine in the T cell proliferation assay. Resting B cells, however, begin to lose presenting function at 1500 rads and have completely lost this activity at 3300 rads. It was also possible to distinguish two distinct T cell clonal phenotypes when resting B cells were used as APCs on the basis of two different assays (T cell proliferation, and B cell proliferation resulting from T cell activation). The majority of T cell clones tested were capable of both proliferating themselves and inducing the proliferation of B cells. Some T cells clones, however, could not proliferate in the presence of antigen and B cell APCs, although they were very good at inducing the proliferation of B cells

  16. Acute Liver Injury Is Independent of B Cells or Immunoglobulin M.

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    James A Richards

    Full Text Available Acute liver injury is a clinically important pathology and results in the release of Danger Associated Molecular Patterns, which initiate an immune response. Withdrawal of the injurious agent and curtailing any pathogenic secondary immune response may allow spontaneous resolution of injury. The role B cells and Immunoglobulin M (IgM play in acute liver injury is largely unknown and it was proposed that B cells and/or IgM would play a significant role in its pathogenesis.Tissue from 3 models of experimental liver injury (ischemia-reperfusion injury, concanavalin A hepatitis and paracetamol-induced liver injury and patients transplanted following paracetamol overdose were stained for evidence of IgM deposition. Mice deficient in B cells (and IgM were used to dissect out the role B cells and/or IgM played in the development or resolution of injury. Serum transfer into mice lacking IgM was used to establish the role IgM plays in injury.Significant deposition of IgM was seen in the explanted livers of patients transplanted following paracetamol overdose as well as in 3 experimental models of acute liver injury (ischemia-reperfusion injury, concanavalin A hepatitis and paracetamol-induced liver injury. Serum transfer into IgM-deficient mice failed to reconstitute injury (p = 0.66, despite successful engraftment of IgM. Mice deficient in both T and B cells (RAG1-/- mice (p<0.001, but not B cell deficient (μMT mice (p = 0.93, were significantly protected from injury. Further interrogation with T cell deficient (CD3εKO mice confirmed that the T cell component is a key mediator of sterile liver injury. Mice deficient in B cells and IgM mice did not have a significant delay in resolution following acute liver injury.IgM deposition appears to be common feature of both human and murine sterile liver injury. However, neither IgM nor B cells, play a significant role in the development of or resolution from acute liver injury. T cells appear to be key

  17. Cytotoxic activity of kenaf (Hibiscus cannabinus L.) seed extract and oil against human cancer cell lines

    Science.gov (United States)

    Wong, Yu Hua; Tan, Wai Yan; Tan, Chin Ping; Long, Kamariah; Nyam, Kar Lin

    2014-01-01

    Objective To examine the cytotoxic properties of both the kenaf (Hibiscus cannabinus L.) seed extract and kenaf seed oil on human cervical cancer, human breast cancer, human colon cancer and human lung cancer cell lines. Methods The in vitro cytotoxic activity of the kenaf (Hibiscus cannabinus L.) seed extract and kenaf seed oil on human cancer cell lines was evaluated by using 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide and sulforhodamine B assays. Cell morphological changes were observed by using an inverted light microscope. Results The kenaf seed extract (KSE) exhibited a lower IC50 than kenaf seed oil (KSO) in all of the cancer cell lines. Morphological alterations in the cell lines after KSE and KSO treatment were observed. KSE and KSO possessed effective cytotoxic activities against all the cell lines been selected. Conclusions KSE and KSO could be potential sources of natural anti-cancer agents. Further investigations on using kenaf seeds for anti-proliferative properties are warranted. PMID:25183141

  18. Expression of Tumor Necrosis Factor Receptor 2 Characterizes TLR9-Driven Formation of Interleukin-10-Producing B Cells

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    Olga Ticha

    2018-01-01

    Full Text Available B cell-derived interleukin-10 (IL-10 production has been described as a hallmark for regulatory function in B lymphocytes. However, there is an ongoing debate on the origin of IL-10-secreting B cells and lack of specific surface markers has turned into an important obstacle for studying human B regulatory cells. In this study, we propose that tumor necrosis factor receptor 2 (TNFR2 expression can be used for enrichment of IL-10-secreting B cells. Our data confirm that IL-10 production can be induced by TLR9 stimulation with CpG ODN and that IL-10 secretion accompanies differentiation of peripheral blood B cells into plasma blasts. We further show that CpG ODN stimulation induces TNFR2 expression, which correlates with IL-10 secretion and terminal differentiation. Indeed, flow cytometric sorting of TNFR2+ B cells revealed that TNFR2+ and TNFR2− fractions correspond to IL-10+ and IL-10− fractions, respectively. Furthermore, CpG-induced TNFR2+ B cells were predominantly found in the IgM+ CD27+ B cell subset and spontaneously released immunoglobulin. Finally, our data corroborate the functional impact of TNFR2 by demonstrating that stimulation with a TNFR2 agonist significantly augments IL-10 and IL-6 production in B cells. Altogether, our data highlight a new role for TNFR2 in IL-10-secreting human B lymphocytes along with the potential to exploit this finding for sorting and isolation of this currently ill-defined B cell subset.

  19. Role of the functional MNS16A VNTR-243 variant of the human telomerase reverse transcriptase gene in progression and response to therapy of patients with non-Hodgkin's B-cell lymphomas.

    Science.gov (United States)

    Wysoczanska, B; Wrobel, T; Dobrzynska, O; Mazur, G; Bogunia-Kubik, K

    2015-04-01

    MNS16A is a functional polymorphic tandem repeat within the human telomerase reverse transcriptase (hTERT) gene. To investigate whether any of the MNS16A repeats represents a genetic risk factor for NHL susceptibility, progression of or response to therapy in 75 patients with non-Hodgkin's lymphomas (NHLs) and 126 healthy individuals were genotyped using the PCR-VNTR technique. A slightly higher frequency of the MNS16A VNTR-243 variant was detected among patients who did not respond to treatment (NR) as compared to patients with complete or partial remission (0.83 vs. 0.51, P = 0.055). NR patients more frequently developed aggressive than indolent type of the disease (0.92 vs. 0.41, P = 0.001). The VNTR-243 allele was more frequently detected among patients with an intermediate-high/high International Prognostic Index (IPI 3-4) score (P = 0.063), especially in patients with advanced age and IPI 3-4 (P = 0.040). In multivariate analysis, higher IPI 3-4 score (OR = 11.364, P = 0.051) and aggressive type of the disease (OR = 18.182, P = 0.012) were found to be independent genetic markers associated with nonresponse to treatment. Presence of the MNS16A VNTR-243 variant also strongly tended to affect the risk of a less favourable response to therapy and was more frequently present among nonresponders (OR = 5.848, P = 0.059). Genetic variation within the hTERT gene may affect the progression and treatment of lymphoproliferative disorders. © 2015 John Wiley & Sons Ltd.

  20. Molecular cloning of Ly-1, a membrane glycoprotein of mouse T lymphocytes and a subset of B cells: molecular homology to its human counterpart Leu-1/T1 (CD5)

    International Nuclear Information System (INIS)

    Huang, H.J.S.; Jones, N.H.; Strominger, J.L.; Herzenberg, L.A.

    1987-01-01

    The authors report the isolation of cDNA clones of the mouse lymphocyte differentiation antigen Ly-1. One of these cDNA clones was confirmed to be full-length by DNA sequencing and by expression of Ly-1 by L cells transfected with this clone. Analysis of the predicted amino acid sequence indicated that the Ly-1 polypeptide is synthesized with a 23 amino acid leader and that the mature protein consists of an amino-terminal region of 347 amino acids, a transmembrane sequence of 30 residues, and a carboxyl-terminal region of 94 amino acids. The amino-terminal region appears to be divided into two subregions by a threonine- and proline-rich sequence of 23 amino acids that is highly conserved between Ly-1 and its human homologue Leu-1 (CD5) in position and amino acid composition. The first amino-terminal subregion of 111 amino acids is predicted to be arranged in a β-pleated sheet structure of six strands. The entire amino-terminal region is rich in cysteine, with all of its 22 cysteine residues conserved between Ly-1 and Leu-1. The carboxyl-terminal region has no cysteins. Ly-1 and Leu-1 are 63% identical, with a gradient of identical residues from 43% for the first amino-terminal to 58% for the second amino-terminal subregion and 90% for the carboxyl-terminal region. The predicted secondary structure of the first amino-terminal subregion and identities of certain conserved residues among most members of the immunoglobulin gene superfamily suggest that Ly-1 and Leu-1 are distant members of this family

  1. The TEL-AML1 fusion protein of acute lymphoblastic leukemia modulates IRF3 activity during early B-cell differentiation.

    Science.gov (United States)

    de Laurentiis, A; Hiscott, J; Alcalay, M

    2015-12-03

    The t(12;21) translocation is the most common genetic rearrangement in childhood acute lymphoblastic leukemia (ALL) and gives rise to the TEL-AML1 fusion gene. Many studies on TEL-AML1 describe specific properties of the fusion protein, but a thorough understanding of its function is lacking. We exploited a pluripotent hematopoietic stem/progenitor cell line, EML1, and generated a cell line (EML-TA) stably expressing the TEL-AML1 fusion protein. EML1 cells differentiate to mature B-cells following treatment with IL7; whereas EML-TA display an impaired differentiation capacity and remain blocked at an early stage of maturation. Global gene expression profiling of EML1 cells at different stages of B-lymphoid differentiation, compared with EML-TA, identified the interferon (IFN)α/β pathway as a primary target of repression by TEL-AML1. In particular, expression and phosphorylation of interferon-regulatory factor 3 (IRF3) was decreased in EML-TA cells; strikingly, stable expression of IRF3 restored the capacity of EML-TA cells to differentiate into mature B-cells. Similarly, IRF3 silencing in EML1 cells by siRNA was sufficient to block B-lymphoid differentiation. The ability of TEL-AML1 to block B-cell differentiation and downregulate the IRF3-IFNα/β pathway was confirmed in mouse and human primary hematopoietic precursor cells (Lin- and CD34+ cells, respectively), and in a patient-derived cell line expressing TEL-AML1 (REH). Furthermore, treatment of TEL-AML1 expressing cells with IFNα/β was sufficient to overcome the maturation block. Our data provide new insight on TEL-AML1 function and may offer a new therapeutic opportunity for B-ALL.

  2. Effect of New Water-Soluble Dendritic Phthalocyanines on Human Colorectal and Liver Cancer Cell Lines

    Directory of Open Access Journals (Sweden)

    Ebru YABAŞ

    2017-08-01

    Full Text Available Human hepatocellular carcinoma (HepG2 cells and colorectal adenocarcinoma (DLD-1 cells were treated with the synthesized water soluble phthalocyanine derivatives to understand the effect of the compounds both on colorectal and liver cancer cells. The compounds inhibited cell proliferation and displayed cytotoxic effect on these cancer cell lines however; the effect of the compounds on healthy control fibroblast cell line was comparatively lower. The compounds can be employed for cancer treatment as anticancer agents.

  3. Perivascular Adipose Tissue Harbors Atheroprotective IgM-Producing B Cells

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    Prasad Srikakulapu

    2017-09-01

    Full Text Available Adipose tissue surrounding major arteries (Perivascular adipose tissue or PVAT has long been thought to exist to provide vessel support and insulation. Emerging evidence suggests that PVAT regulates artery physiology and pathology, such as, promoting atherosclerosis development through local production of inflammatory cytokines. Yet the immune subtypes in PVAT that regulate inflammation are poorly characterized. B cells have emerged as important immune cells in the regulation of visceral adipose tissue inflammation and atherosclerosis. B cell-mediated effects on atherosclerosis are subset-dependent with B-1 cells attenuating and B-2 cells aggravating atherosclerosis. While mechanisms whereby B-2 cells aggravate atherosclerosis are less clear, production of immunoglobulin type M (IgM antibodies is thought to be a major mechanism whereby B-1 cells limit atherosclerosis development. B-1 cell-derived IgM to oxidation specific epitopes (OSE on low density lipoproteins (LDL blocks oxidized LDL-induced inflammatory cytokine production and foam cell formation. However, whether PVAT contains B-1 cells and whether atheroprotective IgM is produced in PVAT is unknown. Results of the present study provide clear evidence that the majority of B cells in and around the aorta are derived from PVAT. Interestingly, a large proportion of these B cells belong to the B-1 subset with the B-1/B-2 ratio being 10-fold higher in PVAT relative to spleen and bone marrow. Moreover, PVAT contains significantly greater numbers of IgM secreting cells than the aorta. ApoE−/− mice with B cell-specific knockout of the gene encoding the helix-loop-helix factor Id3, known to have attenuated diet-induced atherosclerosis, have increased numbers of B-1b cells and increased IgM secreting cells in PVAT relative to littermate controls. Immunostaining of PVAT on human coronary arteries identified fat associated lymphoid clusters (FALCs harboring high numbers of B cells, and flow

  4. A Cross-Talk Between NFAT and NF-κB Pathways is Crucial for Nickel-Induced COX-2 Expression in Beas-2B Cells

    Science.gov (United States)

    Cai, T.; Li, X.; Ding, J.; Luo, W.; Li, J.; Huang, C.

    2013-01-01

    Cyclooxygenase-2 (COX-2) is a critical enzyme implicated in chronic inflammation-associated cancer development. Our studies have shown that the exposure of Beas-2B cells, a human bronchial epithelial cell line, to lung carcinogenic nickel compounds results in increased COX-2 expression. However, the signaling pathways leading to nickel-induced COX-2 expression are not well understood. In the current study, we found that the exposure of Beas-2B cells to nickel compounds resulted in the activation of both nuclear factor of activated T cell (NFAT) and nuclear factor-κB (NF-κB). The expression of COX-2 induced upon nickel exposure was inhibited by either a NFAT pharmacological inhibitor or the knockdown of NFAT3 by specific siRNA. We further found that the activation of NFAT and NF-κB was dependent on each other. Since our previous studies have shown that NF-κB activation is critical for nickel-induced COX-2 expression in Beas-2B cells exposed to nickel compounds under same experimental condition, we anticipate that there might be a cross-talk between the activation of NFAT and NF-κB for the COX-2 induction due to nickel exposure in Beas-2B cells. Furthermore, we showed that the scavenging of reactive oxygen species (ROS) by introduction of mitochondrial catalase inhibited the activation of both NFAT and NF-κB, and the induction of COX-2 due to nickel exposure. Taken together, our results defining the evidence showing a key role of the cross-talk between NFAT and NF-κB pathways in regulating nickel-induced COX-2 expression, further provide insight into the understanding of the molecular mechanisms linking nickel exposure to its lung carcinogenic effects. PMID:21486220

  5. Phenotypic characterization of autoreactive B cells--checkpoints of B cell tolerance in patients with systemic lupus erythematosus.

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    Annett M Jacobi

    Full Text Available DNA-reactive B cells play a central role in systemic lupus erythematosus (SLE; DNA antibodies precede clinical disease and in established disease correlate with renal inflammation and contribute to dendritic cell activation and high levels of type 1 interferon. A number of central and peripheral B cell tolerance mechanisms designed to control the survival, differentiation and activation of autoreactive B cells are thought to be disturbed in patients with SLE. The characterization of DNA-reactive B cells has, however, been limited by their low frequency in peripheral blood. Using a tetrameric configuration of a peptide mimetope of DNA bound by pathogenic anti-DNA antibodies, we can identify B cells producing potentially pathogenic DNA-reactive antibodies. We, therefore, characterized the maturation and differentiation states of peptide, (ds double stranded DNA cross-reactive B cells in the peripheral blood of lupus patients and correlated these with clinical disease activity. Flow cytometric analysis demonstrated a significantly higher frequency of tetramer-binding B cells in SLE patients compared to healthy controls. We demonstrated the existence of a novel tolerance checkpoint at the transition of antigen-naïve to antigen-experienced. We further demonstrate that patients with moderately active disease have more autoreactive B cells in both the antigen-naïve and antigen-experienced compartments consistent with greater impairment in B cell tolerance in both early and late checkpoints in these patients than in patients with quiescent disease. This methodology enables us to gain insight into the development and fate of DNA-reactive B cells in individual patients with SLE and paves the way ultimately to permit better and more customized therapies.

  6. Comparison of EBV DNA viral load in whole blood, plasma, B-cells and B-cell culture supernatant.

    Science.gov (United States)

    Ouedraogo, David Eric; Bollore, Karine; Viljoen, Johannes; Foulongne, Vincent; Reynes, Jacques; Cartron, Guillaume; Vendrell, Jean-Pierre; Van de Perre, Philippe; Tuaillon, Edouard

    2014-05-01

    Epstein-Barr virus (EBV) genome quantitation in whole blood is used widely for therapeutic monitoring of EBV-associated disorders in immunosuppressed individuals and in patients with EBV-associated lymphoma. However, the most appropriate biological material to be used for EBV DNA quantitation remains a subject of debate. This study compare the detection rate and levels of EBV DNA from whole blood, plasma, enriched B-cells, and B-cell short-term culture supernatant using quantitative real-time PCR. Samples were collected from 33 subjects with either HIV infection or B-cell lymphoma. Overall, EBV DNA was detected in 100% of enriched B-cell samples, in 82% of B-cell culture supernatants, in 57% of plasma, and 42% of whole blood samples. A significant correlation for EBV viral load was found between enriched B-cell and B-cell culture supernatant material (ρ = 0.92; P cells (ρ = -0.02; P = 0.89), whole blood and plasma (ρ = 0.24; P = 0.24), or enriched B-cells and plasma (ρ = 0.08; P = 0.77). Testing of enriched B-cells appeared to be the most sensitive method for detection of EBV DNA as well as for exploration of the cellular reservoir. Quantitation of EBV DNA in plasma and B-cell culture supernatant may be of interest to assess EBV reactivation dynamics and response to treatment as well as to decipher EBV host-pathogen interactions in various clinical scenarios. © 2013 Wiley Periodicals, Inc.

  7. Cytogenetic Evolution of Human Ovarian Cell Lines Associated with Chemoresistance and Loss of Tumorigenicity

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    Stéphanie Struski

    2003-01-01

    Full Text Available In order to identify genomic changes associated with a resistant phenotype acquisition, we used comparative genomic hybridization (CGH to compare a human ovarian cell line, Igrov1, and four derived subcell lines, resistant to vincristine and presenting a reversion of malignant properties. Multicolor FISH (Multiplex‐FISH and Spectral Karyotype and conventional FISH are also used to elucidate the karyotype of parental cell line. The drug‐resistant subcell lines displayed many chromosomal abnormalities suggesting the implication of different pathways leading to a multidrug resistance phenotype. However, these cell lines shared two common rearrangements: an unbalanced translocation der(8t(8;13(p22;q? and a deletion of the 11p. These chromosomal imbalances could reflected the acquisition of the chemoresistance (der(8 or the loss of tumorigenicity properties (del(11p. Colour figure can be viewed on http://www.esacp.org/acp/2003/25‐3/struski.htm.

  8. A stable murine-based RD114 retroviral packaging line efficiently transduces human hematopoietic cells.

    Science.gov (United States)

    Ward, Maureen; Sattler, Rose; Grossman, I Robert; Bell, Anthony J; Skerrett, Donna; Baxi, Laxmi; Bank, Arthur

    2003-11-01

    Several barriers exist to high-efficiency transfer of therapeutic genes into human hematopoietic stem cells (HSCs) using complex oncoretroviral vectors. Human clinical trials to date have used Moloney leukemia virus-based amphotropic and gibbon ape leukemia virus-based envelopes in stable retroviral packaging lines. However, retroviruses pseudotyped with these envelopes have low titers due to the inability to concentrate viral supernatants efficiently by centrifugation without damaging the virus and low transduction efficiencies because of low-level expression of viral target receptors on human HSC. The RD114 envelope from the feline endogenous virus has been shown to transduce human CD34+ cells using transient packaging systems and to be concentrated to high titers by centrifugation. Stable packaging systems have potential advantages over transient systems because greater and more reproducible viral productions can be attained. We have, therefore, constructed and tested a stable RD114-expressing packaging line capable of high-level transduction of human CD34+ cells. Viral particles from this cell line were concentrated up to 100-fold (up to 10(7) viral particles/ml) by ultracentrifugation. Human hematopoietic progenitors from cord blood and sickle cell CD34+ cells were efficiently transduced with a Neo(R)-containing vector after a single exposure to concentrated RD114-pseudotyped virus produced from this cell line. Up to 78% of progenitors from transduced cord blood CD34+ cells and 51% of progenitors from sickle cell CD34+ cells expressed the NeoR gene. We also show transfer of a human beta-globin gene into progenitor cells from CD34+ cells from sickle cell patients with this new RD114 stable packaging system. The results indicate that this packaging line may eventually be useful in human clinical trials of globin gene therapy.

  9. Leukocyte-associated immunoglobulin-like receptor-1 is expressed on human megakaryocytes and negatively regulates the maturation of primary megakaryocytic progenitors and cell line

    International Nuclear Information System (INIS)

    Xue, Jiangnan; Zhang, Xiaoshu; Zhao, Haiya; Fu, Qiang; Cao, Yanning; Wang, Yuesi; Feng, Xiaoying; Fu, Aili

    2011-01-01

    Research highlights: → LAIR-1 is expressed on human megakaryocytes from an early stage. → Up-regulation of LAIR-1 negatively regulates megakaryocytic differentiation of cell line. → LAIR-1 negatively regulates the differentiation of primary megakaryocytic progenitors. -- Abstract: Leukocyte-associated immunoglobulin-like receptor-1 (LAIR-1) is an inhibitory collagen receptor which belongs to the immunoglobulin (Ig) superfamily. Although the inhibitory function of LAIR-1 has been extensively described in multiple leukocytes, its role in megakaryocyte (MK) has not been explored so far. Here, we show that LAIR-1 is expressed on human bone marrow CD34 + CD41a + and CD41a + CD42b + cells. LAIR-1 is also detectable in a fraction of human cord blood CD34 + cell-derived MK that has morphological characteristics of immature MK. In megakaryoblastic cell line Dami, the membrane protein expression of LAIR-1 is up-regulated significantly when cells are treated with phorbol ester phorbol 12-myristate 13-acetate (PMA). Furthermore, cross-linking of LAIR-1 in Dami cells with its natural ligand or anti-LAIR-1 antibody leads to the inhibition of cell proliferation and PMA-promoted differentiation when examined by the MK lineage-specific markers (CD41a and CD42b) and polyploidization. In addition, we also observed that cross-linking of LAIR-1 results in decreased MK generation from primary human CD34 + cells cultured in a cytokines cocktail that contains TPO. These results suggest that LAIR-1 is a likely candidate for an early marker of MK differentiation, and provide initial evidence indicating that LAIR-1 serves as a negative regulator of megakaryocytopoiesis.

  10. 78 FR 25091 - Submission for OMB Review; 30-Day Comment Request: Request for Human Embryonic Stem Cell Line To...

    Science.gov (United States)

    2013-04-29

    ...; 30-Day Comment Request: Request for Human Embryonic Stem Cell Line To Be Approved for Use in NIH... Embryonic Stem Cell Line to be Approved for Use in NIH-Funded Research, 0925-0601, Expiration Date 04/30... Information Collection: The form is used by applicants to request that human embryonic stem cell lines be...

  11. [Establishment and characterization of a cell line derived from human ovarian mucinous cystadenocarcinoma].

    Science.gov (United States)

    Wan, Q; Xu, D; Li, Z

    2001-07-01

    To establish a cell line of human ovarian cancer, and study its characterization. The cell line was established by the cultivation of subsides walls, and kept by freezing. The morphology was observed by microscope and electromicroscope. The authors studied its growth and propagation, the agglutination test of phytohemagglutinin (PHA), the chromosome analysis, heterotransplanting, immuno-histochemistry staining, the analysis of hormone, the pollution examination and the test of sensitivity to virus etc. A new human ovarian carcinoma cell line, designated ovarian mucinous cystadenocarcinoma 685 (OMC685), was established from mucinous cystadenocarcinoma. This cell line had subcultured to 91 generations, and some had been frozen for 8 years and revived, still grew well. This cell line possessed the feature of glandular epithelium cancer cell. The cells grew exuberantly, and the agglutinating test of PHA was positive. Karyotype was subtriploid with distortion. Heterotransplantations, alcian blue periobic acid-schiff (AbPAS), mucicarmine, alcian blue stainings, estradiol (E2) and progesterone were all positive. Without being polluted, it was sensitive to polivirus-I, adenovirus 7 and measles virus. OMC685 is a distinct human ovarian tumous cell line.

  12. Establishment of a novel human medulloblastoma cell line characterized by highly aggressive stem-like cells.

    Science.gov (United States)

    Silva, Patrícia Benites Gonçalves da; Rodini, Carolina Oliveira; Kaid, Carolini; Nakahata, Adriana Miti; Pereira, Márcia Cristina Leite; Matushita, Hamilton; Costa, Silvia Souza da; Okamoto, Oswaldo Keith

    2016-08-01

    Medulloblastoma is a highly aggressive brain tumor and one of the leading causes of morbidity and mortality related to childhood cancer. These tumors display differential ability to metastasize and respond to treatment, which reflects their high degree of heterogeneity at the genetic and molecular levels. Such heterogeneity of medulloblastoma brings an additional challenge to the understanding of its physiopathology and impacts the development of new therapeutic strategies. This translational effort has been the focus of most pre-clinical studies which invariably employ experimental models using human tumor cell lines. Nonetheless, compared to other cancers, relatively few cell lines of human medulloblastoma are available in central repositories, partly due to the rarity of these tumors and to the intrinsic difficulties in establishing continuous cell lines from pediatric brain tumors. Here, we report the establishment of a new human medulloblastoma cell line which, in comparison with the commonly used and well-established cell line Daoy, is characterized by enhanced proliferation and invasion capabilities, stem cell properties, increased chemoresistance, tumorigenicity in an orthotopic metastatic model, replication of original medulloblastoma behavior in vivo, strong chromosome structural instability and deregulation of genes involved in neural development. These features are advantageous for designing biologically relevant experimental models in clinically oriented studies, making this novel cell line, named USP-13-Med, instrumental for the study of medulloblastoma biology and treatment.

  13. Comparison of the effect of interferon on two human hepatoma cell lines

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    Crespi, M; Schoub, B D; Lyons, S F; Chiu, M N [University of the Witwatersrand, Johannesburg (South Africa). Dept. of Virology

    1985-06-01

    Two human hepatoma cell lines, the PLC/PRF/5 and the Mahlavu cells, which differ in their production of the hepatitis B surface antigen (HBsAg), responded differently to interferon (IFN). After IFN treatment both cell lines were able to inhibit Sindbis virus replication. Oligo A synthetase (E enzyme) could be activated in the PLC/PRF/5 cells although they were not sensitive to exogenous 2 - 5 oligoadenylic acid (2 - 5 A). In contrast, the Mahlavu cells were sensitive to exogenous 2 - 5 A, but unable to activate the E enzyme. Both cell lines were unable to stimulate phosphorylation of the exogenous initiator factor eIF-2.

  14. Initiation of Antiviral B Cell Immunity Relies on Innate Signals from Spatially Positioned NKT Cells.

    Science.gov (United States)

    Gaya, Mauro; Barral, Patricia; Burbage, Marianne; Aggarwal, Shweta; Montaner, Beatriz; Warren Navia, Andrew; Aid, Malika; Tsui, Carlson; Maldonado, Paula; Nair, Usha; Ghneim, Khader; Fallon, Padraic G; Sekaly, Rafick-Pierre; Barouch, Dan H; Shalek, Alex K; Bruckbauer, Andreas; Strid, Jessica; Batista, Facundo D

    2018-01-25

    B cells constitute an essential line of defense from pathogenic infections through the generation of class-switched antibody-secreting cells (ASCs) in germinal centers. Although this process is known to be regulated by follicular helper T (TfH) cells, the mechanism by which B cells initially seed germinal center reactions remains elusive. We found that NKT cells, a population of innate-like T lymphocytes, are critical for the induction of B cell immunity upon viral infection. The positioning of NKT cells at the interfollicular areas of lymph nodes facilitates both their direct priming by resident macrophages and the localized delivery of innate signals to antigen-experienced B cells. Indeed, NKT cells secrete an early wave of IL-4 and constitute up to 70% of the total IL-4-producing cells during the initial stages of infection. Importantly, the requirement of this innate immunity arm appears to be evolutionarily conserved because early NKT and IL-4 gene signatures also positively correlate with the levels of neutralizing antibodies in Zika-virus-infected macaques. In conclusion, our data support a model wherein a pre-TfH wave of IL-4 secreted by interfollicular NKT cells triggers the seeding of germinal center cells and serves as an innate link between viral infection and B cell immunity. Copyright © 2017 The Authors. Published by Elsevier Inc. All rights reserved.

  15. [Effects of icotinib hydrochloride on the proliferation and apoptosis of human lung cancer cell lines].

    Science.gov (United States)

    Ma, Li; Han, Xiao-hong; Wang, Shuai; Wang, Jian-fei; Shi, Yuan-kai

    2012-09-25

    To explore the effects of icotinib on the proliferation and apoptosis of various lung cancer cell lines. Human lung cancer cell lines HCC827, H1650, H1975, A549 and human epidermal cancer cell line A431 were treated in vitro with icotinib or gefitinib at a concentration gradient of 0 - 40 µmol/L. Their proliferation effects were analyzed by the thiazolyl blue (MTT) assay and the apoptotic effects detected by flow cytometer. The downstream signaling proteins were detected by Western blot. The median inhibitory concentrations (IC(50)) of icotinib for A431 and HCC827 cell lines were (0.04 ± 0.02) and (0.15 ± 0.06) µmol/L respectively. No significant differences existed between the inhibitions of gefitinib and icotinib on A431, HCC827, H1650, H1975 and A549 cell lines (all P > 0.05). Compared with H1650, H1975 and A549 cell lines, icotinib significantly inhibited A431 (P = 0.009, 0.005 and 0.000) and HCC827 (P = 0.001, 0.001 and 0.000) cell lines. And it lowered the expressions of p-AKT, p-ERK and survivin protein expression through the inhibited activity of p-EGFR protein. Icotinib can arrest the proliferation of lung adenocarcinoma cells with EGFR mutation or over-expression by inhibiting the signal pathways of AKT-ERK and survivin.

  16. The emerging roles of B cells as partners and targets in periodontitis.

    Science.gov (United States)

    Zouali, Moncef

    2017-02-01

    Initial studies of periodontal disease suggested that T cell-mediated immunity against oral Gram-negative microorganisms is a key player in the pathogenesis of this inflammatory disease. Recent investigations, however, revealed that B cells are also engaged. Given their chief role in innate-like and adaptive immune responses, B cells could exert protective functions in periodontitis. However, the periodontal bacteria-specific antibody response is generally unable to halt disease progression in affected subjects, suggesting that the antibodies produced could exhibit low anti-bacterial blocking functions or opsonophagocytic potential, and/or unfavorable effects. Moreover, although microbial antigens are involved in the induction of the inflammatory responses in human adult periodontitis, endogenous antigens also may contribute to the chronicity of this common disease. Not only antibodies to self-antigens, such as collagen, are locally produced, but the autoreactivities observed in aggressive periodontitis are more severe and diverse than those observed in chronic periodontitis, suggesting that autoimmune reactivity could play a role in the tissue destruction of periodontal disease. Further support for a pathological role of B cells in periodontitis comes from the finding that B cell-deficient mice are protected from bacterial infection-induced alveolar bone loss. Studies in patients indicate that B cells and plasma cells, together with osteoclastogenic factors (RANKL and osteoprotegerin) and specific cytokines involved in their growth and differentiation (BAFF and APRIL) participate in the induction of the pathological bone loss in periodontitis. This novel insight suggests that selective targeting of B cells could represent a future therapeutic avenue for severe periodontal disease.

  17. Eμ/miR-125b transgenic mice develop lethal B-cell malignancies.

    Science.gov (United States)

    Enomoto, Y; Kitaura, J; Hatakeyama, K; Watanuki, J; Akasaka, T; Kato, N; Shimanuki, M; Nishimura, K; Takahashi, M; Taniwaki, M; Haferlach, C; Siebert, R; Dyer, M J S; Asou, N; Aburatani, H; Nakakuma, H; Kitamura, T; Sonoki, T

    2011-12-01

    MicroRNA-125b-1 (miR-125b-1) is a target of a chromosomal translocation t(11;14)(q24;q32) recurrently found in human B-cell precursor acute lymphoblastic leukemia (BCP-ALL). This translocation results in overexpression of miR-125b controlled by immunoglobulin heavy chain gene (IGH) regulatory elements. In addition, we found that six out of twenty-one BCP-ALL patients without t(11;14)(q24;q32) showed overexpression of miR-125b. Interestingly, four out of nine patients with BCR/ABL-positive BCP-ALL and one patient with B-cell lymphoid crisis that had progressed from chronic myelogenous leukemia overexpressed miR-125b. To examine the role of the deregulated expression of miR-125b in the development of B-cell tumor in vivo, we generated transgenic mice mimicking the t(11;14)(q24;q32) (Eμ/miR-125b-TG mice). Eμ/miR-125b-TG mice overexpressed miR-125b driven by IGH enhancer and promoter and developed IgM-negative or IgM-positive lethal B-cell malignancies with clonal proliferation. B cells obtained from the Eμ/miR-125b-TG mice were resistant to apoptosis induced by serum starvation. We identified Trp53inp1, a pro-apoptotic gene induced by cell stress, as a novel target gene of miR-125b in hematopoietic cells in vitro and in vivo. Our results provide direct evidence that miR-125b has important roles in the tumorigenesis of precursor B cells.

  18. Interleukin (IL) 36 gamma induces mucin 5AC, oligomeric mucus/gel-forming expression via IL-36 receptor-extracellular signal regulated kinase 1 and 2, and p38-nuclear factor kappa-light-chain-enhancer of activated B cells in human airway epithelial cells.

    Science.gov (United States)

    Bae, Chang Hoon; Choi, Yoon Seok; Na, Hyung Gyun; Song, Si-Youn; Kim, Yong-Dae

    2018-03-01

    Mucin 5AC, oligomeric mucus/gel-forming (MUC5AC) expression is significantly increased in allergic and inflammatory airway diseases. Interleukin (IL) 36 gamma is predominantly expressed in airway epithelial cells and plays an important role in innate and adaptive immune responses. IL-36 gamma is induced by many inflammatory mediators, including cytokines and bacterial and viral infections. However, the association between IL-36 gamma and mucin secretion in human airway epithelial cells has not yet been fully investigated. The objective of this study was to determine whether IL-36 gamma might play a role in the regulation of mucin secretion in airway epithelial cells. We investigated the effect and brief signaling pathway of IL-36 gamma on MUC5AC expression in human airway epithelial cells. Enzyme immunoassay, immunoblot analysis, immunofluorescence staining, reverse transcriptase-polymerase chain reaction (PCR), and real-time PCR were performed in mucin-producing human airway epithelial NCI-H292 cells and in human nasal epithelial cells after pretreatment with IL-36 gamma, several specific inhibitors, or small interfering RNAs (siRNA). IL-36 gamma induced MUC5AC expression and activated the phosphorylation of extracellular signal regulated kinase (ERK) 1 and 2, p38, and nuclear factor-kappa-light-chain-enhancer of activated B cells (NF-kappa B). IL-36 receptor antagonist significantly attenuated these effects. The specific inhibitor and siRNA of ERK1, ERK2, p38, and NF-kappa B significantly attenuated IL-36 gamma induced MUC5AC expression. These results indicated that IL-36 gamma induced MUC5AC expression via the IL-36 receptor-mediated ERK1/2 and p38/NF-kappa B pathway in human airway epithelial cells.

  19. Derivation of the human embryonic stem cell line RCe014-A (RC-10

    Directory of Open Access Journals (Sweden)

    P.A. De Sousa

    2016-03-01

    Full Text Available The human embryonic stem cell line RCe014-A (RC-10 was derived from a fresh oocyte voluntarily donated as unsuitable and surplus to fertility requirements following ethics committee approved informed consent under licence from the UK Human Fertilisation and Embryology Authority. The cell line shows normal pluripotency marker expression and differentiation to the three germ layers in vitro. It has a mixed 46XY and 47XY +12 male karyotype and microsatellite PCR identity, HLA and blood group typing data is available.

  20. Derivation of the human embryonic stem cell line RCe006-A (RC-2

    Directory of Open Access Journals (Sweden)

    P.A. De Sousa

    2016-03-01

    Full Text Available The human embryonic stem cell line RCe006-A (RC-2 was derived from a frozen and thawed blastocyst voluntarily donated as surplus to fertility requirements following ethics committee approved informed consent under licence from the UK Human Fertilisation and Embryology Authority. The cell line exhibits expression of expected pluripotency markers and in vitro differentiation potential to three germinal lineage representative cell populations. It has a male trisomy 12 karyotype (47XY, +12. Microsatellite DNA marker identity and HLA and blood group typing data are available.

  1. Derivation of the human embryonic stem cell line RCe010-A (RC-6

    Directory of Open Access Journals (Sweden)

    P.A. De Sousa

    2016-03-01

    Full Text Available The human embryonic stem cell line RCe010-A (RC-6 was derived from a frozen and thawed blastocyst voluntarily donated as unsuitable and surplus to fertility requirements following ethics committee approved informed consent under licence from the UK Human Fertilisation and Embryology Authority. The cell line shows normal pluripotency marker expression and differentiation to the three germ layers in vitro. It has a normal 46XY male karyotype and microsatellite PCR identity, HLA and blood group typing data are available.

  2. Derivation of the human embryonic stem cell line RCe012-A (RC-8

    Directory of Open Access Journals (Sweden)

    P.A. De Sousa

    2016-03-01

    Full Text Available The human embryonic stem cell line RCe012-A (RC-8 was derived from a frozen and thawed day 5 embryo cultivated to the blastocyst stage. The embryo was voluntarily donated as unsuitable and surplus to fertility requirements following ethics committee approved informed consent under licence from the UK Human Fertilisation and Embryology Authority. The cell line shows normal pluripotency marker expression and differentiation to the three germ layers in vitro. It has a normal 46XX female karyotype and microsatellite PCR identity, HLA and blood group typing data is available.

  3. Neutron and photon clonogenic survival curves of two chemotherapy resistant human intermediate-grade non-Hodgkin lymphoma cell lines

    International Nuclear Information System (INIS)

    Aref, Amr; Yudelev, Mark; Mohammad, Ramzi; Choudhuri, Rajani; Orton, Colin; Al-Katib, Ayad

    1999-01-01

    Background: The potential role of neutron therapy in the management of intermediate-grade non-Hodgkin lymphoma (IGNHL) has not been examined because of the belief that the anticipated radiobiological effectiveness (RBE) would be uniformly very low. Purpose: To determine the fast neutron RBE for two chemotherapy-resistant IGNHL cell lines. Methods and Materials: Conventional soft agar clonogenic survival curves following irradiation by 60 Co and fast neutron were established for two IGNHL cell lines. These cell lines, WSU-DLCL2 and SK-DHL2B, were found in previous studies to be able to repair sublethal damage, and were also resistant to L-Pam and doxorubicin chemotherapy. Results: When the surviving fraction after 2 Gy photon was chosen as the biological endpoint, the RBE for WSU-DLCL2 and SK-DHL2B measured 3.34 and 3.06. Similarly, when 10% survival was considered, the RBE for these two cell lines measured 2.54 and 2.59. The RBE, as measured by the ratios α neutron/α photon, for WSU-DLCL2, SK-DHL2B cell lines are 6.67 and 5.65, respectively. These results indicate that the RBE for these IGNHL cell lines is higher than the average RBE for cell lines of other histological types. Conclusion: Fast neutron irradiation may be of potential value in treating selected cases of IGNHL

  4. BETULINIC ACID WAS MORE CYTOTOXIC TOWARDS THE HUMAN BREAST CANCER CELL LINE MDA-MB-231 THAN THE HUMAN PROMYELOCYTIC LEUKAEMIA CELL LINE HL-60

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    LATIFAH SAIFUL YAZAN

    2009-01-01

    Full Text Available Betulinic acid (BA is a pentacyclic triterpene found in several botanical sources that has been shown to cause apoptosis in a number of cell lines. This study was undertaken to determine the in vitro cytotoxic properties of BA towards the human mammary carcinoma cell line MDA-MB-231 and the human promyelocytic leukaemia cell line HL-60 and the mode of the induced cell death. The cytotoxicity and mode of cell death of BA were determined using the MTT assay and DNAfragmentation analysis, respectively. In our study, the compound was found to be cytotoxic to MDA-MB-231 and HL-60 cells with IC50 values of 58 μg/mL and 134 μg/mL, respectively. Cells treated with high concentrations of BA exhibited features characteristic of apoptosis such as blebbing, shrinking and a number of small cytoplasm body masses when viewed under an inverted light microscope after 24 h. The incidence of apoptosis in MDA-MB-231 was further confirmed bythe DNA fragmentation analysis, with the formation of DNA fragments of oligonucleosomal size (180-200 base pairs, giving a ladder-like pattern on agarose gel electrophoresis. BA was more cytotoxic towards MDA-MB-231 than HL-60 cells, and induced apoptosis in MDA-MB-231 cells.

  5. Type II NKT-TFH cells against Gaucher lipids regulate B-cell immunity and inflammation.

    Science.gov (United States)

    Nair, Shiny; Boddupalli, Chandra Sekhar; Verma, Rakesh; Liu, Jun; Yang, Ruhua; Pastores, Gregory M; Mistry, Pramod K; Dhodapkar, Madhav V

    2015-02-19

    Chronic inflammation including B-cell activation is commonly observed in both inherited (Gaucher disease [GD]) and acquired disorders of lipid metabolism. However, the cellular mechanisms underlying B-cell activation in these settings remain to be elucidated. Here, we report that β-glucosylceramide 22:0 (βGL1-22) and glucosylsphingosine (LGL1), 2 major sphingolipids accumulated in GD, can be recognized by a distinct subset of CD1d-restricted human and murine type II natural killer T (NKT) cells. Human βGL1-22- and LGL1-reactive CD1d tetramer-positive T cells have a distinct T-cell receptor usage and genomic and cytokine profiles compared with the classical type I NKT cells. In contrast to type I NKT cells, βGL1-22- and LGL1-specific NKT cells constitutively express T-follicular helper (TFH) phenotype. Injection of these lipids leads to an increase in respective lipid-specific type II NKT cells in vivo and downstream induction of germinal center B cells, hypergammaglobulinemia, and production of antilipid antibodies. Human βGL1-22- and LGL1-specific NKT cells can provide efficient cognate help to B cells in vitro. Frequency of LGL1-specific T cells in GD mouse models and patients correlates with disease activity and therapeutic response. Our studies identify a novel type II NKT-mediated pathway for glucosphingolipid-mediated dysregulation of humoral immunity and increased risk of B-cell malignancy observed in metabolic lipid disorders. © 2015 by The American Society of Hematology.

  6. Overexpression of c-Jun contributes to sorafenib resistance in human hepatoma cell lines.

    Directory of Open Access Journals (Sweden)

    Yuki Haga

    Full Text Available Despite recent advances in treatment strategies, it is still difficult to cure patients with hepatocellular carcinoma (HCC. Sorafenib is the only approved multiple kinase inhibitor for systemic chemotherapy in patients with advanced HCC. The majority of advanced HCC patients are resistant to sorafenib. The mechanisms of sorafenib resistance are still unknown.The expression of molecules involved in the mitogen-activated protein kinase (MAPK signaling pathway in human hepatoma cell lines was examined in the presence or absence of sorafenib. Apoptosis of human hepatoma cells treated with sorafenib was investigated, and the expression of Jun proto-oncogene (c-Jun was measured.The expression and phosphorylation of c-Jun were enhanced in human hepatoma cell lines after treatment with sorafenib. Inhibiting c-Jun enhanced sorafenib-induced apoptosis. The overexpression of c-Jun impaired sorafenib-induced apoptosis. The expression of osteopontin, one of the established AP-1 target genes, was enhanced after treatment with sorafenib in human hepatoma cell lines.The protein c-Jun plays a role in sorafenib resistance in human hepatoma cell lines. The modulation and phosphorylation of c-Jun could be a new therapeutic option for enhancing responsiveness to sorafenib. Modulating c-Jun may be useful for certain HCC patients with sorafenib resistance.

  7. Radiobiological parameters of a human tumor parent line and four tumor clones of a human epidermoid carcinoma

    International Nuclear Information System (INIS)

    Weichselbaum, R.R.; Beckett, M.; Dahlberg, W.

    1987-01-01

    The authors examined the radiobiological parameters of a parent tumor line and four tumor clones of a human squamous cell carcinoma of the skin. The parent line and clones have a tumor morphology, aneuploid karyotype, and the ability to passage continuously in vitro. With the exception of clone F2A, all cell lines form tumors in nude mice. The parent line, SCC-12 has a D/sub o/ of 154 and an n 7.5 In four tumor clones, D/sub o/ ranges from 131 (clone V) to 266 (clone B2); n ranges from 22.8 in clone V to 2.1 in clone B2. PLDR following 1100 rad ranges from 1.7 in clone B2 to 13.1 in clone V. However, PLDR following equitoxic doses of radiation is similar in the parent and all sub-clones. Radiobiological heterogeneity may complicate predictive assays for clinical radiotherapy

  8. B-Cell-Specific Diversion of Glucose Carbon Utilization Reveals a Unique Vulnerability in B Cell Malignancies.

    Science.gov (United States)

    Xiao, Gang; Chan, Lai N; Klemm, Lars; Braas, Daniel; Chen, Zhengshan; Geng, Huimin; Zhang, Qiuyi Chen; Aghajanirefah, Ali; Cosgun, Kadriye Nehir; Sadras, Teresa; Lee, Jaewoong; Mirzapoiazova, Tamara; Salgia, Ravi; Ernst, Thomas; Hochhaus, Andreas; Jumaa, Hassan; Jiang, Xiaoyan; Weinstock, David M; Graeber, Thomas G; Müschen, Markus

    2018-04-05

    B cell activation during normal immune responses and oncogenic transformation impose increased metabolic demands on B cells and their ability to retain redox homeostasis. While the serine/threonine-protein phosphatase 2A (PP2A) was identified as a tumor suppressor in multiple types of cancer, our genetic studies revealed an essential role of PP2A in B cell tumors. Thereby, PP2A redirects glucose carbon utilization from glycolysis to the pentose phosphate pathway (PPP) to salvage oxidative stress. This unique vulnerability reflects constitutively low PPP activity in B cells and transcriptional repression of G6PD and other key PPP enzymes by the B cell transcription factors PAX5 and IKZF1. Reflecting B-cell-specific transcriptional PPP-repression, glucose carbon utilization in B cells is heavily skewed in favor of glycolysis resulting in lack of PPP-dependent antioxidant protection. These findings reveal a gatekeeper function of the PPP in a broad range of B cell malignancies that can be efficiently targeted by small molecule inhibition of PP2A and G6PD. Copyright © 2018 Elsevier Inc. All rights reserved.

  9. B-cell activating factor detected on both naïve and memory B cells in bullous pemphigoid.

    Science.gov (United States)

    Qian, Hua; Kusuhara, Masahiro; Li, Xiaoguang; Tsuruta, Daisuke; Tsuchisaka, Atsunari; Ishii, Norito; Koga, Hiroshi; Hayakawa, Taihei; Ohara, Koji; Karashima, Tadashi; Ohyama, Bungo; Ohata, Chika; Furumura, Minao; Hashimoto, Takashi

    2014-08-01

    B-cell activating factor (BAFF), an important immune regulatory cytokine, is involved in development of autoimmune diseases. Although BAFF is expressed in various cells, including dendritic cells (DCs) and monocytes, BAFF expression on B cells has not been well documented. In the present study, BAFF molecules on DCs and naïve and memory B cells in autoimmune bullous diseases, including pemphigus vulgaris, pemphigus foliaceus and bullous pemphigoid (BP), were analysed by flow cytometry. Compared with healthy controls (HC), BAFF expression on naïve and memory B cells increased significantly in BP. No difference in BAFF receptor expression in naïve and memory B cells was shown among all study groups. Furthermore, BAFF expression in both naïve and memory B cells of BP, but not HC, was detected by confocal microscopic analysis. These results implied that BAFF expressed by B cells may play a pathogenic role in autoimmune bullous diseases, particularly BP. © 2014 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.

  10. 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 thyroiditis (HT), Graves' disease (GD) and healthy controls were incubated with human thyroglobulin (Tg) before adding normal peripheral blood mononuclear cells. The deposition of immunoglobulins and C3 fragments on B cells was then assessed. Inclusion of Tg in serum from HT patients promoted B cell capture...

  11. 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

    's thyroiditis (HT), Graves' disease (GD) and healthy controls were incubated with human thyroglobulin (Tg) before adding normal peripheral blood mononuclear cells. The deposition of immunoglobulins and C3 fragments on B cells was then assessed. Inclusion of Tg in serum from HT patients promoted B cell capture......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...

  12. FLAX OIL FROM TRANSGENIC LINUM USITATISSIMUM SELECTIVELY INHIBITS IN VITRO PROLIFERATION OF HUMAN CANCER CELL LINES.

    Science.gov (United States)

    Gebarowski, Tomasz; Gebczak, Katarzyna; Wiatrak, Benita; Kulma, Anna; Pelc, Katarzyna; Czuj, Tadeusz; Szopa, Jan; Gasiorowski, Kazimierz

    2017-03-01

    Emulsions made of oils from transgenic flaxseeds significantly decreased in vitro proliferation of six tested human cancer cell lines in 48-h cultures, as assessed with the standard sulforhodamine assay. However, the emulsions also increased proliferation rate of normal human dermal fibroblasts and, to a lower extend, of keratinocytes. Both inhibition of in vitro proliferation of human cancer cell lines and stimulation of proliferation of normal dermal fibroblasts and keratinocytes were especially strong with the emulsion type B and with emulsion type M. Oils from seeds of transgenic flax type B and M should be considered as valuable adjunct to standard cytostatic therapy of human cancers and also could be applied to improve the treatment of skin lesions in wound healing.

  13. Antibody and B cell responses to Plasmodium sporozoites

    Directory of Open Access Journals (Sweden)

    Johanna N Dups

    2014-11-01

    Full Text Available Antibodies are capable of blocking infection of the liver by Plasmodium sporozoites. Accordingly the induction of anti-sporozoite antibodies is a major aim of various vaccine approaches to malaria. In recent years our knowledge of the specificity and quantities of antibodies required for protection has been greatly expanded by clinical trials of various whole sporozoite and subunit vaccines. Moreover, the development of humanized mouse models and transgenic parasites have also aided our ability to assess the specificity of antibodies and their ability to block infection. Nonetheless, considerable gaps remain in our knowledge - in particular in understanding what antigens are recognized by infection blocking antibodies and in knowing how we can induce robust, long-lived antibody responses. Maintaining high levels of circulating antibodies is likely to be of primary importance, as antibodies must block infection in the short time it takes for sporozoites to reach the liver from the skin. It is clear that a better understanding of the development of protective B cell-mediated immunity will aid the development and refinement of malaria vaccines.

  14. The MHV68 M2 protein drives IL-10 dependent B cell proliferation and differentiation.

    Directory of Open Access Journals (Sweden)

    Andrea M Siegel

    2008-04-01

    Full Text Available Murine gammaherpesvirus 68 (MHV68 establishes long-term latency in memory B cells similar to the human gammaherpesvirus Epstein Barr Virus (EBV. EBV encodes an interleukin-10 (IL-10 homolog and modulates cellular IL-10 expression; however, the role of IL-10 in the establishment and/or maintenance of chronic EBV infection remains unclear. Notably, MHV68 does not encode an IL-10 homolog, but virus infection has been shown to result in elevated serum IL-10 levels in wild-type mice, and IL-10 deficiency results in decreased establishment of virus latency. Here we show that a unique MHV68 latency-associated gene product, the M2 protein, is required for the elevated serum IL-10 levels observed at 2 weeks post-infection. Furthermore, M2 protein expression in primary murine B cells drives high level IL-10 expression along with increased secretion of IL-2, IL-6, and MIP-1alpha. M2 expression was also shown to significantly augment LPS driven survival and proliferation of primary murine B cells. The latter was dependent on IL-10 expression as demonstrated by the failure of IL10-/- B cells to proliferate in response to M2 protein expression and rescue of M2-associated proliferation by addition of recombinant murine IL-10. M2 protein expression in primary B cells also led to upregulated surface expression of the high affinity IL-2 receptor (CD25 and the activation marker GL7, along with down-regulated surface expression of B220, MHC II, and sIgD. The cells retained CD19 and sIgG expression, suggesting differentiation to a pre-plasma memory B cell phenotype. These observations are consistent with previous analyses of M2-null MHV68 mutants that have suggested a role for the M2 protein in expansion and differentiation of MHV68 latently infected B cells-perhaps facilitating the establishment of virus latency in memory B cells. Thus, while the M2 protein is unique to MHV68, analysis of M2 function has revealed an important role for IL-10 in MHV68 pathogenesis

  15. The MHV68 M2 protein drives IL-10 dependent B cell proliferation and differentiation.

    Science.gov (United States)

    Siegel, Andrea M; Herskowitz, Jeremy H; Speck, Samuel H

    2008-04-04

    Murine gammaherpesvirus 68 (MHV68) establishes long-term latency in memory B cells similar to the human gammaherpesvirus Epstein Barr Virus (EBV). EBV encodes an interleukin-10 (IL-10) homolog and modulates cellular IL-10 expression; however, the role of IL-10 in the establishment and/or maintenance of chronic EBV infection remains unclear. Notably, MHV68 does not encode an IL-10 homolog, but virus infection has been shown to result in elevated serum IL-10 levels in wild-type mice, and IL-10 deficiency results in decreased establishment of virus latency. Here we show that a unique MHV68 latency-associated gene product, the M2 protein, is required for the elevated serum IL-10 levels observed at 2 weeks post-infection. Furthermore, M2 protein expression in primary murine B cells drives high level IL-10 expression along with increased secretion of IL-2, IL-6, and MIP-1alpha. M2 expression was also shown to significantly augment LPS driven survival and proliferation of primary murine B cells. The latter was dependent on IL-10 expression as demonstrated by the failure of IL10-/- B cells to proliferate in response to M2 protein expression and rescue of M2-associated proliferation by addition of recombinant murine IL-10. M2 protein expression in primary B cells also led to upregulated surface expression of the high affinity IL-2 receptor (CD25) and the activation marker GL7, along with down-regulated surface expression of B220, MHC II, and sIgD. The cells retained CD19 and sIgG expression, suggesting differentiation to a pre-plasma memory B cell phenotype. These observations are consistent with previous analyses of M2-null MHV68 mutants that have suggested a role for the M2 protein in expansion and differentiation of MHV68 latently infected B cells-perhaps facilitating the establishment of virus latency in memory B cells. Thus, while the M2 protein is unique to MHV68, analysis of M2 function has revealed an important role for IL-10 in MHV68 pathogenesis-identifying a

  16. B-Cell waste classification sampling and analysis plan

    International Nuclear Information System (INIS)

    HOBART, R.L.

    1999-01-01

    This report documents the methods used to collect and analyze samples to obtain data necessary to verify and/or determine the radionuclide content of the 324 Facility B-Cell decontamination and decommissioning waste stream

  17. Suitability of Yin Yang 1 transcript and protein levels for biomarker studies in B cell non-Hodgkin lymphoma.

    Science.gov (United States)

    Arribas Arranz, Jéssica; Winter, Dalia Nilufar; Drexler, Hans Günter; Eberth, Sonja

    2018-01-01

    Yin Yang 1 (YY1) is a transcription factor that plays an important role during all stages of B cell differentiation. Several studies reported upregulation of YY1 in B cell derived lymphoma, indicating that it might act as an oncogene. Furthermore, aberrant YY1 expression has been associated with survival in some entities of B cell non-Hodgkin lymphoma (B-NHL), suggesting that YY1 could be a valuable biomarker in B-NHL. However, studies are controversial and methodologically disparate, partially because some studies are based on transcript levels while others rely on YY1 protein data. Therefore, we aimed to investigate the dependence of YY1 protein levels on YY1 transcription. A panel of human cell lines representing different B-NHL subtypes was used to test for the correlation of YY1 mRNA and protein levels which were determined by quantitative PCR and immunoblotting. To analyze YY1 mRNA and YY1 protein stability cells were treated with actinomycin-D and cycloheximide, respectively. siRNAs were transfected to knockdown YY1 . Kaplan-Meier survival analyses were performed with data from published patient cohorts. Pearson's correlation analyses were assessed and statistical power was examined by Student's t-test. In the analyzed panel of B-NHL cell lines YY1 transcript levels do not correlate with their cellular protein amounts. YY1 protein levels were unaffected by transient block of transcription or by targeting YY1 mRNA using siRNA. Additionally, global inhibition of translation up to 48 h did not alter protein levels of YY1, indicating that YY1 is a highly stable protein in B-NHL. Furthermore, in a retrospective analysis of two different B-NHL cohorts, YY1 transcript levels had no impact on patients' survival probabilities. Our results point out the necessity to focus on YY1 protein expression to understand the potential role of YY1 as an oncogene and to unravel its suitability as clinical biomarker in B-NHL.

  18. Low-risk susceptibility alleles in 40 human breast cancer cell lines

    International Nuclear Information System (INIS)

    Riaz, Muhammad; Elstrodt, Fons; Hollestelle, Antoinette; Dehghan, Abbas; Klijn, Jan GM; Schutte, Mieke

    2009-01-01

    Low-risk breast cancer susceptibility alleles or SNPs confer only modest breast cancer risks ranging from just over 1.0 to1.3 fold. Yet, they are common among most populations and therefore are involved in the development of essentially all breast cancers. The mechanism by which the low-risk SNPs confer breast cancer risks is currently unclear. The breast cancer association consortium BCAC has hypothesized that the low-risk SNPs modulate expression levels of nearby located genes. Genotypes of five low-risk SNPs were determined for 40 human breast cancer cell lines, by direct sequencing of PCR-amplified genomic templates. We have analyzed expression of the four genes that are located nearby the low-risk SNPs, by using real-time RT-PCR and Human Exon microarrays. The SNP genotypes and additional phenotypic data on the breast cancer cell lines are presented. We did not detect any effect of the SNP genotypes on expression levels of the nearby-located genes MAP3K1, FGFR2, TNRC9 and LSP1. The SNP genotypes provide a base line for functional studies in a well-characterized cohort of 40 human breast cancer cell lines. Our expression analyses suggest that a putative disease mechanism through gene expression modulation is not operative in breast cancer cell lines

  19. Transcriptome variations among human embryonic stem cell lines are associated with their differentiation propensity.

    Directory of Open Access Journals (Sweden)

    Changbin Sun

    Full Text Available Human embryonic stem cells (hESCs have the potential to form any cell type in the body, making them attractive cell sources in drug screening, regenerative medicine, disease and developmental processes modeling. However, not all hESC lines have the equal potency to generate desired cell types in vitro. Significant variations have been observed for the differentiation efficiency of various human ESC lines. The precise underpinning molecular mechanisms are still unclear. In this work, we compared transcriptome variations of four hESC lines H7, HUES1, HUES8 and HUES9. We found that hESC lines have different gene expression profiles, and these differentially expressed genes (DEGs are significantly enriched in developmental processes, such as ectodermal, mesodermal and endodermal development. The enrichment difference between hESC lines was consistent with its lineage bias. Among these DEGs, some pluripotency factors and genes involved in signaling transduction showed great variations as well. The pleiotropic functions of these genes in controlling hESC identity and early lineage specification, implicated that different hESC lines may utilize distinct balance mechanisms to maintain pluripotent state. When the balance is broken in a certain environment, gene expression variation between them could impact on their different lineage specification behavior.

  20. In vitro radiosensitivity of six human cell lines. A comparative study with different statistical models

    International Nuclear Information System (INIS)

    Fertil, B.; Deschavanne, P.J.; Lachet, B.; Malaise, E.P.

    1980-01-01

    The intrinsic radiosensitivity of human cell lines (five tumor and one nontransformed fibroblastic) was studied in vitro. The survival curves were fitted by the single-hit multitarget, the two-hit multitarget, the single-hit multitarget with initial slope, and the quadratic models. The accuracy of the experimental results permitted evaluation of the various fittings. Both a statistical test (comparison of variances left unexplained by the four models) and a biological consideration (check for independence of the fitted parameters vis-a-vis the portion of the survival curve in question) were carried out. The quadratic model came out best with each of them. It described the low-dose effects satisfactorily, revealing a single-hit lethal component. This finding and the fact that the six survival curves displayed a continuous curvature ruled out the adoption of the target models as well as the widely used linear regression. As calculated by the quadratic model, the parameters of the six cell lines lead to the following conclusions: (a) the intrinsic radiosensitivity varies greatly among the different cell lines; (b) the interpretation of the fibroblast survival curve is not basically different from that of the tumor cell lines; and (c) the radiosensitivity of these human cell lines is comparable to that of other mammalian cell lines

  1. Lymphoma and the control of B cell growth and differentiation.

    Science.gov (United States)

    Rui, Lixin; Goodnow, Christopher C

    2006-05-01

    It is now widely accepted that lymphomagenesis is a multistep transformation process. A number of genetic changes and environmental and infectious factors contributing to the development and malignant progression of B-cell lymphoproliferative disorders are well documented. Reciprocal chromosomal translocations involving the immunoglobulin loci are a hallmark of most mature B cell lymphomas and lead to dysregulated expression of proto-oncogenes (c-myc) important for cell proliferation or genes involved in cell cycle progression (cyclin D1), differentiation block (bcl-6, PAX5) and cell survival (bcl-2, NF-kappaB). In addition, genetic alterations that inactivate tumor suppressor genes (p53, p16) have been frequently detected in some lymphoma tissues. Many of these genes are normally regulated by signals from the B cell antigen receptor. The high prevalence of bacterial and viral infection in lymphoma patients supports the hypothesis that infectious agents may play a contributory role in the development and evolution of B cell lymphoproliferative disorders by either directly inducing polyclonal B cell hyperactivation (EBV, HCV), or providing a chronic antigenic stimulus (EBV, HCV, HBV, H. pylori), or mimicking B cell antigen receptor signaling (EBV, HCV, HHV8), although whether these are causative factors or they are secondary to genetic changes in lymphomagenesis remains to be defined. Stimulatory signals from reactive T cells, local cytokines and growth factors can also contribute, to some extent, to the progression of transformation. Modulation of B cell antigen receptor signaling therefore emerges as a potentially powerful strategy for controlling the growth of certain B cell lymphomas.

  2. 'Big bang' of B-cell development revealed.

    Science.gov (United States)

    Murre, Cornelis

    2018-01-15

    Earlier studies have identified transcription factors that specify B-cell fate, but the underlying mechanisms remain to be revealed. Two new studies by Miyai and colleagues (pp. 112-126) and Li and colleagues (pp. 96-111) in this issue of Genes & Development provide new and unprecedented insights into the genetic and epigenetic mechanisms that establish B-cell identity. © 2018 Murre; Published by Cold Spring Harbor Laboratory Press.

  3. 324 Facility B-Cell quality process plan

    International Nuclear Information System (INIS)

    Carlson, J.L.

    1998-01-01

    This report documents the quality process plan for the restart of a hot cell in the B Plant, originally a bismuth phosphate processing facility, but later converted to a waste fractionation plant. B-Cell is currently being cleaned out and deactivated. TPA Milestone M-89-02 dictates that all mixed waste and equipment be removed from B-Cell by 5/31/1999. This report describes the major activities that remain for completion of the TPA milestone

  4. Hematopoietic Overexpression of FOG1 Does Not Affect B-Cells but Reduces the Number of Circulating Eosinophils

    Science.gov (United States)

    Du Roure, Camille; Versavel, Aude; Doll, Thierry; Cao, Chun; Pillonel, Vincent; Matthias, G