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Sample records for activation antigen cd23

  1. Mast cells modulate transport of CD23/IgE/antigen complex across human intestinal epithelial barrier

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    Ping-Chang Yang

    2009-06-01

    Full Text Available Background: Food allergy and chronic intestinal inflammation are common in western countries. The complex of antigen/IgE is taken up into the body from the gut lumen with the aid of epithelial cell-derived CD23 (low affinity IgE receptor II that plays an important role in the pathogenesis of intestinal allergy. This study aimed to elucidate the role of mast cell on modulation of antigen/IgE complex transport across intestinal epithelial barrier. Methods: Human intestinal epithelial cell line HT29 cell monolayer was used as a study platform. Transepithelial electric resistance (TER and permeability to ovalbumin (OVA were used as the markers of intestinal epithelial barrier function that were recorded in response to the stimulation of mast cell-derived chemical mediators. Results: Conditioned media from naïve mast cell line HMC-1 cells or monocyte cell line THP-1 cells significantly upregulated the expression of CD23 and increased the antigen transport across the epithelium. Treatment with stem cell factor (SCF, nerve growth factor (NGF, retinoic acid (RA or dimethyl sulphoxide (DMSO enhanced CD23 expression in HT29 cells. Conditioned media from SCF, NGF or RA-treated HMC-1 cells, and SCF, NGF, DMSO or RA-treated THP-1 cells enhanced immune complex transport via enhancing the expression of the CD23 in HT29 cells and the release of inflammatory mediator TNF-α. Nuclear factor kappa B inhibitor, tryptase and TNF-α inhibited the increase in CD23 in HT29 cells and prevents the enhancement of epithelial barrier permeability. Conclusions: Mast cells play an important role in modulating the intestinal CD23 expression and the transport of antigen/IgE/CD23 complex across epithelial barrier.

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

    OpenAIRE

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

    1989-01-01

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

  3. Serum levels of total IgE and soluble CD23 in bronchial asthma

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    G. Di Lorenzo

    1996-01-01

    Full Text Available The aim of the present study was to compare, during the pollen season, serum levels of total IgE and soluble CD23 (sCD23 from patients with allergic bronchial asthma, with those from healthy subjects. Significantly higher levels of total IgE and sCD23 were found in patients with asthma compared to the control group. Both in normal controls and in asthmatic patients, a significant correlation was shown between the levels of these two molecules. In asthmatic patients, significant correlations were found for both total IgE and sCD23, with lung function measured as bronchial responsiveness to inhaled methacholine. These results suggest that in asthmatic patients, in addition to the study of total serum IgE levels, the assessment of sCD23 serum levels may be helpful in the evaluation of disease activity.

  4. IgE mediates killing of intracellular Toxoplasma gondii by human macrophages through CD23-dependent, interleukin-10 sensitive pathway.

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    Ioannis Vouldoukis

    Full Text Available BACKGROUND: In addition to helminthic infections, elevated serum IgE levels were observed in many protozoal infections, while their contribution during immune response to these pathogens remained unclear. As IgE/antigen immune complexes (IgE-IC bind to human cells through FcεRI or FcεRII/CD23 surface molecules, the present study aimed to identify which functional receptor may be involved in IgE-IC interaction with human macrophages, the major effector cell during parasite infection. METHODOLOGY/PRINCIPAL FINDINGS: Human monocyte-derived macrophages were infected with Toxoplasma gondii before being incubated with IgE-IC. IgE receptors were then identified using appropriate blocking antibodies. The activation of cells and parasiticidal activity were evaluated by mediator quantification and direct counting of infected macrophages. RNAs were extracted and cell supernatants were also collected for their content in tumor necrosis factor (TNF-α, interleukin-10 (IL-10 and nitrites. Sera from symptomatic infected patients were also tested for their content of IgE, IL-10 and nitrites, and compared to values found in healthy donors. Results showed that IgE-IC induced intracellular elimination of parasites by human macrophages. IgE-mediated effect was FcεRI-independent, but required cross-linking of surface FcεRII/CD23, cell activation and the generation of nitric oxide (NO. Although TNF-α was shown to be produced during cell activation, this cytokine had minor contribution in this phenomenon while endogenous and exogenous IL-10 down-regulated parasite killing. Inverse relationship was found between IL-10 and NO expression by infected human macrophages at both mRNA and mediator levels. The relationship between these in vitro data and in vivo levels of various factors in T. gondii infected patients supports the involvement of CD23 antigen and IL-10 expression in disease control. CONCLUSION: Thus, IgE may be considered as immune mediator during

  5. Structural Changes in the Lectin Domain of CD23, the Low-Affinity IgE Receptor, upon Calcium Binding

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    Wurzburg, Beth A.; Tarchevskaya, Svetlana S.; Jardetzky, Theodore S. (NWU)

    2010-03-08

    CD23, the low-affinity receptor for IgE (Fc{var_epsilon}RII), regulates IgE synthesis and also mediates IgE-dependent antigen transport and processing. CD23 is a unique Fc receptor belonging to the C-type lectin-like domain superfamily and binds IgE in an unusual, non-lectin-like manner, requiring calcium but not carbohydrate. We have solved the high-resolution crystal structures of the human CD23 lectin domain in the presence and absence of Ca{sup 2+}. The crystal structures differ significantly from a previously determined NMR structure and show that calcium binding occurs at the principal binding site, but not at an auxiliary site that appears to be absent in human CD23. Conformational differences between the apo and Ca{sup 2+} bound structures suggest how IgE-Fc binding can be both calcium-dependent and carbohydrate-independent.

  6. Differential regulation of monocyte cytokine release by αV and β2 integrins that bind CD23

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    Edkins, Adrienne L; Borland, Gillian; Acharya, Mridu; Cogdell, Richard J; Ozanne, Bradford W; Cushley, William

    2012-01-01

    The human soluble CD23 (sCD23) protein displays highly pleiotropic cytokine-like activity. Monocytic cells express the sCD23-binding integrins αVβ3, αVβ5, αMβ2 and αXβ2, but it is unclear which of these four integrins most acutely regulates sCD23-driven cytokine release. The hypothesis that ligation of different sCD23-binding integrins promoted release of distinct subsets of cytokines was tested. Lipopolysaccharide (LPS) and sCD23 promoted release of distinct groups of cytokines from the THP-1 model cell line. The sCD23-driven cytokine release signature was characterized by elevated amounts of RANTES (CCL5) and a striking increase in interleukin-8 (IL-8; CXCL8) secretion, but little release of macrophage inflammatory protein 1β (MIP-1β; CCL4). Antibodies to αVβ3 or αXβ2 both promoted IL-8 release, consistent with the sCD23-driven pattern, but both also evoked strong MIP-1β secretion; simultaneous ligation of these two integrins further increased cytokine secretion but did not alter the pattern of cytokine output. In both model cell lines and primary tissue, integrin-mediated cytokine release was more pronounced in immature monocyte cells than in mature cells. The capacity of anti-integrin monoclonal antibodies to elicit a cytokine release response is epitope-dependent and also reflects the differentiation state of the cell. Although a pattern of cytokine release identical to that provoked by sCD23 could not be elicited with any individual anti-integrin monoclonal antibody, αXβ2 and αVβ3 appear to regulate IL-8 release, a hallmark feature of sCD23-driven cytokine secretion, more acutely than αMβ2 or αVβ5. PMID:22348662

  7. Antigen-specific active immunotherapy for ovarian cancer

    NARCIS (Netherlands)

    Leffers, N.; Daemen, T.; Helfrich, W.; Boezen, H. M.; Cohlen, B. J.; Melief, Cornelis; Nijman, H. W.

    2010-01-01

    BACKGROUND: Despite advances in chemotherapy, prognosis of ovarian cancer remains poor. Antigen-specific active immunotherapy aims to induce a tumour-antigen-specific anti-tumour immune responses as an alternative treatment for ovarian cancer. OBJECTIVES: To assess feasibility of antigen-specific ac

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

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

    2014-01-01

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

  9. Circulating levels of the innate and humoral immune regulators CD14 and CD23 are associated with adult glioma

    OpenAIRE

    Zhou, Mi; Wiemels, Joseph L.; Bracci, Paige; Wrensch, Margaret R.; McCoy, Lucie; Rice, Terri; Sison, Jennette; Patoka, Joseph; Wiencke, John K.

    2010-01-01

    Allergy history has been consistently inversely associated with glioma risk. Two serologic markers, soluble CD23 (sCD23) and soluble CD14 (sCD14), are part of the innate and adaptive humoral immune systems and modulate allergic responses in opposite directions, with sCD23 enhancing and sCD14 blunting inflammatory responses. We measured sCD23 and sCD14 in serum from blood that was drawn at a single time point from 1079 glioma patients post diagnosis and 736 healthy controls. Glioma was strongl...

  10. Circulating levels of the innate and humoral immune regulators CD14 and CD23 are associated with adult glioma.

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    Zhou, Mi; Wiemels, Joseph L; Bracci, Paige M; Wrensch, Margaret R; McCoy, Lucie S; Rice, Terri; Sison, Jennette D; Patoka, Joseph S; Wiencke, John K

    2010-10-01

    Allergy history has been consistently inversely associated with glioma risk. Two serologic markers, soluble CD23 (sCD23) and soluble CD14 (sCD14), are part of the innate and adaptive humoral immune systems and modulate allergic responses in opposite directions, with sCD23 enhancing and sCD14 blunting inflammatory responses. We measured sCD23 and sCD14 in serum from blood that was drawn at a single time point from 1,079 glioma patients postdiagnosis and 736 healthy controls. Glioma was strongly associated with high sCD14 [highest versus lowest quartile odds ratio (OR), 3.94; 95% confidence interval (95% CI), 2.98-5.21] and low sCD23 (lowest versus highest quartile OR, 2.5; 95% CI, 1.89-3.23). Results were consistent across glioma histologic types and grades, but were strongest for glioblastoma. Whereas temozolomide treatment was not associated with either sCD14 or sCD23 levels among cases, those taking dexamethasone had somewhat lower sCD23 levels than those not taking dexamethasone. However, sCD23 was associated with case status regardless of dexamethasone treatment. These results augment the long-observed association between allergies and glioma and support a role for the innate and adaptive humoral functions of the immune system, in particular immunoregulatory proteins, in gliomagenesis. PMID:20719886

  11. Immune complexes that contain HIV antigens activate peripheral blood T cells.

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    Korolevskaya, L B; Shmagel, K V; Saidakova, E V; Shmagel, N G; Chereshnev, V A

    2016-07-01

    Uninfected donor T cells were treated in vitro by model immune complexes that contained either HIV or hepatitis C virus (HCV) antigens. Unlike HCV antigen-containing complexes, the immune complexes that contained HIV antigens have been shown to activate peripheral blood T cells of uninfected donors under in vitro conditions. Both the antiviral antibodies and HIV antigen were involved in the activation process. The unique properties of the immune complexes formed by HIV antigens and antiviral antibodies are believed to result from the virus-specific antibody properties and molecular conformation of the antigen-antibody complex. PMID:27595830

  12. Murine T-lymphocyte activation by mycobacterial antigens

    International Nuclear Information System (INIS)

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

  13. Evolución de la leucemia linfática crónica: Valor predictivo del inmunofenotipo, el CD23 soluble y la morfología Evolution of chronic lymphocytic leukemia: Predictive value of immunophenotype, sCD23 and morphology

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    Marcela A. Sarmiento

    2002-08-01

    -treated patients were enrolled. We analysed prospectively: morphology (typical, mixed and PL-CLL; immunophenotypic profile (Matutes score; sCD23 plasma levels; clinical stage; lymphocyte doubling time; b2microglobulin and karyotype abnormalities. Disease progression (need of treatment, progression to advanced stages, development of bulky organomegaly and death related to disease were considered as events. Md of follow-up 24 mo. Results: Stage 0: 11/36, PFS 80%; I: 10/36 PFS 90%; II: 13/36; III and IV: 2/36. SLE > II PFS 37%. p= 0.023. Lymphocyte doubling time 12mo. 24/31. PFS 28% vs. 80% p 350 UI/ml: 17/32. PFS 92% vs. 53% p=0.005. Immunophenotype: Score 5: 15/36, Score 4: 19/36, PFS 64%. Score 3: 2/36. p=0.516. Morphology: typical 17/35, mixed 17/35, PFS 81% vs. 57%, p=0.099. PL-CLL 1/35. Conclusions: sCD23 was suitable to predict PFS, specially useful for early stages without additional markers of active disease. Morphology (excluding PL-CLL and immunophenotype, two common tools, were not useful for the study purpose.

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

    OpenAIRE

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

    2013-01-01

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

  15. Signal transduction by HLA class II antigens expressed on activated T cells

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    Ødum, Niels; Martin, P J; Schieven, G L;

    1991-01-01

    Human T cells express HLA class II antigens upon activation. Although activated, class II+ T cells can present alloantigens under certain circumstances, the functional role of class II antigens on activated T cells remains largely unknown. Here, we report that cross-linking of HLA-DR molecules ex...

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

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

    1987-01-01

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

  17. Distinct morphophenotypic features of chronic B-cell leukaemias identified with CD1c and CD23 antibodies.

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    Orazi, A; Cattoretti, G; Polli, N; Delia, D; Rilke, F

    1991-07-01

    Morphological criteria usually applied to diagnose various subtypes of B-cell chronic lymphoid leukaemia are largely subjective. Immunophenotyping of 61 relevant cases using a selected panel of monoclonal antibodies (mAb), showed that CD1c and CD23 mAb were able to separate B-cell chronic lymphocytic leukaemia (B-CLL) from other chronic B-cell lymphoproliferative diseases. Lymphocytes of B-CLL were CD1c-, CD23+, whereas those of other types of chronic B-cell leukaemia were CD1c+/-, CD23-, and CD38/-. Non-B-CLL cases had a significantly higher amount of large peroxidase-negative (unstained) cells analyzed with an automated blood cell counter (Technicon H6000). This type of volumetric assessment allowed a separation between typical and "atypical" B-CLL, which otherwise were both CD1c-, and CD23+. These combinations of phenotypic markers corresponded to well-defined haematopathologic entities, conventionally diagnosed on peripheral blood (PB) and bone marrow smears, and on histologic sections of lymph nodes and spleen.

  18. The role of CD4 in antigen-independent activation of isolated single T lymphocytes

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    Kelso, A; Owens, T

    1988-01-01

    The membrane molecule CD4 (L3T4) is thought to facilitate activation of Class II H-2-restricted T cells by binding to Ia determinants on antigen-presenting cells. Recent reports suggest that CD4 can also contribute to antigen-independent activation by anti-T cell receptor (TCR) antibodies. An assay...

  19. Immune activation by casein dietary antigens in bipolar disorder

    NARCIS (Netherlands)

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

    2010-01-01

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

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

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    Hinrich eAbken

    2013-11-01

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

  1. Regulation of protein synthesis and autophagy in activated dendritic cells: implications for antigen processing and presentation.

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    Argüello, Rafael J; Reverendo, Marisa; Gatti, Evelina; Pierre, Philippe

    2016-07-01

    Antigenic peptides presented in the context of major histocompatibility complex (MHC) molecules originate from the degradation of both self and non-self proteins. T cells can therefore recognize at the surface of surveyed cells, the self-peptidome produced by the cell itself (mostly inducing tolerance) or immunogenic peptides derived from exogenous origins. The initiation of adaptive immune responses by dendritic cells (DCs), through the antigenic priming of naïve T cells, is associated to microbial pattern recognition receptors engagement. Activation of DCs by microbial product or inflammatory cytokines initiates multiple processes that maximize DC capacity to present exogenous antigens and stimulate T cells by affecting major metabolic and membrane traffic pathways. These include the modulation of protein synthesis, the regulation of MHC and co-stimulatory molecules transport, as well as the regulation of autophagy, that, all together promote exogenous antigen presentation while limiting the display of self-antigens by MHC molecules.

  2. Antigen-activated dendritic cells ameliorate influenza A infections

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

    2013-01-01

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

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

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

    2013-06-01

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

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

    Institute of Scientific and Technical Information of China (English)

    HU Jun; PAN Sheng-jun; CAI Zhen-jie; GUAN De-lin; LIU Xiao-cheng

    2006-01-01

    Objective:With the regular mixed lymphocytes culture (MLC) to detect the allograft rejection, the reactivity of the activated lymphocytes (primed lymphocytes) of a recipient shows sometimes increase and sometimes decrease against the antigens from the donor, which is inconsistent with the clinical results. In order to establish a convenient method for testing the specificity of the activated lymphocytes in vitro, so as to know the rejection occurred or not by testing the existence of the specific activated lymphocytes against donor's HLA antigens in the recipient's peripheral blood. Methods: Anti-IL-2 neutralizing monoclonal antibody (anti-IL-2 N-mAb) and immunosuppressors were introduced in this test system in the presence of specific stimulators and activated lymphocytes. Results: When the activated lymphocytes were chosen from the one-way MLC 4 d to undergo re-stimulation by specific stimulators, the activity of activated lymphocytes in the treatment group was suppressed significantly compared with that in the control group. The result of this test method is consistent with the biopsy in the clinical diagnosis of rejection.Conclusion :It suggests that the activated lymphocytes can be inactivated by specific antigens in certain conditions. This can be a useful tool to define the specificity of the activated lymphocytes.

  5. Regulation of murine macrophage Ia-antigen expression by products of activated spleen cells

    OpenAIRE

    1980-01-01

    This investigation examined the effects of mediators derived form activated spleen cells on macrophage Ia-antigen expression and function. Incubation of adherent thioglycollate-induced murine peritoneal macrophages(> 90% Ia-) with concanavalin A (Con A)- stimulated spleen cell supernate (Con A sup) resulted in a dose- dependent increase in the percentage of Ia-containing (Ia+) phagocytic cells, as detected by antiserum-and-complement-mediated cytotoxicity. The Ia-antigen expression of macroph...

  6. Inhibitory activities of microalgal extracts against Epstein-Barr Virus (EBV antigen expression in lymphoblastoid cells

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    Koh Yih Yih

    2014-01-01

    Full Text Available The inhibitory activities of microalgal extracts against the expression of three EBV antigens, latent membrane protein (LMP1, Epstein-Barr nuclear antigen (EBNA1 and Z Epstein-Barr reactivation activator (ZEBRA were assessed by immunocytochemistry. The observation that the methanol extracts and their fractions from Ankistrodesmus convolutus, Synechococcus elongatus and Spirulina platensis exhibited inhibitory activity against EBV proteins in three Burkitt’s lymphoma cell lines at concentrations as low as 20 μg/ml suggests that microalgae could be a potential source of antiviral compounds against EBV.

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

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    Ferenc A Scheeren

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

  8. Monoclonal antibodies to antigens on human neutrophils, activated T lymphocytes, and acute leukemia blast cells

    Energy Technology Data Exchange (ETDEWEB)

    Miterev, G.Yu.; Burova, G.F.; Puzhitskaya, M.S.; Danilevich, S.V.; Bulycheva, T.I.

    1987-11-01

    The authors describe the production of two mouse hybridomas secreting monoclonal antibodies to antigenic determinants of the surface membranes of human neutrophils, activated T lymphocytes, and acute leukemic blast cells. The degree of lymphocyte stimulation was estimated from incorporation of /sup 3/H-thymidine with parallel microculture. Monoclonal antibodies of supernatants of hybridoma cultures shown here reacted in both immunofluorescence test and cytotoxicity test with surface membrane antigens on the majority of neutrophils and PHA-activated peripheral blood lymphocytes from healthy subjects, but did not give positive reactions with unactivated lymphocytes, adherent monocytes, erythrocytes, and alloantigen-stimulated lymphocytes.

  9. Active self-healing encapsulation of vaccine antigens in PLGA microspheres

    Science.gov (United States)

    Desai, Kashappa-Goud H.; Schwendeman, Steven P.

    2013-01-01

    Herein, we describe the detailed development of a simple and effective method to microencapsulate vaccine antigens in poly(lactic-co-glycolic acid) (PLGA) by simple mixing of preformed active self-microencapsulating (SM) PLGA microspheres in a low concentration aqueous antigen solution at modest temperature (10-38 °C). Co-encapsulating protein-sorbing vaccine adjuvants and polymer plasticizers were used to “actively” load the protein in the polymer pores and facilitate polymer self-healing at temperature > hydrated polymer glass transition temperature, respectively. The microsphere formulation parameters and loading conditions to provide optimal active self-healing microencapsulation of vaccine antigen in PLGA was investigated. Active self-healing encapsulation of two vaccine antigens, ovalbumin and tetanus toxoid (TT), in PLGA microspheres was adjusted by preparing blank microspheres containing different vaccine adjuvant (aluminum hydroxide (Al(OH)3) or calcium phosphate). Active loading of vaccine antigen in Al(OH)3-PLGA microspheres was found to: a) increase proportionally with an increasing loading of Al(OH)3 (0.88-3 wt%) and addition of porosigen, b) decrease when the inner Al(OH)3/trehalose phase to 1 mL outer oil phase and size of microspheres was respectively > 0.2 mL and 63 μm, and c) change negligibly by PLGA concentration and initial incubation (loading) temperature. Encapsulation of protein sorbing Al(OH)3 in PLGA microspheres resulted in suppression of self-healing of PLGA pores, which was then overcome by improving polymer chain mobility, which in turn was accomplished by coincorporating hydrophobic plasticizers in PLGA. Active self-healing microencapsulation of manufacturing process-labile TT in PLGA was found to: a) obviate micronization- and organic solvent-induced TT degradation, b) improve antigen loading (1.4-1.8 wt% TT) and encapsulation efficiency (~ 97%), c) provide nearly homogeneous distribution and stabilization of antigen in polymer

  10. Intravenously delivered glucocorticoid liposomes inhibit osteoclast activity and bone erosion in murine antigen-induced arthritis

    NARCIS (Netherlands)

    Hofkens, Wouter; Grevers, Lilyanne C.; Walgreen, Birgitte; de Vries, Teun J.; Leenen, Pieter J. M.; Everts, Vincent; Storm, Gert; van den Berg, Wim B.; van Lent, Peter L.

    2011-01-01

    The objective of this study was to determine the effect of systemic delivery of prednisolone phosphate (PLP) encapsulated within long circulating 'stealth' liposomes on bone erosion and osteoclast activity during experimental antigen-induced arthritis (AIA). Liposomal PLP strongly suppressed knee jo

  11. Enhancement of human natural cytotoxicity by Plasmodium falciparum antigen activated lymphocytes

    DEFF Research Database (Denmark)

    Theander, T G; Pedersen, B K; Bygbjerg, I C;

    1987-01-01

    Mononuclear cells (MNC) isolated from malaria immune donors and from donors never exposed to malaria were stimulated in vitro with soluble purified Plasmodium falciparum antigens (SPag) or PPD. After 7 days of culture the proliferative response and the cytotoxic activity against the natural killer...

  12. Inhibition of Ly-6A antigen expression prevents T cell activation

    OpenAIRE

    1990-01-01

    Antisense oligonucleotides complementary to the 5' end of the mRNA encoding the Ly-6A protein were used to block the expression of that protein. Using this approach we could inhibit the expression of Ly-6A by 60-80% in antigen-primed lymph node (LN) T cells as well as in the D10 T cell clone. Inhibition of Ly-6 expression resulted in the inability to restimulate in vitro, antigen-primed T cells. It also blocked the activation of normal spleen cells by Con A, monoclonal antibody (mAb) to CD3, ...

  13. Tumorigenic activity of Merkel cell polyomavirus T antigens expressed in the stratified epithelium of mice

    Science.gov (United States)

    Spurgeon, Megan E.; Cheng, Jingwei; Bronson, Roderick T.; Lambert, Paul F.; DeCaprio, James A.

    2015-01-01

    Merkel cell polyomavirus (MCPyV) is frequently associated with Merkel cell carcinoma (MCC), a highly aggressive neuroendocrine skin cancer. Most MCC tumors contain integrated copies of the viral genome with persistent expression of the MCPyV large T (LT) and small T (ST) antigen. MCPyV isolated from MCC typically contain wild type ST but truncated forms of LT that retain the N-terminus but delete the C-terminus and render LT incapable of supporting virus replication. To determine the oncogenic activity of MCC tumor-derived T antigens in vivo, a conditional, tissue-specific mouse model was developed. Keratin 14-mediated Cre recombinase expression induced expression of MCPyV T antigens in stratified squamous epithelial cells and Merkel cells of the skin epidermis. Mice expressing MCPyV T antigens developed hyperplasia, hyperkeratosis, and acanthosis of the skin with additional abnormalities in whisker pads, footpads and eyes. Nearly half of the mice also developed cutaneous papillomas. Evidence for neoplastic progression within stratified epithelia included increased cellular proliferation, unscheduled DNA synthesis, increased E2F-responsive genes levels, disrupted differentiation, and presence of a DNA damage response. These results indicate that MCPyV T antigens are tumorigenic in vivo, consistent with their suspected etiological role in human cancer. PMID:25596282

  14. Tumorigenic activity of merkel cell polyomavirus T antigens expressed in the stratified epithelium of mice.

    Science.gov (United States)

    Spurgeon, Megan E; Cheng, Jingwei; Bronson, Roderick T; Lambert, Paul F; DeCaprio, James A

    2015-03-15

    Merkel cell polyomavirus (MCPyV) is frequently associated with Merkel cell carcinoma (MCC), a highly aggressive neuroendocrine skin cancer. Most MCC tumors contain integrated copies of the viral genome with persistent expression of the MCPyV large T (LT) and small T (ST) antigen. MCPyV isolated from MCC typically contains wild-type ST but truncated forms of LT that retain the N-terminus but delete the C-terminus and render LT incapable of supporting virus replication. To determine the oncogenic activity of MCC tumor-derived T antigens in vivo, a conditional, tissue-specific mouse model was developed. Keratin 14-mediated Cre recombinase expression induced expression of MCPyV T antigens in stratified squamous epithelial cells and Merkel cells of the skin epidermis. Mice expressing MCPyV T antigens developed hyperplasia, hyperkeratosis, and acanthosis of the skin with additional abnormalities in whisker pads, footpads, and eyes. Nearly half of the mice also developed cutaneous papillomas. Evidence for neoplastic progression within stratified epithelia included increased cellular proliferation, unscheduled DNA synthesis, increased E2F-responsive genes levels, disrupted differentiation, and presence of a DNA damage response. These results indicate that MCPyV T antigens are tumorigenic in vivo, consistent with their suspected etiologic role in human cancer. PMID:25596282

  15. Attenuated recombinant vaccinia virus expressing oncofetal antigen (tumor-associated antigen) 5T4 induces active therapy of established tumors.

    Science.gov (United States)

    Mulryan, Kate; Ryan, Matthew G; Myers, Kevin A; Shaw, David; Wang, Who; Kingsman, Susan M; Stern, Peter L; Carroll, Miles W

    2002-10-01

    The human oncofetal antigen 5T4 (h5T4) is a transmembrane glycoprotein overexpressed by a wide spectrum of cancers, including colorectal, ovarian, and gastric, but with a limited normal tissue expression. Such properties make 5T4 an excellent putative target for cancer immunotherapy. The murine homologue of 5T4 (m5T4) has been cloned and characterized, which allows for the evaluation of immune intervention strategies in "self-antigen" in vivo tumor models. We have constructed recombinant vaccinia viruses based on the highly attenuated and modified vaccinia virus ankara (MVA strain), expressing h5T4 (MVA-h5T4), m5T4 (MVA-m5T4), and Escherichia coli LacZ (MVA-LacZ). Immunization of BALB/c and C57BL/6 mice with MVA-h5T4 and MVA-m5T4 constructs induced antibody responses to human and mouse 5T4, respectively. C57BL/6 and BALB/c mice vaccinated with MVA-h5T4 were challenged with syngeneic tumor line transfectants, B16 melanoma, and CT26 colorectal cells that express h5T4. MVA-h5T4-vaccinated mice showed significant tumor retardation compared with mice vaccinated with MVA-LacZ or PBS. In active treatment studies, inoculation with MVA-h5T4 was able to treat established CT26-h5T4 lung tumor and to a lesser extent B16.h5T4 s.c. tumors. Additionally, when C57BL/6 mice vaccinated with MVA-m5T4 were challenged with B16 cells expressing m5T4, resulting growth of the tumors was significantly retarded compared with control animals. Furthermore, mice vaccinated with MVA-m5T4 showed no signs of autoimmune toxicity. These data support the use of MVA-5T4 for tumor immunotherapy. PMID:12481437

  16. Effective Delivery of Antigen-Encapsulin Nanoparticle Fusions to Dendritic Cells Leads to Antigen-Specific Cytotoxic T Cell Activation and Tumor Rejection.

    Science.gov (United States)

    Choi, Bongseo; Moon, Hyojin; Hong, Sung Joon; Shin, Changsik; Do, Yoonkyung; Ryu, Seongho; Kang, Sebyung

    2016-08-23

    In cancer immunotherapy, robust and efficient activation of cytotoxic CD8(+) T cell immune responses is a promising, but challenging task. Dendritic cells (DCs) are well-known professional antigen presenting cells that initiate and regulate antigen-specific cytotoxic CD8(+) T cells that kill their target cells directly as well as secrete IFN-γ, a cytokine critical in tumor rejection. Here, we employed recently established protein cage nanoparticles, encapsulin (Encap), as antigenic peptide nanocarriers by genetically incorporating the OT-1 peptide of ovalbumin (OVA) protein to the three different positions of the Encap subunit. With them, we evaluated their efficacy in activating DC-mediated antigen-specific T cell cytotoxicity and consequent melanoma tumor rejection in vivo. DCs efficiently engulfed Encap and its variants (OT-1-Encaps), which carry antigenic peptides at different positions, and properly processed them within phagosomes. Delivered OT-1 peptides were effectively presented by DCs to naïve CD8(+) T cells successfully, resulting in the proliferation of antigen-specific cytotoxic CD8(+) T cells. OT-1-Encap vaccinations in B16-OVA melanoma tumor bearing mice effectively activated OT-1 peptide specific cytotoxic CD8(+) T cells before or even after tumor generation, resulting in significant suppression of tumor growth in prophylactic as well as therapeutic treatments. A large number of cytotoxic CD8(+) T cells that actively produce both intracellular and secretory IFN-γ were observed in tumor-infiltrating lymphocytes collected from B16-OVA tumor masses originally vaccinated with OT-1-Encap-C upon tumor challenges. The approaches we describe herein may provide opportunities to develop epitope-dependent vaccination systems that stimulate and/or modulate efficient and epitope-specific cytotoxic T cell immune responses in nonpathogenic diseases.

  17. Immunopathology of human schistosomiasis mansoni: II. NK activity and stimulation by specific antigen

    Directory of Open Access Journals (Sweden)

    L. K. Benarroch

    1988-12-01

    Full Text Available Sixteen S. mansoni infected and untreated patients (5 with recent infection and 11 with chronic disease were evaluated for their in vitro natural killer (NK activity against the NK sensitive target K562 cell line. NK levels in 9 out of 11 patients (82% with chronic disease were significantly lower (mean = 15 ± 6%,compared with patients recently infected (mean = 41 ± 9% p < 0.001 and with the control group (mean = 38 ± 13% p < 0.001. However, both patients and controls NK activity was stimulated by soluble adult worm antigens (SAWA, indicating that NK function even in the chronic stage of the infection is able to respond to the parasite antigens. These results suggest the possibility of NK cell participation as effector mechanism against S. mansoni.

  18. Soluble Expression and Characterization of Biologically Active Bacillus anthracis Protective Antigen in Escherichia coli

    OpenAIRE

    Nagendra Suryanarayana; Vanlalhmuaka,; Bharti Mankere; Monika Verma; Kulanthaivel Thavachelvam; Urmil Tuteja

    2016-01-01

    Bacillus anthracis secretory protein protective antigen (PA) is primary candidate for subunit vaccine against anthrax. Attempts to obtain large quantity of PA from Escherichia coli expression system often result in the formation of insoluble inclusion bodies. Therefore, it is always better to produce recombinant proteins in a soluble form. In the present study, we have obtained biologically active recombinant PA in small scale E. coli shake culture system using three different expression cons...

  19. Antigenicity and Hemaglutination Activity of a Recombinant Hemagglutinin-Neuraminidase of Paramyxovirus Tianjin Strain

    Institute of Scientific and Technical Information of China (English)

    Mei LI; Li-jun YUAN; Li-ying SHI; Xiao-mian LI; Qing WANG; Wen-xiu WANG

    2008-01-01

    Paramyxovirus Tianjin strain, a new genotype of Sendal virus, was isolated from the lungs of common cotton-eared marmoset that died of severe respiratory infection in the marmoset colonies. The 19.28% IgM positive rate in the young children with acute respiratory tract infection suggested a close relationship between Tianjin strain and humans. Hemagglutinin-neuraminidase (HN) is its major transmembrane glycoprotein responsible for viral attachment, penetration and release. To clear the relationship between HN structure and function of paramyxovirus Tianjin strain, rHN1, rHN2 and rHN3 overlapping the ectodomain of HN protein were expressed. Their antigenicity and hemaglutination activity, as well as cross reactivity to standard antisera against influenza virus type A, type B were analyzed. The results indicated expressed rHNs have the natural antigenicity.The segment rHN2 possesses more linear epitopes exposed on the surface of the native I-IN protein than found in segments rHN3 and rHN1. The hemagglutination activity of segment rHN3 is higher than that of segments rHN2 and rHN1, and partially dependent on the three-dimensional conformation of HN3 protein. Cross-reactivity between rHNs and standard antisera against influenza virus type A, type B suggested that rHNs might not be the best alternative as specific antigens to detect virus in clinicalserum specimens.

  20. Antigen transfer from exosomes to dendritic cells as an explanation for the immune enhancement seen by IgE immune complexes.

    Directory of Open Access Journals (Sweden)

    Rebecca K Martin

    Full Text Available IgE antigen complexes induce increased specific T cell proliferation and increased specific IgG production. Immediately after immunization, CD23(+ B cells capture IgE antigen complexes, transport them to the spleen where, via unknown mechanisms, dendritic cells capture the antigen and present it to T cells. CD23, the low affinity IgE receptor, binds IgE antigen complexes and internalizes them. In this study, we show that these complexes are processed onto B-cell derived exosomes (bexosomes in a CD23 dependent manner. The bexosomes carry CD23, IgE and MHC II and stimulate antigen specific T-cell proliferation in vitro. When IgE antigen complex stimulated bexosomes are incubated with dendritic cells, dendritic cells induce specific T-cell proliferation in vivo, similar to IgE antigen complexes. This suggests that bexosomes can provide the essential transfer mechanism for IgE antigen complexes from B cells to dendritic cells.

  1. Antigen-bound C3b and C4b enhance antigen-presenting cell function in activation of human T-cell clones.

    Science.gov (United States)

    Arvieux, J; Yssel, H; Colomb, M G

    1988-10-01

    The effect of complement fragments C3b and C4b, on the triggering of antigen-specific human T-cell clones by Epstein-Barr virus-transformed human lymphoblastoid B cells (LCL) when these fragments are covalently coupled to the antigen tetanus toxin (TT) is described. TT was chemically cross-linked to purified C3b [(TT-C3b)n], C4b [(TT-C4b)n] or bovine serum albumin [(TT-BSA)n] as a control. T-cell activation was quantified by tritiated thymidine incorporation and 51Cr release. (TT-C3b)n and (TT-C4b)n induced proliferative responses comparable to (TT-BSA)n but at 18-25 and 4-6 lower concentrations, respectively. This enhancing effect required the covalent cross-linking of the complement fragments to the antigen and involved intracellular processing of the latter by LCL. Antigen presentation was similarly enhanced when measuring the cytotoxic activity of a helper T-cell clone against LCL previously pulsed with (TT-C3b)n or (TT-C4b)n compared with (TT-BSA)n. Binding studies, carried out on LCL using TT radiolabelled with 125I before cross-linking, indicated that (TT-C3b)n and (TT-C4b)n gave three- to four-fold more binding than (TT-BSA)n. Addition of antibodies against CR1 and CR2 or proteolytic removal of these complement receptors with trypsin inhibited by about 60% the enhancing effect of TT-bound C3b and C4b in both binding and functional assays. These results indicate that binding of C3b or C4b to antigen enhances antigen-specific proliferative and cytotoxic responses of T cells by targeting opsonized antigen onto complement receptors CR1 and CR2 of LCL. The putative significance of these findings in terms of regulation of immune responses by complement is discussed. PMID:2973431

  2. Antigen-bound C3b and C4b enhance antigen-presenting cell function in activation of human T-cell clones.

    Science.gov (United States)

    Arvieux, J; Yssel, H; Colomb, M G

    1988-10-01

    The effect of complement fragments C3b and C4b, on the triggering of antigen-specific human T-cell clones by Epstein-Barr virus-transformed human lymphoblastoid B cells (LCL) when these fragments are covalently coupled to the antigen tetanus toxin (TT) is described. TT was chemically cross-linked to purified C3b [(TT-C3b)n], C4b [(TT-C4b)n] or bovine serum albumin [(TT-BSA)n] as a control. T-cell activation was quantified by tritiated thymidine incorporation and 51Cr release. (TT-C3b)n and (TT-C4b)n induced proliferative responses comparable to (TT-BSA)n but at 18-25 and 4-6 lower concentrations, respectively. This enhancing effect required the covalent cross-linking of the complement fragments to the antigen and involved intracellular processing of the latter by LCL. Antigen presentation was similarly enhanced when measuring the cytotoxic activity of a helper T-cell clone against LCL previously pulsed with (TT-C3b)n or (TT-C4b)n compared with (TT-BSA)n. Binding studies, carried out on LCL using TT radiolabelled with 125I before cross-linking, indicated that (TT-C3b)n and (TT-C4b)n gave three- to four-fold more binding than (TT-BSA)n. Addition of antibodies against CR1 and CR2 or proteolytic removal of these complement receptors with trypsin inhibited by about 60% the enhancing effect of TT-bound C3b and C4b in both binding and functional assays. These results indicate that binding of C3b or C4b to antigen enhances antigen-specific proliferative and cytotoxic responses of T cells by targeting opsonized antigen onto complement receptors CR1 and CR2 of LCL. The putative significance of these findings in terms of regulation of immune responses by complement is discussed.

  3. Vaccination with Antigen Combined with αβ-ATP as a Vaccine Adjuvant Enhances Antigen-Specific Antibody Production via Dendritic Cell Activation.

    Science.gov (United States)

    Matsuo, Kazuhiko; Nishiuma, Satoshi; Hasegawa, Yuta; Kawabata, Fumika; Kitahata, Kosuke; Nakayama, Takashi

    2016-01-01

    Adjuvants are required to enhance antigen-specific immune responses by vaccines. Extracellular ATP serves as a danger signal to alert the immune system of tissue damage by acting on P2X and P2Y receptors and triggers the activation of dendritic cells (DCs). Here we investigated the in vivo adjuvant efficacy of α,β-methylene-ATP (αβ-ATP), a non-hydrolysable form of ATP. We found that intradermal injection of ovalbumin (OVA), as a model antigen, combined with αβ-ATP, as the adjuvant, enhanced OVA-specific immune responses more than OVA alone. Additionally, DCs in the skin of mice injected with OVA and αβ-ATP had increased expression of major histocompatibility complex class II and co-stimulator molecules, CD40, CD80, and CD86, suggesting that αβ-ATP activated DC. These findings indicate that αβ-ATP functions as a potent vaccine adjuvant. PMID:27251512

  4. Kinetics of T cell-activation molecules in response to Mycobacterium tuberculosis antigens

    Directory of Open Access Journals (Sweden)

    Antas Paulo RZ

    2002-01-01

    Full Text Available The phenotypic features acquired subsequent to antigen-specific stimulation in vitro were evaluated by means of the kinetic expressions of CD69 and CD25 activation molecules on T lymphocytes and assayed by flow cytometry in response to PPD, Ag85B, and ferritin in PPD-positive healthy control individuals. In response to PHA, CD69 staining on both CD4+ and CD8+ T cells became initially marked after 4 h, peaked at 24 h, and quickly decreased after 120 h. For CD25, a latter expression was detected around 8 h, having increased after 96 h. As expected, the response rate to the mycobacterial antigens was much lower than that to the mitogen. Positive staining was high after 96 h for CD25 and after 24 h for CD69. CD69 expression was significantly enhanced (p < 0.05 on CD8+ as compared to CD4+ T cells. High levels were also found between 96-120 h. Regarding Ag85B, CD25+ cells were mostly CD4+ instead of CD8+ T cells. Moreover, in response to ferritin, a lower CD25 expression was noted. The present data will allow further characterization of the immune response to new mycobacterial-specific antigens and their evaluation for possible inclusion in developing new diagnostic techniques for tuberculosis as well in a new vaccine to prevent the disease.

  5. Bacteroides gingivalis antigens and bone resorbing activity in root surface fractions of periodontally involved teeth

    Energy Technology Data Exchange (ETDEWEB)

    Patters, M.R.; Landsberg, R.L.; Johansson, L.A.; Trummel, C.L.; Robertson, P.R. (Department of Periodontology, University of Connecticut, School of Dental Medicine, Farmington, Connecticut, U.S.A.)

    1982-01-01

    Bone resorbing activity and the presence of antigens of Bacteroides gingivalis were assessed in plaque, calculus, cementum, and dentin obtained from roots of teeth previously exposed to periodontitis. Each fraction was obtained by scaling the root surface. The fraction were extracted by stirring and sonication, and the soluble centrifuged, sterilized, dialyzed, and adjusted to equivalent protein concentrations. Cementum and dentin extracts from impacted teeth were prepared similarly and served as controls. Stimulation of bone resorption by each extract was assessed in organ cultures of fetal rat bones by measurement of release of previously-incorporated /sup 45/Ca from the bone into the medium. In some groups of teeth, calculus and cementum were treated with acid prior to scaling. Citric acid washes were recovered and dialyzed. An enzyme-linked immunosorbent assay (ELISA) was used to assess the extracts for the presence of antigens reactive with an antiserum to B. gingivalis. Significant stimulation of bone resorption was found in all calculus and periodontally-involved cementum preparations. ELISA showed significant levels of B.gingivalis antigens in plaque, calculus, and cementum of periodontally-involved teeth, but not in involved dentin nor in cementum or dentin of impact teeth. Treatment with citric acid removed essentially all B.gingivalis antigens from cementum but not calculus. The results suggest that substances which stimulate bone resorption and substances which react with B. gingivalis antiserum are present in surface plaque, calculus, and cementum or periodontally-involved teeth. These substances are not present in cementum and dentin of impacted teeth nor in dentin of periodontally-involved teeth. Treatment by both scaling and citric demineralization will remove most of these substances from cementum of teeth previously exposed to periodontitis.

  6. INDUCEMENT OF ANTITUMOR-IMMUNITY BY DC ACTIVATED BY HSP70-H22 TUMOR ANTIGEN PEPTIDE

    Institute of Scientific and Technical Information of China (English)

    冯作化; 黄波; 张桂梅; 李东; 王洪涛

    2003-01-01

    Objective: To investigate the feasibility of decreasing the dosage of tumor antigen peptides by dendritic cell (DC)-presenting and the characteristics of modification of DC by heat shock protein (Hsp70) and antigen peptides. Methods: Peptides were bound to Hsp70 and used to modify DC in vitro. The metabolism of the modified DC and the cytokines secreted by the modified DC were determined. The activation of lymphocytes by the modified DC and Hsp70-H22 peptides was tested. The cytotoxicity of the activated lymphocytes to H22 tumor cells was analyzed. The inhibitory effect of tumor in mice by the injection of DC and Hsp70-H22 peptides was tested. Results: 0.15μg of H22 peptides bound with Hsp70 could make 2×105 DC mature. 4×103 matured DC could activate 2×106 lymphocytes. The same amount of lymphocytes could be activated to produce similar cytotoxicity to tumor cells by either DC modified by 0.003μg of peptides bound with Hsp70 or by direct stimulation with 0.15μg of peptides bound with Hsp70. The dosage of peptides could be reduced by about 50 folds if the modified DC was used for injection instead of Hsp70-peptides. Peptides from normal hepatocytes, bound with Hsp70, could not make DC mature, nor activate lymphocytes through DC. Conclusion: The dosage of Hsp70-H22 peptides can be reduced significantly by DC-presenting to activate lymphocytes. Peptides from normal cells could not activate lymphocytes by either Hsp70-presenting or DC-presenting and they have little chance to induce autoimmunity.

  7. A Viral Vectored Prime-Boost Immunization Regime Targeting the Malaria Pfs25 Antigen Induces Transmission-Blocking Activity

    OpenAIRE

    Goodman, Anna L.; Blagborough, Andrew M.; Sumi Biswas; Yimin Wu; Hill, Adrian V.; Sinden, Robert E.; Draper, Simon J

    2011-01-01

    The ookinete surface protein Pfs25 is a macrogamete-to-ookinete/ookinete stage antigen of Plasmodium falciparum, capable of exerting high-level anti-malarial transmission-blocking activity following immunization with recombinant protein-in-adjuvant formulations. Here, this antigen was expressed in recombinant chimpanzee adenovirus 63 (ChAd63), human adenovirus serotype 5 (AdHu5) and modified vaccinia virus Ankara (MVA) viral vectored vaccines. Two immunizations were administered to mice in a ...

  8. ACTIVATION OF CELLS FROM HYPOTHALAMIC STRUCTURES AFTER INJECTION OF ANTIGENS DIFFERENT IN THEIR NATURE (by the c-Fos expression

    Directory of Open Access Journals (Sweden)

    S. V. Perekrest

    2006-01-01

    Full Text Available Abstract. Activation levels of hypothalamic structures (AHN, PVH, DMH, VMH, LHA, PH were analyzed within first hours after injection of antigens [lipopolysaccharide (LPS and bovine serum albumine [BSA]. For all investigated hypothalamic structures, an increase of c-Fos positive cells numbers was observed after injection of the antigens, as compared to the controls. LPS injection caused activation of more multiple neuron populations. AHN, PVH, LHA, and PH structures exhibited the highest levels of c-Fos activation upon LPS application. BSA injection induced activation of lesser cells quantities, but their enrichment in c-Fos protein was higher, thus resulting into increased optical density of VMH, LHA and PH neurons. Hence, the present work has shown that activation pattern of hypothalamic structures differs upon application of antigens that are different by their origin.

  9. Surface expression of Mo3e antigen by activated human monocytes and U-937 cells

    Energy Technology Data Exchange (ETDEWEB)

    Todd R.F. III; Bury, M.J.; Liu, D.Y.

    1986-03-05

    The surface expression of a protease-sensitive antigen, Mo3e, by activated human monocytes and U-937 cells is a plasma membrane feature of the activated state. Mo3e, which is an 80 kD protein on Western blot analysis, may represent the surface receptor for migration inhibitory factor (MIF), as evidenced by inhibition of MIF responsiveness produced by anti-Mo3e monoclonal antibody. Mo3e is barely detectable (by surface immunofluorescence) on freshly isolated monocytes but becomes expressed in high antigen density during 18-24 hrs culture in medium containing E. coli lipopolysaccharide (> 1 ng/ml), 4..beta..-phorbol 12-myristate 13-acetate (PMA) (5-10 nM), or muramyl dipeptide (0.1-1 ..mu..M). In U-937 cells, Mo3e surface expression is detectable after 24 hrs exposure to PMA and other pharmacological activators of protein kinase C: 4..beta..-phorbol 12, 13 dibutyrate, 4..beta..-phorbol 12, 13 didecanoate, mezerein, or Sn-1,2-dioctanoylglycerol. The biologically-inactivate phorbol compounds, 4..cap alpha..-phorbol 12, 13 didecanoate and 4/sub ..beta../-phorbol do not stimulate Mo3e expression. The calcium ionophore, ionomycin, has a synergistic effect on Mo3e expression stimulated by PMA; conversely, calcium antagonists block PMA-induced Mo3e expression. These results suggest the involvement of protein kinase C activation and intracellular calcium mobilization in the stimulated expression of Mo3e by activated human mononuclear phagocytes.

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

    Science.gov (United States)

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

    1988-06-28

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

  11. Vitamin D controls T cell antigen receptor signaling and activation of human T cells

    DEFF Research Database (Denmark)

    von Essen, Marina Rode; Kongsbak-Wismann, Martin; Schjerling, Peter;

    2010-01-01

    Phospholipase C (PLC) isozymes are key signaling proteins downstream of many extracellular stimuli. Here we show that naive human T cells had very low expression of PLC-gamma1 and that this correlated with low T cell antigen receptor (TCR) responsiveness in naive T cells. However, TCR triggering...... led to an upregulation of approximately 75-fold in PLC-gamma1 expression, which correlated with greater TCR responsiveness. Induction of PLC-gamma1 was dependent on vitamin D and expression of the vitamin D receptor (VDR). Naive T cells did not express VDR, but VDR expression was induced by TCR...... signaling via the alternative mitogen-activated protein kinase p38 pathway. Thus, initial TCR signaling via p38 leads to successive induction of VDR and PLC-gamma1, which are required for subsequent classical TCR signaling and T cell activation....

  12. A small molecule inhibitor for ATPase activity of Hsp70 and Hsc70 enhances the immune response to protein antigens

    Science.gov (United States)

    Baek, Kyung-Hwa; Zhang, Haiying; Lee, Bo Ryeong; Kwon, Young-Guen; Ha, Sang-Jun; Shin, Injae

    2015-12-01

    The ATPase activities of Hsp70 and Hsc70 are known to be responsible for regulation of various biological processes. However, little is known about the roles of Hsp70 and Hsc70 in modulation of immune responses to antigens. In the present study, we investigated the effect of apoptozole (Az), a small molecule inhibitor of Hsp70 and Hsc70, on immune responses to protein antigens. The results show that mice administered with both protein antigen and Az produce more antibodies than those treated with antigen alone, showing that Az enhances immune responses to administered antigens. Treatment of mice with Az elicits production of antibodies with a high IgG2c/IgG1 ratio and stimulates the release of Th1 and Th2-type cytokines, suggesting that Az activates the Th1 and Th2 immune responses. The observations made in the present study suggest that inhibition of Hsp70 and Hsc70 activities could be a novel strategy designing small molecule-based adjuvants in protein vaccines.

  13. A new role for complement C3: regulation of antigen processing through an inhibitory activity.

    OpenAIRE

    Villiers, Christian; Cretin, François,; Lefebvre, Nicole; Marche, Patrice; Villiers, Marie-Bernadette

    2008-01-01

    International audience Increasing evidence underlines the involvement of complement component C3 in the establishment of acquired immunity which appears to play a complex role and to act at different levels. As antigen proteolysis by antigen presenting cells is a key event in the control of antigen presentation efficiency, and consequently in the quality of the immune response, we investigated whether C3 could modulate this step. Our results demonstrate for the first time that C3 can inter...

  14. Transformation and radiosensitivity of human diploid skin fibroblasts transfected with activated RAS oncogene and SV40 T-antigen

    Energy Technology Data Exchange (ETDEWEB)

    Su, L.-N.; Little, J.B. (Harvard School of Public Health, Boston, MA (United States))

    1992-08-01

    Three normal human diploid cell strains were transfected with an activated Ha-ras oncogene (EJ ras) or SV40 T-antigen. Multiple clones were examined for morphological alterations, growth requirements, ability to grow under anchorage independent conditions, immortality and tumorigenicity in nude mice. Clones expressing SV40 T-antigen alone or in combination with ras protein p21 were significantly radioresistant as compared with their parent cells or clones transfected with the neo gene only. This radioresistant phenotype persisted in post-crisis, immortalized cell lines. These data suggest that expression of the SV40 T-antigen but not activated Ha-ras plays an important role in the radiosensitivity of human diploid cells. The radioresistant phenotype in SV40 T transfected cells was not related to the enhanced level of genetic instability seen in pre-crisis and newly immortalized cells, nor to the process of immortalization itself. (author).

  15. Proteolytic activity of prostate-specific antigen (PSA towards protein substrates and effect of peptides stimulating PSA activity.

    Directory of Open Access Journals (Sweden)

    Johanna M Mattsson

    Full Text Available Prostate-specific antigen (PSA or kallikrein-related peptidase-3, KLK3 exerts chymotrypsin-like proteolytic activity. The main biological function of PSA is the liquefaction of the clot formed after ejaculation by cleavage of semenogelins I and II in seminal fluid. PSA also cleaves several other substrates, which may explain its putative functions in prostate cancer and its antiangiogenic activity. We compared the proteolytic efficiency of PSA towards several protein and peptide substrates and studied the effect of peptides stimulating the activity of PSA with these substrates. An endothelial cell tube formation model was used to analyze the effect of PSA-degraded protein fragments on angiogenesis. We showed that PSA degrades semenogelins I and II much more efficiently than other previously identified protein substrates, e.g., fibronectin, galectin-3 and IGFBP-3. We identified nidogen-1 as a new substrate for PSA. Peptides B2 and C4 that stimulate the activity of PSA towards small peptide substrates also enhanced the proteolytic activity of PSA towards protein substrates. Nidogen-1, galectin-3 or their fragments produced by PSA did not have any effect on endothelial cell tube formation. Although PSA cleaves several other protein substrates, in addition to semenogelins, the physiological importance of this activity remains speculative. The PSA levels in prostate are very high, but several other highly active proteases, such as hK2 and trypsin, are also expressed in the prostate and may cleave protein substrates that are weakly cleaved by PSA.

  16. Soluble Expression and Characterization of Biologically Active Bacillus anthracis Protective Antigen in Escherichia coli

    Directory of Open Access Journals (Sweden)

    Nagendra Suryanarayana

    2016-01-01

    Full Text Available Bacillus anthracis secretory protein protective antigen (PA is primary candidate for subunit vaccine against anthrax. Attempts to obtain large quantity of PA from Escherichia coli expression system often result in the formation of insoluble inclusion bodies. Therefore, it is always better to produce recombinant proteins in a soluble form. In the present study, we have obtained biologically active recombinant PA in small scale E. coli shake culture system using three different expression constructs. The PA gene was cloned in expression vectors bearing trc, T5, and T7 promoters and transformed into their respective E. coli hosts. The growth conditions were optimized to obtain maximum expression of PA in soluble form. The expression construct PA-pET32c in DE3-pLysS E. coli host resulted in a maximum production of soluble PA (15 mg L−1 compared to other combinations. Purified PA was subjected to trypsin digestion and binding assay with lethal factor to confirm the protein’s functionality. Biological activity was confirmed by cytotoxicity assay on J774.1 cells. Balb/c mice were immunized with PA and the immunogenicity was tested by ELISA and toxin neutralization assay. This study highlights the expression of soluble and biologically active recombinant PA in larger quantity using simpler E. coli production platform.

  17. Soluble Expression and Characterization of Biologically Active Bacillus anthracis Protective Antigen in Escherichia coli.

    Science.gov (United States)

    Suryanarayana, Nagendra; Vanlalhmuaka; Mankere, Bharti; Verma, Monika; Thavachelvam, Kulanthaivel; Tuteja, Urmil

    2016-01-01

    Bacillus anthracis secretory protein protective antigen (PA) is primary candidate for subunit vaccine against anthrax. Attempts to obtain large quantity of PA from Escherichia coli expression system often result in the formation of insoluble inclusion bodies. Therefore, it is always better to produce recombinant proteins in a soluble form. In the present study, we have obtained biologically active recombinant PA in small scale E. coli shake culture system using three different expression constructs. The PA gene was cloned in expression vectors bearing trc, T5, and T7 promoters and transformed into their respective E. coli hosts. The growth conditions were optimized to obtain maximum expression of PA in soluble form. The expression construct PA-pET32c in DE3-pLysS E. coli host resulted in a maximum production of soluble PA (15 mg L(-1)) compared to other combinations. Purified PA was subjected to trypsin digestion and binding assay with lethal factor to confirm the protein's functionality. Biological activity was confirmed by cytotoxicity assay on J774.1 cells. Balb/c mice were immunized with PA and the immunogenicity was tested by ELISA and toxin neutralization assay. This study highlights the expression of soluble and biologically active recombinant PA in larger quantity using simpler E. coli production platform. PMID:26966576

  18. Longitudinal microarray analysis of cell surface antigens on peripheral blood mononuclear cells from HIV+ individuals on highly active antiretroviral therapy

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    Wang Bin

    2008-03-01

    Full Text Available Abstract Background The efficacy of highly active antiretroviral therapy (HAART determined by simultaneous monitoring over 100 cell-surface antigens overtime has not been attempted. We used an antibody microarray to analyze changes in the expression of 135 different cell-surface antigens overtime on PBMC from HIV+ patients on HAART. Two groups were chosen, one (n = 6 achieved sustainable response by maintaining below detectable plasma viremia and the other (n = 6 responded intermittently. Blood samples were collected over an average of 3 years and 5–8 time points were selected for microarray assay and statistical analysis. Results Significant trends over time were observed for the expression of 7 cell surface antigens (CD2, CD3epsilon, CD5, CD95, CD36, CD27 and CD28 for combined patient groups. Between groups, expression levels of 10 cell surface antigens (CD11a, CD29, CD38, CD45RO, CD52, CD56, CD57, CD62E, CD64 and CD33 were found to be differential. Expression levels of CD9, CD11a, CD27, CD28 and CD52, CD44, CD49d, CD49e, CD11c strongly correlated with CD4+ and CD8+ T cell counts, respectively. Conclusion Our findings not only detected markers that may have potential prognostic/diagnostic values in evaluating HAART efficacy, but also showed how density of cell surface antigens could be efficiently exploited in an array-like manner in relation to HAART and HIV-infection. The antigens identified in this study should be further investigated by other methods such as flow cytometry for confirmation as biological analysis of these antigens may help further clarify their role during HAART and HIV infection.

  19. Hepatitis B virus e antigen induces activation of rat hepatic stellate cells

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    Zan, Yanlu [Center for Molecular Virology, CAS Key Laboratory of Pathogenic Microbiology and Immunology, Institute of Microbiology, Chinese Academy of Sciences, Beijing 100101 (China); University of Chinese Academy of Sciences, Beijing 100049 (China); Zhang, Yuxia, E-mail: yzhang@wehi.edu.au [Center for Molecular Virology, CAS Key Laboratory of Pathogenic Microbiology and Immunology, Institute of Microbiology, Chinese Academy of Sciences, Beijing 100101 (China); Tien, Po, E-mail: tienpo@sun.im.ac.cn [Center for Molecular Virology, CAS Key Laboratory of Pathogenic Microbiology and Immunology, Institute of Microbiology, Chinese Academy of Sciences, Beijing 100101 (China)

    2013-06-07

    Highlights: •HBeAg expression in HSCs induced production of ECM protein and liver fibrotic markers. •The activation and proliferation of HSCs were mediated by TGF-β. •HBeAg protein purified from cell medium directly activated HSCs. -- Abstract: Chronic hepatitis B virus infection is a major cause of hepatic fibrosis, leading to liver cirrhosis and hepatocellular carcinoma. Hepatitis B virus e antigen (HBeAg) is an accessory protein of HBV, not required for viral replication but important for natural infection in vivo. Hepatic stellate cells (HSCs) are the major producers of excessive extracellular matrix during liver fibrogenesis. Therefore, we examined the influence of HBeAg on HSCs. The rat HSC line HSC-T6 was transfected with HBeAg plasmids, and expression of α-smooth muscle actin, collagen I, transforming growth factor-β1 (TGF-β), and tissue inhibitors of metalloproteinase 1 (TIMP-1) was investigated by quantitative real-time PCR. The proliferation of HSCs was determined by MTS analysis. HBeAg transduction induced up-regulation of these fibrogenic genes and proliferation of HSCs. We found that HBeAg induced TGF-β secretion in HSCs, and the activation of HSCs was prevented by a neutralizing anti-TGF-β antibody. Depletion and addition of HBeAg protein in conditioned medium from HSC-T6 cells transduced with HBeAg indicated that HBeAg directly induced the activation and proliferation of rat primary HSCs. Taken together, HBeAg induces the activation and proliferation of HSCs, mainly mediated by TGF-β, and HBeAg protein purified from cell medium can directly activate HSCs.

  20. Functional activation of the T-cell antigen receptor induces tyrosine phosphorylation of phospholipase C-gamma 1.

    OpenAIRE

    Weiss, A; Koretzky, G; Schatzman, R C; Kadlecek, T

    1991-01-01

    Stimulation of the T-cell antigen receptor (TCR), which itself is not a protein-tyrosine kinase (PTK), activates a PTK and phospholipase C (PLC). Using the human T-cell leukemic line Jurkat and normal peripheral blood lymphocytes, we demonstrate that stimulation of the TCR specifically induces the recovery of PLC activity in eluates from anti-phosphotyrosine immunoprecipitates. Stimulation of the human muscarinic receptor, subtype 1, when expressed in Jurkat activates PLC through a guanine nu...

  1. Transcription Activity of Ectogenic Human Carcinoembryonic Antigen Promoter in Lung Adenocarcinoma Cells A549

    Institute of Scientific and Technical Information of China (English)

    XIONG Weining; FANG Huijuan; XU Yongjian; XIONG Shendao; CAO Yong; SONG Qingfeng; ZENG Daxiong; ZHANG Huilan

    2006-01-01

    The transcription activity of ectogenic human carcinoembryonic antigen (CEA) promoter in lung adenocarcinoma cells A549 was investigated for the further gene-targeting therapy. The reporter gene green fluorescent protein (GFP) driven by CEA promoter and human cytomegalovirus (CMV) promoter were relatively constructed and named plasmid pCEA-EGFP and pCMV-GFP respectively. The intensity of fluorescence was detected by fluorescence microscope and flow cytometry analysis after the pCEA-GFP and pSNAV-GFP plasmids were transfected into A549 cells through liposome respectively. The results showed (4.08±0.63) % of the A549 cells transfected with pCEA-AFP plasmid expressed, significantly lower than that of the A549 cells transfected with pCMV-GFP [(43.27±3.54) %]. It was suggested that ectogenic human CEA promoter in lung adenocarcinoma cells A549 was weakly expressed. The distinct specificity of CEA promoter in CEA high expression cells was regarded as a tool in selective gene therapy, but the transcription activity of ectogenic human CEA promoter was needed to increase in the future.

  2. Merkel Cell Polyomavirus Large T Antigen Has Growth-Promoting and Inhibitory Activities

    OpenAIRE

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

    2013-01-01

    Merkel cell carcinoma (MCC) is a rare and aggressive form of skin cancer. In at least 80% of all MCC, Merkel cell polyomavirus (MCPyV) DNA has undergone clonal integration into the host cell genome, and most tumors express the MCPyV large and small T antigens. In all cases of MCC reported to date, the integrated MCPyV genome has undergone mutations in the large T antigen. These mutations result in expression of a truncated large T antigen that retains the Rb binding or LXCXE motif but deletes...

  3. Induction of human complement activation without cytolysis by mouse monoclonal antibodies to human leukocyte antigens.

    Science.gov (United States)

    Sugita, K; Majdic, O; Stockinger, H; Holter, W; Burger, R; Knapp, W

    1987-04-01

    Ten monoclonal antibodies to human leukocyte subsets that had previously been shown to lyse their respective target cells in the presence of rabbit serum as complement source were evaluated for their cytolytic capacity with human complement. Four of the ten were lytic with human complement. All were of IgM type. Antibodies were also evaluated for their capacity to induce C3 binding to target cells. With this method we could demonstrate that, indeed, 3 of the 6 noncytolytic antibodies had the capacity to initiate the human complement activation process and to induce C3 binding. Two of these 3 antibodies were of IgM class (VIT3 and VIM13), one of IgG3 (562). From the practical point of view the most interesting of these 3 antibodies is the nonmitogenic anti-CD3 pan-T cell antibody VIT3. Therefore, this antibody was analyzed in more detail. VIT3 antibody concentrations needed to induce detectable C3 binding to human T cells are very low (down to 1 ng VIT3/ml). Human serum as complement source can also be considerably (100X) diluted before C3 binding becomes undetectable. Activation of C3 is a prerequesite for VIT3-induced C3 binding, and bound C3 seems to lack the C3a fragment. Bound C3, in contrast to the quickly occuring antigenic modulation of the CD3 complex and the simultaneous disappearance of the antibody coat, remains expressed also after prolonged incubation at 37 degrees C. C3 fragments bound to T cells after activation with VIT3 are also recognized by cells bearing C3 receptors of types CR1 and CR2. PMID:3576673

  4. Preparation, Characterization, and Determination of Immunological Activities of Transfer Factor Specific to Human Sperm Antigen

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    Jianwei Zhou

    2013-01-01

    Full Text Available Objective. The objective of this study was to prepare, characterize, and determine immunological activities of specific transfer factor (STF specific to human sperm antigen (HSA for the preparation of antisperm contraceptive vaccine that can be used as an immunocontraceptive. Methods. HSA-STF was prepared using the spleens of rabbits vaccinated with HSA. The specific immunological activities were examined by lymphocyte proliferation test (LPT, leukocyte adhesion inhibition test (LAIT, and by determining the concentrations of IL-4, γ-IFN, and IL-21. HSA-STF was a helveolous substance, having a pH value of 7.0±0.4 and UV absorption maxima at 258 ± 6 nm. It contained seventeen amino acids; glycine and glutamic acids were the highest in terms of concentrations (38.8 μg/mL and 36.3 μg/mL, resp.. Results. The concentration of polypeptide was 2.34±0.31 mg/mL, and ribose was 0.717±0.043 mg/mL. The stimulation index for lymphocyte proliferation test was 1.84, and the leukocyte adhesion inhibition rate was 37.7%. There was a statistically significant difference between the cultural lymphocytes with HSA-STF and non-HSA-STF for γ-IFN and IL-21 (P0.05. Conclusion. HSA-STF was prepared and characterized successfully. It had immunological activity which could transfer the immune response specific to HSA and prove to be a potential candidate for the development of male immunocontraceptive agents.

  5. Ribonucleotide reductase activity is coupled to DNA synthesis via proliferating cell nuclear antigen.

    Science.gov (United States)

    Salguero, Israel; Guarino, Estrella; Shepherd, Marianne E A; Deegan, Tom D; Havens, Courtney G; MacNeill, Stuart A; Walter, Johannes C; Kearsey, Stephen E

    2012-04-24

    Synthesis of deoxynucleoside triphosphates (dNTPs) is required for both DNA replication and DNA repair and is catalyzed by ribonucleotide reductases (RNR), which convert ribonucleotides to their deoxy forms [1, 2]. Maintaining the correct levels of dNTPs for DNA synthesis is important for minimizing the mutation rate [3-7], and this is achieved by tight regulation of RNR [2, 8, 9]. In fission yeast, RNR is regulated in part by a small protein inhibitor, Spd1, which is degraded in S phase and after DNA damage to allow upregulation of dNTP supply [10-12]. Spd1 degradation is mediated by the activity of the CRL4(Cdt2) ubiquitin ligase complex [5, 13, 14]. This has been reported to be dependent on modulation of Cdt2 levels, which are cell cycle regulated, peaking in S phase, and which also increase after DNA damage in a checkpoint-dependent manner [7, 13]. We show here that Cdt2 level fluctuations are not sufficient to regulate Spd1 proteolysis and that the key step in this event is the interaction of Spd1 with the polymerase processivity factor proliferating cell nuclear antigen (PCNA), complexed onto DNA. This mechanism thus provides a direct link between DNA synthesis and RNR regulation. PMID:22464192

  6. Definition of a physiologic aging autoantigen by using synthetic peptides of membrane protein band 3: localization of the active antigenic sites.

    Science.gov (United States)

    Kay, M M; Marchalonis, J J; Hughes, J; Watanabe, K; Schluter, S F

    1990-08-01

    Senescent cell antigen (SCA), an aging antigen, is a protein that appears on old cells and marks them for removal by the immune system in mammals. It is derived from band 3, a ubiquitous membrane transport protein found in diverse cell types and tissues. We have used synthetic peptides to identify aging antigenic sites on band 3, using a competitive inhibition assay and immunoblotting with IgG directed against the aging antigen on old cells. Results indicate that: (i) the active antigenic sites of the aging antigen reside on membrane protein band 3 residues that are extracellular regions implicated in anion transport (residues 538-554 and 788-827); (ii) a putative ankyrin-binding-region peptide is not involved in SCA activity; and (iii) carbohydrate moieties are not required for the antigenicity or recognition of SCA because synthetic peptides alone abolish binding of senescent cell IgG to erythrocytes. One of the putative transport sites that contributes to the aging antigen is located toward the carboxyl terminus. A model of band 3 is presented. Localization of the active antigenic site on the band 3 molecule facilitates definition of the molecular changes occurring during aging that initiate molecular as well as cellular degeneration. PMID:1696010

  7. Enhanced interferon-γ secretion and antitumor activity of T-lymphocytes activated by dendritic cells loaded with glycoengineered myeloma antigens

    Institute of Scientific and Technical Information of China (English)

    XIONG Hong; WU Qiu-ye; HU Hong-gang; LIU Ban; GUO Zhong-wu; Daniel Man-yuan Sze; HOU Jian

    2007-01-01

    Background Immunotherapy is emerging as a promising cure for cancer. However, a severe problem in this area is the immune tolerance to tumor cells and tumor-associated antigens, as evidenced by the ability of cancer to escape immune surveillance. To overcome this problem this work examined the potential of improving the antigenicity of myeloma by metabolic engineering of its cell surface carbohydrate antigens (i.e., glycoengineering) and presentation of the modified tumor antigens by dendritic cells (DCs) to generate cytotoxic T-lymphocytes (CTLs).Methods CD138+ myeloma cells were isolated from 11 multipe myeloma (MM) patients by the immunomagnetic bead method. The MM cells were treated with N-propionyl-D-mannosamine (ManNPr), a synthetic analog of N-acetyl-D-mannosamine (ManNAc), the natural biosynthetic precursor of N-acetyl sialic acid (NeuNAc), to express unnatural N-propionylated sialoglycans. The giycoengineered cells were then induced to apoptosis, and the apoptotic products were added to cultured functional DCs that could present the unnatural carbohydrate antigens to autologous T-lymphocytes.Results It was found that the resultant DCs could activate CD4+ and CD8+ T-lymphocytes, resulting in increased expression of T cell surface markers, including CD8CD28 and CD4CD29. Moreover, upon stimulation by glycoengineered MM cells, these DC-activated T-lymphocytes could release significantly higher levels of IFN-γ (P<0.05).Lactate dehydrogenase (LDH) assays further showed that the stimulated T-lymphocytes were cytotoxic to glycoengineered MM cells.Conclusions This work demonstrated that glycoengineered myeloma cells were highly antigenic and the CTLs induced by the DCs loaded with the unnatural myeloma antigens were specifically cytotoxic to the glycoengineered myeloma.This may provide a new strategy for overcoming the problem of immune tolerance for the development of effective immunotherapies for MM.

  8. Research Upregulation of CD23 (FcεRII Expression in Human Airway Smooth Muscle Cells (huASMC in Response to IL-4, GM-CSF, and IL-4/GM-CSF

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    Lew D Betty

    2005-05-01

    Full Text Available Abstract Background Airway smooth muscle cells play a key role in remodeling that contributes to airway hyperreactivity. Airway smooth muscle remodeling includes hypertrophy and hyperplasia. It has been previously shown that the expression of CD23 on ASMC in rabbits can be induced by the IgE component of the atopic serum. We examined if other components of atopic serum are capable of inducing CD23 expression independent of IgE. Methods Serum starved huASMC were stimulated with either IL-4, GM-CSF, IL-13, IL-5, PGD2, LTD4, tryptase or a combination of IL-4, IL-5, IL-13 each with GM-CSF for a period of 24 h. CD23 expression was analyzed by flow cytometry, western blot, and indirect immunofluorescence. Results The CD23 protein expression was upregulated in huASMC in response to IL-4, GM-CSF, and IL-4/GM-CSF. The percentage of cells with increased fluorescence intensity above the control was 25.1 ± 4.2% (IL-4, 15.6 ± 2.7% (GM-CSF and 32.9 ± 13.9% (IL-4/GMCSF combination(n = 3. The protein content of IL-4/GMCSF stimulated cells was significantly elevated. Expression of CD23 in response to IL-4, GM-CSF, IL-4/GM-CSF was accompanied by changes in cell morphology including depolymerization of isoactin fibers, cell spreading, and membrane ruffling. Western blot revealed abundant expression of the IL-4Rα and a low level expression of IL-2Rγc in huASMC. Stimulation with IL-4 resulted in the phosphorylation of STAT-6 and an increase in the expression of the IL-2Rγc. Conclusion CD23 on huASMC is upregulated by IL-4, GM-CSF, and IL-4/GM-CSF. The expression of CD23 is accompanied by an increase in cell volume and an increase in protein content per cell, suggesting hypertrophy. Upregulation of CD23 by IL-4/GM-CSF results in phenotypic changes in huASMC that could play a role in cell migration or a change in the synthetic function of the cells. Upregulation of CD23 in huASMC by IL-4 and GM-CSF can contribute to changes in huASMC and may provide an avenue

  9. Research Upregulation of CD23 (FcεRII) Expression in Human Airway Smooth Muscle Cells (huASMC) in Response to IL-4, GM-CSF, and IL-4/GM-CSF

    Science.gov (United States)

    Belleau, Joseph T; Gandhi, Radha K; McPherson, Holly M; Lew, D Betty

    2005-01-01

    Background Airway smooth muscle cells play a key role in remodeling that contributes to airway hyperreactivity. Airway smooth muscle remodeling includes hypertrophy and hyperplasia. It has been previously shown that the expression of CD23 on ASMC in rabbits can be induced by the IgE component of the atopic serum. We examined if other components of atopic serum are capable of inducing CD23 expression independent of IgE. Methods Serum starved huASMC were stimulated with either IL-4, GM-CSF, IL-13, IL-5, PGD2, LTD4, tryptase or a combination of IL-4, IL-5, IL-13 each with GM-CSF for a period of 24 h. CD23 expression was analyzed by flow cytometry, western blot, and indirect immunofluorescence. Results The CD23 protein expression was upregulated in huASMC in response to IL-4, GM-CSF, and IL-4/GM-CSF. The percentage of cells with increased fluorescence intensity above the control was 25.1 ± 4.2% (IL-4), 15.6 ± 2.7% (GM-CSF) and 32.9 ± 13.9% (IL-4/GMCSF combination)(n = 3). The protein content of IL-4/GMCSF stimulated cells was significantly elevated. Expression of CD23 in response to IL-4, GM-CSF, IL-4/GM-CSF was accompanied by changes in cell morphology including depolymerization of isoactin fibers, cell spreading, and membrane ruffling. Western blot revealed abundant expression of the IL-4Rα and a low level expression of IL-2Rγc in huASMC. Stimulation with IL-4 resulted in the phosphorylation of STAT-6 and an increase in the expression of the IL-2Rγc. Conclusion CD23 on huASMC is upregulated by IL-4, GM-CSF, and IL-4/GM-CSF. The expression of CD23 is accompanied by an increase in cell volume and an increase in protein content per cell, suggesting hypertrophy. Upregulation of CD23 by IL-4/GM-CSF results in phenotypic changes in huASMC that could play a role in cell migration or a change in the synthetic function of the cells. Upregulation of CD23 in huASMC by IL-4 and GM-CSF can contribute to changes in huASMC and may provide an avenue for new therapeutic options

  10. Tumor Expression of the Carcinoembryonic Antigen Correlates with High Mitotic Activity and Cell Pleomorphism Index in Lung Carcinoma

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    Rancés Blanco

    2013-01-01

    Full Text Available At present, some research efforts are focusing on the evaluation of a variety of tumor associated antigens (TAAs for a better understanding of tumor biology and genetics of lung tumors. For this reason, we evaluated the tissue expression of carcinoembryonic antigen (CEA and ior C2 (a cell surface O-linked glycoprotein carbohydrate chain TAA in lung carcinomas, as well as its correlation with a variety of clinicopathological features. The tissue expression of CEA was evidenced in 22/43 (51.16% lung carcinomas and it was correlated with mitotic activity, cell pleomorphism indexes, and age of patients. The expression of ior C2 was observed in 15/43 (34.88% tumors but no correlation with the clinicopathological features mentioned above was obtained. No correlation between both CEA and ior C2 antigens expression and the overall survival (OS of non-small-cell lung cancer patients was also observed. However, CEA-negative patients displayed higher OS rates as compared with positive ones (69.74 versus 58.26 months. Our results seem to be in agreement with the role of CEA expression in tumor cell proliferation, inhibition of cell polarizations and tissue architecture distortion. The significance of ior C2 antigen in these malignancies and it potential use in diagnosis, prognosis, and/or immunotherapy must be reevaluated.

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

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    Margaret Veselits

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

  12. [Comparative study of the antigens of Streptococcus group A. Rport I. Comparative characteristics of the immunologic activity of partially purified M-protien and the cytoplasmic protective antigen].

    Science.gov (United States)

    Evseev, V A; Avdeeva, Zh I; Kondrashov, G I

    1975-12-01

    Experiments were conducted on mice. A study was made of the protective properties of the cytoplasmic fraction of streptococcus, group A, Type 1 and of an antigen isolated from it by sedimentation with ammonium sulfate, in comparison with M-protein partially purified by the method of Lancefield and Perlman. Cytoplasmic antigen was not inferior by immunogenicity in comparison with M-protein. In difference from the latter, it was thermolabile and sensitive to the action of hydrochloric acid. The protective antigen was revealed in the cytoplasm not only of the virulent, but also of avirulent strains of streptococcus devoid of M-protein.

  13. Invariant chain as a vehicle to load antigenic peptides on human MHC class I for cytotoxic T-cell activation.

    Science.gov (United States)

    Wälchli, Sébastien; Kumari, Shraddha; Fallang, Lars-Egil; Sand, Kine M K; Yang, Weiwen; Landsverk, Ole J B; Bakke, Oddmund; Olweus, Johanna; Gregers, Tone F

    2014-03-01

    Protective T-cell responses depend on efficient presentation of antigen (Ag) in the context of major histocompatibility complex class I (MHCI) and class II (MHCII) molecules. Invariant chain (Ii) serves as a chaperone for MHCII molecules and mediates trafficking to the endosomal pathway. The genetic exchange of the class II-associated Ii peptide (CLIP) with antigenic peptides has proven efficient for loading of MHCII and activation of specific CD4(+) T cells. Here, we investigated if Ii could similarly activate human CD8(+) T cells when used as a vehicle for cytotoxic T-cell (CTL) epitopes. The results show that wild type Ii, and Ii in which CLIP was replaced by known CTL epitopes from the cancer targets MART-1 or CD20, coprecipitated with HLA-A*02:01 and mediated colocalization in the endosomal pathway. Furthermore, HLA-A*02:01-positive cells expressing CLIP-replaced Ii efficiently activated Ag-specific CD8(+) T cells in a TAP- and proteasome-independent manner. Finally, dendritic cells transfected with mRNA encoding IiMART-1 or IiCD20 primed naïve CD8(+) T cells. The results show that Ii carrying antigenic peptides in the CLIP region can promote efficient presentation of the epitopes to CTLs independently of the classical MHCI peptide loading machinery, facilitating novel vaccination strategies against cancer.

  14. Self-antigen recognition by TGFβ1-deficient T cells causes their activation and systemic inflammation

    OpenAIRE

    Bommireddy, Ramireddy; Pathak, Leena J; Martin, Jennifer; Ormsby, Ilona; Engle, Sandra J; Gregory P. Boivin; Babcock, George F.; Eriksson, Anna U.; Singh, Ram R; DOETSCHMAN, THOMAS

    2006-01-01

    To investigate whether the multifocal inflammatory disease in TGFβ1-deficient mice is caused by self-antigen (self-Ag)-specific autoreactive T cells, or whether it is caused by antigen independent, spontaneous hyperactivation of T cells, we have generated Tgfb1−/− and Tgfb1−/− Rag1−/− mice expressing the chicken OVA-specific TCR transgene (DO11.10). On a Rag1-sufficient background, Tgfb1−/− DO11.10 mice develop a milder inflammation than do Tgfb1−/− mice, and their T cells display a less acti...

  15. Major histocompatibility complex class II (DR) antigen and costimulatory molecules on in vitro and in vivo activated human polymorphonuclear neutrophils

    Science.gov (United States)

    Sandilands, Gavin P; McCrae, Jame; Hill, Kathryn; Perry, Martin; Baxter, Derek

    2006-01-01

    We have previously shown that normal human peripheral blood polymorphonuclear neutrophils (PMNs) contain cytoplasmic ‘stores’ of three key molecules normally associated with antigen presentation and T-cell costimulation, i.e. major histocompatibility complex class II (DR) antigen, CD80 (B7-1) and CD86 (B7-2). These cytoplasmic molecules were found to translocate to the cell surface within a few minutes following cross-linking (X-L) of Mac-1: an early neutrophil activation signal. In this study we have compared X-L of Mac −1 in parallel with four other well documented in vitro neutrophil activators: phorbol myristate acetate, N-formyl methionyl leucyl phenylalanine, lipopolysaccharide, and phagocytosis of immunoglobulin G–Latex particles. In addition, we have used paired samples of neutrophils obtained from peripheral blood (as a control) and synovial fluid from patients with rheumatoid arthritis as a source of in vivo activated cells. With the exception of phagocytosis, all activators resulted in the rapid (within 30 min) generation of two populations of activated neutrophils (designated P1 and P2) based on flow-cytometry measurements of size, granularity and phenotype. Significant up-regulation of DR and costimulatory molecules was observed, predominantly on P2 cells, with all activators except phagocytosis. CD80 and CD86 were noted to respond to the various activation signals in a different pattern suggesting that their intracellular granule location may be different. Dual-staining confocal laser microscopy studies showed that CD80 is largely confined to secretory vesicles (SVs) while CD86 appears to have a much wider distribution being found in SVs and within secondary (specific) and primary (azurophilic) granules. Increased surface expression of these antigens was also observed on P2 synovial fluid neutrophils appearing as large heterogeneous clusters on the cell surface when visualized by confocal laser microscopy. PMID:17034427

  16. Standardization of natural mycolic acid antigen composition and production for use in biomarker antibody detection to diagnose active tuberculosis.

    Science.gov (United States)

    Ndlandla, F L; Ejoh, V; Stoltz, A C; Naicker, B; Cromarty, A D; van Wyngaardt, S; Khati, M; Rotherham, L S; Lemmer, Y; Niebuhr, J; Baumeister, C R; Al Dulayymi, J R; Swai, H; Baird, M S; Verschoor, J A

    2016-08-01

    Mycobacterium tuberculosis, the causative agent of tuberculosis, is characterized by the abundance of species specific, antigenic cell wall lipids called mycolic acids. These wax-like molecules all share an identical, amphiphilic mycolic motif, but have different functional groups in a long hydrophobic hydrocarbon mero-chain that divide them into three main classes: alpha-, keto- and methoxy-mycolic acids. Whereas alpha-mycolic acids constitutively maintain an abundance of around 50%, the ratio of methoxy- to keto-mycolic acid types may vary depending on, among other things, the growth stage of M. tuberculosis. In human patients, antibodies to mycolic acids have shown potential as diagnostic serum biomarkers for active TB. Variations in mycolic acid composition affect the antigenic properties and can potentially compromise the precision of detection of anti-mycolic acids antibodies in patient sera to natural mixtures. We demonstrate this here with combinations of synthetic mycolic acid antigens, tested against TB patient and control sera. Combinations of methoxy- and α-mycolic acids are more antigenic than combinations of keto- and α-mycolic acids, showing the former to give a more sensitive test for TB biomarker antibodies. Natural mixtures of mycolic acids isolated from mature cultures of M. tuberculosis H37Rv give the same sensitivity as that with synthetic methoxy- and α-mycolic acids in combination, in a surface plasmon resonance inhibition biosensor test. To ensure that the antigenic activity of isolates of natural mycolic acids is reproducible, we cultured M. tuberculosis H37Rv on Middlebrook 7H10 solid agar plates to stationary growth phase in a standardized, optimal way. The proportions of mycolic acid classes in various batches of the isolates prepared from these cultures were compared to a commercially available natural mycolic acid isolate. LC-MS/MS and NMR data for quantitation of mycolic acids class compositions show that the variation in batches

  17. Activated human nasal epithelial cells modulate specific antibody response against bacterial or viral antigens.

    Directory of Open Access Journals (Sweden)

    Chiou-Yueh Yeh

    Full Text Available Nasal mucosa is an immune responsive organ evidenced by eliciting both specific local secretory IgA and systemic IgG antibody responses with intra-nasal administration of antigens. Nevertheless, the role of nasal epithelial cells in modulating such responses is unclear. Human nasal epithelial cells (hNECs obtained from sinus mucosa of patients with chronic rhinosinusitis were cultured in vitro and firstly were stimulated by Lactococcus lactis bacterium-like particles (BLPs in order to examine their role on antibody production. Secondly, both antigens of immunodominant protein IDG60 from oral Streptococcus mutans and hemagglutinin (HA from influenza virus were tested to evaluate the specific antibody response. Stimulated hNECs by BLPs exhibited a significant increase in the production of interleukin-6 (IL-6, and thymic stromal lymphopoietin (TSLP. Conditioned medium of stimulated hNECs has effects on enhancing the proliferation of CD4+ T cells together with interferon-γ and IL-5 production, increasing the costimulatory molecules on dendritic cells and augmenting the production of IDG60 specific IgA, HA specific IgG, IgA by human peripheral blood lymphocytes. Such production of antigen specific IgG and IgA is significantly counteracted in the presence of IL-6 and TSLP neutralizing antibodies. In conclusion, properly stimulated hNECs may impart immuno-modulatory effects on the antigen-specific antibody response at least through the production of IL-6 and TSLP.

  18. Research Upregulation of CD23 (FcεRII) Expression in Human Airway Smooth Muscle Cells (huASMC) in Response to IL-4, GM-CSF, and IL-4/GM-CSF

    OpenAIRE

    Lew D Betty; McPherson Holly M; Gandhi Radha K; Belleau Joseph T

    2005-01-01

    Abstract Background Airway smooth muscle cells play a key role in remodeling that contributes to airway hyperreactivity. Airway smooth muscle remodeling includes hypertrophy and hyperplasia. It has been previously shown that the expression of CD23 on ASMC in rabbits can be induced by the IgE component of the atopic serum. We examined if other components of atopic serum are capable of inducing CD23 expression independent of IgE. Methods Serum starved huASMC were stimulated with either IL-4, GM...

  19. Cloning, sequence analysis, and expression of the large subunit of the human lymphocyte activation antigen 4F2

    Energy Technology Data Exchange (ETDEWEB)

    Lumadue, J.A.; Glick, A.B.; Ruddle, F.H.

    1987-12-01

    Among the earliest expressed antigens on the surface of activated human lymphocytes is the surface antigen 4F2. The authors have used DNA-mediated gene transfer and fluorescence-activated cell sorting to obtain cell lines that contain the gene encoding the large subunit of the human 4F2 antigen in a mouse L-cell background. Human DNAs cloned from these cell lines were subsequently used as hybridization probes to isolate a full-length cDNA clone expressing 4F2. Sequence analysis of the coding region has revealed an amino acid sequence of 529 residues. Hydrophobicity plotting has predicted a probable structure for the protein that includes an external carboxyl terminus, an internal leader sequence, a single hydrophobic transmembrane domain, and two possible membrane-associated domains. The 4F2 cDNA detects a single 1.8-kilobase mRNA in T-cell and B-cell lines. RNA gel blot analysis of RNA derived from quiescent and serum-stimulated Swiss 3T3 fibroblasts reveals a cell-cycle modulation of 4F2 gene expression: the mRNA is present in quiescent fibroblasts but increases 8-fold 24-36 hr after stimulation, at the time of maximal DNA synthesis.

  20. Malassezia yeasts activate the NLRP3 inflammasome in antigen-presenting cells via Syk-kinase signalling.

    Science.gov (United States)

    Kistowska, Magdalena; Fenini, Gabriele; Jankovic, Dragana; Feldmeyer, Laurence; Kerl, Katrin; Bosshard, Philipp; Contassot, Emmanuel; French, Lars E

    2014-12-01

    Although being a normal part of the skin flora, yeasts of the genus Malassezia are associated with several common dermatologic conditions including pityriasis versicolour, seborrhoeic dermatitis (SD), folliculitis, atopic eczema/dermatitis (AE/AD) and dandruff. While Malassezia spp. are aetiological agents of pityriasis versicolour, a causal role of Malassezia spp. in AE/AD and SD remains to be established. Previous reports have shown that fungi such as Candida albicans and Aspergillus fumigatus are able to efficiently activate the NLRP3 inflammasome leading to robust secretion of the pro-inflammatory cytokine IL-1β. To date, innate immune responses to Malassezia spp. are not well characterized. Here, we show that different Malassezia species could induce NLRP3 inflammasome activation and subsequent IL-1β secretion in human antigen-presenting cells. In contrast, keratinocytes were not able to secrete IL-1β when exposed to Malassezia spp. Moreover, we demonstrate that IL-1β secretion in antigen-presenting cells was dependent on Syk-kinase signalling. Our results identify Malassezia spp. as potential strong inducers of pro-inflammatory responses when taken up by antigen-presenting cells and identify C-type lectin receptors and the NLRP3 inflammasome as crucial actors in this process. PMID:25267545

  1. Temporal differences in the activation of three classes of non-transmembrane protein tyrosine kinases following B-cell antigen receptor surface engagement.

    OpenAIRE

    Saouaf, S J; Mahajan, S.; Rowley, R B; Kut, S A; Fargnoli, J.; Burkhardt, A L; Tsukada, S; Witte, O. N.; Bolen, J B

    1994-01-01

    We evaluated in WEHI 231 B cells the time-dependent responses of Lyn, Blk, Btk, Syk, and three members of the Jak family of protein tyrosine kinases following antibody-mediated surface engagement of the B-cell antigen receptor. Our results show that the enzyme activities of Lyn and Blk were stimulated within seconds of antigen receptor engagement and correlated with the initial tyrosine phosphorylation of the Ig alpha and Ig beta subunits of the B-cell antigen receptor. Btk enzyme activity wa...

  2. Antigen induced production of υ-interferon ex vivo, in the peripheral blood of patients with active pulmonary tuberculosis

    Directory of Open Access Journals (Sweden)

    Z. M. Zagdyn

    2013-01-01

    Full Text Available Tuberculosis (TB is one of the most significant problems in the Russian Health Care. Russia remains on the list of the 22 countries with a high TB incidence and on the third place in the world with a high prevalence of Drug Resistant TB [1]. It is urgently needed to develop new TB diagnostic methods as well as effective measures of the specific TB prevention, including a development of the novel vaccines, so we have to know better about the most immunogenic antigens of Mycobacterium Tuberculosis. We studied the Interferon-Q production in the whole blood after stimulating immune response with different proteins of Mycobacterium Tuberculosis in patients with active TB. The study results permitted us to evaluate the immunogenicity of the previously known proteins (Ag85a и ESAT-6 in comparison to the recently identified ones (Rv2957, Rv2958c и Rv0447, analyzing simultaneously their relation to tuberculin, as well as to antigens of the different viruses (Human Immunodeficiency Virus, Cytomegalovirus, Epstein-Barr Virus, Influenza Virus. Protein Rv2958c, unlike protein ESAT-6, showed the high immunogenicity in comparison to tuberculin. The expressed immunogenicity of protein Rv2958c might be indicated a possible greatest specificity of immune response to this antigen in TB patients. Meanwhile, bacillary tuberculosis was strongly associated with low immune response to this protein. Also we were found statistical differences in immune responses of patients to the different Mycobacterium Tuberculosis antigens depending on the drug sensitivity. In addition it was interesting to know about a significantly low immune response of patients with Drug Resistant TB to protein pp65 CMV.

  3. Appearance of an inhibitory cell nuclear antigen in rat and human serum during variable degrees of hepatic regenerative activity

    Institute of Scientific and Technical Information of China (English)

    1999-01-01

    AIM To determine whether proliferating cell nuclear antigen (PCNA) is present in the peripheral circulation and whether PCNA levels correlate with enhanced regenerative activity.METHODS In animal studies, adult male Sprague-Dawley rats (n=3-4/ group) were sacrificed at 0, 12, 24, 36, 48, 72 and 96 hours following 70% partial hepatectomy. At each interval, sera were analyzed by Western blot for PCNA by two monoclonal antibodies (PC-10 and 19F-4). In human studies, sera from 4 patients with liver cirrhosis and 4 healthy controls were tested in a similar manner.RESULTS The PC-10 monoclonal antibody identified a protein with a molecular mass of 120 KD which remained stable in rat sera for 24 hours following partial hepatectomy, then increased 1.5-fold at 48 hours prior to returning to baseline at 96 hours after partial hepatectomy. However, it was not detected in the sera of patients with or without liver disease. In the 19F-4 monoclonal antibody, a protein with a molecular mass of approximately 46 KD was found. which was present in rat sera prior to partial hepatectomy and for 12 hours after surgery. Thereafter, levels fell by approximately 50% at 24 hours, 65% at 36 hours and 75% at 48 hours where they remained until 96 hours after partial hepatectomy. The decrease in levels correlated with the extent of partial hepatectomy. In human sera, the appearance of this inhibitory cell nuclear antigen (ICNA) was higher in the sera of patients with cirrhosis than in healthy controls.CONCLUSION The PC-10 monoclonal antibody can detect a protein in the circulation when active hepatic regenerative activity is taking place. The 19F-4 monoclonal antibody, however, identifies a protein in both rat and human sera that inversely correlates with hepatic regenerative activity. This protein which is tentatively referred to as inhibitory cell nuclear antigen (ICNA) may be used in documenting the extent of suppression of hepatic regeneration.

  4. Improved Activation toward Primary Colorectal Cancer Cells by Antigen-Specific Targeting Autologous Cytokine-Induced Killer Cells

    Directory of Open Access Journals (Sweden)

    Claudia Schlimper

    2012-01-01

    Full Text Available Adoptive therapy of malignant diseases with cytokine-induced killer (CIK cells showed promise in a number of trials; the activation of CIK cells from cancer patients towards their autologous cancer cells still needs to be improved. Here, we generated CIK cells ex vivo from blood lymphocytes of colorectal cancer patients and engineered those cells with a chimeric antigen receptor (CAR with an antibody-defined specificity for carcinoembryonic antigen (CEA. CIK cells thereby gained a new specificity as defined by the CAR and showed increase in activation towards CEA+ colon carcinoma cells, but less in presence of CEA− cells, indicated by increased secretion of proinflammatory cytokines. Redirected CIK activation was superior by CAR-mediated CD28-CD3ζ than CD3ζ signaling only. CAR-engineered CIK cells from colon carcinoma patients showed improved activation against their autologous, primary carcinoma cells from biopsies resulting in more efficient tumour cell lysis. We assume that adoptive therapy with CAR-modified CIK cells shows improved selectivity in targeting autologous tumour lesions.

  5. IFN-gamma and prostaglandin E2 inhibit IL-4-induced expression of Fc epsilon R2/CD23 on B lymphocytes through different mechanisms without altering binding of IL-4 to its receptor

    Energy Technology Data Exchange (ETDEWEB)

    Galizzi, J.P.; Cabrillat, H.; Rousset, F.; Menetrier, C.; de Vries, J.E.; Banchereau, J.

    1988-09-15

    Human rIL-4 specifically induces the expression of the low affinity receptor for IgE (Fc epsilon R2/CD23) on normal B cells and on the Burkitt lymphoma cell line Jijoye. IL-4 does not induce the generation of the second messenger cAMP in Jijoye cells. PGE2 (at 10(-7) M) was found to inhibit by 50% the IL-4 mediated Fc epsilon R2/CD23 induction on Jijoye cells. The PGE2 half maximum inhibitory concentration (1 nM) was comparable to that inducing a half maximal increase of intracellular cAMP (4nM PGE2). 8-bromo-cAMP (10(-3) M), forskolin (10(-5) M), and cholera toxin (100 ng/ml), which increase intracellular cAMP levels, also inhibited by 40 to 80% the IL-4 induced Fc epsilon R2/CD23 expression on Jijoye cells. PGE2 8-bromo-cAMP, forskolin, and cholera toxin also inhibited the IL-4-induced Fc epsilon R2/CD23 expression on normal B lymphocytes. Taken together these data suggest that PGE2 inhibits the IL-4 induced Fc epsilon R2/CD23 through an increase of intracellular cAMP. In contrast, IFN-gamma, which strongly inhibits IL-4-mediated Fc epsilon R2/CD23 expression on Jijoye cells, did not increase intracellular cAMP levels and thus probably acts through another mechanism. IFN-gamma and PGE2 did not inhibit binding of IL-4 to its receptor. It could be excluded that IFN-gamma and PGE2 were acting via an alteration/desensitization of the IL-4R inasmuch as 24 h pre-incubation of Jijoye cells with these agents affected neither the affinity of 125I-IL-4 for its receptor (Kd = 0.8 to 1.5 x 10(-10) M) nor the maximal number of binding sites per Jijoye cells (Bmax = 390 to 550). Furthermore, IFN-gamma and PGE2 did not affect the internalization and degradation of 125I-IL-4. These data demonstrate that PGE2 and IFN-gamma inhibit the IL-4-mediated induction of Fc epsilon R2/CD23 on B lymphocytes via different mechanisms that do not alter the interaction of IL-4 with its receptor.

  6. Inhibition of the Antigen-Induced Activation of RBL-2H3 Cells by Gab2 siRNA

    Institute of Scientific and Technical Information of China (English)

    Feng Huang; Xiaoyun Tong; Huaming Deng; Liangqing Fu; Ronghua Zhang

    2008-01-01

    Gab2 plays an important role in FcεRI mediated signal events which lead to degranulation from mast cells. The present study was designed to investigate the effect of the synthetic Gab2 (scaffolding adapter Grb2-associated binder 2) siRNA on the antigen-induced activation of RBL-2H3 cells. A double stranded siRNA against Gab2mRNA was synthesized and transfected into RBL-2H3 cells. After 6 h, cells were then sensitized with dinitrophenyl (DNP)-specific IgE overnight and challenged with dinitrophenyl-human serum albumin (DNP-HSA) to induce mast cell degranulation before supernatants were collected. Effects of Gab2 siRNA on antigen-induced release of β-hexosaminidase and histamine, cytokine production and regulation of the proteins in the pathway were measured by enzymatic assay, EIA, ELISA and Western blotting. Treatment with Gab2 siRNA significantly decreased Gab2 expression, inhibited the FcεRI-mediated mast cell release of β-hexosaminidase and histamine, reduced the production of IL-4 and TNF-α and inhibited the phosphorylation of Akt, PKCδ and p38 mitogen-activated protein kinase (MAPK). Data showed that Gab2 siRNA could suppress the antigen-induced activation of RBL-2H3 cells and suggested a possible mechanism through inhibition of signaling molecules downstream of Gab2 in the FcεRI-mediated Ca2+-independent pathway. Furthermore, potential usefulness of Gab2 knock-down as a method for inhibition of mast cell-mediated allergic reactions was demonstrated. Cellular & Molecular Immunology. 2008;5(6):433-438.

  7. Loss of proliferation and antigen presentation activity following internalization of polydispersed carbon nanotubes by primary lung epithelial cells.

    Directory of Open Access Journals (Sweden)

    Mandavi Kumari

    Full Text Available Interactions between poly-dispersed acid functionalized single walled carbon nanotubes (AF-SWCNTs and primary lung epithelial (PLE cells were studied. Peritoneal macrophages (PMs, known phagocytic cells were used as positive controls in this study. Recovery of live cells from cultures of PLE cells and PMs was significantly reduced in the presence of AF-SWCNTs, in a time and dose dependent manner. Both PLE cells as well as PMs could take up fluorescence tagged AF-SWCNTs in a time dependent manner and this uptake was significantly blocked by cytochalasin D, an agent that blocks the activity of acto-myosin fibers and therefore the phagocytic activity of cells. Confocal microscopic studies confirmed that AF-SWCNTs were internalized by both PLE cells and PMs. Intra-trachially instilled AF-SWCNTs could also be taken up by lung epithelial cells as well as alveolar macrophages. Freshly isolated PLE cells had significant cell division activity and cell cycling studies indicated that treatment with AF-SWCNTs resulted in a marked reduction in S-phase of the cell cycle. In a previously standardized system to study BCG antigen presentation by PLE cells and PMs to sensitized T helper cells, AF-SWCNTs could significantly lower the antigen presentation ability of both cell types. These results show that mouse primary lung epithelial cells can efficiently internalize AF-SWCNTs and the uptake of nanotubes interfered with biological functions of PLE cells including their ability to present BCG antigens to sensitized T helper cells.

  8. Modeling and analysis of early events in T-lymphocyte antigen-activated intracellular-signaling pathways

    Science.gov (United States)

    Zheng, Yanan; Balakrishnan, Venkataramanan; Buzzard, Greg; Geahlen, Robert; Harrison, Marietta; Rundell, Ann

    2005-12-01

    The T-cell antigen-activated signaling pathway is a highly regulated intracellular biochemical system that is crucial for initiating an appropriate adaptive immune response. To improve the understanding of the complex regulatory mechanisms controlling the early events in T-cell signaling, a detailed mathematical model was developed that utilizes ordinary differential equations to describe chemical reactions of the signaling pathway. The model parameter values were constrained by experimental data on the activation of a specific signaling intermediate and indicated an initial rapid cascade of phosphorylation events followed by a comparatively slow signal downregulation. Nonlinear analysis of the model suggested that thresholding and bistability occur as a result of the embedded positive and negative feedback loops within the model. These nonlinear system properties may enhance the T-cell receptor specificity and provide sub-threshold noise filtering with switch-like behavior to ensure proper cell response. Additional analysis using a reduced second-order model led to further understanding of the observed system behavior. Moreover, the interactions between the positive and negative feedback loops enabled the model to exhibit, among a variety of other feasible dynamics, a sustained oscillation that corresponds to a stable limit cycle in the two-dimensional phase plane. Quantitative analysis in this paper has helped identify potential regulatory mechanisms in the early T-cell signaling events. This integrated approach provides a framework to quantify and discover the ensemble of interconnected T-cell antigen-activated signaling pathways from limited experimental data.

  9. Cytochemical localization of ATP diphosphohydrolase from Leishmania (Viannia) braziliensis promastigotes and identification of an antigenic and catalytically active isoform.

    Science.gov (United States)

    Rezende-Soares, F A; Carvalho-Campos, C; Marques, M J; Porcino, G N; Giarola, N L L; Costa, B L S; Taunay-Rodrigues, A; Faria-Pinto, P; Souza, M A; Diniz, V A; Corte-Real, S; Juliano, M A; Juliano, L; Vasconcelos, E G

    2010-04-01

    An ATP diphosphohydrolase (EC 3.6.1.5) activity was identified in a Leishmania (Viannia) braziliensis promastigotes preparation (Lb). Ultrastructural cytochemical microscopy showed this protein on the parasite surface and also stained a possible similar protein at the mitochondrial membrane. Isolation of an active ATP diphosphohydrolase isoform from Lb was obtained by cross-immunoreactivity with polyclonal anti-potato apyrase antibodies. These antibodies, immobilized on Protein A-Sepharose, immunoprecipitated a polypeptide of approximately 48 kDa and, in lower amount, a polypeptide of approximately 43 kDa, and depleted 83% ATPase and 87% of the ADPase activities from detergent-homogenized Lb. Potato apyrase was recognized in Western blots by IgG antibody from American cutaneous leishmaniasis (ACL) patients, suggesting that the parasite and vegetable proteins share antigenic conserved epitopes. Significant IgG seropositivity in serum samples diluted 1:50 from ACL patients (n=20) for Lb (65%) and potato apyrase (90%) was observed by ELISA technique. Significant IgG antibody reactivity was also observed against synthetic peptides belonging to a conserved domain from L. braziliensis NDPase (80% seropositivity) and its potato apyrase counterpart (50% seropositivity), in accordance with the existence of shared antigenic epitopes and demonstrating that in leishmaniasis infection the domain r82-103 from L. braziliensis NDPase is a target for the human immune response. PMID:19961654

  10. Tumorigenic activity of Merkel cell polyomavirus T antigens expressed in the stratified epithelium of mice

    OpenAIRE

    Spurgeon, Megan E.; Cheng, Jingwei; Bronson, Roderick T.; Lambert, Paul F.; DeCaprio, James A.

    2015-01-01

    Merkel cell polyomavirus (MCPyV) is frequently associated with Merkel cell carcinoma (MCC), a highly aggressive neuroendocrine skin cancer. Most MCC tumors contain integrated copies of the viral genome with persistent expression of the MCPyV large T (LT) and small T (ST) antigen. MCPyV isolated from MCC typically contain wild type ST but truncated forms of LT that retain the N-terminus but delete the C-terminus and render LT incapable of supporting virus replication. To determine the oncogeni...

  11. Exosomes Derived from M. Bovis BCG Infected Macrophages Activate Antigen-Specific CD4+ and CD8+ T Cells In Vitro and In Vivo

    OpenAIRE

    Giri, Pramod K.; Schorey, Jeffrey S.

    2008-01-01

    Activation of both CD4(+) and CD8(+) T cells is required for an effective immune response to an M. tuberculosis infection. However, infected macrophages are poor antigen presenting cells and may be spatially separated from recruited T cells, thus limiting antigen presentation within a granuloma. Our previous studies showed that infected macrophages release from cells small membrane-bound vesicles called exosomes which contain mycobacterial lipid components and showed that these exosomes could...

  12. Dectin-2 Recognizes Mannosylated O-antigens of Human Opportunistic Pathogens and Augments Lipopolysaccharide Activation of Myeloid Cells*

    Science.gov (United States)

    Wittmann, Alexandra; Lamprinaki, Dimitra; Bowles, Kristian M.; Katzenellenbogen, Ewa; Knirel, Yuriy A.; Whitfield, Chris; Nishimura, Takashi; Matsumoto, Naoki; Yamamoto, Kazuo; Iwakura, Yoichiro; Saijo, Shinobu; Kawasaki, Norihito

    2016-01-01

    LPS consists of a relatively conserved region of lipid A and core oligosaccharide and a highly variable region of O-antigen polysaccharide. Whereas lipid A is known to bind to the Toll-like receptor 4 (TLR4)-myeloid differentiation factor 2 (MD2) complex, the role of the O-antigen remains unclear. Here we report a novel molecular interaction between dendritic cell-associated C-type lectin-2 (Dectin-2) and mannosylated O-antigen found in a human opportunistic pathogen, Hafnia alvei PCM 1223, which has a repeating unit of [-Man-α1,3-Man-α1,2-Man-α1,2-Man-α1,2-Man-α1,3-]. H. alvei LPS induced higher levels of TNFα and IL-10 from mouse bone marrow-derived dendritic cells (BM-DCs), when compared with Salmonella enterica O66 LPS, which has a repeat of [-Gal-α1,6-Gal-α1,4-[Glc-β1,3]GalNAc-α1,3-GalNAc-β1,3-]. In a cell-based reporter assay, Dectin-2 was shown to recognize H. alvei LPS. This binding was inhibited by mannosidase treatment of H. alvei LPS and by mutations in the carbohydrate-binding domain of Dectin-2, demonstrating that H. alvei LPS is a novel glycan ligand of Dectin-2. The enhanced cytokine production by H. alvei LPS was Dectin-2-dependent, because Dectin-2 knock-out BM-DCs failed to do so. This receptor cross-talk between Dectin-2 and TLR4 involved events including spleen tyrosine kinase (Syk) activation and receptor juxtaposition. Furthermore, another mannosylated LPS from Escherichia coli O9a also bound to Dectin-2 and augmented TLR4 activation of BM-DCs. Taken together, these data indicate that mannosylated O-antigens from several Gram-negative bacteria augment TLR4 responses through interaction with Dectin-2. PMID:27358401

  13. Dectin-2 Recognizes Mannosylated O-antigens of Human Opportunistic Pathogens and Augments Lipopolysaccharide Activation of Myeloid Cells.

    Science.gov (United States)

    Wittmann, Alexandra; Lamprinaki, Dimitra; Bowles, Kristian M; Katzenellenbogen, Ewa; Knirel, Yuriy A; Whitfield, Chris; Nishimura, Takashi; Matsumoto, Naoki; Yamamoto, Kazuo; Iwakura, Yoichiro; Saijo, Shinobu; Kawasaki, Norihito

    2016-08-19

    LPS consists of a relatively conserved region of lipid A and core oligosaccharide and a highly variable region of O-antigen polysaccharide. Whereas lipid A is known to bind to the Toll-like receptor 4 (TLR4)-myeloid differentiation factor 2 (MD2) complex, the role of the O-antigen remains unclear. Here we report a novel molecular interaction between dendritic cell-associated C-type lectin-2 (Dectin-2) and mannosylated O-antigen found in a human opportunistic pathogen, Hafnia alvei PCM 1223, which has a repeating unit of [-Man-α1,3-Man-α1,2-Man-α1,2-Man-α1,2-Man-α1,3-]. H. alvei LPS induced higher levels of TNFα and IL-10 from mouse bone marrow-derived dendritic cells (BM-DCs), when compared with Salmonella enterica O66 LPS, which has a repeat of [-Gal-α1,6-Gal-α1,4-[Glc-β1,3]GalNAc-α1,3-GalNAc-β1,3-]. In a cell-based reporter assay, Dectin-2 was shown to recognize H. alvei LPS. This binding was inhibited by mannosidase treatment of H. alvei LPS and by mutations in the carbohydrate-binding domain of Dectin-2, demonstrating that H. alvei LPS is a novel glycan ligand of Dectin-2. The enhanced cytokine production by H. alvei LPS was Dectin-2-dependent, because Dectin-2 knock-out BM-DCs failed to do so. This receptor cross-talk between Dectin-2 and TLR4 involved events including spleen tyrosine kinase (Syk) activation and receptor juxtaposition. Furthermore, another mannosylated LPS from Escherichia coli O9a also bound to Dectin-2 and augmented TLR4 activation of BM-DCs. Taken together, these data indicate that mannosylated O-antigens from several Gram-negative bacteria augment TLR4 responses through interaction with Dectin-2. PMID:27358401

  14. Different-Sized Gold Nanoparticle Activator/Antigen Increases Dendritic Cells Accumulation in Liver-Draining Lymph Nodes and CD8+ T Cell Responses.

    Science.gov (United States)

    Zhou, Qianqian; Zhang, Yulong; Du, Juan; Li, Yuan; Zhou, Yong; Fu, Qiuxia; Zhang, Jingang; Wang, Xiaohui; Zhan, Linsheng

    2016-02-23

    The lack of efficient antigen and activator delivery systems, as well as the restricted migration of dendritic cells (DCs) to secondary lymph organs, dramatically limits DC-based adoptive immunotherapy. We selected two spherical gold nanoparticle (AuNP)-based vehicles of optimal size for activator and antigen delivery. Their combination (termed the NanoAu-Cocktail) was associated with the dual targeting of CpG oligonucleotides (CpG-ODNs) and an OVA peptide (OVAp) to DC subcellular compartments, inducing enhanced antigen cross-presentation, upregulated expression of costimulatory molecules and elevated secretion of T helper1 cytokines. We demonstrated that the intravenously transfused NanoAu-Cocktail pulsed DCs showed dramatically improved in vivo homing ability to lymphoid tissues and were settled in T cell area. Especially, by tissue-distribution analysis, we found that more than 60% of lymphoid tissues-homing DCs accumulated in liver-draining lymph nodes (LLNs). The improved homing ability of NanoAu-Cocktail pulsed DCs was associated with the high expression of chemokine receptor 7 (CCR7) and rearrangement of the cytoskeletons. In addition, by antigen-specific tetramers detection, NanoAu-Cocktail pulsed DCs were proved able to elicit strong antigen-specific CD8+ T cell responses, which provided enhanced protection from viral invasions. This study highlights the importance of codelivering antigen/adjuvant using different sized gold nanoparticles to improve DC homing and therapy. PMID:26771692

  15. High-Level Expression of Functionally Active Dengue-2 Non-Structural Antigen 1 Production in Escherichia coli

    Directory of Open Access Journals (Sweden)

    S. Gowri Sankar

    2013-01-01

    Full Text Available Detection of nonstructural protein (NS1 is an important diagnostic marker during acute phase of dengue infection. Not only for diagnostic purpose, the protein had important role in vaccine design as well, as a candidate for studying virus assembly and maturation. Various researchers employed different expression systems and strategies for recombinant NS1 protein production. Attempts to express NS1 protein in prokaryotic and yeast expression system result in formation of insoluble protein which needs to undergo refolding to attain native structural and functional forms. Here, we report the production of soluble NS1 protein in E. coli by using appropriate vector and employing suitable culture conditions to maximize protein production. Proteins were purified using metal affinity chromatography. SDS-PAGE and western blot analysis reveal the native structure of NS1 protein. Solid phase ELISA using the recombinantly expressed antigen with positive and negative dengue samples showed that the expressed protein retains its antigenic and immunological properties. To our knowledge, this is the first report on the successful production of functionally active recombinant dengue-2 NS1 protein production without undergoing any in vitro posttranslational modification process.

  16. Expression of Major Antigen Domains of Gene of E2 CSFV and Analysis of its Immunological Activity

    Institute of Scientific and Technical Information of China (English)

    Hong TIAN; Xiang-tao LIU; Jing-yan WU; You-jun SHANG; Tao JIANG; Hai-xue ZHENG; Qing-ge XIE

    2008-01-01

    E2 is an envelope glycoprotein of Classical swine fever virus (CSFV) and contains sequential neutralizing epitopes to induce virus-neutralizing antibodies and mount protective immunity in the natural host. In this study, four antigen domains (ABCD) of the E2 gene was cloned from CSFV Shimen strain into the retroviral vector pBABE puro and expressed in eukaryotic cell (PK15) by an retroviral gene expression system, and the activity of recombinant E2 protein to induce immune responses was evaluated in rabbits. The results indicated that recombinant E2 protein can be recognized by fluorescence antibodies of CSFV and CSFV positive serum (Lanzhou Veterinary Research Institute, Chinese Academy of Agricultural Sciences, Lanzhou, China) using Western blot, indirect immunofluorescence antibody test (IFAT) and ELISA, Furthermore, anti-CSFV specific antibodies and lymphocyte proliferation were elicited and increased by recombinant protein after vaccination. In the challenge test, all of rabbits vaccinated with recombinant protein and Chinese vaccine strain (C-strain) were fully protected from a rabbit spleen virus challenge. These results indicated that a retroviral-based epitope-vaccine carrying the major antigen domains of E2 is able to induce high level of epitope-specific antibodies and exhibits similar protective capability with that induced by the C-strain, and encourages further work towards the development of a vaccine against CSFV infection.

  17. Expression of the K54 and O4 specific antigen has opposite effects on the bactericidal activity of squalamine against an extraintestinal isolate of Escherichia coli.

    Science.gov (United States)

    Russo, T A; Mylotte, D

    1998-05-15

    Squalamine is a novel cationic steroid that possesses potent, broad spectrum, antimicrobial activity. Recent data suggests that squalamine or related compounds may be present and important in host resistance to infection in the urinary tract. Therefore, the role of the K54 capsule and the O4 specific antigen moiety of the lipopolysaccharide in protecting an extraintestinal isolate of Escherichia coli against the bactericidal activity of this novel antimicrobial compound was studied. The O4 specific antigen was important for protection against squalamine. Surprisingly, in contrast, the presence of the K54 antigen enhanced the bactericidal activity of squalamine. This is the first example, to our knowledge, in which an established virulence trait, the K54 capsule, may be detrimental to an infecting pathogen under certain circumstances. PMID:9627966

  18. Activation of Type II Cells into Regenerative Stem Cell Antigen-1+ Cells during Alveolar Repair

    Science.gov (United States)

    Kumar, Varsha Suresh; Zhang, Wei; Rehman, Jalees; Malik, Asrar B.

    2015-01-01

    The alveolar epithelium is composed of two cell types: type I cells comprise 95% of the gas exchange surface area, whereas type II cells secrete surfactant, while retaining the ability to convert into type I cells to induce alveolar repair. Using lineage-tracing analyses in the mouse model of Pseudomonas aeruginosa–induced lung injury, we identified a population of stem cell antigen (Sca)-1–expressing type II cells with progenitor cell properties that mediate alveolar repair. These cells were shown to be distinct from previously reported Sca-1–expressing bronchioalveolar stem cells. Microarray and Wnt reporter studies showed that surfactant protein (Sp)-C+Sca-1+ cells expressed Wnt signaling pathway genes, and inhibiting Wnt/β-catenin signaling prevented the regenerative function of Sp-C+Sca-1+ cells in vitro. Thus, P. aeruginosa–mediated lung injury induces the generation of a Sca-1+ subset of type II cells. The progenitor phenotype of the Sp-C+Sca-1+ cells that mediates alveolar epithelial repair might involve Wnt signaling. PMID:25474582

  19. Interaction between M-CSF and IL-10 on productions of IL-12 and IL-18 and expressions of CD14, CD23, and CD64 by human monocytes

    Institute of Scientific and Technical Information of China (English)

    Xiao-hui JI; Ting YAO; Jun-chuan QIN; Shu-kui WANG; Hui-juan WANG; Kun YAO

    2004-01-01

    AIM: To Study the interaction of macrophage colony-stimulating factor (M-CSF) and interleukin-10 (IL-10) in productions of IL-12 and IL-18 and expressions of CD14, CD23, and CD64 by human monocytes. METHODS:Purified adherent human monocytes were cultured with M-CSF or IL-10 alone, or with M-CSF+IL-10 and 2-3d later, the culture supernatants and cells were separated and collected. IL-12P40 and IL-18 levels in the supernatants were determined by ELISA and the percentages of CD14, CD23, and CD64 positive cells were examined by flow cytometry. RESULTS: (1) IL-10 decreased M-CSF-induced IL-18 levels, while M-CSF further reduced IL-12P40 level in the culture supernatants of IL-10-treated monocytes; (2) IL-10 alone had no effect on the percentage of CD14-positive cells, but further increased the percentage of CD14-positive cells induced by M-CSF; M-CSF alone had no effect on the percentage of CD64-positive cells, but further increased the percentage of CD64-positive cells induced by IL-10; (3) IL-10 decreased the percentage of CD23-positive cells induced by M-CSF.CONCLUSION: Between M-CSF and IL-10, there were antagonistic effects on inducing IL-18 and CD23 expressions by monocytes; there were also synergistic effects on inhibiting IL-12P40 production and inducing CD 14 and CD64 expressions by monocytes.

  20. Functional Analysis of the Glucuronyltransferases GlcAT-P and GlcAT-S of Drosophila melanogaster: Distinct Activities towards the O-linked T-antigen.

    Science.gov (United States)

    Breloy, Isabelle; Schwientek, Tilo; Althoff, Deborah; Holz, Marvin; Koppen, Tim; Krupa, Angelika; Hanisch, Franz-Georg

    2016-01-01

    The Drosophila melanogaster glucuronyltransferases dGlcAT-S and dGlcAT-P were reported to be expressed ubiquitously and results of in vitro activity assays indicate a functional redundancy. We analyzed both transferases in vivo and in vitro and could show significant differences in their activity towards N-and O-glycoproteins in vivo. While GlcAT-P is able to use N-linked N-acetyllactosamine chains and the O-linked T-antigen as a substrate to form non-sulfated HNK1- (GlcAβ1-3Galβ1-4GlcNAcβ1-) and glucuronyl-T-antigens in vivo, GlcAT-S adds glucuronic acid only to N-linked chains, thereby synthesizing only the non-sulfated HNK1-antigen. PMID:26751495

  1. Functional Analysis of the Glucuronyltransferases GlcAT-P and GlcAT-S of Drosophila melanogaster: Distinct Activities towards the O-linked T-antigen

    Directory of Open Access Journals (Sweden)

    Isabelle Breloy

    2016-01-01

    Full Text Available The Drosophila melanogaster glucuronyltransferases dGlcAT-S and dGlcAT-P were reported to be expressed ubiquitously and results of in vitro activity assays indicate a functional redundancy. We analyzed both transferases in vivo and in vitro and could show significant differences in their activity towards N-and O-glycoproteins in vivo. While GlcAT-P is able to use N-linked N-acetyllactosamine chains and the O-linked T-antigen as a substrate to form non-sulfated HNK1- (GlcAβ1-3Galβ1-4GlcNAcβ1- and glucuronyl-T-antigens in vivo, GlcAT-S adds glucuronic acid only to N-linked chains, thereby synthesizing only the non-sulfated HNK1-antigen.

  2. Exosomes derived from M. Bovis BCG infected macrophages activate antigen-specific CD4+ and CD8+ T cells in vitro and in vivo.

    Directory of Open Access Journals (Sweden)

    Pramod K Giri

    Full Text Available Activation of both CD4(+ and CD8(+ T cells is required for an effective immune response to an M. tuberculosis infection. However, infected macrophages are poor antigen presenting cells and may be spatially separated from recruited T cells, thus limiting antigen presentation within a granuloma. Our previous studies showed that infected macrophages release from cells small membrane-bound vesicles called exosomes which contain mycobacterial lipid components and showed that these exosomes could stimulate a pro-inflammatory response in naïve macrophages. In the present study we demonstrate that exosomes stimulate both CD4(+ and CD8(+ splenic T cells isolated from mycobacteria-sensitized mice. Although the exosomes contain MHC I and II as well as costimulatory molecules, maximum stimulation of T cells required prior incubation of exosomes with antigen presenting cells. Exosomes isolated from M. bovis and M. tuberculosis infected macrophages also stimulated activation and maturation of mouse bone marrow-derived dendritic cells. Interestingly, intranasal administration of mice with exosomes isolated from M. bovis BCG infected macrophages induce the generation of memory CD4(+ and CD8(+ T cells. The isolated T cells also produced IFN-gamma upon restimulation with BCG antigens. The release of exosomes from infected macrophages may overcome some of the defects in antigen presentation associated with mycobacterial infections and we suggest that exosomes may be a promising M. tuberculosis vaccine candidate.

  3. Activation of Cdc42 is necessary for sustained oscillations of Ca2+ and PIP2 stimulated by antigen in RBL mast cells

    Directory of Open Access Journals (Sweden)

    Marcus M. Wilkes

    2014-07-01

    Full Text Available Antigen stimulation of mast cells via FcεRI, the high-affinity receptor for IgE, triggers a signaling cascade that requires Ca2+ mobilization for exocytosis of secretory granules during the allergic response. To characterize the role of Rho GTPases in FcεRI signaling, we utilized a mutant RBL cell line, B6A4C1, that is deficient in antigen-stimulated Cdc42 activation important for these processes. Recently the importance of stimulated intracellular oscillations has emerged, and we find that B6A4C1 cells exhibit severely attenuated Ca2+ oscillations in response to antigen, which are restored to wild-type RBL-2H3 levels by expression of constitutively active Cdc42 G12V or by a GEF for Cdc42, DOCK7, but not when the C-terminal di-arginine motif of active Cdc42 is mutated to di-glutamine. We found that antigen-stimulated FcεRI endocytosis, which occurs independently of Ca2+ mobilization, is also defective in B6A4C1 cells, and Cdc42 G12V reconstitutes this response as well. Thus, activation of Cdc42 occurs prior to and is critical for antigen-stimulated pathways leading separately to both Ca2+ mobilization and receptor endocytosis. Accounting for these downstream functional consequences, we show that Cdc42 G12V reconstitutes antigen-stimulated oscillations of phosphatidylinositol 4,5-bisphosphate (PIP2 at the plasma membrane in mutant B6A4C1 cells, pointing to Cdc42 participation in the regulation of stimulated PIP2 synthesis.

  4. High levels of tissue plasminogen activator (tPA antigen precede the development of type 2 diabetes in a longitudinal population study. The Northern Sweden MONICA Study

    Directory of Open Access Journals (Sweden)

    Jansson Jan-Håkan

    2003-12-01

    Full Text Available Abstract Background Impaired fibrinolysis is found in impaired glucose tolerance and type 2 diabetes, associated with components of the metabolic syndrome. There are no data concerning fibrinolysis in subjects with normal glucose tolerance that convert to diabetes. Methods We studied the activities of tissue plasminogen activator (tPA and plasminogen activator inhibitor-1 (PAI-1 and the levels of tPA antigen (a marker of endothelial dysfunction in 551 subjects with normal glucose tolerance in 1990 in relation to incident diabetes during nine years of follow-up. Results Subjects with diabetes at follow-up (n = 15 had significantly lower baseline tPA activity and higher PAI-1 activity and tPA antigen than non-converters. The risk of diabetes increased linearly across quartiles of PAI-activity (p = 0.007 and tPA antigen (p p = 0.026. The risk of diabetes with low tPA activity or high PAI-1 activity persisted after adjustment for age and sex but diminished to a non-significant level after further adjustments. The odds ratio of diabetes for high tPA antigen was 10.4 (95% confidence interval 2.7–40 adjusted for age and sex. After further adjustment for diastolic blood pressure, waist circumference, insulin, triglycerides, fasting and post load glucose the odds ratio was 6.5 (1.3–33, p = 0.024. Conclusions Impaired fibrinolysis and endothelial dysfunction are evident in subjects with normal glucose tolerance who later develop diabetes. High tPA antigen is predictive of future diabetes independent from the metabolic syndrome.

  5. Activation of OX40 Augments Th17 Cytokine Expression and Antigen-Specific Uveitis

    NARCIS (Netherlands)

    Zhong, Wenwei; Hinrichs, David; Wu, Xiumei; Weinberg, Andrew; Hall, Mark; Spencer, Doran; Wegmann, Keith; Rosenbaum, James T.

    2010-01-01

    Uveitis is a major and common cause of visual disability Recent studies have shown that Th17 cells are implicated in the pathogenesis of this serious intraocular disorder Activated T cells express an inducible costimulatory molecule called OX40 and OX40 in turn promotes the activation and proliferat

  6. Structural Insights into the Protease-like Antigen Plasmodium falciparum SERA5 and Its Noncanonical Active-Site Serine

    Energy Technology Data Exchange (ETDEWEB)

    Hodder, Anthony N.; Malby, Robyn L.; Clarke, Oliver B.; Fairlie, W. Douglas; Colman, Peter M.; Crabb, Brendan S.; Smith, Brian J.; (WEHIMR); (Melbourne)

    2009-08-28

    The sera genes of the malaria-causing parasite Plasmodium encode a family of unique proteins that are maximally expressed at the time of egress of parasites from infected red blood cells. These multi-domain proteins are unique, containing a central papain-like cysteine-protease fragment enclosed between the disulfide-linked N- and C-terminal domains. However, the central fragment of several members of this family, including serine repeat antigen 5 (SERA5), contains a serine (S596) in place of the active-site cysteine. Here we report the crystal structure of the central protease-like domain of Plasmodium falciparum SERA5, revealing a number of anomalies in addition to the putative nucleophilic serine: (1) the structure of the putative active site is not conducive to binding substrate in the canonical cysteine-protease manner; (2) the side chain of D594 restricts access of substrate to the putative active site; and (3) the S{sub 2} specificity pocket is occupied by the side chain of Y735, reducing this site to a small depression on the protein surface. Attempts to determine the structure in complex with known inhibitors were not successful. Thus, despite having revealed its structure, the function of the catalytic domain of SERA5 remains an enigma.

  7. Complement receptor type two (CR2,CR21): a target for influencing the humoral immune response and antigen-trapping.

    Science.gov (United States)

    Prodinger, W M

    1999-01-01

    Cellular receptors for complement C3 fragments deposited on antigens are important bricks in the wall defending against microbial pathogens. The part of complement receptor type 2 (CR2; CD21) deals with enhancing humoral immune responses and with long-term trapping of C3d-coated antigen by follicular dendritic cells. CR2 is also pivotal for Epstein-Barr virus (EBV) infection. Here, the current understanding, how CR2 interacts with its ligands C3d, EBV, and CD23 is summarized. The potential to target CR2 for clinical therapy or immunization purposes are discussed. PMID:10741859

  8. GD2-specific CAR T Cells Undergo Potent Activation and Deletion Following Antigen Encounter but can be Protected From Activation-induced Cell Death by PD-1 Blockade.

    Science.gov (United States)

    Gargett, Tessa; Yu, Wenbo; Dotti, Gianpietro; Yvon, Eric S; Christo, Susan N; Hayball, John D; Lewis, Ian D; Brenner, Malcolm K; Brown, Michael P

    2016-06-01

    Chimeric antigen receptor (CAR) T cells have shown great promise in the treatment of hematologic malignancies but more variable results in the treatment of solid tumors and the persistence and expansion of CAR T cells within patients has been identified as a key correlate of antitumor efficacy. Lack of immunological "space", functional exhaustion, and deletion have all been proposed as mechanisms that hamper CAR T-cell persistence. Here we describe the events following activation of third-generation CAR T cells specific for GD2. CAR T cells had highly potent immediate effector functions without evidence of functional exhaustion in vitro, although reduced cytokine production reversible by PD-1 blockade was observed after longer-term culture. Significant activation-induced cell death (AICD) of CAR T cells was observed after repeated antigen stimulation, and PD-1 blockade enhanced both CAR T-cell survival and promoted killing of PD-L1(+) tumor cell lines. Finally, we assessed CAR T-cell persistence in patients enrolled in the CARPETS phase 1 clinical trial of GD2-specific CAR T cells in the treatment of metastatic melanoma. Together, these data suggest that deletion also occurs in vivo and that PD-1-targeted combination therapy approaches may be useful to augment CAR T-cell efficacy and persistence in patients.

  9. GD2-specific CAR T Cells Undergo Potent Activation and Deletion Following Antigen Encounter but can be Protected From Activation-induced Cell Death by PD-1 Blockade.

    Science.gov (United States)

    Gargett, Tessa; Yu, Wenbo; Dotti, Gianpietro; Yvon, Eric S; Christo, Susan N; Hayball, John D; Lewis, Ian D; Brenner, Malcolm K; Brown, Michael P

    2016-06-01

    Chimeric antigen receptor (CAR) T cells have shown great promise in the treatment of hematologic malignancies but more variable results in the treatment of solid tumors and the persistence and expansion of CAR T cells within patients has been identified as a key correlate of antitumor efficacy. Lack of immunological "space", functional exhaustion, and deletion have all been proposed as mechanisms that hamper CAR T-cell persistence. Here we describe the events following activation of third-generation CAR T cells specific for GD2. CAR T cells had highly potent immediate effector functions without evidence of functional exhaustion in vitro, although reduced cytokine production reversible by PD-1 blockade was observed after longer-term culture. Significant activation-induced cell death (AICD) of CAR T cells was observed after repeated antigen stimulation, and PD-1 blockade enhanced both CAR T-cell survival and promoted killing of PD-L1(+) tumor cell lines. Finally, we assessed CAR T-cell persistence in patients enrolled in the CARPETS phase 1 clinical trial of GD2-specific CAR T cells in the treatment of metastatic melanoma. Together, these data suggest that deletion also occurs in vivo and that PD-1-targeted combination therapy approaches may be useful to augment CAR T-cell efficacy and persistence in patients. PMID:27019998

  10. Active immunization with an octa-valent Staphylococcus aureus antigen mixture in models of S. aureus bacteremia and skin infection in mice.

    Directory of Open Access Journals (Sweden)

    Sanne van den Berg

    Full Text Available Proteomic studies with different Staphylococcus aureus isolates have shown that the cell surface-exposed and secreted proteins IsaA, LytM, Nuc, the propeptide of Atl (pro-Atl and four phenol-soluble modulins α (PSMα are invariantly produced by this pathogen. Therefore the present study was aimed at investigating whether these proteins can be used for active immunization against S. aureus infection in mouse models of bacteremia and skin infection. To this end, recombinant His-tagged fusions of IsaA, LytM, Nuc and pro-Atl were isolated from Lactococcus lactis or Escherichia coli, while the PSMα1-4 peptides were chemically synthesized. Importantly, patients colonized by S. aureus showed significant immunoglobulin G (IgG responses against all eight antigens. BALB/cBYJ mice were immunized subcutaneously with a mixture of the antigens at day one (5 μg each, and boosted twice (25 μg of each antigen with 28 days interval. This resulted in high IgG responses against all antigens although the response against pro-Atl was around one log lower compared to the other antigens. Compared to placebo-immunized mice, immunization with the octa-valent antigen mixture did not reduce the S. aureus isolate P load in blood, lungs, spleen, liver, and kidneys in a bacteremia model in which the animals were challenged for 14 days with a primary load of 3 × 10(5 CFU. Discomfort scores and animal survival rates over 14 days did not differ between immunized mice and placebo-immunized mice upon bacteremia with S. aureus USA300 (6 × 10(5 CFU. In addition, this immunization did not reduce the S. aureus isolate P load in mice with skin infection. These results show that the target antigens are immunogenic in both humans and mice, but in the used animal models do not result in protection against S. aureus infection.

  11. Temporal differences in the activation of three classes of non-transmembrane protein tyrosine kinases following B-cell antigen receptor surface engagement.

    Science.gov (United States)

    Saouaf, S J; Mahajan, S; Rowley, R B; Kut, S A; Fargnoli, J; Burkhardt, A L; Tsukada, S; Witte, O N; Bolen, J B

    1994-09-27

    We evaluated in WEHI 231 B cells the time-dependent responses of Lyn, Blk, Btk, Syk, and three members of the Jak family of protein tyrosine kinases following antibody-mediated surface engagement of the B-cell antigen receptor. Our results show that the enzyme activities of Lyn and Blk were stimulated within seconds of antigen receptor engagement and correlated with the initial tyrosine phosphorylation of the Ig alpha and Ig beta subunits of the B-cell antigen receptor. Btk enzyme activity was also transiently stimulated and was maximal at approximately 5 min after B-cell receptor surface binding. Syk activity gradually increased to a maximum at 10-30 min following receptor ligation and was found to parallel the association of Syk with the tyrosine phosphorylated Ig alpha and Ig beta subunits of the receptor. While the specific activities of the Jak1, Jak2, and Tyk2 protein tyrosine kinases were unaltered following B-cell receptor ligation, the abundance of Jak1 and Jak2 were increased 3- to 4-fold within 10 min of receptor engagement. These results demonstrate that multiple families of non-transmembrane protein tyrosine kinases are temporally regulated during the process of B-cell antigen receptor-initiated intracellular signal transduction. PMID:7524079

  12. Wild-type, but not mutant, human p53 proteins inhibit the replication activities of simian virus 40 large tumor antigen.

    OpenAIRE

    Friedman, P N; Kern, S. E.; Vogelstein, B; Prives, C

    1990-01-01

    Murine p53 blocks many of the replication activities of simian virus 40 (SV40) large tumor antigen (T antigen) in vitro. As murine cells do not replicate SV40 DNA, it was of interest to determine how p53 from permissive human cells functions. Recombinant baculoviruses encoding either the wild-type form of human p53 or a mutant p53 cloned from a human tumor cell line were constructed, and p53 proteins were purified from infected insect cells. Surprisingly, we found that wild-type human p53 was...

  13. Immunological recovery and dose evaluation in IFN-alpha treatment of hairy cell leukemia: analysis of leukocyte differentiation antigens, NK and 2',5'-oligoadenylate synthetase activity

    DEFF Research Database (Denmark)

    Nielsen, B; Hokland, M; Justesen, J;

    1989-01-01

    A low-dose interferon (IFN)-alpha regimen for the treatment of hairy cell leukemia (HCL) was evaluated by following changes in leukocyte differentiation antigens (LDA), natural killer cell (NK) and 2',5'-oligoadenylate (2-5A) synthetase activities. Due to hairy cells' (HC) weak expression...... of several antigens positive for T cells, B cells, NK cells and monocytes, the use of a double marker specific for hairy cells was needed to distinguish the different subpopulations. Analysis of LDA in peripheral blood (PB) showed a total normalization of the T cell and monocyte numbers within 90 days...

  14. Non-major histocompatibility complex-restricted cytotoxic activity of blood mononuclear cells stimulated with secreted mycobacterial proteins and other mycobacterial antigens

    DEFF Research Database (Denmark)

    Ravn, P; Pedersen, B K

    1994-01-01

    Several observations indicate that non-major histocompatibility complex (MHC)-restricted cytotoxicity, mediated for example by natural killer cells and lymphokine-activated killer cells, may serve as an important antimicrobial defense mechanism. The purpose of the present study was to investigate...... the influences of different mycobacterial antigens on non-MHC-restricted cytotoxicity and further to investigate the ways by which various lymphocyte subpopulations contribute to the development of this cytotoxicity. Non-MHC-restricted cytotoxicity was induced following stimulation of mononuclear cells......+ cells proliferated and expressed interleukin-2 receptors following stimulation with mycobacterial antigens. Depletion studies after antigen stimulation showed that the cytotoxic effector cells were CD16+ CD56+ and CD4-; the CD4+ cells alone did not mediate non-MHC-restricted cytotoxicity. To evaluate...

  15. Non-major histocompatibility complex-restricted cytotoxic activity of blood mononuclear cells stimulated with secreted mycobacterial proteins and other mycobacterial antigens

    DEFF Research Database (Denmark)

    Ravn, P; Pedersen, B K

    1994-01-01

    Several observations indicate that non-major histocompatibility complex (MHC)-restricted cytotoxicity, mediated for example by natural killer cells and lymphokine-activated killer cells, may serve as an important antimicrobial defense mechanism. The purpose of the present study was to investigate...... the influences of different mycobacterial antigens on non-MHC-restricted cytotoxicity and further to investigate the ways by which various lymphocyte subpopulations contribute to the development of this cytotoxicity. Non-MHC-restricted cytotoxicity was induced following stimulation of mononuclear...... interferon. The CD4+ cells proliferated and expressed interleukin-2 receptors following stimulation with mycobacterial antigens.Depletion studies after antigen stimulation showed that the cytotoxic effector cells were CD16+ CD56+ and CD4-; the CD4+ cells alone did not mediate non-MHC-restricted cytotoxicity...

  16. Activated Complement Component 3 (C3) Is Required for Ultraviolet Induction of Immunosuppression and Antigenic Tolerance

    OpenAIRE

    Hammerberg, Craig; Santosh K. Katiyar; Carroll, Michael C.; Cooper, Kevin D

    1998-01-01

    Complement component 3 (C3), a critical regulator of innate immunity, may also play a role in the regulation of cognate immunity, such as contact sensitivity responses. Because ultraviolet (UV) radiation also activates C3 in the skin, we determined whether the immunosuppressed state that results when a contact sensitizer is applied through UVB-exposed skin requires the presence and activation of C3. This question was addressed through the use of C3-deficient mice, blockade of C3 cleavage to C...

  17. Partial in vitro analysis of toxic and antigenic activities of eleven Peruvian pitviper snake venoms.

    Science.gov (United States)

    Guerra-Duarte, C; Lopes-Peixoto, J; Fonseca-de-Souza, B R; Stransky, S; Oliveira, D; Schneider, F S; Lopes-de-Souza, L; Bonilla, C; Silva, W; Tintaya, B; Yarleque, A; Chávez-Olórtegui, C

    2015-12-15

    This work used eleven Peruvian snake venoms (Bothrops andianus, Bothrops atrox, Bothrops barnetti, Bothrops castelnaudi, Bothriopsis chloromelas, Bothrocophias microphthalmus, Bothrops neuwiedi, Bothriopsis oligolepis, Bothriopsis peruviana, Bothrops pictus and Bothriopsis taeniata) to perform in vitro experimentation and determine its main characteristics. Hyaluronidase (HYAL), phospholipase A2 (PLA2), snake venom metalloproteinase (SVMP), snake venom serine protease (SVSP) and L-amino acid oxidase (LAAO) activities; toxicity by cell viability assays using MGSO3, VERO and HeLa cell lineages; and crossed immunoreactivity with Peruvian (PAV) and Brazilian (BAV) antibothropic polyvalent antivenoms, through ELISA and Western Blotting assays, were determined. Results show that the activities tested in this study were not similar amongst the venoms and each species present their own peculiarities, highlighting the diversity within Bothrops complex. All venoms were capable of reducing cell viability of all tested lineages. It was also demonstrated the crossed recognition of all tested venoms by both antivenoms. PMID:26365916

  18. A viral vectored prime-boost immunization regime targeting the malaria Pfs25 antigen induces transmission-blocking activity.

    Directory of Open Access Journals (Sweden)

    Anna L Goodman

    Full Text Available The ookinete surface protein Pfs25 is a macrogamete-to-ookinete/ookinete stage antigen of Plasmodium falciparum, capable of exerting high-level anti-malarial transmission-blocking activity following immunization with recombinant protein-in-adjuvant formulations. Here, this antigen was expressed in recombinant chimpanzee adenovirus 63 (ChAd63, human adenovirus serotype 5 (AdHu5 and modified vaccinia virus Ankara (MVA viral vectored vaccines. Two immunizations were administered to mice in a heterologous prime-boost regime. Immunization of mice with AdHu5 Pfs25 at week 0 and MVA Pfs25 at week 10 (Ad-MVA Pfs25 resulted in high anti-Pfs25 IgG titers, consisting of predominantly isotypes IgG1 and IgG2a. A single priming immunization with ChAd63 Pfs25 was as effective as AdHu5 Pfs25 with respect to ELISA titers at 8 weeks post-immunization. Sera from Ad-MVA Pfs25 immunized mice inhibited the transmission of P. falciparum to the mosquito both ex vivo and in vivo. In a standard membrane-feeding assay using NF54 strain P. falciparum, oocyst intensity in Anopheles stephensi mosquitoes was significantly reduced in an IgG concentration-dependent manner when compared to control feeds (96% reduction of intensity, 78% reduction in prevalence at a 1 in 5 dilution of sera. In addition, an in vivo transmission-blocking effect was also demonstrated by direct feeding of immunized mice infected with Pfs25DR3, a chimeric P. berghei line expressing Pfs25 in place of endogenous Pbs25. In this assay the density of Pfs25DR3 oocysts was significantly reduced when mosquitoes were fed on vaccinated as compared to control mice (67% reduction of intensity, 28% reduction in prevalence and specific IgG titer correlated with efficacy. These data confirm the utility of the adenovirus-MVA vaccine platform for the induction of antibodies with transmission-blocking activity, and support the continued development of this alternative approach to transmission-blocking malaria subunit

  19. Structural parameterization and functional prediction of antigenic polypeptome sequences with biological activity through quantitative sequence-activity models (QSAM) by molecular electronegativity edge-distance vector (VMED)

    Institute of Scientific and Technical Information of China (English)

    2007-01-01

    Only from the primary structures of peptides, a new set of descriptors called the molecular electronegativity edge-distance vector (VMED) was proposed and applied to describing and characterizing the molecular structures of oligopeptides and polypeptides, based on the electronegativity of each atom or electronic charge index (ECI) of atomic clusters and the bonding distance between atom-pairs. Here, the molecular structures of antigenic polypeptides were well expressed in order to propose the automated technique for the computerized identification of helper T lymphocyte (Th) epitopes. Furthermore, a modified MED vector was proposed from the primary structures of polypeptides, based on the ECI and the relative bonding distance of the fundamental skeleton groups. The side-chains of each amino acid were here treated as a pseudo-atom. The developed VMED was easy to calculate and able to work. Some quantitative model was established for 28 immunogenic or antigenic polypeptides (AGPP) with 14 (1―14) Ad and 14 other restricted activities assigned as "1"(+) and "0"(-), respectively. The latter comprised 6 Ab(15-20), 3 Ak(21-23), 2 Ek(24-26), 2 H-2k(27 and 28) restricted sequences. Good results were obtained with 90% correct classification (only 2 wrong ones for 20 training samples) and 100% correct prediction (none wrong for 8 testing samples); while contrastively 100% correct classification (none wrong for 20 training samples) and 88% correct classification (1 wrong for 8 testing samples). Both stochastic samplings and cross validations were performed to demonstrate good performance. The described method may also be suitable for estimation and prediction of classes I and II for major histocompatibility antigen (MHC) epitope of human. It will be useful in immune identification and recognition of proteins and genes and in the design and development of subunit vaccines. Several quantitative structure activity relationship (QSAR) models were developed for various

  20. Iron oxide superparamagnetic nanoparticles conjugated with a conformationally blocked α-Tn antigen mimetic for macrophage activation

    Science.gov (United States)

    Manuelli, Massimo; Fallarini, Silvia; Lombardi, Grazia; Sangregorio, Claudio; Nativi, Cristina; Richichi, Barbara

    2014-06-01

    Among new therapies to fight tumors, immunotherapy is still one of the most promising and intriguing. Thanks to the ongoing structural elucidation of several tumor antigens and the development of innovative antigen carriers, immunotherapy is in constant evolution and it is largely used either alone or in synergy with chemotherapy/radiotherapy. With the aim to develop fully synthetic immunostimulants we have recently developed a mimetic of the α-Tn mucin antigen, a relevant tumor antigen. The 4C1 blocked mimetic 1, unique example of an α-Tn mimetic antigen, was functionalized with an ω-phosphonate linker and used to decorate iron oxide superparamagnetic nanoparticles (MNPs), employed as multivalent carriers. MNPs, largely exploited for supporting and carrying biomolecules, like antibodies, drugs or antigens, consent to combine in the same nanometric system the main features of an inorganic magnetic core with a bioactive organic coating. The superparamagnetic glyconanoparticles obtained, named GMNPs, are indeed biocompatible and immunoactive, and they preserve suitable characteristics for use as heat mediators in the magnetic fluid hyperthermia (MFH) treatment of tumors. All together these properties make GMNPs attracting devices for innovative tumor treatment.Among new therapies to fight tumors, immunotherapy is still one of the most promising and intriguing. Thanks to the ongoing structural elucidation of several tumor antigens and the development of innovative antigen carriers, immunotherapy is in constant evolution and it is largely used either alone or in synergy with chemotherapy/radiotherapy. With the aim to develop fully synthetic immunostimulants we have recently developed a mimetic of the α-Tn mucin antigen, a relevant tumor antigen. The 4C1 blocked mimetic 1, unique example of an α-Tn mimetic antigen, was functionalized with an ω-phosphonate linker and used to decorate iron oxide superparamagnetic nanoparticles (MNPs), employed as multivalent

  1. Inhibition of Replication and Transcription Activator and Latency-Associated Nuclear Antigen of Kaposi’s Sarcoma-Associated Herpesvirus by Morpholino Oligomers

    OpenAIRE

    Zhang, Yan-Jin; Wang, Kai-Yu; Stein, David A.; Patel, Deendayal; Watkins, Rheba; Moulton, Hong M.; Iversen, Patrick L.; Matson, David O.

    2006-01-01

    Kaposi’s sarcoma-associated herpesvirus (KSHV) is associated with Kaposi’s sarcoma and primary effusion lymphoma (PEL). The KSHV replication and transcription activator (RTA) and latency-associated nuclear antigen (LANA) play key roles in activating KSHV lytic replication and maintaining KSHV latency, respectively. Phosphorodiamidate morpholino oligomers (PMO) are similar to short single-stranded DNA oligomers, but possess a modified backbone that confers highly specific binding and resistanc...

  2. Proliferating cell nuclear antigen (PCNA) activity in hepatocellular carcinoma, benign peri-neoplastic and normal liver.

    Science.gov (United States)

    Mun, Kein-Seong; Cheah, Phaik-Leng; Baharudin, Nurul Bahiyah; Looi, Lai-Meng

    2006-12-01

    Hepatocellular carcinoma (HCC) is among the ten most common cancers in Malaysian males. As cellular proliferation is an important feature of malignant transformation, we studied the proliferation pattern of normal and benign perineoplastic liver versus hepatocellular carcinoma in an attempt to further understand the tumour transformation process. 39 HCC (21 with accompanying and 18 without cirrhosis) histologically diagnosed at the Department of Pathology, University of Malaya Medical Centre between January 1992 and December 2003 were immunohistochemically studied using a monoclonal antibody to PCNA (Clone PC10: Dako). 20 livers from cases who had succumbed to traumatic injuries served as normal liver controls (NL). PCNA labeling index (PCNA-LI) was determined by counting the number of immunopositive cells in 1000 contiguous HCC, benign cirrhotic perineoplastic liver (BLC), benign perineoplastic non-cirrhotic (BLNC) and NL cells and conversion to a percentage. The PCNA-LI was also expressed as Ojanguren et al's grades. PCNA was expressed in 10% NL, 38.9% BLNC, 76.2% BLC and 71.8% HCC with BLNC, BLC and HCC showing significantly increased (p cells being immunopositive) immunoreactivity on Ojanguren et al's grading system and only HCC demonstrated immunoreactivity which ranged up to grade 3 (75% of cells). From this study, there appears to be a generally increasing trend of proliferative activity from NL to BLNC to BLC and HCC. Nonetheless, BLNC and BLC, like NL, retained low PCNA-LI and only HCC had a significantly increased PCNA-LI compared with the benign categories. This is probably related to the malignant nature of HCC and may reflect the uncontrolled proliferation of the neoplastic hepatocytes. PMID:18376794

  3. Antigen Export Reduces Antigen Presentation and Limits T Cell Control of M. tuberculosis.

    Science.gov (United States)

    Srivastava, Smita; Grace, Patricia S; Ernst, Joel D

    2016-01-13

    Persistence of Mycobacterium tuberculosis results from bacterial strategies that manipulate host adaptive immune responses. Infected dendritic cells (DCs) transport M. tuberculosis to local lymph nodes but activate CD4 T cells poorly, suggesting bacterial manipulation of antigen presentation. However, M. tuberculosis antigens are also exported from infected DCs and taken up and presented by uninfected DCs, possibly overcoming this blockade of antigen presentation by infected cells. Here we show that the first stage of this antigen transfer, antigen export, benefits M. tuberculosis by diverting bacterial proteins from the antigen presentation pathway. Kinesin-2 is required for antigen export and depletion of this microtubule-based motor increases activation of antigen-specific CD4 T cells by infected cells and improves control of intracellular infection. Thus, although antigen transfer enables presentation by bystander cells, it does not compensate for reduced antigen presentation by infected cells and represents a bacterial strategy for CD4 T cell evasion.

  4. Loss of phosphoinositide 3-kinase γ decreases migration and activation of phagocytes but not T cell activation in antigen-induced arthritis

    Directory of Open Access Journals (Sweden)

    Wetzker Reinhard

    2010-04-01

    Full Text Available Abstract Background Phosphoinositide 3-kinase γ (PI3Kγ has been depicted as a major regulator of inflammatory processes, including leukocyte activation and migration towards several chemokines. This study aims to explore the role of PI3Kγ in the murine model of antigen-induced arthritis (AIA. Methods Development of AIA was investigated in wildtype and PI3Kγ-deficient mice as well as in mice treated with a specific inhibitor of PI3Kγ (AS-605240 in comparison to untreated animals. Inflammatory reactions of leukocytes, including macrophage and T cell activation, and macrophage migration, were studied in vivo and in vitro. Results Genetic deletion or pharmacological inhibition of PI3Kγ induced a marked decrease of clinical symptoms in early AIA, together with a considerably diminished macrophage migration and activation (lower production of NO, IL-1β, IL-6. Also, macrophage and neutrophil infiltration into the knee joint were impaired in vivo. However, T cell functions, measured by cytokine production (TNFα, IFNγ, IL-2, IL-4, IL-5, IL-17 in vitro and DTH reaction in vivo were not altered, and accordingly, disease developed normally at later timepoints Conclusion PI3Kγ specifically affects phagocyte function in the AIA model but has no impact on T cell activation.

  5. Quantitative RT-PCR assays for the determination of urokinase-type plasminogen activator and plasminogen activator inhibitor type 1 mRNA in primary tumor tissue of breast cancer patients: comparison to antigen quantification by ELISA.

    NARCIS (Netherlands)

    Biermann, J.C.; Holzscheiter, L.; Kotzsch, M.; Luther, T.; Kiechle-Bahat, M.; Sweep, F.C.; Span, P.N.; Schmitt, M.; Magdolen, V.

    2008-01-01

    Urokinase-type plasminogen activator (uPA) and its inhibitor plasminogen activator inhibitor type 1 (PAI-1) play a key role in tumor-associated processes such as the degradation of extracellular matrix proteins, tissue remodeling, cell adhesion, migration, and invasion. High antigen levels of uPA an

  6. Immunophenotyping of Waldenstroms macroglobulinemia cell lines reveals distinct patterns of surface antigen expression: potential biological and therapeutic implications.

    Directory of Open Access Journals (Sweden)

    Aneel Paulus

    Full Text Available Waldenströms macroglobulinemia (WM is a subtype of Non-Hodgkin's lymphoma in which the tumor cell population is markedly heterogeneous, consisting of immunoglobulin-M secreting B-lymphocytes, plasmacytoid lymphocytes and plasma cells. Due to rarity of disease and scarcity of reliable preclinical models, many facets of WM molecular and phenotypic architecture remain incompletely understood. Currently, there are 3 human WM cell lines that are routinely used in experimental studies, namely, BCWM.1, MWCL-1 and RPCI-WM1. During establishment of RPCI-WM1, we observed loss of the CD19 and CD20 antigens, which are typically present on WM cells. Intrigued by this observation and in an effort to better define the immunophenotypic makeup of this cell line, we conducted a more comprehensive analysis for the presence or absence of other cell surface antigens that are present on the RPCI-WM1 model, as well as those on the two other WM cell lines, BCWM.1 and MWCL-1. We examined expression of 65 extracellular and 4 intracellular antigens, comprising B-cell, plasma cell, T-cell, NK-cell, myeloid and hematopoietic stem cell surface markers by flow cytometry analysis. RPCI-WM1 cells demonstrated decreased expression of CD19, CD20, and CD23 with enhanced expression of CD28, CD38 and CD184, antigens that were differentially expressed on BCWM.1 and MWCL-1 cells. Due to increased expression of CD184/CXCR4 and CD38, RPCI-WM1 represents a valuable model in which to study the effects anti-CXCR4 or anti-CD38 targeted therapies that are actively being developed for treatment of hematologic cancers. Overall, differences in surface antigen expression across the 3 cell lines may reflect the tumor clone population predominant in the index patients, from whom the cell lines were developed. Our analysis defines the utility of the most commonly employed WM cell lines as based on their immunophenotype profiles, highlighting unique differences that can be further studied for

  7. Antibodies against a class II HLA-peptide complex raised by active immunization of mice with antigen mimicking peptides

    DEFF Research Database (Denmark)

    Dam-Tuxen, R; Riise, Erik Skjold

    2009-01-01

    Multiple sclerosis (MS) is an autoimmune disease linked to the human leucocyte antigen (HLA) class II genes DRB1*1501, DRB5*0101 and DQB1*0602. T cells reactive towards the DRB1*1501 in complex with various peptides derived from myelin basic protein (MBP), which is the major component of myelin...

  8. Establishment and Characterization of a Cell Based Artificial Antigen-Presenting Cell for Expansion and Activation of CD8+ T Cells Ex Vivo

    Institute of Scientific and Technical Information of China (English)

    Weijuan Gong; Mingchun Ji; Zhengfeng Cao; Liheng Wang; Yayun Qian; Maozhi Hu; Li Qian; Xingyuan Pan

    2008-01-01

    Atificial antigen-presenting cells are expected to stimulate the expansion and acquisition of optimal therapeutic features of T cells before infusion. Here CD32 that binds to a crystallizable fragment of IgG monoclonal antibody was genetically expressed on human K562 leukemia cells to provide a ligand for T-cell receptor. CD86 and 4-1BBL, which are ligands of CO. stimulating receptors of CD28 and 4-1BB. respectively, were also expressed on K562 cells. Then we accomplished the artificial antigen-presenting cells by coupling K32, CD86/4-IBBL cell with OKT3 monoclonal antibody against CD3.named K32/CD86/4-lBBL/OKT3 cells. These artificial modified cells had the abilities of inducing CD8+ T cell activation. promoting CD8+ T cell proliferation, division, and long-term growth, inhibiting CD8+ T cell apoptosis, and enhancing CD8+ T cell secretion of IFN-Y and perforin. Furthermore, antigen. secific cytotoxic T lymphocytes could be retained in the culture stimulated with K32/CD86/4-1BBL/OKT3 cells at least within 28 day This approach was robust, simple, reproducible and economical for expansion and activation of CD8+ T cells and may have important therapeutic implications for adoptive immunotherapy. Cellular & Molecular Immunology.2007;5(1):47-53.

  9. [Antigenic response against PPD and antigen 60 in tubercular patients: single antigen versus the combined test].

    Science.gov (United States)

    Máttar, S; Broquetas, J M; Gea, J; Aran, X; el-Banna, N; Sauleda, J; Torres, J M

    1992-05-01

    We analyze serum samples from 70 patients with pulmonary tuberculosis and 50 healthy individuals. The antigenic activity (IgG) against protein purified antigen (PPD) and antigen 60 (A60) from M. tuberculosis. Thirteen patients were also HIV infected, and three patients had AIDS defined by the presence of disseminated tuberculosis. The test using antigen alone showed a 77% sensitivity and 74% specificity when PPD is used. When A60 was used, both values improved (81% sensitivity, 94% specificity). The use of a combined test (PPD and A60) improves the sensitivity (89%) but reduces the specificity (82%). The HIV infected patients showed similar responses to those of other patients. The combined use of different antigens might be useful for diagnosing tuberculosis. PMID:1390996

  10. Stable solid-phase Rh antigen.

    Science.gov (United States)

    Yared, M A; Moise, K J; Rodkey, L S

    1997-12-01

    Numerous investigators have attempted to isolate the Rh antigens in a stable, immunologically reactive form since the discovery of the Rh system over 56 years ago. We report here a successful and reproducible approach to solubilizing and adsorbing the human Rh antigen(s) to a solid-phase matrix in an antigenically active form. Similar results were obtained with rabbit A/D/F red blood cell antigens. The antigen preparation was made by dissolution of the red blood cell membrane lipid followed by fragmentation of the residual cytoskeleton in an EDTA solution at low ionic strength. The antigenic activity of the soluble preparations was labile in standard buffers but was stable in zwitterionic buffers for extended periods of time. Further studies showed that the antigenic activity of these preparations was enhanced, as was their affinity for plastic surfaces, in the presence of acidic zwitterionic buffers. Adherence to plastic surfaces at low pH maintained antigenic reactivity and specificity for antibody was retained. The data show that this approach yields a stable form of antigenically active human Rh D antigen that could be used in a red blood cell-free assay for quantitative analysis of Rh D antibody and for Rh D antibody immunoadsorption and purification.

  11. An Echinococcus multilocularis Antigen B3 Proteoform That Shows Specific Antibody Responses to Active-Stage Alveolar Echinococcosis.

    Science.gov (United States)

    Ahn, Chun-Seob; Cai, Huixia; Kim, Jeong-Geun; Han, Xiumin; Ma, Xiao; Bae, Young-An; Yang, Hyun-Jong; Kang, Insug; Wang, Hu; Kong, Yoon

    2015-10-01

    Alveolar echinococcosis (AE), caused by the Echinococcus multilocularis metacestode, represents one of the most frequently fatal zoonoses. Early diagnosis significantly reduces morbidity and mortality associated with AE. Diagnosis of AE largely depends on a combination of imaging and serological tests due to its minimal clinical manifestations. Several antigens derived from the whole worm and protoscolex have been targeted for AE serodiagnosis, while the antigenic properties of E. multilocularis hydatid fluid (EmHF) are unclear. We observed two AE-specific 6- and 8-kDa antigen proteoforms through an immunoproteome array of the EmHF. We identified these proteins as representing an E. multilocularis antigen B3 (EmAgB3) isoform, and the proteins were shown to be encoded by the same gene. We cloned the gene and expressed the recombinant EmAgB3 protein (rEmAgB3) in Escherichia coli. rEmAgB3 exhibited sensitivity of 90.9% (80/88 cases) and specificity of 98.5% (597/606 samples) by immunoblotting. The positive and negative predictive values were 89.9% and 98.6%, respectively. The protein did not show antibody responses to 33 AE sera collected during posttreatment follow-up monitoring. Mouse sera experimentally infected with AE protoscoleces began to demonstrate specific antibody responses to native and recombinant EmAgB3 6 months after infection. At that stage, fully mature metacestode vesicles that harbored the brood capsule, primary cell, and protoscolex were observed within an AE mass(es). The response declined along with worm degeneration. Our results demonstrate that the immune responses to this EmAgB3 isoform were highly correlated with worm viability accompanied with AE progression. rEmAgB3 is a promising biomarker for serological assessment of AE patients. PMID:26269620

  12. A new scorpion venom toxin paralytic to insects that affects Na+ channel activation. Purification, structure, antigenicity and mode of action.

    OpenAIRE

    Borchani, L.; Mansuelle, P.; Stankiewicz, M; Grolleau, F.; Cestèle, S.; Karoui, H.; Lapied, B; Rochat, H; Pelhate, M.; El Ayeb, M.

    1996-01-01

    International audience A new toxin, BotIT2, with a unique mode of action on the isolated giant axon of the cockroach Periplaneta americana and DUM (dorsal unpaired median) neurons, has been purified from the venom of the scorpion Buthus occitanus tunetanus. Its structural, antigenic and pharmacological properties are compared to those of three other groups of neurotoxins found in Buthidae scorpion venoms. Like excitatory, depressant and alpha-type insect-selective neurotoxins, BotIT2 is to...

  13. Transfection of B7-1 cDNA empowers antigen presentation of blood malignant cells for activation of anti-tumor T cells

    Institute of Scientific and Technical Information of China (English)

    克晓燕; 贾丽萍; 王晶; 王德炳

    2003-01-01

    Objective To define roles of B7-1 co-stimulation factor expressed in human malignant cell lines in mediating anti-tumor T cell immune responses. Methods Examining human leucocyte antigen (HLA) and B7 expressions on 8 human blood malignancies cell lines by flow cytometry. Transfecting B7-1 gene to B7-1 negative (B7*!-) Raji and B7*!- Jurkat cell lines by liposome, and comparing the potencies of blood malignant cell lines in the induction of T cell activation by examination of T cell cytokine mRNAs before and after transfection using semi-quantitative reverse transcription polymerase chain reaction (RT-PCR). Results High level of HLA Ⅰ and Ⅱ molecules were expressed in most human blood malignant cell lines examined, and the co-stimulatory factor B7-2 was also highly expressed. In contrast, another member of B7 family: B7-1 was either not expressed or very limitedly expressed in most of these hematopoietic malignant cell lines. Most importantly, transfection of B7-1 gene to B7*!-. Raji and B7*!-. Jurkat cell lines made these cell lines better antigen presenting cells for stimulation of anti-tumor T cell activation, which was demonstrated by up regulation of expression of T cell cytokines IL-2, IL-4 and INF-γ mRNAs after incubation of these tumor cells with T cells for 24 h. Conclusions B7 co-stimulation plays an important role in anti-tumor immunity. Transfection of B7-1 gene to the human hematopoietic malignant cell lines that are deficient in the B7-1 expression empowers their antigen presentation potency for activation of anti-tumor T cells. Our results suggested that repairing the deficiency of B7-1 co-stimulatory pathway in tumor cells might be a novel immunotherapeutic approach for human hematopoietic malignancies.

  14. Interleukin-2 (rIL-2)-induced lymphokine-activated killer (LAK) cells and their precursors express the VGO1 antigen

    International Nuclear Information System (INIS)

    Precursor and effector cells of recombinant interleukin-2 (r-IL-2)-induced lymphokine-activated killer (LAK) activity were investigated for their expression of VGO1. Peripheral blood lymphocytes (PBL) from normal donors were purified and separated in a FACS 420 into VGO1+- and VGO1- cell fractions before and after culture for 96 hr with 100 U/ml of r-IL-2. Their lytic activity against K 562 and Daudi cells was measured in a 51Cr release assay. The majority, if not all, of the LAK effector and precursor cells was VGO1+ lymphocytes. The expression of VGO1 by LAK precursor cells remained stable under the culture conditions used in our experiments. VGO1- lymphocytes cultured with r-IL-2 demonstrated neither LAK-induced activity nor expression of VGO1 antigen

  15. Resveratrol Suppresses Cytokine Production Linked to FcεRI-MAPK Activation in IgE-Antigen Complex-Exposed Basophilic Mast Cells and Mice.

    Science.gov (United States)

    Han, Seon-Young; Choi, Yean-Jung; Kang, Min-Kyung; Park, Jung Han Yoon; Kang, Young-Hee

    2015-01-01

    A complicated interplay between resident mast cells and other recruited inflammatory cells contributes to the development and progression of allergic inflammation entailing the promotion of T helper 2 (Th2) cytokine responses. The current study examined whether resveratrol suppressed the production of inflammatory Th2 cytokines in cultured rat basophilic leukemia RBL-2H3 cells. Cells pre-treated with resveratrol nontoxic at 1–25 μM were sensitized with anti-dinitrophenyl (anti-DNP), and subsequently stimulated by dinitrophenyl-human serum albumin (DNP–HSA) antigen. Resveratrol dose-dependently diminished the secretion of interleukin (IL)-3, IL-4, IL-13 as well as tumor necrosis factor (TNF)-α by the antigen stimulation from sensitized cells. It was found that resveratrol mitigated the phosphorylation of p38 MAPK, ERK, and JNK elevated in mast cells exposed to Fc epsilon receptor I (FcεRI)-mediated immunoglobulin E (IgE)-antigen complex. The FcεRI aggregation was highly enhanced on the surface of mast cells following the HSA stimulation, which was retarded by treatment with 1–25 μM resveratrol. The IgE-receptor engagement rapidly induced tyrosine phosphorylation of c-Src-related focal adhesion protein paxillin involved in the cytoskeleton rearrangement. The FcεRI-mediated rapid activation of c-Src and paxillin was attenuated in a dose-dependent manner. In addition, the paxillin activation entailed p38 MAPK and ERK-responsive signaling, but the JNK activation was less involved. Consistently, oral administration of resveratrol reduced the tissue level of phosphorylated paxillin in the dorsal skin of DNP–HSA-challenged mice. The other tyrosine kinase Tyk2-STAT1 signaling was activated in the dorsal epidermis of antigen-exposed mice, which was associated with allergic inflammation. These results showed that resveratrol inhibited Th2 cytokines- and paxillin-linked allergic responses dependent upon MAPK signaling. Therefore, resveratrol may possess the

  16. OX48, a monoclonal antibody against a 70,000 MW rat activation antigen expressed by T cells bearing the high-affinity interleukin-2 receptor.

    Science.gov (United States)

    Somoza, C; Fernández-Ruiz, E; Rebollo, A; Sanz, E; Ramírez, F; Silva, A

    1990-01-01

    The monoclonal antibody (mAb) OX48 recognizes a 70,000 MW cell-surface protein present in a small percentage of activated rat T cells and in CD8+ rat x BW5147 interleukin-2 (IL-2)-dependent T-cell hybridomas, but not in resting spleen cells or in IL-2-independent T-cell hybrids. OX48 antibody added simultaneously with concanavalin A (Con A) to resting spleen cells inhibits the cell proliferation and reduces the IL-2 production. However, addition of IL-2 does not restore the mitogenic response. Growth of rat blast T cells or IL-2-dependent hybrids is not affected by the OX48 antibody. There is a close correlation between the expression of high-affinity IL-2 receptors (IL-2R) and the OX48 antigen in T-cell hybridomas. In spite of this striking correlation, OX48 mAb does not inhibit the binding of 125I-IL-2 to the IL-2-dependent hybrids, and is unable to immunoprecipitate any of the proteins chemically cross-linked to 125I-IL-2. Therefore, the OX48 molecule represents a new rat activation antigen, undefined in other species, and probably involved in the early steps of T-cell activation. Images Figure 5 Figure 7 PMID:2373518

  17. Transiently reduced PI3K/Akt activity drives the development of regulatory function in antigen-stimulated Naive T-cells.

    Directory of Open Access Journals (Sweden)

    Eloho Etemire

    Full Text Available Regulatory T-cells (Tregs are central for immune homeostasis and divided in thymus-derived natural Tregs and peripherally induced iTreg. However, while phenotype and function of iTregs are well known, a remarkable lack exists in knowledge about signaling mechanisms leading to their generation from naïve precursors in peripheral tissues. Using antigen specific naïve T-cells from mice, we investigated CD4+ CD25+ FoxP3- iTreg induction during antigen-specific T-cell receptor (TCR stimulation with weak antigen presenting cells (APC. We show that early signaling pathways such as ADAM-17-activation appeared similar in developing iTreg and effector cells (Teff and both initially shedded CD62-L. But iTreg started reexpressing CD62-L after 24 h while Teff permanently downmodulated it. Furthermore, between 24 and 72 hours iTreg presented with significantly lower phosphorylation levels of Akt-S473 suggesting lower activity of the PI3K/Akt-axis. This was associated with a higher expression of the Akt hydrophobic motif-specific phosphatase PHLPP1 in iTreg. Importantly, the lack of costimulatory signals via CD28 from weak APC was central for the development of regulatory function in iTreg but not for the reappearance of CD62-L. Thus, T-cells display a window of sensitivity after onset of TCR triggering within which the intensity of the PI3K/Akt signal controls entry into either effector or regulatory pathways.

  18. The IgG antibody reactivity of sera from patients with active chronic hepatitis to a crude liver antigen and liver specific protein (LSP): analysis by ELISA and immunoblotting.

    Science.gov (United States)

    Sundin, U; Heigl, Z; Sundqvist, K G

    1988-11-01

    The antibody reactivity to liver specific protein (LSP) and a crude liver antigen of sera from patients with chronic active hepatitis (CAH) were studied along with other related diseases and healthy individuals. CAH sera containing liver reacting antibodies were selected using an ELISA with a crude liver preparation as antigen and subsequently the specificity was analysed by immunoblotting of SDS-PAGE-separated LSP. The incidence of IgG antibodies to the crude liver antigen and LSP in sera from 15 patients with CAH was 94% and 55% respectively. In the healthy control group (n = 30) the corresponding figures were 3% and 17%. Sera from patients with other autoimmune conditions with considerable reactivity in the crude liver ELISA test were those with antibodies against extractable nuclear antigens (ENA) and thyroid gland antigens, while the anti-nuclear antibody (ANA) group as a whole did not differ from the control group. In immunoblotting of SDS-PAGE-separated crude liver and LSP antigens, the IgG binding pattern of ELISA IgG positive CAH sera and sera from patients with thyroid disease was distinct, with bands corresponding to antigens of molecular weights of 38, 45 and 50 kD which were not observed in ELISA negative CAH sera or in sera from patients with other diseases and among healthy controls.

  19. Targeted delivery of lipid antigen to macrophages via the CD169/sialoadhesin endocytic pathway induces robust invariant natural killer T cell activation

    OpenAIRE

    Kawasaki, Norihito; Vela, Jose Luis; Nycholat, Corwin M.; Rademacher, Christoph; Khurana, Archana; van Rooijen, Nico; Crocker, Paul R.; Kronenberg, Mitchell; Paulson, James C.

    2013-01-01

    Invariant natural killer T (iNKT) cells induce a protective immune response triggered by foreign glycolipid antigens bound to CD1d on antigen-presenting cells (APCs). A limitation of using glycolipid antigens to stimulate immune responses in human patients has been the inability to target them to the most effective APCs. Recent studies have implicated phagocytic CD169+ macrophages as major APCs in lymph nodes for priming iNKT cells in mice immunized with glycolipid antigen in particulate form...

  20. Specificity of mimotope-induced anti-high molecular weight-melanoma associated antigen (HMW-MAA antibodies does not ensure biological activity.

    Directory of Open Access Journals (Sweden)

    Julia Latzka

    Full Text Available Vaccines based on peptide mimics (mimotopes of conformational tumor antigen epitopes have been investigated for a variety of human tumors including breast cancer, tumors expressing the carcinoembryonic antigen, B cell lymphoma, neuroblastoma, and melanoma. In our previous work, we designed a vaccine based on a mimotope of the high molecular weight-melanoma associated antigen (HMW-MAA that elicited HMW-MAA-specific antibodies (Abs with anti-tumor activity in vitro and in vivo. In this study, we aimed to identify mimotopes of additional distinct HMW-MAA epitopes, since they could be used to construct a polymimotope melanoma vaccine. For this purpose, random peptide phage libraries were screened with the anti-HMW-MAA monoclonal antibodies (mAbs VT80.12 and VF1-TP43 yielding one peptide ligand for each mAb. Both peptides inhibited the binding of the corresponding mAb to the HMW-MAA. Furthermore, when coupled to the carrier protein keyhole limpet hemocyanin (KLH, both HMW-MAA mimotopes elicited peptide-specific Abs in rabbits or BALB/c mice, but only the mimotope isolated with the mAb VT80.12 elicited HMW-MAA-specific Abs and only in mice. However, the latter Abs had no detectable effect on HMW-MAA expressing human melanoma cells in vitro. These results describe limitations related to the phage display technique and emphasize the need to characterize the functional properties of the mAb utilized to isolate mimotopes of the corresponding epitopes.

  1. Tumor Antigen Specific Activation of Primary Human T-Cells Expressing a Virally Encoded Chimeric T-Cell Receptor Specific for p185HER2

    Institute of Scientific and Technical Information of China (English)

    杨建民; MichaelSFRIEDMAN; ChristopherMREYNOLDS; MarianneTHUBEN; LeeWILKE; JenniferFULLER; 李桥; ZeligESHHAR; JamesJMULE; KevimTMCDONAGH

    2004-01-01

    We have developed and tested chimeric T-cell receptors (TCR) specific for p185HER2. In these experiments,retroviral vectors expressing the N297 or N29ξ receptors were constructed in pRET6. Amphotropic viral producer cells were established in the GALV-based PG13 packaging cell line. Ficoll purified human peripheral blood lymphocytes (PBL) were vitally transduced using an optimized protocol incorporating activation with immobilized anti-CD3/anti-CD28 monoclonal antibodies, followed by viral infection in the presence of fibronectin fragment CH296. Transduced cells were co-cultured with human tumor cell lines that overexpress (SK-OV-3) or underexpress (MCF7) p185HER2 to assay for antigen specific immune responses. Both CD4+ and CD8+ T-cells transduced with the N297 or N29ξ chTCR demonstrated HER2-specific antigen responses, as determined by release of Th1 like cytokines, and cellular cytotoxicity assays. Our results support the feasibility of adoptive immunothempy with genetically modified T-cells expressing a chTCR specific for p185HER2.

  2. T-antigen binding lectin with antibacterial activity from marine invertebrate, sea cucumber (Holothuria scabra): possible involvement in differential recognition of bacteria.

    Science.gov (United States)

    Gowda, Nagaraj M; Goswami, Usha; Khan, M Islam

    2008-10-01

    In invertebrates, cellular and humoral components are evolved to maintain their body immunity and integrity. Both these factors respond to different antigens such as microorganisms, vertebrate erythrocytes and foreign proteins. In this article, we report a study of a lectin (HSL) involved in immune response in the echinoderm, sea cucumber (Holothuria scabra). Correlative studies indicate that the expression of this defensive lectin is induced by bacterial challenge, wherein cell wall glycoconjugates of bacteria are involved in lectin induction. HSL showed strong broad spectrum antibacterial activity against both gram-positive and gram-negative bacteria. Under in vitro conditions, purified HSL mediate agglutination of the test bacteria, there by indicating a possible mode of action in physiological situation. PMID:18501924

  3. Chemopreventive effect of resveratrol, sesamol, sesame oil and sunflower oil in the Epstein-Barr virus early antigen activation assay and the mouse skin two-stage carcinogenesis.

    Science.gov (United States)

    Kapadia, Govind J; Azuine, Magnus A; Tokuda, Harukuni; Takasaki, Midori; Mukainaka, Teruo; Konoshima, Takao; Nishino, Hoyoku

    2002-06-01

    Resveratrol, sesamol, sesame oil and sunflower oil are known natural dietary components with intrinsic cancer chemopreventive potentials. As a part of our study of dietary constituents as potential cancer chemopreventive agents, we have assessed the anti-cancer potentials of these products in the promotion stage of cancer development employing the in vitro Epstein-Barr virus early antigen activation assay induced by the tumor promoter 12-O-tetradecanoylphorbol 13-acetate (TPA). Further, we studied the activities of these compounds in the brine shrimp cytotoxicity assay as well as on the stable 1,1-diphenyl-2-picrylhydrazyl (DPPH) free radical scavenging bioassay with a view to comparing some of the mechanisms of their anti-cancer activity. Finally, we compared the observed chemoprotective capabilities of the four products in the in vivo 7,12 dimethylbenz(a)anthracene initiated and TPA-promoted mouse skin two-stage carcinogenesis protocols. All the products tested showed a profound inhibitory effect on the Epstein-Barr virus early antigen induction using Raji cells. Comparatively, sesame oil was the most potent followed by sesamol and then resveratrol. Only sesamol and resveratrol showed a remarkable cytotoxic activity in the brine shrimp lethality assays as well as profound free radical scavenging activity in the DPPH bioassay. In both test systems, sesamol exhibited a more remarkable activity than resveratrol while sesame oil and sunflower oil did not exhibit any appreciable activity even at the highest concentrations tested (4000 microg ml(-1) ). In our in vivo assay at a 50-fold molar ratio to TPA, sesamol offered 50% reduction in mouse skin papillomas at 20 weeks after promotion with TPA. Under an identical molar ratio to TPA, resveratrol offered a 60% reduction in the papillomas in mouse at 20 weeks. Thus sesamol seems to be an almost equally potent chemopreventive agent. Sesame oil and sunflower oil offered 20 and 40% protection, respectively, in the mouse

  4. Surface antigen expression in chronic lymphocytic leukemia: clustering analysis, interrelationships and effects of chromosomal abnormalities.

    Science.gov (United States)

    Hulkkonen, J; Vilpo, L; Hurme, M; Vilpo, J

    2002-02-01

    Chronic lymphocytic leukemia (CLL) is a phenotypically distinguishable form of B-lymphoid leukemias. The regularity of surface membrane antigen expression patterns, their interrelationships as well as the effects of the three frequent chromosomal aberrations, ie 11q deletion, 13q deletion and trisomy 12, were investigated in 35 classic CLL cases by flow cytometry. The two-way cluster analysis of 31 individual antigens revealed three expression patterns: (1) most cells in most cases positive (CD5, CD19, CD20, CD23, CD27, CD40, CD45, CD45RA); (2) most cells in most cases negative (CD10, CD14, CD34, CD122, CD154, mIgG); and (3) a mixed pattern with a variable number of positive cases and a variable percentage of positive cells in individual cases (CD11c, CD21, CD22, CD25, CD38, CD45RO, CD79b, CD80, CD95, CD124, CD126, CD130, FMC7, mIgD, mIgkappa, mIglambda, mIgM). The expressions of several antigens were strongly interdependent, even when antigens belonged to entirely different gene families. Such antigen pairs were: CD11c/CD21; CD19/CD45; CD19/CD79b; CD22/CD45RA; CD23/Igkappa; CD25/mIgM; CD27/CD45; CD45/CD79b; CD45RA/Igkappa. In contrast, the expression of some antigens was mutually exclusive, the best examples being CD45RA/CD45RO, CD38/CD80 and CD45RA/CD80. Deletion of chromosome arm 11q attenuated expression of splicing variant CD45RA, but enhanced CD45RO expression. In contrast, cases of trisomy 12 were associated with enhanced CD45RA and attenuated CD45RO expression. Similarly, trisomy 12 was associated with enhanced CD27 and mIgkappa expression. The variable levels of signaling surface membrane antigens, their interactions and interference by genetic aberrations are likely to affect the clinical progression and drug response of CLL. PMID:11840283

  5. The 15 SCR flexible extracellular domains of human complement receptor type 2 can mediate multiple ligand and antigen interactions.

    Science.gov (United States)

    Gilbert, Hannah E; Asokan, Rengasamy; Holers, V Michael; Perkins, Stephen J

    2006-10-01

    Complement receptor type 2 (CR2, CD21) is a cell surface protein that links the innate and adaptive immune response during the activation of B cells. The extracellular portion of CR2 comprises 15 or 16 short complement regulator (SCR) domains, for which the overall arrangement in solution is unknown. This was determined by constrained scattering and ultracentrifugation modelling. The radius of gyration of CR2 SCR 1-15 was determined to be 11.5 nm by both X-ray and neutron scattering, and that of its cross-section was 1.8 nm. The distance distribution function P(r) showed that the overall length of CR2 SCR 1-15 was 38 nm. Sedimentation equilibrium curve fits gave a mean molecular weight of 135,000 (+/- 13,000) Da, in agreement with a fully glycosylated structure. Velocity experiments using the g*(s) derivative method gave a sedimentation coefficient of 4.2 (+/- 0.1) S. In order to construct a model of CR2 SCR 1-15 for constrained fitting, homology models for the 15 SCR domains were combined with randomised linker peptides generated by molecular dynamics simulations. Using an automated procedure, the analysis of 15,000 possible CR2 SCR 1-15 models showed that only those models in which the 15 SCR domains were flexible but partially folded back accounted for the scattering and sedimentation data. The best-fit CR2 models provided a visual explanation for the versatile interaction of CR2 with four ligands C3d, CD23, gp350 and IFN-alpha. The flexible location of CR2 SCR 1-2 is likely to facilitate interactions of C3d-antigen complexes with the B cell receptor.

  6. Histocompatibility antigen test

    Science.gov (United States)

    ... more common in certain autoimmune diseases . For example, HLA-B27 antigen is found in many people (but not ... More Ankylosing spondylitis Autoimmune disorders Bone marrow transplant HLA-B27 antigen Kidney transplant Reactive arthritis Update Date 2/ ...

  7. Prospective evaluation of a whole-blood test using Mycobacterium tuberculosis-specific antigens ESAT-6 and CFP-10 for diagnosis of active tuberculosis

    DEFF Research Database (Denmark)

    Ravn, Pernille; Munk, Martin E; Andersen, Ase B;

    2005-01-01

    in a specificity of 60%. However, 80% (8/10) of these had risk-factors for TB, indicating latent infection in this group. In healthy controls, only 3% (1/39) were QFT-RD1 positive. In conclusion, the QFT-RD1 test is sensitive for diagnosis of TB, especially in patients with negative microscopy and culture......A new immunodiagnostic test based on the Mycobacterium tuberculosis-specific antigens CFP-10/ESAT-6(QFT-RD1) has been launched as an aid in the diagnosis of latent tuberculosis (TB) infection (LTBI). The aim of this study was to evaluate this test for the diagnosis of active TB. Eighty-two patients...... with suspicion of TB and 39 healthy BCG-vaccinated persons were enrolled. Forty-eight had active TB, 25 did not, and 9 were excluded. Sensitivity and specificity of the test for active TB were evaluated in a prospective blinded manner in patients suspected of TB. The sensitivity of the QFT-RD1 was 85% (40...

  8. Prospective evaluation of a whole-blood test using Mycobacterium tuberculosis-specific antigens ESAT-6 and CFP-10 for diagnosis of active tuberculosis

    DEFF Research Database (Denmark)

    Ravn, Pernille; Munk, Martin E; Andersen, Ase B;

    2005-01-01

    A new immunodiagnostic test based on the Mycobacterium tuberculosis-specific antigens CFP-10/ESAT-6(QFT-RD1) has been launched as an aid in the diagnosis of latent tuberculosis (TB) infection (LTBI). The aim of this study was to evaluate this test for the diagnosis of active TB. Eighty-two patients...... with suspicion of TB and 39 healthy BCG-vaccinated persons were enrolled. Forty-eight had active TB, 25 did not, and 9 were excluded. Sensitivity and specificity of the test for active TB were evaluated in a prospective blinded manner in patients suspected of TB. The sensitivity of the QFT-RD1 was 85......% (40/48; confidence interval [CI], 75 to 96), and it was higher than the sensitivity of microscopy, 42% (20/48; CI, 27 to 56; P = 0.001), and culture, 59% (27/46; CI, 44 to 73; P = 0.009). Of patients with extrapulmonary TB, 92% (12/13) were QFT-RD1 positive, whereas only 31% (4/13) were positive by...

  9. Clonality Analysis of Immunoglobulin Gene Rearrangement by Next-Generation Sequencing in Endemic Burkitt Lymphoma Suggests Antigen Drive Activation of BCR as Opposed to Sporadic Burkitt Lymphoma

    Science.gov (United States)

    Amato, Teresa; Abate, Francesco; Piccaluga, Pierpaolo; Iacono, Michele; Fallerini, Chiara; Renieri, Alessandra; De Falco, Giulia; Ambrosio, Maria Raffaella; Mourmouras, Vaselious; Ogwang, Martin; Calbi, Valeria; Rabadan, Roul; Hummel, Michael; Pileri, Stefano; Bellan, Cristiana

    2016-01-01

    Objectives: Recent studies using next-generation sequencing (NGS) analysis disclosed the importance of the intrinsic activation of the B-cell receptor (BCR) pathway in the pathogenesis of sporadic Burkitt lymphoma (sBL) due to mutations of TCF3/ID3 genes. Since no definitive data are available on the genetic landscape of endemic Burkitt (eBL), we first assessed the mutation frequency of TCF3/ID3 in eBL compared with sBL and subsequently the somatic hypermutation status of the BCR to answer whether an extrinsic activation of BCR signaling could also be demonstrated in Burkitt lymphoma. Methods: We assessed the mutations of TCF3/ID3 by RNAseq and the BCR status by NGS analysis of the immunoglobulin genes (IGs). Results: We detected mutations of TCF3/ID3 in about 30% of the eBL cases. This rate is significantly lower than that detected in sBL (64%). The NGS analysis of IGs revealed intraclonal diversity, suggesting an active targeted somatic hypermutation process in eBL compared with sBL. Conclusions: These findings support the view that the antigenic pressure plays a key role in the pathogenetic pathways of eBL, which may be partially distinct from those driving sBL development. PMID:26712879

  10. Comparison Study of Cyfra 21-1, Carcinoembryonic Antigen and Telomerase Activity between Non Small Cell and Small Cell Lung Cancer Patients

    Directory of Open Access Journals (Sweden)

    B. Alani

    2008-10-01

    Full Text Available Introduction & Objective: The roles of tumor markers in the prognosis and diagnosis of lung cancer is under investigation. The aim of this study was to examine correlation between serum levels of Carcinoembryonic Antigen (CEA and Cyfra 21-1 with telomerase activity on biopsy samples in patients with lung cancer and controls.Materials & Methods: We studied 50 malignant lung cancer patients (mean age 68.3±13.8 years and 20 normal individuals (mean age 64.9±11.4 years. Serum levels of Cyfra21-1 and CEA were measured with available enzyme immunoassay kits. Telomerase activity was measured by TRAP assay based on PCR-ELISA in lung tumor biopsy. To compare quantitative means of the two groups, t-independent and Man-Whitney analysis were applied.Results: The mean serum levels of CEA and Cyfra21-1 concentration together telomerase activity of biopsy samples in lung cancer patients were significantly higher than controls. There were a strong correlation between Cyfra21-1 and CEA, Telomerase and CEA and Telomerase and Cyfra21-1 in patient group. In patients with small cell carcinoma, the mean serum levels of Cyfra21-1 and CEA were significantly higher than non small cell carcinoma patients. Telomerase activities in biopsy samples of small cell carcinoma were significantly lower than non small cell carcinoma patients.Conclusion: It is speculated that based on this findings, telomerase activity in biopsy samples for small cell carcinoma patients and serum levels of Cyfra 21-1 and CEA are the most useful tumor markers for diagnosis of squamous and adenocarcinoma of non small cell cancers respectively.

  11. Coordinated activation of candidate proto-oncogenes and cancer testes antigens via promoter demethylation in head and neck cancer and lung cancer.

    Directory of Open Access Journals (Sweden)

    Ian M Smith

    Full Text Available BACKGROUND: Epigenetic alterations have been implicated in the pathogenesis of solid tumors, however, proto-oncogenes activated by promoter demethylation have been sporadically reported. We used an integrative method to analyze expression in primary head and neck squamous cell carcinoma (HNSCC and pharmacologically demethylated cell lines to identify aberrantly demethylated and expressed candidate proto-oncogenes and cancer testes antigens in HNSCC. METHODOLOGY/PRINCIPAL FINDINGS: We noted coordinated promoter demethylation and simultaneous transcriptional upregulation of proto-oncogene candidates with promoter homology, and phylogenetic footprinting of these promoters demonstrated potential recognition sites for the transcription factor BORIS. Aberrant BORIS expression correlated with upregulation of candidate proto-oncogenes in multiple human malignancies including primary non-small cell lung cancers and HNSCC, induced coordinated proto-oncogene specific promoter demethylation and expression in non-tumorigenic cells, and transformed NIH3T3 cells. CONCLUSIONS/SIGNIFICANCE: Coordinated, epigenetic unmasking of multiple genes with growth promoting activity occurs in aerodigestive cancers, and BORIS is implicated in the coordinated promoter demethylation and reactivation of epigenetically silenced genes in human cancers.

  12. Activation of CD8-dependent cytotoxic T lymphocyte adhesion and degranulation by peptide class I antigen complexes.

    Science.gov (United States)

    Kane, K P; Mescher, M F

    1993-06-01

    Activation of CTL requires engagement of both the TCR and the CD8 coreceptor. Immobilized class I proteins and in vitro-formed peptide class I Ag complexes have been used to examine the relative contributions of TCR and CD8 to the adhesion and response of cloned, class I-restricted CTL. The extent of degranulation was found to be directly proportional to the concentration of peptide used to pulse class I, suggesting that activation is a direct function of TCR occupancy level. In contrast, activation of degranulation as a function of the amount of class I on the surface displayed a marked threshold density dependence. Essentially the same density dependence was found for the response of CTL to fluid phase anti-TCR mAb and non-Ag class I, indicating that CD8-class I interaction must exceed a threshold before effective cosignaling can occur. Adhesion and degranulation of CTL was minimal in response to in vitro peptide-class I complexes prepared at a class I density below the threshold. However, the same density of peptide class I initiated both adhesion and response if additional non-Ag class I was coimmobilized on the same surface at levels above threshold. Thus, when surface levels of peptide class I complex are low, as is likely to be the case under physiologic conditions, the level of TCR occupancy achieved is, by itself, insufficient to mediate cell adhesion or activate degranulation. The results demonstrate, however, that low TCR occupancy is sufficient to provide the signal to prime CD8. Provided that the surface density of class I is sufficiently high, CD8 then mediates strong adhesion and provides the costimulatory signal(s) to activate response.

  13. Allo-antigen stimulated CD8+ T-cells suppress NF-κB and Ets-1 DNA binding activity, and inhibit phosphorylated NF-κB p65 nuclear localization in CD4+ T-cells.

    Science.gov (United States)

    Nagashima, Ryuichi; Kawakami, Fumitaka; Takahashi, Shinichiro; Obata, Fumiya; Kubo, Makoto

    2014-08-01

    CD8+ T-cells of asymptomatic HIV-1 carriers (AC) suppress human immunodeficiency virus type 1 (HIV-1) replication in a class I major histocompatibility complex (MHC-I)-restricted and -unrestricted manner. In order to investigate the mechanism of MHC-I-unrestricted CD8+ T-cell-mediated HIV-1 suppression, we previously established allo-antigen stimulated CD8+T-cells from HIV-1-uninfected donors. These allo-antigen stimulated CD8+ T-cells suppressed HIV-1 replication in acutely infected autologous CD4+ T-cells when directly co-cultured. To elucidate the mechanism of HIV-1 replication suppression, we analyzed DNA-binding activity and phosphorylation of transcriptional factors associated with HIV-1 replication by electrophoresis mobility shift assay and Western blotting. When CD4+ T-cells were cultured with allo-antigen stimulated CD8+ T-cells, the reduction of NF-κB and Ets-1 DNA-binding activity was observed. Nuclear localization of NF-κB p65 and Ets-1 was suppressed in CD4+ T-cells. Although NF-κB p65 and Ets-1 are known to be regulated by protein kinase A (PKA), no difference was observed in the expression and phosphorylation of the PKA catalytic subunit in CD4+ T-cells cultured with PHA-treated CD8+ T-cells or allo-antigen stimulated CD8+ T-cells. Cyclic AMP is also known to enter through gap junctions, but the suppression of HIV-1 replication mediated by allo-antigen stimulated CD8+ T-cells was not affected by the gap junction inhibitor. The nuclear transport of phosphorylated NF-κB p65 (Ser276) was inhibited only in CD4+ T-cells cultured with allo-antigen stimulated CD8+ T-cells. Our results indicate that allo-antigen stimulated CD8+ T-cells suppress the transcriptional activity of NF-κB p65 or Ets-1 in an antigen-nonspecific manner, and inhibit the nuclear transport of phosphorylated NF-κB p65 (Ser276).

  14. Fibrosarcoma versus fibromatoses and cellular nodular fasciitis. A comparative study of their proliferative activity using proliferating cell nuclear antigen, DNA flow cytometry, and p53.

    Science.gov (United States)

    Oshiro, Y; Fukuda, T; Tsuneyoshi, M

    1994-07-01

    We analyzed the proliferative activities, immunoreactivity of the p53 protein, and aneuploidy in patients with benign and malignant fibrous lesions, including 19 with nodular fasciitis (cellular type) (6-88 years old, mean 42.9), 11 with abdominal fibromatoses (22-74 years old, mean 37.9), 13 with extraabdominal fibromatoses (2-38 years old, mean 19.5), and 23 with fibrosarcomas (adult type: 16-71 years old, mean 47.3; infantile type: 3 months to 9 years, mean 2.9) using immunohistochemistry to determine proliferating cell nuclear antigen (PC10) and p53 protein (CM1) as well as performing DNA flow cytometry. The proliferating cell nuclear antigen (PCNA) score was measured as the ratio of PCNA-positive nuclear size/total nuclear size determined by an image analysis computer system. The distribution pattern of the PCNA-positive cells was uneven in each instance of nodular fasciitis, in contrast to the distribution in abdominal fibromatosis, extraabdominal fibromatosis, and fibrosarcoma. Both fibrosarcoma (28.4 +/- 20.0) and nodular fasciitis (33.6 +/- 20.9) exhibited a larger value and a greater variation in the PCNA score than did either abdominal (13.5 +/- 14.5) or extraabdominal fibromatosis (19.9 +/- 21.5). Abdominal fibromatosis exhibited a smaller value and less variation in the score. In short, the PCNA score did not correlate with the malignant potential. The proliferative index (S + G2 + M fraction) in fibrosarcoma was significantly higher than in either nodular fasciitis or abdominal fibromatosis. Aneuploidy was detected in five cases (26%) of fibrosarcoma, while six (26%) fibrosarcomas showed p53 positivity. Furthermore, p53-positive patients had a worse survival (0.01 < p < 0.05), and p53 positivity correlated with the proliferative index (p < 0.01). In conclusion, the PCNA score simply indicates the proliferative activity independent of malignant potential. On the other hand, p53 positivity, proliferative index, and aneuploidy are all indicators of

  15. JC virus small T antigen binds phosphatase PP2A and Rb family proteins and is required for efficient viral DNA replication activity.

    Directory of Open Access Journals (Sweden)

    Brigitte Bollag

    Full Text Available BACKGROUND: The human polyomavirus, JC virus (JCV produces five tumor proteins encoded by transcripts alternatively spliced from one precursor messenger RNA. Significant attention has been given to replication and transforming activities of JCV's large tumor antigen (TAg and three T' proteins, but little is known about small tumor antigen (tAg functions. Amino-terminal sequences of tAg overlap with those of the other tumor proteins, but the carboxy half of tAg is unique. These latter sequences are the least conserved among the early coding regions of primate polyomaviruses. METHODOLOGY AND FINDINGS: We investigated the ability of wild type and mutant forms of JCV tAg to interact with cellular proteins involved in regulating cell proliferation and survival. The JCV P99A tAg is mutated at a conserved proline, which in the SV40 tAg is required for efficient interaction with protein phosphatase 2A (PP2A, and the C157A mutant tAg is altered at one of two newly recognized LxCxE motifs. Relative to wild type and C157A tAgs, P99A tAg interacts inefficiently with PP2A in vivo. Unlike SV40 tAg, JCV tAg binds to the Rb family of tumor suppressor proteins. Viral DNAs expressing mutant t proteins replicated less efficiently than did the intact JCV genome. A JCV construct incapable of expressing tAg was replication-incompetent, a defect not complemented in trans using a tAg-expressing vector. CONCLUSIONS: JCV tAg possesses unique properties among the polyomavirus small t proteins. It contributes significantly to viral DNA replication in vivo; a tAg null mutant failed to display detectable DNA replication activity, and a tAg substitution mutant, reduced in PP2A binding, was replication-defective. Our observation that JCV tAg binds Rb proteins, indicates all five JCV tumor proteins have the potential to influence cell cycle progression in infected and transformed cells. It remains unclear how these proteins coordinate their unique and overlapping functions.

  16. Association of Single Nucleotide Polymorphisms in the ST3GAL4 Gene with VWF Antigen and Factor VIII Activity.

    Science.gov (United States)

    Song, Jaewoo; Xue, Cheng; Preisser, John S; Cramer, Drake W; Houck, Katie L; Liu, Guo; Folsom, Aaron R; Couper, David; Yu, Fuli; Dong, Jing-Fei

    2016-01-01

    VWF is extensively glycosylated with biantennary core fucosylated glycans. Most N-linked and O-linked glycans on VWF are sialylated. FVIII is also glycosylated, with a glycan structure similar to that of VWF. ST3GAL sialyltransferases catalyze the transfer of sialic acids in the α2,3 linkage to termini of N- and O-glycans. This sialic acid modification is critical for VWF synthesis and activity. We analyzed genetic and phenotypic data from the Atherosclerosis Risk in Communities (ARIC) study for the association of single nucleotide polymorphisms (SNPs) in the ST3GAL4 gene with plasma VWF levels and FVIII activity in 12,117 subjects. We also analyzed ST3GAL4 SNPs found in 2,535 subjects of 26 ethnicities from the 1000 Genomes (1000G) project for ethnic diversity, SNP imputation, and ST3GAL4 haplotypes. We identified 14 and 1,714 ST3GAL4 variants in the ARIC GWAS and 1000G databases respectively, with 46% being ethnically diverse in their allele frequencies. Among the 14 ST3GAL4 SNPs found in ARIC GWAS, the intronic rs2186717, rs7928391, and rs11220465 were associated with VWF levels and with FVIII activity after adjustment for age, BMI, hypertension, diabetes, ever-smoking status, and ABO. This study illustrates the power of next-generation sequencing in the discovery of new genetic variants and a significant ethnic diversity in the ST3GAL4 gene. We discuss potential mechanisms through which these intronic SNPs regulate ST3GAL4 biosynthesis and the activity that affects VWF and FVIII. PMID:27584569

  17. Treponema pallidum (syphilis) antigen TpF1 induces angiogenesis through the activation of the IL-8 pathway

    OpenAIRE

    Tommaso Pozzobon; Nicola Facchinello; Fleur Bossi; Nagaja Capitani; Marisa Benagiano; Giulietta Di Benedetto; Cristina Zennaro; Nicole West; Gaia Codolo; Marialina Bernardini; Cosima Tatiana Baldari; Mario Milco D’Elios; Luca Pellegrini; Francesco Argenton; Marina de Bernard

    2016-01-01

    Over 10 million people every year become infected by Treponema pallidum and develop syphilis, a disease with broad symptomatology that, due to the difficulty to eradicate the pathogen from the highly vascularized secondary sites of infection, is still treated with injections of penicillin. Unlike most other bacterial pathogens, T. pallidum infection produces indeed a strong angiogenic response whose mechanism of activation, however, remains unknown. Here, we report that one of the major antig...

  18. Novel CD47: SIRPα dependent mechanism for the activation of STAT3 in antigen-presenting cell.

    Directory of Open Access Journals (Sweden)

    Natan Toledano

    Full Text Available Cell surface CD47 interacts with its receptor, signal-regulatory-protein α (SIRPα that is expressed predominantly on macrophages, to inhibit phagocytosis of normal, healthy cells. This "don't eat me" signal is mediated through tyrosine phosphorylation of SIRPα at the cytoplasmic ITIM motifs and the recruitment of the phosphatase, SHP-1. We previously revealed a novel mechanism for the activation of the STAT3 pathway and the regulation of human APC maturation and function that is based on cell:cell interaction. In this study, we present evidence supporting the notion that CD47:SIRPα serves as a cell surface receptor: ligand pair involved in this contact-dependent STAT3 activation and regulation of APC maturation. We show that upon co-culturing APC with various primary and tumor cell lines STAT3 phosphorylation and IL-10 expression are induced, and such regulation could be suppressed by specific CD47 siRNAs and shRNAs. Significantly, >50% reduction in CD47 expression abolished the contact-dependent inhibition of T cell activation. Furthermore, co-immunoprecipitation experiments revealed a physical association between SIRPα and STAT3. Thus, we suggest that in addition to signaling through the ITIM-SHP-1 complex that transmit an anti-phagocytotic, CD47:SIRPα also triggers STAT3 signaling that is linked to an immature APC phenotype and peripheral tolerance under steady state and pathological conditions.

  19. A Neoglycoconjugate Containing the Human Milk Sugar LNFPIII Drives Anti-Inflammatory Activation of Antigen Presenting Cells in a CD14 Dependent Pathway.

    Science.gov (United States)

    Tundup, Smanla; Srivastava, Leena; Norberg, Thomas; Watford, Wendy; Harn, Donald

    2015-01-01

    The milk pentasaccharide LNFPIII has therapeutic action for metabolic and autoimmune diseases and prolongs transplant survival in mice when presented as a neoglycoconjugate. Within LNFPIII is the Lewisx trisaccharide, expressed by many helminth parasites. In humans, LNFPIII is found in human milk and also known as stage-specific embryonic antigen-1. LNFPIII-NGC drives alternative activation of macrophages and dendritic cells via NFκB activation in a TLR4 dependent mechanism. However, the connection between LNFPIII-NGC activation of APCs, TLR4 signaling and subsequent MAP kinase signaling leading to anti-inflammatory activation of APCs remains unknown. In this study we determined that the innate receptor CD14 was essential for LNFPIII-NGC induction of both ERK and NFkB activation in APCs. Induction of ERK activation by LNFPIII-NGC was completely dependent on CD14/TLR4-Ras-Raf1/TPL2-MEK axis in bone marrow derived dendritic cells (BMDCs). In addition, LNFPIII-NGC preferentially induced the production of Th2 "favoring" chemokines CCL22 and matrix metalloprotease protein-9 in a CD14 dependent manner in BMDCs. In contrast, LNFPIII-NGC induces significantly lower levels of Th1 "favoring" chemokines, MIP1α, MIP1β and MIP-2 compared to levels in LPS stimulated cells. Interestingly, NGC of the identical human milk sugar LNnT, minus the alpha 1-3 linked fucose, failed to activate APCs via TLR4/MD2/CD14 receptor complex, suggesting that the alpha 1-3 linked fucose in LNFPIII and not on LNnT, is required for this process. Using specific chemical inhibitors of the MAPK pathway, we found that LNFPIII-NGC induction of CCL22, MMP9 and IL-10 production was dependent on ERK activation. Over all, this study suggests that LNFPIII-NGC utilizes CD14/TLR4-MAPK (ERK) axis in modulating APC activation to produce anti-inflammatory chemokines and cytokines in a manner distinct from that seen for the pro-inflammatory PAMP LPS. These pathways may explain the in vivo therapeutic effect of

  20. A Neoglycoconjugate Containing the Human Milk Sugar LNFPIII Drives Anti-Inflammatory Activation of Antigen Presenting Cells in a CD14 Dependent Pathway.

    Directory of Open Access Journals (Sweden)

    Smanla Tundup

    Full Text Available The milk pentasaccharide LNFPIII has therapeutic action for metabolic and autoimmune diseases and prolongs transplant survival in mice when presented as a neoglycoconjugate. Within LNFPIII is the Lewisx trisaccharide, expressed by many helminth parasites. In humans, LNFPIII is found in human milk and also known as stage-specific embryonic antigen-1. LNFPIII-NGC drives alternative activation of macrophages and dendritic cells via NFκB activation in a TLR4 dependent mechanism. However, the connection between LNFPIII-NGC activation of APCs, TLR4 signaling and subsequent MAP kinase signaling leading to anti-inflammatory activation of APCs remains unknown. In this study we determined that the innate receptor CD14 was essential for LNFPIII-NGC induction of both ERK and NFkB activation in APCs. Induction of ERK activation by LNFPIII-NGC was completely dependent on CD14/TLR4-Ras-Raf1/TPL2-MEK axis in bone marrow derived dendritic cells (BMDCs. In addition, LNFPIII-NGC preferentially induced the production of Th2 "favoring" chemokines CCL22 and matrix metalloprotease protein-9 in a CD14 dependent manner in BMDCs. In contrast, LNFPIII-NGC induces significantly lower levels of Th1 "favoring" chemokines, MIP1α, MIP1β and MIP-2 compared to levels in LPS stimulated cells. Interestingly, NGC of the identical human milk sugar LNnT, minus the alpha 1-3 linked fucose, failed to activate APCs via TLR4/MD2/CD14 receptor complex, suggesting that the alpha 1-3 linked fucose in LNFPIII and not on LNnT, is required for this process. Using specific chemical inhibitors of the MAPK pathway, we found that LNFPIII-NGC induction of CCL22, MMP9 and IL-10 production was dependent on ERK activation. Over all, this study suggests that LNFPIII-NGC utilizes CD14/TLR4-MAPK (ERK axis in modulating APC activation to produce anti-inflammatory chemokines and cytokines in a manner distinct from that seen for the pro-inflammatory PAMP LPS. These pathways may explain the in vivo

  1. The carboxyl-terminal domain of large T antigen rescues SV40 host range activity in trans independent of acetylation.

    Science.gov (United States)

    Poulin, Danielle L; DeCaprio, James A

    2006-05-25

    The host range activity of SV40 has been described as the inability of mutant viruses with deletions in the C terminal region of large T Ag to replicate in certain types of African green monkey kidney cells. We constructed new mutant viruses expressing truncated T Ag proteins and found that these mutant viruses exhibited the host range phenotype. The host range phenotype was independent of acetylation of T Ag at lysine 697. Co-expression of the C terminal domain of T Ag (aa 627-708) in trans increased both T Ag and VP1 mRNA as well as protein levels for host range mutant viruses in the restrictive cell type. In addition, the T Ag 627-708 fragment promoted the productive lytic infection of host range mutant viruses in the nonpermissive cell type. The carboxyl-terminal region of T Ag contains a biological function essential for the SV40 viral life cycle. PMID:16510165

  2. DNA-based vaccination induces humoral and cellular immune responses against hepatitis B virus surface antigen in mice without activation of Cmyc

    Institute of Scientific and Technical Information of China (English)

    Lian San Zhao; Shan Qin; Tao You Zhou; Hong Tang; Li Liu; Bing Jun Lei

    2000-01-01

    AIM To develop a safe and effective DNA vaccine for inducing humoral and cellular immunological responses against hepatitis B virus surface antigen (HBsAg). METHODS BALB/c mice were inoculated with NV-HB/s, a recombinant plasmid that had been inserted S gene of hepatitis B virus genome and could express HBsAg in eukaryotes. HBsAg expression was measured by ABC immunohistochemical assay, generation of anti-HBs by ELISA and cytotoxic T lymphocyte (CTL), by MTT method, existence of vaccine DNA by Southern blot hybridization and activation of oncogene C-myc by in situ hybridization.RESULTS With NV-HB/s vaccination by intramuscular injection, anti-HBs was initially positive 2 weeks after inoculation while all mice tested were HBsAg positive in the muscles. The titers and seroconversion rate of anti-HBs were steadily increasing as time went on and were dose-dependent. All the mice inoculated with 100 μg NV-HB/ s were anti-HBs positive one month after inoculation, the titer was 1:1024 or more. The humoral immune response was similar induced by either intramuscular or intradermal injection. CTL activities were much stronger (45.26%) in NV-HB/s DNA immunized mice as compared with those (only 6%) in plasmaderived HBsAg vaccine immunized mice. Two months after inoculation, all muscle samples were positive by Southern-blot hybridization for NV-HB/s DNA detection, but decreased to 25%and all were undetectable by in situ hybridization after 6 months. No oncogene Cmyc activation was found in the muscle of inoculation site. CONCLUSION NV-HB/s could generate humoral and cellular immunological responses against HBsAg that had been safely expressed in situ by NV-HB/s vaccination.

  3. Growth delay of human bladder cancer cells by Prostate Stem Cell Antigen downregulation is associated with activation of immune signaling pathways

    International Nuclear Information System (INIS)

    Prostate stem cell antigen (PSCA) is a glycosylphosphatidylinositol (GPI) anchored protein expressed not only in prostate but also in pancreas and bladder cancer as shown by immunohistochemistry and mRNA analysis. It has been targeted by monoclonal antibodies in preclinical animal models and more recently in a clinical trial in prostate cancer patients. The biological role played in tumor growth is presently unknown. In this report we have characterized the contribution of PSCA expression to tumor growth. A bladder cell line was engineered to express a doxycycline (dox) regulated shRNA against PSCA. To shed light on the PSCA biological role in tumor growth, microarray analysis was carried out as a function of PSCA expression. Expression of gene set of interest was further analyzed by qPCR Down regulation of the PSCA expression was associated with reduced cell proliferation in vitro and in vivo. Mice bearing subcutaneous tumors showed a reduced tumor growth upon treatment with dox, which effectively induced shRNA against PSCA as revealed by GFP expression. Pathway analysis of deregulated genes suggests a statistical significant association between PSCA downregulation and activation of genes downstream of the IFNα/β receptor. These experiments established for the first time a correlation between the level of PSCA expression and tumor growth and suggest a role of PSCA in counteracting the natural immune response

  4. Combined effects of 5-Fluorouracil, Folinic acid and Oxaliplatin on the expression of carcinoembryonic antigen in human colon cancer cells: pharmacological basis to develop an active antitumor immunochemotherapy

    Directory of Open Access Journals (Sweden)

    De Vecchis Liana

    2008-05-01

    Full Text Available Abstract Background Five-fluorouracil (FU, mainly associated with leucovorin (L, plays an essential role in chemotherapy of colorectal carcinoma. Moreover, FU ± L has been found to increase the expression of tumor-associated carcinoembryonic antigen (CEA, that may be an important target in therapeutic protocols of active specific immunotherapy. FU + L (FUL are frequently combined with oxaliplatin (OXA in advanced colon cancer patients. Thus, we investigated whether FUL in combination with OXA according to 2 different schedules may influence CEA expression in human colon cancer cells in vitro. Methods CEA protein expression was evaluated by cytofluorimetric and western blot analysis. Relative quantification of CEA mRNA was assessed by real time RT-PCR analysis. Results Levels of CEA protein and transcript were found to be higher in FUL-treated cells than in controls. However, when target cells were exposed to OXA before but not after FUL treatment, the up-regulation of CEA was partially inhibited. Conclusion These results suggest that target cells must be exposed to OXA after but not before treatment with the fluoropyrimidine in order to exploit drug-induced up-regulation of CEA. This finding appears to provide useful information to design chemo-immunotherapy protocols based on FUL + OXA, combined with host's immunity against CEA directed cancer vaccines.

  5. Understanding the Impact of ErbB Activating Events and Signal Transduction on Antigen Processing and Presentation: MHC Expression as a Model

    Science.gov (United States)

    Kersh, Anna E.; Sasaki, Maiko; Cooper, Lee A.; Kissick, Haydn T.; Pollack, Brian P.

    2016-01-01

    Advances in molecular pathology have changed the landscape of oncology. The ability to interrogate tissue samples for oncogene amplification, driver mutations, and other molecular alterations provides clinicians with an enormous level of detail about their patient’s cancer. In some cases, this information informs treatment decisions, especially those related to targeted anti-cancer therapies. However, in terms of immune-based therapies, it is less clear how to use such information. Likewise, despite studies demonstrating the pivotal role of neoantigens in predicting responsiveness to immune checkpoint blockade, it is not known if the expression of neoantigens impacts the response to targeted therapies despite a growing recognition of their diverse effects on immunity. To realize the promise of ‘personalized medicine’, it will be important to develop a more integrated understanding of the relationships between oncogenic events and processes governing anti-tumor immunity. One area of investigation to explore such relationships centers on defining how ErbB/HER activation and signal transduction influences antigen processing and presentation. PMID:27729860

  6. Antitumor activities of dFv-LDP-AE: An enediyne-energized fusion protein targeting tumor-associated antigen gelatinases.

    Science.gov (United States)

    Zhong, Gen-Shen; Wu, Min-Na; Guo, Xiao-Fang; Zhang, Sheng-Hua; Miao, Qing-Fang; Zhen, Yong-Su

    2012-10-01

    Gelatinases play an important role in tumor growth and metastasis, and overexpression of these molecules is strongly correlated with poor prognosis in a variety of malignant tumors. Lidamycin is an enediyne antitumor antibiotic with potent cytotoxicity. We previously reported that a tandem scFv format (dFv-LDP-AE) showed enhanced binding ability with gelatinases compared with the scFv-lidamycin conjugate (Fv-LDP-AE). In this study, the antitumor activities of dFv-LDP-AE on hepatocellular carcinoma (HCC) were evaluated in vitro and in vivo. By SDS-PAGE analysis, it was found that partial fusion protein dFv-LDP existed as dimer; the results of ELISA and immunofluorescence demonstrated that the fusion protein dFv-LDP could efficiently bind to hepatoma cells in vitro. The apparent arrest of cell cycle at G2/M phase and induction of apoptosis at nanomole levels indicated that the dFv-LDP-AE was very potent against HCC. In in vivo experiments, dFv-LDP-AE shown enhanced cytotoxic effects compared to those of LDM. Administration at mouse tolerable dosage level, the inhibition rate of tumor growth was 89.5% of dFv-LDP-AE vs. 73.6% of LDM on transplantable H22 in mice (Penediyne-energized fusion protein dFv-LDP-AE showed potential application as a new agent for therapeutic appications in HCC. PMID:22797730

  7. Centrosomal localisation of the cancer/testis (CT) antigens NY-ESO-1 and MAGE-C1 is regulated by proteasome activity in tumour cells.

    Science.gov (United States)

    Pagotto, Anna; Caballero, Otavia L; Volkmar, Norbert; Devalle, Sylvie; Simpson, Andrew J G; Lu, Xin; Christianson, John C

    2013-01-01

    The Cancer/Testis (CT) antigen family of genes are transcriptionally repressed in most human tissues but are atypically re-expressed in many malignant tumour types. Their restricted expression profile makes CT antigens ideal targets for cancer immunotherapy. As little is known about whether CT antigens may be regulated by post-translational processing, we investigated the mechanisms governing degradation of NY-ESO-1 and MAGE-C1 in selected cancer cell lines. Inhibitors of proteasome-mediated degradation induced the partitioning of NY-ESO-1 and MAGE-C1 into a detergent insoluble fraction. Moreover, this treatment also resulted in increased localisation of NY-ESO-1 and MAGE-C1 at the centrosome. Despite their interaction, relocation of either NY-ESO-1 or MAGE-C1 to the centrosome could occur independently of each other. Using a series of truncated fragments, the regions corresponding to NY-ESO-1(91-150) and MAGE-C1(900-1116) were established as important for controlling both stability and localisation of these CT antigens. Our findings demonstrate that the steady state levels of NY-ESO-1 and MAGE-C1 are regulated by proteasomal degradation and that both behave as aggregation-prone proteins upon accumulation. With proteasome inhibitors being increasingly used as front-line treatment in cancer, these data raise issues about CT antigen processing for antigenic presentation and therefore immunogenicity in cancer patients.

  8. Plasmids enriched with CpG motifs activate human peripheral blood mononuclear cells in vitro and enhance th-1 immune responses to hepatitis B surface antigen in mice.

    Science.gov (United States)

    Chen, Zhihui; Cao, Jie; Liao, Xiaoling; Ke, Jinshan; Zhu, Shiying; Zhao, Ping; Qi, Zhongtian

    2011-06-01

    T helper-1 (Th-1)-type immune responses play an important role in viral clearance during infection with hepatitis B virus (HBV). Unmethylated CpG motifs present in bacterial DNA can activate toll-like receptor 9 (TLR9) signals and act as potent adjuvants to induce Th-1-type immune responses. Here, a mini-plasmid with 812 base pairs in length was constructed and used as a vector to prepare a series of plasmids containing 3-21 copies of D-type CpG motifs. In vitro, these CpG-enriched plasmids strongly stimulated proliferation of human peripheral blood mononuclear cells (PBMCs) and enhanced secretion of interferon-γ (IFN-γ) and interleukin-12 (IL-12). The responses of the PBMCs from healthy individuals to the plasmids were stronger than those obtained from HBV-infected individuals. Contrary to the strong Th-2-biased response induced by surface antigen of hepatitis B virus (HBsAg) plus alum adjuvant, immunization of BALB/c mice with HBsAg plus these plasmids induced a strong Th-1-biased response. The plasmids increased the titers of HBsAg-specific total immunoglobulin G (IgG) and IgG(2a). HBsAg-specific IL-2 and IFN-γ production and cytotoxic activity were also enhanced in the presence of the plasmids. The strength of the immune responses positively correlated with the number of CpG motifs in the plasmids. These results indicate that the use of CpG-enriched plasmids as an adjuvant to recombinant HBsAg could provide a promising and cost-effective approach for the development of efficacious therapeutic vaccines against HBV infection. PMID:21668361

  9. Expression of an antigen homologous to the human CO17-1A/GA733 colon cancer antigen in animal tissues.

    OpenAIRE

    Zaloudik, J; Basak, S.; Nesbit, M.; Speicher, D W; Wunner, W H; Miller, E.; Ernst-Grotkowski, C.; Kennedy, R; Bergsagel, L. P.; Koido, T.; Herlyn, D

    1997-01-01

    The CO17-1A/GA733 antigen is associated with human carcinomas and some normal epithelial tissues. This antigen has shown promise as a target in approaches to passive and active immunotherapy of colorectal cancer. The relevance of animal models for studies of immunotherapy targeting this antigen in patients is dependent on the expression of the antigen on normal animal tissues. Immunohistoperoxidase staining with polyclonal rabbit antibodies to the human antigen revealed the human homologue on...

  10. Structural Characterization of Humanized Nanobodies with Neutralizing Activity against the Bordetella pertussis CyaA-Hemolysin: Implications for a Potential Epitope of Toxin-Protective Antigen.

    Science.gov (United States)

    Malik, Aijaz Ahmad; Imtong, Chompounoot; Sookrung, Nitat; Katzenmeier, Gerd; Chaicumpa, Wanpen; Angsuthanasombat, Chanan

    2016-04-01

    Previously, the 126-kDa CyaA-hemolysin (CyaA-Hly) fragment cloned from Bordetella pertussis--the causative agent of whooping cough--and functionally expressed in Escherichia coli was revealed as a key determinant for CyaA-mediated hemolysis against target erythrocytes. Here, phagemid-transfected E. coli clones producing nanobodies capable of binding to CyaA-Hly were selected from a humanized-camel VH/VHH phage-display library. Subsequently verified for binding activities by indirect ELISA and Western blotting, four CyaA-Hly-specific nanobodies were obtained and designated according to the presence/absence of VHH-hallmark amino acids as VHH2, VH5, VH18 and VHH37. In vitro neutralization assay revealed that all four ~17-kDa His-tagged VH/VHH nanobodies, in particular VHH37, which were over-expressed as inclusions and successfully unfolded-refolded, were able to effectively inhibit CyaA-Hly-mediated hemolysis. Phage-mimotope searching revealed that only peptides with sequence homologous to Linker 1 connecting Blocks I and II within the CyaA-RTX subdomain were able to bind to these four CyaA-Hly-specific nanobodies. Structural analysis of VHH37 via homology modeling and intermolecular docking confirmed that this humanized nanobody directly interacts with CyaA-RTX/Linker 1 through multiple hydrogen and ionic bonds. Altogether, our present data demonstrate that CyaA-RTX/Linker 1 could serve as a potential epitope of CyaA-protective antigen that may be useful for development of peptide-based pertussis vaccines. Additionally, such toxin-specific nanobodies have a potential for test-driven development of a ready-to-use therapeutic in passive immunization for mitigation of disease severity.

  11. Separation of uncompromised whole blood mixtures for single source STR profiling using fluorescently-labeled human leukocyte antigen (HLA) probes and fluorescence activated cell sorting (FACS).

    Science.gov (United States)

    Dean, Lee; Kwon, Ye Jin; Philpott, M Katherine; Stanciu, Cristina E; Seashols-Williams, Sarah J; Dawson Cruz, Tracey; Sturgill, Jamie; Ehrhardt, Christopher J

    2015-07-01

    Analysis of biological mixtures is a significant problem for forensic laboratories, particularly when the mixture contains only one cell type. Contributions from multiple individuals to biologic evidence can complicate DNA profile interpretation and often lead to a reduction in the probative value of DNA evidence or worse, its total loss. To address this, we have utilized an analytical technique that exploits the intrinsic immunological variation among individuals to physically separate cells from different sources in a mixture prior to DNA profiling. Specifically, we applied a fluorescently labeled antibody probe to selectively bind to one contributor in a mixture through allele-specific interactions with human leukocyte antigen (HLA) proteins that are expressed on the surfaces of most nucleated cells. Once the contributor's cells were bound to the probe, they were isolated from the mixture using fluorescence activated cell sorting (FACS)-a high throughput technique for separating cell populations based on their optical properties-and then subjected to STR analysis. We tested this approach on two-person and four-person whole blood mixtures where one contributor possessed an HLA allele (A*02) that was not shared by other contributors to the mixture. Results showed that hybridization of the mixture with a fluorescently-labeled antibody probe complimentary to the A*02 allele's protein product created a cell population with a distinct optical profile that could be easily differentiated from other cells in the mixture. After sorting the cells with FACS, genetic analysis showed that the STR profile of this cell population was consistent with that of the contributor who possessed the A*02 allele. Minor peaks from the A*02 negative contributor(s) were observed but could be easily distinguished from the profile generated from A*02 positive cells. Overall, this indicates that HLA antibody probes coupled to FACS may be an effective approach for generating STR profiles of

  12. Haemolytic activities of plant saponins and adjuvants. Effect of Periandra mediterranea saponin on the humoral response to the FML antigen of Leishmania donovani.

    Science.gov (United States)

    Santos, W R; Bernardo, R R; Peçanha, L M; Palatnik, M; Parente, J P; Palatnik de Sousa, C B

    1997-06-01

    An 87.7% (P saponin combined to the fucose-mannose ligand of Leishmania donovani (FML). However, an undesirable haemolytic effect was described for several saponins. Aiming to improve the formulation with FML/saponin, we comparatively analysed the haemolytic potential of recently characterized plant saponins and currently used adjuvants. The haemolytic activity of steroidic saponins from Agave sisalana; Smilax officinalis as well as commercial saponin (Riedel De Haën's), was higher than that of triterpenoid ones (Bredemeyera floribunda; Periandra mediterranea) and the Freund's complete adjuvant. The concentration resulting in 50% haemolysis was 500 micrograms ml-1 for aluminum hydroxide. The low haemolytic effect of P. mediterranea saponin was abolished by removal of its glycidic moiety and its sapogenin fraction as well as the Freund's Incomplete Adjuvant were non-haemolytic within this range. Furthermore, the adjuvant effect of three doses of P. mediterranea saponin injected with the FML antigen of L. donovani, was assayed in mice, either by the intraperitoneal (i.p.) or the subcutaneous (s.c.) route. The anti-FML IgG antibody levels increased and detectable levels were observed up to 3 months in the s.c. group. The response was expanded in both groups after an injection with a fourth vaccine dose. The IgG response showed increased levels of IgG2a only in the i.p. group, while IgG2b and IgG1 but not IgG3 antibodies were higher than controls in both groups. In conclusion, the results suggest that the recently described triterpenoid fractions of P. mediterranea can be safely used as adjuvant with low or non-haemolytic effect.

  13. Immunization with antigenic peptides complexed with β-glucan induces potent cytotoxic T-lymphocyte activity in combination with CpG-ODNs.

    Science.gov (United States)

    Mochizuki, Shinichi; Morishita, Hiromi; Kobiyama, Kouji; Aoshi, Taiki; Ishii, Ken J; Sakurai, Kazuo

    2015-12-28

    The induction of antigen-specific immune responses requires immunization with not only antigens, but also adjuvants. CpG oligonucleotides (CpG-ODNs) are well-known ligands for Toll-like receptor 9 and a potent adjuvant that induces both Th1-type humoral and cellular immune responses including cytotoxic T-lymphocyte responses. We previously demonstrated that β-glucan schizophyllan (SPG) can form complexes with CpG-ODNs with attached dA40 (CpG-dA/SPG), which can accumulate in macrophages in the draining inguinal lymph nodes and induce strong immune responses by co-administration of antigenic proteins, namely ovalbumin (OVA). Immunization with antigenic peptides, OVA257-264, did not induce these antigen-specific immune responses even in combination with CpG-dA/SPG, indicating that peptides require a carrier to antigen presenting cells. In this study, we prepared conjugates comprising OVA257-264 and dA40, and made complexes with SPG. Immunization with OVA257-264-dA/SPG induced peptide-specific immune responses in combination with CpG-dA regardless of complexation with SPG both in vitro and in vivo. When splenocytes from immunized mice were incubated with E.G7-OVA tumor model cells presenting OVA peptides, the number of cells drastically decreased after 24h. Furthermore, mice pre-immunized with OVA257-264-dA/SPG and CpG-ODNs exhibited a long delay in tumor growth after tumor inoculation. Therefore, these peptide-dA/SPG and CpG-dA/SPG complexes could be used as a potent vaccine for the treatment of cancers and infectious diseases. PMID:26562685

  14. CD85/LIR-1/ILT2 and CD152 (cytotoxic T lymphocyte antigen 4) inhibitory molecules down-regulate the cytolytic activity of human CD4+ T-cell clones specific for Mycobacterium tuberculosis.

    Science.gov (United States)

    Merlo, A; Saverino, D; Tenca, C; Grossi, C E; Bruno, S; Ciccone, E

    2001-10-01

    Antigen-specific cytolytic CD4+ T lymphocytes control Mycobacterium tuberculosis infection by secreting cytokines and by killing macrophages that have phagocytosed the pathogen. However, lysis of the latter cells promotes microbial dissemination, and other macrophages engulf the released bacteria. Subsequently, CD4+ T-cell-mediated killing of macrophages goes on, and this persistent process may hamper control of infection, unless regulatory mechanisms maintain a subtle balance between lysis of macrophages by cytolytic CD4+ cells and activation of cytolytic CD4+ cells by infected macrophages. We asked whether inhibitory molecules expressed by CD4+ cytolytic T lymphocytes could play a role in such a balance. To this end, human CD4+ T-cell clones specific for M. tuberculosis were produced that displayed an autologous major histocompatibility complex class II-restricted lytic ability against purified protein derivative (PPD)-pulsed antigen-presenting cells. All T-cell clones expressed CD152 (cytotoxic T-lymphocyte antigen 4 [CTLA-4]) and CD85/leukocyte immunoglobulin-like receptor 1 (LIR-1)/immunoglobulin-like transcript 2 (ILT2) inhibitory receptors, but not CD94 and the killer inhibitory receptor (or killer immunoglobulin-like receptor [KIR]) p58.2. CD3-mediated activation of the clones was inhibited in a redirected killing assay in which CD152 and CD85/LIR-1/ILT2 were cross-linked. Specific antigen-mediated proliferation of the clones was also sharply reduced when CD152 and CD85/LIR-1/ILT2 were cross-linked by specific monoclonal antibody (MAb) followed by goat anti-mouse antiserum. In contrast, blockade of the receptors by specific MAb only increased their proliferation. Production of interleukin 2 (IL-2) and gamma interferon (IFN-gamma) by the T-cell clones was also strongly reduced when CD152 and CD85/LIR-1/ILT2 were cross-linked. The lytic activity of the T-cell clones against PPD-pulsed autologous monocytes or Epstein-Barr virus-activated B cells was increased

  15. A liquid crystal of ascorbyl palmitate, used as vaccine platform, provides sustained release of antigen and has intrinsic pro-inflammatory and adjuvant activities which are dependent on MyD88 adaptor protein.

    Science.gov (United States)

    Sánchez Vallecillo, María F; Minguito de la Escalera, María M; Aguirre, María V; Ullio Gamboa, Gabriela V; Palma, Santiago D; González-Cintado, Leticia; Chiodetti, Ana L; Soldano, Germán; Morón, Gabriel; Allemandi, Daniel A; Ardavín, Carlos; Pistoresi-Palencia, María C; Maletto, Belkys A

    2015-09-28

    Modern subunit vaccines require the development of new adjuvant strategies. Recently, we showed that CpG-ODN formulated with a liquid crystal nanostructure formed by self-assembly of 6-O-ascorbyl palmitate (Coa-ASC16) is an attractive system for promoting an antigen-specific immune response to weak antigens. Here, we showed that after subcutaneous injection of mice with near-infrared fluorescent dye-labeled OVA antigen formulated with Coa-ASC16, the dye-OVA was retained at the injection site for a longer period than when soluble dye-OVA was administered. Coa-ASC16 alone elicited a local inflammation, but how this material triggers this response has not been described yet. Although it is known that some materials used as a platform are not immunologically inert, very few studies have directly focused on this topic. In this study, we explored the underlying mechanisms concerning the interaction between Coa-ASC16 and the immune system and we found that the whole inflammatory response elicited by Coa-ASC16 (leukocyte recruitment and IL-1β, IL-6 and IL-12 production) was dependent on the MyD88 protein. TLR2, TLR4, TLR7 and NLRP3-inflammasome signaling were not required for induction of this inflammatory response. Coa-ASC16 induced local release of self-DNA, and in TLR9-deficient mice IL-6 production was absent. In addition, Coa-ASC16 revealed an intrinsic adjuvant activity which was affected by MyD88 and IL-6 absence. Taken together these results indicate that Coa-ASC16 used as a vaccine platform is effective due to the combination of the controlled release of antigen and its intrinsic pro-inflammatory activity. Understanding how Coa-ASC16 works might have significant implications for rational vaccine design. PMID:26188153

  16. Relationship of spermatoscopy, prostatic acid phosphatase activity and prostate-specific antigen (p30) assays with further DNA typing in forensic samples from rape cases.

    Science.gov (United States)

    Romero-Montoya, Lydia; Martínez-Rodríguez, Hugo; Pérez, Miguel Antonio; Argüello-García, Raúl

    2011-03-20

    In the forensic laboratory the biological analyses for rape investigation commonly include vaginal swabs as sample material combined to biochemical tests including sperm cytology (SC) and detection of acid phosphatase activity (AP) and prostate-specific antigen (PSA, p30) for the conclusive identification of semen components. Most reports comparing these tests relied on analysis of semen samples or donor swabs taken under controlled conditions; however their individual or combined efficacy under real live sampling conditions in different laboratories is largely unknown. We carried out SC, APA and PSA analyses in vaginal swabs collected from casework rapes submitted to Mexican Forensic Laboratories at Texcoco and Toluca. On the basis of positive and negative results from each assay and sample, data were classified into eight categories (I-VIII) and compared with those obtained in the two only similar studies reported in Toronto, Canada and Hong Kong, China. SC and APA assays had the higher overall positivity in Toluca and Texcoco samples respectively and otherwise PSA had a lower but very similar positivity between these two laboratories. When compared to the previous studies some similarities were found, namely similar frequencies (at a ratio of approximately 1 out of 3) of samples being positive or negative by all techniques (Categories I and VI respectively) and a comparable overall positivity of APA and SC but higher than that of PSA. Indeed the combined results of using SC, APA and PSA tests was considered as conclusive for semen detection from approximately 1 out of 3 cases (Category I) to approximately 1 out of 2 cases in a scenario where at least SC is positive, strongly presumptive in 2 out of 3 cases (with at least one test positive) and the remainder 1 out of 3 cases (Category VI) suggested absence of semen. By determining Y-STR polymorphisms (12-loci) in additional samples obtained at Toluca laboratory, complete DNA profiles were determined from all

  17. Humanized Affinity-matured Monoclonal Antibody 8H9 Has Potent Antitumor Activity and Binds to FG Loop of Tumor Antigen B7-H3.

    Science.gov (United States)

    Ahmed, Mahiuddin; Cheng, Ming; Zhao, Qi; Goldgur, Yehuda; Cheal, Sarah M; Guo, Hong-fen; Larson, Steven M; Cheung, Nai-kong V

    2015-12-11

    B7-H3 (CD276) is both an inhibitory ligand for natural killer cells and T cells and a tumor antigen that is widely expressed among human solid tumors. Anti-B7-H3 mouse monoclonal antibody 8H9 has been successfully used for radioimmunotherapy for patients with B7-H3(+) tumors. We present the humanization, affinity maturation, and epitope mapping of 8H9 based on structure determination, modeling, and yeast display methods. The crystal structure of ch8H9 Fab fragment was solved to 2.5-Å resolution and used as a template for humanization. By displaying the humanized 8H9 single chain Fv (scFv) on the surface of yeast, the affinity was matured by sequential random mutagenesis and fluorescence-activated cell sorting. Six mutations (three in the complementarity-determining region and three in the framework regions) were identified and incorporated into an affinity-matured humanized 8H9 construct (hu8H9-6m) and an affinity-matured chimeric 8H9 construct (ch8H9-6m). The hu8H9-6m scFv had a 160-fold improvement in affinity (0.9 nm KD) compared with parental hu8H9 scFv (144 nm KD). The IgG formats of ch8H9-6m and hu8H9-6m (nanomolar to subnanomolar KD) had 2-9-fold enhancements in affinity compared with their parental forms, potent in vitro antibody-dependent cell-mediated cytotoxicity (0.1-0.3 μg/ml EC50), and high tumor uptake in mouse xenografts. Based on in silico docking studies and experimental validation, the molecular epitope of 8H9 was determined to be dependent on the FG loop of B7-H3, a region critical to its function in immunologic blockade and unique among anti-B7-H3 antibodies published to date. PMID:26487718

  18. COMPARISON OF EXCRETORY SECRETORY ANTIGENS OF ASCARIS LUMBRICOLDES AND TOXOCARA CATI 2ND. STAGE LARVAE WITH BIOCHEMICAL METHODES

    OpenAIRE

    F. Maleky; Massoud, J.

    1995-01-01

    Purification of parasitic antigens is a major activity in immunoparasitology because of its application in immunodiagnosis, vaccination analysis of immunopathology, preparation of monocolomal antibody and finding out the cross reactive antigens versus specific antigens of a parasite. In this survey F2 ultrafiltration excretory secretory antigens of Ascaris lumbricoides and Toxocara cati were separated by gel chromatography on sephacryl S-200 into several antigenic and non antigenic fractions....

  19. Wegener's granulomatosis and autoantibodies to neutrophil antigens

    OpenAIRE

    McCluskey, D R; Maxwell, A. P.; Watt, L

    1988-01-01

    We report five cases of Wegener's granulomatosis all of whom had clinical and histological evidence of disease activity at presentation and in whom autoantibodies to neutrophil antigens were detected. This test may prove useful for the diagnosis of this serious condition and help to monitor disease activity during treatment.

  20. Common acute lymphoblastic leukemia antigen (CALLA) is active neutral endopeptidase 24.11 ("enkephalinase"): direct evidence by cDNA transfection analysis.

    OpenAIRE

    Shipp, M A; Vijayaraghavan, J.; Schmidt, E V; Masteller, E L; D'Adamio, L; Hersh, L.B.; Reinherz, E L

    1989-01-01

    The common acute lymphoblastic leukemia antigen (CALLA) is a 749-amino acid type II integral membrane protein expressed by most acute lymphoblastic leukemias, certain other lymphoid malignancies with an immature phenotype, and normal lymphoid progenitors. A computer search against the most recent GenBank release (no. 56) indicates that human CALLA cDNA encodes a protein nearly identical to the rat and rabbit neutral endopeptidase 24.11 ("enkephalinase;" EC 3.4.24.11). This zinc metalloendopep...

  1. Similarities and differences in signal transduction by interleukin 4 and interleukin 13: analysis of Janus kinase activation.

    Science.gov (United States)

    Keegan, A D; Johnston, J A; Tortolani, P J; McReynolds, L J; Kinzer, C; O'Shea, J J; Paul, W E

    1995-08-15

    The cytokines interleukin (IL) 4 and IL-13 induce many of the same biological responses, including class switching to IgE and induction of major histocompatibility complex class II antigens and CD23 on human B cells. It has recently been shown that IL-4 induces the tyrosine phosphorylation of a 170-kDa protein, a substrate called 4PS, and of the Janus kinase (JAK) family members JAK1 and JAK3. Because IL-13 has many functional effects similar to those of IL-4, we compared the ability of IL-4 and IL-13 to activate these signaling molecules in the human multifactor-dependent cell line TF-1. In this report we demonstrate that both IL-4 and IL-13 induced the tyrosine phosphorylation of 4PS and JAK1. Interestingly, although IL-4 induced the tyrosine phosphorylation of JAK3, we did not detect JAK3 phosphorylation in response to IL-13. These data suggest that IL-4 and IL-13 signal in similar ways via the activation of JAK1 and 4PS. However, our data further indicate that there are significant differences because IL-13 does not activate JAK3.

  2. Production of soluble recombinant proteins with Kell, Duffy and Lutheran blood group antigen activity, and their use in screening human sera for Kell, Duffy and Lutheran antibodies.

    Science.gov (United States)

    Ridgwell, K; Dixey, J; Scott, M L

    2007-10-01

    The aim of this study was to show that soluble recombinant (sr) proteins can mimic blood group antigens and be used to screen human sera for blood-group-specific antibodies. The blood of all pregnant women and pretransfusion patients should be screened for blood-group-specific antibodies to identify and monitor pregnancies at risk of haemolytic disease of the foetus and newborn (HDFN), and to prevent haemolytic transfusion reactions. Current antibody screening and identification methods use human red blood cell panels, which can complicate antibody identification if more than one antibody specificity is present. COS-7 cells were transfected to produce sr forms of the extracellular domains of the red blood cell membrane proteins that express Kell, Duffy or Lutheran blood group antigens. These sr proteins were used to screen for and identify anti-Kell, anti-Duffy or anti-Lutheran blood-group-specific allo-antibodies in human sera by haemagglutination inhibition and in solid-phase enzyme-linked immunosorbent assays (ELISAs). There is a positive correlation (correlation coefficient 0.605, P value 0.002) between antibody titre by standard indirect antiglobulin test (IAT) and signal intensity in the ELISA test. This work shows that sr proteins can mimic blood group antigens and react with human allogeneic antibodies, and that such proteins could be used to develop solid-phase, high-throughput blood group antibody screening and identification platforms. PMID:17725551

  3. Novel selective inhibitors of aminopeptidases that generate antigenic peptides.

    Science.gov (United States)

    Papakyriakou, Athanasios; Zervoudi, Efthalia; Theodorakis, Emmanuel A; Saveanu, Loredana; Stratikos, Efstratios; Vourloumis, Dionisios

    2013-09-01

    Endoplasmic reticulum aminopeptidases, ERAP1 and ERAP2, as well as Insulin regulated aminopeptidase (IRAP) play key roles in antigen processing, and have recently emerged as biologically important targets for manipulation of antigen presentation. Taking advantage of the available structural and substrate-selectivity data for these enzymes, we have rationally designed a new series of inhibitors that display low micromolar activity. The selectivity profile for these three highly homologous aminopeptidases provides a promising avenue for modulating intracellular antigen processing. PMID:23916253

  4. Acute Lymphoid Leukemia Cells with Greater Stem Cell Antigen-1 (Ly6a/Sca-1) Expression Exhibit Higher Levels of Metalloproteinase Activity and Are More Aggressive In Vivo

    OpenAIRE

    Yu-Chiao Hsu; Kurt Mildenstein; Kordell Hunter; Olena Tkachenko; Mullen, Craig A.

    2014-01-01

    Stem cell antigen-1 (Ly6a/Sca-1) is a gene that is expressed in activated lymphocytes, hematopoietic stem cells and stem cells of a variety of tissues in mice. Despite decades of study its functions remain poorly defined. These studies explored the impact of expression of this stem cell associated gene in acute lymphoid leukemia. Higher levels of Ly6a/Sca-1 expression led to more aggressive leukemia growth in vivo and earlier death of hosts. Leukemias expressing higher levels of Ly6a/Sca-1 ex...

  5. Simian virus 40 T antigen can transcriptionally activate and mediate viral DNA replication in cells which lack the retinoblastoma susceptibility gene product.

    OpenAIRE

    Trifillis, P; Picardi, J; Alwine, J C

    1990-01-01

    Simian virus 40 T antigen is a multifunctional protein which has recently been shown to form a complex with the retinoblastoma susceptibility gene product (Rb protein) (J.A. DeCaprio, J.W. Ludlow, J. Figge, J.-Y. Shaw, C.-M. Huang, W.-H. Lee, E. Marsilio, E. Paucha, and D.M. Livingston, Cell 54:275-283, 1988; P. Whyte, K.J. Buchkovich, J.M. Horowitz, S.H. Friend, M. Raybuck, R.A. Weinberg, and E. Harlow, Nature (London) 334:124-129, 1988). This interaction may facilitate some of the functions...

  6. Murine antigen-induced arthritis.

    NARCIS (Netherlands)

    Berg, W.B. van den; Joosten, L.A.B.; Lent, P.L.E.M. van

    2007-01-01

    Antigen induced arthritis is a unilateral T-cell driven model caused by direct injection of an antigen into the knee joint of a FCA preimmunized animal. The chronicity is determined by antigen retention in avascular structures of the joint through charge mediated binding or antibody mediated trappin

  7. The highly antigenic 53/25 kDa Taenia solium protein fraction with cathepsin-L like activity is present in the oncosphere/cysticercus and induces non-protective IgG antibodies in pigs.

    Science.gov (United States)

    Zimic, Mirko; Pajuelo, Mónica; Gilman, Robert H; Gutiérrez, Andrés H; Rueda, Luis D; Flores, Myra; Chile, Nancy; Verástegui, Manuela; Gonzalez, Armando; García, Héctor H; Sheen, Patricia

    2012-01-15

    Cathepsin L-like proteases are secreted by several parasites including Taenia solium. The mechanism used by T. solium oncospheres to degrade and penetrate the intestine and infect the host is incompletely understood. It is assumed that intestinal degradation is driven by the proteolytic activity of enzymes secreted by the oncosphere. Blocking the proteolytic activity by an antibody response would prevent the oncosphere penetration and further infection. Serine and cysteine proteases including chymotrypsin, trypsin, elastase, and cathepsin L, are secreted by T. solium and Taenia saginata oncospheres when cultured in vitro, being potential vaccine candidates. However, the purification of a sufficient quantity of proteases secreted by oncospheres to conduct a vaccine trial is costly and lengthy. A 53/25 kDa cathepsin L-like fraction partially purified from T. solium cyst fluid was described previously as an important antigen for immunodiagnostics. In this study we found that this antigen is present in the T. solium oncosphere and is also secreted by the cysticercus. This protein fraction was tested for its ability to protect pigs against an oral challenge with T. solium oncospheres in a vaccine trial. IgG antibodies against the 53/25 kDa cathepsin L-like protein fraction were elicited in the vaccinated animals but did not confer protection.

  8. Heat Shock Enhances the Expression of the Human T Cell Leukemia Virus Type-I (HTLV-I) Trans-Activator (Tax) Antigen in Human HTLV-I Infected Primary and Cultured T Cells.

    Science.gov (United States)

    Kunihiro, Marie; Fujii, Hideki; Miyagi, Takuya; Takahashi, Yoshiaki; Tanaka, Reiko; Fukushima, Takuya; Ansari, Aftab A; Tanaka, Yuetsu

    2016-07-11

    The environmental factors that lead to the reactivation of human T cell leukemia virus type-1 (HTLV-I) in latently infected T cells in vivo remain unknown. It has been previously shown that heat shock (HS) is a potent inducer of HTLV-I viral protein expression in long-term cultured cell lines. However, the precise HTLV-I protein(s) and mechanisms by which HS induces its effect remain ill-defined. We initiated these studies by first monitoring the levels of the trans-activator (Tax) protein induced by exposure of the HTLV-I infected cell line to HS. HS treatment at 43 °C for 30 min for 24 h led to marked increases in the level of Tax antigen expression in all HTLV-I-infected T cell lines tested including a number of HTLV-I-naturally infected T cell lines. HS also increased the expression of functional HTLV-I envelope gp46 antigen, as shown by increased syncytium formation activity. Interestingly, the enhancing effect of HS was partially inhibited by the addition of the heat shock protein 70 (HSP70)-inhibitor pifithlin-μ (PFT). In contrast, the HSP 70-inducer zerumbone (ZER) enhanced Tax expression in the absence of HS. These data suggest that HSP 70 is at least partially involved in HS-mediated stimulation of Tax expression. As expected, HS resulted in enhanced expression of the Tax-inducible host antigens, such as CD83 and OX40. Finally, we confirmed that HS enhanced the levels of Tax and gp46 antigen expression in primary human CD4⁺ T cells isolated from HTLV-I-infected humanized NOD/SCID/γc null (NOG) mice and HTLV-I carriers. In summary, the data presented herein indicate that HS is one of the environmental factors involved in the reactivation of HTLV-I in vivo via enhanced Tax expression, which may favor HTLV-I expansion in vivo.

  9. Heat Shock Enhances the Expression of the Human T Cell Leukemia Virus Type-I (HTLV-I Trans-Activator (Tax Antigen in Human HTLV-I Infected Primary and Cultured T Cells

    Directory of Open Access Journals (Sweden)

    Marie Kunihiro

    2016-07-01

    Full Text Available The environmental factors that lead to the reactivation of human T cell leukemia virus type-1 (HTLV-I in latently infected T cells in vivo remain unknown. It has been previously shown that heat shock (HS is a potent inducer of HTLV-I viral protein expression in long-term cultured cell lines. However, the precise HTLV-I protein(s and mechanisms by which HS induces its effect remain ill-defined. We initiated these studies by first monitoring the levels of the trans-activator (Tax protein induced by exposure of the HTLV-I infected cell line to HS. HS treatment at 43 °C for 30 min for 24 h led to marked increases in the level of Tax antigen expression in all HTLV-I-infected T cell lines tested including a number of HTLV-I-naturally infected T cell lines. HS also increased the expression of functional HTLV-I envelope gp46 antigen, as shown by increased syncytium formation activity. Interestingly, the enhancing effect of HS was partially inhibited by the addition of the heat shock protein 70 (HSP70-inhibitor pifithlin-μ (PFT. In contrast, the HSP 70-inducer zerumbone (ZER enhanced Tax expression in the absence of HS. These data suggest that HSP 70 is at least partially involved in HS-mediated stimulation of Tax expression. As expected, HS resulted in enhanced expression of the Tax-inducible host antigens, such as CD83 and OX40. Finally, we confirmed that HS enhanced the levels of Tax and gp46 antigen expression in primary human CD4+ T cells isolated from HTLV-I-infected humanized NOD/SCID/γc null (NOG mice and HTLV-I carriers. In summary, the data presented herein indicate that HS is one of the environmental factors involved in the reactivation of HTLV-I in vivo via enhanced Tax expression, which may favor HTLV-I expansion in vivo.

  10. Heat Shock Enhances the Expression of the Human T Cell Leukemia Virus Type-I (HTLV-I) Trans-Activator (Tax) Antigen in Human HTLV-I Infected Primary and Cultured T Cells.

    Science.gov (United States)

    Kunihiro, Marie; Fujii, Hideki; Miyagi, Takuya; Takahashi, Yoshiaki; Tanaka, Reiko; Fukushima, Takuya; Ansari, Aftab A; Tanaka, Yuetsu

    2016-01-01

    The environmental factors that lead to the reactivation of human T cell leukemia virus type-1 (HTLV-I) in latently infected T cells in vivo remain unknown. It has been previously shown that heat shock (HS) is a potent inducer of HTLV-I viral protein expression in long-term cultured cell lines. However, the precise HTLV-I protein(s) and mechanisms by which HS induces its effect remain ill-defined. We initiated these studies by first monitoring the levels of the trans-activator (Tax) protein induced by exposure of the HTLV-I infected cell line to HS. HS treatment at 43 °C for 30 min for 24 h led to marked increases in the level of Tax antigen expression in all HTLV-I-infected T cell lines tested including a number of HTLV-I-naturally infected T cell lines. HS also increased the expression of functional HTLV-I envelope gp46 antigen, as shown by increased syncytium formation activity. Interestingly, the enhancing effect of HS was partially inhibited by the addition of the heat shock protein 70 (HSP70)-inhibitor pifithlin-μ (PFT). In contrast, the HSP 70-inducer zerumbone (ZER) enhanced Tax expression in the absence of HS. These data suggest that HSP 70 is at least partially involved in HS-mediated stimulation of Tax expression. As expected, HS resulted in enhanced expression of the Tax-inducible host antigens, such as CD83 and OX40. Finally, we confirmed that HS enhanced the levels of Tax and gp46 antigen expression in primary human CD4⁺ T cells isolated from HTLV-I-infected humanized NOD/SCID/γc null (NOG) mice and HTLV-I carriers. In summary, the data presented herein indicate that HS is one of the environmental factors involved in the reactivation of HTLV-I in vivo via enhanced Tax expression, which may favor HTLV-I expansion in vivo. PMID:27409630

  11. Heat Shock Enhances the Expression of the Human T Cell Leukemia Virus Type-I (HTLV-I) Trans-Activator (Tax) Antigen in Human HTLV-I Infected Primary and Cultured T Cells

    Science.gov (United States)

    Kunihiro, Marie; Fujii, Hideki; Miyagi, Takuya; Takahashi, Yoshiaki; Tanaka, Reiko; Fukushima, Takuya; Ansari, Aftab A.; Tanaka, Yuetsu

    2016-01-01

    The environmental factors that lead to the reactivation of human T cell leukemia virus type-1 (HTLV-I) in latently infected T cells in vivo remain unknown. It has been previously shown that heat shock (HS) is a potent inducer of HTLV-I viral protein expression in long-term cultured cell lines. However, the precise HTLV-I protein(s) and mechanisms by which HS induces its effect remain ill-defined. We initiated these studies by first monitoring the levels of the trans-activator (Tax) protein induced by exposure of the HTLV-I infected cell line to HS. HS treatment at 43 °C for 30 min for 24 h led to marked increases in the level of Tax antigen expression in all HTLV-I-infected T cell lines tested including a number of HTLV-I-naturally infected T cell lines. HS also increased the expression of functional HTLV-I envelope gp46 antigen, as shown by increased syncytium formation activity. Interestingly, the enhancing effect of HS was partially inhibited by the addition of the heat shock protein 70 (HSP70)-inhibitor pifithlin-μ (PFT). In contrast, the HSP 70-inducer zerumbone (ZER) enhanced Tax expression in the absence of HS. These data suggest that HSP 70 is at least partially involved in HS-mediated stimulation of Tax expression. As expected, HS resulted in enhanced expression of the Tax-inducible host antigens, such as CD83 and OX40. Finally, we confirmed that HS enhanced the levels of Tax and gp46 antigen expression in primary human CD4+ T cells isolated from HTLV-I-infected humanized NOD/SCID/γc null (NOG) mice and HTLV-I carriers. In summary, the data presented herein indicate that HS is one of the environmental factors involved in the reactivation of HTLV-I in vivo via enhanced Tax expression, which may favor HTLV-I expansion in vivo. PMID:27409630

  12. The role of Ia molecules in the activation of T lymphocytes. III. Antigen-specific, Ia-restricted, interleukin 2-producing T cell hybridomas with detectable affinity for the restricting I-A molecule

    OpenAIRE

    1983-01-01

    Antigen-specific I region-restricted, interleukin 2-producing T cell hybridomas were produced by fusing GAT-specific T cell blasts with BW5147. Two antigen-specific phenotypes were identified, one autoreactive and one nonautoreactive. All of the antigen-specific and autoreactive clones were H-2 restricted, mapping to the IA subregion by genetic analysis and monoclonal antibody inhibition. Both the antigen- specific and autoreactive stimulation are the property of a single cell and required no...

  13. Overexpression of linker for activated T cells, cyclooxygenase-2, CD1a, CD68 and myeloid/histiocyte antigen in an inflamed seborrheic keratosis

    Directory of Open Access Journals (Sweden)

    Ana Maria Abreu-Velez

    2011-01-01

    Full Text Available Context: Inflamed seborrheic keratoses are generally associated with the accumulation of variable numbers of lymphocytes and histiocytes in the superficial dermis. The precise immunologic mechanism of this histologic phenomenon is not known Case Report: A 62-year-old male presented with a patch on the right neck with additional features of inflammation. Skin biopsies for hematoxylin and eosin examination, as well as for immunohistochemistry analysis were performed. Results: H&E staining demonstrated classic features of an inflamed seborrheic keratosis. Overexpression of LAT, COX-2, CD1a, and CD68 was noticed in the inflammatory infiltrate. A strong presence of CD1a was also seen in the epidermis suprajacent to the inflammation. Myeloid/histiocyte antigen was strongly expressed by the keratinocytes . Conclusion : A complex immune response seems to be involved in the pathophysiology of an inflamed seborrheic keratosis.

  14. Human antigen-presenting cells respond differently to gut-derived probiotic bacteria but mediate similar strain-dependent NK and T cell activation

    DEFF Research Database (Denmark)

    Fink, Lisbeth Nielsen; Zeuthen, Louise Hjerrild; Ferlazzo, Guido;

    2007-01-01

    (APC) was compared; blood myeloid dendritic cells (DC), monocyte-derived DC and monocytes, and the effector response of natural killer cells and naïve T cells was characterized. Maturation induced by gut-derived bacteria differed between APC, with blood DC and monocytes responding with the production...... of IL-6 and tumour necrosis factor-alpha to bacteria, which elicited mainly IL-10 in monocyte-derived DC. In contrast, comparable IFN-gamma production patterns were found in both natural killer cells and T cells induced by all bacteria-matured APC. An inhibitory effect of certain strains on this IFN......, in vitro assessment of the immunomodulatory effects of distinct strains may depend strongly on the cell type used as a model. To select the most appropriate model for screening of beneficial bacteria in human cells, the response to strains of intestinal bacteria of three types of antigen-presenting cells...

  15. A Role For Mitochondria In Antigen Processing And Presentation.

    Science.gov (United States)

    Bonifaz, Lc; Cervantes-Silva, Mp; Ontiveros-Dotor, E; López-Villegas, Eo; Sánchez-García, Fj

    2014-09-23

    Immune synapse formation is critical for T lymphocyte activation, and mitochondria have a role in this process, by localizing close to the immune synapse, regulating intracellular calcium concentration, and providing locally required ATP. The interaction between antigen presenting cells (APCs) and T lymphocytes is a two-way signaling process. However, the role of mitochondria in antigen presenting cells during this process remains unknown. For APCs to be able to activate T lymphocytes, they must first engage in an antigen-uptake, -processing, and -presentation process. Here we show that HEL-loaded B lymphocytes, as a type of APCs, undergo a small but significant mitochondrial depolarization by 1-2 h following antigen exposure thus suggesting an increase in their metabolic demands. Inhibition of ATP synthase (oligomycin) or mitochondrial Ca(2+) uniporter (MCU) (Ruthenium red) had no effect on antigen uptake. Therefore, antigen processing and antigen presentation were further analyzed. Oligomycin treatment reduced the amount of specific MHC-peptide complexes but not total MHC II on the cell membrane of B lymphocytes which correlated with a decrease in antigen presentation. However, oligomycin also reduced antigen presentation by B lymphocytes that endogenously express HEL and by B lymphocytes loaded with the HEL48-62 peptide, although to a lesser extent. ATP synthase inhibition and MCU inhibition had a clear inhibitory effect on antigen processing (DQ-OVA). Taking together these results suggest that ATP synthase and MCU are relevant for antigen processing and presentation. Finally, APCs mitochondria were found to re-organize towards the APC-T immune synapse. This article is protected by copyright. All rights reserved.

  16. Focal radiation therapy combined with 4-1BB activation and CTLA-4 blockade yields long-term survival and a protective antigen-specific memory response in a murine glioma model.

    Directory of Open Access Journals (Sweden)

    Zineb Belcaid

    Full Text Available BACKGROUND: Glioblastoma (GBM is the most common malignant brain tumor in adults and is associated with a poor prognosis. Cytotoxic T lymphocyte antigen -4 (CTLA-4 blocking antibodies have demonstrated an ability to generate robust antitumor immune responses against a variety of solid tumors. 4-1BB (CD137 is expressed by activated T lymphocytes and served as a co-stimulatory signal, which promotes cytotoxic function. Here, we evaluate a combination immunotherapy regimen involving 4-1BB activation, CTLA-4 blockade, and focal radiation therapy in an immune-competent intracranial GBM model. METHODS: GL261-luciferace cells were stereotactically implanted in the striatum of C57BL/6 mice. Mice were treated with a triple therapy regimen consisted of 4-1BB agonist antibodies, CTLA-4 blocking antibodies, and focal radiation therapy using a small animal radiation research platform and mice were followed for survival. Numbers of brain-infiltrating lymphocytes were analyzed by FACS analysis. CD4 or CD8 depleting antibodies were administered to determine the relative contribution of T helper and cytotoxic T cells in this regimen. To evaluate the ability of this immunotherapy to generate an antigen-specific memory response, long-term survivors were re-challenged with GL261 glioma en B16 melanoma flank tumors. RESULTS: Mice treated with triple therapy had increased survival compared to mice treated with focal radiation therapy and immunotherapy with 4-1BB activation and CTLA-4 blockade. Animals treated with triple therapy exhibited at least 50% long-term tumor free survival. Treatment with triple therapy resulted in a higher density of CD4+ and CD8+ tumor infiltrating lymphocytes. Mechanistically, depletion of CD4+ T cells abrogated the antitumor efficacy of triple therapy, while depletion of CD8+ T cells had no effect on the treatment response. CONCLUSION: Combination therapy with 4-1BB activation and CTLA-4 blockade in the setting of focal radiation therapy

  17. Antigen pulsed CpG-ODN activated dendritic cells induce host-protective immune response by regulating the T regulatory cell functioning in Leishmania donovani-infected mice: critical role of CXCL10

    Directory of Open Access Journals (Sweden)

    Saikat eMajumder

    2014-06-01

    Full Text Available Visceral leishmaniasis (VL, caused by Leishmania donovani, is a systemic infection of reticulo-endothelial system. There is currently no protective vaccine against VL and chemotherapy is increasingly limited due to appearance of drug resistance to first line drugs such as antimonials and amphotericin B. In the present study, by using a murine model of leishmaniasis we evaluated the function played by soluble leishmanial antigen (SLA pulsed-CpG-ODN stimulated dendritic cells (SLA-CpG-DCs in restricting the intracellular parasitic growth. We establish that a single dose of SLA-CpG-DCs vaccination is sufficient in rendering complete protection against Leishmania donovani infection. In probing the possible mechanism, we observed that SLA-CpG-DCs vaccination results in the significant decrease in Foxp3+GITR+CTLA4+CD4+CD25+ Treg cell population in Leishmania-infected mice. Vaccination with these antigen stimulated dendritic cells results in the decrease in the secretion of TGF-β by these Treg cells by possible regulation of the SMAD signalling. Moreover, we demonstrated that a CXC chemokine, IFN-γ-inducible protein 10 (IP-10, has a direct role in the regulation of CD4+CD25+ Treg cells in SLA-CpG-DCs vaccinated parasitized mice as Treg cells isolated from IP-10 depleted vaccinated mice showed significantly increased TGF-β production and suppressive activity.

  18. Carcino-Embryonic Antigen

    International Nuclear Information System (INIS)

    Tumour marker analysis has increased our understanding of the presence of tumours in the body. Carcino-embryonic antigen, CEA, is one of the best studied tumour markers and has proved an ideal diagnostic adjuvant. It has helped in quantifying the amount of disease present in a patient and thence to make accurate prognosis on the various diagnosed ailments. At UCH, it is observed that there is an increase in cancer related ailments and therefore the need for early diagnosis is more compelling in our environment to mitigate future cost of managing advanced manifestation

  19. Human antigen-presenting cells respond differently to gut-derived probiotic bacteria but mediate similar strain-dependent NK and T cell activation.

    Science.gov (United States)

    Fink, Lisbeth N; Zeuthen, Louise H; Ferlazzo, Guido; Frøkiaer, Hanne

    2007-12-01

    The intestinal microbiota is essential for homeostasis of the local and systemic immune system, and particularly strains of lactic acid bacteria and Escherichia coli have been shown to have balancing effects on inflammatory conditions such as allergy and inflammatory bowel disease. However, in vitro assessment of the immunomodulatory effects of distinct strains may depend strongly on the cell type used as a model. To select the most appropriate model for screening of beneficial bacteria in human cells, the response to strains of intestinal bacteria of three types of antigen-presenting cells (APC) was compared; blood myeloid dendritic cells (DC), monocyte-derived DC and monocytes, and the effector response of natural killer cells and naïve T cells was characterized. Maturation induced by gut-derived bacteria differed between APC, with blood DC and monocytes responding with the production of IL-6 and tumour necrosis factor-alpha to bacteria, which elicited mainly IL-10 in monocyte-derived DC. In contrast, comparable IFN-gamma production patterns were found in both natural killer cells and T cells induced by all bacteria-matured APC. An inhibitory effect of certain strains on this IFN-gamma production was also mediated by all types of APC. The most potent responses were induced by monocyte-derived DC, which thus constitute a sensitive screening model. PMID:17903206

  20. Enhancement of cytotoxic T lymphocyte activity by dendritic cells loaded with Tat-protein transduction domain-fused hepatitis B virus core antigen

    Institute of Scientific and Technical Information of China (English)

    2008-01-01

    The protein transduction domain (PTD) of human immuno-deficiency virus-1-Tat protein has a unique potency to pen-etrate the cellular membranes. To synthesize the sequence of Tat-PTD and hepatitis B virus core antigen (HBcAg), we spliced these sequences and linked a fusion gene into the pMAL-c2x vector. The fusion proteins were purified by affin-ity chromatography and pulsed with bone marrow -derived den-dritic cells (DCs), and the transduction of recombinant pro-tein was detected by immunofluorescence antibody assay.Results showed that recombinant PTD-HBcAg could pen-etrate into DC cytoplasm while recombinant HBcAg was de-tected on the surface of cells. The percentage of DC surface molecules, such as CD80, CD86 and major histocompatibii-ity complex Ⅱ, and production of cytokine (IL-12pT0) induced by recombinant PTD-HBcAg were significantly higher than those induced by recombinant HBcAg or tumor necrosis fac-tor-α. DCs treated with PTD-HBcAg induced T cells to dif-ferentiate into specific cytotoxic T lymphocytes (CTLs) and enhanced the CTL killing response. In conclusion, the ex-pressed and purified PTD-HBcAg fusion protein could pen-etrate into cells through the plasma membrane, promote DC maturation, and enhance T cells response to generate HBcAg-specific CTLs efficiently.

  1. Cancer testis antigen and immunotherapy

    Directory of Open Access Journals (Sweden)

    Krishnadas DK

    2013-04-01

    Full Text Available Deepa Kolaseri Krishnadas, Fanqi Bai, Kenneth G Lucas Department of Pediatrics, Division of Hematology/Oncology, University of Louisville, KY, USA Abstract: The identification of cancer testis (CT antigens has been an important advance in determining potential targets for cancer immunotherapy. Multiple previous studies have shown that CT antigen vaccines, using both peptides and dendritic cell vaccines, can elicit clinical and immunologic responses in several different tumors. This review details the expression of melanoma antigen family A, 1 (MAGE-A1, melanoma antigen family A, 3 (MAGE-A3, and New York esophageal squamous cell carcinoma-1 (NY-ESO-1 in various malignancies, and presents our current understanding of CT antigen based immunotherapy. Keywords: cancer testis antigens, immunotherapy, vaccine

  2. Antigen receptor signaling: integration of protein tyrosine kinase functions.

    Science.gov (United States)

    Tamir, I; Cambier, J C

    1998-09-17

    Antigen receptors on T and B cells function to transduce signals leading to a variety of biologic responses minimally including antigen receptor editing, apoptotic death, developmental progression, cell activation, proliferation and survival. The response to antigen depends upon antigen affinity and valence, involvement of coreceptors in signaling and differentiative stage of the responding cell. The requirement that these receptors integrate signals that drive an array of responses may explain their evolved structural complexity. Antigen receptors are composed of multiple subunits compartmentalized to provide antigen recognition and signal transduction function. In lieu of on-board enzymatic activity these receptors rely on associated Protein Tyrosine Kinases (PTKs) for their signaling function. By aggregating the receptors, and hence their appended PTKs, antigens induce PTK transphosphorylation, activating them to phosphorylate the receptor within conserved motifs termed Immunoreceptor Tyrosine-based Activation Motifs (ITAMs) found in transducer subunits. The tyrosyl phosphorylated ITAMs then interact with Src Homology 2 (SH2) domains within the PTKs leading to their further activation. As receptor phosphorylation is amplified, other effectors, such as Shc, dock by virtue of SH2 binding, and serve, in-turn, as substrates for these PTKs. This sequence of events not only provides a signal amplification mechanism by combining multiple consecutive steps with positive feedback, but also allows for signal diversification by differential recruitment of effectors that provide access to distinct parallel downstream signaling pathways. The subject of antigen receptor signaling has been recently reviewed in depth (DeFranco, 1997; Kurosaki, 1997). Here we discuss the biochemical basis of antigen receptor signal transduction, using the B cell receptor (BCR) as a paradigm, with specific emphasis on the involved PTKs. We review several specific mechanisms by which responses

  3. Use of antigenic cartography in vaccine seed strain selection.

    Science.gov (United States)

    Fouchier, Ron A M; Smith, Derek J

    2010-03-01

    Human influenza A viruses are classic examples of antigenically variable pathogens that have a seemingly endless capacity to evade the host's immune response. The viral hemagglutinin (HA) and neuraminidase (NA) proteins are the main targets of our antibody response to combat infections. HA and NA continuously change to escape from humoral immunity, a process known as antigenic drift. As a result of antigenic drift, the human influenza vaccine is updated frequently. The World Health Organization (WHO) coordinates a global influenza surveillance network that, by the hemagglutination inhibition (HI) assay, routinely characterizes the antigenic properties of circulating strains in order to select new seed viruses for such vaccine updates. To facilitate a quantitative interpretation and easy visualization of HI data, a new computational technique called "antigenic cartography" was developed. Since its development, antigenic cartography has been applied routinely to assist the WHO with influenza surveillance activities. Until recently, antigenic variation was not considered a serious issue with influenza vaccines for poultry. However, because of the diversification of the Asian H5N1 lineage since 1996 into multiple genetic clades and subclades, and because of the long-term use of poultry vaccines against H5 in some parts of the world, this issue needs to be re-addressed. The antigenic properties of panels of avian H5N1 viruses were characterized by HI assay, using mammalian or avian antisera, and analyzed using antigenic cartography methods. These analyses revealed antigenic differences between circulating H5N1 viruses and the H5 viruses used in poultry vaccines. Considerable antigenic variation was also observed within and between H5N1 clades. These observations have important implications for the efficacy and long-term use of poultry vaccines.

  4. Human leucocyte antigens in tympanosclerosis.

    Science.gov (United States)

    Dursun, G; Acar, A; Turgay, M; Calgüner, M

    1997-02-01

    This study was designed to evaluate the association between certain HLA antigens and tympanosclerosis. The serum concentrations of HLA antigens were measured by a microlymphocytotoxicity technique in patients with tympanosclerosis and compared with a healthy control group. The serum levels of HLA-B35 and -DR3 were significantly higher in the patients with tympanosclerosis. This result suggests that certain types of HLA antigens may play an important role as an indicator or mediator in the pathogenesis of tympanosclerosis. PMID:9088683

  5. Structure-activity relationships of vanillic acid ester analogs in inhibitory effect of antigen-mediated degranulation in rat basophilic leukemia RBL-2H3 cells.

    Science.gov (United States)

    Ishimata, Nao; Ito, Hideyuki; Tai, Akihiro

    2016-08-01

    Methyl vanillate (1) showed strong degranulation inhibitory activity among vanillin derivatives tested. In order to find structure-activity relationships for developing anti-allergic agents with simple structures and potent activity, we synthesized several vanillic acid (VA) ester derivatives with C1-C4 and C8 alkyl chains and evaluated their degranulation inhibitory activities. The most active compound of VA ester derivatives was derivative 5 with a C4 straight alkyl chain, and derivative 5 exhibited approximately three-fold greater inhibitory activity than that of 1. Moreover, we designed 8 types of analogs based on 5, and we found that the minimum structure for potent degranulation inhibitory activity requires direct connection of the butyl ester moiety on the benzene ring and at least one hydroxyl group on the benzene ring. Butyl meta or para hydroxyl benzoate (10 or 11) has a simpler structure than that of 5 and exhibited more potent degranulation inhibitory activity than that of 5. PMID:27324979

  6. Antigen antibody interactions

    CERN Document Server

    DeLisi, Charles

    1976-01-01

    1. 1 Organization of the Immune System One of the most important survival mechanisms of vertebrates is their ability to recognize and respond to the onslaught of pathogenic microbes to which they are conti- ously exposed. The collection of host cells and molecules involved in this recognition­ 12 response function constitutes its immune system. In man, it comprises about 10 cells 20 (lymphocytes) and 10 molecules (immunoglobulins). Its ontogenic development is c- strained by the requirement that it be capable of responding to an almost limitless variety of molecular configurations on foreign substances, while simultaneously remaining inert to those on self components. It has thus evolved to discriminate, with exquisite precision, between molecular patterns. The foreign substances which induce a response, called antigens, are typically large molecules such as proteins and polysaccharides. The portions of these with which immunoglobulins interact are called epitopes or determinants. A typical protein epitope m...

  7. Molecular mimics of the tumour antigen MUC1.

    Directory of Open Access Journals (Sweden)

    Tharappel C James

    Full Text Available A key requirement for the development of cancer immunotherapy is the identification of tumour-associated antigens that are differentially or exclusively expressed on the tumour and recognized by the host immune system. However, immune responses to such antigens are often muted or lacking due to the antigens being recognized as "self", and further complicated by the tumour environment and regulation of immune cells within. In an effort to circumvent the lack of immune responses to tumour antigens, we have devised a strategy to develop potential synthetic immunogens. The strategy, termed mirror image phage display, is based on the concept of molecular mimicry as demonstrated by the idiotype/anti-idiotype paradigm in the immune system. Here as 'proof of principle' we have selected molecular mimics of the well-characterised tumour associated antigen, the human mucin1 protein (MUC1 from two different peptide phage display libraries. The putative mimics were compared in structure and function to that of the native antigen. Our results demonstrate that several of the mimic peptides display T-cell stimulation activity in vitro when presented by matured dendritic cells. The mimic peptides and the native MUC1 antigenic epitopes can cross-stimulate T-cells. The data also indicate that sequence homology and/or chemical properties to the original epitope are not the sole determining factors for the observed immunostimulatory activity of the mimic peptides.

  8. Active treatment of murine tumors with a highly attenuated vaccinia virus expressing the tumor associated antigen 5T4 (TroVax) is CD4+ T cell dependent and antibody mediated.

    Science.gov (United States)

    Harrop, Richard; Ryan, Matthew G; Myers, Kevin A; Redchenko, Irina; Kingsman, Susan M; Carroll, Miles W

    2006-09-01

    5T4 is a tumor associated antigen that is expressed on the surface of a wide spectrum of human adenocarcinomas. The highly attenuated virus, modified vaccinia Ankara, has been engineered to express human 5T4 (h5T4). In a pre-clinical murine model, the recombinant virus (TroVax) induces protection against challenge with CT26-h5T4 (a syngeneic tumor line expressing h5T4). Anti-tumor activity is long lived, with protection still evident 6 months after the final vaccination. In a therapeutic setting, injection of mice with TroVax results in a reduction in tumor burden of >90%. Depletion of CD8+ T cells has no effect upon therapy in the active treatment model, whereas depletion of CD4+ T cells completely abrogates anti-tumor activity. In a prophylactic setting, depletion of CD4+ and CD8+ T cells after the induction of a h5T4 immune response has no deleterious effect on protection following challenge with CT26-h5T4. In light of these studies, the role of antibodies in protection against tumor challenge was investigated. 5T4 specific polyclonal serum decreased tumor burden by approximately 70%. Thus, we conclude that CD4+ T cells are essential for the induction of a protective immune response and that antibodies are the likely effector moiety in this xenogeneic murine tumor model. PMID:16311730

  9. COLONOSCOPY AND CARCINOEMBRYONIC ANTIGEN VARIATIONS

    Directory of Open Access Journals (Sweden)

    Rita G SOUSA

    2014-03-01

    Full Text Available Context Colonoscopy is essential for synchronous and metachronous cancer detection. Carcinoembryonic antigen is a colorectal cancer tumor marker, important as a follow-up tool in patients with previous colorectal cancer. False-positive carcinoembryonic antigen elevation results in multiples exams and in patient anxiety. In literature, there is reference to transient carcinoembryonic antigen increase with colonoscopy. Objective To evaluate the influence of bowel preparation and colonoscopy in carcinoembryonic antigen blood levels. Methods We prospectively studied subjects that underwent routine colonoscopy in our institution. Blood samples were collected (1 before bowel cleaning, (2 before colonoscopy and (3 immediately after colonoscopy. Blood carcinoembryonic antigen levels were determined by “Sandwich” immunoassay. The statistical methods used were the paired t-test and ANOVA. Results Thirty-seven patients (22M/15F were included; age range 28-84 (mean 56 years. Mean carcinoembryonic antigen values were 1.9, 2 and 1.8 for (1, (2 and (3, respectively. An increase in value (2 compared with (1 was observed in 20/37 patients (P = 0.018, mainly in younger patients and in patients requiring more endoluminal interventions. In 29/37 patients, the CEA value decreased from (2 to (3 (P = 1.3x10-7. Conclusions A trend for carcinoembryonic antigen increase after bowel cleaning was observed, especially in younger patients and in patients with more endoluminal interventions, but without clinical meaning.

  10. Circulating precursor CCR7(lo)PD-1(hi) CXCR5⁺ CD4⁺ T cells indicate Tfh cell activity and promote antibody responses upon antigen reexposure.

    Science.gov (United States)

    He, Jing; Tsai, Louis M; Leong, Yew Ann; Hu, Xin; Ma, Cindy S; Chevalier, Nina; Sun, Xiaolin; Vandenberg, Kirsten; Rockman, Steve; Ding, Yan; Zhu, Lei; Wei, Wei; Wang, Changqi; Karnowski, Alexander; Belz, Gabrielle T; Ghali, Joanna R; Cook, Matthew C; Riminton, D Sean; Veillette, André; Schwartzberg, Pamela L; Mackay, Fabienne; Brink, Robert; Tangye, Stuart G; Vinuesa, Carola G; Mackay, Charles R; Li, Zhanguo; Yu, Di

    2013-10-17

    Follicular B helper T (Tfh) cells support high affinity and long-term antibody responses. Here we found that within circulating CXCR5⁺ CD4⁺ T cells in humans and mice, the CCR7(lo)PD-1(hi) subset has a partial Tfh effector phenotype, whereas CCR7(hi)PD-1(lo) cells have a resting phenotype. The circulating CCR7(lo)PD-1(hi) subset was indicative of active Tfh differentiation in lymphoid organs and correlated with clinical indices in autoimmune diseases. Thus the CCR7(lo)PD-1(hi) subset provides a biomarker to monitor protective antibody responses during infection or vaccination and pathogenic antibody responses in autoimmune diseases. Differentiation of both CCR7(hi)PD-1(lo) and CCR7(lo)PD-1(hi) subsets required ICOS and BCL6, but not SAP, suggesting that circulating CXCR5⁺ helper T cells are primarily generated before germinal centers. Upon antigen reencounter, CCR7(lo)PD-1(hi) CXCR5⁺ precursors rapidly differentiate into mature Tfh cells to promote antibody responses. Therefore, circulating CCR7(lo)PD-1(hi) CXCR5⁺ CD4⁺ T cells are generated during active Tfh differentiation and represent a new mechanism of immunological early memory.

  11. Modulation of Th1/Th2 Immune Responses by Killed Propionibacterium acnes and Its Soluble Polysaccharide Fraction in a Type I Hypersensitivity Murine Model: Induction of Different Activation Status of Antigen-Presenting Cells

    Science.gov (United States)

    Mussalem, Juliana Sekeres; Ishimura, Mayari Eika; Longo-Maugéri, Ieda Maria

    2015-01-01

    Propionibacterium acnes (P. acnes) is a gram-positive anaerobic bacillus present in normal human skin microbiota, which exerts important immunomodulatory effects, when used as heat- or phenol-killed suspensions. We previously demonstrated that heat-killed P. acnes or its soluble polysaccharide (PS), extracted from the bacterium cell wall, suppressed or potentiated the Th2 response to ovalbumin (OVA) in an immediate hypersensitivity model, depending on the treatment protocol. Herein, we investigated the mechanisms responsible for these effects, using the same model and focusing on the activation status of antigen-presenting cells (APCs). We verified that higher numbers of APCs expressing costimulatory molecules and higher expression levels of these molecules are probably related to potentiation of the Th2 response to OVA induced by P. acnes or PS, while higher expression of toll-like receptors (TLRs) seems to be related to Th2 suppression. In vitro cytokines production in cocultures of dendritic cells and T lymphocytes indicated that P. acnes and PS seem to perform their effects by acting directly on APCs. Our data suggest that P. acnes and PS directly act on APCs, modulating the expression of costimulatory molecules and TLRs, and these differently activated APCs drive distinct T helper patterns to OVA in our model. PMID:25973430

  12. Antigen-specific helper activity in serum of mice primed with sheep red cells I. Definition of the test system and comparison with other systems

    NARCIS (Netherlands)

    Dijk, H. van; Rademaker, P.M.; Slotboom, A.; Willers, J.M.

    1980-01-01

    An adoptive transfer system is described to measure serum helper activity in the primary antibody response to sheep red blood cells (SRBC). Mice injected with a high dose of cyclophosphamide and reconstituted with rabbit anti-thymocyte serum-treated spleen cells were used as recipients. Serum obtain

  13. Coexpressed Catalase Protects Chimeric Antigen Receptor-Redirected T Cells as well as Bystander Cells from Oxidative Stress-Induced Loss of Antitumor Activity.

    Science.gov (United States)

    Ligtenberg, Maarten A; Mougiakakos, Dimitrios; Mukhopadhyay, Madhura; Witt, Kristina; Lladser, Alvaro; Chmielewski, Markus; Riet, Tobias; Abken, Hinrich; Kiessling, Rolf

    2016-01-15

    Treatment of cancer patients by adoptive T cell therapy has yielded promising results. In solid tumors, however, T cells encounter a hostile environment, in particular with increased inflammatory activity as a hallmark of the tumor milieu that goes along with abundant reactive oxygen species (ROS) that substantially impair antitumor activity. We present a strategy to render antitumor T cells more resilient toward ROS by coexpressing catalase along with a tumor specific chimeric Ag receptor (CAR) to increase their antioxidative capacity by metabolizing H2O2. In fact, T cells engineered with a bicistronic vector that concurrently expresses catalase, along with the CAR coexpressing catalase (CAR-CAT), performed superior over CAR T cells as they showed increased levels of intracellular catalase and had a reduced oxidative state with less ROS accumulation in both the basal state and upon activation while maintaining their antitumor activity despite high H2O2 levels. Moreover, CAR-CAT T cells exerted a substantial bystander protection of nontransfected immune effector cells as measured by CD3ζ chain expression in bystander T cells even in the presence of high H2O2 concentrations. Bystander NK cells, otherwise ROS sensitive, efficiently eliminate their K562 target cells under H2O2-induced oxidative stress when admixed with CAR-CAT T cells. This approach represents a novel means for protecting tumor-infiltrating cells from tumor-associated oxidative stress-mediated repression.

  14. Dichotomy of the T cell response to Leishmania antigens in patients suffering from cutaneous leishmaniasis; absence or scarcity of Th1 activity is associated with severe infections

    DEFF Research Database (Denmark)

    Gaafar, A; Kharazmi, A; Ismail, A;

    1995-01-01

    The T cell response was studied in 25 patients suffering from cutaneous leishmaniasis caused by Leishmania major with severe (n = 10) and mild (n = 15) disease manifestations. Peripheral blood mononuclear cells (PBMC) from the patients were activated by sonicates of Leishmania promastigotes (LMP...

  15. The prognostic value of oncogenic antigen 519 (OA-519) expression and proliferative activity detected by antibody MIB-1 in node-negative breast cancer

    DEFF Research Database (Denmark)

    Jensen, V; Ladekarl, M; Holm-Nielsen, P;

    1995-01-01

    of invasion of skin or deep fascia (= T1N0M0 and T2N0M0). The median follow-up time was 104 months (range 5-143 months). Immunohistochemical analysis of OA-519 expression was performed on formalin-fixed, paraffin-embedded tissue. The proliferative activity was estimated using a Ki-67 equivalent monoclonal...

  16. Minocycline modulates antigen-specific CTL activity through inactivation of mononuclear phagocytes in patients with HTLV-I associated neurologic disease

    Directory of Open Access Journals (Sweden)

    Enose-Akahata Yoshimi

    2012-02-01

    Full Text Available Abstract Background The activation of mononuclear phagocytes (MPs, including monocytes, macrophages and dendritic cells, contributes to central nervous system inflammation in various neurological diseases. In HTLV-I-associated myelopathy/tropical spastic paraparesis (HAM/TSP, MPs are reservoirs of HTLV-I, and induce proinflammatory cytokines and excess T cell responses. The virus-infected or activated MPs may play a role in immuneregulation and disease progression in patients with HTLV-I-associated neurological diseases. Results Phenotypic analysis of CD14+ monocytes in HAM/TSP patients demonstrated high expression of CX3CR1 and HLA-DR in CD14lowCD16+ monocytes, compared to healthy normal donors (NDs and asymptomatic carriers (ACs, and the production of TNF-α and IL-1β in cultured CD14+ cells of HAM/TSP patients. CD14+ cells of HAM/TSP patients also showed acceleration of HTLV-I Tax expression in CD4+ T cells. Minocycline, an inhibitor of activated MPs, decreased TNF-α expression in CD14+ cells and IL-1β release in PBMCs of HAM/TSP patients. Minocycline significantly inhibited spontaneous lymphoproliferation and degranulation/IFN-γ expression in CD8+ T cells of HAM/TSP patients. Treatment of minocycline also inhibited IFN-γ expression in CD8+ T cells of HAM/TSP patients after Tax11-19 stimulation and downregulated MHC class I expression in CD14+ cells. Conclusion These results demonstrate that minocycline directly inhibits the activated MPs and that the downregulation of MP function can modulate CD8+ T cells function in HAM/TSP patients. It is suggested that activated MPs may be a therapeutic target for clinical intervention in HAM/TSP.

  17. Classification of human leukocyte antigen (HLA) supertypes

    DEFF Research Database (Denmark)

    Wang, Mingjun; Claesson, Mogens H

    2014-01-01

    Identification of new antigenic peptides, derived from infectious agents or cancer cells, which bind to human leukocyte antigen (HLA) class I and II molecules, is of importance for the development of new effective vaccines capable of activating the cellular arm of the immune response. However, the...... barrier to the development of peptide-based vaccines with maximum population coverage is that the restricting HLA genes are extremely polymorphic resulting in a vast diversity of peptide-binding HLA specificities and a low population coverage for any given peptide-HLA specificity. One way to reduce this...... complexity is to group thousands of different HLA molecules into several so-called HLA supertypes: a classification that refers to a group of HLA alleles with largely overlapping peptide binding specificities. In this chapter, we focus on the state-of-the-art classification of HLA supertypes including HLA...

  18. Antibody-antigen-adjuvant conjugates enable co-delivery of antigen and adjuvant to dendritic cells in cis but only have partial targeting specificity.

    Directory of Open Access Journals (Sweden)

    Martin Kreutz

    Full Text Available Antibody-antigen conjugates, which promote antigen-presentation by dendritic cells (DC by means of targeted delivery of antigen to particular DC subsets, represent a powerful vaccination approach. To ensure immunity rather than tolerance induction the co-administration of a suitable adjuvant is paramount. However, co-administration of unlinked adjuvant cannot ensure that all cells targeted by the antibody conjugates are appropriately activated. Furthermore, antigen-presenting cells (APC that do not present the desired antigen are equally strongly activated and could prime undesired responses against self-antigens. We, therefore, were interested in exploring targeted co-delivery of antigen and adjuvant in cis in form of antibody-antigen-adjuvant conjugates for the induction of anti-tumour immunity. In this study, we report on the assembly and characterization of conjugates consisting of DEC205-specific antibody, the model antigen ovalbumin (OVA and CpG oligodeoxynucleotides (ODN. We show that such conjugates are more potent at inducing cytotoxic T lymphocyte (CTL responses than control conjugates mixed with soluble CpG. However, our study also reveals that the nucleic acid moiety of such antibody-antigen-adjuvant conjugates alters their binding and uptake and allows delivery of the antigen and the adjuvant to cells partially independently of DEC205. Nevertheless, antibody-antigen-adjuvant conjugates are superior to antibody-free antigen-adjuvant conjugates in priming CTL responses and efficiently induce anti-tumour immunity in the murine B16 pseudo-metastasis model. A better understanding of the role of the antibody moiety is required to inform future conjugate vaccination strategies for efficient induction of anti-tumour responses.

  19. Activation of antigen-exposed iMC-DCs at the "right place" and "right time" promotes potent anti-tumor immunity.

    Science.gov (United States)

    Spencer, David M

    2012-05-01

    To better control the "licensing" of pro-Th1 dendritic cells (DCs), Spencer and colleagues have developed a synthetic ligand-inducible chimeric receptor, iMyD88/CD40 (iMC), incorporating synergistic Toll-like receptor (TLR) and costimulatory signaling elements, permitting DC regulation in vivo within the context of an immunological synapse. This novel technology results in potent anti-cancer activity.

  20. DNA binding and antigene activity of a daunomycin-conjugated triplex-forming oligonucleotide targeting the P2 promoter of the human c-myc gene

    OpenAIRE

    Giuseppina M. Carbone; McGuffie, Eileen; Napoli, Sara; Flanagan, Courtney E.; Dembech, Chiara; Negri, Umberto; Arcamone, Federico; Capobianco, Massimo L.; Catapano, Carlo V

    2004-01-01

    Triplex-forming oligonucleotides (TFO) that bind DNA in a sequence-specific manner might be used as selective repressors of gene expression and gene-targeted therapeutics. However, many factors, including instability of triple helical complexes in cells, limit the efficacy of this approach. In the present study, we tested whether covalent linkage of a TFO to daunomycin, which is a potent DNA-intercalating agent and anticancer drug, could increase stability of the triple helix and activity of ...

  1. ISCOMATRIX Adjuvant Combines Immune Activation with Antigen Delivery to Dendritic Cells In Vivo Leading to Effective Cross-Priming of CD8+ T Cells

    Science.gov (United States)

    Duewell, Peter; Kisser, Ulrich; Heckelsmiller, Klaus; Hoves, Sabine; Stoitzner, Patrizia; Koernig, Sandra; Morelli, Adriana B.; Clausen, Björn E.; Dauer, Marc; Eigler, Andreas; Anz, David; Bourquin, Carole; Maraskovsky, Eugene; Endres, Stefan; Schnurr, Max

    2014-01-01

    Cancer vaccines aim to induce CTL responses against tumors. Challenges for vaccine design are targeting Ag to dendritic cells (DCs) in vivo, facilitating cross-presentation, and conditioning the microenvironment for Th1 type immune responses. In this study, we report that ISCOM vaccines, which consist of ISCOMATRIX adjuvant and protein Ag, meet these challenges. Subcutaneous injection of an ISCOM vaccine in mice led to a substantial influx and activation of innate and adaptive immune effector cells in vaccine site-draining lymph nodes (VDLNs) as well as IFN-γ production by NK and NKT cells. Moreover, an ISCOM vaccine containing the model Ag OVA (OVA/ISCOM vaccine) was efficiently taken up by CD8α+ DCs in VDLNs and induced their maturation and IL-12 production. Adoptive transfer of transgenic OT-I T cells revealed highly efficient cross-presentation of the OVA/ISCOM vaccine in vivo, whereas cross-presentation of soluble OVA was poor even at a 100-fold higher concentration. Cross-presenting activity was restricted to CD8α+ DCs in VDLNs, whereas Langerin+ DCs and CD8α− DCs were dispensable. Remarkably, compared with other adjuvant systems, the OVA/ISCOM vaccine induced a high frequency of OVA-specific CTLs capable of tumor cell killing in different tumor models. Thus, ISCOM vaccines combine potent immune activation with Ag delivery to CD8α+ DCs in vivo for efficient induction of CTL responses. PMID:21613613

  2. Activation of human antigen-presenting cells by the mycobacterial cord factor and its glycolipid adjuvant analogue trehalose-6,6’-dibehenate

    OpenAIRE

    Ostrop, Jenny

    2015-01-01

    The mycobacterial cord factor trehalose-6,6’-dimycolate (TDM) is an abundant cell wall glycolipid of Mycobacterium tuberculosis and other mycobacteria. It causes inflammation and adjuvanticity, but it is also a major virulence factor of M. tuberculosis. Its synthetic analogue trehalose-6,6’-dibehenate (TDB) has robust adjuvant activity and induces a Th1/Th17 T cell response in animal models. The TDB-containing liposomal adjuvant formulation Caf01 has entered phase I clinical studies in humans...

  3. Control of T cell antigen reactivity via programmed TCR downregulation.

    Science.gov (United States)

    Gallegos, Alena M; Xiong, Huizhong; Leiner, Ingrid M; Sušac, Bože; Glickman, Michael S; Pamer, Eric G; van Heijst, Jeroen W J

    2016-04-01

    The T cell antigen receptor (TCR) is unique in that its affinity for ligand is unknown before encounter and can vary by orders of magnitude. How the immune system regulates individual T cells that display very different reactivity to antigen remains unclear. Here we found that activated CD4(+) T cells, at the peak of clonal expansion, persistently downregulated their TCR expression in proportion to the strength of the initial antigen recognition. This programmed response increased the threshold for cytokine production and recall proliferation in a clone-specific manner and ultimately excluded clones with the highest antigen reactivity. Thus, programmed downregulation of TCR expression represents a negative feedback mechanism for constraining T cell effector function with a suitable time delay to thereby allow pathogen control while avoiding excess inflammatory damage. PMID:26901151

  4. The Antigen Presenting Cells Instruct Plasma Cell Differentiation

    Directory of Open Access Journals (Sweden)

    Wei eXu

    2014-01-01

    Full Text Available The professional antigen presenting cells (APCs, including many subsets of dendritic cells and macrophages, not only mediate prompt but nonspecific response against microbes, but also bridge the antigen-specific adaptive immune response through antigen presentation. In the latter, typically activated B cells acquire cognate signals from T helper cells in the germinal center of lymphoid follicles to differentiate into plasma cells, which generate protective antibodies. Recent advances have revealed that many APC subsets provide not only signal 1 (the antigen, but also signal 2 to directly instruct the differentiation process of plasma cells in a T cell-independent manner. Herein, the different signals provided by these APC subsets to direct B cell proliferation, survival, class switching and terminal differentiation are discussed. We furthermore propose that the next generation of vaccines for boosting antibody response could be designed by targeting APCs.

  5. Immune response of sheep to Haemonchus contortus: serum antibodies against cross reacting antigens between parasites.

    Science.gov (United States)

    Charley, J; Bourdieu, C; Luffau, G; Pery, P

    1981-01-01

    Normal and H. contortus infected sera were studied by ELISA technique against different stages of the parasites. In all cases antibody activity was detected. This activity in serum is partially eliminated after absorption with an adult worm extract of N brasiliensis. N. brasiliensis and H. contortus antigens were analysed by TCIEP with a rabbit anti-N. brasiliensis serum to examine shared antigens of H. contortus. A minimum of seven cross reacting antigens were detected. H. contortus adult worm extract was absorbed by the rabbit anti-N. brasiliensis serum. After absorption all cross reacting antigens were removed but at least one antigen reacting with a rabbit serum anti-H. Contortus is maintained. When this antigen is tested in elisa technique only a weak antibody activity is found in normal serum.

  6. Activation of human monocytes by streptococcal rhamnose glucose polymers is mediated by CD14 antigen, and mannan binding protein inhibits TNF-alpha release.

    Science.gov (United States)

    Soell, M; Lett, E; Holveck, F; Schöller, M; Wachsmann, D; Klein, J P

    1995-01-15

    The present work was initiated to define mechanisms that account for the binding on human monocytes of streptococcal cell wall polysaccharides formed by rhamnose glucose polymers (RGPs), and subsequent stimulatory activities. We show here that RGPs bind to and stimulate human monocytes to produce TNF-alpha in a dose-dependent manner. To detect cell surface RGPs binding proteins, intact monocytes were biotinylated before lysis with Nonidet P-40 and solubilized proteins were incubated with RGPs Affi-Prep beads. One major membrane protein of 55 kDa was specifically detected and identified as CD14 because it reacted with anti-CD14 mAbs. Furthermore, anti-CD14 mAbs were able to perform a dose-dependent inhibition of RGPs binding, and suppressed TNF-alpha release from RGPs-stimulated monocytes. Moreover, we demonstrated that RGPs also bind to CD11b; however, this binding is not implicated in synthesis of TNF-alpha. Interestingly, RGPs binding to monocytes was enhanced by human normal serum (HNS) whereas HNS inhibits the TNF-alpha-stimulating activity of RGPs. Western blotting analysis of HNS proteins purified on RGPs Affi-prep beads revealed three specific bands of 75, 55, and 32 kDa reactive with anti-C3 Abs, anti-CD14 mAbs (TUK4), and anti-human mannan binding protein (hMBP)-derived peptide IgG, respectively. These results suggest that C3, soluble CD14, and hMBP form complexes that are probably active in enhancing the binding of RGPs to monocytes. Additional studies have shown that hMBP that recognizes RGPs prevents, unlike the LPS binding protein, TNF-alpha release by inhibiting the binding of RGPs to CD14 Ag. By incubating cells with a constant amount of RGPs-hMBP complexes in the presence or absence of increasing concentrations of C1q, we also demonstrated that C1q receptor mediates the binding and probably the uptake of RGPs-hMBP complexes by human monocytes. PMID:7529289

  7. Proliferating cell nuclear antigen (PCNA) interacts with a meiosis-specific RecA homologues, Lim15/Dmc1, but does not stimulate its strand transfer activity

    International Nuclear Information System (INIS)

    PCNA is a multi-functional protein that is involved in various nuclear events. Here we show that PCNA participates in events occurring during early meiotic prophase. Analysis of protein-protein interactions using surface plasmon resonance indicates that Coprinus cinereus PCNA (CoPCNA) specifically interacts with a meiotic specific RecA-like factor, C. cinereus Lim15/Dmc1 (CoLim15) in vitro. The binding efficiency increases with addition of Mg2+ ions, while ATP inhibits the interaction. Co-immunoprecipitation experiments indicate that the CoLim15 protein interacts with the CoPCNA protein in vitro and in the cell extracts. Despite the interaction between these two factors, no enhancement of CoLim15-dependent strand transfer activity by CoPCNA was found in vitro. We propose that the interaction between Lim15/Dmc1 and PCNA mediates the recombination-associated DNA synthesis during meiosis

  8. Continuous activation of the CD122/STAT-5 signaling pathway during selection of antigen-specific regulatory T cells in the murine thymus.

    Directory of Open Access Journals (Sweden)

    Jérémie D Goldstein

    Full Text Available Signaling events affecting thymic selection of un-manipulated polyclonal natural CD25(+foxp3(+ regulatory T cells (nTreg have not been established ex vivo. Here, we report a higher frequency of phosphorylated STAT-5 (pSTAT-5 in nTreg cells in the adult murine thymus and to a lesser extent in the periphery, compared to other CD4(+CD8(- subsets. In the neonatal thymus, the numbers of pSTAT-5(+ cells in CD25(+foxp3(- and nTreg cells increased in parallel, suggesting that pSTAT-5(+CD25(+foxp3(- cells might represent the precursors of foxp3(+ regulatory T cells. This "specific" pSTAT-5 expression detected in nTreg cells ex vivo was likely due to a very recent signal given by IL-2/IL-15 cytokines in vivo since (i it disappeared rapidly if cells were left unstimulated in vitro and (ii was also observed if total thymocytes were stimulated in vitro with saturating amounts of IL-2 and/or IL-15 but not IL-7. Interestingly, STAT-5 activation upon IL-2 stimulation correlated better with foxp3 and CD122 than with CD25 expression. Finally, we show that expression of an endogenous superantigen strongly affected the early Treg cell repertoire but not the proportion of pSTAT-5(+ cells within this repertoire. Our results reveal that continuous activation of the CD122/STAT-5 signaling pathway characterize regulatory lineage differentiation in the murine thymus.

  9. Icariin inhibits oxidized low-density lipoprotein-induced proliferation of vascular smooth muscle cells by suppressing activation of extracellular signal-regulated kinase 1/2 and expression of proliferating cell nuclear antigen.

    Science.gov (United States)

    Hu, Yanwu; Liu, Kai; Yan, Mengtong; Zhang, Yang; Wang, Yadi; Ren, Liqun

    2016-03-01

    Icariin, a flavonoid isolated from the traditional Chinese herbal medicine Epimedium brevicornum Maxim, has been shown to possess anti-inflammatory, anti‑oxidant and anti-atherosclerotic activities in vivo and in vitro. The aim of the present study was to investigate the effects of icariin on oxidized low‑density lipoprotein (ox-LDL)-induced proliferation of vascular smooth muscle cells (VSMCs) and the possible underlying mechanism. VSMCs were cultured and pre‑treated with various concentrations of icariin (0, 10, 20 or 40 µm) prior to stimulation by ox‑LDL (50 µg/ml). Cell proliferation was evaluated by an MTT assay. Flow cytometry was used to study the influence of icariin on the cell cycle. Proliferating cell nuclear antigen (PCNA) expression and phosphorylation levels of extracellular signal-regulated kinase (ERK)1/2 were detected by western blot analysis. The results indicated that icariin significantly inhibited ox‑LDL‑induced proliferation of VSMCs and phosphorylation of ERK1/2. Furthermore, icariin also blocked the ox‑LDL‑induced cell‑cycle progression at G1/S‑interphase and downregulated the expression of PCNA in VSMCs. In conclusion, the present study indicated for the first time that icariin reduced the amount of ox‑LDL‑induced proliferation of VSMCs through suppression of PCNA expression and inactivation of ERK1/2.

  10. Mouse bone marrow-derived dendritic cells can phagocytize the Sporothrix schenckii, and mature and activate the immune response by secreting interleukin-12 and presenting antigens to T lymphocytes.

    Science.gov (United States)

    Kusuhara, Masahiro; Qian, Hua; Li, Xiaoguang; Tsuruta, Daisuke; Tsuchisaka, Atsunari; Ishii, Norito; Ohata, Chika; Furumura, Minao; Hashimoto, Takashi

    2014-05-01

    In sporotrichosis, dermal dendritic cells were considered to participate in induction of the immune responses against Sporothrix schenckii infection. However, it is still unclear whether and how dermal dendritic cells were involved in the progress. To clarify the pathogenic role of dermal dendritic cells (DC) in sporotrichosis, we examined the phagocytosis, maturation stages, cytokine production and antigen-presenting ability of mouse bone marrow-derived DC after stimulation with S. schenckii. By analysis of flow cytometry, electron microscope and confocal microscope, mouse bone marrow-derived DC were proved to be able to phagocytize the S. schenckii. The increased expression of CD40, CD80 and CD86 on the surface of S. schenckii-pulsed mouse bone marrow-derived DC was detected by flow cytometer, indicating that the S. schenckii-pulsed mouse bone marrow-derived DC underwent the maturation program. The secretory enhancement of interleukin (IL)-12, but not IL-4, was found in S. schenckii-pulsed mouse bone marrow-derived DC, suggesting the possible activation of T-helper 1 prone immune responses. Furthermore, S. schenckii-pulsed mouse bone marrow-derived DC were demonstrated to be capable of inducing the proliferation of T lymphocytes from BALB/c mice that were pre-sensitized with S. schenckii. Together, all the results implied that dermal DC may participate in the induction of immune responses against S. schenckii infection in sporotrichosis.

  11. The Impacts of Helicobacter Pylori Antigen Positivity on Ankylosing Spondylitis

    Directory of Open Access Journals (Sweden)

    Esra Erkol Ižnal

    2014-12-01

    Full Text Available Aim: We aimed to clarify the impacts of H. pylori infection on disease activity and clinical findings of AS. Material and Method: Forty-eight patients with AS were included in this study. The demographic data including age, sex, durations of the disease and medication of the patients were recorded. The laboratory analysis comprised Erythrocyte sedimentation rate (ESR, C-reactive protein (CRP and H. pylori antigen determination in gaita. The disease activity, functional disability and clinical status were assessed using the Bath Ankylosing Spondylitis Disease Activity Index (BASDAI, The Bath Ankylosing Spondylitis Functional index (BASFI and The Bath Ankylosing Spondylitis Metrology Index (BASMI respectively. We divided patients according to H. pylori antigen positivity in gaita as H. pylori positive and negative patients.Results: The mean age of patients was 41.9 11.8. CRP levels were slightly but not significantly higher in patients with positive H. pylori antigen compared to those in patients without H. pylori antigen in gaita (p=0.08. There was no significant difference in terms of ESR levels, BASDAI, BASFI and BASMI scores in patients with positive H. pylori antigen compared to those in patients with negative H. pylori antigen in gaita (p-values were >0.05 for all. In regression model BASDAI score was found to have no relationship with H. pylori antigen positivity, ESR and CRP levels (p-values were >0.05 for all. Discussion: H. pylori seemed to have probable impacts on the disease activity of AS. Studies with greater patient population and longer follow-up periods are warranted to enlighten this issue.

  12. Cytokine-Independent Detection of Antigen-Specific Germinal Center T Follicular Helper Cells in Immunized Nonhuman Primates Using a Live Cell Activation-Induced Marker Technique.

    Science.gov (United States)

    Havenar-Daughton, Colin; Reiss, Samantha M; Carnathan, Diane G; Wu, Jennifer E; Kendric, Kayla; Torrents de la Peña, Alba; Kasturi, Sudhir Pai; Dan, Jennifer M; Bothwell, Marcella; Sanders, Rogier W; Pulendran, Bali; Silvestri, Guido; Crotty, Shane

    2016-08-01

    A range of current candidate AIDS vaccine regimens are focused on generating protective HIV-neutralizing Ab responses. Many of these efforts rely on the rhesus macaque animal model. Understanding how protective Ab responses develop and how to increase their efficacy are both major knowledge gaps. Germinal centers (GCs) are the engines of Ab affinity maturation. GC T follicular helper (Tfh) CD4 T cells are required for GCs. Studying vaccine-specific GC Tfh cells after protein immunizations has been challenging, as Ag-specific GC Tfh cells are difficult to identify by conventional intracellular cytokine staining. Cytokine production by GC Tfh cells may be intrinsically limited in comparison with other Th effector cells, as the biological role of a GC Tfh cell is to provide help to individual B cells within the GC, rather than secreting large amounts of cytokines bathing a tissue. To test this idea, we developed a cytokine-independent method to identify Ag-specific GC Tfh cells. RNA sequencing was performed using TCR-stimulated GC Tfh cells to identify candidate markers. Validation experiments determined CD25 (IL-2Rα) and OX40 to be highly upregulated activation-induced markers (AIM) on the surface of GC Tfh cells after stimulation. In comparison with intracellular cytokine staining, the AIM assay identified >10-fold more Ag-specific GC Tfh cells in HIV Env protein-immunized macaques (BG505 SOSIP). CD4 T cells in blood were also studied. In summary, AIM demonstrates that Ag-specific GC Tfh cells are intrinsically stingy producers of cytokines, which is likely an essential part of their biological function. PMID:27335502

  13. Construction and Screening of Antigen Targeted Immune Yeast Surface Display Antibody Libraries

    Energy Technology Data Exchange (ETDEWEB)

    Miller, Keith D.; Pefaur, Noah B.; Baird, Cheryl L.

    2008-07-01

    These protocols describe a yeast surface display-based process for the rapid selection of antibodies from immunized mice, eliminating the need for creating and screening hybridoma fusions. A yeast surface display library of single-chain antibody fragments (scFvs) is created from antigen-binding B cells from the splenocytes of immunized mice. The antigen targeted library is then screened for antigen specific scFv by magnetic activated cell sorting (MACS) and fluorescence activated cell sorting (FACS). Library construction and screening can be accomplished in as little as 2 weeks resulting in a panel of scFvs specific for the target antigen.

  14. Double-antibody radioimmunoassay for factor VIII-related antigen

    International Nuclear Information System (INIS)

    A plasma protein required for the support of ristocetin-induced platelet aggregation was isolated from antihemophilic factor concentrate and radiolabeled with 125I. A double-antibody radioimmunoassay was developed, with use of specific rabbit anti-VIII related antigen serum and goat anti-rabbit globulin. The assay is sensitive, reproducible, and technically simple to perform. Values obtained in normal subjects ranged from 0.65 to 1.53 units, similar to our normal range for VIII coagulant activity (0.67 to 1.43 units). However, normal or increased values of VIII-related antigen were observed in VIII coagulant-deficient hemophiliacs. Also, concentrations of VIII-related antigen significantly exceeded coagulant concentrations in several patients with liver disease or disseminated intravascular coagulation, or both. Of a broad selection of congenital coagulation disorders examined, only patients with von Willebrand's disease had decreased VIII-related antigen concentrations, and these corresponded to the lowered concentration of ristocetin cofactor in the patients. In three transfused patients, VIII-related antigen values correlated with the concentration of the cofactor. Our results suggest that the radioimmunoassay of VIII-related antigen is a simple and valuable adjunct in the study of patients with clotting abnormalities

  15. The role of FcRn in antigen presentation

    Directory of Open Access Journals (Sweden)

    Kristi eBaker

    2014-08-01

    Full Text Available Immunoglobulins are unique molecules capable of simultaneously recognizing a diverse array of antigens and themselves being recognized by a broad array of receptors. The abundance specifically of the IgG subclass and the variety of signaling receptors to which it binds render this an important immunomodulatory molecule. In addition to the classical Fcγ receptors (FcγR which bind IgG at the cell surface, the neonatal Fc receptor (FcRn is a lifelong resident of the endolysosomal system of most hematopoietic cells where it determines the intracellular fate of both IgG and IgG-containing immune complexes (IgG IC. Crosslinking of FcRn by multivalent IgG IC within antigen presenting cells such as dendritic cells (DC initiates specific mechanisms which result in trafficking of the antigen-bearing IgG IC into compartments from which the antigen can successfully be processed into peptide epitopes compatible with loading onto both MHC class I and II molecules. In turn, this enables the synchronous activation of both CD4+ and CD8+ T cell responses against the cognate antigen, thereby bridging the gap between the humoral and cellular branches of the adaptive immune response. Critically, FcRn-driven T cell priming is efficient at very low doses of antigen due to the exquisite sensitivity of the IgG-mediated antigen delivery system through which it operates. FcRn-mediated antigen presentation has important consequences in tissue compartments replete with IgG and serves not only to determine homeostatic immune activation at a variety of sites but also to induce inflammatory responses upon exposure to antigens perceived as foreign. Therapeutically targeting the pathway by which FcRn enables T cell activation in response to IgG IC is thus a highly attractive prospect not only for the treatment of diseases that are driven by immune complexes but also for manipulating local immune responses against defined antigens such as those present during infections and

  16. Enhancing the recognition of tumor associated antigens

    OpenAIRE

    Restifo, Nicholas P; Irvine, Kari R.; Minev, Boris R.; Taggarse, Akash S.; McFariand, Barbra J.; Wang, Michael

    1994-01-01

    Activated CD8+ T cells (TCD8+) can directly recognize malignant cells because processed fragments of tumor associated antigens (TAA), 8-10 amino acids in length and complexed with MHC class I molecules, are displayed on tumor cell surfaces. Tumor cells have been genetically modified in a variety of ways in efforts to enhance the immune recognition of TAA. An alternative strategy is the expression of TAA in recombinant or synthetic form. This has been made possible by the recent cloning of TAA...

  17. Cell Wall-Associated Protein Antigens of Streptococcus salivarius: Purification, Properties, and Function in Adherence

    Science.gov (United States)

    Weerkamp, Anton H.; Jacobs, Ton

    1982-01-01

    Three cell wall-associated protein antigens (antigens b, c, and d) were isolated from mutanolysin-solubilized cell walls of Streptococcus salivarius HB and purified to apparent homogeneity by a combination of ion-exchange chromatography, gel filtration, and immunoadsorption chromatography. Antigens b and c were also isolated from culture supernatants. Antigen b consisted of more than 80% protein and had an apparent molecular weight as determined by sodium dodecyl sulfate-polyacrylamide gel electrophoresis of 320,000. Antigen c consisted of 57% protein, about 30% neutral sugar, and about 13% amino sugar, and its glycoprotein nature was confirmed by specific staining techniques. During sodium dodecyl sulfate-polyacrylamide gel electrophoresis antigen c resolved into two or more bands, depending on the source or the isolation procedure, in the molecular weight range from 220,000 to 280,000. Antigen d consisted of 95% protein and was observed in sodium dodecyl sulfate-polyacrylamide gel electrophoresis as two bands with molecular weights of 129,000 and 121,000. Under nondenaturing conditions all three antigens had molecular weights in the range from 1 × 106 to 3 × 106 as determined by gel filtration. The amino acid compositions of antigens b, c, and d were characterized by low amounts of basic amino acids and relatively high levels of nonpolar amino acids. Among oral streptococcal species antigens b and c were virtually restricted to strains of S. salivarius and most often to serotype I strains. Antigen b was recognized as the factor that mediates coaggregation of S. salivarius with Veillonella strains. The purified protein retained its biological activity. Antigen c could be linked to functions relating to adhesion of the streptococci to host tissues on the basis of its absence in mutant strains and blocking by specific antisera. The purified molecule had no detectable biological activity. Antigen d could not be linked to an established adhesion function. Images

  18. [Farmer's lung antigens in Germany].

    Science.gov (United States)

    Sennekamp, J; Joest, M; Sander, I; Engelhart, S; Raulf-Heimsoth, M

    2012-05-01

    Recent studies suggest that besides the long-known farmer's lung antigen sources Saccharopolyspora rectivirgula (Micropolyspora faeni), Thermoactinomyces vulgaris, and Aspergillus fumigatus, additionally the mold Absidia (Lichtheimia) corymbifera as well as the bacteria Erwinia herbicola (Pantoea agglomerans) and Streptomyces albus may cause farmer's lung in Germany. In this study the sera of 64 farmers with a suspicion of farmer's lung were examined for the following further antigens: Wallemia sebi, Cladosporium herbarum, Aspergillus versicolor, and Eurotium amstelodami. Our results indicate that these molds are not frequent causes of farmer's lung in Germany. PMID:22477566

  19. [Mucose associated lymphoid tissue. Antigen presenting cells].

    Science.gov (United States)

    Luzardo-Baptista, Mario J; Luzardo, José Rafael

    2013-12-01

    We studied samples of normal and abnormal human mucosae, including oral tissue and uterine cervix, using electron microscopy. Special attention was given to the functions and mechanisms of defense carried out by the epithelial (EC) and dendritic cells (DC). Activated epithelial cells posses the capacity to uptake and process antigens, in order to present them, subsequently, to the dendritic cells. The structures and elements of the cells intervening on this function are: micropinocytic vesicles, multivesicular bodies, lysosomes, phagosomes, clathrin-covered vesicles, dense granules covered by a unit membrane, granules with onion likes leaves, microbodies, and dense granules with acid phosphatase activity. When they first arrive within the epithelial layers, the DC are clear with long cytoplasmic projections, which later become short, and the density of their cytoplasm increases. They possess mycropinocytic vesicles, some clathrine-covered vesicles, lysososmes and Birbeck granules. At this moment, they are known as Langerhans cells. EC and DC present many surface folds rich in micropynocytic vesicles. Between EC and DC there are many contacts (close junctions or tight junctions), through which antigens, phagocitized and processed by the EC, are given to the DC. These cells join them to major histocompatibility complex molecules or to other molecules with similar functions (CD1). Then the Langerhans cells travel to the lymphatic node to activate T cells and continue the immunologic task. So, in this way, both the EC and the DC are a link between the natural and the acquired immunological mechanisms. PMID:24502183

  20. Participation of L3T4 in T cell activation in the absence of class II major histocompatibility complex antigens. Inhibition by anti-L3T4 antibodies is a function both of epitope density and mode of presentation of anti-receptor antibody

    DEFF Research Database (Denmark)

    Owens, T; Fazekas de St Groth, B

    1987-01-01

    The recognition of many class II major histocompatibility complex (MHC)-associated antigens by T cells requires the participation of the L3T4 molecule. It has been proposed that this molecule acts to stabilize low affinity binding to antigen in association with MHC and thereby increases the avidity...... two monoclonal antibodies, KJ16-133.18 and F23.1, that recognize a determinant encoded by the T cell receptor V beta 8 gene family. These antibodies were used to select two clones of T cells with surface phenotype Thy-1.2+, L3T4+, Lyt-2-, KJ16-133.18+, F23.1+, IA-, IE-. One of these clones (E9.D4...... the formation of TCR complexes and so prevent activation. However, by increasing the epitope density of the activating ligand, the avidity of the T cell/ligand interaction can be increased sufficiently to prevent this disruption.(ABSTRACT TRUNCATED AT 400 WORDS)...

  1. New Concepts in Tumor Antigens: Their Significance in Future Immunotherapies for Tumors

    Institute of Scientific and Technical Information of China (English)

    Fan Yang; Xiao-Feng Yang

    2005-01-01

    The identification and molecular characterization of self-antigens expressed by human malignancies that are capable of elicitation of anti-tumor immune responses in patients have been an active field in tumor immunology.More than 2,000 tumor antigens have been identified, and most of these antigens are self-antigens. These significant progresses have led to the renaissance of tumor immunology and studies on anti-tumor immunotherapy.However, despite of the progress in the identification of self-tumor antigens, current antigen-specific immunotherapies for tumors are far less satisfied than expected, which reflects the urgent need to improve our understanding on self-tumor antigens. In order to develop more effective antigen specific anti-tumor immunotherapies and to monitor the responses to these immunotherapies in patients with tumors, many important fundamental questions need to be addressed. We propose for the first time that the studies in addressing the characteristics of self-tumor antigens and autoantigens are grouped as a new subject termed "antigenology". In this brief review, we would outline the progress in the identification of tumor antigens in solid tumors and hematologic malignancies, and overview the new concepts and principles of antigenology and their significance for future immunotherapies to these malignancies. Cellular & Molecular Immunology.

  2. Characterization of oligosaccharides from an antigenic mannan of Saccharomyces cerevisiae.

    Science.gov (United States)

    Young, M; Davies, M J; Bailey, D; Gradwell, M J; Smestad-Paulsen, B; Wold, J K; Barnes, R M; Hounsell, E F

    1998-08-01

    Mannans of the yeast Saccharomyces cerevisiae have been implicated as containing the allergens to which bakers and brewers are sensitive and also the antigen recognized by patients with Crohn's disease. A fraction of S. cerevisiae mannan, Sc500, having high affinity for antibodies in Crohn's patients has been characterized by NMR spectroscopy followed by fragmentation using alkaline elimination, partial acid hydrolysis and acetolysis. The released oligosaccharides were separated by gel filtration on a Biogel P4 column and analyzed by fluorescence labeling, HPLC and methylation analysis. The relationship between structure and antigen activity was measured by competitive ELISA. The antigenic activity of the original high molecular weight mannan could be ascribed to terminal Manalpha1-->3Manalpha1-->2 sequences which are rarely found in human glycoproteins but were over-represented in Sc500 compared to other yeast mannans.

  3. Surface membrane antigen expression changes induced in vitro by exogenous growth factors in chronic lymphocytic leukemia cells.

    Science.gov (United States)

    Vilpo, J; Hulkkonen, J; Hurme, M; Vilpo, L

    2002-09-01

    The factors determining the growth and survival of cells in B chronic lymphocytic leukemia (CLL) have remained poorly understood. We investigated the effects of optimal mitogen combinations (OMCs) on the expression of 26 surface membrane antigens among 33 CLL patients. The seven OMCs used were selected after pre-testing 14 combinations of (1) S. aureus Cowan I (SAC), (2) interleukin-2 (IL-2), (3) tumor necrosis factor alpha (TNF-alpha) and (4) 12-O-tetradecanoylphorbol 13-acetate (TPA; also known as phorbol 12-myristate 13-acetate or PMA). In flow cytometry we revealed that OMCs induced statistically highly significant upregulation of the expression of CD5, CD11c, CD19, CD22, CD23, CD25, CD38, CD40, CD45, CD45RO, CD95, CD126, CD130 and FMC7, and downregulation of CD20 and CD124 expression. Interestingly, the expression of CD27, CD45RA, CD79b, CD80, CD122 and that of the immunoglobulin gene superfamily members CD21, Ig-kappa, Ig-lambda, Ig-delta and Ig-micro were not significantly affected under similar conditions. The expression of several antigens was co-regulated, suggesting common regulatory pathways. These antigens include CD11c/CD5, CD11c/CD22, CD11c/CD126, CD11c/FMC7 as well as CD27/CD45, CD27/CD45RA and CD27/CD79b. Upregulation of surface antigen expression, induced by OMCs, should be applicable in antibody therapy in vitro and in vivo, and in negative stem cell selection for autotransplantation. Furthermore, the current strategy to enhance cell surface antigen expression may be a versatile tool to raise humoral and cell-mediated host defense against CLL cells. Upregulation of proteins mediating positive growth signals (eg CD25, CD40) and negative signals or apoptosis (eg CD95) may be used to sensitize cells to chemotherapy and programmed cell death. PMID:12200683

  4. ONCOLYTIC VIRUS-MEDIATED REVERSAL OF IMPAIRED TUMOR ANTIGEN PRESENTATION

    Directory of Open Access Journals (Sweden)

    Shashi Ashok Gujar

    2014-04-01

    Full Text Available Anti-tumor immunity can eliminate existing cancer cells and also maintain a constant surveillance against possible relapse. Such an antigen-specific adaptive response begins when tumor-specific T cells become activated. T cell activation requires two signals on antigen presenting cells (APCs: antigen presentation through MHC molecules and co-stimulation. In the absence of one or both of these signals, T cells remain inactivated or can even become tolerized. Cancer cells and their associated microenvironment strategically hinder the processing and presentation of tumor antigens and consequently prevent the development of anti-tumor immunity. Many studies, however, demonstrate that interventions that overturn tumor-associated immune evasion mechanisms can establish anti-tumor immune responses of therapeutic potential. One such intervention is oncolytic virus (OV-based anti-cancer therapy. Here we discuss how OV-induced immunological events override tumor-associated antigen presentation impairment and promote appropriate T cell:APC interaction. Detailed understanding of this phenomenon is pivotal for devising the strategies that will enhance the efficacy of OV-based anti-cancer therapy by complementing its inherent oncolytic

  5. Natural selection promotes antigenic evolvability

    NARCIS (Netherlands)

    Graves, C.J.; Ros, V.I.D.; Stevenson, B.; Sniegowski, P.D.; Brisson, D.

    2013-01-01

    The hypothesis that evolvability - the capacity to evolve by natural selection - is itself the object of natural selection is highly intriguing but remains controversial due in large part to a paucity of direct experimental evidence. The antigenic variation mechanisms of microbial pathogens provide

  6. Oncogenic cancer/testis antigens

    DEFF Research Database (Denmark)

    Gjerstorff, Morten F; Andersen, Mads H; Ditzel, Henrik J

    2015-01-01

    /testis antigens are immunogenic, highly cancer-specific, and frequently expressed in various types of cancer, which make them promising candidate targets for cancer immunotherapy, including cancer vaccination and adoptive T-cell transfer with chimeric T-cell receptors. Our current understanding of tumor...

  7. Construction and Screening of Antigen Targeted Immune Yeast Surface Display Antibody Libraries

    Energy Technology Data Exchange (ETDEWEB)

    Miller, Keith D.; Pefaur, Noah B.; Baird, Cheryl L.

    2009-08-02

    These protocols describe a yeast surface display-based process for the rapid selection of antibodies from immunized mice, eliminating the need for creating and screening hybridoma fusions. A yeast surface display library of single-chain antibody fragments (scFvs) is created from antigen-binding B cells from the splenocytes of immunized mice. The antigen targeted library is then screened for antigen specific scFv by magneticactivated cell sorting (MACS) and fluorescence-activated cell sorting (FACS). Library construction and screening can be accomplished in as little as 2 weeks, resulting in a panel of scFvs specific for the target antigen.

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

    Science.gov (United States)

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

    2016-07-01

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

  9. Effects of fucosylated milk of goat and mouse on Helicobacter pylori binding to Lewis b antigen

    Institute of Scientific and Technical Information of China (English)

    Hong-Tao Xu; Ning Li; Lennart Hammarstr(o)m; Thomas Borén; Rolf Sj(o)str(o)m; Yao-Feng Zhao; Zheng-Xing Lian; Bao-Liang Fan; Zhi-Hui Zhao; Shu-Yang Yu; Yun-Ping Dai; Li-Li Wang; Hui-Ling Niu

    2004-01-01

    AIM: To evaluate the effects of animal milk containing fucosylated antigens on Helicobacter pylori (Hpylori) binding to Lewis b antigen.METHODS: A mammary gland expression vector containing human α1-3/4-fucosyltransferase cDNA sequences was constructed. Transient expression of human α1-3/4-fucosyltransferase cDNA in goat mammary cell and establishment of transgenic mice were performed. The adhesion inhibitory properties of milk samples were analyzed by using H pylori RESULTS: Goat milk samples were found to inhibit bacterial binding to Lewis b antigen. The highest inhibition was observed 42 h after injection of the plasmid. The binding activity of Hpylori to Lewis b antigen reduced mostly, by 83%, however milk samples from transgenic mice did not inhibit H pylori binding to Lewis b antigen.CONCLUSION: The use of "humanized" animal milk produced by the transgenic introduction of fucosylated antigen can perhaps provide an alternative therapy and preventive measure for H pylori infection.

  10. Liposome-Based Adjuvants for Subunit Vaccines: Formulation Strategies for Subunit Antigens and Immunostimulators

    DEFF Research Database (Denmark)

    Schmidt, Signe Tandrup; Foged, Camilla; Korsholm, Karen Smith;

    2016-01-01

    be classified into delivery systems or immunostimulators. Liposomes are versatile delivery systems for antigens, and they can carefully be customized towards desired immune profiles by combining them with immunostimulators and optimizing their composition, physicochemical properties and antigen-loading mode...... for which no effective vaccines exist. The subunit vaccine technology exploits pathogen subunits as antigens, e.g., recombinant proteins or synthetic peptides, allowing for highly specific immune responses against the pathogens. However, such antigens are usually not sufficiently immunogenic to induce...... protective immunity, and they are often combined with adjuvants to ensure robust immune responses. Adjuvants are capable of enhancing and/or modulating immune responses by exposing antigens to antigen-presenting cells (APCs) concomitantly with conferring immune activation signals. Few adjuvant systems have...

  11. Concepts and applications for influenza antigenic cartography

    Science.gov (United States)

    Cai, Zhipeng; Zhang, Tong; Wan, Xiu-Feng

    2011-01-01

    Influenza antigenic cartography projects influenza antigens into a two or three dimensional map based on immunological datasets, such as hemagglutination inhibition and microneutralization assays. A robust antigenic cartography can facilitate influenza vaccine strain selection since the antigenic map can simplify data interpretation through intuitive antigenic map. However, antigenic cartography construction is not trivial due to the challenging features embedded in the immunological data, such as data incompleteness, high noises, and low reactors. To overcome these challenges, we developed a computational method, temporal Matrix Completion-Multidimensional Scaling (MC-MDS), by adapting the low rank MC concept from the movie recommendation system in Netflix and the MDS method from geographic cartography construction. The application on H3N2 and 2009 pandemic H1N1 influenza A viruses demonstrates that temporal MC-MDS is effective and efficient in constructing influenza antigenic cartography. The web sever is available at http://sysbio.cvm.msstate.edu/AntigenMap. PMID:21761589

  12. Genome Scale Identification of Treponema pallidum Antigens

    OpenAIRE

    McKevitt, Matthew; Brinkman, Mary Beth; McLoughlin, Melanie; Perez, Carla; Howell, Jerrilyn K.; Weinstock, George M.; Norris, Steven J; Palzkill, Timothy

    2005-01-01

    Antibody responses for 882 of the 1,039 proteins in the proteome of Treponema pallidum were examined. Sera collected from infected rabbits were used to systematically identify 106 antigenic proteins, including 22 previously identified antigens and 84 novel antigens. Additionally, sera collected from rabbits throughout the course of infection demonstrated a progression in the breadth and intensity of humoral immunoreactivity against a representative panel of T. pallidum antigens.

  13. Towards efficient cancer immunotherapy: advances in developing artificial antigen-presenting cells

    NARCIS (Netherlands)

    Eggermont, L.J.; Paulis, L.E.M.; Tel, J.; Figdor, C.G.

    2014-01-01

    Active anti-cancer immune responses depend on efficient presentation of tumor antigens and co-stimulatory signals by antigen-presenting cells (APCs). Therapy with autologous natural APCs is costly and time-consuming and results in variable outcomes in clinical trials. Therefore, development of artif

  14. Peripheral blood lymphocytes from low-grade squamous intraepithelial lesions patients recognize vaccine antigens in the presence of activated dendritic cells, and produced high levels of CD8 + IFNγ + T cells and low levels of IL-2 when induced to proliferate

    Directory of Open Access Journals (Sweden)

    Hernández-Montes Jorge

    2012-05-01

    Full Text Available Abstract Background Most infections with human papillomavirus (HPV are resolved without clinical intervention, but a minority evolves into chronic lesions of distinct grades, including cervical-uterine cancer. It is known that in most cases the immune system mediates elimination of HPV infection. However, the mechanism of immune evasion leading to HPV persistence and development of early cervical lesions is not fully understood. The aim of the present work was to evaluate the potential of peripheral blood leukocytes (PBL from low-grade squamous intraepithelial lesions (LSIL patients to be activated ex-vivo by vaccine antigens, the participation of cytotoxic lymphocytes and regulatory T cells, and to determine the secretion of Th1 and Th2 cytokines mediated by stimulation of T cell receptors. Results We found that PBL from LSIL patients showed a significantly lower proliferation rate to vaccine antigens as compared to that of healthy donors, even though there was not a difference in the presence of antibodies to those antigens in sera from both groups. We did not find differences in either the frequency of CD4 + CD25 + FoxP3+ in PBL, or the levels of IL-4, IL-5 and IL-10 in plasma or conditioned media from PBL incubated with TcR agonists in vitro, between the two groups. However, we detected a lower production of IL-2 and a higher proportion of CD8 + IFNγ + cells in PBL from LSIL patients as compared with PBL from normal donors. We also observed that PBL from patients infected by HPV-16 and −18 were not able to proliferate in the presence of soluble HPV antigens added to the culture; however, a high level of proliferation was attained when these antigens were presented by activated dendritic cells. Conclusions Our results suggest that the immunodeficiency reported in LSIL patients could be due to the inability of specific cytotoxic T lymphocytes that for some unknown reason are present but unable to mount a response when

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

    NARCIS (Netherlands)

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

    2000-01-01

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

  16. Analysis of the antigen- and mitogen-induced differentiation of B lymphocytes from asymptomatic human immunodeficiency virus-seropositive male homosexuals. Discrepancy between T cell-dependent and T cell-independent activation.

    NARCIS (Netherlands)

    V.J.P. Teeuwsen; T. Logtenberg (Ton); C.H.J. Siebelink (Kees); J. Goudsmit (Jaap); F.G.C.M. Uytdehaag (Fons); A.D.M.E. Osterhaus (Ab); J.M.A. Lange (Joep)

    1987-01-01

    textabstractFive asymptomatic human immunodeficiency virus (HIV)-seropositive ; male homosexuals were immunized with the recall antigens tetanus toxoid (TT) and the three types of poliovirus present in diphtheria, tetanus, and polio vaccine. Four weeks after immunization, the in vivo response to boo

  17. Immunoprotective activity of a Salmonid Alphavirus Vaccine: comparison of the immune responses induced by inactivated whole virus antigen formulations based on CpG class B oligonucleotides and poly I:C alone or combined with an oil adjuvant.

    Science.gov (United States)

    Thim, Hanna L; Iliev, Dimitar B; Christie, Karen E; Villoing, Stéphane; McLoughlin, Marian F; Strandskog, Guro; Jørgensen, Jorunn B

    2012-07-01

    CpG oligonucleotides and polyinosinic:polycytidylic acid (poly I:C) are toll-like receptor (TLR) agonists that mimic the immunostimulatory properties of bacterial DNA and double-stranded viral RNA respectively, and which have exhibited potential to serve as vaccine adjuvants in previous experiments. Here, a combination of CpGs and poly I:C together with water- or oil-formulated Salmonid Alphavirus (SAV) antigen preparations has been used for a vaccine in Atlantic salmon and tested for protection in SAV challenge trial. The results demonstrate that vaccination with a high dose of the SAV antigen induced protection against challenge with SAV which correlated with production of neutralizing antibodies (NAbs). As the high antigen dose alone induced full protection, no beneficial effect from the addition of CpG and poly I:C could be observed. Nevertheless, these TLR ligands significantly enhanced the levels of NAbs in serum of vaccinated fish. Interestingly, gene expression analysis demonstrated that while addition of oil suppressed the CpG/poly I:C-induced expression of IFN-γ, the upregulation of IFNa1 was substantially enhanced. A low dose of the SAV antigen combined with oil did not induce any detectable levels of NAbs either with or without TLR ligands present, however the addition of CpG and poly I:C to the low SAV antigen dose formulation significantly enhanced the protection against SAV suggesting that CpG/poly I:C may have enhanced a cytotoxic response - a process which is dependent on the up-regulation of type I IFN. These results highlight the immunostimulatory properties of the tested TLR ligands and will serve as a ground for further, more detailed studies aimed to investigate their capacity to serve as adjuvants in vaccine formulations for Atlantic salmon. PMID:22634299

  18. State of the Art in Tumor Antigen and Biomarker Discovery

    Directory of Open Access Journals (Sweden)

    Patrick Chames

    2011-06-01

    Full Text Available Our knowledge of tumor immunology has resulted in multiple approaches for the treatment of cancer. However, a gap between research of new tumors markers and development of immunotherapy has been established and very few markers exist that can be used for treatment. The challenge is now to discover new targets for active and passive immunotherapy. This review aims at describing recent advances in biomarkers and tumor antigen discovery in terms of antigen nature and localization, and is highlighting the most recent approaches used for their discovery including “omics” technology.

  19. State of the Art in Tumor Antigen and Biomarker Discovery

    Energy Technology Data Exchange (ETDEWEB)

    Even-Desrumeaux, Klervi; Baty, Daniel; Chames, Patrick, E-mail: patrick.chames@inserm.fr [INSERM U624, 163 avenue de Luminy, 13288 Marseille Cedex 09 (France)

    2011-06-09

    Our knowledge of tumor immunology has resulted in multiple approaches for the treatment of cancer. However, a gap between research of new tumors markers and development of immunotherapy has been established and very few markers exist that can be used for treatment. The challenge is now to discover new targets for active and passive immunotherapy. This review aims at describing recent advances in biomarkers and tumor antigen discovery in terms of antigen nature and localization, and is highlighting the most recent approaches used for their discovery including “omics” technology.

  20. INVESTIGATION OF INDUCING EFFECT OF SPECIFIC CYTOTOXICITY OF CTLS BY ANTIGEN PEPTIDES FROM T LYMPHOCYTIC LEUKEMIA CELLS

    Institute of Scientific and Technical Information of China (English)

    张桂梅; 黄波; 李东; 王洪涛; 冯作化

    2003-01-01

    Objective: To investigate the characteristics of specific antitumor immunity induced by antigen peptides mixture from T lymphocytic leukemia cells. Method: Antigen peptides mixtures were prepared from different leukemia cell lines and then bound with Hsp70 in vitro. Human peripheral blood mononuclear cells (PBMC) were cultured in vitro, and activated with Hsp70-antigen peptides. The activated PBMC was cultured continuously in vitro, and used as effector cells in vitro test of cytotoxicity to different target cells. Results: The antigen peptides from different leukemia cell lines were peptides mixture and could activate PBMC effectively if they were presented by Hsp70. The activated PBMC could proliferate in the presence of IL-2 and Hsp70-antigen peptides. The proliferative PBMC had specific cytotoxicity to leukemia cells corresponding to the antigen peptides. PBMC activated by antigen peptides from T lymphocytic leukemia cell lines could effectively kill T lymphocytic leukemia cells, and the cytotoxicity of these PBMC to T lymphocytic leukemia cells was significantly stronger than that of PBMC activated by antigen peptides from other leukemia cells (P < 0.05). PBMC activated by either Hut78-peptides or Molt 4-peptides could effectively kill Jurkat cells. And the cytotoxicity of PBMC activated by Hut78/Molt-4-peptides to Jurkat cells was significantly stronger than that of PBMC activated by either Hut78-peptides or Molt-4-peptides alone (P<0.05).Conclusion: Antigen peptides mixture from T lymphocytic leukemia cell lines can induce specific cytotoxic effect to T lymphocytic leukemia cells. There exists cross-reactivity among antigen peptides mixture from different T lymphocytic leukemia cell lines. The cross-reactivity could be amplified by blending of different antigen peptides from different T lymphocytic leukemia cell lines, suggesting that it is possible to prepare broad-spectrum antigen peptide vaccine against T lymphocytic leukemia by using multiple leukemia

  1. Rationally designed inhibitor targeting antigen-trimming aminopeptidases enhances antigen presentation and cytotoxic T-cell responses.

    Science.gov (United States)

    Zervoudi, Efthalia; Saridakis, Emmanuel; Birtley, James R; Seregin, Sergey S; Reeves, Emma; Kokkala, Paraskevi; Aldhamen, Yasser A; Amalfitano, Andrea; Mavridis, Irene M; James, Edward; Georgiadis, Dimitris; Stratikos, Efstratios

    2013-12-01

    Intracellular aminopeptidases endoplasmic reticulum aminopeptidases 1 and 2 (ERAP1 and ERAP2), and as well as insulin-regulated aminopeptidase (IRAP) process antigenic epitope precursors for loading onto MHC class I molecules and regulate the adaptive immune response. Their activity greatly affects the antigenic peptide repertoire presented to cytotoxic T lymphocytes and as a result can regulate cytotoxic cellular responses contributing to autoimmunity or immune evasion by viruses and cancer cells. Therefore, pharmacological regulation of their activity is a promising avenue for modulating the adaptive immune response with possible applications in controlling autoimmunity, in boosting immune responses to pathogens, and in cancer immunotherapy. In this study we exploited recent structural and biochemical analysis of ERAP1 and ERAP2 to design and develop phosphinic pseudopeptide transition state analogs that can inhibit this family of enzymes with nM affinity. X-ray crystallographic analysis of one such inhibitor in complex with ERAP2 validated our design, revealing a canonical mode of binding in the active site of the enzyme, and highlighted the importance of the S2' pocket for achieving inhibitor potency. Antigen processing and presentation assays in HeLa and murine colon carcinoma (CT26) cells showed that these inhibitors induce increased cell-surface antigen presentation of transfected and endogenous antigens and enhance cytotoxic T-cell responses, indicating that these enzymes primarily destroy epitopes in those systems. This class of inhibitors constitutes a promising tool for controlling the cellular adaptive immune response in humans by modulating the antigen processing and presentation pathway. PMID:24248368

  2. Development of the Artificial Antigens for the Organophosphorus Insecticide chlorpyrifos

    Institute of Scientific and Technical Information of China (English)

    ZHU Guo-nian; WU Gang; WU Hui-ming

    2004-01-01

    This study reported that the hapten of the organophosphorus insecticide chlorpyrifos,O,Odiethyl-O-[3,5-dichloro-6-(2-carboxyethyl)thio-2-pyridyl]phosphorothioate(named AR) was synthesized by using technical grade chlorpyrifos reacted with 3-marcapropanoic acid in hot alkaline solution.The hapten was conjugated to bovine serum albumin (BSA) with the modified active ester method to form artificial immune antigen.The ratio of AR:BSA was 39:1.The artificial coating antigen for chlorpyrifos was synthesized by conjugating AR to ovalbumin (OVA) with the mixed-anhydride method,and the ratio was 13:1.The anti-chlorpyrifos polyclonal antibodies were obtained by using the artificial immune antigen (AR-BSA) to immune in the rabbits.

  3. Artificial Loading of ASC Specks with Cytosolic Antigens.

    Directory of Open Access Journals (Sweden)

    Ali Can Sahillioğlu

    Full Text Available Inflammasome complexes form upon interaction of Nod Like Receptor (NLR proteins with pathogen associated molecular patterns (PAPMS inside the cytosol. Stimulation of a subset of inflammasome receptors including NLRP3, NLRC4 and AIM2 triggers formation of the micrometer-sized spherical supramolecular complex called the ASC speck. The ASC speck is thought to be the platform of inflammasome activity, but the reason why a supramolecular complex is preferred against oligomeric platforms remains elusive. We observed that a set of cytosolic proteins, including the model antigen ovalbumin, tend to co-aggregate on the ASC speck. We suggest that co-aggregation of antigenic proteins on the ASC speck during intracellular infection might be instrumental in antigen presentation.

  4. Delayed type hypersensitivity to allogeneic mouse epidermal cell antigens, 2

    International Nuclear Information System (INIS)

    A low dose of ultraviolet B radiation impairs the effectiveness of epidermal cell antigens. We studied the effect of ultraviolet B radiation on the delayed type hypersensitivity induced by allogeneic epidermal cell antigen. The delayed type hypersensitivity response was assayed by footpad swelling in mice. When epidermal cells were exposed to ultraviolet B radiation (660 J/m2), their ability to induce T cells of delayed type hypersensitivity activation was markedly inhibited in any combination of recipient mice and allogeneic epidermal cells. The effect of ultraviolet B radiation on epidermal cells was observed before immunization and challenge. Ultraviolet B treated epidermal cells did not induce suppressor T cells in mice. These results indicate that ultraviolet B radiation destroys the antigenicity of epidermal cells. (author)

  5. Antigenic Variation in Plasmodium falciparum.

    Science.gov (United States)

    Petter, Michaela; Duffy, Michael F

    2015-01-01

    Plasmodium falciparum is the protozoan parasite that causes most malaria-associated morbidity and mortality in humans with over 500,000 deaths annually. The disease symptoms are associated with repeated cycles of invasion and asexual multiplication inside red blood cells of the parasite. Partial, non-sterile immunity to P. falciparum malaria develops only after repeated infections and continuous exposure. The successful evasion of the human immune system relies on the large repertoire of antigenically diverse parasite proteins displayed on the red blood cell surface and on the merozoite membrane where they are exposed to the human immune system. Expression switching of these polymorphic proteins between asexual parasite generations provides an efficient mechanism to adapt to the changing environment in the host and to maintain chronic infection. This chapter discusses antigenic diversity and variation in the malaria parasite and our current understanding of the molecular mechanisms that direct the expression of these proteins. PMID:26537377

  6. Efficient induction of CD25- iTreg by co-immunization requires strongly antigenic epitopes for T cells

    Directory of Open Access Journals (Sweden)

    Li Jinyao

    2011-05-01

    Full Text Available Abstract Background We previously showed that co-immunization with a protein antigen and a DNA vaccine coding for the same antigen induces CD40low IL-10high tolerogenic DCs, which in turn stimulates the expansion of antigen-specific CD4+CD25-Foxp3+ regulatory T cells (CD25- iTreg. However, it was unclear how to choose the antigen sequence to maximize tolerogenic antigen presentation and, consequently, CD25- iTreg induction. Results In the present study, we demonstrated the requirement of highly antigenic epitopes for CD25- iTreg induction. Firstly, we showed that the induction of CD25- iTreg by tolerogenic DC can be blocked by anti-MHC-II antibody. Next, both the number and the suppressive activity of CD25- iTreg correlated positively with the overt antigenicity of an epitope to activate T cells. Finally, in a mouse model of dermatitis, highly antigenic epitopes derived from a flea allergen not only induced more CD25- iTreg, but also more effectively prevented allergenic reaction to the allergen than did weakly antigenic epitopes. Conclusions Our data thus indicate that efficient induction of CD25- iTreg requires highly antigenic peptide epitopes. This finding suggests that highly antigenic epitopes should be used for efficient induction of CD25- iTreg for clinical applications such as flea allergic dermatitis.

  7. Production of Functionally Active and Immunogenic Non-Glycosylated Protective Antigen from Bacillus anthracis in Nicotiana benthamiana by Co-Expression with Peptide-N-Glycosidase F (PNGase F) of Flavobacterium meningosepticum.

    Science.gov (United States)

    Mamedov, Tarlan; Chichester, Jessica A; Jones, R Mark; Ghosh, Ananya; Coffin, Megan V; Herschbach, Kristina; Prokhnevsky, Alexey I; Streatfield, Stephen J; Yusibov, Vidadi

    2016-01-01

    Bacillus anthracis has long been considered a potential biological warfare agent, and therefore, there is a need for a safe, low-cost and highly efficient anthrax vaccine with demonstrated long-term stability for mass vaccination in case of an emergency. Many efforts have been made towards developing an anthrax vaccine based on recombinant protective antigen (rPA) of B. anthracis, a key component of the anthrax toxin, produced using different expression systems. Plants represent a promising recombinant protein production platform due to their relatively low cost, rapid scalability and favorable safety profile. Previous studies have shown that full-length rPA produced in Nicotiana benthamiana (pp-PA83) is immunogenic and can provide full protection against lethal spore challenge; however, further improvement in the potency and stability of the vaccine candidate is necessary. PA of B. anthracis is not a glycoprotein in its native host; however, this protein contains potential N-linked glycosylation sites, which can be aberrantly glycosylated during expression in eukaryotic systems including plants. This glycosylation could affect the availability of certain key epitopes either due to masking or misfolding of the protein. Therefore, a non-glycosylated form of pp-PA83 was engineered and produced in N. benthamiana using an in vivo deglycosylation approach based on co-expression of peptide-N-glycosidase F (PNGase F) from Flavobacterium meningosepticum. For comparison, versions of pp-PA83 containing point mutations in six potential N-glycosylation sites were also engineered and expressed in N. benthamiana. The in vivo deglycosylated pp-PA83 (pp-dPA83) was shown to have in vitro activity, in contrast to glycosylated pp-PA83, and to induce significantly higher levels of toxin-neutralizing antibody responses in mice compared with glycosylated pp-PA83, in vitro deglycosylated pp-PA83 or the mutated versions of pp-PA83. These results suggest that pp-dPA83 may offer advantages

  8. [HLA antigens in juvenile rheumatoid arthritis].

    Science.gov (United States)

    Rumba, I V; Sochnev, A M; Kukaĭne, E M; Burshteĭn, A M; Benevolenskaia, L I

    1990-01-01

    Antigens of I class HLA system (locus A and B) were investigated in 67 patients of Latvian nationality suffering from juvenile rheumatoid arthritis (JRA). Associations of HLA antigens with juvenile rheumatoid arthritis partially coincided with the ones revealed earlier. Typing established an increased incidence of antigen B27 (p less than 0.01) and gaplotype A2, B40 (p less than 0.01). Antigen B15 possessed a protective action with respect to JRA. Interlocus combinations demonstrated a closer association with the disease than a single antigen. The authors also revealed markers of various clinico-anatomical variants of JRA.

  9. Immunoregulation by Taenia crassiceps and Its Antigens

    Directory of Open Access Journals (Sweden)

    Alberto N. Peón

    2013-01-01

    Full Text Available Taenia crassiceps is a cestode parasite of rodents (in its larval stage and canids (in its adult stage that can also parasitize immunocompromised humans. We have studied the immune response elicited by this helminth and its antigens in mice and human cells, and have discovered that they have a strong capacity to induce chronic Th2-type responses that are primarily characterized by high levels of Th2 cytokines, low proliferative responses in lymphocytes, an immature and LPS-tolerogenic profile in dendritic cells, the recruitment of myeloid-derived suppressor cells and, specially, alternatively activated macrophages. We also have utilized the immunoregulatory capabilities of this helminth to successfully modulate autoimmune responses and the outcome of other infectious diseases. In the present paper, we review the work of others and ourselves with regard to the immune response induced by T. crassiceps and its antigens, and we compare the advances in our understanding of this parasitic infection model with the knowledge that has been obtained from other selected models.

  10. CD28 and T cell antigen receptor signal transduction coordinately regulate interleukin 2 gene expression in response to superantigen stimulation

    OpenAIRE

    1992-01-01

    Activation of an immune response requires intercellular contact between T lymphocytes and antigen-presenting cells (APC). Interaction of the T cell antigen receptor (TCR) with antigen in the context of major histocompatibility molecules mediates signal transduction, but T cell activation appears to require the induction of a second costimulatory signal transduction pathway. Recent studies suggest that interaction of CD28 with B7 on APC might deliver such a costimulatory signal. To investigate...

  11. Detection of Avian Antigen-Specific T Cells Induced by Viral Vaccines

    DEFF Research Database (Denmark)

    Dalgaard, Tina Sørensen; Norup, Liselotte Rothmann; Juul-Madsen, Helle Risdahl

    2016-01-01

    Live attenuated viral vaccines are widely used in commercial poultry production, but the development of new effective inactivated/subunit vaccines is needed. Studies of avian antigen-specific T cells are primarily based on analyses ex vivo after activating the cells with recall antigen. There is ......Live attenuated viral vaccines are widely used in commercial poultry production, but the development of new effective inactivated/subunit vaccines is needed. Studies of avian antigen-specific T cells are primarily based on analyses ex vivo after activating the cells with recall antigen...... in the cells even throughout division. This leads to daughter cells containing half the fluorescence of their parents. When lymphocytes are loaded with CFSE prior to ex vivo stimulation with specific antigen, the measurement of serial halving of its fluorescence by flow cytometry identifies the cells...

  12. Evaluation of Intracellular Signaling Downstream Chimeric Antigen Receptors.

    Directory of Open Access Journals (Sweden)

    Hannah Karlsson

    Full Text Available CD19-targeting CAR T cells have shown potency in clinical trials targeting B cell leukemia. Although mainly second generation (2G CARs carrying CD28 or 4-1BB have been investigated in patients, preclinical studies suggest that third generation (3G CARs with both CD28 and 4-1BB have enhanced capacity. However, little is known about the intracellular signaling pathways downstream of CARs. In the present work, we have analyzed the signaling capacity post antigen stimulation in both 2G and 3G CARs. 3G CAR T cells expanded better than 2G CAR T cells upon repeated stimulation with IL-2 and autologous B cells. An antigen-driven accumulation of CAR+ cells was evident post antigen stimulation. The cytotoxicity of both 2G and 3G CAR T cells was maintained by repeated stimulation. The phosphorylation status of intracellular signaling proteins post antigen stimulation showed that 3G CAR T cells had a higher activation status than 2G. Several proteins involved in signaling downstream the TCR were activated, as were proteins involved in the cell cycle, cell adhesion and exocytosis. In conclusion, 3G CAR T cells had a higher degree of intracellular signaling activity than 2G CARs which may explain the increased proliferative capacity seen in 3G CAR T cells. The study also indicates that there may be other signaling pathways to consider when designing or evaluating new generations of CARs.

  13. BIOCHEMICAL STUDIES ON SO-CALLED SYPHILIS ANTIGEN.

    Science.gov (United States)

    Noguchi, H; Bronfenbrenner, J

    1911-01-01

    The liver tissues of man and certain animals (dogs, rabbits, guinea pigs, etc.) yield, upon alcoholic extraction, various substances which may be divided by their physical and chemical properties into several groups. While many substances are present in the alcoholic extract, the ones possessing antigenic properties are comparatively few. The latter are responsible for the antigenic properties exhibited by the whole alcoholic extract. The substances extracted with alcohol were fractionated into the following four groups. (a) Substances Insoluble in Ether and Hot Alcohol.-These are chiefly proteins and salts. The proteins are probably the minute particles of larger molecules held in apparent suspension in alcohol until all other substances are removed. The water extracted from the tissues and admixed with alcohol is also an essential factor in extracting these particles in an alcoholic solution. The salts present are the usual physiological constituents of the liver, notably, sodium chloride. The quantity of these substances extracted with alcohol varies greatly with different specimens. Biologically considered, they are neither markedly hemolytic nor anticomplementary and possess no antigenic property for the Wassermann reaction. It is important, however, to note that the proteins bind complement when mixed with certain active human sera. For this reason a preparation of antigen containing this group of substances is unsuitable for use in combination with an active serum, and should, therefore, be rejected. (b) Substances Insoluble in Ether and Soluble in Hot Alcohol.-This group contains soaps, cleavage products of proteins, and small amounts of the bile salts. Soaps and bile salts are very strongly hemolytic and are absolutely unfit for use as antigen. Moreover, their antigenic properties are very slight. It is best to eliminate this group of substances from the preparation of antigen. The quantity of the substances of this group extracted from different specimens

  14. Surface antigen expression and correlation with variable heavy-chain gene mutation status in chronic lymphocytic leukemia.

    Science.gov (United States)

    Vilpo, Juhani; Tobin, Gerard; Hulkkonen, Janne; Hurme, Mikko; Thunberg, Ulf; Sundström, Christer; Vilpo, Leena; Rosenquist, Richard

    2003-01-01

    Recent studies have demonstrated that B-cell chronic lymphocytic leukemia (CLL) consists of two clinical entities with either somatically hypermutated (M-CLL) or unmutated (UM-CLL) immunoglobulin variable heavy-chain (VH) regions. In view of the fact that the cellular biology of these two subsets of disease is currently unexplored, we performed an extensive analysis of the surface antigen expression and correlated this with the VH gene mutation status in a cohort of 32 CLL patients. Using polymerase chain reaction amplification and nucleotide sequencing, the VH genes were shown to be mutated in 10 cases (31%) and unmutated in 22 (69%). The expression of 27 surface membrane antigens in peripheral blood leukemic cells was analyzed by flow cytometry, measuring both the percentage of positive cells as well as the geometric mean fluorescence intensity (GMF). Most of the surface membrane antigens (CD5, CD11c, CD19, CD20, CD21, CD22, CD23, CD25, CD40, CD45, VD79b, CD80, CD95, CD122, CD124, CD126, CD130, CD154, IgM, and IgD) showed a similar expression pattern in both UM-CLL and M-CLL patients. The similarity of M-CLL and UM-CLL, as demonstrated here for the first time with many protein markers, indicates a considerably homogeneous phenotype in both subsets. Furthermore, CD27 was strongly expressed in all cases, which may suggest a memory cell phenotype for both M-CLL and UM-CLL. More positive cells in the UM-CLL group were observed regarding CD38, but CD38 was not a good predictor of VH gene mutation status. Seventy percent of the M-CLL cases, but only 36% of UM-CLL cases, were Ig-lambda+. The most striking differential expression, however, was observed in the two slicing variants of the common leukocyte antigen CD45, namely CD45RO and CD45RA. CD45RO expression was significantly associated with M-CLL, whereas the GMF intensity of CD45RA tended to be associated with UM-CLL. The role of these CD45 splicing variants in the pathogenesis of CLL deserves further investigation

  15. Anti-tuberculosis activity of TCR gene-modified T cells specific for the M.tuberculosis 38 kD antigen%38kD抗原特异TCR基因修饰T细胞的抗结核抗原活性研究

    Institute of Scientific and Technical Information of China (English)

    罗微; 张笑冰; 黄永塔; 郝佩佩; 姜振民; 温茜; 周明乾; 马骊

    2012-01-01

    Recent advances in T-cell receptor (TCR) gene modification methods have enabled researchers to design T-cells specific for target antigens, e.g., tumor-associated or viral antigens. These genetically redirected T-cells display similar anti-tumor or anti-viral activity as the cell clones from which the TCR genes were isolated. However, there is no data describing the activity of T-cells with genetically modified TCR genes specific for intracellular bacterial antigens (e.g., tuberculosis antigens). This study enriched T-cells specific for the mycobacteria! 38 kD glycolipoprotein by repeated in vitro stimulation with antigen-pulsed dendritic cells. Magnetic beads were used to separate CD4+ and CD8+ T-cells. Spectratype analysis was then used to identify and characterize the third complimentarily-determining regions (CDR3) of the antigen-specific human leukocyte antigen (HLA) class Ⅱ- and Ⅰ-restricted TCRs derived from specific CD4+ or CDS+ T-cells, respectively. Full-length TCR genes were inserted into retroviral vectors and transferred into autologous, primary CD4+ and CD8+ T-cells. Both redirected CD4+ and CD8+ T-cells expressed exogenous TCRs with the ability to recognize specific antigens and mediate effector functions. Thus, the method presented in this study generated TCR gene-modified T-cells that can treat multidrug-resistant Mycobaclerium tuberculosis infections.%抗原特异T细胞受体(T cell receptor,TCR)基因修饰T细胞已被应用于肿瘤、病毒感染过继免疫治疗研究,取得了鼓舞人心的结果,但在结核上未见报道.本研究利用结核分枝杆菌38 kD抗原刺激人CD4+,CD8+T细胞,通过分析刺激前后T细胞TCR互补决定区3(complementarity determining region 3,CDR3)谱型,从CD4+,CD8+T细胞中分别筛选出38 kD抗原特异的TCR Vα11,Vβ8和Vα3,vβ8基因家族T细胞,聚合酶链式反应(polymerase chain reaction,PCR)扩增获得其α,β链全长基因并插入逆转录病毒载体,构建重组载体pMX-β8

  16. CD85/LIR-1/ILT2 and CD152 (Cytotoxic T Lymphocyte Antigen 4) Inhibitory Molecules Down-Regulate the Cytolytic Activity of Human CD4+ T-Cell Clones Specific for Mycobacterium tuberculosis

    OpenAIRE

    Merlo, Andrea; Saverino, Daniele; Tenca, Claudya; Grossi, Carlo Enrico; Bruno, Silvia; Ciccone, Ermanno

    2001-01-01

    Antigen-specific cytolytic CD4+ T lymphocytes control Mycobacterium tuberculosis infection by secreting cytokines and by killing macrophages that have phagocytosed the pathogen. However, lysis of the latter cells promotes microbial dissemination, and other macrophages engulf the released bacteria. Subsequently, CD4+ T-cell-mediated killing of macrophages goes on, and this persistent process may hamper control of infection, unless regulatory mechanisms maintain a subtle balance between lysis o...

  17. Evaluation of Alum-Naltrexone Adjuvant Activity, on Efficacy of Anti-Leishmania Immunization with Autoclaved Leishmania major (MRHO/IR/75/ER Antigens in BALB/C Mice.

    Directory of Open Access Journals (Sweden)

    Habib Mohammadzadeh Hajipirloo

    2014-09-01

    Full Text Available Naltrexone, an opioid receptor antagonist shifts the immune response toward a Th1 profile. In the current study, we evaluated the efficacy of the mixture of NTX and alum, as a new adjuvant, to enhance immune response and induce protection against Leishmania major in a mouse model.BALB/c mice were immunized three times either autoclaved L. major promastigotes' antigens alone or in combination with the adjuvant alum, naltrexone or the alum-naltrexone mixture. Both humoral and cellular immune responses were assessed two weeks after the last immunization and compared with control mice.The administration of alum- NTX in combination with the parasite antigen, significantly increased production of IFN-γ IFN-γ /IL-5 ratio, lymphocyte proliferation and improved DTH response against L. major. There was no significant difference in survival following challenge among groups.Immunization with the alum- naltrexone mixture as an adjuvant, in combination with the autoclaved L. major promastigotes antigens, can enhance cellular immunity and shift the immune responses to a Th1 pattern.

  18. THE SEARCH OF OPTIMAL COMBINATION OF ANTIGENS FOR SEROLOGICAL DIAGNOSTICS OF TUBERCULOSIS

    Directory of Open Access Journals (Sweden)

    E. V. Vasilyeva

    2013-01-01

    Full Text Available Abstract. The four chimeric recombinant antigens CBD-CFP10, CBD-ESAT6, ESAT6-CFP10 and CBD-P38 contained aminoacid sequences of full-size proteins ESAT6, CFP10 and matured protein P38 of M. tuberculosis, joined with aminoacid sequences of cellulose bind domain of endogluconase A (CBD from Cellumonas fimi have been obtained by gene engineering methods. Recombinant proteins were purified by affine chromatography in column with Ni-NTA-sepharose 6В-CL and as PPDN-3 were used for detection of their antigenic activity in indirect ELISA for TB serological diagnostics. The sera from patients with lung tuberculosis (n = 321, from persons who had professional contacts with TB patients (n = 42, from healthy blood donors (n = 366 and from patients with lung diseases of non-TB etiology were tested. It was detected that there was positive correlation between antibodies level for all studied antigens compared by pair. It has been demonstrated that although antigens were different by antigenic and immunobiological characteristics they add each other in the content of antigenic diagnostics compositions. Thus, all these antigens can be used in the test kits for serological diagnostics of TB. Using of these antigens will allow to detect persons infected by TB and patients with active tuberculosis. 

  19. Engineering antigen-specific immunological tolerance.

    Energy Technology Data Exchange (ETDEWEB)

    Kontos, Stephan; Grimm, Alizee J.; Hubbell, Jeffrey A.

    2015-05-01

    Unwanted immunity develops in response to many protein drugs, in autoimmunity, in allergy, and in transplantation. Approaches to induce immunological tolerance aim to either prevent these responses or reverse them after they have already taken place. We present here recent developments in approaches, based on engineered peptides, proteins and biomaterials, that harness mechanisms of peripheral tolerance both prophylactically and therapeutically to induce antigenspecific immunological tolerance. These mechanisms are based on responses of B and T lymphocytes to other cells in their immune environment that result in cellular deletion or ignorance to particular antigens, or in development of active immune regulatory responses. Several of these approaches are moving toward clinical development, and some are already in early stages of clinical testing.

  20. Antigen-Specific Antibody Glycosylation Is Regulated via Vaccination.

    Science.gov (United States)

    Mahan, Alison E; Jennewein, Madeleine F; Suscovich, Todd; Dionne, Kendall; Tedesco, Jacquelynne; Chung, Amy W; Streeck, Hendrik; Pau, Maria; Schuitemaker, Hanneke; Francis, Don; Fast, Patricia; Laufer, Dagna; Walker, Bruce D; Baden, Lindsey; Barouch, Dan H; Alter, Galit

    2016-03-01

    Antibody effector functions, such as antibody-dependent cellular cytotoxicity, complement deposition, and antibody-dependent phagocytosis, play a critical role in immunity against multiple pathogens, particularly in the absence of neutralizing activity. Two modifications to the IgG constant domain (Fc domain) regulate antibody functionality: changes in antibody subclass and changes in a single N-linked glycan located in the CH2 domain of the IgG Fc. Together, these modifications provide a specific set of instructions to the innate immune system to direct the elimination of antibody-bound antigens. While it is clear that subclass selection is actively regulated during the course of natural infection, it is unclear whether antibody glycosylation can be tuned, in a signal-specific or pathogen-specific manner. Here, we show that antibody glycosylation is determined in an antigen- and pathogen-specific manner during HIV infection. Moreover, while dramatic differences exist in bulk IgG glycosylation among individuals in distinct geographical locations, immunization is able to overcome these differences and elicit antigen-specific antibodies with similar antibody glycosylation patterns. Additionally, distinct vaccine regimens induced different antigen-specific IgG glycosylation profiles, suggesting that antibody glycosylation is not only programmable but can be manipulated via the delivery of distinct inflammatory signals during B cell priming. These data strongly suggest that the immune system naturally drives antibody glycosylation in an antigen-specific manner and highlights a promising means by which next-generation therapeutics and vaccines can harness the antiviral activity of the innate immune system via directed alterations in antibody glycosylation in vivo.  .

  1. Antigen-Specific Antibody Glycosylation Is Regulated via Vaccination.

    Directory of Open Access Journals (Sweden)

    Alison E Mahan

    2016-03-01

    Full Text Available Antibody effector functions, such as antibody-dependent cellular cytotoxicity, complement deposition, and antibody-dependent phagocytosis, play a critical role in immunity against multiple pathogens, particularly in the absence of neutralizing activity. Two modifications to the IgG constant domain (Fc domain regulate antibody functionality: changes in antibody subclass and changes in a single N-linked glycan located in the CH2 domain of the IgG Fc. Together, these modifications provide a specific set of instructions to the innate immune system to direct the elimination of antibody-bound antigens. While it is clear that subclass selection is actively regulated during the course of natural infection, it is unclear whether antibody glycosylation can be tuned, in a signal-specific or pathogen-specific manner. Here, we show that antibody glycosylation is determined in an antigen- and pathogen-specific manner during HIV infection. Moreover, while dramatic differences exist in bulk IgG glycosylation among individuals in distinct geographical locations, immunization is able to overcome these differences and elicit antigen-specific antibodies with similar antibody glycosylation patterns. Additionally, distinct vaccine regimens induced different antigen-specific IgG glycosylation profiles, suggesting that antibody glycosylation is not only programmable but can be manipulated via the delivery of distinct inflammatory signals during B cell priming. These data strongly suggest that the immune system naturally drives antibody glycosylation in an antigen-specific manner and highlights a promising means by which next-generation therapeutics and vaccines can harness the antiviral activity of the innate immune system via directed alterations in antibody glycosylation in vivo.  .

  2. Anti-Ia antibody in the sera of normal subjects after in vivo antigenic stimulation

    OpenAIRE

    1982-01-01

    We showed that sera from normal subjects after antigenic challenge with intradermal PPD or Candida antigens or with subcutaneous tetanus vaccine contain a factor that blocks the binding of mouse monoclonal anti-Ia antibody to Ia-positive T cells or to B35 M cells, an Ia- positive human B cell line. The blocking activity appears 48 to 72 h after antigenic challenge and is gone by day 7. The appearance of the anti-Ia blocking activity coincided with a drop in the percentage of Ia- positive T ce...

  3. Effect of particulate adjuvant on the anthrax protective antigen dose required for effective nasal vaccination.

    Science.gov (United States)

    Bento, Dulce; Staats, Herman F; Borges, Olga

    2015-07-17

    Successful vaccine development is dependent on the development of effective adjuvants since the poor immunogenicity of modern subunit vaccines typically requires the use of potent adjuvants and high antigen doses. In recent years, adjuvant formulations combining both immunopotentiators and delivery systems have emerged as a promising strategy to develop effective and improved vaccines. In this study we investigate if the association of the mast cell activating adjuvant compound 48/80 (C48/80) with chitosan nanoparticles would promote an antigen dose sparing effect when administered intranasally. Even though the induction of strong mucosal immunity required higher antigen doses, incorporation of C48/80 into nanoparticles provided significant dose sparing when compared to antigen and C48/80 in solution with no significant effect on serum neutralizing antibodies titers. These results suggest the potential of this novel adjuvant combination to improve the immunogenicity of a vaccine and decrease the antigen dose required for vaccination. PMID:26087299

  4. Antigen Incorporation on Cryptosporidium parvum Oocyst Walls

    OpenAIRE

    Entrala Emilio; Sbihi Younes; Sánchez-Moreno Manuel; Mascaró Carmen

    2001-01-01

    Cryptosporidium parvum oocysts are the infective stages responsible for transmission and survival of the organism in the environment. In the present work we show that the oocyst wall, far from being a static structure, is able to incorporate antigens by a mechanism involving vesicle fusion with the wall, and the incorporation of the antigen to the outer oocyst wall. Using immunoelectron microscopy we show that the antigen recognized by a monoclonal antibody used for diagnosis of cryptosporidi...

  5. Histocompatibility antigens in coal miners with pneumoconiosis.

    OpenAIRE

    Soutar, C A; Coutts, I.; Parkes, W R; Dodi, I. A.; Gauld, S; Castro, J E; Turner-Warwick, M

    1983-01-01

    Twenty-five histocompatibility antigens have been measured in 100 coal miners with pneumoconiosis attending a pneumoconiosis medical panel and the results compared with a panel of 200 normal volunteers not exposed to dust. Chest radiographs were read independently by three readers according to the ILO U/C classification. On a combined score, 40 men were thought to have simple pneumoconiosis and 60 men complicated pneumoconiosis. The number of antigens tested and associations between antigens ...

  6. Concerted activity of IgG1 antibodies and IL-4/IL-25-dependent effector cells trap helminth larvae in the tissues following vaccination with defined secreted antigens, providing sterile immunity to challenge infection.

    Directory of Open Access Journals (Sweden)

    James P Hewitson

    2015-03-01

    Full Text Available Over 25% of the world's population are infected with helminth parasites, the majority of which colonise the gastrointestinal tract. However, no vaccine is yet available for human use, and mechanisms of protective immunity remain unclear. In the mouse model of Heligmosomoides polygyrus infection, vaccination with excretory-secretory (HES antigens from adult parasites elicits sterilising immunity. Notably, three purified HES antigens (VAL-1, -2 and -3 are sufficient for effective vaccination. Protection is fully dependent upon specific IgG1 antibodies, but passive transfer confers only partial immunity to infection, indicating that cellular components are also required. Moreover, immune mice show greater cellular infiltration associated with trapping of larvae in the gut wall prior to their maturation. Intra-vital imaging of infected intestinal tissue revealed a four-fold increase in extravasation by LysM+GFP+ myeloid cells in vaccinated mice, and the massing of these cells around immature larvae. Mice deficient in FcRγ chain or C3 complement component remain fully immune, suggesting that in the presence of antibodies that directly neutralise parasite molecules, the myeloid compartment may attack larvae more quickly and effectively. Immunity to challenge infection was compromised in IL-4Rα- and IL-25-deficient mice, despite levels of specific antibody comparable to immune wild-type controls, while deficiencies in basophils, eosinophils or mast cells or CCR2-dependent inflammatory monocytes did not diminish immunity. Finally, we identify a suite of previously uncharacterised heat-labile vaccine antigens with homologs in human and veterinary parasites that together promote full immunity. Taken together, these data indicate that vaccine-induced immunity to intestinal helminths involves IgG1 antibodies directed against secreted proteins acting in concert with IL-25-dependent Type 2 myeloid effector populations.

  7. Analysis of expression profiles of MAGE-A antigens in oral squamous cell carcinoma cell lines

    Directory of Open Access Journals (Sweden)

    Reichert Torsten E

    2009-04-01

    Full Text Available Abstract Background The immunological response to solid tumours is insufficient. Therefore, tumour specific antigens have been explored to facilitate the activation of the immune system. The cancer/testis antigen class of MAGE-A antigens is a possible target for vaccination. Their differential expression profiles also modulate the course of the cancer disease and its response to antineoplastic drugs. Methods The expression profiles of MAGE-A2, -A3, -A4, -A6 and -A10 in five own oral squamous cell carcinoma cell lines were characterised by rt-PCR, qrt-PCR and immunocytochemistry with a global MAGE-A antibody (57B and compared with those of an adult keratinocyte cell line (NHEK. Results All tumour cell lines expressed MAGE-A antigens. The antigens were expressed in groups with different preferences. The predominant antigens expressed were MAGE-A2, -A3 and -A6. MAGE-A10 was not expressed in the cell lines tested. The MAGE-A gene products detected in the adult keratinocyte cell line NHEK were used as a reference. Conclusion MAGE-A antigens are expressed in oral squamous cell carcinomas. The expression profiles measured facilitate distinct examinations in forthcoming studies on responses to antineoplastic drugs or radiation therapy. MAGE-A antigens are still an interesting aim for immunotherapy.

  8. A Review of Intra- and Extracellular Antigen Delivery Systems for Virus Vaccines of Finfish.

    Science.gov (United States)

    Munang'andu, Hetron Mweemba; Evensen, Øystein

    2015-01-01

    Vaccine efficacy in aquaculture has for a long time depended on evaluating relative percent survival and antibody responses after vaccination. However, current advances in vaccine immunology show that the route in which antigens are delivered into cells is deterministic of the type of adaptive immune response evoked by vaccination. Antigens delivered by the intracellular route induce MHC-I restricted CD8+ responses while antigens presented through the extracellular route activate MHC-II restricted CD4+ responses implying that the route of antigen delivery is a conduit to induction of B- or T-cell immune responses. In finfish, different antigen delivery systems have been explored that include live, DNA, inactivated whole virus, fusion protein, virus-like particles, and subunit vaccines although mechanisms linking these delivery systems to protective immunity have not been studied in detail. Hence, in this review we provide a synopsis of different strategies used to administer viral antigens via the intra- or extracellular compartments. Further, we highlight the differences in immune responses induced by antigens processed by the endogenous route compared to exogenously processed antigens. Overall, we anticipate that the synopsis put together in this review will shed insights into limitations and successes of the current vaccination strategies used in finfish vaccinology.

  9. A Review of Intra- and Extracellular Antigen Delivery Systems for Virus Vaccines of Finfish

    Directory of Open Access Journals (Sweden)

    Hetron Mweemba Munang’andu

    2015-01-01

    Full Text Available Vaccine efficacy in aquaculture has for a long time depended on evaluating relative percent survival and antibody responses after vaccination. However, current advances in vaccine immunology show that the route in which antigens are delivered into cells is deterministic of the type of adaptive immune response evoked by vaccination. Antigens delivered by the intracellular route induce MHC-I restricted CD8+ responses while antigens presented through the extracellular route activate MHC-II restricted CD4+ responses implying that the route of antigen delivery is a conduit to induction of B- or T-cell immune responses. In finfish, different antigen delivery systems have been explored that include live, DNA, inactivated whole virus, fusion protein, virus-like particles, and subunit vaccines although mechanisms linking these delivery systems to protective immunity have not been studied in detail. Hence, in this review we provide a synopsis of different strategies used to administer viral antigens via the intra- or extracellular compartments. Further, we highlight the differences in immune responses induced by antigens processed by the endogenous route compared to exogenously processed antigens. Overall, we anticipate that the synopsis put together in this review will shed insights into limitations and successes of the current vaccination strategies used in finfish vaccinology.

  10. The uptake of soluble and particulate antigens by epithelial cells in the mouse small intestine.

    Science.gov (United States)

    Howe, Savannah E; Lickteig, Duane J; Plunkett, Kyle N; Ryerse, Jan S; Konjufca, Vjollca

    2014-01-01

    Intestinal epithelial cells (IECs) overlying the villi play a prominent role in absorption of digested nutrients and establish a barrier that separates the internal milieu from potentially harmful microbial antigens. Several mechanisms by which antigens of dietary and microbial origin enter the body have been identified; however whether IECs play a role in antigen uptake is not known. Using in vivo imaging of the mouse small intestine, we investigated whether epithelial cells (enterocytes) play an active role in the uptake (sampling) of lumen antigens. We found that small molecular weight antigens such as chicken ovalbumin, dextran, and bacterial LPS enter the lamina propria, the loose connective tissue which lies beneath the epithelium via goblet cell associated passageways. However, epithelial cells overlying the villi can internalize particulate antigens such as bacterial cell debris and inert nanoparticles (NPs), which are then found co-localizing with the CD11c+ dendritic cells in the lamina propria. The extent of NP uptake by IECs depends on their size: 20-40 nm NPs are taken up readily, while NPs larger than 100 nm are taken up mainly by the epithelial cells overlying Peyer's patches. Blocking NPs with small proteins or conjugating them with ovalbumin does not inhibit their uptake. However, the uptake of 40 nm NPs can be inhibited when they are administered with an endocytosis inhibitor (chlorpromazine). Delineating the mechanisms of antigen uptake in the gut is essential for understanding how tolerance and immunity to lumen antigens are generated, and for the development of mucosal vaccines and therapies.

  11. Development of recombinant antigen array for simultaneous detection of viral antibodies.

    Directory of Open Access Journals (Sweden)

    Yi Liu

    Full Text Available Protein microarrays have been developed to study antibody reactivity against a large number of antigens, demonstrating extensive perspective for clinical application. We developed a viral antigen array by spotting four recombinant antigens and synthetic peptide, including glycoprotein G of herpes simplex virus (HSV type 1 and 2, phosphoprotein 150 of cytomegalovirus (CMV, Rubella virus (RV core plus glycoprotein E1 and E2 as well as a E1 peptide with the optimal concentrations on activated glass slides to simultaneously detect IgG and IgM against HSV1, HSV2, CMV and RV in clinical specimens of sera and cerebrospinal fluids (CSFs. The positive reference sera were initially used to measure the sensitivity and specificity of the array with the optimal conditions. Then clinical specimens of 144 sera and 93 CSFs were tested for IgG and IgM antibodies directed against HSV1, HSV2, CMV and RV by the antigen array. Specificity of the antigen array for viral antibodies detection was satisfying compared to commercial ELISA kits but sensitivity of the array varied relying on quality and antigenic epitopes of the spotting antigens. In short, the recombinant antigen array has potential to simultaneous detect multiple viral antibodies using minute amount (3 µl of samples, which holds the particularly advantage to detect viral antibodies in clinical CSFs being suspicious of neonatal meningitis and encephalitis.

  12. Activation of a heterogeneous hepatitis B (HB) core and e antigen-specific CD4+ T-cell population during seroconversion to anti-HBe and anti-HBs in hepatitis B virus infection.

    OpenAIRE

    Jung, M C; Diepolder, H M; Spengler, U; Wierenga, E A; Zachoval, R; Hoffmann, R M; Eichenlaub, D; Frösner, G; Will, H; Pape, G. R.

    1995-01-01

    Overcoming hepatitis B virus infection essentially depends on the appropriate immune response of the infected host. Among the hepatitis B virus antigens, the core (HBcAg) and e (HBeAg) proteins appear highly immunogenic and induce important lymphocyte effector functions. In order to investigate the importance of HBcAg/HBeAg-specific T lymphocytes in patients with acute and chronic hepatitis B and to identify immunodominant epitopes within the HBcAg/HBeAg, CD4+ T-cell responses to hepatitis B ...

  13. A computational framework for influenza antigenic cartography.

    Directory of Open Access Journals (Sweden)

    Zhipeng Cai

    Full Text Available Influenza viruses have been responsible for large losses of lives around the world and continue to present a great public health challenge. Antigenic characterization based on hemagglutination inhibition (HI assay is one of the routine procedures for influenza vaccine strain selection. However, HI assay is only a crude experiment reflecting the antigenic correlations among testing antigens (viruses and reference antisera (antibodies. Moreover, antigenic characterization is usually based on more than one HI dataset. The combination of multiple datasets results in an incomplete HI matrix with many unobserved entries. This paper proposes a new computational framework for constructing an influenza antigenic cartography from this incomplete matrix, which we refer to as Matrix Completion-Multidimensional Scaling (MC-MDS. In this approach, we first reconstruct the HI matrices with viruses and antibodies using low-rank matrix completion, and then generate the two-dimensional antigenic cartography using multidimensional scaling. Moreover, for influenza HI tables with herd immunity effect (such as those from Human influenza viruses, we propose a temporal model to reduce the inherent temporal bias of HI tables caused by herd immunity. By applying our method in HI datasets containing H3N2 influenza A viruses isolated from 1968 to 2003, we identified eleven clusters of antigenic variants, representing all major antigenic drift events in these 36 years. Our results showed that both the completed HI matrix and the antigenic cartography obtained via MC-MDS are useful in identifying influenza antigenic variants and thus can be used to facilitate influenza vaccine strain selection. The webserver is available at http://sysbio.cvm.msstate.edu/AntigenMap.

  14. A computational framework for influenza antigenic cartography.

    Science.gov (United States)

    Cai, Zhipeng; Zhang, Tong; Wan, Xiu-Feng

    2010-10-07

    Influenza viruses have been responsible for large losses of lives around the world and continue to present a great public health challenge. Antigenic characterization based on hemagglutination inhibition (HI) assay is one of the routine procedures for influenza vaccine strain selection. However, HI assay is only a crude experiment reflecting the antigenic correlations among testing antigens (viruses) and reference antisera (antibodies). Moreover, antigenic characterization is usually based on more than one HI dataset. The combination of multiple datasets results in an incomplete HI matrix with many unobserved entries. This paper proposes a new computational framework for constructing an influenza antigenic cartography from this incomplete matrix, which we refer to as Matrix Completion-Multidimensional Scaling (MC-MDS). In this approach, we first reconstruct the HI matrices with viruses and antibodies using low-rank matrix completion, and then generate the two-dimensional antigenic cartography using multidimensional scaling. Moreover, for influenza HI tables with herd immunity effect (such as those from Human influenza viruses), we propose a temporal model to reduce the inherent temporal bias of HI tables caused by herd immunity. By applying our method in HI datasets containing H3N2 influenza A viruses isolated from 1968 to 2003, we identified eleven clusters of antigenic variants, representing all major antigenic drift events in these 36 years. Our results showed that both the completed HI matrix and the antigenic cartography obtained via MC-MDS are useful in identifying influenza antigenic variants and thus can be used to facilitate influenza vaccine strain selection. The webserver is available at http://sysbio.cvm.msstate.edu/AntigenMap.

  15. An efficient antigene activity and antiproliferative effect by targeting the Bcl-2 or survivin gene with triplex forming oligonucleotides containing a W-shaped nucleoside analogue (WNA-βT).

    Science.gov (United States)

    Taniguchi, Yosuke; Sasaki, Shigeki

    2012-10-01

    Triplex forming oligonucleotides (TFOs) are some of the most promising tools in the antigene strategy for the development of gene targeting therapeutics. However, the stable triplex formation is restricted to the homopurine sequences consisting of purine nucleosides, dG and dA. Therefore, the T or dC nucleoside in the homopurine strand inhibits the stable triplex formation. We have developed W-shaped nucleoside analogues (WNAs) for the formation of the unnatural type triplex DNA, with sequences containing the interrupting site in an antiparallel triplex formation. In the present study, we tested the antigene effect of TFOs having WNA-βT, which increased the stability of the triplex formation with a target sequence including the TA interrupting site. We designed the GU TFO (WNA) and GU TFO (natural) for targeting sequences of the Bcl-2 or survivin oncogene. The gel shift assay showed that the TFO (WNA) formed more stable triplexes than the natural TFO. Remarkably, the Bcl-2- or survivin-targeted TFO (WNA) inhibited the cell proliferation and induced a caspase-dependent apoptosis. It was confirmed that the survivin-targeted TFO (WNA) more effectively decreased the number of survivin products in the A549 cell than the natural TFOs. PMID:22987068

  16. P System antigenic determiners expression in Ascaris lumbricoides

    Directory of Open Access Journals (Sweden)

    Ponce De León Patricia

    2003-01-01

    Full Text Available The P System antigens have been detected in numerous parasites, bacterias and viruses, nevertheless the clinical significance is still unknown. The aim was to study the presence of P1 antigenic determiners in A. lumbricoides extracts by means of the use of 6 different monoclonal antibodies of well-known concentrations and Ig class. We worked with 14 A. lumbricoides extracts. Inhibition Agglutination Test was made in a bromelin enzymatic medium and 4 masculineC temperature. Titre, Score and Sensitivity Parameter were determined for each monoclonal antibody against red cells suspension used as revealing system. Ten extracts inhibited the agglutination of all anti P1 monoclonal antibodies. The 4 remaining extracts only inhibited the agglutination of some of them. It is demonstrated that the extracts have P1 activity. This activity is independent of titre, Score, Sensitivity Parameter, concentration and Ig class and it depends on the epitope at which the monoclonal antibody is directed.

  17. Virosomes for antigen and DNA delivery

    NARCIS (Netherlands)

    Daemen, T; de Mare, A; Bungener, L; de Jonge, J; Huckriede, A; Wilschut, J

    2005-01-01

    Specific targeting and delivery as well as the display of antigens on the surface of professional antigen-presenting cells (APCs) are key issues in the design and development of new-generation vaccines aimed at the induction of both humoral and cell-mediated immunity. Prophylactic vaccination agains

  18. Biological function and molecular mapping of M antigen in yeast phase of Histoplasma capsulatum.

    Directory of Open Access Journals (Sweden)

    Allan Jefferson Guimarães

    Full Text Available Histoplasmosis, due to the intracellular fungus Histoplasma capsulatum, can be diagnosed by demonstrating the presence of antibodies specific to the immunodominant M antigen. However, the role of this protein in the pathogenesis of histoplasmosis has not been elucidated. We sought to structurally and immunologically characterize the protein, determine yeast cell surface expression, and confirm catalase activity. A 3D-rendering of the M antigen by homology modeling revealed that the structures and domains closely resemble characterized fungal catalases. We generated monoclonal antibodies (mAbs to the protein and determined that the M antigen is present on the yeast cell surface and in cell wall/cell membrane preparations. Similarly, we found that the majority of catalase activity was in extracts containing fungal surface antigens and that the M antigen is not significantly secreted by live yeast cells. The mAbs also identified unique epitopes on the M antigen. The localization of the M antigen to the cell surface of H. capsulatum yeast and the characterization of the protein's major epitopes have important implications since it demonstrates that although the protein may participate in protecting the fungus against oxidative stress it is also accessible to host immune cells and antibody.

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

    Science.gov (United States)

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

    2012-01-01

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

  20. Protein antigen delivery by gene gun-mediated epidermal antigen incorporation (EAI).

    Science.gov (United States)

    Scheiblhofer, Sandra; Ritter, Uwe; Thalhamer, Josef; Weiss, Richard

    2013-01-01

    The gene gun technology can not only be employed for efficient transfer of gene vaccines into upper layers of the skin, but also for application of protein antigens. As a tissue rich in professional antigen presenting cells, the skin represents an attractive target for immunizations. In this chapter we present a method for delivery of the model antigen ovalbumin into the skin of mice termed epidermal antigen incorporation and describe in detail how antigen-specific proliferation in draining lymph nodes can be followed by flow cytometry.

  1. Adsorption of multimeric T cell antigens on carbon nanotubes

    DEFF Research Database (Denmark)

    Fadel, Tarek R; Li, Nan; Shah, Smith;

    2013-01-01

    Antigen-specific activation of cytotoxic T cells can be enhanced up to three-fold more than soluble controls when using functionalized bundled carbon nanotube substrates ((b) CNTs). To overcome the denaturing effects of direct adsorption on (b) CNTs, a simple but robust method is demonstrated...... to stabilize the T cell stimulus on carbon nanotube substrates through non-covalent attachment of the linker neutravidin....

  2. INNOVATIVE STRATEGIES TO IDENTIFY M. TUBERCULOSIS ANTIGENS AND EPITOPES USING GENOME-WIDE ANALYSES

    Directory of Open Access Journals (Sweden)

    Annemieke eGeluk

    2014-06-01

    Full Text Available In view of the fact that only a small part of the Mtb expressome has been explored for identification of antigens capable of activating human T-cell responses, which is critically required for the design of better TB vaccination strategies, more emphasis should be placed on innovative ways to discover new Mtb antigens and explore their function at the several stages of infection. Better protective antigens for TB vaccines are urgently needed, also in view of the disappointing results of the MVA85 vaccine which failed to induce additional protection in BCG vaccinated infants [54]. Moreover, immune responses to relevant antigens may be useful to identify TB-specific biomarker signatures. Here we describe the potency of novel tools and strategies to reveal such Mtb antigens. Using proteins specific for different Mtb infection phases, many new antigens of the latency-associated Mtb DosR regulon as well as Rpf proteins, associated with resuscitating TB, were discovered that were recognized by CD4+ and CD8+ T-cells. Furthermore, by employing MHC binding algorithms and bioinformatics combined with high throughput human T-cell screens and tetramers, HLA-class Ia restricted poly-functional CD8+ T-cells were identified in TB patients. Comparable methods, led to the identification of HLA-E-restricted Mtb epitopes recognized by CD8+ T-cells. A genome-wide unbiased antigen discovery approach was applied to analyse the in vivo Mtb gene expression profiles in the lungs of mice, resulting in the identification of IVE-TB antigens, which are expressed during infection in the lung, the main target organ of Mtb. IVE-TB antigens induce strong T cell responses in long-term latently Mtb infected individuals, and represent an interesting new group of TB antigens for vaccination. In summary, new tools have helped expand our view on the Mtb antigenome involved in human cellular immunity and provided new candidates for TB vaccination.

  3. O-antigen protects gram-negative bacteria from histone killing.

    Directory of Open Access Journals (Sweden)

    Catherine Chaput

    Full Text Available Beyond their traditional role of wrapping DNA, histones display antibacterial activity to Gram-negative and -positive bacteria. To identify bacterial components that allow survival to a histone challenge, we selected resistant bacteria from homologous Escherichia coli libraries that harbor plasmids carrying pieces of the chromosome in different sizes. We identified genes required for exopolysaccharide production and for the synthesis of the polysaccharide domain of the lipopolysaccharide, called O-antigen. Indeed, O-antigen and exopolysaccharide conferred further resistance to histones. Notably, O-antigen also conferred resistance to histones in the pathogens Shigella flexneri and Klebsiella pneumoniae.

  4. Expression of I-A and I-E,C region-coded Ia antigens on functional B cell subpopulations.

    Science.gov (United States)

    Frelinger, J A; Hibbler, F J; Hill, S W

    1978-12-01

    Ia antigens from specific subregions have been examined on functional B cell populations. Expression of both I-A and I-E,C region antigens was demonstrated on cells required for both lipopolysaccharide mitogenesis and polyclonal activation. Similar I-A and I-E,C subregion expression was found on cells required for response to the T-independent antigen, polyvinylpyrrolidone. TNP-specific IgM and hen egg lysozyme-specific IgG plaque-forming cells also express I-A and I-E,C region antigens. No evidence was found for an Ia- population responsive in the systems tested. Further, no evidence of preferential expression of I-A or I-E,C region antigens was observed in any system examined. Therefore, it appears that B cells express both I-A and I-E,C region-coded Ia antigens.

  5. Selective expression and immunogenicity of the cancer/testis antigens SP17, AKAP4 and PTTG1 in non-small cell lung cancer: new candidates for active immunotherapy.

    Science.gov (United States)

    Chiriva-Internati, Maurizio; Mirandola, Leonardo; Figueroa, Jose A; Yu, Yuefei; Grizzi, Fabio; Kim, Minji; Jenkins, Marjorie; Cobos, Everardo; Jumper, Cynthia; Alalawi, Raed

    2014-05-01

    ABSTRACT BACKGROUND. Lung cancer is the leading cause of cancer deaths in both genders worldwide, with an incidence only second to prostate cancer in men and breast cancer in women. The lethality of the disease highlights the urgent need for innovative therapeutic options. Immunotherapy can afford efficient and specific targeting of tumor cells, improving efficacy and reducing the side effects of current therapies. We have previously reported the aberrant expression of cancer/testis antigens (CTAs) in tumors of unrelated histological origin. In this study we investigated the expression and immunogenicity of the cancer/testis antigens (CTAs) Sperm Protein 17 (SP17), A-kinase anchor protein 4 (AKAP4) and Pituitary Tumor Transforming Gene 1 (PTTG1) in human non-small cell lung cancer (NSCLC) cell lines and primary tumors. METHODS. We used RT-PCR, immunofluorescence, flow cytometry, ELISA and cytotoxicity assays to determine the expression levels and immunogenicity of SP17, AKAP4 and PTTG1 in human NSCLC cell lines and primary tumors. RESULTS. We found that SP17, AKAP4 and PTTG1 are aberrantly expressed in NSCLC cancer cell lines and primary tumor tissues from patients, compared to normal lung cell lines and tissues. We established the immunogenicity of these CTAs by measuring CTA-specific autoantibodies in patients' sera and generating CTA-specific autologous cytotoxic lymphocytes (CTLs) from patients' peripheral blood mononuclear cells (PBMCs). CONCLUSIONS. Our results provide proof of principle that the CTAs SP17/AKAP4/PTTG1 are expressed in both human NSCLC cell lines and primary tumors and can elicit an immunogenic response in NSCLC patients. Based on our findings, further studies are warranted to explore the feasibility of developing CTA-specific immunotherapeutic strategies for NSCLC patients. PMID:24811938

  6. Further characterization of filarial antigens by SDS polyacrylamide gel electrophoresis

    OpenAIRE

    Dissanayake, S.; Galahitiyawa, S. C.; Ismail, M. M.

    1983-01-01

    SDS (sodium dodecyl sulfate)-polyacrylamide gel electrophoresis of an antigen isolated from sera of Wuchereria bancrofti-infected patients and Setaria digitata antigen SD2-4 is reported. Both antigens showed carbohydrate (glycoprotein) staining. The W. bancrofti antigen had an apparent relative molecular mass of 35 000 while the S. digitata antigen SD2-4 migrated at the marker dye position on SDS-polyacrylamide gel electrophoresis. SDS treatment of these antigens did not abolish the precipita...

  7. Enhancement of DNA vaccine potency through linkage of antigen to filamentous bacteriophage coat protein III domain I

    DEFF Research Database (Denmark)

    Cuesta, Àngel M; Suárez, Eduardo; Larsen, Martin;

    2006-01-01

    immune pathways by adding immune-activating genes to the tumour antigen sequence. In this work, we converted a model non-immunogenic antigen into a vaccine by fusing it to domain I of the filamentous bacteriophage coat protein III gene. Vaccination with a DNA construct encoding the domain I fusion...

  8. Neurofibromatosis type 2 tumor suppressor protein, NF2, induces proteasome-mediated degradation of JC virus T-antigen in human glioblastoma.

    Directory of Open Access Journals (Sweden)

    Sarah Beltrami

    Full Text Available Neurofibromatosis type 2 protein (NF2 has been shown to act as tumor suppressor primarily through its functions as a cytoskeletal scaffold. However, NF2 can also be found in the nucleus, where its role is less clear. Previously, our group has identified JC virus (JCV tumor antigen (T-antigen as a nuclear binding partner for NF2 in tumors derived from JCV T-antigen transgenic mice. The association of NF2 with T-antigen in neuronal origin tumors suggests a potential role for NF2 in regulating the expression of the JCV T-antigen. Here, we report that NF2 suppresses T-antigen protein expression in U-87 MG human glioblastoma cells, which subsequently reduces T-antigen-mediated regulation of the JCV promoter. When T-antigen mRNA was quantified, it was determined that increasing expression of NF2 correlated with an accumulation of T-antigen mRNA; however, a decrease in T-antigen at the protein level was observed. NF2 was found to promote degradation of ubiquitin bound T-antigen protein via a proteasome dependent pathway concomitant with the accumulation of the JCV early mRNA encoding T-antigen. The interaction between T-antigen and NF2 maps to the FERM domain of NF2, which has been shown previously to be responsible for its tumor suppressor activity. Co-immunoprecipitation assays revealed a ternary complex among NF2, T-antigen, and the tumor suppressor protein, p53 within a glioblastoma cell line. Further, these proteins were detected in various degrees in patient tumor tissue, suggesting that these associations may occur in vivo. Collectively, these results demonstrate that NF2 negatively regulates JCV T-antigen expression by proteasome-mediated degradation, and suggest a novel role for NF2 as a suppressor of JCV T-antigen-induced cell cycle regulation.

  9. Tumor antigen specific iTreg accumulate in the tumor microenvironment and suppress therapeutic vaccination

    OpenAIRE

    Schreiber, Taylor H; Wolf, Dietlinde; Bodero, Maria; Podack, Eckhard

    2012-01-01

    Tumor specific antigens (TSA) provide an opportunity to mobilize therapeutic immune responses against cancer. To evade such responses, tumor development in immunocompetent hosts is accompanied by acquisition of both active and passive mechanisms of immune suppression, including recruitment of CD4+FoxP3+ regulatory T cells (Treg). Thymic derived Treg (nTreg) may recognize self-antigens in the tumor microenvironment, while peripherally induced Treg (iTreg) may preferentially recognize the same ...

  10. Characterization of the Cryptosporidium antigens from sporulated oocysts of Cryptosporidium parvum.

    OpenAIRE

    Luft, B J; Payne, D.; Woodmansee, D; Kim, C. W.

    1987-01-01

    The antigenic constituents of sporulated Cryptosporidium parvum oocyst antigens were characterized with antisera from mice immunized against C. parvum. Sodium dodecyl sulfate-polyacrylamide gel electrophoresis followed by silver staining defined the major proteins. Six of seven lectins used recognized as many as 15 bands. The lectins concanavalin A, Dolichos biflorus, and wheat germ agglutinin showed strong activity against the same eight bands with molecular weights ranging from 72,000 to gr...

  11. Meningococcal vaccine antigen diversity in global databases.

    Science.gov (United States)

    Brehony, Carina; Hill, Dorothea M; Lucidarme, Jay; Borrow, Ray; Maiden, Martin C

    2015-01-01

    The lack of an anti-capsular vaccine against serogroup B meningococcal disease has necessitated the exploration of alternative vaccine candidates, mostly proteins exhibiting varying degrees of antigenic variation. Analysis of variants of antigen-encoding genes is facilitated by publicly accessible online sequence repositories, such as the Neisseria PubMLST database and the associated Meningitis Research Foundation Meningococcus Genome Library (MRF-MGL). We investigated six proposed meningococcal vaccine formulations by deducing the prevalence of their components in the isolates represented in these repositories. Despite high diversity, a limited number of antigenic variants of each of the vaccine antigens were prevalent, with strong associations of particular variant combinations with given serogroups and genotypes. In the MRF-MGL and globally, the highest levels of identical sequences were observed with multicomponent/multivariant vaccines. Our analyses further demonstrated that certain combinations of antigen variants were prevalent over periods of decades in widely differing locations, indicating that vaccine formulations containing a judicious choice of antigen variants have potential for long-term protection across geographic regions. The data further indicated that formulations with multiple variants would be especially relevant at times of low disease incidence, as relative diversity was higher. Continued surveillance is required to monitor the changing prevalence of these vaccine antigens. PMID:26676305

  12. Diminished Memory T-Cell Expansion Due to Delayed Kinetics of Antigen Expression by Lentivectors.

    Directory of Open Access Journals (Sweden)

    Karina Furmanov

    Full Text Available Memory CD8(+ T lymphocytes play a central role in protective immunity. In attempt to increase the frequencies of memory CD8(+ T cells, repeated immunizations with viral vectors are regularly explored. Lentivectors have emerged as a powerful vaccine modality with relatively low pre-existing and anti-vector immunity, thus, thought to be ideal for boosting memory T cells. Nevertheless, we found that lentivectors elicited diminished secondary T-cell responses that did not exceed those obtained by priming. This was not due to the presence of anti-vector immunity, as limited secondary responses were also observed following heterologous prime-boost immunizations. By dissecting the mechanisms involved in this process, we demonstrate that lentivectors trigger exceptionally slow kinetics of antigen expression, while optimal activation of lentivector-induced T cells relays on durable expression of the antigen. These qualities hamper secondary responses, since lentivector-encoded antigen is rapidly cleared by primary cytotoxic T cells that limit its presentation by dendritic cells. Indeed, blocking antigen clearance by cytotoxic T cells via FTY720 treatment, fully restored antigen presentation. Taken together, while low antigen expression is expected during secondary immunization with any vaccine vector, our results reveal that the intrinsic delayed expression kinetics of lentiviral-encoded antigen, further dampens secondary CD8(+ T-cell expansion.

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

    Science.gov (United States)

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

    2012-09-21

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

  14. Antigen incorporation on Cryptosporidium parvum oocyst walls

    Directory of Open Access Journals (Sweden)

    Entrala Emilio

    2001-01-01

    Full Text Available Cryptosporidium parvum oocysts are the infective stages responsible for transmission and survival of the organism in the environment. In the present work we show that the oocyst wall, far from being a static structure, is able to incorporate antigens by a mechanism involving vesicle fusion with the wall, and the incorporation of the antigen to the outer oocyst wall. Using immunoelectron microscopy we show that the antigen recognized by a monoclonal antibody used for diagnosis of cryptosporidiosis (Merifluor®, Meridian Diagnostic Inc. could be found associated with vesicles in the space between the sporozoites and the oocysts wall, and incorporated to the outer oocyst wall by an unknown mechanism.

  15. A rapid radioimmunoassay for determination of tumor antigens with reference to carcinoembryonic antigen

    International Nuclear Information System (INIS)

    A novel experimental procedure for the determination of carcinoembryonic antigen (CEA) by a solid phase microradioimmunoassay has been developed. Goat anti-CEA antibodies were immobilized by coupling to cyanogen bromide activated filter paper discs. The inhibition of binding of radioiodinated CEA to the discs was proportional to the amount of unlabelled CEA present in the test sample. The experimental procedure involved two steps: (i) the test material containing unlabelled CEA was allowed to react with the antibody coated discs at 37 deg C for 2 hrs., and (ii) a standard amount of 125I-CEA was added to the reaction mixture and incubated for 24 hours at 37 deg C or room temperature. The discs were then washed 4 times and the radioactivity of each disc was determined. The sensitivity of the test in its present state of development was 2.5 ng/ml. (author)

  16. Hepatitis B virus antigens impair NK cell function.

    Science.gov (United States)

    Yang, Yinli; Han, Qiuju; Zhang, Cai; Xiao, Min; Zhang, Jian

    2016-09-01

    An inadequate immune response of the host is thought to be a critical factor causing chronic hepatitis B virus (CHB) infection. Natural killer (NK) cells, as one of the key players in the eradication and control of viral infections, were functionally impaired in CHB patients, which might contribute to viral persistence. Here, we reported that HBV antigens HBsAg and HBeAg directly inhibited NK cell function. HBsAg and/or HBeAg blocked NK cell activation, cytokine production and cytotoxic granule release in human NK cell-line NK-92 cells, which might be related to the downregulation of activating receptors and upregulation of inhibitory receptor. Furthermore, the underlying mechanisms likely involved the suppression of STAT1, NF-κB and p38 MAPK pathways. These findings implicated that HBV antigen-mediated inhibition of NK cells might be an efficient strategy for HBV evasion, targeting the early antiviral responses mediated by NK cells and resulting in the establishment of chronic virus infection. Therefore, this study revealed the relationship between viral antigens and human immune function, especially a potential important interaction between HBV and innate immune responses. PMID:27341035

  17. Conjugating influenza a (H1N1) antigen to n-trimethylaminoethylmethacrylate chitosan nanoparticles improves the immunogenicity of the antigen after nasal administration.

    Science.gov (United States)

    Liu, Qingfeng; Zheng, Xiaoyao; Zhang, Chi; Shao, Xiayan; Zhang, Xi; Zhang, Qizhi; Jiang, Xinguo

    2015-11-01

    As one of the most serious infectious respiratory diseases, influenza A (H1N1) is a great threat to human health, and it has created an urgent demand for effective vaccines. Nasal immunization can induce both systemic and mucosal immune responses against viruses, and it can serve as an ideal route for vaccination. However, the low immunogenicity of antigens on nasal mucosa is a high barrier for the development of nasal vaccines. In this study, we covalently conjugated an influenza A (H1N1) antigen to the surface of N-trimethylaminoethylmethacrylate chitosan (TMC) nanoparticles (H1N1-TMC/NP) through thioester bonds to increase the immunogenicity of the antigen after nasal administration. SDS-PAGE revealed that most of the antigen was conjugated on TMC nanoparticles, and an in vitro biological activity assay confirmed the stability of the antigen after conjugation. After three nasal immunizations, the H1N1-TMC/NP induced significantly higher levels of serum IgG and mucosal sIgA compared with free antigen. A hemagglutination inhibition assay showed that H1N1-TMC/NP induced much more protective antibodies than antigen-encapsulated nanoparticles or alum-precipitated antigen (I.M.). In the mechanistic study, H1N1-TMC/NP was shown to stimulate macrophages to produce IL-1β and IL-6 and to stimulate spleen lymphocytes to produce IL-2 and IFN-γ. These results indicated that H1N1-TMC/NP may be an effective vaccine against influenza A (H1N1) viruses for use in nasal immunization.

  18. Targeting tumor antigens to secreted membrane vesicles in vivo induces efficient antitumor immune responses.

    NARCIS (Netherlands)

    Zeelenberg, I.S.; Ostrowski, M.; Krumeich, S.; Bobrie, A.; Jancic, C.; Boissonnas, A.; Delcayre, A.; Pecq, JB Le; Combadiere, B.; Amigorena, S.; Thery, C.

    2008-01-01

    Expression of non-self antigens by tumors can induce activation of T cells in vivo, although this activation can lead to either immunity or tolerance. CD8+ T-cell activation can be direct (if the tumor expresses MHC class I molecules) or indirect (after the capture and cross-presentation of tumor an

  19. [ELISA method for the determination of factor VII antigen].

    Science.gov (United States)

    Jorquera, J I; Aznar, J A; Monteagudo, J; Montoro, J M; Casaña, P; Pascual, I; Bañuls, E; Curats, R; Llopis, F

    1989-12-01

    The low plasma concentration of clotting factor VII makes it difficult to assay its antigenic fraction by the conventional methods of precipitation with specific antigens. Simple and peroxidase-conjugated antisera are currently available from commercial sources, thus allowing one to determine F VII:Ag by enzyme immunoassay. An ELISA method has been developed in this laboratory which provides sensitivity limits about 0.1% of the plasma concentration of F VII and correlates significantly with its functional activity (r = 0.603, n = 44, p less than 0.001). This technique can be highly helpful in characterising molecular variants of F VII, as well as in detecting acquired deficiencies of this factor.

  20. Metal based nanoparticles as cancer antigen delivery vehicles for macrophage based antitumor vaccine.

    Science.gov (United States)

    Chattopadhyay, Sourav; Dash, Sandeep Kumar; Mandal, Debasis; Das, Balaram; Tripathy, Satyajit; Dey, Aditi; Pramanik, Panchanan; Roy, Somenath

    2016-02-10

    In the present study, we would like to evaluate the efficacy of modified metal oxide nanoparticles (NPs) as cancer antigen delivery vehicles for macrophage (MФs) based antitumor vaccine. The cobalt oxide nanoparticles (CoO NPs) were promising tools for delivery of antigens to antigen presenting cells and have induced an antitumor immune response. Synthesized CoO NPs were modified by N-phosphonomethyliminodiacetic acid (PMIDA), facilitated the conjugation of lysate antigen, i.e. cancer antigen derived from lysis of cancer cells. The cancer cell lysate antigen conjugated PMIDA-CoO NPs (Ag-PMIDA-CoO NPs) successfully activated macrophage (MФ) evident by the increasing the serum IFN-γ and TNF-α level. Immunization of mice with the Ag-PMIDA-CoO NPs constructed an efficient immunological adjuvant induced anticancer IgG responses, and increased the antibody dependent cellular cytotoxicity (ADCC) response than only lysate antigen treated group to combat the cancer cell. The nanocomplexes enhanced the anticancer CD4(+)T cell response in mice. The result showed that Ag-PMIDA-CoO NPs can stimulate the immune responses over only lysate antigens, which are the most important findings in this study. These NP-mediated Ag deliveries may significantly improve the anticancer immune response by activating MФs and may act as adjuvant and will balance the pro-inflammatory and anti-inflammatory immunoresponse. The crosstalk between the activated MФ with other immune competent cells will be monitored by measuring the cytokines which illustrate the total immunological network setups.

  1. Proliferating cell nuclear antigen in neutrophil fate.

    Science.gov (United States)

    Witko-Sarsat, Véronique; Ohayon, Delphine

    2016-09-01

    The life span of a neutrophil is a tightly regulated process as extended survival is beneficial for pathogen elimination and cell death necessary to prevent cytotoxic content release from activated neutrophils at the inflammatory site. Therefore, the control between survival and death must be a dynamic process. We have previously described that proliferating cell nuclear antigen (PCNA) which is known as a nuclear protein pivotal in DNA synthesis, is a key element in controlling neutrophil survival through its association with procaspases. Contrary to the dogma which asserted that PCNA has a strictly nuclear function, in mature neutrophils, PCNA is present exclusively within the cytosol due to its nuclear export at the end of the granulocytic differentiation. More recent studies are consistent with the notion that the cytosolic scaffold of PCNA is aimed at modulating neutrophil fate rather than simply preventing death. Ultimately, targeting neutrophil survival might have important applications not just in the field of immunology and inflammation, but also in hematology and transfusion. The neutrophil emerges as a unique and powerful cellular model to unravel the basic mechanisms governing the cell cycle-independent functions of PCNA and should be considered as a leader of the pack. PMID:27558345

  2. CD8+ T cell priming by dendritic cell vaccines requires antigen transfer to endogenous antigen presenting cells.

    Directory of Open Access Journals (Sweden)

    Alice W Yewdall

    Full Text Available Immunotherapeutic strategies to stimulate anti-tumor immunity are promising approaches for cancer treatment. A major barrier to their success is the immunosuppressive microenvironment of tumors, which inhibits the functions of endogenous dendritic cells (DCs that are necessary for the generation of anti-tumor CD8+ T cells. To overcome this problem, autologous DCs are generated ex vivo, loaded with tumor antigens, and activated in this non-suppressive environment before administration to patients. However, DC-based vaccines rarely induce tumor regression.We examined the fate and function of these DCs following their injection using murine models, in order to better understand their interaction with the host immune system. Contrary to previous assumptions, we show that DC vaccines have an insignificant role in directly priming CD8+ T cells, but instead function primarily as vehicles for transferring antigens to endogenous antigen presenting cells, which are responsible for the subsequent activation of T cells.This reliance on endogenous immune cells may explain the limited success of current DC vaccines to treat cancer and offers new insight into how these therapies can be improved. Future approaches should focus on creating DC vaccines that are more effective at directly priming T cells, or abrogating the tumor induced suppression of endogenous DCs.

  3. Anti-tumor effect of CTLs activated by dendritic cells pulsed with K-ras mutant peptide and whole tumor antigen on pancreatic cancer%K-ras突变多肽与全细胞抗原致敏DCs诱导CTLs对胰腺癌的杀伤活性研究

    Institute of Scientific and Technical Information of China (English)

    Guang Tan; Zhongyu Wang; Xin Zhang; Zhengang Cai; Junkai Zhang

    2010-01-01

    Objective:We studied the role of specific cytotoxic T lymphocytes(CTLs)activated by dendritic cells(DCs)presenting cationic nanoparticles with the K-ras(12-Val)mutant peptide and whole tumor antigen in the killing of different pancreatic cancer cell lines in vitro and in vitro.Methods:Peripheral blood DCs were induced by rhGM-CSF and IL-4 and cultured.DCs were sensitized by whole antigen of a pancreatic cancer cell line(PANC-1)with expression of K-ras mutant,K-ras mutant peptide(K-ras+peptide)and cationic nanoparticles with K-ras mutant peptide(K-ras+peptide-CNP),respectively.Cell surface markers were measured by flow cytometry.Lymphocyte proliferation was detected by the 3H-TdR test,and ELISA was performed to detect IFN-γ secretion.125I-UdR was used to measure the killing effect of CTLs.We also evaluated the antitumor activity of CTLs in vivo in a tumor-bearing nude mouse model prepared with the PANC-1(K-ras+)and SW1990(K-ras-)cell lines.Results:Compared with K-ras+peptide,low concentration K-ras+peptide-CNP can be effectively presented by DCs(P0.05)on SW1990 cell lines(P>0.05).Conclusion:Cationic nanoparticles with K-res(12-Val)mutant peptide can be effectively presented by DCs at a low concentration in a short time.CTLs induced by K-ras+peptide-CNP had specific killing activity for the pancreatic cancer cell line with the K-ras(12-Val)mutant and could significantly inhibit tumor growth and increase the survival time of tumor-bearing nude mice.

  4. Humoral response to low molecular weight antigens of Mycobacterium tuberculosis by tuberculosis patients and contacts

    Directory of Open Access Journals (Sweden)

    Beck S.T.

    2005-01-01

    Full Text Available Much effort has been devoted to the identification of immunologically important antigens of Mycobacterium tuberculosis and to the combination of target antigens to which antibodies from serum of tuberculous patients could react specifically. We searched for IgG antibodies specific for antigens of 45 to 6 kDa obtained after sonication of the well-characterized wild M. tuberculosis strain in order to detect differences in the antibody response to low molecular weight antigens from M. tuberculosis between patients with pulmonary tuberculosis and contacts. Specific IgG antibodies for these antigens were detected by Western blot analysis of 153 serum samples collected from 51 patients with confirmed pulmonary tuberculosis. Three samples were collected from each patient: before therapy, and after 2 and 6 months of treatment. We also analyzed 25 samples obtained from contacts, as well as 30 samples from healthy individuals with known tuberculin status, 50 samples from patients with other lung diseases and 200 samples from healthy blood donors. The positive predictive value for associated IgG reactivity against the 6-kDa and 16-kDa antigens, 6 and 38 kDa, and 16 and 38 kDa was 100% since simultaneous reactivity for these antigens was absent in healthy individuals and individuals with other lung diseases. This association was observed in 67% of the patients, but in only 8% of the contacts. The humoral response against antigens of 16 and 6 kDa seems to be important for the detection of latent tuberculosis since the associated reactivity to these antigens is mainly present in individuals with active disease.

  5. Prostate-specific antigen (PSA) blood test

    Science.gov (United States)

    Prostate-specific antigen; Prostate cancer screening test; PSA ... PSA testing is an important tool for detecting prostate cancer, but it is not foolproof. Other conditions can cause a rise in PSA, including: A larger prostate ...

  6. Mapping Epitopes on a Protein Antigen by the Proteolysis of Antigen-Antibody Complexes

    Science.gov (United States)

    Jemmerson, Ronald; Paterson, Yvonne

    1986-05-01

    A monoclonal antibody bound to a protein antigen decreases the rate of proteolytic cleavage of the antigen, having the greatest effect on those regions involved in antibody contact. Thus, an epitope can be identified by the ability of the antibody to protect one region of the antigen more than others from proteolysis. By means of this approach, two distinct epitopes, both conformationally well-ordered, were characterized on horse cytochrome c.

  7. Tales of Antigen Evasion from CAR Therapy.

    Science.gov (United States)

    Sadelain, Michel

    2016-06-01

    Both T cells bearing chimeric antigen receptors and tumor-specific antibodies can successfully target some malignancies, but antigen escape can lead to relapse. Two articles in this issue of Cancer Immunology Research explore what effective countermeasures may prevent it. Cancer Immunol Res; 4(6); 473-473. ©2016 AACRSee articles by Zah et al., p. 498, and Rufener et al., p. 509. PMID:27252092

  8. Characterization of an antigenically distinct porcine rotavirus.

    OpenAIRE

    Bridger, J C; Clarke, I. N.; McCrae, M A

    1982-01-01

    A porcine virus with rotavirus morphology, which was antigenically unrelated to previously described rotaviruses, is described. Particles with an outer capsid layer measured 75 nm and those lacking the outer layer were 63 nm in diameter. Particles which resembled cores were also identified. The virus was shown to be antigenically distinct from other rotaviruses as judged by immunofluorescence and immune electron microscopy, and it failed to protect piglets from challenge with porcine rotaviru...

  9. Defining the antigenic diversity of Plasmodium falciparum apical membrane antigen 1 and the requirements for a multi-allele vaccine against malaria.

    Directory of Open Access Journals (Sweden)

    Damien R Drew

    Full Text Available Apical Membrane Antigen 1 (AMA1 is a leading malaria vaccine candidate and a target of naturally-acquired human immunity. Plasmodium falciparum AMA1 is polymorphic and in vaccine trials it induces strain-specific protection. This antigenic diversity is a major roadblock to development of AMA1 as a malaria vaccine and understanding how to overcome it is essential. To assess how AMA1 antigenic diversity limits cross-strain growth inhibition, we assembled a panel of 18 different P. falciparum isolates which are broadly representative of global AMA1 sequence diversity. Antibodies raised against four well studied AMA1 alleles (W2Mef, 3D7, HB3 and FVO were tested for growth inhibition of the 18 different P. falciparum isolates in growth inhibition assays (GIA. All antibodies demonstrated substantial cross-inhibitory activity against different isolates and a mixture of the four different AMA1 antibodies inhibited all 18 isolates tested, suggesting significant antigenic overlap between AMA1 alleles and limited antigenic diversity of AMA1. Cross-strain inhibition by antibodies was only moderately and inconsistently correlated with the level of sequence diversity between AMA1 alleles, suggesting that sequence differences are not a strong predictor of antigenic differences or the cross-inhibitory activity of anti-allele antibodies. The importance of the highly polymorphic C1-L region for inhibitory antibodies and potential vaccine escape was assessed by generating novel transgenic P. falciparum lines for testing in GIA. While the polymorphic C1-L epitope was identified as a significant target of some growth-inhibitory antibodies, these antibodies only constituted a minor proportion of the total inhibitory antibody repertoire, suggesting that the antigenic diversity of inhibitory epitopes is limited. Our findings support the concept that a multi-allele AMA1 vaccine would give broad coverage against the diversity of AMA1 alleles and establish new tools to

  10. The potential for induction of autoimmune disease by a randomly-mutated self-antigen

    DEFF Research Database (Denmark)

    Pedersen, Anders Elm

    2007-01-01

    The pathology of most autoimmune diseases is well described. However, the exact event that triggers the onset of the inflammatory cascade leading to disease is less certain and most autoimmune diseases are complex idiopathic diseases with no single gene known to be causative. In many cases, a...... relation to an infectious disease is described, and it is thought that microbes can play a direct role in induction of autoimmunity, for instance by molecular mimicry or bystander activation of autoreactive T cells. In contrast, less attention has been given to the possibility that modified self......-antigens can be immunogenic and lead to autoimmunity against wildtype self-antigens. In theory, modified self-antigens can arise by random errors and mutations during protein synthesis and would be recognized as foreign antigens by naïve B and T lymphocytes. Here, it is postulated that the initial auto...

  11. PATHWAYS OF RECOGNITION OF FOREIGN ANTIGENS IN THE ADAPTIVE IMMUNE RESPONSE TO ALLOGENEIC ORGAN TRANSPLANTATION

    Directory of Open Access Journals (Sweden)

    A. S. Berkos

    2015-01-01

    Full Text Available Immunological allograft rejection is based on the process of recognition of donor antigens by T-cell receptors. Antigens are generally represented by molecules of major hystocompatibility complex (MHC, expressed by the donor organ tissue cells. Immune response to allogeneic transplant in its activity is greatly higher than response to conventional infection wherein the basic mechanisms of recognition of infectious antigens and alloantigens seem to be similar. In this review we have analyzed three main pathways of recognition of alloantigens. Those pathways are differentiated by the nature of antigen presenting cells, kinetics and duration of its influence on the formation of alloimmunity: direct, indirect and semidirect. Establishing the relationship between mechanisms of allorecognition and manifestations of graft rejection is of great clinical importance. 

  12. Design, automated synthesis and immunological evaluation of NOD2-ligand–antigen conjugates

    Directory of Open Access Journals (Sweden)

    Marian M. J. H. P. Willems

    2014-06-01

    Full Text Available The covalent attachment of an innate immune system stimulating agent to an antigen can provide active vaccine modalities capable of eliciting a potent immune response against the incorporated antigen. Here we describe the design, automated synthesis and immunological evaluation of a set of four muramyl dipeptide–peptide antigen conjugates. Muramyl dipeptide (MDP represents a well-known ligand for the intracellular NOD2 receptor and our study shows that covalently linking an MDP-moiety to an antigenic peptide can lead to a construct that is capable of stimulating the NOD2 receptor if the ligand is attached at the anomeric center of the muramic acid. The constructs can be processed by dendritic cells (DCs and the conjugation does not adversely affect the presentation of the incorporated SIINFEKL epitope on MHC-I molecules. However, stimulation of the NOD2 receptor in DCs was not sufficient to provide a strong immunostimulatory signal.

  13. Features of glycoproteins distribution in the pancreatic structures of newborn rats after prenatal antigenic influence

    Directory of Open Access Journals (Sweden)

    Grinivetska N.V.

    2014-03-01

    Full Text Available Background. Abnormalities of digestion and absorption are the most common syndromes associated with the digestive system diseases in children. Pancreatic enzyme failure leads to violation of different metabolic processes, especially in neonates. Exocrine pancreas is sensitive to a variety of factors, including virus. It emphasizes the importance of investigation of pancreatic secretory activity in children who was born from mothers exposed to viral infection during pregnancy. Objective. The purpose is to determine the features of glycoproteins distribution in the pancreatic structures of newborn rats after prenatal antigenic influence. Methods. Animals were divided into four groups: the 1st – intact, the 2nd – intrafetal injection of the antigen, the 3rd – animals administered with antigen into the amniotic fluid, and the 4th group – control (intrafetal injection of a normal salt solution. As antigen we used Vaxigrip vaccine. Results. It was revealed that on the14th day after birth antigen-exposed animals were characterized by the increase of glycoproteins (++ in a connective tissue capsule of pancreas and decreased content of glycoproteins in the cytoplasm of acinar cells and ducts comparing with the intact group (++/+. Taken together this data evidence the reduction of the synthetic activity of acinar cells after intrafetal administration of the antigen. This may cause a predisposition for the development of dyspepsia and food allergy. Further work is planned to investigate the dynamics of glycosaminoglycans redistribution in different parts of pancreas after prenatal antigenic administrations. Citation: Grinivetska NV. [Features of glycoproteins distribution in the pancreatic structures of newborn rats after prenatal antigenic influence]. Morphologia. 2014;8(1:30-4. Ukrainian.

  14. Immunosensor Based on Surface Plasmon Resonance for Antigen Recognition

    Institute of Scientific and Technical Information of China (English)

    2008-01-01

    A novel immunosensor based on surface plasmon resonance(SPR)has been developed for the recognition of antigen.The sensor was designed on the basis of the fixed angle of incidence and measuring the reflected intensities in a wavelength range of 430-750 nm in real-time. An ultra-bright white light-emitting diode(LED)was used as the light source. Molecular self-assembling in solution was used to form the sensing membrane on gold substrate. It has been seen that the sensitivity of the SPR sensor with 3-mercaptopropionic acid(MPA)/protein A(SPA) sensing membrane is considerably higher than that with MPA or SPA modified Sensing membrane. The kinetic processes on the sensing membrane were studied. The human B factor(Bf), an activator of complement 3(C3), was recognized among the other antigens. This sensor can also be used for other antigen/antibody or adaptor/receptor recognition. Under optimized experimental conditions, the sensor has good selectivity, repeatability, and reversibility.

  15. Transplant immuno-diagnostics: crossmatch and antigen detection.

    Science.gov (United States)

    South, Andrew M; Grimm, Paul C

    2016-06-01

    Identifying and monitoring donor-directed anti-human leukocyte antigen antibodies are a rapidly evolving area of solid organ transplantation. Donor-specific antibodies dictate pre-transplant donor choice and donor-recipient matching and underlie much acute and chronic allograft rejection and loss. The evolution of available technology has driven this progress. Early, labor-intensive, whole-cell assays based on complement-dependent cytotoxicity suffered from poor sensitivity and specificity, technical challenges and lack of precision. Sequential improvement in assay performance included anti-human immunoglobulin-enhanced, complement-dependent cytotoxicity techniques followed by cell-based flow cytometry. However, variable specificity and sensitivity inherent in cell-based testing continued to limit flow cytometry. The introduction of solid-phase assays led to a second revolution in histocompatibility testing with the use of purified antigens bound to artificial surfaces rather than whole cells. These techniques augmented sensitivity and specificity to detect even low-titer antibodies to previously undetected antigens. Identification of complement-activating antibodies is being introduced, but current technology is in the developmental stage. While the detection of alloantibodies has improved dramatically, our comprehension of their importance remains imperfect. Variability in methodology and a lack of standardization limits the clinical application of these tests. In spite of the hurdles that remain, antibody-mediated rejection has become a key target to improve graft survival. PMID:26139577

  16. Gold nanoparticle-based fluorescence immunoassay for malaria antigen detection.

    Science.gov (United States)

    Guirgis, Bassem S S; Sá e Cunha, Cláudia; Gomes, Inês; Cavadas, Miguel; Silva, Isabel; Doria, Gonçalo; Blatch, Gregory L; Baptista, Pedro V; Pereira, Eulália; Azzazy, Hassan M E; Mota, Maria M; Prudêncio, Miguel; Franco, Ricardo

    2012-01-01

    The development of rapid detection assays for malaria diagnostics is an area of intensive research, as the traditional microscopic analysis of blood smears is cumbersome and requires skilled personnel. Here, we describe a simple and sensitive immunoassay that successfully detects malaria antigens in infected blood cultures. This homogeneous assay is based on the fluorescence quenching of cyanine 3B (Cy3B)-labeled recombinant Plasmodium falciparum heat shock protein 70 (PfHsp70) upon binding to gold nanoparticles (AuNPs) functionalized with an anti-Hsp70 monoclonal antibody. Upon competition with the free antigen, the Cy3B-labeled recombinant PfHsp70 is released to solution resulting in an increase of fluorescence intensity. Two types of AuNP-antibody conjugates were used as probes, one obtained by electrostatic adsorption of the antibody on AuNPs surface and the other by covalent bonding using protein cross-linking agents. In comparison with cross-linked antibodies, electrostatic adsorption of the antibodies to the AuNPs surfaces generated conjugates with increased activity and linearity of response, within a range of antigen concentration from 8.2 to 23.8 μg.mL(-1). The estimated LOD for the assay is 2.4 μg.mL(-1) and the LOQ is 7.3 μg.mL(-1). The fluorescence immunoassay was successfully applied to the detection of antigen in malaria-infected human blood cultures at a 3% parasitemia level, and is assumed to detect parasite densities as low as 1,000 parasites.μL(-1).

  17. 肺癌相关抗原RNA转染树突状细胞诱导细胞因子诱导的杀伤细胞杀伤活力的体外研究%Study of lung cancer associa ted antigen RNA-transfected DC-induced cell killing activity of CIK in vitro

    Institute of Scientific and Technical Information of China (English)

    刘伯轩; 戴北鸿; 向启德; 廖东承; 高细强; 吴玉兰; 周频

    2015-01-01

    Objective To investigate the NCI-H1975 human lung adenocarcinoma cell line associated antigen RNA-transfected human dendritic cells(DCs) in vitro cytokine-induced killer(CIK) activity induced killer cells to cancer cells. Methods Using blood cell apheresis machine collecting peripheral blood mononuclear cells (PBMCs) and using mononuclear cells by density gradient centrifugation purified, cell culture flasks adherent mononuclear cells, adding recombinant human granulocyte-macrophage colony-stimulating factor(rhGM-CSF) and recombinant human interleukin -4 (rhIL-4) to induce immature DCs, extraction of total RNA NCI-H1975 cells were transfected DCs induced to mature DCs. After transfection induce mature DCs and PBMCs cultured CIK cells induced mixed culture , mixed culture with target cells again to observe the destruction of vitality. The experimental points associated antigen RNA-transfected DC-CIK group of lung cancer, non-transfected DC-CIK group and transfected with empty liposomes DC-CIK group were detected by flow cytometry analysis DC and CIK cell surface antigen expression , using CSFE/PI double staining was used to detect differences in anti-tumor activity of the three groups. Results RNA-associated antigen transfected group, DC untransfected group and transfected with empty liposome group and CIK cell surface antigen expression was significantly associated antigen RNA transfection on the group turned in primary cell killing activity and the other two groups significantly increased lung cancer. Conclusion Lung cancer associated antigen RNA-transfected DC induced killer cells to target the vitality of CIK cells was significantly enhanced for the clinical application of individual DC-CIK cells had adoptive immunotherapy offers a new approach.%目的:研究NCI-H1975人肺腺癌细胞株相关抗原RNA转染的人树突状细胞(DCs)在体外诱导细胞因子诱导的杀伤细胞(CIK)对人肺腺癌细胞的杀伤活力。方法使用血细胞单

  18. Tandem CAR T cells targeting HER2 and IL13Rα2 mitigate tumor antigen escape.

    Science.gov (United States)

    Hegde, Meenakshi; Mukherjee, Malini; Grada, Zakaria; Pignata, Antonella; Landi, Daniel; Navai, Shoba A; Wakefield, Amanda; Fousek, Kristen; Bielamowicz, Kevin; Chow, Kevin K H; Brawley, Vita S; Byrd, Tiara T; Krebs, Simone; Gottschalk, Stephen; Wels, Winfried S; Baker, Matthew L; Dotti, Gianpietro; Mamonkin, Maksim; Brenner, Malcolm K; Orange, Jordan S; Ahmed, Nabil

    2016-08-01

    In preclinical models of glioblastoma, antigen escape variants can lead to tumor recurrence after treatment with CAR T cells that are redirected to single tumor antigens. Given the heterogeneous expression of antigens on glioblastomas, we hypothesized that a bispecific CAR molecule would mitigate antigen escape and improve the antitumor activity of T cells. Here, we created a CAR that joins a HER2-binding scFv and an IL13Rα2-binding IL-13 mutein to make a tandem CAR exodomain (TanCAR) and a CD28.ζ endodomain. We determined that patient TanCAR T cells showed distinct binding to HER2 or IL13Rα2 and had the capability to lyse autologous glioblastoma. TanCAR T cells exhibited activation dynamics that were comparable to those of single CAR T cells upon encounter of HER2 or IL13Rα2. We observed that TanCARs engaged HER2 and IL13Rα2 simultaneously by inducing HER2-IL13Rα2 heterodimers, which promoted superadditive T cell activation when both antigens were encountered concurrently. TanCAR T cell activity was more sustained but not more exhaustible than that of T cells that coexpressed a HER2 CAR and an IL13Rα2 CAR, T cells with a unispecific CAR, or a pooled product. In a murine glioblastoma model, TanCAR T cells mitigated antigen escape, displayed enhanced antitumor efficacy, and improved animal survival. Thus, TanCAR T cells show therapeutic potential to improve glioblastoma control by coengaging HER2 and IL13Rα2 in an augmented, bivalent immune synapse that enhances T cell functionality and reduces antigen escape.

  19. Tandem CAR T cells targeting HER2 and IL13Rα2 mitigate tumor antigen escape.

    Science.gov (United States)

    Hegde, Meenakshi; Mukherjee, Malini; Grada, Zakaria; Pignata, Antonella; Landi, Daniel; Navai, Shoba A; Wakefield, Amanda; Fousek, Kristen; Bielamowicz, Kevin; Chow, Kevin K H; Brawley, Vita S; Byrd, Tiara T; Krebs, Simone; Gottschalk, Stephen; Wels, Winfried S; Baker, Matthew L; Dotti, Gianpietro; Mamonkin, Maksim; Brenner, Malcolm K; Orange, Jordan S; Ahmed, Nabil

    2016-08-01

    In preclinical models of glioblastoma, antigen escape variants can lead to tumor recurrence after treatment with CAR T cells that are redirected to single tumor antigens. Given the heterogeneous expression of antigens on glioblastomas, we hypothesized that a bispecific CAR molecule would mitigate antigen escape and improve the antitumor activity of T cells. Here, we created a CAR that joins a HER2-binding scFv and an IL13Rα2-binding IL-13 mutein to make a tandem CAR exodomain (TanCAR) and a CD28.ζ endodomain. We determined that patient TanCAR T cells showed distinct binding to HER2 or IL13Rα2 and had the capability to lyse autologous glioblastoma. TanCAR T cells exhibited activation dynamics that were comparable to those of single CAR T cells upon encounter of HER2 or IL13Rα2. We observed that TanCARs engaged HER2 and IL13Rα2 simultaneously by inducing HER2-IL13Rα2 heterodimers, which promoted superadditive T cell activation when both antigens were encountered concurrently. TanCAR T cell activity was more sustained but not more exhaustible than that of T cells that coexpressed a HER2 CAR and an IL13Rα2 CAR, T cells with a unispecific CAR, or a pooled product. In a murine glioblastoma model, TanCAR T cells mitigated antigen escape, displayed enhanced antitumor efficacy, and improved animal survival. Thus, TanCAR T cells show therapeutic potential to improve glioblastoma control by coengaging HER2 and IL13Rα2 in an augmented, bivalent immune synapse that enhances T cell functionality and reduces antigen escape. PMID:27427982

  20. Tresyl-Based Conjugation of Protein Antigen to Lipid Nanoparticles Increases Antigen Immunogencity

    Science.gov (United States)

    Jain, Anekant; Yan, Weili; Miller, Keith R.; O'Carra, Ronan; Woodward, Jerold G.; Mumper, Russell J.

    2010-01-01

    The present studies were aimed at investigating the engineering of NPs with protein-conjugated-surfactant at their surface. In order to increase the immunogenicity of a protein antigen, Brij 78 was functionalized by tresyl chloride and then further reacted with the primary amine of the model proteins ovalbumin (OVA) or horseradish peroxide (HRP). The reaction yielded Brij 78-OVA and Brij 78-HRP conjugates which were then used directly to form NP-OVA or NP-HRP using a one-step warm oil-in-water microemulsion precursor method with emulsifying wax as the oil phase, and Brij 78 and the Brij 78-OVA or Brij 78-HRP conjugate as surfactants. Similarly, Brij 700 was conjugated to HIV p24 antigen to yield Brij 700-p24 conjugate. The utility of these NPs for enhancing the immune responses to protein-based vaccines was evaluated in vivo using ovalbumin (OVA) as model protein and p24 as a relevant HIV antigen. In separate in vivo studies, female BALB/c mice were immunized by subcutaneous (s.c.) injection with NP-OVA and NP-p24 formulations along with several control formulations. These results suggested that with multiple antigens, covalent attachment of the antigen to the NP significantly enhanced antigen-specific immune responses. This facile covalent conjugation and incorporation method may be utilized to further incorporate other protein antigens, even multiple antigens, into an enhanced vaccine delivery system. PMID:20837122

  1. Tresyl-based conjugation of protein antigen to lipid nanoparticles increases antigen immunogenicity.

    Science.gov (United States)

    Jain, Anekant; Yan, Weili; Miller, Keith R; O'Carra, Ronan; Woodward, Jerold G; Mumper, Russell J

    2010-11-30

    The present studies were aimed at investigating the engineering of NPs with protein-conjugated-surfactant at their surface. In order to increase the immunogenicity of a protein antigen, Brij 78 was functionalized by tresyl chloride and then further reacted with the primary amine of the model proteins ovalbumin (OVA) or horseradish peroxide (HRP). The reaction yielded Brij 78-OVA and Brij 78-HRP conjugates which were then used directly to form NP-OVA or NP-HRP using a one-step warm oil-in-water microemulsion precursor method with emulsifying wax as the oil phase, and Brij 78 and the Brij 78-OVA or Brij 78-HRP conjugate as surfactants. Similarly, Brij 700 was conjugated to HIV p24 antigen to yield Brij 700-p24 conjugate. The utility of these NPs for enhancing the immune responses to protein-based vaccines was evaluated in vivo using ovalbumin (OVA) as model protein and p24 as a relevant HIV antigen. In separate in vivo studies, female BALB/c mice were immunized by subcutaneous (s.c.) injection with NP-OVA and NP-p24 formulations along with several control formulations. These results suggested that with multiple antigens, covalent attachment of the antigen to the NP significantly enhanced antigen-specific immune responses. This facile covalent conjugation and incorporation method may be utilized to further incorporate other protein antigens, even multiple antigens, into an enhanced vaccine delivery system. PMID:20837122

  2. A new virion precipitation test for oncovirus envelope antigens which detects common antigenic determinants in mammalian type-C viruses and Mason-Pfizer monkey virus.

    Science.gov (United States)

    Altstein, A D; Zakharova, L G; Zhdanov, V M

    1979-03-15

    A method for the study of oncovirus envelope antigens was developed, bases on the precipitation of intact virions by a double antibody technique. The amount of precipitated virus was then measured as reverse transcriptase activity. The method was designated the virion precipitation test (VPT). It has been used for titration of antibodies to envelope antigens of oncoviruses. The study of envelop antigens of 11 different oncoviruses permitted their differentiation into the following groups: (1) murine type-C viruses: (2) feline type-C viruses; (3) simian type-C viruses; (4) the RD-114/BEV group; (5) Mason-Pfizer monkey virus (M-PMV); (6) bovine leukemia virus; (7) avian type-C viruses; (8) mouse mammary tumor virus. No common antigenic determinants were detected in the last three groups. Mammalian type-C viruses (RD-114, NIH-MuLV, G-MuLV) had common antigenic determinants in the envelope, as demonstrated with an anti-RD-114 serum. Mammalian type-C viruses also shared antigenic determinants with M-PMV. The relationship of type-C viruses to M-PMV decreased in the following order: RD-114--NIH-MuLV--G-MuLV. It was also shown that the endogenous xenotropic feline RD-114 virus was more closely related to xenotropic NIH-MuLV than to ecotropic G-MuLV. The nature of the common antigenic determinants, as demonstrated by VPT on the surface of mammalian type-C viruses and M-PMV, and their significance for the concept of oncovirus evolution are discussed.

  3. Screening and characterization of early diagnostic antigens in excretory-secretory proteins from Trichinella spiralis intestinal infective larvae by immunoproteomics.

    Science.gov (United States)

    Liu, Ruo Dan; Jiang, Peng; Wen, Hui; Duan, Jiang Yang; Wang, Li Ang; Li, Jie Feng; Liu, Chun Ying; Sun, Ge Ge; Wang, Zhong Quan; Cui, Jing

    2016-02-01

    The excretory-secretory (ES) antigens from Trichinella spiralis muscle larvae are the most commonly used diagnostic antigens for trichinellosis, but specific IgG antibodies were not detected in early stage of infection. The aim of this study was to identify early diagnostic antigens from ES proteins of intestinal infective larvae (IIL), the first invasive stage of T. spiralis. Six bands (92, 52, 45, 35, 32, and 29 kDa) of IIL ES proteins were recognized by infection sera in Western blotting as early as 10 days post infection. Total of 54 T. spiralis proteins in six bands were identified by shotgun LC-MS/MS, 30 proteins were annotated, and 27 had hydrolase activity. Several proteins (serine protease, putative trypsin, deoxyribonuclease II family protein, etc.) could be considered as the potential early diagnostic antigens for trichinellosis. Our study provides new insights for screening early diagnostic antigens from intestinal worms of T. spiralis. PMID:26468148

  4. Caracterización y análisis de la actividad antigénica de proteínas de la membrana germinal de Echinococcus granulosus Characterization and antigenic activity of Echinococcus granulosus germinal layer proteins

    Directory of Open Access Journals (Sweden)

    L.H. Prieto

    2006-12-01

    Full Text Available Proteínas presentes en un extracto de membrana germinal de Echinococcus granulosus fueron separadas en una fase acuosa y en una fase detergente mediante el tratamiento con Triton X-114. La caracterización mediante electroforesis en gel de poliacrilamida en presencia de dodecil sulfato de sodio, mostró que proteínas con pesos moleculares aproximados de 102, 45, 28.5-28 y 15.5-15 kDa se presentaron en el extracto y en la fase acuosa (proteínas hidrofílicas. La caracterización inmunológica de proteínas del extracto mediante inmunotransferencia frente a suero ovino no inmunizado, frente a suero ovino inmunizado con antígenos de E. granulosus y frente a suero anti-inmunoglobulina ovina, mostró patrones de reactividad similares. El ensayo inmunodetección en un punto, de proteínas del extracto previamente tratadas con Triton X-114 frente a sueros humanos incubados con el detergente iónico dodecil sulfato de sodio, mostró reactividad frente a sueros humanos con equinococosis quística y no mostró reactividad frente a sueros humanos sin equinococosis quística ni frente a un suero humano infectado con la filaria Mansonella ozzardi.Proteins present in an extract of Echinococcus granulosus germinal layer were separated in hydrophobic and hydrophilic phases using Triton X-114. The characterization carried out by sodium dodecyl sulphate polyacrylamide gel electrophoresis showed that proteins of apparent molecular weights of 102, 45, 28.5-28 and 15.5-15 kDa were present in the extract and in the aqueous phase (hydrophilic proteins. The immunological characterization of extract proteins performed by immunoblotting against to no immunized sheep serum, against to immunized with E. granulosus antigens sheep serum and against to anti-sheep immunoglobulin serum, showed similar reaction patterns. The dot immunobinding assay of extract proteins after Triton X-114 treatment against to human sera incubated in the presence of sodium dodecil sulphate

  5. MHC-restricted antigen presentation and recognition: constraints on gene, recombinant and peptide vaccines in humans

    Directory of Open Access Journals (Sweden)

    Cunha-Neto E.

    1999-01-01

    Full Text Available The target of any immunization is to activate and expand lymphocyte clones with the desired recognition specificity and the necessary effector functions. In gene, recombinant and peptide vaccines, the immunogen is a single protein or a small assembly of epitopes from antigenic proteins. Since most immune responses against protein and peptide antigens are T-cell dependent, the molecular target of such vaccines is to generate at least 50-100 complexes between MHC molecule and the antigenic peptide per antigen-presenting cell, sensitizing a T cell population of appropriate clonal size and effector characteristics. Thus, the immunobiology of antigen recognition by T cells must be taken into account when designing new generation peptide- or gene-based vaccines. Since T cell recognition is MHC-restricted, and given the wide polymorphism of the different MHC molecules, distinct epitopes may be recognized by different individuals in the population. Therefore, the issue of whether immunization will be effective in inducing a protective immune response, covering the entire target population, becomes an important question. Many pathogens have evolved molecular mechanisms to escape recognition by the immune system by variation of antigenic protein sequences. In this short review, we will discuss the several concepts related to selection of amino acid sequences to be included in DNA and peptide vaccines.

  6. Exposed and concealed antigens as vaccine targets for controlling ticks and tick-borne diseases.

    Science.gov (United States)

    Nuttall, P A; Trimnell, A R; Kazimirova, M; Labuda, M

    2006-04-01

    Tick vaccines derived from Bm86, a midgut membrane-bound protein of the cattle tick, Boophilus microplus, are currently the only commercially available ectoparasite vaccines. Despite its introduction to the market in 1994, and the recognized need for alternatives to chemical pesticides, progress in developing effective antitick vaccines (and ectoparasite vaccines in general) is slow. The primary rate-limiting step is the identification of suitable antigenic targets for vaccine development. Two sources of candidate vaccine antigens have been identified: 'exposed' antigens that are secreted in tick saliva during attachment and feeding on a host and 'concealed' antigens that are normally hidden from the host. Recently, a third group of antigens has been distinguished that combines the properties of both exposed and concealed antigens. This latter group offers the prospect of a broad-spectrum vaccine effective against both adults and immature stages of a wide variety of tick species. It also shows transmission-blocking and protective activity against a tick-borne pathogen. With the proliferation of molecular techniques and their application to vaccine development, there are high hopes for new and effective antitick vaccines that also control tick-borne diseases. PMID:16542317

  7. Regeneration of tumor antigen-specific CTLs utilizing iPS technology.

    Science.gov (United States)

    Maeda, Takuya; Masuda, Kyoko; Kawamoto, Hiroshi

    2016-08-01

    Tumor immunotherapy, especially tumor antigen specific T cell therapy, is currently attracting attention. However, a critical issue still awaits resolution; it is difficult to efficiently expand tumor antigen-specific T cells. To solve this problem, we are now utilizing iPS cell technology. When iPS cells are established from tumor antigen specific T cells, T cells regenerated from these iPS cells are expected to express the same TCRs as the original T cells. In line with this concept, we succeeded in regenerating tumor antigen specific cytotoxic T cells. The regenerated T cells exhibited TCR specific killing activity comparable to that of the original cells, and were able to kill leukemia cells in an antigen-specific manner. We are currently endeavoring to apply this method clinically. In the future, we intend to establish an allogeneic transfusion system, in which various tumor antigen specific T-iPS cells from a wide range of HLA haplotype homozygous donors will be lined up as a "T-iPS cell bank", with the aim of making off-the-shelf tumor immunotherapy a reality. PMID:27599425

  8. Added value of antigen ELISA in the diagnosis of neurocysticercosis in resource poor settings.

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    Sarah Gabriël

    Full Text Available BACKGROUND: Neurocysticercosis (NCC is the most common cause of acquired epilepsy in Taenia solium endemic areas, primarily situated in low-income countries. Diagnosis is largely based upon the "Del Brutto diagnostic criteria" using the definitive/probable/no NCC diagnosis approach. Neuroimaging and specific T. solium cysticercosis antibody detection results are at the mainstay of this diagnosis, while antigen detection in serum has never been included. This study aimed at evaluating the addition of antigen detection as a major diagnostic criterion, especially in areas where neuroimaging is absent. METHODS: The B158/B60 monoclonal antibody-based enzyme-linked immunosorbent assay (ELISA for the detection of circulating cysticercus antigen was carried out retrospectively on serum samples collected during a hospital-based study from 83 people with epilepsy (PWE in an endemic area. RESULTS: The addition of antigen results as a major criterion allowed the correct diagnosis of definitive NCC in 10 out of 17 patients as opposed to 0/17 without antigen results in the absence of neuroimaging. A sensitivity of 100% and a specificity of 84% were determined for the diagnosis of active NCC using antigen ELISA. While the use of a higher cutoff improves the specificity of the test to 96%, it decreases its sensitivity to 83%. CONCLUSIONS: In areas where neuroimaging is absent, NCC diagnosis according to the existing criteria is problematic. Taking into account its limitations for diagnosis of inactive NCC, antigen detection can be of added value for diagnosing NCC in PWE by supporting diagnostic and treatment decisions. Therefore, we recommend a revision of the "Del Brutto diagnostic criteria" for use in resource poor areas and suggest the inclusion of serum antigen detection as a major criterion.

  9. Liposome-Based Adjuvants for Subunit Vaccines: Formulation Strategies for Subunit Antigens and Immunostimulators

    Directory of Open Access Journals (Sweden)

    Signe Tandrup Schmidt

    2016-03-01

    Full Text Available The development of subunit vaccines has become very attractive in recent years due to their superior safety profiles as compared to traditional vaccines based on live attenuated or whole inactivated pathogens, and there is an unmet medical need for improved vaccines and vaccines against pathogens for which no effective vaccines exist. The subunit vaccine technology exploits pathogen subunits as antigens, e.g., recombinant proteins or synthetic peptides, allowing for highly specific immune responses against the pathogens. However, such antigens are usually not sufficiently immunogenic to induce protective immunity, and they are often combined with adjuvants to ensure robust immune responses. Adjuvants are capable of enhancing and/or modulating immune responses by exposing antigens to antigen-presenting cells (APCs concomitantly with conferring immune activation signals. Few adjuvant systems have been licensed for use in human vaccines, and they mainly stimulate humoral immunity. Thus, there is an unmet demand for the development of safe and efficient adjuvant systems that can also stimulate cell-mediated immunity (CMI. Adjuvants constitute a heterogeneous group of compounds, which can broadly be classified into delivery systems or immunostimulators. Liposomes are versatile delivery systems for antigens, and they can carefully be customized towards desired immune profiles by combining them with immunostimulators and optimizing their composition, physicochemical properties and antigen-loading mode. Immunostimulators represent highly diverse classes of molecules, e.g., lipids, nucleic acids, proteins and peptides, and they are ligands for pattern-recognition receptors (PRRs, which are differentially expressed on APC subsets. Different formulation strategies might thus be required for incorporation of immunostimulators and antigens, respectively, into liposomes, and the choice of immunostimulator should ideally be based on knowledge regarding the

  10. Clearance of depot vaccine SPIO-labeled antigen and substrate visualized using MRI.

    Science.gov (United States)

    Brewer, Kimberly D; Lake, Kerry; Pelot, Nicole; Stanford, Marianne M; DeBay, Drew R; Penwell, Andrea; Weir, Genevieve M; Karkada, Mohan; Mansour, Marc; Bowen, Chris V

    2014-12-01

    Immunotherapies, including peptide-based vaccines, are a growing area of cancer research, and understanding their mechanism of action is crucial for their continued development and clinical application. Exploring the biodistribution of vaccine components may be key to understanding this action. This work used magnetic resonance imaging (MRI) to characterize the in vivo biodistribution of the antigen and oil substrate of the vaccine delivery system known as DepoVax(TM). DepoVax uses a novel adjuvanted lipid-in-oil based formulation to solubilise antigens and promote a depot effect. In this study, antigen or oil were tagged with superparamagnetic iron oxide (SPIO), making them visible on MR images. This enables tracking of individual vaccine components to determine changes in biodistribution. Mice were injected with SPIO-labeled antigen or SPIO-labeled oil, and imaged to examine clearance of labeled components from the vaccine site. The SPIO-antigen was steadily cleared, with nearly half cleared within two months post-vaccination. In contrast, the SPIO-oil remained relatively unchanged. The biodistribution of the SPIO-antigen component within the vaccine site was heterogeneous, indicating the presence of active clearance mechanisms, rather than passive diffusion or drainage. Mice injected with SPIO-antigen also showed MRI contrast for several weeks post-vaccination in the draining inguinal lymph node. These results indicate that MRI can visualize the in vivo longitudinal biodistribution of vaccine components. The sustained clearance is consistent with antigen up-take and trafficking by immune cells, leading to accumulation in the draining lymph node, which corresponds to the sustained immune responses and reduced tumor burden observed in vaccinated mice. PMID:25444822

  11. Nonprostatic sources of prostate-specific antigen.

    Science.gov (United States)

    Diamandis, E P; Yu, H

    1997-05-01

    The name prostate-specific antigen has been given to a protein that now is known not to be prostate-specific; however, prostatic tissue does produces extremely high levels of PSA and secrets it into the seminal plasma. Seminal plasma contains about 1 million micrograms/L of PSA and is the richest source of PSA reported. The biologic fluid with the second highest PSA concentration, however, is nipple aspirate fluid from the female breast (up to about 5000 micrograms/L), and the third is milk from lactating women (up to 300 micrograms/L). Male serum PSA is usually less than 4 micrograms/L. In nonprostatic tissues, PSA exists mainly in its free molecular form, but PSA-ACT complex is also present in most of the fluids that contain PSA, such as breast secretions and amniotic fluid. The gene expression and protein production of PSA in nonprostatic tissues are under the regulation of steroid hormones via their receptors. Androgens, glucocorticoids, and progestins up-regulate the PSA gene expression, resulting in an increase of protein production. Estrogen by itself seems to have no effect on PSA regulation, but it can impair PSA production induced by androgen. It remains unknown whether PSA is enzymatically active and what is the physiologic role of PSA in nonprostatic tissues. It is speculated that PSA may be involved in the regulation of growth factors. Measuring PSA in breast cancer cytosol, breast-nipple aspirate fluid, and female serum may have potential clinical utilities, including breast cancer prognosis, breast cancer risk assessment, and evaluation of androgen excess. Further studies are needed to identify the exact function and regulation of PSA in nonprostatic tissues and to explore the clinical application of this protein. PMID:9126224

  12. Melanocyte antigen triggers autoimmunity in human psoriasis.

    Science.gov (United States)

    Arakawa, Akiko; Siewert, Katherina; Stöhr, Julia; Besgen, Petra; Kim, Song-Min; Rühl, Geraldine; Nickel, Jens; Vollmer, Sigrid; Thomas, Peter; Krebs, Stefan; Pinkert, Stefan; Spannagl, Michael; Held, Kathrin; Kammerbauer, Claudia; Besch, Robert; Dornmair, Klaus; Prinz, Jörg C

    2015-12-14

    Psoriasis vulgaris is a common T cell-mediated inflammatory skin disease with a suspected autoimmune pathogenesis. The human leukocyte antigen (HLA) class I allele, HLA-C*06:02, is the main psoriasis risk gene. Epidermal CD8(+) T cells are essential for psoriasis development. Functional implications of HLA-C*06:02 and mechanisms of lesional T cell activation in psoriasis, however, remained elusive. Here we identify melanocytes as skin-specific target cells of an HLA-C*06:02-restricted psoriatic T cell response. We found that a Vα3S1/Vβ13S1 T cell receptor (TCR), which we had reconstituted from an epidermal CD8(+) T cell clone of an HLA-C*06:02-positive psoriasis patient specifically recognizes HLA-C*06:02-positive melanocytes. Through peptide library screening, we identified ADAMTS-like protein 5 (ADAMTSL5) as an HLA-C*06:02-presented melanocytic autoantigen of the Vα3S1/Vβ13S1 TCR. Consistent with the Vα3S1/Vβ13S1-TCR reactivity, we observed numerous CD8(+) T cells in psoriasis lesions attacking melanocytes, the only epidermal cells expressing ADAMTSL5. Furthermore, ADAMTSL5 stimulation induced the psoriasis signature cytokine, IL-17A, in CD8(+) T cells from psoriasis patients only, supporting a role as psoriatic autoantigen. This unbiased analysis of a TCR obtained directly from tissue-infiltrating CD8(+) T cells reveals that in psoriasis HLA-C*06:02 directs an autoimmune response against melanocytes through autoantigen presentation. We propose that HLA-C*06:02 may predispose to psoriasis via this newly identified autoimmune pathway.

  13. A Lipid Based Antigen Delivery System Efficiently Facilitates MHC Class-I Antigen Presentation in Dendritic Cells to Stimulate CD8+ T Cells

    Science.gov (United States)

    Maji, Mithun; Mazumder, Saumyabrata; Bhattacharya, Souparno; Choudhury, Somsubhra Thakur; Sabur, Abdus; Shadab, Md.; Bhattacharya, Pradyot; Ali, Nahid

    2016-06-01

    The most effective strategy for protection against intracellular infections such as Leishmania is vaccination with live parasites. Use of recombinant proteins avoids the risks associated with live vaccines. However, due to low immunogenicity, they fail to trigger T cell responses particularly of CD8+ cells requisite for persistent immunity. Previously we showed the importance of protein entrapment in cationic liposomes and MPL as adjuvant for elicitation of CD4+ and CD8+ T cell responses for long-term protection. In this study we investigated the role of cationic liposomes on maturation and antigen presentation capacity of dendritic cells (DCs). We observed that cationic liposomes were taken up very efficiently by DCs and transported to different cellular sites. DCs activated with liposomal rgp63 led to efficient presentation of antigen to specific CD4+ and CD8+ T cells. Furthermore, lymphoid CD8+ T cells from liposomal rgp63 immunized mice demonstrated better proliferative ability when co-cultured ex vivo with stimulated DCs. Addition of MPL to vaccine enhanced the antigen presentation by DCs and induced more efficient antigen specific CD8+ T cell responses when compared to free and liposomal antigen. These liposomal formulations presented to CD8+ T cells through TAP-dependent MHC-I pathway offer new possibilities for a safe subunit vaccine.

  14. Antigenic typing Polish isolates of canine parvovirus

    Energy Technology Data Exchange (ETDEWEB)

    Mizak, B. [National Veterinary Research Institute, Pulawy (Poland); Plucienniczak, A. [Polish Academy ofd Sciences. Microbiology and Virology Center, Lodz (Poland)

    1995-12-31

    Polish strains of canine parvovirus isolated between 1982 and 1993 were examined to determine the extent to which the virus has evolved antigenically and genetically over eleven years. Two CPV isolates obtained in Warsaw in 1982 and Pulawy in 1993, were examined using monoclonal antibody typing, restriction analysis and sequencing VP-2 protein gene. Five other isolates from Warsaw and Pulawy were tested with the panel of monoclonal antibodies specific to CPV-2, CPV-2a and common for canine parvovirus, feline panleukopenia virus and milk enteritis virus. Results of the studies demonstrated that all isolates tested represented CPV-2a antigenic type. Rapid antigenic strain replacement recorded by Parrish and Senda in the U.S.A and Japan was not confirmed in Poland. (author). 30 refs, 2 tabs.

  15. Antigen sampling in the fish intestine.

    Science.gov (United States)

    Løkka, Guro; Koppang, Erling Olaf

    2016-11-01

    Antigen uptake in the gastrointestinal tract may induce tolerance, lead to an immune response and also to infection. In mammals, most pathogens gain access to the host though the gastrointestinal tract, and in fish as well, this route seems to be of significant importance. The epithelial surface faces a considerable challenge, functioning both as a barrier towards the external milieu but simultaneously being the site of absorption of nutrients and fluids. The mechanisms allowing antigen uptake over the epithelial barrier play a central role for maintaining the intestinal homeostasis and regulate appropriate immune responses. Such uptake has been widely studied in mammals, but also in fish, a number of experiments have been reported, seeking to reveal cells and mechanisms involved in antigen sampling. In this paper, we review these studies in addition to addressing our current knowledge of the intestinal barrier in fish and its anatomical construction. PMID:26872546

  16. Idiopathic focal segmental glomerulosclerosis and HLA antigens.

    Science.gov (United States)

    Gerbase-DeLima, M; Pereira-Santos, A; Sesso, R; Temin, J; Aragão, E S; Ajzen, H

    1998-03-01

    The objective of the present study was to investigate a possible association between HLA class II antigens and idiopathic focal segmental glomerulosclerosis (FSGS). HLA-A, -B, -DR and -DQ antigens were determined in 19 Brazilian patients (16 white subjects and three subjects of Japanese origin) with biopsy-proven FSGS. Comparison of the HLA antigen frequencies between white patients and white local controls showed a significant increase in HLA-DR4 frequency among FSGS patients (37.7 vs 17.2%, P < 0.05). In addition, the three patients of Japanese extraction, not included in the statistical analysis, also presented HLA-DR4. In conclusion, our data confirm the association of FSGS with HLA-DR4 previously reported by others, thus providing further evidence for a role of genes of the HLA complex in the susceptibility to this disease. PMID:9698788

  17. Idiopathic focal segmental glomerulosclerosis and HLA antigens

    Directory of Open Access Journals (Sweden)

    M. Gerbase-DeLima

    1998-03-01

    Full Text Available The objective of the present study was to investigate a possible association between HLA class II antigens and idiopathic focal segmental glomerulosclerosis (FSGS. HLA-A, -B, -DR and -DQ antigens were determined in 19 Brazilian patients (16 white subjects and three subjects of Japanese origin with biopsy-proven FSGS. Comparison of the HLA antigen frequencies between white patients and white local controls showed a significant increase in HLA-DR4 frequency among FSGS patients (37.7 vs 17.2%, P<0.05. In addition, the three patients of Japanese extraction, not included in the statistical analysis, also presented HLA-DR4. In conclusion, our data confirm the association of FSGS with HLA-DR4 previously reported by others, thus providing further evidence for a role of genes of the HLA complex in the susceptibility to this disease

  18. Differential Recognition and Hydrolysis of Host Carbohydrate Antigens by Streptococcus pneumoniae Family 98 Glycoside Hydrolases

    Energy Technology Data Exchange (ETDEWEB)

    Higgins, M.; Whitworth, G; El Warry, N; Randriantsoa, M; Samain, E; Burke, R; Vocadlo, D; Boraston, A

    2009-01-01

    The presence of a fucose utilization operon in the Streptococcus pneumoniae genome and its established importance in virulence indicates a reliance of this bacterium on the harvesting of host fucose-containing glycans. The identities of these glycans, however, and how they are harvested is presently unknown. The biochemical and high resolution x-ray crystallographic analysis of two family 98 glycoside hydrolases (GH98s) from distinctive forms of the fucose utilization operon that originate from different S. pneumoniae strains reveal that one enzyme, the predominant type among pneumococcal isolates, has a unique endo-{beta}-galactosidase activity on the LewisY antigen. Altered active site topography in the other species of GH98 enzyme tune its endo-{beta}-galactosidase activity to the blood group A and B antigens. Despite their different specificities, these enzymes, and by extension all family 98 glycoside hydrolases, use an inverting catalytic mechanism. Many bacterial and viral pathogens exploit host carbohydrate antigens for adherence as a precursor to colonization or infection. However, this is the first evidence of bacterial endoglycosidase enzymes that are known to play a role in virulence and are specific for distinct host carbohydrate antigens. The strain-specific distribution of two distinct types of GH98 enzymes further suggests that S. pneumoniae strains may specialize to exploit host-specific antigens that vary from host to host, a factor that may feature in whether a strain is capable of colonizing a host or establishing an invasive infection.

  19. 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...... of IgG and C3 fragments. Furthermore, the binding of Tg to B cells in preparations of normal blood cells was higher in HT serum than in serum from controls and correlated positively with the serum anti-Tg activity, as did the B and CD4+ T cell proliferation. Disruption of the three-dimensional structure...

  20. Properties of glycolipid-enriched membrane rafts in antigen presentation.

    Science.gov (United States)

    Rodgers, William; Smith, Kenneth

    2005-01-01

    Presentation of antigen to T cells represents one of the central events in the engagement of the immune system toward the defense of the host against pathogens. Accordingly, understanding the mechanisms by which antigen presentation occurs is critical toward our understanding the properties of host defense against foreign antigen, as well as insight into other features of the immune system, such as autoimmune disease. The entire antigen-presentation event is complex, and many features of it remain poorly understood. However, recent studies have provided evidence showing that glycolipid-enriched membrane rafts are important for efficient antigen presentation; the studies suggest that one such function of rafts is trafficking of antigen-MHC II complexes to the presentation site on the surface of the antigen-presenting cell. Here, we present a critical discussion of rafts and their proposed functions in antigen presentation. Emerging topics of rafts and antigen presentation that warrant further investigation are also highlighted.

  1. Antigen processing and remodeling of the endosomal pathway: requirements for antigen cross-presentation.

    Directory of Open Access Journals (Sweden)

    Ewoud Bernardus Compeer

    2012-03-01

    Full Text Available The cross-presentation of endocytosed antigen as peptide/class I MHC complexes plays a central role in the elicitation of CD8+ T cell clones that mediate anti-viral and anti-tumor immune responses. While it has been clear that there are specific subsets of professional antigen presenting cells (APC capable of antigen cross-presentation, description of mechanisms involved is still ongoing. Especially amongst dendritic cells (DC, there are specialized subsets that are highly proficient at antigen cross-presentation. We here present a focused survey on the cell biological processes in the endosomal pathway that support antigen cross-presentation. This review highlight DC-intrinsic mechanisms that facilitate the cross-presentation of endocytosed antigen, including receptor-mediated uptake, recycling and maturation including the sorting of membrane proteins, dynamic remodeling of endosomal structures and cell-surface directed endosomal trafficking. We will conclude with description of pathogen-induced deviation of endosomal processing, and discuss how immune evasion strategies pertaining endosomal trafficking may preclude antigen cross-presentation.

  2. Cytotoxicity of tumor antigen specific human T cells is unimpaired by arginine depletion.

    Directory of Open Access Journals (Sweden)

    Markus Munder

    Full Text Available Tumor-growth is often associated with the expansion of myeloid derived suppressor cells that lead to local or systemic arginine depletion via the enzyme arginase. It is generally assumed that this arginine deficiency induces a global shut-down of T cell activation with ensuing tumor immune escape. While the impact of arginine depletion on polyclonal T cell proliferation and cytokine secretion is well documented, its influence on chemotaxis, cytotoxicity and antigen specific activation of human T cells has not been demonstrated so far. We show here that chemotaxis and early calcium signaling of human T cells are unimpaired in the absence of arginine. We then analyzed CD8(+ T cell activation in a tumor peptide as well as a viral peptide antigen specific system: (i CD8(+ T cells with specificity against the MART-1aa26-35*A27L tumor antigen expanded with in vitro generated dendritic cells, and (ii clonal CMV pp65aa495-503 specific T cells and T cells retrovirally transduced with a CMV pp65aa495-503 specific T cell receptor were analyzed. Our data demonstrate that human CD8(+ T cell antigen specific cytotoxicity and perforin secretion are completely preserved in the absence of arginine, while antigen specific proliferation as well as IFN-γ and granzyme B secretion are severely compromised. These novel results highlight the complexity of antigen specific T cell activation and demonstrate that human T cells can preserve important activation-induced effector functions in the context of arginine deficiency.

  3. Antigen-Antibody Interaction Database (AgAbDb): a compendium of antigen-antibody interactions.

    Science.gov (United States)

    Kulkarni-Kale, Urmila; Raskar-Renuse, Snehal; Natekar-Kalantre, Girija; Saxena, Smita A

    2014-01-01

    Antigen-Antibody Interaction Database (AgAbDb) is an immunoinformatics resource developed at the Bioinformatics Centre, University of Pune, and is available online at http://bioinfo.net.in/AgAbDb.htm. Antigen-antibody interactions are a special class of protein-protein interactions that are characterized by high affinity and strict specificity of antibodies towards their antigens. Several co-crystal structures of antigen-antibody complexes have been solved and are available in the Protein Data Bank (PDB). AgAbDb is a derived knowledgebase developed with an objective to compile, curate, and analyze determinants of interactions between the respective antigen-antibody molecules. AgAbDb lists not only the residues of binding sites of antigens and antibodies, but also interacting residue pairs. It also helps in the identification of interacting residues and buried residues that constitute antibody-binding sites of protein and peptide antigens. The Antigen-Antibody Interaction Finder (AAIF), a program developed in-house, is used to compile the molecular interactions, viz. van der Waals interactions, salt bridges, and hydrogen bonds. A module for curating water-mediated interactions has also been developed. In addition, various residue-level features, viz. accessible surface area, data on epitope segment, and secondary structural state of binding site residues, are also compiled. Apart from the PDB numbering, Wu-Kabat numbering and explicit definitions of complementarity-determining regions are provided for residues of antibodies. The molecular interactions can be visualized using the program Jmol. AgAbDb can be used as a benchmark dataset to validate algorithms for prediction of B-cell epitopes. It can as well be used to improve accuracy of existing algorithms and to design new algorithms. AgAbDb can also be used to design mimotopes representing antigens as well as aid in designing processes leading to humanization of antibodies. PMID:25048123

  4. Combinational targeting offsets antigen escape and enhances effector functions of adoptively transferred T cells in glioblastoma.

    Science.gov (United States)

    Hegde, Meenakshi; Corder, Amanda; Chow, Kevin K H; Mukherjee, Malini; Ashoori, Aidin; Kew, Yvonne; Zhang, Yi Jonathan; Baskin, David S; Merchant, Fatima A; Brawley, Vita S; Byrd, Tiara T; Krebs, Simone; Wu, Meng Fen; Liu, Hao; Heslop, Helen E; Gottschalk, Stephen; Gottachalk, Stephen; Yvon, Eric; Ahmed, Nabil

    2013-11-01

    Preclinical and early clinical studies have demonstrated that chimeric antigen receptor (CAR)-redirected T cells are highly promising in cancer therapy. We observed that targeting HER2 in a glioblastoma (GBM) cell line results in the emergence of HER2-null tumor cells that maintain the expression of nontargeted tumor-associated antigens. Combinational targeting of these tumor-associated antigens could therefore offset this escape mechanism. We studied the single-cell coexpression patterns of HER2, IL-13Rα2, and EphA2 in primary GBM samples using multicolor flow cytometry and immunofluorescence, and applied a binomial routine to the permutations of antigen expression and the related odds of complete tumor elimination. This mathematical model demonstrated that cotargeting HER2 and IL-13Rα2 could maximally expand the therapeutic reach of the T cell product in all primary tumors studied. Targeting a third antigen did not predict an added advantage in the tumor cohort studied. We therefore generated bispecific T cell products from healthy donors and from GBM patients by pooling T cells individually expressing HER2 and IL-13Rα2-specific CARs and by making individual T cells to coexpress both molecules. Both HER2/IL-13Rα2-bispecific T cell products offset antigen escape, producing enhanced effector activity in vitro immunoassays (against autologous glioma cells in the case of GBM patient products) and in an orthotopic xenogeneic murine model. Further, T cells coexpressing HER2 and IL-13Rα2-CARs exhibited accentuated yet antigen-dependent downstream signaling and a particularly enhanced antitumor activity.

  5. Detection and manipulation of live antigen-expressing cells using conditionally stable nanobodies.

    Science.gov (United States)

    Tang, Jonathan Cy; Drokhlyansky, Eugene; Etemad, Behzad; Rudolph, Stephanie; Guo, Binggege; Wang, Sui; Ellis, Emily G; Li, Jonathan Z; Cepko, Constance L

    2016-01-01

    The ability to detect and/or manipulate specific cell populations based upon the presence of intracellular protein epitopes would enable many types of studies and applications. Protein binders such as nanobodies (Nbs) can target untagged proteins (antigens) in the intracellular environment. However, genetically expressed protein binders are stable regardless of antigen expression, complicating their use for applications that require cell-specificity. Here, we created a conditional system in which the stability of an Nb depends upon an antigen of interest. We identified Nb framework mutations that can be used to rapidly create destabilized Nbs. Fusion of destabilized Nbs to various proteins enabled applications in living cells, such as optogenetic control of neural activity in specific cell types in the mouse brain, and detection of HIV-infected human cells by flow cytometry. These approaches are generalizable to other protein binders, and enable the rapid generation of single-polypeptide sensors and effectors active in cells expressing specific intracellular epitopes. PMID:27205882

  6. Radioimmunoassay in the detection of the hepatitis Be antigen/antibody system in asymptomatic carriers of hepatitis B surface antigen

    International Nuclear Information System (INIS)

    A radioimmunoassay for hepatitis e antigen (HBeAg) and antibody to e (anti-HBe) was developed and sera of 71 asymptomatic chronic carriers of hepatitis B surface antigen (HBsAg), in 44 of whom liver biopsy was obtained, were tested. In addition, testing for Dane particle associated DNA polymerase activity was performed in all sera. HBeAg was detected in 14 subjects (19.7%) and anti-HBe in 46 (64.8%). The highest proportion of HBeAg positivity (40%) was found among carriers with histological evidence of chronic hepatitis, whereas anti-HBe was present in 80% of carriers with normal liver histology, in 58% of carriers with non-specific reactive hepatitis and in 60% of carriers with chronic liver lesions. DNA polymerase activity was present in 92.8% of sera positive for HBeAg, in 13% of sera positive for anti HBe, and in 9% of sera negative for both markers. Our results demonstrate that not all HBsAg carriers reactive to HBeAg show evidence of chronic hepatitis nor, conversely, that anti-HBe is invariably associated with the healthy carrier state of HBsAg. Finally, circulating Dane particles, as revealed by the presence of serum specific DNA polymerase activity, may also be present in anti-HBe positive sera other than those of some HBsAg carriers lacking both HBeAg and anti-HBe. (orig.)

  7. Hypersensitivity pneumonitis due to metalworking fluids: how to find the antigens.

    Science.gov (United States)

    Merget, R; Sander, I; van Kampen, V; Raulf-Heimsoth, M; Rabente, T; Kolk, A; Brüning, Th

    2013-01-01

    Most surveys of outbreaks of hypersensitivity pneumonitis (HP) in subjects with occupational exposure to water-based metalworking fluids (MWFs) were unable to detect a clear link between symptoms and the precise causative agents. We studied the case of a male 41-year-old industrial knife grinder with exposure to water-based MWFs since 12 years. The diagnosis of HP was made by typical work-related symptoms, the demonstration of high lymphocyte numbers in bronchoalveolar lavage and elevated IgG antibody concentrations to various molds in the patient's serum, and complete recovery after early exposure cessation. Whereas an environmental survey showed only low numbers of mold contamination in one sump sample, high antigenic activity was demonstrated in the same sample by antigen-specific IgG inhibition tests. We conclude that the detection of antigenic molds in water-based MWFs by culture methods may be limited. The link between occupational exposure to specific molds in MWFs and hypersensitivity pneumonitis can be established by the demonstration of antigenic activity by antigen-specific IgG inhibition tests. PMID:23835995

  8. [Presence of Australia antigen in blood donors].

    Science.gov (United States)

    Gota, F

    1980-01-01

    The differential diagnosis of type A and B viral hepatitis is discussed and guidelines for the prevention of post-transfusional hospital hepatitis are proposed. Methods for the immunological demonstration of HBs antigen are illustrated, together with the respective positivity percentages in blood donors.

  9. HLA antigens and asthma in Greeks.

    Science.gov (United States)

    Apostolakis, J; Toumbis, M; Konstantopoulos, K; Kamaroulias, D; Anagnostakis, J; Georgoulias, V; Fessas, P; Zervas, J

    1996-04-01

    HLA-A and -B antigens were determined in a group of 76 Greek asthmatic patients: 35 children (1.5-15 years) and 41 adults (18-73 years). The results were compared to those of 400 healthy unrelated controls from the same population. The standard NIH lymphocytotoxicity test was applied. When all 76 patients were compared to the controls, a statistically significant lower frequency of HLA-B5 and -B35 antigens was noted. When adults were analysed alone, an increased frequency of HLA-B8 was found. On the other hand, in the asthmatic children sub-group, the HLA-A10 antigen was significantly higher and the HLA-B5 was significantly lower than in the controls. These data imply that different HLA antigens may be involved in the pathogenesis of several clinical forms of asthma and that, in order to study the role of immunogenetic factor(s) in the pathogenesis of this disease, more adequate grouping criteria are needed.

  10. Antigen dynamics of follicular dendritic cells

    NARCIS (Netherlands)

    Heesters, B.A.

    2015-01-01

    Stromal-derived follicular dendritic cells (FDCs) are a major depot for antigen that are essential for formation of germinal centers, the site where memory and effector B cells differentiate and high-affinity antibody production takes place. Historically, FDCs have been characterized as ‘accessory’

  11. Circulating filarial antigen detection in brugian filariasis.

    Science.gov (United States)

    Tripathi, Praveen Kumar; Mahajan, Ramesh Chander; Malla, Nancy; Mewara, Abhishek; Bhattacharya, Shailja Misra; Shenoy, Ranganatha Krishna; Sehgal, Rakesh

    2016-03-01

    Human lymphatic filariasis (LF) is a major cause of disability globally. The success of global elimination programmes for LF depends upon effectiveness of tools for diagnosis and treatment. In this study on stage-specific antigen detection in brugian filariasis, L3, adult worm (AW) and microfilarial antigenaemia were detected in around 90-95% of microfilariae carriers (MF group), 50-70% of adenolymphangitis (ADL) patients, 10-25% of chronic pathology (CP) patients and 10-15% of endemic normal (EN) controls. The sensitivity of the circulating filarial antigen (CFA) detection in serum samples from MF group was up to 95%. In sera from ADL patients, unexpectedly, less antigen reactivity was observed. In CP group all the CFA positive individuals were from CP grade I and II only and none from grade III or IV, suggesting that with chronicity the AWs lose fecundity and start to disintegrate and die. Amongst EN subject, 10-15% had CFA indicating that few of them harbour filarial AWs, thus they might not be truly immune as has been conventionally believed. The specificity for antigen detection was 100% when tested with sera from various other protozoan and non-filarial helminthic infections.

  12. Antigenic characterisation of lyssaviruses in South Africa

    Directory of Open Access Journals (Sweden)

    Ernest Ngoepe

    2014-02-01

    Full Text Available There are at least six Lyssavirus species that have been isolated in Africa, which include classical rabies virus, Lagos bat virus, Mokola virus, Duvenhage virus, Shimoni bat virus and Ikoma lyssavirus. In this retrospective study, an analysis of the antigenic reactivity patterns of lyssaviruses in South Africa against a panel of 15 anti-nucleoprotein monoclonal antibodies was undertaken. A total of 624 brain specimens, collected between 2005 and 2009, confirmed as containing lyssavirus antigen by direct fluorescent antibody test, were subjected to antigenic differentiation. The lyssaviruses were differentiated into two species, namely rabies virus (99.5% and Mokola virus (0.5%. Furthermore, rabies virus was further delineated into two common rabies biotypes in South Africa: canid and mongoose. Initially, it was found that the canid rabies biotype had two reactivity patterns; differential staining was observed with just one monoclonal antibody. This difference was likely to have been an artefact related to sample quality, as passage in cell culture restored staining. Mongoose rabies viruses were more heterogeneous, with seven antigenic reactivity patterns detected. Although Mokola viruses were identified in this study, prevalence and reservoir host species are yet to be established. These data demonstrate the usefulness of monoclonal antibody typing panels in lyssavirus surveillance with reference to emergence of new species or spread of rabies biotypes to new geographic zones.

  13. Lea blood group antigen on human platelets

    International Nuclear Information System (INIS)

    One- and two-stage radioligand assays were used to determine if human platelets possess the Lea antigen. Goat IgG anti-Lea antibody was purified by multiple adsorptions with Le(a-b-) human red blood cells, followed by affinity chromatography with synthetic Lea substance and labeling with 125I. Human IgG anti-Lea antibody was used either in a two stage radioassay with 125I-labeled mouse monoclonal IgG anti-human IgG as the second antibody or, alternatively, purified by Staph protein A chromatography, labeled with 125I, and used in a one-stage radioassay. Platelets from donors of appropriate red blood cell phenotypes were incubated with the antisera, centrifuged through phthalate esters, and assayed in a gamma scintillation counter. Dose response and saturation curve analysis demonstrate the presence of Lewis a antigen on platelets from Lea+ donors. Furthermore, platelets from an Le(a-b-) donor incubated in Le (a+b-) plasma adsorb Lea antigen in a similar manner to red blood cells. The clinical significance of these antigens in platelet transfusion remains undefined

  14. Ultraviolet radiation (UVR) induces cell-surface Ro/SSA antigen expression by human keratinocytes in vitro: a possible mechanism for the UVR induction of cutaneous lupus lesions

    Energy Technology Data Exchange (ETDEWEB)

    Jones, S.K. (Bristol Royal Infirmary (United Kingdom))

    1992-06-01

    Antinuclear antibodies are useful markers of connective tissue disease. In this study, UVB but not UVA induced the expression of Ro/SSA antigen on keratinocyte surfaces in vitro. This expression was also found with the extractable nuclear antigens RnP and Sm, but not with single or double-stranded DNA. The expression was prevented by blocking protein synthesis, suggesting that it was an active process. The results suggest that UVB exposure may result in the expression of Ro/SSA antigen on the surfaces of basal keratinocytes in vivo. This antigen could then bind circulating antibody leading to the cutaneous lesions in neonatal and subacute cutaneous lupus erythematosus. (Author).

  15. Effect of anticancer therapy on Tn antigen exposure on the leucocyte membranes in patients with leukemia

    Directory of Open Access Journals (Sweden)

    G. S. Maslak

    2014-08-01

    Full Text Available Tn-antigen (Thomsen-nouvelle antigen is tumor-associated carbohydrate antigen with only one GalNAc residue attached to serine or threonine of polypeptide chain. There is not enough data about the expression of this glycotope in hematologic processes. But the correlations between increasing Tn-antigen expression on the cell surface and tumor growth progression, invasion, and activation of cell migration are well known. Therefore, the currently important area of modern research is studying of the impact of anticancer therapy by expression of this carbohydrate antigen in the onco-proliferative process. There are two types of cytostatic therapies in clinical hospitals of Ukraine: COP-therapy (cyclophosphamide, vincristine, prednisone and FC-therapy (fludarabine, cyclophosphamide, which are the most popular due to their effectiveness and low price. The aim of our study was to investigate Tn-antigen exposure on the surface of lymphocytes, monocytes and granulocytes in polycythemia vera and subleukemic myelosis; to examine the influence of COP- and FC-therapies on Tn-antigen exponation in patients with chronic lymphocytic leukemia. The objects of the study were blood cells of patients with chronic lymphocytic leukemia (n = 25, polycythemia vera (n = 15 and subleukemic myelosis (n = 15 aged 58–66 years. Healthy hematologic volunteers (n = 15 aged 55 to 65 years were in the control group. Lymphocytes of patients with chronic lymphocytic leukemia (n = 25 were also studied after the chemotherapy treatment of patients divided into two groups: those who took COP-therapy (n = 13; and those who treated with FC-therapy (n = 12. Tn-antigen exposure on lymphocytes, monocytes and granulocytes was investigated by Beckman Сoulter EPICS flow cytometer with primary monoclonal Tn-antigen anybodies (Institute of Immunology, Moscow, Russia and secondary fluorescein isothiocyanate labeled antybodies (Millipore, USA. The number of dead cells was monitored by binding

  16. Human leukocyte Antigen-DM polymorphisms in autoimmune diseases.

    Science.gov (United States)

    Alvaro-Benito, Miguel; Morrison, Eliot; Wieczorek, Marek; Sticht, Jana; Freund, Christian

    2016-08-01

    Classical MHC class II (MHCII) proteins present peptides for CD4(+) T-cell surveillance and are by far the most prominent risk factor for a number of autoimmune disorders. To date, many studies have shown that this link between particular MHCII alleles and disease depends on the MHCII's particular ability to bind and present certain peptides in specific physiological contexts. However, less attention has been paid to the non-classical MHCII molecule human leucocyte antigen-DM, which catalyses peptide exchange on classical MHCII proteins acting as a peptide editor. DM function impacts the presentation of both antigenic peptides in the periphery and key self-peptides during T-cell development in the thymus. In this way, DM activity directly influences the response to pathogens, as well as mechanisms of self-tolerance acquisition. While decreased DM editing of particular MHCII proteins has been proposed to be related to autoimmune disorders, no experimental evidence for different DM catalytic properties had been reported until recently. Biochemical and structural investigations, together with new animal models of loss of DM activity, have provided an attractive foundation for identifying different catalytic efficiencies for DM allotypes. Here, we revisit the current knowledge of DM function and discuss how DM function may impart autoimmunity at the organism level.

  17. Cysteine proteases as potential antigens in antiparasitic DNA vaccines

    DEFF Research Database (Denmark)

    Jørgensen, Louise von Gersdorff; Buchmann, Kurt

    2011-01-01

    En litteraturgennemgang af muligheder for at bruge cystein proteaser som antigener i antiparasitære vacciner.......En litteraturgennemgang af muligheder for at bruge cystein proteaser som antigener i antiparasitære vacciner....

  18. Dengue viruses cluster antigenically but not as discrete serotypes

    NARCIS (Netherlands)

    L. Katzelnick (Leah); J.M. Fonville (Judith); G.D. Gromowski (Gregory D.); J.B. Arriaga (Jose Bustos); A. Green (Angela); S.L. James (Sarah ); L. Lau (Louis); M. Montoya (Magelda); C. Wang (Chunling); L.A. Van Blargan (Laura A.); C.A. Russell (Colin); H.M. Thu (Hlaing Myat); T.C. Pierson (Theodore C.); P. Buchy (Philippe); J.G. Aaskov (John G.); J.L. Muñoz-Jordán (Jorge L.); N. Vasilakis (Nikos); R.V. Gibbons (Robert V.); R.B. Tesh (Robert B.); A.D.M.E. Osterhaus (Albert); R.A.M. Fouchier (Ron); A. Durbin (Anna); C.P. Simmons (Cameron P.); E.C. Holmes (Edward C.); E. Harris (Eva); S.S. Whitehead (Stephen S.); D.R. Smith (Derek Richard)

    2015-01-01

    textabstractThe four genetically divergent dengue virus (DENV) types are traditionally classified as serotypes. Antigenic and genetic differences among the DENV types influence disease outcome, vaccine-induced protection, epidemic magnitude, and viral evolution.We scharacterized antigenic diversity

  19. Comparison of E and NS1 antigens capture ELISA to detect dengue viral antigens from mosquitoes

    Directory of Open Access Journals (Sweden)

    Day-Yu Chao

    2015-01-01

    Interpretation & conclusion: With the future potential of antigen capture ELISA to be used in the resource deprived regions, the study showed that E-ELISA has similar sensitivity and antigen stability as NS1 Ag kit to complement the current established virological surveillance in human. The improvement of the sensitivity in detecting DENV-3/4 will be needed to incorporate this method into routine mosquito surveillance system.

  20. [Identification of serological antigens in excretory-secretory antigens of Trichinella spiralis muscle larvae].

    Science.gov (United States)

    Huang, Xuegui; He, Lifang; Yuan, Shishan; Liu, Hui; Wang, Xin

    2016-05-01

    Objective To isolate and identify serological antigens in the excretory-secretory antigens of Trichinella spiralis muscle larvae by the combination of co-immunoprecipitation and mass spectrometric technology. Methods The serum IgG of New Zealand rabbits infected with Trichinella spiralis was isolated by ammonium sulfate precipitation. Muscle larvaes were isolated from the infected muscle, and then purified and cultured to collect excretory-secretory antigens. Serological antigens in excretory-secretory antigens were isolated by co-immunoprecipitation and SDS-PAGE, and analyzed by Western blotting. Moreover, the protein bands in New Zealand rabbit sera infected with Trichinella spiralis were identified by mass spectrometric technology. Results Indirect ELISA showed that the titer of serum antibody of New Zealand rabbits infected with Trichinella spiralis was 1:6400. The rabbit serum IgG was effectively isolated by ammonium sulfate precipitation. A total of four clear protein bands of the excretory-secretory antigens of Trichinella spiralis were obtained by electrophoresis. Among them, three clear protein bands with relative molecular mass (Mr) being 40 kDa, 50 kDa and 83 kDa were recognized by the rabbit sera infected with Trichinella spiralis but not recognized by the normal rabbit sera. The obtained four protein molecules were confirmed as serine protease, specific serine protease of muscle larvae, 43 kDa secreted glycoprotein and 53 kDa excretory-secretory antigen. Conclusion Four proteins were obtained from the excretory-secretory antigens of Trichinella spiralis muscle larvae by combination of co-immunoprecipitation and mass spectrometric technique analysis, which provided new sources and insights for the diagnosis and vaccine candidates of Trichinellosis. PMID:27126943

  1. A novel strategy for the development of selective active-site inhibitors of the protein tyrosine phosphatase-like proteins islet-cell antigen 512 (IA-2) and phogrin (IA-2 beta)

    DEFF Research Database (Denmark)

    Drake, P.G.; Peters, Günther H.j.; Andersen, H.S.;

    2003-01-01

    . Thus molecular modelling of this classical, competitive inhibitor in the catalytic site of wild-type IA-2beta identified two residues (Ser(762) and ASP(933)) that offer the possibility for unique interaction with an appropriately modified 'Compound 6'. Such a compound has the potential to be a highly...... indicate that IA-2 may be involved in granule trafficking and exocytosis. To further 9 understand their function, we have embarked upon developing low-molecular-mass inhibitors of IA-2 and IA-2. Previously, we have shown that a general PTP inhibitor, 2-(oxalylamino)benzoic acid (OBA), can be developed...... into highly selective and potent inhibitors of PTP1B. However. since wild-type IA-2 and IA-2beta lack conventional PTP activity, a novel strategy was designed whereby catalytically active species were generated by 'back-mutating key non-consensus catalytic region residues to those of PTP1B. These mutants were...

  2. A prospective study of serum tumour markers carcinoembryonic antigen, carbohydrate antigens 50 and 242, tissue polypeptide antigen and tissue polypeptide specific antigen in the diagnosis of pancreatic cancer with special reference to multivariate diagnostic score.

    OpenAIRE

    Pasanen, P. A.; Eskelinen, M.; Partanen, K.; Pikkarainen, P; Penttilä, I.; Alhava, E

    1994-01-01

    The aim of this study was to assess by a stepwise multivariate discriminant analysis the value of four current serum tumour markers - carcinoembryonic antigen (CEA), carbohydrate antigen (CA) 50 and CA 242 and tissue polypeptide antigen (TPA) - and a new serum tumour marker, tissue polypeptide specific antigen (TPS), in the diagnosis of pancreatic cancer. The serum values were measured in a prospective series of patients with jaundice, with unjaundiced cholestasis and with a suspicion of chro...

  3. Antigenic community between Schistosoma mansoni and Biomphalaria glabrata: on the search of candidate antigens for vaccines

    Directory of Open Access Journals (Sweden)

    N Chacón

    2002-10-01

    Full Text Available We have previously confirmed the presence of common antigens between Schistosoma mansoni and its vector, Biomphalaria glabrata. Cross-reactive antigens may be important as possible candidates for vaccine and diagnosis of schistosomiasis. Sera from outbred mice immunized with a soluble Biomphalaria glabrata antigen (SBgA of non-infected B. glabrata snails recognized molecules of SBgA itself and S. mansoni AWA by Western blot. Recognition of several molecules of the SBgA were inhibited by pre-incubation with AWA (16, 30, 36, 60 and 155 kDa. The only specific molecule of AWA, inhibited by SBgA, was a 120 kDa protein. In order to determine which epitopes of SBgA were glycoproteins, the antigen was treated with sodium metaperiodate and compared with non-treated antigen. Molecules of 140, 60 and 24 kDa in the SBgA appear to be glycoproteins. Possible protective effects of the SBgA were evaluated immunizing outbred mice in two different experiments using Freund's Adjuvant. In the first one (12 mice/group, we obtained a significant level of protection (46% in the total worm load, with a high variability in worm recovery. In the second experiment (22 mice/group, no significant protection was observed, neither in worm load nor in egg production per female. Our results suggest that SBgA constitutes a rich source of candidate antigens for diagnosis and prophylactic studies.

  4. Antigenic profiling of Yersinia pestis infection in the Wyoming coyote (Canis latrans)

    Science.gov (United States)

    Vernati, G.; Edwards, W.H.; Rocke, T.E.; Little, S.F.; Andrews, G.P.

    2011-01-01

    Although Yersinia pestis is classified as a "high-virulence" pathogen, some host species are variably susceptible to disease. Coyotes (Canis latrans) exhibit mild, if any, symptoms during infection, but antibody production occurs postinfection. This immune response has been reported to be against the F1 capsule, although little subsequent characterization has been conducted. To further define the nature of coyote humoral immunity to plague, qualitative serology was conducted to assess the antiplague antibody repertoire. Humoral responses to six plasmid-encoded Y. pestis virulence factors were first examined. Of 20 individual immune coyotes, 90% were reactive to at least one other antigen in the panel other than F1. The frequency of reactivity to low calcium response plasmid (pLcr)-encoded Yersinia protein kinase A (YpkA) and Yersinia outer protein D (YopD) was significantly greater than that previously observed in a murine model for plague. Additionally, both V antigen and plasminogen activator were reactive with over half of the serum samples tested. Reactivity to F1 was markedly less frequent in coyotes (35%). Twenty previously tested antibody-negative samples were also examined. While the majority were negative across the panel, 15% were positive for 1-3 non-F1 antigens. In vivo-induced antigen technology employed to identify novel chromosomal genes of Y. pestis that are up-regulated during infection resulted in the identification of five proteins, including a flagellar component (FliP) that was uniquely reactive with the coyote serum compared with immune serum from two other host species. Collectively, these data suggest that humoral immunity to pLcr-encoded antigens and the pesticin plasmid (pPst)-encoded Pla antigen may be relevant to plague resistance in coyotes. The serologic profile of Y. pestis chromosomal antigens up-regulated in vivo specific to C. latrans may provide insight into the differences in the pathogen-host responses during Y. pestis infection.

  5. Blood group antigen studies using CdTe quantum dots and flow cytometry.

    Science.gov (United States)

    Cabral Filho, Paulo E; Pereira, Maria I A; Fernandes, Heloise P; de Thomaz, Andre A; Cesar, Carlos L; Santos, Beate S; Barjas-Castro, Maria L; Fontes, Adriana

    2015-01-01

    New methods of analysis involving semiconductor nanocrystals (quantum dots [QDs]) as fluorescent probes have been highlighted in life science. QDs present some advantages when compared to organic dyes, such as size-tunable emission spectra, broad absorption bands, and principally exceptional resistance to photobleaching. Methods applying QDs can be simple, not laborious, and can present high sensibility, allowing biomolecule identification and quantification with high specificity. In this context, the aim of this work was to apply dual-color CdTe QDs to quantify red blood cell (RBC) antigen expression on cell surface by flow cytometric analysis. QDs were conjugated to anti-A or anti-B monoclonal antibodies, as well as to the anti-H (Ulex europaeus I) lectin, to investigate RBCs of A1, B, A1B, O, A2, and Aweak donors. Bioconjugates were capable of distinguishing the different expressions of RBC antigens, both by labeling efficiency and by flow cytometry histogram profile. Furthermore, results showed that RBCs from Aweak donors present fewer amounts of A antigens and higher amounts of H, when compared to A1 RBCs. In the A group, the amount of A antigens decreased as A1 > A3 > AX = Ael, while H antigens were AX = Ael > A1. Bioconjugates presented stability and remained active for at least 6 months. In conclusion, this methodology with high sensibility and specificity can be applied to study a variety of RBC antigens, and, as a quantitative tool, can help in achieving a better comprehension of the antigen expression patterns on RBC membranes. PMID:26185442

  6. Targeting of tumor-associated antigens (TAA) in experimental immunotherapy

    International Nuclear Information System (INIS)

    We have previously shown the superiority of tumor-associated antigens (TAA) to function as effective immunogens when administered with bilayer membrane vesicles called liposomes. The ability of liposomes to target TAA to host antigen-presenting cells is analyzed here. 1-Butanol extracted TAA from two syngeneic rat colon cancer tumors (WB 2054 and W 1756) was radioiodinated (131I-TAA). Free 131I and 131I-TAA (2.8 X 10(7) cpm and 75 micrograms TAA per rat) were used as tracers, with or without incorporation into liposomes (composition: sphingomyelin, cholesterol, dicetyl phosphate at 70:24:6 molar ratio). Six groups of male rats (BN X WF for WB2054 and Wistar/Furth for W1756, n = 18 each group) were injected iv with either free tracers or the tracers incorporated into liposomes. Whole blood clearance curve was biphasic (half-life alpha = 5 min; half life beta = 12 hr), suggesting a two-compartmental model of distribution. Seven animals from each group were sacrificed at set times (15 min to 48 hr), organs harvested and cpm/g of tissue estimated. Liposome 131I and liposome 131I-TAA were targeted to and retained preferentially in liver and spleen. Four animals from each group were imaged serially using a gamma camera. Matched pair analysis of regions showed persistently higher activity in liver-spleen area when liposomes were used (P less than 0.001). The uptake of radiolabeled antigens by plastic adherent mononuclear cells in liver and spleen was significantly higher when presented with liposomes (macrophage uptake index: liver = 1.65 vs 0.55; spleen = 5.85 vs 1.15; with and without liposomes, respectively)

  7. Performance of calibration standards for antigen quantitation with flow cytometry.

    Science.gov (United States)

    Lenkei, R; Gratama, J W; Rothe, G; Schmitz, G; D'hautcourt, J L; Arekrans, A; Mandy, F; Marti, G

    1998-10-01

    In the frame of the activities initiated by the Task Force for Antigen Quantitation of the European Working Group on Clinical Cell Analysis (EWGCCA), an experiment was conducted to evaluate microbead standards used for quantitative flow cytometry (QFCM). An unified window of analysis (UWA) was established on three different instruments (EPICS XL [Coulter Corporation, Miami, FL], FACScan and FACS Calibur [Becton Dickinson, San Jose, CA]) with QC3 microbeads (FCSC, PR). By using this defined fluorescence intensity scale, the performance of several monoclonal antibodies directed to CD3, CD4, and CD8 (conjugated and unconjugated), from three manufacturers (BDIS, Coulter [Immunotech], and DAKO) was tested. In addition, the QIFI system (DAKO) and QuantiBRITE (BDIS), and a method of relative fluorescence intensity (RFI, method of Giorgi), were compared. mAbs reacting with three more antigens, CD16, CD19, and CD38 were tested on the FACScan instrument. Quantitation was carried out using a single batch of cryopreserved peripheral blood leukocytes, and all tests were performed as single color analyses. Significant correlations were observed between the antibody-binding capacity (ABC) values of the same CD antigen measured with various calibrators and with antibodies differing in respect to vendor, labeling and possible epitope recognition. Despite the significant correlations, the ABC values of most monoclonal antibodies differed by 20-40% when determined by the different fluorochrome conjugates and different calibrators. The results of this study indicate that, at the present stage of QFCM consistent ABC values may be attained between laboratories provided that a specific calibration system is used including specific calibrators, reagents, and protocols.

  8. Lipid motif of a bacterial antigen mediates immune responses via TLR2 signaling.

    Directory of Open Access Journals (Sweden)

    Amit A Lugade

    Full Text Available The cross-talk between the innate and the adaptive immune system is facilitated by the initial interaction of antigen with dendritic cells. As DCs express a large array of TLRs, evidence has accumulated that engagement of these molecules contributes to the activation of adaptive immunity. We have evaluated the immunostimulatory role of the highly-conserved outer membrane lipoprotein P6 from non-typeable Haemophilus influenzae (NTHI to determine whether the presence of the lipid motif plays a critical role on its immunogenicity. We undertook a systematic analysis of the role that the lipid motif plays in the activation of DCs and the subsequent stimulation of antigen-specific T and B cells. To facilitate our studies, recombinant P6 protein that lacked the lipid motif was generated. Mice immunized with non-lipidated rP6 were unable to elicit high titers of anti-P6 Ig. Expression of the lipid motif on P6 was also required for proliferation and cytokine secretion by antigen-specific T cells. Upregulation of T cell costimulatory molecules was abrogated in DCs exposed to non-lipidated rP6 and in TLR2(-/- DCs exposed to native P6, thereby resulting in diminished adaptive immune responses. Absence of either the lipid motif on the antigen or TLR2 expression resulted in diminished cytokine production from stimulated DCs. Collectively, our data suggest that the lipid motif of the lipoprotein antigen is essential for triggering TLR2 signaling and effective stimulation of APCs. Our studies establish the pivotal role of a bacterial lipid motif on activating both innate and adaptive immune responses to an otherwise poorly immunogenic protein antigen.

  9. 21 CFR 866.3402 - Plasmodium species antigen detection assays.

    Science.gov (United States)

    2010-04-01

    ... 21 Food and Drugs 8 2010-04-01 2010-04-01 false Plasmodium species antigen detection assays. 866... Plasmodium species antigen detection assays. (a) Identification. A Plasmodium species antigen detection assay... malaria caused by the four malaria species capable of infecting humans: Plasmodium falciparum,...

  10. Immunity to intracellular Salmonella depends on surface-associated antigens.

    Directory of Open Access Journals (Sweden)

    Somedutta Barat

    Full Text Available Invasive Salmonella infection is an important health problem that is worsening because of rising antimicrobial resistance and changing Salmonella serovar spectrum. Novel vaccines with broad serovar coverage are needed, but suitable protective antigens remain largely unknown. Here, we tested 37 broadly conserved Salmonella antigens in a mouse typhoid fever model, and identified antigen candidates that conferred partial protection against lethal disease. Antigen properties such as high in vivo abundance or immunodominance in convalescent individuals were not required for protectivity, but all promising antigen candidates were associated with the Salmonella surface. Surprisingly, this was not due to superior immunogenicity of surface antigens compared to internal antigens as had been suggested by previous studies and novel findings for CD4 T cell responses to model antigens. Confocal microscopy of infected tissues revealed that many live Salmonella resided alone in infected host macrophages with no damaged Salmonella releasing internal antigens in their vicinity. In the absence of accessible internal antigens, detection of these infected cells might require CD4 T cell recognition of Salmonella surface-associated antigens that could be processed and presented even from intact Salmonella. In conclusion, our findings might pave the way for development of an efficacious Salmonella vaccine with broad serovar coverage, and suggest a similar crucial role of surface antigens for immunity to both extracellular and intracellular pathogens.

  11. 21 CFR 660.40 - Hepatitis B Surface Antigen.

    Science.gov (United States)

    2010-04-01

    ... 21 Food and Drugs 7 2010-04-01 2010-04-01 false Hepatitis B Surface Antigen. 660.40 Section 660.40...) BIOLOGICS ADDITIONAL STANDARDS FOR DIAGNOSTIC SUBSTANCES FOR LABORATORY TESTS Hepatitis B Surface Antigen § 660.40 Hepatitis B Surface Antigen. (a) Proper name and definition. The proper name of this...

  12. Mapping epitopes and antigenicity by site-directed masking

    Science.gov (United States)

    Paus, Didrik; Winter, Greg

    2006-06-01

    Here we describe a method for mapping the binding of antibodies to the surface of a folded antigen. We first created a panel of mutant antigens (-lactamase) in which single surface-exposed residues were mutated to cysteine. We then chemically tethered the cysteine residues to a solid phase, thereby masking a surface patch centered on each cysteine residue and blocking the binding of antibodies to this region of the surface. By these means we mapped the epitopes of several mAbs directed to -lactamase. Furthermore, by depleting samples of polyclonal antisera to the masked antigens and measuring the binding of each depleted sample of antisera to unmasked antigen, we mapped the antigenicity of 23 different epitopes. After immunization of mice and rabbits with -lactamase in Freund's adjuvant, we found that the antisera reacted with both native and denatured antigen and that the antibody response was mainly directed to an exposed and flexible loop region of the native antigen. By contrast, after immunization in PBS, we found that the antisera reacted only weakly with denatured antigen and that the antibody response was more evenly distributed over the antigenic surface. We suggest that denatured antigen (created during emulsification in Freund's adjuvant) elicits antibodies that bind mainly to the flexible regions of the native protein and that this explains the correlation between antigenicity and backbone flexibility. Denaturation of antigen during vaccination or natural infections would therefore be expected to focus the antibody response to the flexible loops. backbone flexibility | Freund's adjuvant | conformational epitope | antisera

  13. ERAP1 functions override the intrinsic selection of specific antigens as immunodominant peptides, thereby altering the potency of antigen-specific cytolytic and effector memory T-cell responses.

    Science.gov (United States)

    Rastall, David P W; Aldhamen, Yasser A; Seregin, Sergey S; Godbehere, Sarah; Amalfitano, Andrea

    2014-12-01

    Endoplasmic reticulum aminopeptidase 1 (ERAP1) is a critical component of the adaptive immune system that has been shown to increase or decrease the presentation of specific peptides on MHC class I molecules. Here, we have demonstrated that ERAP1 functions are not only important during the presentation of antigen-derived peptides, but these functions can also completely change which antigen-derived peptides ultimately become selected as immunodominant T-cell epitopes. Our results suggest that ERAP1 may do this by destroying epitopes that would otherwise become immunodominant in the absence of adequate ERAP1 functionality. We further establish that ERAP1-mediated influences on T-cell functions are both qualitative and quantitative, by demonstrating that loss of ERAP1 function redirects CTL killing toward a different set of antigen-derived epitopes and increases the percent of antigen-specific memory T cells elicited by antigen exposure. As a result, our studies suggest that normal ERAP1 activity can act to suppress the numbers of T effector memory cells that respond to a given antigen. This unique finding may shed light on why certain ERAP1 single nucleotide polymorphisms are associated with several autoimmune diseases, for example, by significantly altering the robustness and quality of CD8+ T-cell memory responses to antigen-derived peptides. PMID:25087231

  14. Seoul virus suppresses NF-κB-mediated inflammatory responses of antigen presenting cells from Norway rats

    OpenAIRE

    Au, Rebecca Y.; Jedlicka, Anne E.; Li, Wei; Pekosz, Andrew; Klein, Sabra L.

    2010-01-01

    Hantavirus infection reduces antiviral defenses, increases regulatory responses, and causes persistent infection in rodent hosts. To address whether hantaviruses alter the maturation and functional activity of antigen presenting cells (APCs), rat bone marrow-derived dendritic cells (BMDCs) and macrophages (BMDMs) were generated and infected with Seoul virus (SEOV) or stimulated with TLR ligands. SEOV infected both DCs and macrophages, but copies of viral RNA, viral antigen, and infectious vir...

  15. Antigens of Brucella abortus S19 immunodominant for bovine lymphocytes as identified by one- and two-dimensional cellular immunoblotting.

    OpenAIRE

    Brooks-Worrell, B M; Splitter, G A

    1992-01-01

    Cellular immune responses are influential for protection against intracellular bacteria such as brucellae. Therefore, identification of Brucella abortus antigens that activate primed bovine lymphocytes is fundamental for discerning the breadth of cellular response in bovine brucellosis. Potentially antigenic components of B. abortus S19 were isolated by sodium dodecyl sulfate-polyacrylamide gel electrophoresis followed by nitrocellulose blotting. Specific one-dimensional blot segments induced...

  16. Vaccination with intestinal tract antigens does not induce protective immunity in a permissive model of filariasis.

    Science.gov (United States)

    Morris, C Paul; Torrero, Marina N; Larson, David; Evans, Holly; Shi, Yinghui; Cox, Rachel T; Mitre, Edward

    2013-09-01

    Antigens obtained from the intestinal tract of filarial nematodes have been proposed as potential safe and effective vaccine candidates. Because they may be 'hidden' from the immune response during natural infection, yet accessible by antibodies induced by vaccination, intestinal antigens may have a low potential for eliciting allergic responses when vaccinating previously infected individuals. Despite prior promising data, vaccination with intestinal antigens has yet to be tested in a permissive model of filariasis. In this study we investigated the efficacy of vaccination with filarial intestinal antigens in the permissive Litomosoides sigmodontis BALB/c model of filariasis, and we evaluated the extent to which these antigens are recognized by the immune system during and after infection. Infected BALB/c mice developed lower IgG antibody responses to soluble intestinal antigens (GutAg) than to soluble antigens of whole worms (LsAg). Similarly, GutAg induced less proliferation and less production of IL-4 and IFNγ from splenocytes of infected mice than LsAg. In contrast to these differences, active infection resulted in equivalent levels of circulating GutAg-specific IgE and LsAg-specific IgE levels. Consistent with this, basophil activation, as assessed by flow cytometric staining of intracellular basophil IL-4 expression, was equivalent in response to GutAg and LsAg. Vaccination with GutAg adsorbed to CpG/alum induced GutAg specific IgG1 and IgG2A production, with GutAg specific IgG titers greater than 5-fold higher than those measured in previously infected animals. Despite this response to GutAg vaccination, vaccinated mice harbored similar parasite burdens 8 weeks post infection when compared to non-vaccinated controls. These studies demonstrate that soluble antigens obtained from the intestinal tracts of L. sigmodontis have some qualities of 'hidden' antigens, but they still sensitize mice to allergic reactions and fail to protect against future infection

  17. Expression cloning and chromosomal mapping of the leukocyte activation antigen CD97, a new seven-span transmembrane molecule of the secretin receptor superfamily with an unusual extracellular domain

    Energy Technology Data Exchange (ETDEWEB)

    Hamann, J. [Univ. of Amsterdam (Netherlands)]|[Max Planck Society, Berlin-Buch (Germany); Hamann, D.; Lier, R.A.W. [Univ. of Amsterdam (Netherlands)] [and others

    1995-08-15

    CD97 is a monomeric glycoprotein of 75 to 85 kDa that is induced rapidly on the surface of most leukocytes upon activation. We herein report the isolation of a cDNA encoding human CD97 by expression cloning in COS cells. The 3-kb cDNA clone encodes a mature polypeptide chain of 722 amino acids with a predicted molecular mass of 79 kDa. Within the C-terminal part of the protein, a region with seven hydrophobic segments was identified, suggesting that CD97 is a seven-span transmembrane molecule. Sequence comparison indicates that CD97 is the first leukocyte Ag in a recently described superfamily that includes the receptors for secretin, calcitonin, and other mammalian and insect peptide hormones. Different from these receptors, CD97 has an extended extracellular region of 433 amino acids that possesses three N-terminal epidermal growth factor-like domains, two of them with a calcium-binding site, and single Arg-Gly-Asp (RGD) motif. The existence of structural elements characteristic for extracellular matrix proteins in a seven-span transmembrane molecule makes CD97 a receptor potentially involved in both adhesion and signaling processes early after leukocyte activation. The gene encoding CD97 is localized on chromosome 19 (19p13.12-13.2).

  18. Discovering naturally processed antigenic determinants that confer protective T cell immunity

    DEFF Research Database (Denmark)

    Gilchuk, Pavlo; Spencer, Charles T; Conant, Stephanie B;

    2013-01-01

    CD8+ T cells (TCD8) confer protective immunity against many infectious diseases, suggesting that microbial TCD8 determinants are promising vaccine targets. Nevertheless, current T cell antigen identification approaches do not discern which epitopes drive protective immunity during active infectio...

  19. Tolerization of an established αb-crystallin-reactive T-cell response by intravenous antigen

    NARCIS (Netherlands)

    Verbeek, R.; Mark, K. van der; Wawrousek, E.F.; Plomp, A.C.; Noort, J.M. van

    2007-01-01

    Tolerance induction to prevent activation of a naïve T-cell repertoire has been well documented in rodents and can be readily achieved by intravenous, oral or intranasal administration of antigen in the absence of adjuvants. In autoimmune diseases such as multiple sclerosis (MS) the presence of an e

  20. Purification and characterization of fetal hematopoietic cells that express the common acute lymphoblastic leukemia antigen (CALLA)

    DEFF Research Database (Denmark)

    Hokland, P; Rosenthal, P; Griffin, J D;

    1983-01-01

    Fetal hematopoietic cells that express the common acute lymphoblastic leukemia antigen (CALLA) were purified from both fetal liver and fetal bone marrow by immune rosetting with sheep erythrocytes coated with rabbit anti-mouse immunoglobulin and by fluorescence-activated cell sorting. Dual fluore...... that these cells are relatively immature lymphoid cells, CALLA+ cells do not appear to contain either myeloid precursor cells (CFU-G/M) or the earliest lymphoid stem cells. Udgivelsesdato: 1983-Jan-1......Fetal hematopoietic cells that express the common acute lymphoblastic leukemia antigen (CALLA) were purified from both fetal liver and fetal bone marrow by immune rosetting with sheep erythrocytes coated with rabbit anti-mouse immunoglobulin and by fluorescence-activated cell sorting. Dual...... fluorescence techniques disclosed that these cells were heterogenous with respect to the expression of a series of differentiation and activation antigens defined by monoclonal antibodies. Thus, whereas all CALLA+ cells were Ia+ and expressed two activation antigens, J2 and T10, only 30-50% expressed B1...

  1. Composition of inflammatory infiltrate and its correlation with HBV/HCV antigen expression

    Institute of Scientific and Technical Information of China (English)

    Bozena Walewska-Zielecka; Kazimierz Madalinski; Joanna Jablonska; Paulina Godzik; Joanna Cielecka-Kuszyk; Bogumila Litwinska

    2008-01-01

    AIM: To study the composition of liver inflammatory infiltrate in biopsy material from patients chronically infected with hepatotropic viruses and to evaluate the correlation of inflammatory infiltrate with hepatitis B virus (HBV) and hepatitis C virus (HCV) viral antigen expression in chronic B and C hepatitis.METHODS: The phenotype of inflammatory cells was evaluated by the EnVision system, using a panel of monoclonal antibodies. HBV and HCV antigens were detected with the use of monoclonal anti-HBs, poly-clonal anti-HBc and anti-HCV antibodies, respectively.RESULTS: The cellular composition of liver inflammatory infiltrate was similar in the patients with B and C hepatitis: ~50%-60% of cells were T helper lymphooltes. Approximately 25% were T cytotoxic lymphocytes; B lymphocytes comprised 15% of inflammatory infiltrate; other cells, including NK, totalled 10%. Expression of HLA antigens paralleled inflammatory activity. Portal lymphadenoplasia was found more often in hepatitis C (54.5%) than in hepatitis B (30.6%). Expression of HB-cAg was found more often in chronic B hepatitis of moderate or severe activity. Overall inflammatory activity in HBV-infected cases did not correlate with the intensity of HBsAg expression in hepatooltes. Inflammatory infiltrates accompanied the focal expression of HCV anti-gens. A direct correlation between antigen expression and inflammatory reaction in situ was noted more often in hepatitis C than B.CONCLUSION: Irrespective of the etiology and activity of hepatitis, components of the inflammatory infiltrate in liver were similar. Overall inflammatory activity did not correlate with the expression of HBsAg and HCVAg; HBcAg expression, however, accompanied chronic hepatitis B of moderate and severe activity.

  2. Monoclonal antibodies against human Ia antigens stimulate monocytes to secrete interleukin 1.

    OpenAIRE

    Palacios, R

    1985-01-01

    The monoclonal antibodies (mAb) DA6.147, DA6.164, and HIG.48 against human Ia antigens, but not the W6/32 mAb against human class I major histocompatibility complex antigens or the anti-monocyte OKM1 and 63D3 mAb, stimulated monocytes to secrete interleukin 1 (IL-1). IL-1 was measured by its property of promoting the production of interleukin 2 (IL-2) by phytohemagglutinin-treated LBRM-33 clone 1A5 cells. IL-1 activity induced by anti-Ia antibodies could be detected 24 hr after initiation of ...

  3. Rapid production of antigen-specific monoclonal antibodies from a variety of animals

    Directory of Open Access Journals (Sweden)

    Kurosawa Nobuyuki

    2012-09-01

    Full Text Available Abstract Background Although a variety of animals have been used to produce polyclonal antibodies against antigens, the production of antigen-specific monoclonal antibodies from animals remains challenging. Results We propose a simple and rapid strategy to produce monoclonal antibodies from a variety of animals. By staining lymph node cells with an antibody against immunoglobulin and a fluorescent dye specific for the endoplasmic reticulum, plasma/plasmablast cells were identified without using a series of antibodies against lineage markers. By using a fluorescently labeled antigen as a tag for a complementary cell surface immunoglobulin, antigen-specific plasma/plasmablast cells were sorted from the rest of the cell population by fluorescence-activated cell sorting. Amplification of cognate pairs of immunoglobulin heavy and light chain genes followed by DNA transfection into 293FT cells resulted in the highly efficient production of antigen-specific monoclonal antibodies from a variety of immunized animals. Conclusions Our technology eliminates the need for both cell propagation and screening processes, offering a significant advantage over hybridoma and display strategies.

  4. Immunoregulatory adherent cells in human tuberculosis: radiation-sensitive antigen-specific suppression by monocytes

    Energy Technology Data Exchange (ETDEWEB)

    Kleinhenz, M.E.; Ellner, J.J.

    1985-07-01

    In human tuberculosis, adherent mononuclear cells (AMC) selectively depress in vitro responses to the mycobacterial antigen tuberculin purified protein derivative (PPD). The phenotype of this antigen-specific adherent suppressor cell was characterized by examining the functional activity of adherent cells after selective depletion of sheep erythrocyte-rosetting T cells or OKM1-reactive monocytes. Adherent cell suppression was studied in the (/sup 3/H)thymidine-incorporation microculture assay by using T cells rigorously depleted of T cells with surface receptors for the Fc portion of IgG (T gamma cells) as antigen-responsive cells. PPD-induced (/sup 3/H)thymidine incorporation by these non gamma T cells was uniformly reduced (mean, 42% +/- 10% (SD)) when autologous AMC were added to non gamma T cells at a ratio of 1:2. Antigen-specific suppression by AMC was not altered by depletion of sheep erythrocyte-rosetting T cells or treatment with indomethacin. However, AMC treated with OKM1 and complement or gamma irradiation (1,500 rads) no longer suppressed tuberculin responses in vitro. These studies identify the antigen-specific adherent suppressor cell in tuberculosis as an OKM1-reactive, non-erythrocyte-rosetting monocyte. The radiosensitivity of this monocyte immunoregulatory function may facilitate its further definition.

  5. INTRATHYMIC INOCULATION OF LIVER SPECIFIC ANTIGEN ALLEVIATES LIVER TRANSPLANT REJECTION

    Institute of Scientific and Technical Information of China (English)

    贾长库; 郑树森; 朱有法

    2004-01-01

    Objective To study the effects of liver specific antigen (LSA) on liver allotransplantation rejection. Methods Orthotopic liver transplantation was performed in this study. Group Ⅰ: syngeneic control (Wistar-to-Wistar); Group Ⅱ: acute rejection (SD-to-Wistar). Group Ⅲ: thymic inoculation of SD rat LSA day 7 before transplantation. The observation of general condition and survival time, rejection grades and the NF-κB activity of splenocytes were used to analyze severity of acute rejection and immune state of animals in different groups. Results The general condition of group Ⅰ was fair post transplantation with no sign of rejection. All recipients of group Ⅱ died within days 9 to 13 post transplantation with median survival time of 10.7 ±1.37 days. As for group Ⅲ, 5 out of 6 recipients survived for a long period with remarkably better general condition than that of group Ⅱ. Its rejection grades were significantly lower than group Ⅱ (P< 0.05).NF-κB activity was only detected in group Ⅰ between days 5 and 7 after transplantation, whereas high activity of NF-κB was detected at all points in group Ⅱ and low NF-κB activity was detected in group Ⅲ which was significantly lower than that of group Ⅱ (P < 0.05). Conclusions LSA is an important transplantation antigen directly involved in the immunorejection of liver transplantation. Intrathymic inoculation of LSA can alleviate the rejection of liver allotransplantation,grafts survive for a period of time thereby, allowing a novel way to liver transplantation immunotolerance.

  6. Delivery of Exogenous Antigens to Induce Cytotoxic CD8+ T Lymphocyte Responses

    Directory of Open Access Journals (Sweden)

    Julia Kim

    2010-01-01

    Full Text Available Vaccines intended to induce a cytotoxic CD8+ T-cell response are highly sought after. However, some of these vaccines can be problematic if they replicate in the host. An alternative strategy is to exploit cross-presentation of exogenous antigens to express peptides on major histocompatibility complex (MHC class I molecules. During cross-presentation, the delivered exogenous antigen can be taken up and processed through diverse mechanisms. Here, we will discuss the recent advances regarding the complex nature of the cross-priming process and the models that reflect its relevance in vivo. Moreover, we summarize current data that explore potential adjuvants and vaccine vectors that deliver antigens to activate CD8+ T cells relying on cross-presentation.

  7. Follow-up of neurocysticercosis patients after treatment using an antigen detection ELISA

    Directory of Open Access Journals (Sweden)

    Nguekam

    2003-03-01

    Full Text Available Seven patients with active neurocysticercosis (NCC received an eight days treatment with albendazole and were followed up using computed tomography (CT-scan and a monoclonal antibody based ELISA for the detection of circulating antigen (Ag-ELISA. Only three patients were cured as was shown by CT-scan and by the disappearance of circulating antigens one month after treatment. After a second course of albendazole therapy, two other patients became seronegative. CT-scan showed the disappearance of viable cysts in all persons who became seronegative whereas patients who were not cured remained seropositive. These preliminary results show that this Ag-ELISA is a promising technique for monitoring the success of treatment of NCC patients because of the excellent correlation between the presence of circulating antigens and of viable brain cysts.

  8. INTERACTION OF ANTIGEN AND ANTIBODY ON CORE-SHELL POLYMERIC MICROSPHERES

    Institute of Scientific and Technical Information of China (English)

    Gang Li; Qing-bin Meng; Zhan-yong Li; Ying-li An; Xiao-xia Zhu

    2011-01-01

    Monodispersed microspheres with polystyrene as the core and poly(acrylamide-co-N-acryloxysuccinimide) as the shell were synthesized by a two-step surfactant-free emulsion copolymerization. The core-shell morphology of the microspheres was shown by scanning electron microscopy and transmission electron microscopy. Rabbit immunoglobulin G (as antigen) was covalently coupled onto the microspheres by the reaction between succinimide-activated ester groups on the shell of the microspheres and amino groups of the antigen molecules. The size of particles was characterized by dynamic light scattering technique and was found to vary upon bioconjugation and interaction with proteins. The binding process was shown to be specific to goat anti-rabbit immunoglobulin G (as antibody) and reversible upon the addition of free antigen into the system.

  9. Clinical application of active B-cell antigen receptor signaling as novel therapeutic target in B-cell non-Hodgkin lymphoma%靶向B细胞抗原受体信号转导通路小分子抑制剂在B细胞非霍奇金淋巴瘤中的临床应用

    Institute of Scientific and Technical Information of China (English)

    丁宁

    2013-01-01

    The chronic B-cell antigen receptor (BCR) pathway plays an important role in malignant proliferation of B-cell lymphoma.Several tyrosine kinase inhibitors have shown impressive anti-tumor activity in relapsed and refractory B-cell non-Hodgkin lymphoma.This review discussed essential tyrosine kinases in BCR signaling pathway and present data from clinical usage of these related novel target agents.%B细胞抗原受体(BCR)信号转导通路对于B细胞非霍奇金淋巴瘤(B-NHL)恶性增殖的调控作用日益受到关注.针对BCR信号转导通路关键分子所研发的酪氨酸激酶抑制剂,已经在临床研究中初步显现出对B细胞淋巴瘤的良好疗效.文章就BCR信号转导通路中的关键酪氨酸激酶及相应靶向药物的临床应用状况作一综述.

  10. Radionuclide-labelled antigens in serological epidemiology

    International Nuclear Information System (INIS)

    The feasibility of tests using radionuclide-labelled antigens in serological surveys was studied, with particular attention to the likely availability of facilities and personnel in the tropics and arctics, where measurements may be disturbed by climatic influences. The methodology required was to be simple, rapid and suitable for examining large numbers of sera, as for epidemological surveys. In the introduction, limitations of labelled antigen tests are discussed, the choice of radionuclide and measurement methods, test procedures and evaluation of results. Collection, preservation and shipment of speciments (serum, faeces, cerebrospinal fluid, sputum, etc.) are described. Experiments with bacteria and bacterial toxins (Enterobacteriaceae, vibrios, staphylococci, meningococci, etc.), with protozoa and metazoa (Entamoeba hystolytica, Schistosoma mansoni, Trypanosoma cruzi, Plasmodia and other parasites), with viruses (vaccinia, adeno-, polio-, and influenza viruses, etc.), and with fungi are discussed

  11. A competitive-inhibiton radioimmunoassay for influenza virus envelope antigens

    International Nuclear Information System (INIS)

    A double-antibody competitive-inhibition radioimmunoassay for influenza virus envelope antigens is described. A viral antigen preparation from influenza A virus recombinant MRC11 [antigenically identical to A/Port Chalmers/1/73 (H3N2)] consisting of haemagglutinin and neuraminidase was labelled with radioiodine. Rabbit antisera were allowed to react with the labelled antigen and the resultant antigen-antibody complexes were precipitated with the appropriate antiglobulin. The competitive-inhibition radioimmunoassay very sensitively elucidated differences even among closely related influenza virus strains. Attempts have been made to eliminate neuraminidase from radioimmunoprecipitation to obtain a competitive-inhibition radioimmunoassay system for haemagglutinin alone. (author)

  12. CD4+ T cell-mediated presentation of non-infectious HIV-1virion antigens to HIV-specific CD8+ T cells

    Institute of Scientific and Technical Information of China (English)

    XU Jian-qing; Franco Lori; Julianna Lisziewicz

    2006-01-01

    Background The mechanism of chronic immune activation and impairment of HIV-specific immune responses during chronic infection is not fully understood. However, it is known that high immune activation leads to more rapid progression to AIDS. We hypothesize that CD4+ T cell-mediated viral antigen presentation contributes to this pathologic immune activation in HIV-infected individuals.Methods HIV-specific T cells, responding to noninfectious HIV-1 virions as antigen, were measured by flow cytometric assays. These experimental conditions reflect the in vivo condition where noninfectious HIV-1 represents more than 99% of the antigens.Results CD4+ T cells purified from HIV-infected individuals were capable of cross presenting exogenous noninfectious HIV-1 virions to HIV-1-specific CD8+ T cells. Cross presentation required the entry of HIV-1 to CD4+ T cells and antigen translocation from endoplasmic reticulum to the Golgi complex. Blocking CD4+mediated activation of HIV-specific CD8+ T cells and redirecting the viral antigens to antigen presenting cells improved HIV-specific T cell responses.Conclusions One possible cause of chronic immune activation and impairment of HIV-1 specific T cell responses is represented by HIV-1 harboring CD4+ T cells cross presenting HIV-1 antigen to activate CD8+ T cells. This new mechanism provides the first evidence that cross presentation of noninfectious HIV-1. Virions play a role in the immunopathogenesis of HIV-1 infection.

  13. Class II HLA antigens in multiple sclerosis.

    Science.gov (United States)

    Miller, D H; Hornabrook, R W; Dagger, J; Fong, R

    1989-01-01

    HLA typing in Wellington revealed a stronger association of multiple sclerosis with DR2 than with DQw1. The association with DQw1 appeared to be due to linkage disequilibrium of this antigen with DR2. These results, when considered in conjunction with other studies, are most easily explained by the hypothesis that susceptibility to multiple sclerosis is influenced by multiple risk factors, with DR2 being an important risk factor in Caucasoid populations. PMID:2732726

  14. Yeast retrotransposon particles as antigen delivery systems.

    Science.gov (United States)

    Kingsman, A J; Burns, N R; Layton, G T; Adams, S E

    1995-05-31

    The development of technologies to produce recombinant proteins for use in the pharmaceutical industry has made substantial advances, in particular in the area of generating antigens containing multiple copies of important immunological regions. One such antigen-carrier system is based on the ability of a protein encoded by the yeast retrotransposon, Ty, to self-assemble into virus-like particles. Ty-fusion proteins retain this ability to form particles, and a range of hybrid VLPs carrying a variety of heterologous antigens have been produced and shown to induce potent immune responses. In particular, hybrid VLPs carrying the core protein p24 of HIV (p24-VLPs) have been shown to induce antibody and T-cell proliferative responses in both experimental animals and human volunteers, and immunization of rabbits with VLPs carrying the principal neutralizing determinant of HIV (V3-VLPs) resulted in the induction of neutralizing antibody responses and T-cell proliferation. Further studies with V3-VLPs have shown that this particulate antigen stimulates enhanced V3-specific lymphoproliferative responses as compared to whole recombinant gp120 or to V3 peptide conjugated to albumin. The V3-VLPs also induce potent CTL responses following immunization of mice in the absence of adjuvant. These responses are MHC class I restricted and are mediated by CD8-positive cells. These observations therefore demonstrate that hybrid Ty-VLPs induce both humoral and cellular immune responses against HIV and suggest that these immunogens may be important in combatting AIDS and other infections. PMID:7625653

  15. Antigenicity of low molecular weight surfactant species.

    OpenAIRE

    Strayer, D. S.; Merritt, T A; Makunike, C.; Hallman, M

    1989-01-01

    The authors tested the antigenicity of human lung surfactant isolated from amniotic fluid. Mice and rabbits were immunized. Rabbit polyclonal antisera to these surfactant preparations were absorbed with normal human plasma proteins. Polyclonal antisera reacted with both high molecular weight (35 kd) surfactant apoprotein and to lower molecular weight species, both 18 kd and 9 kd. Mice were used to generate monoclonal antibodies to surfactant. Enzyme-linked immunosorbant assay was used to iden...

  16. Rationalisation of Legionella Urinary Antigen Testing.

    OpenAIRE

    Lynch, Breda

    2014-01-01

    Introduction: Legionnaires’ is a severe pneumonia, the diagnosis of which can be confirmed by a positive Legionella Urinary Antigen (LUA) test. The British Thoracic Society has specific guidelines for its use. Incorrect LUA test requests can result in false-positive results while accumulating costs. Aims and Objectives: The aim is the rationalisation of LUA testing. The first objective is to educate clinicians on indications for testing reducing unnecessary orders. The second is to develop...

  17. Tumor-Associated Antigens for Specific Immunotherapy of Prostate Cancer

    Energy Technology Data Exchange (ETDEWEB)

    Kiessling, Andrea [Biologics Safety and Disposition, Preclinical Safety, Translational Sciences, Novartis Institutes for BioMedical Research, Novartis Pharma AG, Werk Klybeck, Klybeckstraße 141, Basel CH-4057 (Switzerland); Wehner, Rebekka [Institute of Immunology, Medical Faculty, University of Technology Dresden, Fetscherstraße 74, Dresden 01307 (Germany); Füssel, Susanne [Department of Urology, Medical Faculty, University of Technology Dresden, Fetscherstraße 74, Dresden 01307 (Germany); Bachmann, Michael [Institute of Immunology, Medical Faculty, University of Technology Dresden, Fetscherstraße 74, Dresden 01307 (Germany); Wirth, Manfred P. [Department of Urology, Medical Faculty, University of Technology Dresden, Fetscherstraße 74, Dresden 01307 (Germany); Schmitz, Marc, E-mail: marc.schmitz@tu-dresden.de [Institute of Immunology, Medical Faculty, University of Technology Dresden, Fetscherstraße 74, Dresden 01307 (Germany)

    2012-02-22

    Prostate cancer (PCa) is the most common noncutaneous cancer diagnosis and the second leading cause of cancer-related deaths among men in the United States. Effective treatment modalities for advanced metastatic PCa are limited. Immunotherapeutic strategies based on T cells and antibodies represent interesting approaches to prevent progression from localized to advanced PCa and to improve survival outcomes for patients with advanced disease. CD8{sup +} cytotoxic T lymphocytes (CTLs) efficiently recognize and destroy tumor cells. CD4{sup +} T cells augment the antigen-presenting capacity of dendritic cells and promote the expansion of tumor-reactive CTLs. Antibodies mediate their antitumor effects via antibody-dependent cellular cytotoxicity, activation of the complement system, improving the uptake of coated tumor cells by phagocytes, and the functional interference of biological pathways essential for tumor growth. Consequently, several tumor-associated antigens (TAAs) have been identified that represent promising targets for T cell- or antibody-based immunotherapy. These TAAs comprise proteins preferentially expressed in normal and malignant prostate tissues and molecules which are not predominantly restricted to the prostate, but are overexpressed in various tumor entities including PCa. Clinical trials provide evidence that specific immunotherapeutic strategies using such TAAs represent safe and feasible concepts for the induction of immunological and clinical responses in PCa patients. However, further improvement of the current approaches is required which may be achieved by combining T cell- and/or antibody-based strategies with radio-, hormone-, chemo- or antiangiogenic therapy.

  18. Tumor-Associated Antigens for Specific Immunotherapy of Prostate Cancer

    International Nuclear Information System (INIS)

    Prostate cancer (PCa) is the most common noncutaneous cancer diagnosis and the second leading cause of cancer-related deaths among men in the United States. Effective treatment modalities for advanced metastatic PCa are limited. Immunotherapeutic strategies based on T cells and antibodies represent interesting approaches to prevent progression from localized to advanced PCa and to improve survival outcomes for patients with advanced disease. CD8+ cytotoxic T lymphocytes (CTLs) efficiently recognize and destroy tumor cells. CD4+ T cells augment the antigen-presenting capacity of dendritic cells and promote the expansion of tumor-reactive CTLs. Antibodies mediate their antitumor effects via antibody-dependent cellular cytotoxicity, activation of the complement system, improving the uptake of coated tumor cells by phagocytes, and the functional interference of biological pathways essential for tumor growth. Consequently, several tumor-associated antigens (TAAs) have been identified that represent promising targets for T cell- or antibody-based immunotherapy. These TAAs comprise proteins preferentially expressed in normal and malignant prostate tissues and molecules which are not predominantly restricted to the prostate, but are overexpressed in various tumor entities including PCa. Clinical trials provide evidence that specific immunotherapeutic strategies using such TAAs represent safe and feasible concepts for the induction of immunological and clinical responses in PCa patients. However, further improvement of the current approaches is required which may be achieved by combining T cell- and/or antibody-based strategies with radio-, hormone-, chemo- or antiangiogenic therapy

  19. Toxicities of chimeric antigen receptor T cells: recognition and management.

    Science.gov (United States)

    Brudno, Jennifer N; Kochenderfer, James N

    2016-06-30

    Chimeric antigen receptor (CAR) T cells can produce durable remissions in hematologic malignancies that are not responsive to standard therapies. Yet the use of CAR T cells is limited by potentially severe toxicities. Early case reports of unexpected organ damage and deaths following CAR T-cell therapy first highlighted the possible dangers of this new treatment. CAR T cells can potentially damage normal tissues by specifically targeting a tumor-associated antigen that is also expressed on those tissues. Cytokine release syndrome (CRS), a systemic inflammatory response caused by cytokines released by infused CAR T cells can lead to widespread reversible organ dysfunction. CRS is the most common type of toxicity caused by CAR T cells. Neurologic toxicity due to CAR T cells might in some cases have a different pathophysiology than CRS and requires different management. Aggressive supportive care is necessary for all patients experiencing CAR T-cell toxicities, with early intervention for hypotension and treatment of concurrent infections being essential. Interleukin-6 receptor blockade with tocilizumab remains the mainstay pharmacologic therapy for CRS, though indications for administration vary among centers. Corticosteroids should be reserved for neurologic toxicities and CRS not responsive to tocilizumab. Pharmacologic management is complicated by the risk of immunosuppressive therapy abrogating the antimalignancy activity of the CAR T cells. This review describes the toxicities caused by CAR T cells and reviews the published approaches used to manage toxicities. We present guidelines for treating patients experiencing CRS and other adverse events following CAR T-cell therapy. PMID:27207799

  20. Study of serum Helicobacter pylori soluble antigen

    Institute of Scientific and Technical Information of China (English)

    吴勤动; 朱永良

    2002-01-01

    Objective: to explore a new serological method for detecting Helicobac ter pylori ( H. pylori ) infection. Methods: Serum soluble antigen of H. p ylor i was detected by using avidin-biotin ELISA technique to evaluate the status of H. pylori infection and for comparison with rapid urease test ( RUT ), histo logi c examination and serology. Results: The sensitivity, specificity, positive pred ictive value and negative predictive value were 77.46%, 91.07%, 91.67% a nd 76.12 %, respectively. The prevalence rate of serum H. pylori soluble antigen in 138 patients undergoing endoscopy was similar to the rate obtained by 14 C-UBT met hods ( P>0.05 ). Conclusions: The detection of serum H. pylori solub le antigen( HpSAg) could be used as a new serological method which is accurate, and convenie nt, not affected by the memorizing reaction of serum antibody; is more sensitive , m ore specific and suitable for clinical diagnosis, and evaluation of eradication and for follow-up of H. pylori as well as for detection in children and pre gnant women.

  1. Study of serum Helicobacter pylori soluble antigen

    Institute of Scientific and Technical Information of China (English)

    吴勤动; 朱永良

    2002-01-01

    Objective:to explore a new serological method for detecting Helicobacter pylori(H.pylori) infection.Methods:Serum soluble antigen of H.pylori was detected by using avidin-biotin ELISA technique to evaluate the status of H.pylori infection and for comparison with rapid urease test(RUT).histologic examination and serology,Results:The sensitivity,specificity,positive predictive value and negative predictive value were 77.46% ,91.07%,91.67% and 76.12%,respectively.The prevalence rate of werum H. pylori soluble antigen in 138 patients undergong endoscopy was similar to the rate obtained by 14 C-UBT methods(P>0.05).Conclusions:The detection of serum H.pylori soluble antigen(HpSAg) could be used as a new serological method which is accurate,and convenient,not affected by the memorizing raction of serum antibody;is more sensitive,more specific and suitable for dinical diagriosis,and evaluation of eradication and for follow-up of H.pylori as well as for detection in children and pregnant women.

  2. A Novel Chimeric Antigen Receptor Against Prostate Stem Cell Antigen Mediates Tumor Destruction in a Humanized Mouse Model of Pancreatic Cancer

    Science.gov (United States)

    Lagisetty, Kiran H.; Tran, Eric; Zheng, Zhili; Gattinoni, Luca; Yu, Zhiya; Burns, William R.; Miermont, Anne M.; Teper, Yaroslav; Rudloff, Udo; Restifo, Nicholas P.; Feldman, Steven A.; Rosenberg, Steven A.; Morgan, Richard A.

    2014-01-01

    Abstract Despite advances in the understanding of its molecular pathophysiology, pancreatic cancer remains largely incurable, highlighting the need for novel therapies. We developed a chimeric antigen receptor (CAR) specific for prostate stem cell antigen (PSCA), a glycoprotein that is overexpressed in pancreatic cancer starting at early stages of malignant transformation. To optimize the CAR design, we used antigen-recognition domains derived from mouse or human antibodies, and intracellular signaling domains containing one or two T cell costimulatory elements, in addition to CD3zeta. Comparing multiple constructs established that the CAR based on human monoclonal antibody Ha1-4.117 had the greatest reactivity in vitro. To further analyze this CAR, we developed a human pancreatic cancer xenograft model and adoptively transferred CAR-engineered T cells into animals with established tumors. CAR-engineered human lymphocytes induced significant antitumor activity, and unlike what has been described for other CARs, a second-generation CAR (containing CD28 cosignaling domain) induced a more potent antitumor effect than a third-generation CAR (containing CD28 and 41BB cosignaling domains). While our results provide evidence to support PSCA as a target antigen for CAR-based immunotherapy of pancreatic cancer, the expression of PSCA on selected normal tissues could be a source of limiting toxicity. PMID:24694017

  3. Conformational dynamics and antigenicity in the disordered malaria antigen merozoite surface protein 2.

    Directory of Open Access Journals (Sweden)

    Christopher A MacRaild

    Full Text Available Merozoite surface protein 2 (MSP2 of Plasmodium falciparum is an abundant, intrinsically disordered protein that is GPI-anchored to the surface of the invasive blood stage of the malaria parasite. Recombinant MSP2 has been trialled as a component of a malaria vaccine, and is one of several disordered proteins that are candidates for inclusion in vaccines for malaria and other diseases. Nonetheless, little is known about the implications of protein disorder for the development of an effective antibody response. We have therefore undertaken a detailed analysis of the conformational dynamics of the two allelic forms of MSP2 (3D7 and FC27 using NMR spectroscopy. Chemical shifts and NMR relaxation data indicate that conformational and dynamic properties of the N- and C-terminal conserved regions in the two forms of MSP2 are essentially identical, but significant variation exists between and within the central variable regions. We observe a strong relationship between the conformational dynamics and the antigenicity of MSP2, as assessed with antisera to recombinant MSP2. Regions of increased conformational order in MSP2, including those in the conserved regions, are more strongly antigenic, while the most flexible regions are minimally antigenic. This suggests that modifications that increase conformational order may offer a means to tune the antigenicity of MSP2 and other disordered antigens, with implications for vaccine design.

  4. Conformational Dynamics and Antigenicity in the Disordered Malaria Antigen Merozoite Surface Protein 2

    Science.gov (United States)

    Andrew, Dean; Krishnarjuna, Bankala; Nováček, Jiří; Žídek, Lukáš; Sklenář, Vladimír; Richards, Jack S.; Beeson, James G.; Anders, Robin F.; Norton, Raymond S.

    2015-01-01

    Merozoite surface protein 2 (MSP2) of Plasmodium falciparum is an abundant, intrinsically disordered protein that is GPI-anchored to the surface of the invasive blood stage of the malaria parasite. Recombinant MSP2 has been trialled as a component of a malaria vaccine, and is one of several disordered proteins that are candidates for inclusion in vaccines for malaria and other diseases. Nonetheless, little is known about the implications of protein disorder for the development of an effective antibody response. We have therefore undertaken a detailed analysis of the conformational dynamics of the two allelic forms of MSP2 (3D7 and FC27) using NMR spectroscopy. Chemical shifts and NMR relaxation data indicate that conformational and dynamic properties of the N- and C-terminal conserved regions in the two forms of MSP2 are essentially identical, but significant variation exists between and within the central variable regions. We observe a strong relationship between the conformational dynamics and the antigenicity of MSP2, as assessed with antisera to recombinant MSP2. Regions of increased conformational order in MSP2, including those in the conserved regions, are more strongly antigenic, while the most flexible regions are minimally antigenic. This suggests that modifications that increase conformational order may offer a means to tune the antigenicity of MSP2 and other disordered antigens, with implications for vaccine design. PMID:25742002

  5. The radiocontrast molecule in anaphylaxis: a surprising antigen.

    Science.gov (United States)

    Lasser, Elliott C

    2004-01-01

    X-ray contrast media are individually injected into human blood vessels in greater quantities than any other pharmacological substance. Adverse reactions to these substances have heretofore been considered anaphylactoid in nature. Others and we have demonstrated that the mechanisms involved are multifactorial and may involve activation of mast cells and basophils, activation of the complement system, activation of the contact system, and the conversion of L-arginine into nitric oxide. Appropriate pretreatment with corticosteriods will diminish the incidence of reactions. Most recently we have demonstrated that the contrast media function as 'pseudoantigens' (PsA). They can combine with antibodies, but cannot themselves produce antibodies. This property appears to be dependent on aggregation in high concentrations and varies with the individual media. It furthermore appears to be non-specific in relation to antibodies, and suggests that binding occurs to the Fc portion of immunoglobulins. We have now demonstrated that the least toxic of current media demonstrate the best antibody binding and in sufficient concentration can inhibit contrast induced mast cell activation and/or non-contrast antigen induced mast cell activation, apparently due to in vivo pseudoantigen excess. In lesser concentrations and/or lesser binding, the media can trigger mast cell activation. PMID:15025400

  6. Interferon-induced changes in expression of antigens defined by monoclonal antibodies on malignant and nonmalignant mononuclear hematopoietic cells

    DEFF Research Database (Denmark)

    Hokland, M; Ritz, J; Hokland, P

    1983-01-01

    The effect of alpha interferon (alpha IFN) on the expression of histocompatibility--as well as differentiation antigens on normal and malignant hematopoietic mononuclear cells--were investigated by cell cytofluorometry using a panel of monoclonal antibodies (MoAbs). An increase in the expression...... of HLA-antigens detected by beta 2-Microglobulin (beta 2-M) could be demonstrated for peripheral blood mononuclear cells, non-T cells, Null cells, activated T cells, fetal thymocytes, adherent cells, and on four malignant non-T lymphoblastoid cell lines. In contrast, no significant differences were...... observed in the expression of antigens specific for B-lymphocytes (B1), T-lymphocytes (T3, T4, T6, T8, T11), NK-cells (901) and adherent cells (Mo1-4). Likewise, the expression of Ia-antigens remained unaltered on non-T cells, Null cells, and monocytes. The only other effect of IFN was an increase...

  7. Use of radiolabeled antibodies to carcinoembryonic antigen for the detection and localization of diverse cancers by external photoscanning

    International Nuclear Information System (INIS)

    To determine whether tumors containing carcinoembryonic antigen could be detected by administration of a radiolabeled, affinity-purified, goat IgG having 70% immunoreactivity against carcinoembryonic antigen, 18 patients with a history of cancer of diverse histopathology received an average total dose of 1.0 mCi of 131I-labeled IgG. Total-body photoscans were performed with a gamma scintillation camera at various intervals after administration of the radioactive antibody. Ordinary photoscans proved difficult to interpret because of blood-pool background radioactivity, thus necessitating the computer subtraction of radioactive blood-pool agents from the antibody's 131I activity. Tumor location could be demonstrated at 48 hours after injection in almost all cases studied. The scans were negative in patients without demonstrable tumors or with tumors apparently devoid of carcinoembryonic antigen. Circulating antigen levels of up to 350 ng per milliliter did not prevent successful tumor imaging after injection of the radioantibody

  8. Modulation of antigenicity of mycelial antigens during developmental cycle of Karnal bunt (Tilletia indica) of wheat.

    Science.gov (United States)

    Rai, G; Kumar, A; Singh, A; Garg, G K

    2000-05-01

    Indirect enzyme linked immunosorbent assays (ELISA) were developed using polyclonal antibodies against soluble cytoplasmic (SCA) and insoluble cell wall antigens (ICWA) for monitoring modulation of mycelial antigens during growth cycle of T. indica. With SCA, continuous decrease in ELISA reactivity was observed in maturing fungus cultures, suggesting that SCA were expressed predominantly during early vegetative phase and their decreasing role was apparent as the fungus matures possibly towards sporogenous mycelium. In case of ICWA, the reaction profile showed an increase up to exponential phase of growth probably due to increase in the cell division and branching of mycelium. But later, ICWA antibody reactivity was decreased which may be due to conversion of mycelial phase to sporogenous phase, a quiescent stage of growth. Characterization of changes in antigenic configuration during developmental cycle of Tilletia indica by these antibodies could prove to be useful in identification of developmentally related and virulence marker(s).

  9. New diagnostic antigens for early trichinellosis: the excretory-secretory antigens of Trichinella spiralis intestinal infective larvae.

    Science.gov (United States)

    Sun, Ge Ge; Liu, Ruo Dan; Wang, Zhong Quan; Jiang, Peng; Wang, Li; Liu, Xiao Lin; Liu, Chun Yin; Zhang, Xi; Cui, Jing

    2015-12-01

    The excretory-secretory (ES) antigens from Trichinella spiralis muscle larvae (ML) are the most commonly used diagnostic antigens for trichinellosis, but anti-Trichinella IgG antibodies cannot be detected until 2-3 weeks after infection; there is an obvious window period between Trichinella infection and antibody positivity. Intestinal infective larvae (IIL) are the first invasive stage during Trichinella infection, and their ES antigens are firstly exposed to the immune system and might be the early diagnostic markers of trichinellosis. The aim of this study was to evaluate the early diagnostic values of IIL ES antigens for trichinellosis. The IIL were collected from intestines of infected mice at 6 h postinfection (hpi), and IIL ES antigens were prepared by incubation for 18 h. Anti-Trichinella IgG antibodies in mice infected with 100 ML were detectable by ELISA with IIL ES antigens as soon as 10 days postinfection (dpi), but ELISA with ML ES antigens did not permit detection of infected mice before 12 dpi. When the sera of patients with trichinellosis at 19 dpi were assayed, the sensitivity (100 %) of ELISA with IIL ES antigens was evidently higher than 75 % of ELISA with ML ES antigens (P < 0.05) The specificity (96.86 %) of ELISA with IIL ES antigens was also higher than 89.31 % of ELISA with ML ES antigens (P < 0.05). The IIL ES antigens provided a new source of diagnostic antigens and could be considered as a potential early diagnostic antigen for trichinellosis. PMID:26342828

  10. Identification and characterization of an immunogenic antigen, enolase 2, among excretory/secretory antigens (ESA) of Toxoplasma gondii.

    Science.gov (United States)

    Jiang, Wei; Xue, Jun-Xin; Liu, Ying-Chun; Li, Tao; Han, Xian-Gan; Wang, Shao-Hui; Chen, Yong-Jun; Qi, Jingjing; Yu, Sheng-Qing; Wang, Quan

    2016-11-01

    An immunogenic protein, enolase 2, was identified among the secreted excretory/secretory antigens (ESAs) from Toxoplasma gondii strain RH using immunoproteomics based on matrix-assisted laser desorption/ionization time-of-flight mass spectrometry. Enolase 2 was cloned, sequenced, and heterologously expressed. BLAST analysis revealed 75-96% similarity with enolases from other parasites. Immunoblotting demonstrated good immunoreactivity of recombinant T. gondii enolase (Tg-enolase 2) to T. gondii-infected animal serum. Purified Tg-enolase 2 was found to catalyze dehydration of 2-phospho-d-glycerate to phosphoenolpyruvate. In vitro studies revealed maximal activity at pH 7.5 and 37 °C, and activity was inhibited by K(+), Ni(2+), Al(3+), Na(+), Cu(2+) and Cr(3+). A monoclonal antibody against Tg-enolase 2 was prepared, 1D6, with the isotype IgG2a/κ. Western blotting revealed that 1D6 reacts with Tg-enolase 2 and native enolase 2, present among T. gondii ESAs. The indirect immunofluorescence assays showed that enolase 2 could be specifically detected on the growing T. gondii tachyzoites. Immunoelectron microscopy revealed the surface and intracellular locations of enolase 2 on T. gondii cells. In conclusion, our results clearly show that the enzymatic activity of T. gondii enolase 2 is ion dependent and that it could be influenced by environmental factors. We also provide evidence that enolase 2 is an important immunogenic protein of ESAs from T. gondii and that it is a surface-exposed protein with strong antigenicity and immunogenicity. Our findings indicate that enolase 2 could play important roles in metabolism, immunogenicity and pathogenicity and that it may serve as a novel drug target and candidate vaccine against T. gondii infection. PMID:27450536

  11. Architecture of the DNA polymerase B-proliferating cell nuclear antigen (PCNA)-DNA ternary complex

    OpenAIRE

    Mayanagi, Kouta; Kiyonari, Shinichi; Nishida, Hirokazu; Saito, Mihoko; Kohda, Daisuke; Ishino, Yoshizumi; Shirai, Tsuyoshi; Morikawa, Kosuke

    2011-01-01

    DNA replication in archaea and eukaryotes is executed by family B DNA polymerases, which exhibit full activity when complexed with the DNA clamp, proliferating cell nuclear antigen (PCNA). This replication enzyme consists of the polymerase and exonuclease moieties responsible for DNA synthesis and editing (proofreading), respectively. Because of the editing activity, this enzyme ensures the high fidelity of DNA replication. However, it remains unclear how the PCNA-complexed enzyme temporally ...

  12. Abnormal antigens in breast cancer tissues and production of monoclonal antibodies against one of these antigens

    International Nuclear Information System (INIS)

    Breast cancer is associated with up regulation, down regulation of normal antigens or abnormal antigens. These antigens are very useful candidates as targets for the different breast cancer therapies and for vaccination trials. This study was done to characterize abnormal antigens, extract one of them and to produce monoclonal antibodies against the extracted antigen. One hundred and twenty Sudanese female patients were included in this study after informed consent. The mean age was 47. 2 years (16-80). Two tissue samples were obtained from each patient and they were confirmed as normal and cancerous breast tissues microscopically. 2D PAGE was used to analyze the protein content of samples. LC/MS and nr. fast a database search were used for separation and indentification of the abnormal proteins. Three different patterns of 2D Page results were obtained, the first pattern involved detection of four abnormal proteins in 26.7% of the patient cancerous tissues while they were undetected in the normal tissues of the same patients. In the second 2D PAGE result pattern the cancerous and the normal tissues of 67.5% patients were identical and they did not contain the four abnormal proteins while the third 2D PAGE pattern involved the presence of two abnormal antigens (from the four) in the cancerous tissues of 5.8% of the patients and they were absent from the normal tissues of the same patients. The four abnormal proteins were identified as, human Thioredoxin (D60nmutant), x-ray crystal structure of human galectin-1, retrocopy of tropomyosin 3(rc TPM3) and beta-tropomyosin (isoform 2). The primary and the secondary structures were obtained from the SWISSPROT and the PDB databases. Beta tropomyosin spot was extracted and used as antigen for monoclonal antibody production. Monoclonal antibody against beta- tropomyosin with a concentration of 0.35 mg/ml and a G11 anti beta-tropomyosin hybridoma cell line were produced. The monoclonal antibody was with single bad and

  13. The T-cell anergy induced by Leishmania amazonensis antigens is related with defective antigen presentation and apoptosis

    Directory of Open Access Journals (Sweden)

    Roberta O. Pinheiro

    2004-09-01

    Full Text Available Leishmania amazonensis is the main agent of diffuse cutaneous leishmaniasis, a disease associated with anergic immune responses. In this study we show that the crude antigen of Leishmania amazonensis (LaAg but not L. braziliensis promastigotes (LbAg contains substances that suppress mitogenic and spontaneous proliferative responses of T cells. The suppressive substances in LaAg are thermoresistant (100ºC/1h and partially dependent on protease activity. T cell anergy was not due to a decreased production of growth factors as it was not reverted by addition of exogenous IL-2, IL-4, IFN-gamma or IL-12. LaAg did not inhibit anti-CD3-induced T cell activation, suggesting that anergy was due to a defect in antigen presentation. It was also not due to cell necrosis, but was accompanied by expressive DNA fragmentation in lymph node cells, indicative of apoptosis. Although pre-incubation of macrophages with LaAg prevented their capacity to present antigens, this effect was not due to apoptosis of the former. These results suggest that the T cell anergy found in diffuse leishmaniasis may be the result of parasite antigen-driven apoptosis of those cells following defective antigen presentation.A Leishmania amazonensis é o principal agente etiológico da leishmaniose cutânea difusa, uma doença associada a respostas imunes anérgicas. Neste estudo nós mostramos que o extrato bruto de promastigotas de Leishmania amazonensis (LaAg, mas não de L. braziliensis (LbAg, contém substâncias que suprimem respostas proliferativas, espontâneas e mitogênicas, de células T. As substâncias supressoras no LaAg são termo-resistentes (100°C/1h e parcialmente dependentes da atividade de proteases. A anergia de células T não foi devida à diminuição na produção de fatores de crescimento, uma vez que não foi revertida pela adição de: IL-2, IL-4, IFN-gama ou IL-12. O LaAg não inibiu a ativação de células T induzida por anti-CD3, sugerindo que a anergia

  14. Raman spectroscopy of HIV-1 antigen and antibody

    Science.gov (United States)

    Zinin, Pavel V.; Hu, Ningjie; Kamemoto, Lori E.; Yu, Qigui; Misra, Anupam K.; Sharma, Shiv K.

    2011-05-01

    Raman spectra of anti-HIV-1 antibody, HIV-1 antigen (p24), and HIV-1 antibody-antigen complex have been measured in near-infrared and UV regions: 785 nm; 830 nm; and 244 nm laser excitations. The spectrum of the HIV-1 antigen was excited with an infrared laser and contains numerous Raman peaks. The most prominent peaks are broad bands at 1343, 1449, 1609 and 1655 cm-1, which are characteristic of the Raman spectra of biological cells. The UV Raman spectrum of the HIV-1 antigen has a completely different structure. It has two strong peaks at 1613 cm-1 and 1173 cm-1. The peak at 1613 cm-1 is associated with vibrations of the aromatic amino acids tyrosine (Tyr) and tryptophan (Try). The second strongest peak at 1173 cm-1 is associated with the vibration of Tyr. The Raman peak pattern of the HIV-1 antigen-antibody complex is very similar to that of the HIV-1 antigen. The only difference is that the peak at 1007 cm-1 of the Raman spectrum of the HIV-1 antigen-antibody complex is slightly enhanced compared to that of the HIV-1 antigen. This indicates that the peaks of the HIV-1 antigen dominate the Raman spectrum of the HIV-1 antigen-antibody complex.

  15. Protamine-based nanoparticles as new antigen delivery systems.

    Science.gov (United States)

    González-Aramundiz, José Vicente; Peleteiro Olmedo, Mercedes; González-Fernández, África; Alonso Fernández, María José; Csaba, Noemi Stefánia

    2015-11-01

    The use of biodegradable nanoparticles as antigen delivery vehicles is an attractive approach to overcome the problems associated with the use of Alum-based classical adjuvants. Herein we report, the design and development of protamine-based nanoparticles as novel antigen delivery systems, using recombinant hepatitis B surface antigen as a model viral antigen. The nanoparticles, composed of protamine and a polysaccharide (hyaluronic acid or alginate), were obtained using a mild ionic cross-linking technique. The size and surface charge of the nanoparticles could be modulated by adjusting the ratio of the components. Prototypes with optimal physicochemical characteristics and satisfactory colloidal stability were selected for the assessment of their antigen loading capacity, antigen stability during storage and in vitro and in vivo proof-of-concept studies. In vitro studies showed that antigen-loaded nanoparticles induced the secretion of cytokines by macrophages more efficiently than the antigen in solution, thus indicating a potential adjuvant effect of the nanoparticles. Finally, in vivo studies showed the capacity of these systems to trigger efficient immune responses against the hepatitis B antigen following intramuscular administration, suggesting the potential interest of protamine-polysaccharide nanoparticles as antigen delivery systems.

  16. Specific Lipids Modulate the Transporter Associated with Antigen Processing (TAP)

    DEFF Research Database (Denmark)

    Scholz, C.; Parcej, D.; Ejsing, C. S.;

    2011-01-01

    The transporter associated with antigen processing (TAP) plays a key role in adaptive immunity by translocating proteasomal degradation products from the cytosol into the endoplasmic reticulum lumen for subsequent loading onto major histocompatibility (MHC) class I molecules. For functional...... and structural analysis of this ATP-binding cassette complex, we established the overexpression of TAP in the methylotrophic yeast Pichia pastoris. Screening of optimal solubilization and purification conditions allowed the isolation of the heterodimeric transport complex, yielding 30 mg of TAP/liter of culture....... Detailed analysis of TAP function in the membrane, solubilized, purified, and reconstituted states revealed a direct influence of the native lipid environment on activity. TAP-associated phospholipids, essential for function, were profiled by liquid chromatography Fourier transform mass spectrometry...

  17. Quantitating protein synthesis, degradation, and endogenous antigen processing.

    Science.gov (United States)

    Princiotta, Michael F; Finzi, Diana; Qian, Shu-Bing; Gibbs, James; Schuchmann, Sebastian; Buttgereit, Frank; Bennink, Jack R; Yewdell, Jonathan W

    2003-03-01

    Using L929 cells, we quantitated the macroeconomics of protein synthesis and degradation and the microeconomics of producing MHC class I associated peptides from viral translation products. To maintain a content of 2.6 x 10(9) proteins, each cell's 6 x 10(6) ribosomes produce 4 x 10(6) proteins min(-1). Each of the cell's 8 x 10(5) proteasomes degrades 2.5 substrates min(-1), creating one MHC class I-peptide complex for each 500-3000 viral translation products degraded. The efficiency of complex formation is similar in dendritic cells and macrophages, which play a critical role in activating T cells in vivo. Proteasomes create antigenic peptides at different efficiencies from two distinct substrate pools: rapidly degraded newly synthesized proteins that clearly represent defective ribosomal products (DRiPs) and a less rapidly degraded pool in which DRiPs may also predominate. PMID:12648452

  18. A role for NADPH oxidase in antigen presentation

    Directory of Open Access Journals (Sweden)

    Gail J Gardiner

    2013-09-01

    Full Text Available The nicotinamide adenine dinucleotide phosphate (NADPH oxidase expressed in phagocytes is a multi-subunit enzyme complex that generates superoxide (O2.-. This radical is an important precursor of hydrogen peroxide (H2O2 and other reactive oxygen species (ROS needed for microbicidal activity during innate immune responses. Inherited defects in NADPH oxidase give rise to chronic granulomatous disease (CGD, a primary immunodeficiency characterized by recurrent infections and granulomatous inflammation. Interestingly, CGD, CGD carrier status, and oxidase gene polymorphisms have all been associated with autoinflammatory and autoimmune disorders, suggesting a potential role for NADPH oxidase in regulating adaptive immune responses. Here, NADPH oxidase function in antigen processing and presentation is reviewed. NADPH oxidase influences dendritic cell (DC crosspresentation by major histocompatibility complex class I molecules (MHC-I through regulation of the phagosomal microenvironment, while in B lymphocytes, NADPH oxidase alters epitope selection by major histocompatibility complex class II molecules (MHC-II.

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

    NARCIS (Netherlands)

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

    2014-01-01

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

  20. Antigenic characterization of Brazilian bovine viral diarrhea virus isolates by monoclonal antibodies and cross-neutralization

    Directory of Open Access Journals (Sweden)

    Botton S.A.

    1998-01-01

    Full Text Available Nineteen Brazilian isolates of bovine viral diarrhea virus (BVDV were characterized antigenically with a panel of 19 monoclonal antibodies (mAbs (Corapi WV, Donis RO and Dubovi EJ (1990 American Journal of Veterinary Research, 55: 1388-1394. Eight isolates were further characterized by cross-neutralization using sheep monospecific antisera. Analysis of mAb binding to viral antigens by indirect immunofluorescence revealed distinct patterns of reactivity among the native viruses. Local isolates differed from the prototype Singer strain in recognition by up to 14 mAbs. Only two mAbs - one to the non-structural protein NS23/p125 and another to the envelope glycoprotein E0/gp48 - recognized 100% of the isolates. No isolate was recognized by more than 14 mAbs and twelve viruses reacted with 10 or less mAbs. mAbs to the major envelope glycoprotein E2/gp53 revealed a particularly high degree of antigenic variability in this glycoprotein. Nine isolates (47.3% reacted with three or less of 10 E2/gp53 mAbs, and one isolate was not recognized by any of these mAbs. Virus-specific antisera to eight isolates plus three standard BVDV strains raised in lambs had virus-neutralizing titers ranging from 400 to 3200 against the homologous virus. Nonetheless, many antisera showed significantly reduced neutralizing activity when tested against heterologous viruses. Up to 128-fold differences in cross-neutralization titers were observed for some pairs of viruses. When the coefficient of antigenic similarity (R was calculated, 49 of 66 comparisons (74.24% between viruses resulted in R values that antigenically distinguish strains. Moreover, one isolate had R values suggesting that it belongs to a distinct serologic group. The marked antigenic diversity observed among Brazilian BVDV isolates should be considered when planning diagnostic and immunization strategies.

  1. Viral Escape Mutant Epitope Maintains TCR Affinity for Antigen yet Curtails CD8 T Cell Responses.

    Directory of Open Access Journals (Sweden)

    Shayla K Shorter

    Full Text Available T cells have the remarkable ability to recognize antigen with great specificity and in turn mount an appropriate and robust immune response. Critical to this process is the initial T cell antigen recognition and subsequent signal transduction events. This antigen recognition can be modulated at the site of TCR interaction with peptide:major histocompatibility (pMHC or peptide interaction with the MHC molecule. Both events could have a range of effects on T cell fate. Though responses to antigens that bind sub-optimally to TCR, known as altered peptide ligands (APL, have been studied extensively, the impact of disrupting antigen binding to MHC has been highlighted to a lesser extent and is usually considered to result in complete loss of epitope recognition. Here we present a model of viral evasion from CD8 T cell immuno-surveillance by a lymphocytic choriomeningitis virus (LCMV escape mutant with an epitope for which TCR affinity for pMHC remains high but where the antigenic peptide binds sub optimally to MHC. Despite high TCR affinity for variant epitope, levels of interferon regulatory factor-4 (IRF4 are not sustained in response to the variant indicating differences in perceived TCR signal strength. The CD8+ T cell response to the variant epitope is characterized by early proliferation and up-regulation of activation markers. Interestingly, this response is not maintained and is characterized by a lack in IL-2 and IFNγ production, increased apoptosis and an abrogated glycolytic response. We show that disrupting the stability of peptide in MHC can effectively disrupt TCR signal strength despite unchanged affinity for TCR and can significantly impact the CD8+ T cell response to a viral escape mutant.

  2. Chitosan based nanoparticles as protein carriers for efficient oral antigen delivery.

    Science.gov (United States)

    Gao, Ping; Xia, Guixue; Bao, Zixian; Feng, Chao; Cheng, Xiaojie; Kong, Ming; Liu, Ya; Chen, Xiguang

    2016-10-01

    This study aimed to investigate the efficacy of nanoparticles based on chitosan as a vehicle for oral antigen delivery in fish vaccination. Carboxymethyl chitosan/chitosan nanoparticles (CMCS/CS-NPs) loaded extracellular products (ECPs) of Vibrio anguillarum were successfully developed by ionic gelation method. The prepared ECPs-loaded CMCS/CS-NPs were characterized for various parameters including morphology, particle size (312±7.18nm), zeta potential (+17.4±0.38mV), loading efficiency (57.8±2.54%) and stability under the simulated gastrointestinal (GI) tract conditions in turbot. The in vitro profile showed that the cumulative release of ECPs from nanoparticles was higher in pH 7.4 (58%) than in pH 2.0 (37%) and pH 4.5 (29%) after 48h. Fluorescein isothiocyanate-labeled bovine serum albumin (FITC-BSA) was used as model protein antigen and encapsulated in CMCS/CS-NPs for investigating the biodistribution of antigen after oral delivery to turbot in 24h. Oral immunization of ECPs-loaded CMCS/CS-NPs group in turbot showed elevated specific antibody and higher concentrations of lysozyme activity and complement activity in fish serum than ECPs solution. CMCS/CS-NPs loaded with ECPs could enhance both adaptive and innate immune responses than the group treated with ECPs solution and suggested to be a potential antigen delivery system. PMID:27287772

  3. Precise conditional immortalization of mouse cells using tetracycline-regulated SV40 large T-antigen.

    Science.gov (United States)

    Anastassiadis, Konstantinos; Rostovskaya, Maria; Lubitz, Sandra; Weidlich, Stefanie; Stewart, A Francis

    2010-04-01

    Cellular immortalization provides a way for expansion and subsequent molecular characterization of rare cell types. Ideally, immortalization can be achieved by the reversible expression of immortalizing proteins. Here, we describe the use of conditional immortalization based on a modified tetracycline-regulated system for the expression of SV40 large T-antigen in embryonic stem (ES) cells and mice. The modified system relies on a codon improved reverse tetracycline transactivator (irtTA) fused to the ligand-binding domain (LBD) of the androgen receptor (irtTA-ABD) or of a mutated glucocorticoid receptor (irtTA-GBD*). Induction of T-antigen is conferred only after addition of two ligands, one to activate the LBD (mibolerone for irtTA-ABD or dexamethasone for irtTA-GBD*) and one to activate the tetracycline transactivator (doxycycline). In ES cells, changes in gene expression upon large T induction were limited and reversible upon deinduction. Similarly, expression of T-antigen was very tightly regulated in mice. We have isolated and expanded bone marrow mesenchymal stem cells that could be genetically manipulated and maintained their differentiation properties after several passages of expansion under conditions that induce the expression of large T-antigen. PMID:20146354

  4. Identification and characterization of Ixodes scapularis antigens that elicit tick immunity using yeast surface display.

    Science.gov (United States)

    Schuijt, Tim J; Narasimhan, Sukanya; Daffre, Sirlei; DePonte, Kathleen; Hovius, Joppe W R; Van't Veer, Cornelis; van der Poll, Tom; Bakhtiari, Kamran; Meijers, Joost C M; Boder, Eric T; van Dam, Alje P; Fikrig, Erol

    2011-01-05

    Repeated exposure of rabbits and other animals to ticks results in acquired resistance or immunity to subsequent tick bites and is partially elicited by antibodies directed against tick antigens. In this study we demonstrate the utility of a yeast surface display approach to identify tick salivary antigens that react with tick-immune serum. We constructed an Ixodes scapularis nymphal salivary gland yeast surface display library and screened the library with nymph-immune rabbit sera and identified five salivary antigens. Four of these proteins, designated P8, P19, P23 and P32, had a predicted signal sequence. We generated recombinant (r) P8, P19 and P23 in a Drosophila expression system for functional and immunization studies. rP8 showed anti-complement activity and rP23 demonstrated anti-coagulant activity. Ixodes scapularis feeding was significantly impaired when nymphs were fed on rabbits immunized with a cocktail of rP8, rP19 and rP23, a hall mark of tick-immunity. These studies also suggest that these antigens may serve as potential vaccine candidates to thwart tick feeding.

  5. Identification and characterization of Ixodes scapularis antigens that elicit tick immunity using yeast surface display.

    Directory of Open Access Journals (Sweden)

    Tim J Schuijt

    Full Text Available Repeated exposure of rabbits and other animals to ticks results in acquired resistance or immunity to subsequent tick bites and is partially elicited by antibodies directed against tick antigens. In this study we demonstrate the utility of a yeast surface display approach to identify tick salivary antigens that react with tick-immune serum. We constructed an Ixodes scapularis nymphal salivary gland yeast surface display library and screened the library with nymph-immune rabbit sera and identified five salivary antigens. Four of these proteins, designated P8, P19, P23 and P32, had a predicted signal sequence. We generated recombinant (r P8, P19 and P23 in a Drosophila expression system for functional and immunization studies. rP8 showed anti-complement activity and rP23 demonstrated anti-coagulant activity. Ixodes scapularis feeding was significantly impaired when nymphs were fed on rabbits immunized with a cocktail of rP8, rP19 and rP23, a hall mark of tick-immunity. These studies also suggest that these antigens may serve as potential vaccine candidates to thwart tick feeding.

  6. Antigen-induced and non-antigen-induced histamine release from rat mast cells sensitized with mouse antiserum.

    Directory of Open Access Journals (Sweden)

    Kurose,Masao

    1981-10-01

    Full Text Available Marked IgE-mediated histamine release from rat mast cells sensitized in vitro with mouse antiserum occurs in the presence of added Ca++ and phosphatidylserine (PS, although a considerable degree of antigen-induced histamine release which may utilize intracellular or cell-bound calcium is also observed. The decay in the responsiveness to Ca++ of the sensitized cells stimulated by antigen in Ca++-free medium in the presence of PS is relatively slow, and maximum release is produced by Ca++ added 1 min after antigen. Histamine release also occurs when Ca++ is added after PS in the absence of antigen to the sensitized cells suspended in Ca++-free medium. Unlike the antigen-induced release, the intensity of this non-antigen-induced release varies depending on both mast-cell and antiserum pools. A heat-labile factor(s, which is different from antigen-specific IgE antibody and is also contained in normal mouse serum, is involved in this reaction. In the antigen-nondependent (PS + Ca++-induced release, no decay in the responsiveness to Ca++ is observed after PS addition. Both the antigen-induced and non-antigen-induced release are completed fairly rapidly and are dependent of temperature, pH and energy.

  7. 猪骨髓基质抗原2基因的克隆、表达及生物学活性研究%Molecular Cloning and Expression of Bone Marrow Stromal Antigen-2 and Detection of Its Biological Activity

    Institute of Scientific and Technical Information of China (English)

    张廷虹; 赵协; 曹广明; 张振杰; 常维山

    2012-01-01

    研究目的是克隆猪骨髓基质抗原2 (Bonc marrow stromal antigen 2,BST- 2)基因并进行真核表达,获得具有抗病毒活性的重组BST-2蛋白.研究通过RT-PCR从猪瘟弱毒疫苗和聚肌胞(Poly Ⅰ:C)刺激的猪肾细胞(PK-15)中扩增出猪BST-2 cDNA,克隆至真核表达载体pcDNA3.1/V5-His,构建重组表达质粒pcDNA-BST-2,转染HEK293T细胞,纯化BST-2蛋白并经Bradford法定量,Western blot检测,研究BST-2抗病毒生物学活性.酶切鉴定和核酸序列测定证实pcDNA-BST-2真核表达质粒构建成功,转染HEK293T细胞后,经间接免疫荧光能够检测到绿色荧光.生物学活性测定重组BST-2蛋白具有一定的抗水泡性口炎病毒(Vesicular stomatitis virus,VSV)、H9禽流感病毒(Avian influenza virus,AIV)及猪繁殖与呼吸综合症病毒(Porcine reproductive and respiratory syndrome virus,PRRSV)的活性.结果表明,重组BST-2具有一定抗病毒生物功能,为进一步研究重组BST 2蛋白的活性以及BST-2抗病毒药物研究奠定了基础.%To clone porcine bone marrow stromal antigen-2 (BST-2) gene, construct its recombinant eu-karyotic expression plasmid and induce the expression of the fusion antiviral protein, we amplified BST-2 gene by RT-PCR from the total RNA extracted from PK15 cells. The recombinant expression plasmid pcDNA-BST-2 was constructed and then was transfected into HEK293T cells to expresse the BST-2 fusion protein. Western blot and indirect immunofluorescence assay ( IFA) were performed, and the biological activity was detected. The results showed that the construction of recombinant plasmid pcDNA-BST-2 was confirmed by restriction enzyme digestion and sequencing. The expressed product had antiviral activity a-gainst Vesicular stomatitis virus (VSV), Avian influenza virus (AIV) and Porcine reproductive and respiratory syndrome virus (PRRSV). In conclusion, the research paves the way for further research on bioac-tivity assayand antiviral medication.

  8. Structure of ganglioside with CAD blood group antigen activity

    Energy Technology Data Exchange (ETDEWEB)

    Gillard, B.K.; Blanchard, D.; Cartron, J.P.; van Kuik, G.A.; Vliegenthart, J.F.G.; Marcus, D.M.

    1986-05-01

    The novel erythrocyte ganglioside which carries the blood group Cad determinant has been isolated, and its structure has been determined. The ganglioside contained Glu:Gal:GalNAc:GlcNAc in a molar ratio of 1.00:1.94:0.93:0.95. The ganglioside binds Helix pomatia lectin and its chromatographic mobility is similar to G/sub D3/. After treatment with ..beta..-hexosaminidase (human placenta HexA) the product migrated with sialosylparagloboside (SPG), no longer binds Helix lectin, and binds a human anti-SPG antibody. Treatment of this material with neuraminidase (V. cholera) yielded a product with the mobility of paragloboside that bound monoclonal antibody 1B2. NMR analysis revealed that the terminal GalNAc is linked ..beta..1-4 to Gal, and confirms the structure proposed previously: GalNAc..beta..1-4(NeuAc..cap alpha..2-3)Gal..beta..1-4GlcNAc..beta..1-3Gal..beta..1-4Glc-Cer. This structure is consistent with the previous demonstration that a compound with the same chromatographic mobility as the Cad ganglioside could be synthesized by enzymatic transfer of GalNAc to sialosylparagloboside.

  9. Simple mucin-type carbohydrate antigens in major salivary glands

    DEFF Research Database (Denmark)

    Therkildsen, M H; Mandel, U; Thorn, J;

    1994-01-01

    , on the other hand, expressed A, H, and inconstantly sialosyl-T, Tn, and sialosyl-Tn antigens in major salivary glands, whereas serous cells of minor (labial) salivary glands expressed H exclusively, Tn and sialosyl-T antigens inconstantly, but never sialosyl-Tn and A antigens. The difference may be related...... to a more simple cytodifferentiation of serous cells of minor (labial) salivary glands as compared with major salivary glands. Duct cells in major salivary glands expressed A, H, and inconstantly T, sialosyl-T, and Tn antigens, whereas minor (labial) salivary glands ducts exclusively expressed H, T...... and sialosyl-T antigens, differences that may be related to dissimilarities in the duct system. Myoepithelial cells and basal cells exclusively expressed T and sialosyl-T antigens, which may prove useful in studies of salivary gland tumors, since these cells are known to play a key role in the histological...

  10. Tissue distribution of histo-blood group antigens

    DEFF Research Database (Denmark)

    Ravn, V; Dabelsteen, Erik

    2000-01-01

    The introduction of immunohistochemical techniques and monoclonal antibodies to specific carbohydrate epitopes has made it possible to study in detail the tissue distribution of histo-blood group antigens and related carbohydrate structures. The present paper summarizes the available data...... concerning the histological distribution of histo-blood group antigens and their precursor structures in normal human tissues. Studies performed have concentrated on carbohydrate antigens related to the ABO, Lewis, and TTn blood group systems, i.e. histo-blood group antigens carried by type 1, 2, and 3 chain...... carrier carbohydrate chains. Histo-blood group antigens are found in most epithelial tissues. Meanwhile, several factors influence the type, the amount, and the histological distribution of histoblood group antigens, i.e. the ABO, Lewis, and saliva-secretor type of the individual, and the cell- and tissue...

  11. Monoclonal antibody-defined human pancreatic cancer-associated antigens.

    Science.gov (United States)

    Schmiegel, W H; Kalthoff, H; Arndt, R; Gieseking, J; Greten, H; Klöppel, G; Kreiker, C; Ladak, A; Lampe, V; Ulrich, S

    1985-03-01

    Three pancreatic cancer-associated antigens were characterized by use of monoclonal antibodies in immunobinding studies with various cellular and soluble target antigens, in immunoprecipitation, and in immunoperoxidase staining. C54-0 represents a tumor-associated Mr 122,000 antigen, which appears to be widely distributed on various epithelial tumors and to a lower extent on normal tissue. C1-N3 antigen exhibited a more restricted distribution, reacting with pancreatic and various gastrointestinal tract tumors as well as with chronically inflamed pancreatic tissue. The most specific antigen expression was observed for C1-P83 antigen, found on all exocrine tumors of the pancreas, but not on normal or chronically inflamed pancreatic tissue.

  12. Formaldehyde scavengers function as novel antigen retrieval agents

    OpenAIRE

    Craig T. Vollert; Moree, Wilna J; Steven Gregory; Bark, Steven J.; Eriksen, Jason L.

    2015-01-01

    Antigen retrieval agents improve the detection of formaldehyde-fixed proteins, but how they work is not well understood. We demonstrate that formaldehyde scavenging represents a key characteristic associated with effective antigen retrieval; under controlled temperature and pH conditions, scavenging improves the typical antigen retrieval process through reversal of formaldehyde-protein adduct formation. This approach provides a rational framework for the identification and development of more...

  13. T-cell recognition of a cross-reactive antigen(s) in erythrocyte stages of Plasmodium falciparum and Plasmodium yoelii: inhibition of parasitemia by this antigen(s).

    OpenAIRE

    Lucas, B.; Engels, A; Camus, D; Haque, A.

    1993-01-01

    In the current study, we investigated the presence of a cross-reactive antigen(s) in the erythrocyte stage from Plasmodium yoelii (265 BY strain) and Plasmodium falciparum through recognition by T cells primed in vivo with antigens from each of these parasites. BALB/c mice are naturally resistant to P. falciparum but are susceptible to P. yoelii infection. Mice that had recovered from P. yoelii primary infection became resistant to a second infection. A higher in vitro proliferative response ...

  14. Characterization of Ewing sarcoma associated cancer/testis antigens

    OpenAIRE

    Mahlendorf, Dorothea E.; Staege, Martin Sebastian

    2013-01-01

    The prognosis of patients suffering from tumors of the Ewing family (EFT) is still poor. Immunotherapy strategies are pursued and EFT-specific antigens have to be identified as targets for cytotoxic T-lymphocytes (CTL). Due to the lack of expression of cancer/testis antigens (CTA) in normal tissues, these antigens are partially able to induce immune responses in cancer patients. Therefore, they are promising targets for immunotherapy. EFT are characterized by chromosomal rearrangements involv...

  15. Pneumocystis carinii antigen detection in rat serum and lung lavage.

    OpenAIRE

    McNabb, S J; Graves, D C; Kosanke, S.D.; Moyer, M J; Ivey, M H

    1988-01-01

    We developed a modified double-antibody sandwich enzyme-linked immunosorbent assay (ELISA) that detected relatively low concentrations of known Pneumocystis carinii antigen added to buffer or rat sera. Artificial immunization-derived polyclonal rabbit anti-P. carinii antibody was used on the solid phase to capture the antigen. Infection-derived (after P. carinii pneumonia) polyclonal rat anti-P. carinii antibody or a mixture of five murine monoclonal antibodies was used as the antigen detecto...

  16. Identification, characterization and immunogenicity of an O-antigen capsular polysaccharide of Francisella tularensis.

    Directory of Open Access Journals (Sweden)

    Michael A Apicella

    Full Text Available Capsular polysaccharides are important factors in bacterial pathogenesis and have been the target of a number of successful vaccines. Francisella tularensis has been considered to express a capsular antigen but none has been isolated or characterized. We have developed a monoclonal antibody, 11B7, which recognizes the capsular polysaccharide of F. tularensis migrating on Western blot as a diffuse band between 100 kDa and 250 kDa. The capsule stains poorly on SDS-PAGE with silver stain but can be visualized using ProQ Emerald glycoprotein stain. The capsule appears to be highly conserved among strains of F. tularensis as antibody 11B7 bound to the capsule of 14 of 14 F. tularensis type A and B strains on Western blot. The capsular material can be isolated essentially free of LPS, is phenol and proteinase K resistant, ethanol precipitable and does not dissociate in sodium dodecyl sulfate. Immunoelectron microscopy with colloidal gold demonstrates 11B7 circumferentially staining the surface of F. tularensis which is typical of a polysaccharide capsule. Mass spectrometry, compositional analysis and NMR indicate that the capsule is composed of a polymer of the tetrasaccharide repeat, 4-alpha-D-GalNAcAN-(1->4-alpha-D-GalNAcAN-(1->3-beta-D-QuiNAc-(1->2-beta-D-Qui4NFm-(1-, which is identical to the previously described F. tularensis O-antigen subunit. This indicates that the F. tularensis capsule can be classified as an O-antigen capsular polysaccharide. Our studies indicate that F. tularensis O-antigen glycosyltransferase mutants do not make a capsule. An F. tularensis acyltransferase and an O-antigen polymerase mutant had no evidence of an O-antigen but expressed a capsular antigen. Passive immunization of BALB/c mice with 75 microg of 11B7 protected against a 150 fold lethal challenge of F. tularensis LVS. Active immunization of BALB/c mice with 10 microg of capsule showed a similar level of protection. These studies demonstrate that F. tularensis

  17. Blood group antigen studies using CdTe quantum dots and flow cytometry

    Directory of Open Access Journals (Sweden)

    Cabral Filho PE

    2015-07-01

    Full Text Available Paulo E Cabral Filho,1 Maria IA Pereira,1 Heloise P Fernandes,2 Andre A de Thomaz,3 Carlos L Cesar,3 Beate S Santos,4 Maria L Barjas-Castro,2 Adriana Fontes1 1Departamento de Biofísica e Radiobiologia, Universidade Federal de Pernambuco, Recife, Pernambuco, 2Centro de Hematologia e Hemoterapia, Universidade Estadual de Campinas, Instituto Nacional de Ciência e Tecnologia do Sangue, Campinas, São Paulo, 3Departamento de Eletrônica Quântica, Instituto de Física Gleb Wataghin, Universidade Estadual de Campinas, Campinas, São Paulo, 4Departamento de Ciências Farmacêuticas, Universidade Federal de Pernambuco, Recife, PE, Brazil Abstract: New methods of analysis involving semiconductor nanocrystals (quantum dots [QDs] as fluorescent probes have been highlighted in life science. QDs present some advantages when compared to organic dyes, such as size-tunable emission spectra, broad absorption bands, and principally exceptional resistance to photobleaching. Methods applying QDs can be simple, not laborious, and can present high sensibility, allowing biomolecule identification and quantification with high specificity. In this context, the aim of this work was to apply dual-color CdTe QDs to quantify red blood cell (RBC antigen expression on cell surface by flow cytometric analysis. QDs were conjugated to anti-A or anti-B monoclonal antibodies, as well as to the anti-H (Ulex europaeus I lectin, to investigate RBCs of A1, B, A1B, O, A2, and Aweak donors. Bioconjugates were capable of distinguishing the different expressions of RBC antigens, both by labeling efficiency and by flow cytometry histogram profile. Furthermore, results showed that RBCs from Aweak donors present fewer amounts of A antigens and higher amounts of H, when compared to A1 RBCs. In the A group, the amount of A antigens decreased as A1 > A3 > AX = Ael, while H antigens were AX = Ael > A1. Bioconjugates presented stability and remained active for at least 6 months. In conclusion

  18. Failure of expression of class I major histocompatibility antigens to alter tumor immunogenicity of a spontaneous murine carcinoma.

    Science.gov (United States)

    Carlow, D A; Kerbel, R S; Elliott, B E

    1989-05-10

    We have shown previously that clonal immunogenic variants of murine mammary adenocarcinoma 10.1 can be isolated after treatment in vitro with the DNA-hypomethylating agent 5-azacytidine (5-aza). Such immunogenic variants frequently express elevated class I major histocompatibility complex antigens relative to the level of expression in the parent tumor and are rejected in syngeneic mice by a T-cell-dependent process. To ascertain whether elevated immunogenicity is a function of increased class I antigen expression, we isolated high class I antigen expressors from 5-aza-treated 10.1 cells by using the fluorescence-activated cell sorter. Clonal variants displaying any increase in class I antigen expression were more efficient stimulators of allo-class I antigen-specific cytolytic T-cell precursors. However, these variants displayed unaltered tumorigenicity in immunocompetent syngeneic mice. Thus, phenotypic changes other than, or in addition to, elevated class I antigen expression cause the reduced tumorigenicity of immunogenic clones of 10.1 cells isolated after 5-aza treatment.

  19. Lymphocyte proliferation to mycobacterial antigens is detectable across a spectrum of HIV-associated tuberculosis

    Directory of Open Access Journals (Sweden)

    Bakari Muhammad

    2009-02-01

    Full Text Available Abstract Background Identifying novel TB diagnostics is a major public health priority. We explored the diagnostic characteristics of antimycobacterial lymphocyte proliferation assays (LPA in HIV-infected subjects with latent or active TB. Methods HIV-infected subjects with bacille Calmette Guérin (BCG scars and CD4 counts ≥ 200 cells/mm3 entering a TB booster vaccine trial in Tanzania had baseline in vivo and in vitro immune tests performed: tuberculin skin tests (TST, LPA and five day assays of interferon gamma (IFN-γ release. Assay antigens were early secreted antigenic target 6 (ESAT-6, antigen 85 (Ag85, and Mycobacterium tuberculosis whole cell lysate (WCL. Subjects were screened for active TB at enrollment by history, exam, sputum smear and culture. We compared antimycobacterial immune responses between subjects with and without latent or active TB at enrollment. Results Among 1885 subjects screened, 635 had latent TB and 13 had active TB. Subjects with latent TB were more likely than subjects without TB to have LPA responses to ESAT-6 (13.2% vs. 5.5%, P Conclusion Lymphoproliferative responses to mycobacteria are detectable during HIV-associated active TB, and are less sensitive but more specific than TST. Trial registration ClinicalTrials.gov Identifier NCT00052195.

  20. Antigen epitope of Helicobacter pylorivacuolating cytotoxin A

    Institute of Scientific and Technical Information of China (English)

    Xiu-Li Liu; Shu-Qin Li; Chun-Jie Liu; Hao-Xia Tao; Zhao-Shan Zhang

    2004-01-01

    AIM: To construct and select antigen epitopes of vacuolating cytotoxin A (VacA) for nontoxic VacA vaccine against Helicobacter pylori (H pylori) infection.METHODS: Eleven VacA epitopes were predicted according to VacA antigenic bioinformatics. Three candidates of VacA epitope were constructed through different combined epitopes. The candidate was linked with E. coli heat-labile enterotoxin B (LTB) by a linker of 7 amino acids, and cloned into plasmid pQE-60 in which fusion LTB-VacA epitope was efficiently expressed. To test the antigencity of the candidate, 6 BALB/c mice were treated with the fusion LTB-VacA epitope through intraperitoneal injection. To explore the ability of inhibiting the toxicity of VacA, cantiserum against the candidate was used to counteract VacA that induced HeLa cells to produce cell vacuoles in vitro.RESULTS: Serum IgG against the candidate was induced in the BALB/c mice. In vitro, the three antisera against the candidate efficiently counteracted the toxicity of VacA, and decreased the number of cell vacuoles by 14.17%, 20.20%and 30.41% respectively.CONCLUSION: Two of the three candidates, LZ-VacA1and LZ-VacA2, can be used to further study the mechanism of vacuolating toxicity of VacA, and to construct nontoxic VacA vaccine against H pylori infection.

  1. Detection of peste des petits ruminants virus antigen using immunofiltration and antigen-competition ELISA methods.

    Science.gov (United States)

    Raj, G Dhinakar; Rajanathan, T M C; Kumar, C Senthil; Ramathilagam, G; Hiremath, Geetha; Shaila, M S

    2008-06-22

    Peste des petits ruminants (PPR) is one of the most economically important diseases affecting sheep and goats in India. An immunofiltration-based test has been developed using either mono-specific serum/monoclonal antibodies (mAb) prepared against a recombinant truncated nucleocapsid protein of rinderpest virus (RPV) cross-reactive with PPR virus. This method consists of coating ocular swab eluate from suspected animals onto a nitrocellulose membrane housed in a plastic module, which is allowed to react with suitable dilutions of a mAb or a mono-specific polyclonal antibody. The antigen-antibody complex formed on the membrane is then detected by protein A-colloidal gold conjugate, which forms a pink colour. In the immunofiltration test, concordant results were obtained using either PPRV mAb or mono-specific serum. Another test, an antigen-competition ELISA which relies on the competition between plate-coated recombinant truncated 'N' protein of RPV and the PPRV 'N' protein present in ocular swab eluates (sample) for binding to the mono-specific antibody against N protein of RPV (in liquid phase) was developed. The cut-off value for this test was established using reverse transcription polymerase chain reaction (RT-PCR) positive and negative oculo-nasal swab samples. Linear correlation between percent inhibition (PI) values in antigen-competition ELISA and virus infectivity titres was 0.992. Comparison of the immunofiltration test with the antigen-competition ELISA yielded a sensitivity of 80% and specificity of 100%. These two tests can serve as a screening (immunofiltration) and confirmatory (antigen-competition ELISA) test, respectively, in the diagnosis of PPR in sheep or goats. PMID:18182256

  2. Antigen-Specific versus Non-Antigen-Specific Immunoadsorption in ABO-Incompatible Renal Transplantation.

    Directory of Open Access Journals (Sweden)

    Gerold Thölking

    Full Text Available ABO-incompatible (ABOi renal transplantation (RTx from living donors is an established procedure to expand the donor pool for patients with end stage renal disease. Immunoadsorption (IA is a standard procedure for the removal of preformed antibodies against the allograft. In this study, antigen-specific and non-antigen-specific IA in ABOi RTx were compared.10 patients underwent antigen-specific IA (Glycosorb group and 13 patients non-antigen-specific IA (Immunosorba group. The effects of both procedures regarding antibody reduction, number of treatments, complications, costs, as well as the allograft function and patient survival were compared between both groups.Although the IgG levels were reduced equally by both procedures (p=0.82, the reduction of the IgM level was more effective in the Glycosorb group (p=0.0172. Patients in both groups required a median number of 6 IA before ABOi RTx. Allograft function at one year after AB0i RTx was similar in both groups (estimated glomerular filtration rate: 66 vs. 64 ml/min/1.73m² respectively, with a death-censored graft survival of 90.0% and 92.3% respectively. Complication rates did not differ between procedures. Due to the reuse of non-antigen-specific Immunosorba columns, costs were considerably lower in this group; however, the use of the Immunosorba-based IA was less time-efficient.Considering upcoming alternatives as simultaneous performance of dialysis and IA or a possible reuse of Glycosorb columns, this might become less relevant in the future.

  3. Regulation of T cell response to leishmania antigens by determinants of histocompatibility leukocyte class I and II molecules

    Directory of Open Access Journals (Sweden)

    Bacellar O.

    1998-01-01

    Full Text Available It has been shown that HLA class I molecules play a significant role in the regulation of the proliferation of T cells activated by mitogens and antigens. We evaluated the ability of mAb to a framework determinant of HLA class I molecules to regulate T cell proliferation and interferon gamma (IFN-g production against leishmania, PPD, C. albicans and tetanus toxoid antigens in patients with tegumentary leishmaniasis and healthy subjects. The anti-major histocompatibility complex (MHC mAb (W6/32 suppressed lymphocyte proliferation by 90% in cultures stimulated with aCD3, but the suppression was variable in cultures stimulated with leishmania antigen. This suppression ranged from 30-67% and was observed only in 5 of 11 patients. IFN-g production against leishmania antigen was also suppressed by anti-HLA class I mAb. In 3 patients IFN-g levels were suppressed by more than 60%, while in the other 2 cultures IFN-g levels were 36 and 10% lower than controls. The suppression by HLA class I mAb to the proliferative response in leishmaniasis patients and in healthy controls varied with the antigens and the patients or donors tested. To determine whether the suppression is directed at antigen presenting cells (APCs or at the responding T cells, experiments with antigen-primed non-adherent cells, separately incubated with W6/32, were performed. Suppression of proliferation was only observed when the W6/32 mAb was added in the presence of T cells. These data provide evidence that a mAb directed at HLA class I framework determinants can suppress proliferation and cytokine secretion in response to several antigens.

  4. Breed differences in the frequency of bovine lymphocyte antigens.

    Science.gov (United States)

    Stear, M J; Brown, S C; Dimmock, C K; Dufty, J H; Hetzel, D J; Mackie, J T; Nicholas, F W; Tierney, T J; Wetherall, J D

    1987-01-01

    Lymphocytes from 1,564 cattle of 18 breeds and cross-bred groups in Australia were tested for major histocompatibility system class 1 antigens. Gene frequencies were calculated for the Angus, Belmont Red, Brahman, Hereford and Holstein-Friesian breeds. There were substantial differences among these breeds in antigen and gene frequency. There were striking differences among all 18 breeds in the presence or absence of certain antigens. Two antigens, CA13 and CA36, were strongly associated in Hereford cattle but occurred independently of each other in the other breeds. PMID:3273412

  5. Identification of protective antigens for vaccination against systemic salmonellosis

    Directory of Open Access Journals (Sweden)

    Dirk eBumann

    2014-08-01

    Full Text Available There is an urgent medical need for improved vaccines with broad serovar coverage and high efficacy against systemic salmonellosis. Subunit vaccines offer excellent safety profiles but require identification of protective antigens, which remains a challenging task. Here, I review crucial properties of Salmonella antigens that might help to narrow down the number of potential candidates from more than 4000 proteins encoded in Salmonella genomes, to a more manageable number of 50-200 most promising antigens. I also discuss complementary approaches for antigen identification and potential limitations of current pre-clinical vaccine testing.

  6. Cancer-germline antigen vaccines and epigenetic enhancers

    DEFF Research Database (Denmark)

    Gjerstorff, Morten Frier; Burns, Jorge; Ditzel, Henrik Jorn

    2010-01-01

    can be achieved using epigenetic modifiers. AREAS COVERED IN THIS REVIEW: We provide an overview of the potential of CG antigens as targets for cancer immunotherapy, including advantages and disadvantages. We also discuss the current state of development of CG antigen vaccines, and the potential...... synergistic effect of combining CG antigen immunotherapeutic strategies with epigenetic modifiers. WHAT THE READER WILL GAIN: The reader will gain an overview of the past, present and future role of CG antigens in cancer immunotherapy. TAKE HOME MESSAGE: Chemoimmunotherapy using epigenetic drugs and CG...

  7. СAPSULAR ANTIGEN OF YERSINIA PESTIS

    Directory of Open Access Journals (Sweden)

    L. A. Kadnikova

    2015-01-01

    Full Text Available Plague is a zoonosis caused by gram-negative bacteria Yersinia pestis, which, as a rule, is transmitted to humans from septicemic rodents by the bites of infected fleas. This microbe killed more people than all of the wars in the human history. Y. pestis circulation in the natural plague foci is ensured by the whole number of pathogenicity factors with differing functional orientation. This review is devoted to one of them, Y. pestis capsular antigen (F1 or Caf1. The history of its discovery and studying of its genetic control, biosynthesis, isolation and purification, and physicochemical properties are reviewed. Its roles in plague pathogenesis and its application as a main component of plague vaccines are also discussed. Y. pestis capsule under light microscopy is visually amorphous, while high-resolution electron microscopy displays the structure formed from separate fimbria-like cords up to 200 nm long, diverging from the bacterial surface in different directions. At 37°C Y. pestis produce 800–1000 times more capsular antigen than at 28°C. Genes coding for 17.6-kD Caf1 protein, which contains 170 amino acids, are located in caf1 operon of pFra plasmid. Analysis of caf1 operon nucleotide sequence testified its close phylogenetic relationship with the gene clusters coding for pilus adhesins that were secreted with the help of chaperone/usher systems in enterobacteria including six additional adhesins in Y. pestis. Y. pestis multiplication within macrophages is the obligatory stage of plague pathogenesis, and the plague pathogen virulence correlates not with resistance to phagocyte ingesting but with bacterial ability to survive and multiply within phagolysosomes of phagocytes due to neutralization of antibacterial functions of eukaryotic cells. The capsule formed out of the Caf1 aggregates protects Y. pestis from ingestion by naïve host’s phagocytes and prevents from initiation of the alternative pathway of the complement system

  8. In vitro antigenic stimulation of peripheral blood and lymph node lymphocytes of sensitized guinea-pigs: the effect of a second administration of antigen in vivo

    Science.gov (United States)

    Housley, J.; Gell, P. G. H.

    1969-01-01

    Stimulation of DNA synthesis by a guinea-pig albumin—orthanilic acid conjugate (AO) and by tuberculin purified protein derivative (PPD) was obtained in in vitro cultures of peripheral blood and lymph node lymphocytes from guinea-pigs with delayed hypersensitivity to these antigens. Animals sensitized to both AO and PPD were given a further injection of 5 μg AO, intravenously, 8 hours before killing for in vitro studies. In these guinea-pigs, peripheral blood cultures, but not lymph node cultures, showed greater DNA synthesis in response to both AO and PPD than cultures from controls not given a further injection of AO. It is suggested that the further increase in DNA synthesis was due to non-specific lymphocyte `activation' following the interaction of antigen and specifically sensitized lymphocytes. PMID:5352364

  9. Antigen-Specific CD4+ T Cells Recognize Epitopes of Protective Antigen following Vaccination with an Anthrax Vaccine

    OpenAIRE

    Laughlin, Elsa M.; Miller, Joseph D.; James, Eddie; Fillos, Dimitri; Ibegbu, Chris C.; Mittler, Robert S.; Akondy, Rama; Kwok, William; Ahmed, Rafi; Nepom, Gerald,

    2007-01-01

    Detection of antigen-specific CD4+ T cells is facilitated by the use of fluorescently labeled soluble peptide-major histocompatibility complex (MHC) multimers which mirror the antigen specificity of T-cell receptor recognition. We have used soluble peptide-MHC class II tetramers containing peptides from the protective antigen (PA) of Bacillus anthracis to detect circulating T cells in peripheral blood of subjects vaccinated with an anthrax vaccine. PA-specific HLA class II-restricted T lympho...

  10. Cellular gene expression induced by parasite antigens and allergens in neonates from parasite-infected mothers.

    Science.gov (United States)

    Soboslay, Peter T; Orlikowsky, Thorsten; Huang, Xiangsheng; Gille, Christian; Spring, Bärbel; Kocherscheidt, Lars; Agossou, Abram; Banla, Meba; Bonin, Michael; Köhler, Carsten

    2016-05-01

    Prenatal exposure to parasite antigens or allergens will influence the profile and strength of postnatal immune responses, such contact may tolerize and increase susceptibility to future infections or sensitize to environmental allergens. Exposure in utero to parasite antigens will distinctly alter cellular gene expression in newborns. Gene microarrays were applied to study gene expression in umbilical cord blood cell (UCBC) from parasite-exposed (Para-POS) and non-exposed (Para-NEG) neonates. UCBC were activated with antigens of helminth (Onchocerca volvulus), amoeba (Entamoeba histolytica) or allergens of mite (Dermatophagoides farinae). When UCBC from Para-POS and Para-NEG newborns were exposed to helminth antigens or allergens consistent differences occurred in the expression of genes encoding for MHC class I and II alleles, signal transducers of activation and transcription (STATs), cytokines, chemokines, immunoglobulin heavy and light chains, and molecules associated with immune regulation (SOCS, TLR, TGF), inflammation (TNF, CCR) and apoptosis (CASP). Expression of genes associated with innate immune responses were enhanced in Para-NEG, while in Para-POS, the expression of MHC class II and STAT genes was reduced. Within functional gene networks for cellular growth, proliferation and immune responses, Para-NEG neonates presented with significantly higher expression values than Para-POS. In Para-NEG newborns, the gene cluster and pathway analyses suggested that gene expression profiles may predispose for the development of immunological, hematological and dermatological disorders upon postnatal helminth parasite infection or allergen exposure. Thus, prenatal parasite contact will sensitize without generating aberrant inflammatory immune responses, and increased pro-inflammatory but decreased regulatory gene expression profiles will be present in those neonates lacking prenatal parasite antigen encounter. PMID:27062712

  11. Facts on the fragmentation of antigens in presenting cells, on the association of antigen fragments with MHC molecules in cell-free systems, and speculation on the cell biology of antigen processing

    DEFF Research Database (Denmark)

    Werdelin, O; Mouritsen, S; Petersen, B L;

    1988-01-01

    The processing of a protein antigen is a multi-step event taking place in antigen-presenting cells. Processing is a prerequisite for the recognition of most antigens by T lymphocytes. The antigen is ingested by endocytosis, transported to an acid cellular compartment and subjected to proteolytic ...

  12. Thyrotropin Receptor Epitope and Human Leukocyte Antigen in Graves' Disease.

    Science.gov (United States)

    Inaba, Hidefumi; De Groot, Leslie J; Akamizu, Takashi

    2016-01-01

    Graves' disease (GD) is an organ-specific autoimmune disease, and thyrotropin (TSH) receptor (TSHR) is a major autoantigen in this condition. Since the extracellular domain of human TSHR (TSHR-ECD) is shed into the circulation, TSHR-ECD is a preferentially immunogenic portion of TSHR. Both genetic factors and environmental factors contribute to development of GD. Inheritance of human leukocyte antigen (HLA) genes, especially HLA-DR3, is associated with GD. TSHR-ECD protein is endocytosed into antigen-presenting cells (APCs), and processed to TSHR-ECD peptides. These peptide epitopes bind to HLA-class II molecules, and subsequently the complex of HLA-class II and TSHR-ECD epitope is presented to CD4+ T cells. The activated CD4+ T cells secrete cytokines/chemokines that stimulate B-cells to produce TSAb, and in turn hyperthyroidism occurs. Numerous studies have been done to identify T- and B-cell epitopes in TSHR-ECD, including (1) in silico, (2) in vitro, (3) in vivo, and (4) clinical experiments. Murine models of GD and HLA-transgenic mice have played a pivotal role in elucidating the immunological mechanisms. To date, linear or conformational epitopes of TSHR-ECD, as well as the molecular structure of the epitope-binding groove in HLA-DR, were reported to be related to the pathogenesis in GD. Dysfunction of central tolerance in the thymus, or in peripheral tolerance, such as regulatory T cells, could allow development of GD. Novel treatments using TSHR antagonists or mutated TSHR peptides have been reported to be effective. We review and update the role of immunogenic TSHR epitopes and HLA in GD, and offer perspectives on TSHR epitope specific treatments. PMID:27602020

  13. Antigenic characterization of avian influenza H9 subtype isolated from desi and zoo birds

    Directory of Open Access Journals (Sweden)

    Farrukh Saleem

    2011-08-01

    Full Text Available Avian influenza is a viral infection which affects mainly the respiratory system of birds. The H9N2 considered as low pathogenic avian influenza (LPAI virus and continuously circulating in poultry flocks causing enormous economic losses to poultry industry of Pakistan. As these viruses have RNA genome and their RNA polymerase enzyme lacks proof reading activity which resulted in spontaneous mutation in surface glycoproteins (HA and NA and reassortment of their genomic segments results in escape from host immune response produced by the vaccine. Efforts made for the isolation and identification of avian influenza virus from live desi and zoo birds of Lahore and performed antigenic characterization. The local vaccines although gives a little bit less titer when we raise the antisera against these vaccines but their antisera have more interaction with the local H9 subtype antigen so it gives better protective immune response. Infected chicken antisera are more reactive as compare to rabbit antisera. This shows that our isolates have highest similarity with the currently circulating viruses. These results guided us to devise a new control strategy against avian influenza viral infections. The antigenic characterization of these avian influenza isolates helped us to see the antigenic differences between the isolates of this study and H9 subtype avian influenza viruses used in vaccines. Therefore, this study clearly suggests that a new local H9 subtype avian influenza virus should be used as vaccinal candidate every year for the effective control of influenza viral infections of poultry.

  14. IgG Responses to Tissue-Associated Antigens as Biomarkers of Immunological Treatment Efficacy

    Directory of Open Access Journals (Sweden)

    Heath A. Smith

    2011-01-01

    Full Text Available We previously demonstrated that IgG responses to a panel of 126 prostate tissue-associated antigens are common in patients with prostate cancer. In the current report we questioned whether changes in IgG responses to this panel might be used as a measure of immune response, and potentially antigen spread, following prostate cancer-directed immune-active therapies. Sera were obtained from prostate cancer patients prior to and three months following treatment with androgen deprivation therapy (=34, a poxviral vaccine (=31, and a DNA vaccine (=21. Changes in IgG responses to individual antigens were identified by phage immunoblot. Patterns of IgG recognition following three months of treatment were evaluated using a machine-learned Bayesian Belief Network (ML-BBN. We found that different antigens were recognized following androgen deprivation compared with vaccine therapies. While the number of clinical responders was low in the vaccine-treated populations, we demonstrate that ML-BBN can be used to develop potentially predictive models.

  15. Lymphocyte Proliferation Response to S Antigen in Patients with Uveitis and Optic Neuritis

    Institute of Scientific and Technical Information of China (English)

    PeixianRen; XiuzhenYan

    1995-01-01

    Purpose:To evaluate the autoimmunity which may play a major role in the etiolo-gy of certain forms of uveitis and optic neuritis.Methods:lymphocyte proliferation response to retinal soluble antigen in vitro by gy of certain forms of uveitis and optic neuritis.Methods:Lymphocyte proliferation response toretinal soluble antigen in vitro by incoperation3H-thymidine withDNA was tested in 115patients with anterior u-veitis,posterior/pan-uveitis,optic neuritis,and 50volunteers with unrelated diseases such as congenital ptosis,strabismus,or completely healthy persons as control.Results:The positive rate of lymphocyte stimulation was34%(18/53)in anteri-or uveitis,41.5%(17/41)in posterior/pan-uveitis,and57.1%(12/21)in optic euritis,The results in the experimental groups were significantly different from those of the control group(x2=14.76,P<0.05,x2=19.14P<0.005,x2=26.38,P<0.005,respectively).Conclusion:The autoimmunity plays a role in the patogenesis in certain forms of uveitis and optic neuritis,Such immune responses may be secondary to the expo-sition or release of retinal antigens by various causes,leading to activation or augmentation of meager or low-affinity S antigen specific lymphocytes which may preexist in the circulation and starting the pathogenic autoimmune process.Eye Science 1995;11:120-123.

  16. Advances in alfalfa mosaic virus-mediated expression of anthrax antigen in planta.

    Science.gov (United States)

    Brodzik, R; Bandurska, K; Deka, D; Golovkin, M; Koprowski, H

    2005-12-16

    Plant viruses show great potential for production of pharmaceuticals in plants. Such viruses can harbor a small antigenic peptide(s) as a part of their coat proteins (CP) and elicit an antigen-specific immune response. Here, we report the high yield and consistency in production of recombinant alfalfa mosaic virus (AlMV) particles for specific presentation of the small loop 15 amino acid epitope from domain-4 of the Bacillus anthracis protective antigen (PA-D4s). The epitope was inserted immediately after the first 25 N-terminal amino acids of AlMV CP to retain genome activation and binding of CP to viral RNAs. Recombinant AlMV particles were efficiently produced in tobacco, easily purified for immunological analysis, and exhibited extended stability and systemic proliferation in planta. Intraperitional injections of mice with recombinant plant virus particles harboring the PA-D4s epitope elicited a distinct immune response. Western blotting and ELISA analysis showed that sera from immunized mice recognized both native PA antigen and the AlMV CP. PMID:16236249

  17. Advances in alfalfa mosaic virus-mediated expression of anthrax antigen in planta

    International Nuclear Information System (INIS)

    Plant viruses show great potential for production of pharmaceuticals in plants. Such viruses can harbor a small antigenic peptide(s) as a part of their coat proteins (CP) and elicit an antigen-specific immune response. Here, we report the high yield and consistency in production of recombinant alfalfa mosaic virus (AlMV) particles for specific presentation of the small loop 15 amino acid epitope from domain-4 of the Bacillus anthracis protective antigen (PA-D4s). The epitope was inserted immediately after the first 25 N-terminal amino acids of AlMV CP to retain genome activation and binding of CP to viral RNAs. Recombinant AlMV particles were efficiently produced in tobacco, easily purified for immunological analysis, and exhibited extended stability and systemic proliferation in planta. Intraperitional injections of mice with recombinant plant virus particles harboring the PA-D4s epitope elicited a distinct immune response. Western blotting and ELISA analysis showed that sera from immunized mice recognized both native PA antigen and the AlMV CP

  18. Human antigen-specific regulatory T cells generated by T cell receptor gene transfer.

    Directory of Open Access Journals (Sweden)

    Todd M Brusko

    Full Text Available BACKGROUND: Therapies directed at augmenting regulatory T cell (Treg activities in vivo as a systemic treatment for autoimmune disorders and transplantation may be associated with significant off-target effects, including a generalized immunosuppression that may compromise beneficial immune responses to infections and cancer cells. Adoptive cellular therapies using purified expanded Tregs represents an attractive alternative to systemic treatments, with results from animal studies noting increased therapeutic potency of antigen-specific Tregs over polyclonal populations. However, current methodologies are limited in terms of the capacity to isolate and expand a sufficient quantity of endogenous antigen-specific Tregs for therapeutic intervention. Moreover, FOXP3+ Tregs fall largely within the CD4+ T cell subset and are thus routinely MHC class II-specific, whereas class I-specific Tregs may function optimally in vivo by facilitating direct tissue recognition. METHODOLOGY/PRINCIPAL FINDINGS: To overcome these limitations, we have developed a novel means for generating large numbers of antigen-specific Tregs involving lentiviral T cell receptor (TCR gene transfer into in vitro expanded polyclonal natural Treg populations. Tregs redirected with a high-avidity class I-specific TCR were capable of recognizing the melanoma antigen tyrosinase in the context of HLA-A*0201 and could be further enriched during the expansion process by antigen-specific reactivation with peptide loaded artificial antigen presenting cells. These in vitro expanded Tregs continued to express FOXP3 and functional TCRs, and maintained the capacity to suppress conventional T cell responses directed against tyrosinase, as well as bystander T cell responses. Using this methodology in a model tumor system, murine Tregs designed to express the tyrosinase TCR effectively blocked antigen-specific effector T cell (Teff activity as determined by tumor cell growth and luciferase reporter

  19. Survival and signaling changes in antigen presenting cell subsets after radiation

    Science.gov (United States)

    Parker, Jennifer Janell

    Radiation therapy is a widely used cancer treatment that has the potential to influence anti-tumor immune responses. Both myeloablative and non-myeloablative radiation are often used as part of preparatory regimens for hematopoetic stem cell transplantation, in combination with other chemotherapy or immuno-modulatory (e.g. Anti-thymocyte globulin (ATG)) therapies for both cytotoxic and immune modulatory purposes. However, the mechanisms responsible for the effect of radiation on antigen presenting cell (APC) responsiveness and radioresistance are poorly understood. The first studies described in this thesis were designed to identify and characterize early radiation-induced signaling changes in antigen presenting cells and to determine the effects of these signaling changes on APC receptor expression and function. The NFkappaB pathway in antigen presenting cells was chosen for study because it is activated by radiation in a wide range of other cell types and plays a vital role in the maintenance and regulation of the immune system. The effects of therapeutically relevant doses radiation (2 and 20 Gy) were compared at various timepoints in the human monocytic cell line (U937) using phospho-flow cytometry staining methods and cytometric analysis. These studies demonstrated that radiation-induced changes in the phosphorylation state of NFkappaB family members that were p53 independent. However, these changes were dependent upon activation of ATM in response to single or double-stranded breaks in DNA, as shown in experiments using an inhibitor of ATM and ATM siRNA knockdown U937 cells. In addition, studies examining the effect of radiation on co-stimulatory receptors with and without inhibition of the NFkappaB pathway via phospho-flow cytometry revealed that radiation-induced phosphorylation of NEMO promoted the activation and functional maturation of U937 cells. Furthermore, functional studies using both phospho-flow cytometry and/or mixed lymphocyte reactions to

  20. Anti-cytotoxic T lymphocyte antigen-4 antibodies in melanoma

    Directory of Open Access Journals (Sweden)

    Tosti G

    2013-10-01

    Full Text Available Giulio Tosti, Emilia Cocorocchio, Elisabetta PennacchioliDivisione Melanomi e Sarcomi, Istituto Europeo di Oncologia, Milano, ItalyAbstract: Approaches aimed at enhancement of the tumor specific response have provided proof for the rationale of immunotherapy in cancer, both in animal models and in humans. Ipilimumab, an anti-cytotoxic T lymphocyte antigen-4 (CTLA-4 antibody, is a new generation immunotherapeutic agent that has shown activity in terms of disease free and overall survival in metastatic melanoma patients. Its use was approved by the US Food and Drug Administration in March 2011 to treat patients with late stage melanoma that has spread or that cannot be removed by surgery. The mechanism of action of CTLA-4 antibodies in the activation of an antitumor immune response and selected clinical studies of ipilimumab in advanced melanoma patients are discussed. Ipilimumab treatment has been associated with immune related adverse events due to T-cell activation and proliferation. Most of these serious adverse effects are associated with the gastrointestinal tract and include severe diarrhea and colitis. The relationship between immune related adverse events and antitumor activity associated with ipilimumab was explored in clinical studies. Potential biomarkers predictive for clinical response and survival in patients treated with anti-CTLA-4 therapy are presently under investigation. Besides the conventional patterns of response and stable disease as defined by standard Response Evaluation Criteria in Solid Tumors criteria, in subsets of patients, ipilimumab has shown patterns of delayed clinical activity which were associated with an improved overall survival. For this reason a new set of response criteria for tumor immunotherapy has been proposed, which was termed immune related response criteria. These new criteria are presently used to better analyze clinical activity of immunotherapeutic regimens. Ipilimumab is currently under

  1. Alloantibody Generation and Effector Function Following Sensitization to Human Leukocyte Antigen

    Science.gov (United States)

    Hickey, Michelle J.; Valenzuela, Nicole M.; Reed, Elaine F.

    2016-01-01

    Allorecognition is the activation of the adaptive immune system to foreign human leukocyte antigen (HLA) resulting in the generation of alloantibodies. Due to a high polymorphism, foreign HLA is recognized by the immune system following transplant, transfusion, or pregnancy resulting in the formation of the germinal center and the generation of long-lived alloantibody-producing memory B cells. Alloantibodies recognize antigenic epitopes displayed by the HLA molecule on the transplanted allograft and contribute to graft damage through multiple mechanisms, including (1) activation of the complement cascade resulting in the formation of the MAC complex and inflammatory anaphylatoxins, (2) transduction of intracellular signals leading to cytoskeletal rearrangement, growth, and proliferation of graft vasculature, and (3) immune cell infiltration into the allograft via FcγR interactions with the FC portion of the antibody. This review focuses on the generation of HLA alloantibody, routes of sensitization, alloantibody specificity, and mechanisms of antibody-mediated graft damage. PMID:26870045

  2. Elementary study on the antigenicity of SARS-CoV Mc region

    Institute of Scientific and Technical Information of China (English)

    ZU QIONG HU; WEI ZHAO; WEN BING ZHANG; HUI JUN YAN; LI FANG JIANG; ZHI WEI LIU; ZHI HUA LIU; BEI GUO LONG

    2006-01-01

    To test the antigenic activity of M protein (Mc protein) in the inner membrane of SARS-CoV,SARS-CoV Mc protein's bases locating inside the membrane were cloned, the His-fusion protein was expressed in E. coli and analyzed for its antigenic activity. Among those 7 clinically diagnosed patients' sera, there were 5 positive and 2 negative in reaction with His-fusion protein. All of the 20 healthy persons'sera and rabbit anti-OC43 and 229E were of negative reaction with His-fusion protein. The animals immunized with His-fusion protein have produced multi-clonal antibody. The His-fusion protein could specially react with clinically diagnosed SARS patients' sera and the animals immunized with His-fusion protein could produce specifically multi-clonal antibody, but it could not react with the sera of healthy persons and the rabbit anti-OC43 and 229E.

  3. The perivascular phagocyte of the mouse pineal gland: An antigen-presenting cell

    DEFF Research Database (Denmark)

    Møller, Morten; Rath, Martin F; Klein, David C

    2006-01-01

    The perivascular space of the rat pineal gland is known to contain phagocytic cells that are immunoreactive for leukocyte antigens, and thus they appear to belong to the macrophage/microglial cell line. These cells also contain MHC class II proteins. We investigated this cell type in the pineal...... gland of mice. Actively phagocytosing cells with a prominent lysosomal system were found in the pericapillary spaces of the mouse pineal gland following intravenous injection of horseradish peroxidase. The cells also exhibited strong acid phosphatase activity. Perivascular cells were immunopositive for...... MHC class II protein and for CD68, a marker of monocytes/phagocytes. This study verifies that perivascular phagocytes with antigen-presenting properties are present in the mouse pineal gland....

  4. An Overview of B-1 Cells as Antigen-Presenting Cells

    Science.gov (United States)

    Popi, Ana F.; Longo-Maugéri, Ieda M.; Mariano, Mario

    2016-01-01

    The role of B cells as antigen-presenting cells (APCs) has been extensively studied, mainly in relation to the activation of memory T cells. Considering the B cell subtypes, the role of B-1 cells as APCs is beginning to be explored. Initially, it was described that B-1 cells are activated preferentially by T-independent antigens. However, some reports demonstrated that these cells are also involved in a T-dependent response. The aim of this review is to summarize information about the ability of B-1 cells to play a role as APCs and to briefly discuss the role of the BCR and toll-like receptor signals in this process. Furthermore, some characteristics of B-1 cells, such as natural IgM production and phagocytic ability, could interfere in the participation of these cells in the onset of an adaptive response. PMID:27148259

  5. Pneumocystis carinii from pigs and humans are antigenically distinct

    DEFF Research Database (Denmark)

    Christensen, C B; Settnes, Osvald Peter; Bille-Hansen, Vivi;

    1996-01-01

    The antigens of Pneumocystis carinii cysts isolated from pigs and humans were compared by the Western immunoblotting technique. Convalescent pig serum reacted with two antigens (approximately 78 kDa and 32.5 kDa) of porcine P. carinii cysts, whereas convalescent serum from humans did not react wi...

  6. Protein antigen adsorption to the DDA/TDB liposomal adjuvant

    DEFF Research Database (Denmark)

    Hamborg, Mette; Jorgensen, Lene; Bojsen, Anders Riber;

    2013-01-01

    Understanding the nature of adjuvant-antigen interactions is important for the future design of efficient and safe subunit vaccines, but remains an analytical challenge. We studied the interactions between three model protein antigens and the clinically tested cationic liposomal adjuvant composed...

  7. 9 CFR 113.408 - Avian mycoplasma antigen.

    Science.gov (United States)

    2010-01-01

    ... requirements. A 2.5 ml sample of completed antigen shall be diluted with 2.5 ml of buffer solution formulated... with 9 CFR 114.8. If phenol is used, a direct titration with a standardized bromide-bromate solution... tested against the antigens diluted 1:4 in buffer solution formulated in the same manner as the...

  8. Expression and immunoactivity of chimeric particulate antigens of receptor binding site-core antigen of hepatitis B virus

    Institute of Scientific and Technical Information of China (English)

    Hai-Jie Yang; Ning-Shao Xia; Min Chen; Tong Cheng; Shui-Zhen He; Shao-Wei Li; Bao-Quan Guan; Zi-Heng Zhu; Ying Gu; Jun Zhang

    2005-01-01

    AIM: To improve the immunogenicity of receptor binding site of hepatitis B virus (HBV) on preS1 antigen using HBV core antigen as an immuno-carrier.METHODS: One to 6 tandem copies of HBV preS1 (21-47)fragment were inserted into HBcAg at the sites of aa 78 and 82, and expressed in E. coli. ELISA, Western blot and animal immunization were used to analyze the antigenicity and immmunogenicity of purified particulate antigens. The ability to capture HBV by antibodies elicited by chimeric partides was detected with immuno-capture PCR.RESULTS: Recombinant antigens CⅠ, CⅡ, CⅢ carrying 1-3 copies of HBV preS1 (21-47) individually could form viruslike particles (VLPs), similar to HBcAg in morphology. But recombinant antigens carrying 4-6 copies of HBV preS1 (21-47) were poorly expressed in E.coli. Chimeric antigens were lacking of immunoreactivity with anti-HBc monoclonal antibodies (McAbs), but still reserved good immunoreactivity with anti-HBe McAbs. CⅠ, CⅡ, CⅢ could strongly react with anti-preS1 McAb, suggesting that preS1 (21-47) fragment was well exposed on the surface of chimeric VLPs. Three chimeric VLP antigens (CⅠ, CⅡ and CⅢ) could stimulate mice to produce high-level antibody responses, and their immunogenicity was stronger than non-particulate antigen 21-47*6, containing 6 copies of preS1 (21-47). Mouse antibodies to CⅠ, CⅡ and CⅢ were able to capture HBV virions in immuno-capture PCR assay in vitro.CONCLUSION: Chimeric particulate antigens of receptor binding site-core antigen of HBV can elicit strong antibody responses to preS1. They have a potential to be developed into prophylactic or therapeutic vaccines against HBV infection.

  9. Exploiting in situ antigen generation and immune modulation to enhance chemotherapy response in advanced melanoma: A combination nanomedicine approach.

    Science.gov (United States)

    Lu, Yao; Wang, Yuhua; Miao, Lei; Haynes, Matthew; Xiang, Guangya; Huang, Leaf

    2016-08-28

    Therapeutic anticancer vaccine development must address a number of barriers to achieve successful tumor specific killing, including effective antigen presentation and antigen-specific T-cell activation to mediate cytotoxic cellular effects, inhibition of an immune-suppressive tumor microenvironment in order to facilitate and enhance CTL activity, and induction of memory T-cells to prolong tumor rejection. While traditional as well as modern vaccines rely upon delivery of both antigen and adjuvant, a variety of clinically relevant cancers lack ideal immunogenic antigens. Building upon recent efforts, we instead chose to exploit chemotherapy-induced apoptosis to allow for in situ antigen generation in a combination, nanomedicine-based approach. Specifically, lipid-coated cisplatin nanoparticles (LPC) and CpG-encapsulated liposomes (CpG-Lipo) were prepared for the temporally-controlled and multifaceted treatment of an advanced in vivo model of melanoma. Such combination therapy established strong synergistic effects, both in apoptotic extent and subsequent abrogation of tumor growth, which were due largely to both an enhanced cytotoxic T-cell recruitment and a reduction of immune-suppressive mediators in the microenvironments of both spleens and tumor. These results underlie a prolonged host lifespan in the combination approach (45 days) as compared with control (25 days, p < 0.02), providing promise toward a personalized approach to nanomedicine by establishing effect synergy in host-specific immunotherapy following chemotherapy. PMID:27235608

  10. Study of Immunization against Anthrax with the Purified Recombinant Protective Antigen of Bacillus anthracis

    OpenAIRE

    Singh,Yogendra; Ivins, Bruce E.; Leppla, Stephen H.

    1998-01-01

    Protective antigen (PA) of anthrax toxin is the major component of human anthrax vaccine. Currently available human vaccines in the United States and Europe consist of alum-precipitated supernatant material from cultures of toxigenic, nonencapsulated strains of Bacillus anthracis. Immunization with these vaccines requires several boosters and occasionally causes local pain and edema. We previously described the biological activity of a nontoxic mutant of PA expressed in Bacillus subtilis. In ...

  11. Adjuvant effects of liposomes containing lipid A: enhancement of liposomal antigen presentation and recruitment of macrophages.

    OpenAIRE

    Verma, J N; Rao, M.; Amselem, S; Krzych, U; Alving, C R; Green, S J; Wassef, N M

    1992-01-01

    Liposomes containing lipid A induced potent humoral immune responses in mice against an encapsulated malaria antigen (R32NS1) containing NANP epitopes. The immune response was not enhanced by lipid A alone or by empty liposomes containing lipid A. Experiments to investigate the adjuvant mechanisms of liposomes and lipid A revealed that liposome-encapsulated R32NS1 was actively presented by bone marrow-derived macrophages to NANP-specific cloned T cells. The degree of presentation was related ...

  12. Human lung tumor-associated antigen identified as an extracellular matrix adhesion molecule

    OpenAIRE

    1991-01-01

    A single chain glycoprotein with an estimated molecular mass of 160 kD (gp160) was previously identified as a human lung tumor-associated antigen. This tumor marker is shown here to be associated noncovalently with a second 130-kD protein. Sequential immunoprecipitation studies of surface iodinated lung tumor cell lysates reveal that this heterodimeric complex is indistinguishable serologically and structurally from the integrin VLA-2, found originally on activated T lymphocytes and platelets...

  13. Transcriptional Regulation of the Borrelia burgdorferi Antigenically Variable VlsE Surface Protein

    OpenAIRE

    Bykowski, Tomasz; Babb, Kelly; von Lackum, Kate; Riley, Sean P.; Norris, Steven J; Stevenson, Brian

    2006-01-01

    The Lyme disease agent Borrelia burgdorferi can persistently infect humans and other animals despite host active immune responses. This is facilitated, in part, by the vls locus, a complex system consisting of the vlsE expression site and an adjacent set of 11 to 15 silent vls cassettes. Segments of nonexpressed cassettes recombine with the vlsE region during infection of mammalian hosts, resulting in combinatorial antigenic variation of the VlsE outer surface protein. We now demonstrate that...

  14. Overexpression and Enzymatic Assessment of Antigenic Fragments of Hyaluronidase Recombinant Protein From Streptococcus pyogenes

    OpenAIRE

    Sadoogh Abbasian, Shabnam; Ghaznavi Rad, Ehsanollah; Akbari, Neda; Zolfaghari, Mohammad Reza; Pakzad, Iraj; ABTAHI, Hamid

    2014-01-01

    Background: Hyaluronidase catalyzes the hydrolysis of hyaluronan polymers to N-acetyl-D-glucosamine and D-glucuronic acid. This enzyme is a dimer of identical subunits. Hyaluronidase has different pharmaceutical and medical applications. Previously, we produced a recombinant hyaluronidase antigenic fragment of Streptococcus pyogenes. Objectives: This study aimed to improve the protein production and purity of hyaluronidase recombinant protein from S. pyogenes. In addition, the enzymatic activ...

  15. Involvement of proliferating cell nuclear antigen (cyclin) in DNA replication in living cells.

    OpenAIRE

    Zuber, M; Tan, E M; Ryoji, M

    1989-01-01

    Proliferating cell nuclear antigen (PCNA) (also called cyclin) is known to stimulate the activity of DNA polymerase delta but not the other DNA polymerases in vitro. We injected a human autoimmune antibody against PCNA into unfertilized eggs of Xenopus laevis and examined the effects of this antibody on the replication of injected plasmid DNA as well as egg chromosomes. The anti-PCNA antibody inhibited plasmid replication by up to 67%, demonstrating that PCNA is involved in plasmid replicatio...

  16. Isoniazid induces expression of the antigen 85 complex in Mycobacterium tuberculosis.

    OpenAIRE

    Garbe, T R; Hibler, N S; Deretic, V

    1996-01-01

    Exposure to isoniazid induced the expression of several secreted proteins in Mycobacterium tuberculosis H37Rv. Two-dimensional gel electrophoresis and immunoblot analyses indicated that two of the prominent isonicotinic acid hydrazide-inducible polypeptides were members of the antigen 85 complex, recently demonstrated to have mycolyltransferase activity. We postulate the existence of an intermediate, whose production is inhibited by isonicotinic acid hydrazide, which plays a negative feedback...

  17. Degranulation of human mast cells induces an endothelial antigen central to leukocyte adhesion.

    OpenAIRE

    Klein, L M; Lavker, R M; Matis, W L; Murphy, G F

    1989-01-01

    To understand better the role of mast cell secretory products in the genesis of inflammation, a system was developed for in vitro degranulation of human mast cells in skin organ cultures. Within 2 hr after morphine sulfate-induced degranulation, endothelial cells lining microvessels adjacent to affected mast cells expressed an activation antigen important for endothelial-leukocyte adhesion. Identical results were obtained when other mast cell secretagogues (anti-IgE, compound 48/80, and calci...

  18. A Carcinoembryonic Antigen-Secreting Adenocarcinoma Arising in Tailgut Cyst : Clinical Implications of Carcinoembryonic Antigen

    OpenAIRE

    Cho, Byoung Chul; Kim, Nam Kyu; Lim, Beom Jin; Kang, Sang Ook; Sohn, Ju Hyuk; Roh, Jae Kyung; Choi, Sang Tae; Kim, Sung Ai; Park, Se Eun

    2005-01-01

    Tailgut cysts (TGCs) are rare congenital cysts that occur in the retrorectal or presacral spaces. Although most tailgut cysts have been reported as benign, there have been at least 9 cases associated with malignant change. We report herein on an unusual case of a 40-year-old woman with a carcinoembryonic antigen (CEA)-producing adenocarcinoma arising within a TGC who underwent surgical resection and local radiation therapy. Despite the complete resection, metastatic adenocarcinoma developed f...

  19. [HLA and keloids: antigenic frequency and therapeutic response].

    Science.gov (United States)

    Rossi, A; Bozzi, M

    1989-01-01

    Twenty keloid subjects were typed for class 1 (HLA-A, B and C) and class 2 (HLA-DR and DQ) histocompatibility antigens. Their frequencies were compared to those found in control populations. Of all the antigens belonging to class 1, B 21 was more prevalent in patients. The findings regarding class 2 antigens were noteworthy: in keloid patients there was a significant prevalence of DR 5 (RR = 3.54 and 7.93 respectively for the two control groups) and DQw 3 (RR = 16.8). The patients typed for HLA-antigens were treated with corticosteroid infiltrations. The responses to the treatments were no related to the histocompatibility antigens. PMID:2628278

  20. Mosaic VSGs and the scale of Trypanosoma brucei antigenic variation.

    Directory of Open Access Journals (Sweden)

    James P J Hall

    Full Text Available A main determinant of prolonged Trypanosoma brucei infection and transmission and success of the parasite is the interplay between host acquired immunity and antigenic variation of the parasite variant surface glycoprotein (VSG coat. About 0.1% of trypanosome divisions produce a switch to a different VSG through differential expression of an archive of hundreds of silent VSG genes and pseudogenes, but the patterns and extent of the trypanosome diversity phenotype, particularly in chronic infection, are unclear. We applied longitudinal VSG cDNA sequencing to estimate variant richness and test whether pseudogenes contribute to antigenic variation. We show that individual growth peaks can contain at least 15 distinct variants, are estimated computationally to comprise many more, and that antigenically distinct 'mosaic' VSGs arise from segmental gene conversion between donor VSG genes or pseudogenes. The potential for trypanosome antigenic variation is probably much greater than VSG archive size; mosaic VSGs are core to antigenic variation and chronic infection.