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Sample records for lymphocyte ctl clones

  1. ELISPOT Assay for Monitoring Cytotoxic T Lymphocytes (CTL Activity in Cancer Vaccine Clinical Trials

    Directory of Open Access Journals (Sweden)

    Thomas J. Sayers

    2012-05-01

    Full Text Available The profiling and monitoring of immune responses are key elements in the evaluation of the efficacy and development of new biotherapies, and a number of assays have been introduced for analyzing various immune parameters before, during, and after immunotherapy. The choice of immune assays for a given clinical trial depends on the known or suggested immunomodulating mechanisms associated with the tested therapeutic modality. Cell-mediated cytotoxicity represents a key mechanism in the immune response to various pathogens and tumors. Therefore, the selection of monitoring methods for the appropriate assessment of cell-mediated cytotoxicity is thought to be crucial. Assays that can detect both cytotoxic T lymphocytes (CTL frequency and function, such as the IFN-γ enzyme-linked immunospot assay (ELISPOT have gained increasing popularity for monitoring clinical trials and in basic research. Results from various clinical trials, including peptide and whole tumor cell vaccination and cytokine treatment, have shown the suitability of the IFN-γ ELISPOT assay for monitoring T cell responses. However, the Granzyme B ELISPOT assay and Perforin ELISPOT assay may represent a more direct analysis of cell-mediated cytotoxicity as compared to the IFN-γ ELISPOT, since Granzyme B and perforin are the key mediators of target cell death via the granule-mediated pathway. In this review we analyze our own data and the data reported by others with regard to the application of various modifications of ELISPOT assays for monitoring CTL activity in clinical vaccine trials.

  2. Murine leukemia RL male 1 and sarcoma Meth A antigens recognized by cytotoxic T lymphocytes (CTL).

    Science.gov (United States)

    Uenaka, Akiko; Nakayama, Eiichi

    2003-11-01

    Peptide elution and expression cloning methods have been used to identify T cell-recognized antigens for which no molecular information is available. We identified a unique tumor antigen peptide pRL1a, IPGLPLSL that is recognized by CTL on BALB/c RL male 1 leukemia by peptide elution. The sequence of the peptide corresponded to the normally untranslated 5' region of akt. Cytotoxicity was generated in BALB/c spleen cells by in vivo and in vitro sensitization with pRL1a peptide in the form of multiple antigen peptide (MAP), but not the original form. pRL1a MAP immunization had a significant growth-inhibitory effect. pRL1a MAP was mostly internalized into the endosomal compartment of antigen-presenting cells, leaked to the cytosol, and degraded, and the pRL1a peptide produced was presented through the MHC class I pathway. In vivo depletion of CD4 T cells from tumor-inoculated BALB/c mice caused RL male 1 regression. Overexpression of the RLakt molecule seemed to induce CD4 immunoregulatory cells, which resulted in progressive RL male 1 growth in BALB/c mice. In vivo administration of anti-CD25 mAb (PC61) caused regression of RL male 1, suggesting that CD4(+) CD25(+) immunoregulatory cells were involved in the tumor growth. Recently, we improved the sensitivity and the efficacy of T cell antigen cloning from cDNA expression libraries by using large- and small-scale ELISPOT assays. Using the IFN-gamma ELISPOT method, we obtained a cDNA clone S35 of 937 bp recognized by AT-1 CTL on BALB/c Meth A sarcoma. S35 was a part of the retinoic acid-regulated nuclear matrix-associated protein (ramp). AT-1 CTL recognized the peptide LGAEAIFRL, which was derived from a newly created open reading frame due to the exon 14 extension.

  3. Platelets prevent IFN-alpha/beta-induced lethal hemorrhage promoting CTL-dependent clearance of lymphocytic choriomeningitis virus.

    Science.gov (United States)

    Iannacone, Matteo; Sitia, Giovanni; Isogawa, Masanori; Whitmire, Jason K; Marchese, Patrizia; Chisari, Francis V; Ruggeri, Zaverio M; Guidotti, Luca G

    2008-01-15

    We found that mice infected with different isolates of lymphocytic choriomeningitis virus (LCMV) develop a mild hemorrhagic anemia, which becomes severe and eventually lethal in animals depleted of platelets or lacking integrin beta3. Lethal hemorrhagic anemia is mediated by virus-induced IFN-alpha/beta that causes platelet dysfunction, mucocutaneous blood loss and suppression of erythropoiesis. In addition to the life-threatening hemorrhagic anemia, platelet-depleted mice fail to mount an efficient cytotoxic T lymphocyte (CTL) response and cannot clear LCMV. Transfusion of functional platelets into these animals reduces hemorrhage, prevents death and restores CTL-induced viral clearance in a manner partially dependent on CD40 ligand (CD40L). These results indicate that, upon activation, platelets expressing integrin beta3 and CD40L are required for protecting the host against the induction of an IFN-alpha/beta-dependent lethal hemorrhagic diathesis and for clearing LCMV infection through CTLs.

  4. Attachment of an anti-receptor antibody to non-target cells renders them susceptible to lysis by a clone of cytotoxic T lymphocytes.

    OpenAIRE

    Kranz, D M; Tonegawa, S.; Eisen, H N

    1984-01-01

    The molecular basis for the dependence of antigen recognition by T cells on products of the major histocompatibility complex (MHC) is unknown, and the antigenic structures that are actually bound by T-cell receptors are ill-defined. In this study, we asked whether a monoclonal antibody (mAb) that reacts with the T-cell receptor of a clone of murine cytotoxic T lymphocytes (CTL) and not with the receptors of other CTL clones can substitute for that clone's natural ligand in specific cytolytic ...

  5. Attachment of an anti-receptor antibody to non-target cells renders them susceptible to lysis by a clone of cytotoxic T lymphocytes.

    Science.gov (United States)

    Kranz, D M; Tonegawa, S; Eisen, H N

    1984-12-01

    The molecular basis for the dependence of antigen recognition by T cells on products of the major histocompatibility complex (MHC) is unknown, and the antigenic structures that are actually bound by T-cell receptors are ill-defined. In this study, we asked whether a monoclonal antibody (mAb) that reacts with the T-cell receptor of a clone of murine cytotoxic T lymphocytes (CTL) and not with the receptors of other CTL clones can substitute for that clone's natural ligand in specific cytolytic reactions. To answer the question, a mAb (1B2) to the receptor of a CTL clone (2C) was attached covalently to 51Cr-labeled cells that were not otherwise susceptible to lysis by clone 2C, and the cells thus modified were then tested as targets for clone 2C and other CTL clones of similar specificity. All labeled cells modified in this way, including a murine cell line that expresses no cell-surface MHC class I molecules and a human cell line, were lysed by clone 2C but not by other CTL clones. If, however, instead of attaching the mAb to the receptor of clone 2C, the cells were modified by attaching to them mAbs to other surface antigens on CTL [lymphocyte function-associated antigen (LFA-1), Thy-1.2], they were not lysed. In cytolytic titrations, the cells that had been converted by attachment of mAb 1B2 into specific targets for clone 2C were just as susceptible to lysis by that clone as the clone's natural H-2d targets (e.g., P815 cells). However, some accessory surface molecules (LFA-1, Lyt-2) that are required for clone 2C to lyse its natural H-2d targets seemed not to be required for this clone to lyse the mAb-converted target cells. By demonstrating that a variety of different cell types can be thus converted into target cells for CTL, the approach described in this study may provide opportunities to analyze further the mechanisms by which CTL destroy target cells.

  6. Spontaneous human squamous cell carcinomas are killed by a human cytotoxic T lymphocyte clone recognizing a wild-type p53-derived peptide

    DEFF Research Database (Denmark)

    Röpke, M; Hald, J; Guldberg, Per

    1996-01-01

    A cytotoxic T lymphocyte (CTL) clone generated in vitro from the peripheral blood of a healthy HLA-A2-positive individual against a synthetic p53 protein-derived wild-type peptide (L9V) was shown to kill squamous carcinoma cell lines derived from two head and neck carcinomas, which expressed mutant...

  7. Exhaustion of CTL memory and recrudescence of viremia in lymphocytic choriomeningitis virus-infected MHC class II-deficient mice and B cell-deficient mice

    DEFF Research Database (Denmark)

    Thomsen, Allan Randrup; Johansen, J; Marker, O

    1996-01-01

    To study the contribution of CD4+ T cells and B cells to antiviral immunity and long term virus control, MHC class II-deficient and B cell-deficient mice were infected with lymphocytic choriomeningitis virus. In class II-deficient mice, which lack CD4+ T cells, the primary CTL response is virtually...... this phenomenon could reflect participation of B cells and/or Abs in long term virus control, similar experiments were performed with mice that do not have mature B cells because of a disrupted membrane exon of the mu chain gene. In these mice, the cell-mediated immune response was slightly delayed, but transient...... and that in their absence, the virus-specific CTL potential becomes exhausted. Together our results indicate that while CD8+ cells play a dominant role in acute virus control, all three major components of the immune system are required for long term virus control....

  8. Regulatory B cells inhibit cytotoxic T lymphocyte (CTL activity and elimination of infected CD4 T cells after in vitro reactivation of HIV latent reservoirs.

    Directory of Open Access Journals (Sweden)

    Basile Siewe

    Full Text Available During HIV infection, IL-10/IL-10 receptor and programmed death-1 (PD-1/programmed death-1-ligand (PD-L1 interactions have been implicated in the impairment of cytotoxic T lymphocyte (CTL activity. Despite antiretroviral therapy (ART, attenuated anti-HIV CTL functions present a major hurdle towards curative measures requiring viral eradication. Therefore, deeper understanding of the mechanisms underlying impaired CTL is crucial before HIV viral eradication is viable. The generation of robust CTL activity necessitates interactions between antigen-presenting cells (APC, CD4+ and CD8+ T cells. We have shown that in vitro, IL-10hiPD-L1hi regulatory B cells (Bregs directly attenuate HIV-specific CD8+-mediated CTL activity. Bregs also modulate APC and CD4+ T cell function; herein we characterize the Breg compartment in uninfected (HIVNEG, HIV-infected "elite controllers" (HIVEC, ART-treated (HIVART, and viremic (HIVvir, subjects, and in vitro, assess the impact of Bregs on anti-HIV CTL generation and activity after reactivation of HIV latent reservoirs using suberoylanilide hydroxamic acid (SAHA. We find that Bregs from HIVEC and HIVART subjects exhibit comparable IL-10 expression levels significantly higher than HIVNEG subjects, but significantly lower than HIVVIR subjects. Bregs from HIVEC and HIVART subjects exhibit comparable PD-L1 expression, significantly higher than in HIVVIR and HIVNEG subjects. SAHA-treated Breg-depleted PBMC from HIVEC and HIVART subjects, displayed enhanced CD4+ T-cell proliferation, significant upregulation of antigen-presentation molecules, increased frequency of CD107a+ and HIV-specific CD8+ T cells, associated with efficient elimination of infected CD4+ T cells, and reduction in integrated viral DNA. Finally, IL-10-R and PD-1 antibody blockade partially reversed Breg-mediated inhibition of CD4+ T-cell proliferation. Our data suggest that, possibly, via an IL-10 and PD-L1 synergistic mechanism; Bregs likely inhibit APC

  9. Definition of an epitope on NS3 recognized by human CD4+ cytotoxic T lymphocyte clones cross-reactive for dengue virus types 2, 3, and 4.

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    Kurane, I; Zeng, L; Brinton, M A; Ennis, F A

    1998-01-20

    The role of dengue virus-specific serotype-cross-reactive T lymphocytes in recovery from and pathogenesis of dengue virus infections is not known. In the present paper, we have defined a dengue serotype-cross-reactive epitope recognized by two CD4+ CD8- cytotoxic T lymphocyte (CTL) clones, JK36 and JK46. These T cell clones were established from the peripheral blood T lymphocytes of a dengue-3-immune donor, using a limiting dilution method. JK36 and JK46 were cross-reactive for dengue virus types 2, 3, and 4, but not for type 1, and recognized the NS3 protein. The smallest synthetic peptide recognized by JK36 was an 8-amino acid peptide that contains amino acids (aa) 226 to 233 (VVAAEMEE) of NS3. The smallest peptide recognized by JK46 was an 11-amino acid peptide that contains aa 224 to 234 (TRVVAAEMEEA). HLA-DR15 was the restriction allele for recognition of these peptides by both JK36 and JK46. This is the first epitope to be defined that is recognized by human CD4+ CTL cross-reactive for dengue virus types 2, 3, and 4.

  10. Flavivirus-cross-reactive, HLA-DR15-restricted epitope on NS3 recognized by human CD4+ CD8- cytotoxic T lymphocyte clones.

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    Kurane, I; Okamoto, Y; Dai, L C; Zeng, L L; Brinton, M A; Ennis, F A

    1995-09-01

    The role of flavivirus-cross-reactive T lymphocytes in recovery from and pathogenesis of flavivirus infections is not known. In the present paper, we have defined a flavivirus-cross-reactive epitope recognized by two CD4+ CD8- cytotoxic T lymphocyte (CTL) clones, JK4 and JK43. The T cell clones were established from the peripheral blood T lymphocytes of a dengue-4-immune donor, using a limiting-dilution method with dengue-4 antigen. These two T cell clones were cross-reactive for dengue virus types 1, 2, 3 and 4, yellow fever virus and West Nile virus, and recognized NS3 protein. The smallest synthetic peptide recognized by these T cell clones was an identical 9 amino acid peptide which contains amino acids 146 to 154 (VIGLYGNGV) of dengue-4 NS3. HLA-DR15 was the restriction allele for recognition of this epitope by JK4 and JK43. JK4 and JK43 both used T cell receptor V alpha 8, but JK4 used V beta 8 and JK43 used V beta 2. This result indicates that this epitope is recognized by two flavivirus-cross-reactive CD4+ T cell clones which originated from different T cells in vivo.

  11. Sequence conservation of subdominant HLA-A2-binding CTL epitopes in HIV-1 clinical isolates and CD8+ T-lymphocyte cross-recognition may explain the immune reaction in infected individuals

    DEFF Research Database (Denmark)

    Thorn, Mette; Tang, Sheila; Therrien, Dominic;

    2007-01-01

    Cytotoxic T-lymphocytes (CTL) are critical for immune control of infection with human immunodeficiency virus type-1 (HIV-1) and searches for relevant CTL epitopes for immune therapy are ongoing. Recently, we identified 28 HLA-A2-binding HIV-1 CTL epitopes (1). In this follow-up study we fully...... genome sequenced HIV-1 from 11 HLA-A2(+) patients to examine the sequence variation of these natural epitopes and compared them with the patient's CD8(+) T-cell recall response. Often the epitope was conserved but only a few patients showed a CD8(+) T-cell recall response. This infrequent targeting may...... be explained by immune subdominance. CD8(+) T-cell recall response to a natural epitope could be measured despite sequence differences in the patient's virus. T-cell cross-reaction between such variants could be demonstrated in HLA-A2 transgenic mice. Nine infrequently targeted but conserved or cross...

  12. Nicotine inhibits memory CTL programming.

    Directory of Open Access Journals (Sweden)

    Zhifeng Sun

    Full Text Available Nicotine is the main tobacco component responsible for tobacco addiction and is used extensively in smoking and smoking cessation therapies. However, little is known about its effects on the immune system. We confirmed that multiple nicotinic receptors are expressed on mouse and human cytotoxic T lymphocytes (CTLs and demonstrated that nicotinic receptors on mouse CTLs are regulated during activation. Acute nicotine presence during activation increases primary CTL expansion in vitro, but impairs in vivo expansion after transfer and subsequent memory CTL differentiation, which reduces protection against subsequent pathogen challenges. Furthermore, nicotine abolishes the regulatory effect of rapamycin on memory CTL programming, which can be attributed to the fact that rapamycin enhances expression of nicotinic receptors. Interestingly, naïve CTLs from chronic nicotine-treated mice have normal memory programming, which is impaired by nicotine during activation in vitro. In conclusion, simultaneous exposure to nicotine and antigen during CTL activation negatively affects memory development.

  13. Nicotine inhibits memory CTL programming.

    Science.gov (United States)

    Sun, Zhifeng; Smyth, Kendra; Garcia, Karla; Mattson, Elliot; Li, Lei; Xiao, Zhengguo

    2013-01-01

    Nicotine is the main tobacco component responsible for tobacco addiction and is used extensively in smoking and smoking cessation therapies. However, little is known about its effects on the immune system. We confirmed that multiple nicotinic receptors are expressed on mouse and human cytotoxic T lymphocytes (CTLs) and demonstrated that nicotinic receptors on mouse CTLs are regulated during activation. Acute nicotine presence during activation increases primary CTL expansion in vitro, but impairs in vivo expansion after transfer and subsequent memory CTL differentiation, which reduces protection against subsequent pathogen challenges. Furthermore, nicotine abolishes the regulatory effect of rapamycin on memory CTL programming, which can be attributed to the fact that rapamycin enhances expression of nicotinic receptors. Interestingly, naïve CTLs from chronic nicotine-treated mice have normal memory programming, which is impaired by nicotine during activation in vitro. In conclusion, simultaneous exposure to nicotine and antigen during CTL activation negatively affects memory development.

  14. Acute myeloid leukemia (AML)-reactive cytotoxic T lymphocyte clones rapidly expanded from CD8(+) CD62L((high)+) T cells of healthy donors prevent AML engraftment in NOD/SCID IL2Rgamma(null) mice.

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    Distler, Eva; Wölfel, Catherine; Köhler, Sylvia; Nonn, Marion; Kaus, Nina; Schnürer, Elke; Meyer, Ralf G; Wehler, Thomas C; Huber, Christoph; Wölfel, Thomas; Hartwig, Udo F; Herr, Wolfgang

    2008-04-01

    Current in vitro techniques for isolating leukemia-reactive cytotoxic T lymphocytes (CTLs) from healthy donors are of relatively low efficiency and yield responder populations with unknown biological significance. This study aimed at the development of a more reliable approach, allowing generation and expansion of acute myeloid leukemia (AML)-reactive CTLs using primary in vitro stimulation. We established allogeneic mini-mixed lymphocyte-leukemia cultures (mini-MLLCs) by stimulating donor CD8(+) T cells with human leukocyte antigen (HLA) class I-matched AML blasts in microtiter plates. Before culture, CD8(+) T cells were separated into CD62L((high)+) and CD62L((low)+/neg) subsets enriched for naive/central memory and effector memory cells, respectively. In eight different related and unrelated donor/AML pairs, numerous CTL populations were isolated that specifically lysed myeloid leukemias in association with various HLA-A, -B, or -C alleles. These CTLs expressed T-cell receptors of single Vbeta-chain families, indicating their clonal origin. The majority of CTL clones were obtained from mini-MLLCs initiated with CD62L((high)+) cells. Using antigen-specific stimulation, multiple CTL populations were amplified to 10(8)-10(10) cells within 6 to 8 weeks. Three of four representative CTL clones were capable of completely preventing engraftment of human primary AML blasts in nonobese diabetic/severe combined immune deficient IL2Rgamma(null) mice. The mini-MLLC approach allows the efficient in vitro expansion of AML-reactive CTL clones from CD8(+)CD62L((high)+) precursors of healthy donors. These CTLs can inhibit leukemia engraftment in immunodeficient mice, suggesting their potential biological relevance.

  15. Identification of two epitopes on the dengue 4 virus capsid protein recognized by a serotype-specific and a panel of serotype-cross-reactive human CD4+ cytotoxic T-lymphocyte clones.

    OpenAIRE

    Gagnon, S J; Zeng, W.; Kurane, I; Ennis, F A

    1996-01-01

    We analyzed the CD4+ T-lymphocyte response of a donor who had received an experimental live-attenuated dengue 4 virus (D4V) vaccine. Bulk culture proliferative responses of peripheral blood mononuclear cells (PBMC) to noninfectious dengue virus (DV) antigens showed the highest proliferation to D4V antigen, with lesser, cross-reactive proliferation to D2V antigen. We established CD4+ cytotoxic T-lymphocyte clones (CTL) by stimulation with D4 antigen. Using recombinant baculovirus antigens, we ...

  16. CTL ELISPOT assay.

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    Ranieri, Elena; Popescu, Iulia; Gigante, Margherita

    2014-01-01

    Enzyme-linked immune absorbent spot (Elispot) is a quantitative method for measuring relevant parameters of T cell activation. The sensitivity of Elispot allows the detection of low-frequency antigen-specific T cells that secrete cytokines and effector molecules, such as granzyme B and perforin. Cytotoxic T cell (CTL) studies have taken advantage with this high-throughput technology by providing insights into quantity and immune kinetics. Accuracy, sensitivity, reproducibility, and robustness of Elispot resulted in a wide range of applications in research as well as in diagnostic field. Actually, CTL monitoring by Elispot is a gold standard for the evaluation of antigen-specific T cell immunity in clinical trials and vaccine candidates where the ability to detect rare antigen-specific T cells is of relevance for immune diagnostic. The most utilized Elispot assay is the interferon-gamma (IFN-γ) test, a marker for CD8(+) CTL activation, but Elispot can also be used to distinguish different subsets of activated T cells by using other cytokines such as T-helper (Th) 1-type cells (characterized by the production of IFN-γ, IL-2, IL-6, IL-12, IL-21, and TNF-α), Th2 (producing cytokines like IL-4, IL-5, IL-10, and IL-13), and Th17 (IL-17) cells. The reliability of Elispot-generated data, by the evaluation of T cell frequency recognizing individual antigen/peptide, is the core of this method currently applied widely to investigate specific immune responses in cancer, infections, allergies, and autoimmune diseases. The Elispot assay is competing with other methods measuring single-cell cytokine production, e.g., intracellular cytokine by FACS or Miltenyi cytokine secretion assay. Other types of lymphocyte frequency and function assays include limiting dilution assay (LDA), cytotoxic T cell assay (CTL), and tetramer staining. Compared with respect to sensitivity the Elispot assay is outranking other methods to define frequency of antigen-specific lymphocytes. The method

  17. Escape of leukemia blasts from HLA-specific CTL pressure in a recipient of HLA one locus-mismatched bone marrow transplantation.

    Science.gov (United States)

    Kato, Tomonori; Terakura, Seitaro; Murata, Makoto; Sugimoto, Kyoko; Murase, Miho; Iriyama, Chisako; Tomita, Akihiro; Abe, Akihiro; Suzuki, Momoko; Nishida, Tetsuya; Naoe, Tomoki

    2012-01-01

    A case of leukemia escape from an HLA-specific cytotoxic T lymphocyte (CTL) response in a recipient of bone marrow transplantation is presented. Only the expression of HLA-B51, which was a mismatched HLA locus in the graft-versus-host direction, was down-regulated in post-transplant leukemia blasts compared with that in pre-transplant blasts. All CTL clones, that were isolated from the recipient's blood when acute graft-versus-host disease developed, recognized the mismatched B(∗)51:01 molecule in a peptide-dependent manner. The pre-transplant leukemia blasts were lysed by CTL clones, whereas the post-transplant leukemia blasts were not lysed by any CTL clones. The IFN-γ ELISPOT assay revealed that B(∗)51:01-reactive T lymphocytes accounted for the majority of the total alloreactive T lymphocytes in the blood just before leukemia relapse. These data suggest that immune escape of leukemia blasts from CTL pressure toward a certain HLA molecule can lead to clinical relapse after bone marrow transplantation. Copyright © 2012 Elsevier Inc. All rights reserved.

  18. Nicotine Inhibits Memory CTL Programming

    OpenAIRE

    Zhifeng Sun; Kendra Smyth; Karla Garcia; Elliot Mattson; Lei Li; Zhengguo Xiao

    2013-01-01

    Nicotine is the main tobacco component responsible for tobacco addiction and is used extensively in smoking and smoking cessation therapies. However, little is known about its effects on the immune system. We confirmed that multiple nicotinic receptors are expressed on mouse and human cytotoxic T lymphocytes (CTLs) and demonstrated that nicotinic receptors on mouse CTLs are regulated during activation. Acute nicotine presence during activation increases primary CTL expansion in vitro, but imp...

  19. Broad CTL Response in Early HIV Infection Drives Multiple Concurrent CTL Escapes.

    Science.gov (United States)

    Leviyang, Sivan; Ganusov, Vitaly V

    2015-10-01

    Recent studies have highlighted the ability of HIV to escape from cytotoxic T lymphocyte (CTL) responses that concurrently target multiple viral epitopes. Yet, the viral dynamics involved in such escape are incompletely understood. Previous analyses have made several strong assumptions regarding HIV escape from CTL responses such as independent or non-concurrent escape from individual CTL responses. Using experimental data from evolution of HIV half genomes in four patients we observe concurrent viral escape from multiple CTL responses during early infection (first 100 days of infection), providing confirmation of a recent result found in a study of one HIV-infected patient. We show that current methods of estimating CTL escape rates, based on the assumption of independent escapes, are biased and perform poorly when CTL escape proceeds concurrently at multiple epitopes. We propose a new method for analyzing longitudinal sequence data to estimate the rate of CTL escape across multiple epitopes; this method involves few parameters and performs well in simulation studies. By applying our novel method to experimental data, we find that concurrent multiple escapes occur at rates between 0.03 and 0.4 day(-1), a relatively broad range that reflects uncertainty due to sparse sampling and wide ranges of parameter values. However, we show that concurrent escape at rates 0.1-0.2 day(-1) across multiple epitopes is consistent with our patient datasets.

  20. T-cell-receptor engagement and tumor ICAM-1 up-regulation are required to by-pass low susceptibility of melanoma cells to autologous CTL-mediated lysis.

    Science.gov (United States)

    Anichini, A; Mortarini, R; Alberti, S; Mantovani, A; Parmiani, G

    1993-04-01

    Tumor-specific and non-specific CD3+, TcR alpha beta+, CD8+ cytotoxic T-cell (CTL) clones, isolated from tumor-infiltrating lymphocytes (TIL) or peripheral blood lymphocytes (PBL) of a melanoma patient and allogeneic LAK cells, were used to investigate the requirements for bypassing the low lysability of some melanoma clones derived from an s.c. metastasis from which highly lysable clones were also obtained. Cytofluorimetric analysis showed that all melanoma clones expressed ICAM-1, although to different extents, reaching a 10-fold difference in fluorescence units, while HLA class-I antigens were similarly expressed. The differences in expression of ICAM-1 among tumor clones correlated with differences in lysability, by both specific and non-specific CTL, but were not large enough to affect lymphocyte-tumor conjugate formation. Cytokine- or gene-transfer-mediated up-regulation of ICAM-1 did not induce de novo lysis of ICAM-1low tumor cells; however, it markedly enhanced a low level of killing of the same cells by tumor-specific, TcR-dependent and HLA-restricted CTL clones but not by non-specific, TcR-independent effectors. In addition, lysis of melanoma clones by any effector was similarly inhibited by anti-ICAM-1 and anti-LFA-1 antibodies. This indicates that by-pass of low lysability of ICAM-1low melanoma clones by CTL clones, after ICAM-1 up-regulation, is possible only if simultaneous LFA-1 and TcR engagement takes place. In addition, these results suggest that the constitutive high level of expression of ICAM-1 on the subset of ICAM-1high melanoma cells must be only one of the factors contributing to the high lysability of these cells by any effector.

  1. In vitro characterization of human AML-reactive CTL clones generated from the naive subset of healthy donors and adoptive transfer into leukemia-engrafted NSG mice

    OpenAIRE

    Albrecht, Jana

    2012-01-01

    Acute myeloid leukemia (AML) is a very aggressive cancer of the hematopoietic system. Chemotherapy and immunotherapeutical approaches including hematopoietic stem cell transplantation (HSCT) and donor lymphocyte infusion (DLI) are the only curative options available. The beneficial graft-versus-leukemia (GVL) effect of cellular immunotherapy is mostly mediated by donor-derived CD8+ T lymphocytes that recognize minor histocompatibility antigens (mHags) and leukemia-associated antigens (LAAs) p...

  2. Study on the Cytotoxic T Lymphocytes Clone Specific for the Nucleocapsid Protein of Hantaan Virus from Peripheral Blood in Patients with Hemorrhagic Fever with Renal Syndrome

    Institute of Scientific and Technical Information of China (English)

    潘蕾; 白雪帆; 黄长形; 李光玉

    2003-01-01

    In order to elucidate the molecular and immunological mechanisms as well as the pathogenesis of hemorrhagic fever with renal syndrome (HFRS), the CD8+ cytotoxic T lymphecytes (CTL) clone was established directly from peripheral blood mononuclear cells (PBMC) of patients with HFRS. The activities of CTL were detected as usual with EBV-transformed lymphoblastoid cell line (BLCL) as target cells. The results showed that the CTL clone could recognized and killed the targetcells with specificity of nucleocapsid protein of Hantaan virus (HTNVNP) with the cytotoxicity percentages of 50.2%,25.4% and 39.0% respectively. These results demonstrated that the antigenic epitopes of HTNVNP mainly located on the C-temainal of the viral nucleocapsid protein.

  3. Cytotoxic T-lymphocyte-mediated killing of human pancreatic islet cells in vitro.

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    Campbell, Peter D; Estella, Eugene; Dudek, Nadine L; Jhala, Gaurang; Thomas, Helen E; Kay, Thomas W H; Mannering, Stuart I

    2008-09-01

    Cytotoxic T lymphocytes (CTL) are believed to play an essential role in beta-cell destruction leading to development of type 1 diabetes and allogeneic islet graft failure. We aimed to identify the mechanisms used by CTL to kill human beta cells. CTL clones that recognize epitopes from influenza virus and Epstein-Barr virus restricted by human leukocyte antigen (HLA)-A0201 and -B0801, respectively, were used to investigate the susceptibility of human beta cells to CTL. In a short-term (5-hour) assay, CTL killed human islet cells of the appropriate major histocompatibility complex (MHC) class I type that had been pulsed with viral peptides. Killing was increased by pretreating islets with interferon gamma that increases MHC class I on target cells. Killing was abolished by incubation of CTL with the perforin inhibitor concanamycin A. The Fas pathway did not contribute to killing because blocking with neutralizing anti-Fas ligand antibody did not significantly reduce beta-cell killing. In conclusion, we report a novel way of investigating the interaction between CTL and human islets. Human islets were rapidly killed in vitro by MHC class I-restricted CTL predominantly by the granule exocytosis pathway.

  4. In vitro generation and characterization of acute myeloid leukemia-reactive CD8 + cytotoxic T-lymphocyte clones from healthy donors

    OpenAIRE

    Distler, Eva

    2007-01-01

    Donor-derived CD8+ cytotoxic T lymphocytes (CTLs) eliminating host leukemic cells mediate curative graft-versus-leukemia (GVL) reactions after allogeneic hematopoietic stem cell transplantation (HSCT). The leukemia-reactive CTLs recognize hematopoiesis-restricted or broadly expressed minor histocompatibility and leukemia-associated peptide antigens that are presented by human leukocyte antigen (HLA) class I molecules on recipient cells. The development of allogeneic CTL therapy in acute myelo...

  5. Molecular cloning and characterization of lymphocyte cell kinase from humphead snapper (Lutjanus sanguineus).

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    Huang, Y; Cai, J; Wang, B; Tang, J-F; Jian, J-C; Wu, Z-H; Gan, Z; Lu, Y-S

    2016-07-01

    Lymphocyte cell kinase (LCK) belongs to the Src family of tyrosine kinases, which involves in the proliferation control of lymphocytes. In this study, we cloned the LCK gene of humphead snapper (Lutjanus sanguineus) (designed as LsLCK). Sequence analysis showed that the full-length cDNA of LsLCK was 2279 bp, contained a 1506-bp open reading frame (ORF), encoding a polypeptide of 501 amino acids. The deduced amino acid possessed the typical structural features of known LCK proteins, including four Src homology (SH) domains arranged as the SH1 domain followed by a regulatory C-terminal tail (COOH-domain), SH2 and SH3 adapter domains and SH4 domain which required for membrane attachment and CD4/CD8 binding. Fluorescent quantitative real-time PCR analysis indicated that LsLCK transcripts were expressed mainly in thymus, spleen and head kidney in healthy fish. Moreover, the mRNA expressions in these tissues were significantly up-regulated after challenge with Vibrio harveyi. The results of immunohistochemistry showed that LsLCK protein localized distinctly in cytoplasm of cell in thymus, spleen and head kidney. Taken together, these findings indicated that LsLCK may play an important role in the immune response of humphead snapper against bacterial infection.

  6. Investigating CTL mediated killing with a 3D cellular automaton.

    Directory of Open Access Journals (Sweden)

    Frederik Graw

    2009-08-01

    Full Text Available Cytotoxic T lymphocytes (CTLs are important immune effectors against intra-cellular pathogens. These cells search for infected cells and kill them. Recently developed experimental methods in combination with mathematical models allow for the quantification of the efficacy of CTL killing in vivo and, hence, for the estimation of parameters that characterize the effect of CTL killing on the target cell populations. It is not known how these population-level parameters relate to single-cell properties. To address this question, we developed a three-dimensional cellular automaton model of the region of the spleen where CTL killing takes place. The cellular automaton model describes the movement of different cell populations and their interactions. Cell movement patterns in our cellular automaton model agree with observations from two-photon microscopy. We find that, despite the strong spatial nature of the kinetics in our cellular automaton model, the killing of target cells by CTLs can be described by a term which is linear in the target cell frequency and saturates with respect to the CTL levels. Further, we find that the parameters describing CTL killing on the population level are most strongly impacted by the time a CTL needs to kill a target cell. This suggests that the killing of target cells, rather than their localization, is the limiting step in CTL killing dynamics given reasonable frequencies of CTL. Our analysis identifies additional experimental directions which are of particular importance to interpret estimates of killing rates and could advance our quantitative understanding of CTL killing.

  7. Screening of YAC clones and building a map of the chromosome 13 region often deleted during chronic B-cell lymphocytic leucosis

    NARCIS (Netherlands)

    Brodyanskii, VM; Sulimova, GE; Udina, IG; Aitova, SS; Shaikhaev, GO; Sharikova, OA; Zakharev, VM; Fedorova, LI; Zelenin, AV; Eikhorn, S; Baush, C; Laland, M; Ross, M; Yankovskii, NK

    1995-01-01

    Pools of YAC clones from the ICRF library were analyzed by PCR using PBKpt, MGG15, and D13S25 markers that flank the chromosome 13 region often deleted during chronic lymphocytic leukemia. Ten clones were found and described. Nine mega-YAC clones from the CEPH library flanking the region of interest

  8. Cytotoxic T-lymphocyte clones, established by stimulation with the HLA-A2 binding p5365-73 wild type peptide loaded on dendritic cells In vitro, specifically recognize and lyse HLA-A2 tumour cells overexpressing the p53 protein

    DEFF Research Database (Denmark)

    Barfoed, Annette Malene; Petersen, T R; Kirkin, A F;

    2000-01-01

    to carry identical T-cell receptors. The CTL clone, 2D9, was shown to specifically lyse the HLA-A*0201+ squamous carcinoma cell line SCC9 and the breast cancer cell line MDA-MB-468. Our data demonstrate that human peripheral blood lymphocytes from normal healthy individuals comprise T cells capable...... of recognizing p53 derived wild type (self) peptides. Furthermore, the capacity of R9V specific T cell clones to exert HLA restricted cytotoxicity, argues that the R9V peptide is naturally presented on certain cancer cells. This supports the view that p53 derived wild type peptides might serve as candidate...... target antigens for the immunotherapeutic treatment of cancer....

  9. Cytotoxic T lymphocyte responses against melanocytes and melanoma

    Directory of Open Access Journals (Sweden)

    Schwartz Erich J

    2011-07-01

    Full Text Available Abstract Background Vitiligo is a common toxicity associated with immunotherapy for melanoma. Cytotoxic T lymphocytes (CTLs against melanoma commonly target melanoma-associated antigens (MAAs which are also expressed by melanocytes. To uncouple vitiligo from melanoma destruction, it is important to understand if CTLs can respond against melanoma and melanocytes at different levels. Methods To understand the dichotomous role of MAA-specific CTL, we characterized the functional reactivities of established CTL clones directed to MAAs against melanoma and melanocyte cell lines. Results CTL clones generated from melanoma patients were capable of eliciting MHC-restricted, MAA-specific lysis against melanocyte cell lines as well as melanoma cells. Among the tested HLA-A*0201-restricted CTL clones, melanocytes evoked equal to slightly higher degranulation and cytolytic responses as compared to melanoma cells. Moreover, MAA-specific T cells from vaccinated patients responded directly ex vivo to melanoma and melanocytes. Melanoma cells express slightly higher levels of MART-1 and gp100 than melanocytes as measured by quantitative reverse-transcriptase polymerase chain reaction (qRT-PCR and immunohistochemistry. Conclusions Our data suggest that CTLs respond to melanoma and melanocytes equally in vitro and directly ex vivo.

  10. Oligoclonal CD8 lymphocytes from persons with asymptomatic human immunodeficiency virus (HIV) type 1 infection inhibit HIV-1 replication.

    Science.gov (United States)

    Toso, J F; Chen, C H; Mohr, J R; Piglia, L; Oei, C; Ferrari, G; Greenberg, M L; Weinhold, K J

    1995-10-01

    CD8 lymphocytes from asymptomatic human immunodeficiency virus (HIV) type 1-infected patients can suppress virus production from infected CD4 cells. Suppressive activity is separate and distinct from cytotoxic T lymphocyte (CTL) reactivities and is likely mediated by a soluble factor(s). The majority of HIV-1 suppression studies have been done in the context of bulk CD8 cell cultures. In this study, viral suppression was characterized by clonal populations of CD8 cells derived from HIV-1-infected patients. Most of the suppressive clones were devoid of detectable CTL reactivity against env-, gag-, pol-, and nef-expressing targets. Among the suppressive clones derived from an individual patient, a marked heterogeneity was evident with respect to phenotypic markers, cytokine production, and T cell receptor V beta expression. These results suggest that noncytolytic virus suppression is oligoclonal in nature. Clones provide tools for future studies aimed at understanding the mechanism of suppression and identifying the suppressive factor.

  11. Antimelanoma CTL recognizes peptides derived from an ORF transcribed from the antisense strand of the 3′ untranslated region of TRIT1

    Directory of Open Access Journals (Sweden)

    Rolf K Swoboda

    2014-01-01

    Full Text Available Noncoding regions of the genome play an important role in tumorigenesis of cancer. Using expression cloning, we have identified a cytotoxic T lymphocyte (CTL–defined antigen that recognizes a protein sequence derived from an open reading frame transcribed from the reverse strand in the 3′ untranslated region of tRNA isopentenyltransferase 1 (TRIT1. A peptide derived from this open reading frame (ORF sequence and predicted to bind to HLA-B57, sensitized HLA-B57+ tumor cells to lysis by CTL793. The peptide also induced a CTL response in peripheral blood mononuclear cells (PBMC of patient 793 and in two other melanoma patients. The CTL lysed peptide-pulsed HLA-B57+ target cells and melanoma cells with endogenous antigen expression. The recognition of this antigen is not limited to HLA-B57-restricted CTLs. An HLA-A2 peptide derived from the ORF was able to induce CTLs in PBMC of 2 HLA-A2+ patients. This study describes for the first time a CTL-defined melanoma antigen that is derived from an ORF on the reverse strand of the putative tumor suppressor gene TRIT1. This antigen has potential use as a vaccine or its ability to induce CTLs in vitro could be used as a predictive biomarker.

  12. Selective Growth, In Vitro and In Vivo, of Individual T Cell Clones from Tumor-Infiltrating Lymphocytes Obtained from Patients with Melanoma

    Science.gov (United States)

    Zhou, Juhua; Dudley, Mark E.; Rosenberg, Steven A.; Robbins, Paul F.

    2007-01-01

    In recent clinical trials in patients with metastatic melanoma, adoptive transfer of tumor-reactive lymphocytes mediated the regression of metastatic tumor deposits. To better understand the role of individual T cell clones in mediating tumor regression, a 5′ RACE technique was used to determine the distribution of TCR β-chain V region sequences expressed in the transferred cells as well as in tumor samples and circulating lymphocytes from melanoma patients following adoptive cell transfer. We found that dominant T cell clones were present in the in vitro-expanded and transferred tumor-infiltrating lymphocyte samples and certain T cell clones including the dominant T cell clones persisted at relatively high levels in the peripheral blood of the patients that demonstrated clinical responses to adoptive immunotherapy. However, these dominant clones were either undetected or present at a very low level in the resected tumor samples used for tumor-infiltrating lymphocyte generation. These data demonstrated that there was selective growth and survival, both in vitro and in vivo, of individual T cell clones from a relatively small number of T cells in the original tumor samples. These results suggest that the persistent T cell clones played an active role in mediating tumor regression and that 5′ RACE analysis may provide an important tool for the analysis of the role of individual T cell clones in mediating tumor regression. A similar analysis may also be useful for monitoring autoimmune responses. PMID:15585890

  13. Multi Dimensional CTL and Stratified Datalog

    Directory of Open Access Journals (Sweden)

    Theodore Andronikos

    2010-02-01

    Full Text Available In this work we define Multi Dimensional CTL (MD-CTL in short by extending CTL which is thedominant temporal specification language in practice. The need for Multi Dimensional CTL is mainlydue to the advent of semi-structured data. The common path nature of CTL and XPath which provides asuitable model for semi-structured data, has caused the emergence of work on specifying a relation amongthem aiming at exploiting the nice properties of CTL. Although the advantages of such an approach havealready been noticed [36, 26, 5], no formal definition of MD-CTL has been given. The goal of this workis twofold; a we define MD-CTL and prove that the “nice” properties of CTL (linear model checking andbounded model property transfer also to MD-CTL, b we establish new results on stratified Datalog. Inparticular, we define a fragment of stratified Datalog called Multi Branching Temporal (MBT in shortprograms that has the same expressive power as MD-CTL. We prove that by devising a linear translationbetween MBT and MD-CTL. We actually give the exact translation rules for both directions. We furtherbuild on this relation to prove that query evaluation is linear and checking satisfiability, containment andequivalence are EXPTIME–complete for MBT programs. The class MBT is the largest fragment of stratifiedDatalog for which such results exist in the literature.

  14. DC 荷载α-GalCer 联合肿瘤特异性 CTL 对小鼠 Heps 肝癌移植瘤生长的影响%Effects of DC load α-GalCer combined with tumor specific cytotoxic T lymphocyte on the growth of transplanted Heps hepatoma in mice

    Institute of Scientific and Technical Information of China (English)

    王鹏; 栾智勇; 张南征; 吴亮群; 刘军权; 杭敏; 潘进

    2016-01-01

    目的:探讨树突状细胞(DC)荷载α-半乳糖神经酰胺(α-GalCer)联合肿瘤特异性细胞毒 T 淋巴细胞(CTL)对小鼠 Heps 肝癌移植瘤生长的抑制作用。方法:诱导扩增小鼠骨髓来源的 DC 细胞和 T 淋巴细胞,培养成为具有肿瘤特异性的 CTL,DC 细胞体外荷载α-GalCer。建立 Heps 肝癌移植瘤模型,将36只模型鼠随机分为4组(n =9),分别尾静脉给予生理盐水(对照组)、CTL(CTL 组)、DC 荷载α-GalCer(DC 荷载α-GalCer 组)、DC 细胞荷载α-GalCer +CTL(联合治疗组)输注。每隔两天测量移植瘤体积,观察体积变化。于治疗后第14d,处死小鼠,取眼球血及瘤体组织。流式细胞术(FCM)检测外周血 CD4+、CD8+ T 细胞比例。免疫组织化学染色观察肿瘤组织中 Bcl -2/Bax 凋亡蛋白的表达。结果:与对照组相比,各治疗组均可抑制肿瘤的生长,差异具有统计学意义(P <0.01);联合治疗组较 DC 荷载α-GalCer 组及 CTL 组肿瘤体积明显减小(P <0.05)。联合治疗组小鼠外周血 CD4+、CD8+ T 细胞数量较另外三组显著升高(P <0.05);对照组、CTL组、DC 荷载α-GalCer 组小鼠外周血中 CD4+、CD8+ T 细胞含量无明显差异(P >0.05)。与对照组相比,各治疗组移植瘤内 Bax 阳性细胞数量明显增加(P <0.05),Bcl -2阳性细胞数量明显减少(P <0.05);与 CTL组和α-GalCer 组相比,联合治疗组移植瘤内 Bax 阳性细胞明显增加(P <0.05),Bcl -2阳性细胞明显下降(P <0.05)。结论:DC 荷载α-GalCer 与肿瘤特异性 CTL 联合应用能够对小鼠 Heps 肝癌移植瘤具有协同杀伤作用。%Objective:To investigate the inhibitory effects of dendritic cells(DC)load alpha -Galactosylceramide (α-GalCer)combined with tumor specific cytotoxic T lymphocyte(CTL)on the growth of

  15. Common antiviral cytotoxic t-lymphocyte epitope for diverse arenaviruses.

    Science.gov (United States)

    Oldstone, M B; Lewicki, H; Homann, D; Nguyen, C; Julien, S; Gairin, J E

    2001-07-01

    Members of the Arenaviridae family have been isolated from mammalian hosts in disparate geographic locations, leading to their grouping as Old World types (i.e., lymphocytic choriomeningitis virus [LCMV], Lassa fever virus [LFV], Mopeia virus, and Mobala virus) and New World types (i.e., Junin, Machupo, Tacaribe, and Sabia viruses) (C. J. Peters, M. J. Buchmeier, P. E. Rollin, and T. G. Ksiazek, p. 1521-1551, in B. N. Fields, D. M. Knipe, and P. M. Howley [ed.], Fields virology, 3rd ed., 1996; P. J. Southern, p. 1505-1519, in B. N. Fields, D. M. Knipe, and P. M. Howley [ed.], Fields virology, 3rd ed., 1996). Several types in both groups-LFV, Junin, Machupo, and Sabia viruses-cause severe and often lethal human diseases. By sequence comparison, we noted that eight Old World and New World arenaviruses share several amino acids with the nucleoprotein (NP) that consists of amino acids (aa) 118 to 126 (NP 118-126) (RPQASGVYM) of LCMV that comprise the immunodominant cytotoxic T-lymphocyte (CTL) epitope for H-2(d) mice (32). This L(d)-restricted epitope constituted >97% of the total bulk CTLs produced in the specific antiviral or clonal responses of H-2(d) BALB mice. NP 118-126 of the Old World arenaviruses LFV, Mopeia virus, and LCMV and the New World arenavirus Sabia virus bound at high affinity to L(d). The primary H-2(d) CTL anti-LCMV response as well as that of a CTL clone responsive to LCMV NP 118-126 recognized target cells coated with NP 118-126 peptides derived from LCMV, LFV, and Mopeia virus but not Sabia virus, indicating that a common functional NP epitope exists among Old World arenaviruses. Use of site-specific amino acid exchanges in the NP CTL epitope among these arenaviruses identified amino acids involved in major histocompatibility complex binding and CTL recognition.

  16. Monoclonal antibody against IFN-gamma inhibits Moloney murine sarcoma virus-specific cytotoxic T lymphocyte differentiation

    Energy Technology Data Exchange (ETDEWEB)

    Zanovello, P.; Vallerani, E.; Biasi, G.; Landolfo, S.; Collavo, D.

    1988-02-15

    The role of autochthonous IFN- production was evaluated in immune reactions to Moloney murine sarcoma virus (M-MSV)-induced tumors which are characterized by spontaneous regression mainly caused by virus-specific CTL activity. A functional IFN- depletion, induced by repeated administration of mAb anti-IFN- at the site of virus inoculation, prevented tumor regression in M-MSV-injected mice. Moreover, this antibody inhibited in vitro both proliferation and differentiation of M-MSV-specific T lymphocytes obtained in bulk cultures, but not growth and lytic activity of the already differentiated virus-specific CTL clone CHM-14 stimulated with rIL-2 and relevant tumor Ag. In addition, in mice receiving mAb treatment the frequency of M-MSV-specific CTL precursors, evaluated by means of limiting dilution analysis, was strongly reduced in comparison with that of control mice injected only with virus. Because CTL secrete IFN- following antigenic stimulation, the possibility that non-T effector cells recruited by this lymphokine might mediate tumor regression was also considered. Adoptive immunotherapy experiments, performed in T cell-deficient (Tx + BM) and in sublethally irradiated mice, demonstrated that transfer of CHM-14 CTL clone, which secretes IFN-, was able to counteract M-MSV tumor growth despite the local mAb anti-IFN- treatment which may have prevented host cell recruitment. Moreover, repeated local rIFN- inoculations in Tx + BM mice did not counteract M-MSV tumor progression, thus confirming that other IFN- properties such as non-T cell recruitment, antiviral or anti-proliferative IFN- activities have little or no relevance when M-MSV-specific CTL are lacking. On the whole, these results indicate that in M-MSV-injected mice, tumor enhancement after mAb anti-IFN- treatment is principally caused by impaired differentiation of virus-specific CTL precursors.

  17. A dynamical perspective of CTL cross-priming and regulation: implications for cancer immunology.

    Science.gov (United States)

    Wodarz, Dominik; Jansen, Vincent A A

    2003-05-01

    Cytotoxic T lymphocytes (CTL) responses are required to fight many diseases such as viral infections and tumors. At the same time, they can cause disease when induced inappropriately. Which factors regulate CTL and decide whether they should remain silent or react is open to debate. The phenomenon called cross-priming has received attention in this respect. That is, CTL expansion occurs if antigen is recognized on the surface of professional antigen presenting cells (APCs). This is in contrast to direct presentation where antigen is seen on the surface of the target cells (e.g. infected cells or tumor cells). Here we introduce a mathematical model, which takes the phenomenon of cross-priming into account. We propose a new mechanism of regulation which is implicit in the dynamics of the CTL: According to the model, the ability of a CTL response to become established depends on the ratio of cross-presentation to direct presentation of the antigen. If this ratio is relatively high, CTL responses are likely to become established. If this ratio is relatively low, tolerance is the likely outcome. The behavior of the model includes a parameter region where the outcome depends on the initial conditions. We discuss our results with respect to the idea of self/non-self discrimination and the danger signal hypothesis. We apply the model to study the role of CTL in cancer initiation, cancer evolution/progression, and therapeutic vaccination against cancers.

  18. Wnt signaling inhibits CTL memory programming.

    Science.gov (United States)

    Xiao, Zhengguo; Sun, Zhifeng; Smyth, Kendra; Li, Lei

    2013-12-01

    Induction of functional CTLs is one of the major goals for vaccine development and cancer therapy. Inflammatory cytokines are critical for memory CTL generation. Wnt signaling is important for CTL priming and memory formation, but its role in cytokine-driven memory CTL programming is unclear. We found that wnt signaling inhibited IL-12-driven CTL activation and memory programming. This impaired memory CTL programming was attributed to up-regulation of eomes and down-regulation of T-bet. Wnt signaling suppressed the mTOR pathway during CTL activation, which was different to its effects on other cell types. Interestingly, the impaired memory CTL programming by wnt was partially rescued by mTOR inhibitor rapamycin. In conclusion, we found that crosstalk between wnt and the IL-12 signaling inhibits T-bet and mTOR pathways and impairs memory programming which can be recovered in part by rapamycin. In addition, direct inhibition of wnt signaling during CTL activation does not affect CTL memory programming. Therefore, wnt signaling may serve as a new tool for CTL manipulation in autoimmune diseases and immune therapy for certain cancers.

  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. Definition of the region on NS3 which contains multiple epitopes recognized by dengue virus serotype-cross-reactive and flavivirus-cross-reactive, HLA-DPw2-restricted CD4+ T cell clones.

    Science.gov (United States)

    Okamoto, Y; Kurane, I; Leporati, A M; Ennis, F A

    1998-04-01

    The epitopes recognized by six CD4+ CD8- cytotoxic T lymphocyte (CTL) clones established from a dengue-3 virus-immune donor were defined. (i) Three CTL clones, JK10, JK34 and JK39, were cross-reactive for dengue virus types 1-4. (ii) One clone, JK28, was cross-reactive for dengue virus types 1-4 and West Nile virus. (iii) Two clones, JK26 and JK49, were cross-reactive for dengue virus types 1-4, West Nile virus and yellow fever virus. The clones, except for JK49, recognized the same epitope on NS3 in an HLA-DPw2-restricted fashion. The smallest synthetic peptide recognized by the five CTL clones was a 10 aa peptide which comprises aa 255-264 on dengue virus NS3. JK49 recognized the overlapping epitope which comprises aa 257-266 in an HLA-DPw2-restricted fashion. Analysis of T cell receptor (TCR) usage by these T cell clones revealed that (i) JK10 and JK34 use V alpha11, and JK34 and JK28 use V beta23, and (ii) the amino acid sequences of the V(D)J junctional region of the TCR were different among these five CTL clones. There were, however, single amino acid conservations among TCRs of some of these T cell clones. These results indicate that the region on NS3 which comprises aa 255-266 contains multiple epitopes recognized by dengue serotype-cross-reactive and flavivirus-cross-reactive CD4+ CTL in an HLA-DPw2-restricted fashion and that a single epitope can be recognized by T cells which have heterogeneous virus specificities.

  1. Bounded Model Checking of CTL

    Institute of Scientific and Technical Information of China (English)

    Zhi-Hong Tao; Cong-Hua Zhou; Zhong Chen; Li-Fu Wang

    2007-01-01

    Bounded Model Checking has been recently introduced as an efficient verification method for reactive systems.This technique reduces model checking of linear temporal logic to propositional satisfiability.In this paper we first present how quantified Boolean decision procedures can replace BDDs.We introduce a bounded model checking procedure for temporal logic CTL* which reduces model checking to the satisfiability of quantified Boolean formulas.Our new technique avoids the space blow up of BDDs, and extends the concept of bounded model checking.

  2. Cloning and Sequencing of Protein Kinase cDNA from Harbor Seal (Phoca vitulina Lymphocytes

    Directory of Open Access Journals (Sweden)

    Jennifer C. C. Neale

    2004-01-01

    Full Text Available Protein kinases (PKs play critical roles in signal transduction and activation of lymphocytes. The identification of PK genes provides a tool for understanding mechanisms of immunotoxic xenobiotics. As part of a larger study investigating persistent organic pollutants in the harbor seal and their possible immunomodulatory actions, we sequenced harbor seal cDNA fragments encoding PKs. The procedure, using degenerate primers based on conserved motifs of human protein tyrosine kinases (PTKs, successfully amplified nine phocid PK gene fragments with high homology to human and rodent orthologs. We identified eight PTKs and one dual (serine/threonine and tyrosine kinase. Among these were several PKs important in early signaling events through the B- and T-cell receptors (FYN, LYN, ITK and SYK and a MAP kinase involved in downstream signal transduction. V-FGR, RET and DDR2 were also expressed. Sequential activation of protein kinases ultimately induces gene transcription leading to the proliferation and differentiation of lymphocytes critical to adaptive immunity. PKs are potential targets of bioactive xenobiotics, including persistent organic pollutants of the marine environment; characterization of these molecules in the harbor seal provides a foundation for further research illuminating mechanisms of action of contaminants speculated to contribute to large-scale die-offs of marine mammals via immunosuppression.

  3. Differential requirements for CTL generation by novel immunostimulants: APC tropism, use of the TAP-independent processing pathway, and dependency on CD80/CD86 costimulation.

    NARCIS (Netherlands)

    Sheikh, NA; Attard, GS; Rooijen, van N.; Rajananthanan, P; Hariharan, K; Yang, YW; Morrow, WJ

    2003-01-01

    A major drawback of subunit vaccines is their inability to generate cytolytic T lymphocytes (CTL), a deficit attributed to segregation of the class I and class II antigen-processing pathways. We sought to understand processes involved in CTL induction by three proprietary adjuvants: Tomatine, PROVAX

  4. Cloning

    Science.gov (United States)

    ... copies of whole animals Therapeutic cloning, which creates embryonic stem cells. Researchers hope to use these cells to grow healthy tissue to replace injured or diseased tissues in the human body. NIH: National Human Genome Research Institute

  5. Frequency analysis of simian virus 40-specific cytotoxic T lymphocyte precursors in the high responder C57BL/6 mouse strain.

    Science.gov (United States)

    Jennings, S R; Fresa, K L; Lippe, P A; Milici, J E; Tevethia, S S

    1988-10-01

    Studies in this laboratory have shown that long term simian virus 40 (SV40)-specific cytolytic T lymphocyte (CTL) cultures established from the spleens of high responder C57BL/6 (B6; H-2b) mice exhibit a preference for the selection of H-2Db-restricted CTL clones. In this study, we have investigated the basis for this selection. Limiting dilution cultures were established using responder cells from the popliteal lymph nodes and the spleens of B6 mice immunized subcutaneously in the hind footpads or via the intraperitoneal route, respectively, with syngeneic SV40-transformed cells expressing a full length (1 to 708 amino acid residues) SV40 large T antigen. The relative frequency of CTL precursors (CTLp) able to expand in vitro in the presence of SV40-transformed stimulator cells and interleukin 2 and exhibit lytic activity against H-2b cells expressing full length T antigen ranged from 1/1900 to 1/15,000 in the popliteal lymph node and from 1/8000 to 1/55,000 in the spleen. In these two experimental systems, CTLp restricted to H-2Kb were apparently present at higher frequency than H-2Db-restricted CTLp. Furthermore, CTLp recognizing determinants within the amino-terminal or carboxy-terminal halves of T antigen were generated in approximately equal numbers. The relative affinity of SV40-specific CTL, assessed by inhibition with anti-Lyt 2 monoclonal antibody, indicated that CTL restricted to H-2Db interacted with their target with greater affinity than CTL restricted to H-2Kb. These data suggest that the predominance of isolation of H-2Db-restricted CTL clones from long term in vitro cultures may be a function of the relative affinity of this population as a whole, rather than due to the immunodominance of this subpopulation during the in vivo response to SV40 T antigen.

  6. In vitro priming of tumor-specific cytotoxic T lymphocytes using allogeneic dendritic cells derived from the human MUTZ-3 cell line.

    Science.gov (United States)

    Santegoets, Saskia J A M; Schreurs, Marco W J; Masterson, Allan J; Liu, Ying Poi; Goletz, Steffen; Baumeister, Hans; Kueter, Esther W M; Lougheed, Sinéad M; van den Eertwegh, Alfons J M; Scheper, Rik J; Hooijberg, Erik; de Gruijl, Tanja D

    2006-12-01

    The adoptive transfer of in vitro-induced and expanded tumor-specific cytotoxic T lymphocytes (CTL) presents a promising immunotherapeutic approach for the treatment of cancer. The in vitro induction of tumor-reactive CTL requires repeated stimulation of CTL precursors with dendritic cells (DC). To circumvent problems like scarcity of blood DC precursors and donor variability, it would be attractive to use DC from a non-autologous, unlimited source. DCs derived from the human acute myeloid leukemia (AML) cell line MUTZ-3 are attractive candidates since these DCs closely resemble monocyte-derived DC (MoDC) in terms of phenotype and T cell stimulatory capacity. Here we demonstrate that functional CTL clones could be generated against multiple tumor-associated antigens, i.e., human telomerase reverse transcriptase (hTERT), ErbB3-binding protein-1 (Ebp1), carcinoembryonic antigen (CEA) and Her-2/neu, by stimulating CD8beta(+) CTL precursors with peptide-loaded allogeneic, HLA-A2-matched MUTZ-3-derived DC. A consistent induction capacity, as determined by MHC tetramer-binding, was found in multiple donors and comparable to autologous peptide-loaded MoDC. Functional characterization at the clonal level revealed the priming of CTL that recognized endogenously processed epitopes on tumor cell lines in an HLA-A2-restricted fashion. Our data indicate that MUTZ-3-derived DC can be used as stimulator cells for in vitro priming and expansion of functional TAA-specific effector CTL. MUTZ-3-derived DCs thus represent a ready and standardized source of allogeneic DC to generate CTL for therapeutic adoptive transfer strategies.

  7. Persistent virus infection despite chronic cytotoxic T-lymphocyte activation in gamma interferon-deficient mice infected with lymphocytic choriomeningitis virus

    DEFF Research Database (Denmark)

    Bartholdy, C; Christensen, Jan Pravsgaard; Wodarz, D;

    2000-01-01

    ). While wild-type mice rapidly cleared the infection, IFN-gamma -/- mice became chronically infected. Virus persistence in the latter mice did not reflect failure to generate cytotoxic T-lymphocyte (CTL) effectors, as an unimpaired primary CTL response was observed. Furthermore, while ex vivo CTL activity...

  8. CTL Model Update for System Modifications

    CERN Document Server

    Ding, Yulin; Zhang, Yan; Zhang, Y; 10.1613/jair.2420

    2011-01-01

    Model checking is a promising technology, which has been applied for verification of many hardware and software systems. In this paper, we introduce the concept of model update towards the development of an automatic system modification tool that extends model checking functions. We define primitive update operations on the models of Computation Tree Logic (CTL) and formalize the principle of minimal change for CTL model update. These primitive update operations, together with the underlying minimal change principle, serve as the foundation for CTL model update. Essential semantic and computational characterizations are provided for our CTL model update approach. We then describe a formal algorithm that implements this approach. We also illustrate two case studies of CTL model updates for the well-known microwave oven example and the Andrew File System 1, from which we further propose a method to optimize the update results in complex system modifications.

  9. Plasmodium falciparum CS protein - prime malaria vaccine candidate: definition of the human CTL domain and analysis of its variation

    Directory of Open Access Journals (Sweden)

    Denise L. Doolan

    1992-01-01

    Full Text Available Studies in mice have shown that immunity to malaria sporozoites is mediated primarily by citotoxic T lymphocytes (CTL specific for epitopes within the circumsporozoite (CS protein. Humans, had never been shown to generate CTL against any malaria or other parasite protein. The design of a sub-unit vaccine for humans ralies on the epitopes recognized by CTL being identified and polymorphisms therein being defined. We have developed a novel technique using an entire series of overlapping synthetic peptides to define the epitopes of the Plasmodium falciparum CS protein recognized by human CTL and have analyzed the sequence variation of the protein with respect to the identified CTL epitopic domain. We have demonstrated that some humans can indeed generate CTL. against the P. falciparum CS protein. Furthermore, the extent of variation observed for the CTL recognition domain is finite and the combination of peptides necessary for inclusion in a polyvalent vaccine may be small. If ways can be found to increase immune responsiveness, then a vaccine designed to stimulate CS protein-specific CTL activity may prevent malaria.

  10. Antileukemic HLA-restricted T-cell clones generated with naturally processed peptides eluted from acute myeloblastic leukemia blasts.

    Science.gov (United States)

    Ostankovitch, M; Buzyn, A; Bonhomme, D; Connan, F; Bouscary, D; Heshmati, F; Dreyfus, F; Choppin, J; Guillet, J G

    1998-07-01

    Recent studies have shown that transfusions of HLA-compatible donor lymphocytes may induce complete remission in marrow-grafted patients with relapses of acute myeloblastic leukemia (AML). We investigated the in vitro generation of antileukemia T-cell clones obtained from the peripheral blood mononuclear cells of a partially HLA-compatible donor (HLA-A2 and B7 molecules in common with the leukemic blasts) after stimulation with a pool of naturally processed peptides extracted from leukemic blast cells collected at diagnosis from a patient with hyperleucocytosis AML. We recovered a significant quantity of peptides that bound to the HLA-A2 or HLA-B7 molecules that were able to induce cytolytic T-lymphocyte (CTL) lines and clones specific for the eluted AML peptides and restricted to the HLA-A2 or B7 molecules. Such CTL line did not recognize the patient's nonleukemic cells, and one clone was able to interact with the leukemic blasts from which the naturally processed peptides had been eluted. Such T-cell clones might provide a rationale for the development of adoptive immunotherapy and could be used to improve the efficiency of HLA-compatible T-lymphocyte transfusions and the graft-versus-leukemia response in patients with AML.

  11. Single-Cell Analysis and Next-Generation Immuno-Sequencing Show That Multiple Clones Persist in Patients with Chronic Lymphocytic Leukemia.

    Directory of Open Access Journals (Sweden)

    Jitra Kriangkum

    Full Text Available The immunoglobulin heavy chain (IGH gene rearrangement in chronic lymphocytic leukemia (CLL provides a unique molecular signature; however, we demonstrate that 26/198 CLL patients (13% had more than one IGH rearrangement, indicating the power of molecular technology over phenotypic analysis. Single-cell PCR analysis and next-generation immuno-sequencing identified IGH-defined clones. In 23% (18/79 of cases whose clones carried unmutated immunoglobulin heavy chain variable (IGHV genes (U-CLL, IGH rearrangements were bialleic with one productive (P and one non-productive (NP allele. Two U-CLL were biclonal, each clone being monoallelic (P. In 119 IGHV-mutated (M-CLL cases, one had biallelic rearrangements in their CLL (P/NP and five had 2-4 distinct clones. Allelic exclusion was maintained in all B-clones analyzed. Based on single-cell PCR analysis, 5/11 partner clones (45% reached levels of >5x10(9 cells/L, suggesting second CLL clones. Partner clones persisted over years. Conventional IGH characterization and next-generation sequencing of 13 CLL, 3 multiple myeloma, 2 Waldenstrom's macroglobulinemia and 3 age-matched healthy donors consistently identified the same rearranged IGH sequences. Most multiple clones occurred in M-CLL, perhaps indicative of weak clonal dominance, thereby associating with a good prognosis. In contrast, biallelic CLL occurred primarily in U-CLL thus being associated with poor prognosis. Extending beyond intra-clonal diversity, molecular analysis of clonal evolution and apparent subclones in CLL may also reflect inter-clonal diversity.

  12. Large-scale validation of methods for cytotoxic T-lymphocyte epitope prediction

    DEFF Research Database (Denmark)

    Larsen, Mette Voldby; Lundegaard, Claus; Lamberth, K.

    2007-01-01

    BACKGROUND: Reliable predictions of Cytotoxic T lymphocyte (CTL) epitopes are essential for rational vaccine design. Most importantly, they can minimize the experimental effort needed to identify epitopes. NetCTL is a web-based tool designed for predicting human CTL epitopes in any given protein....... of the other methods achieved a sensitivity of 0.64. The NetCTL-1.2 method is available at http://www.cbs.dtu.dk/services/NetCTL.All used datasets are available at http://www.cbs.dtu.dk/suppl/immunology/CTL-1.2.php....

  13. Large-scale validation of methods for cytotoxic T-lymphocyte epitope prediction

    DEFF Research Database (Denmark)

    Larsen, Mette Voldby; Lundegaard, Claus; Lamberth, K.;

    2007-01-01

    BACKGROUND: Reliable predictions of Cytotoxic T lymphocyte (CTL) epitopes are essential for rational vaccine design. Most importantly, they can minimize the experimental effort needed to identify epitopes. NetCTL is a web-based tool designed for predicting human CTL epitopes in any given protein....... of the other methods achieved a sensitivity of 0.64. The NetCTL-1.2 method is available at http://www.cbs.dtu.dk/services/NetCTL.All used datasets are available at http://www.cbs.dtu.dk/suppl/immunology/CTL-1.2.php....

  14. Islet-specific CTL cloned from a type 1 diabetes patient cause beta-cell destruction after engraftment into HLA-A2 transgenic NOD/scid/IL2RG null mice.

    Directory of Open Access Journals (Sweden)

    Wendy W J Unger

    Full Text Available Despite increasing evidence that autoreactive CD8 T-cells are involved in both the initiation of type 1 diabetes (T1D and the destruction of beta-cells, direct evidence for their destructive role in-vivo is lacking. To address a destructive role for autoreactive CD8 T-cells in human disease, we assessed the pathogenicity of a CD8 T-cell clone derived from a T1D donor and specific for an HLA-A2-restricted epitope of islet-specific glucose-6-phosphatase catalytic-subunit related protein (IGRP. HLA-A2/IGRP tetramer staining revealed a higher frequency of IGRP-specific CD8 T-cells in the peripheral blood of recent onset human individuals than of healthy donors. IGRP(265-273-specific CD8 T-cells that were cloned from the peripheral blood of a recent onset T1D individual were shown to secrete IFNγ and Granzyme B after antigen-specific activation and lyse HLA-A2-expressing murine islets in-vitro. Lytic capacity was also demonstrated in-vivo by specific killing of peptide-pulsed target cells. Using the HLA-A2 NOD-scid IL2rγ(null mouse model, HLA-A2-restricted IGRP-specific CD8 T-cells induced a destructive insulitis. Together, this is the first evidence that human HLA-restricted autoreactive CD8 T-cells target HLA-expressing beta-cells in-vivo, demonstrating the translational value of humanized mice to study mechanisms of disease and therapeutic intervention strategies.

  15. Identification and cloning of a prethymic precursor T lymphocyte from a population of common acute lymphoblastic leukemia antigen (CALLA)-positive fetal bone marrow cells

    DEFF Research Database (Denmark)

    Hokland, P; Hokland, M; Daley, J

    1987-01-01

    We have cloned common acute lymphoblastic leukemia (CALLA)-positive cells from human fetal bone marrow containing less than 1 in 10,000 E-RFC in round-bottomed microtiter wells (one cell per well) using the autocloning unit of an EPICS-V cell sorter. Expansion of such cells (with IL-2 and heavily...... irradiated autologous thymocytes as feeder cells) resulted in growth in 6-14% of the wells (mean, 11%) with cells with mature T lymphocyte phenotype. Two-color fluorescence analysis of outgrowing cultures furthermore ascertained that these cells had differentiated through a phase of simultaneous expression...... of T4 and T8 antigens and at the same time expression of the thymocyte-associated T6 antigens. Thus, given the fact that 10-20% of T cell acute lymphoblastic leukemia (T-ALLs) are CALLA+, we have been able to identify a human prethymic T lymphocyte population that might be the normal counterpart...

  16. The CD68 protein as a potential target for leukaemia-reactive CTL.

    Science.gov (United States)

    Sadovnikova, E; Parovichnikova, E N; Savchenko, V G; Zabotina, T; Stauss, H J

    2002-10-01

    CD68, a haematopoietic differentiation marker of the monocyte-macrophage lineage, is expressed in various human malignancies including chronic and acute myeloid leukaemia (AML). While the majority of normal CD34(+) cells are negative for CD68 expression, CD34(+) cells from AML patients produce elevated amounts of this protein. The purpose of this study was to identify CTL epitopes in the human CD68 protein. Mouse CD68 was also analysed to search for epitopes that could be used in murine tumor model. Peptides binding to murine H2(b) class I molecules were identified and used to stimulate CTL responses from allogeneic donor mice to avoid immunological tolerance. High avidity CTL clones specific for three different peptide epitopes did not kill CD68-expressing murine target cells, indicating that endogenous antigen processing failed to produce sufficient amounts of these peptides. In contrast, allo-restricted human CTL specific for an HLA-A2-binding peptide of CD68 recognised not only picomolar concentrations of peptide, but also displayed low levels of killing against HLA-A2-positive K562 and THP-1 leukemia cell lines and blast cells from AML patients. These data suggest that human leukaemia cells express limited amounts of CD68-derived peptides, and that high avidity CTL capable of recognising sub-picomolar concentrations of peptides are required for efficient killing of leukaemia cells.

  17. SARS CTL vaccine candidates; HLA supertype-, genome-wide scanning and biochemical validation

    DEFF Research Database (Denmark)

    Sylvester-Hvid, C.; Nielsen, Morten; Lamberth, K.

    2004-01-01

    An effective Severe Acute Respiratory Syndrome (SARS) vaccine is likely to include components that can induce specific cytotoxic T-lymphocyte (CTL) responses. The specificities of such responses are governed by human leukocyte antigen (HLA)-restricted presentation of SARS-derived peptide epitopes...... of the HLA supertypes and identified almost 100 potential vaccine candidates. These should be further validated in SARS survivors and used for vaccine formulation. We suggest that immunobioinformatics may become a fast and valuable tool in rational vaccine design....

  18. Most influenza A virus-specific memory cytotoxic T lymphocytes react with antigenic epitopes associated with internal virus determinants

    OpenAIRE

    1984-01-01

    This paper shows that most murine (C57BL/6) influenza A virus-specific memory cytotoxic T lymphocyte (CTL) clones tested in limiting dilution did not react with the influenza A virus surface glycoproteins, hemagglutinin (HA) and neuraminidase (NA). This lysis of syngeneic target cells infected with the influenza A virus strains, Aichi (H3N2), PR8 (H1N1), or recombinant strain X31 (H3N2) indicates that most antigenic epitopes recognized are associated with internal virus determinants. X31 and ...

  19. Recognition of clonogenic leukemic cells, remission bone marrow and HLA-identical donor bone marrow by CD8+ or CD4+ minor histocompatibility antigen-specific cytotoxic T lymphocytes.

    Science.gov (United States)

    Faber, L M; van der Hoeven, J; Goulmy, E; Hooftman-den Otter, A L; van Luxemburg-Heijs, S A; Willemze, R; Falkenburg, J H

    1995-08-01

    We investigated whether minor histocompatibility (mH) antigen-specific cytotoxic T lymphocytes (CTL) can discriminate between leukemic hematopoietic progenitor cells (leukemic-HPC) from AML or CML patients, the HPC from their remission bone marrow (remission-HPC), and normal HPC from their HLA-identical sibling bone marrow donor (donor-HPC). Specific lysis by CD8+ CTL clones was observed not only of the leukemic-HPC but also of the donor-HPC in 3/4 patient/donor combinations expressing mH antigen HA-1, 3/5 combinations expressing mH antigen HA-2, 2/3 combinations expressing mH antigen HA-3, and 2/2 combinations expressing mH antigen HY-A1. In four patient/donor combinations the recognition of the donor-HPC was clearly less than of the leukemic-HPC, indicating differential susceptibility to lysis by these mH CTL clones. In addition, differential recognition of leukemic-HPC and remission-HPC within seven patients was analyzed. In one patient expressing the HA-2 antigen on the leukemic cells the recognition of the remission-HPC was clearly less than of the leukemic-HPC. One CD4+ CTL clone showed specific lysis of the leukemic-HPC from an AML patient and a CML patient as well as of normal remission-HPC and donor-HPC. These results illustrate that in general CD8+ and CD4+ mH antigen specific CTL clones do not differentially recognize leukemic-HPC and normal-HPC. However, differences in susceptibility to lysis of malignant versus normal cells may contribute to a differential GVL effect.

  20. Single-channel and whole-cell recordings of mitogen-regulated inward currents in human cloned helper T lymphocytes.

    Science.gov (United States)

    Kuno, M; Goronzy, J; Weyand, C M; Gardner, P

    Cytoplasmic free Ca2+ [( Ca2+]i) appears to be an important signal for DNA synthesis in early stages of lymphocyte activation. In spite of many experimental studies which employ fluorescent Ca2+ indicator dye to demonstrate an early increase of [Ca2+]i in T-lymphocytes after stimulation with lectins, specific antigens, and monoclonal antibodies to T-lymphocyte receptors, the mechanism responsible for the rise of [Ca2+]i is unknown. We have used the extracellular patch clamp technique to investigate this mechanism. Unitary inward currents, mediated by Ca2+ or Ba2+, were recorded in the membrane of T-lymphocytes. The inward current channel was characterized by a conductance of 7 pS and extrapolated reversal potential (Erev) 110 mV positive to resting potential (Vr). While gating kinetic parameters were not affected by membrane potential changes, the probability of channel opening markedly increased upon activation of the T-lymphocyte by the mitogenic lectin, phytohaemagglutinin (PHA). PHA also evoked a cadmium-sensitive, inward Ba2+ current on whole-cell clamp. We suggest that this mitogen-regulated channel introduces Ca2+ into the cytoplasm upon activation and represents a new class of voltage-independent Ca2+ channels.

  1. [Towards an industrial control of the cloning of lymphocytes B human for the manufacturing of monoclonal antibodies stemming from the human repertoire].

    Science.gov (United States)

    Guillot-Chene, P; Lebecque, S; Rigal, D

    2009-05-01

    Monoclonal antibodies (mAbs) are efficient drugs for treating infectious, inflammatory and cancer diseases. Antibodies secreted by human lymphocytes that have been isolated from either peripheral blood or tissues present the definite interest of being part of the physiological or disease-related response to antigens present in the human body. However, attempts to generate hybridomas with human B cells have been largely unsuccessful, and cloning of human B cells has been achieved only via their inefficient immortalization with Epstein Barr Virus (EBV). However, recent progress in our understanding of the molecular mechanisms of polyclonal B cell activation has dramatically increased the capacity to clone human B cells. In particular, activation of human naïve and memory B cells through CD40 or memory B cells only through TLR9 was shown to greatly facilitate their immortalization by EBV. Industrial development based on these observations will soon provide large collections of high affinity human mAbs of every isotype directly selected by the human immune system directed to recognize epitopes relevant for individual patients. Moreover, after CD40 activation, these mAbs will cover the full human repertoire, including the natural auto-immune repertoire. Full characterization of the biological activity of these mAbs will in turn bring useful information for selecting vaccine epitopes. This breakthrough in human B cell cloning opens the way into new areas for therapeutic use of mAbs.

  2. CTL responses to Leishmania mexicana gp63-cDNA vaccine in a murine model.

    Science.gov (United States)

    Ali, S A; Rezvan, H; McArdle, S E; Khodadadi, A; Asteal, F A; Rees, R C

    2009-07-01

    Immunity to Leishmania is believed to be strongly dependent upon the activation of Th1 immune responses, although the exact role of cytotoxic T lymphocytes (CTLs) has not yet been determined. The aims of this study were to establish a suitable cytotoxicity assay to measure CTL activity and to compare immunity induced by Leishmania mexicana gp63 cDNA via i.m. injection and gene gun immunization in the BALB/c mouse model. The CTL activity was evaluated by short-term (51)Cr-release cytotoxicity assays against CT26 tumour cells transfected with L. mexicana gp63 cDNA and dendritic cells (DCs) loaded with soluble Leishmania antigen (SLA) as targets. The results clearly demonstrated that higher protection to L. mexicana infection was induced by gene gun DNA-immunization vs. i.m. injection. Cytotoxic T lymphocyte activity of splenocytes was observed in mice immunized either with L. mexicana gp63 cDNA or SLA and long-lived CTL activity was observed in immunized and/or re-challenged mice but not naïve mice infected with the parasite.

  3. Shell Delays CTL Venture with Shenhua

    Institute of Scientific and Technical Information of China (English)

    2009-01-01

    @@ Royal Dutch Shell has recently made a decision to postpone a coal-to-liquids (CTL) joint project with Shenhua Group in the Ningxia Hui Autonomous Region."In a period of economic downturn, we have postponed the project due to a number of reasons," Lira Haw Kuang,Executive Chairman of Shell Companies in China, told the news media in mid April without elaborating. But the company will keep playing an active role in China's coal gasification sector, Lim added.

  4. Triggering DTH and CTL Activity by fd Filamentous Bacteriophages: Role of CD4+ T Cells in Memory Responses

    Directory of Open Access Journals (Sweden)

    Giovanna Del Pozzo

    2010-01-01

    Full Text Available The ability of fd bacteriophage particles to trigger different arms of the immune system has been previously shown by us with particular emphasis on the ability of phages to raise CTL responses in vitro and in vivo. Here we show that fd virions in the absence of adjuvants are able to evoke a DTH reaction mediated by antigen specific CD8+ T cells. In addition, we analyzed the induction of CTL responses in mice depleted of CD4+ T cells, and we observed that short-term secondary CTL responses were induced in the absence of CD4+ T cells while induction of long-term memory CTLs required the presence of CD4+ T lymphocytes. These results examine the cellular mechanism at the basis of fd efficiency and provide new elements to further validate the use of fd particles for eliciting and monitoring antigen-specific CTLs.

  5. SARS CTL vaccine candidates; HLA supertype-, genome-wide scanning and biochemical validation

    DEFF Research Database (Denmark)

    Sylvester-Hvid, C; Nielsen, M; Lamberth, K;

    2004-01-01

    of the HLA supertypes and identified almost 100 potential vaccine candidates. These should be further validated in SARS survivors and used for vaccine formulation. We suggest that immunobioinformatics may become a fast and valuable tool in rational vaccine design.......An effective Severe Acute Respiratory Syndrome (SARS) vaccine is likely to include components that can induce specific cytotoxic T-lymphocyte (CTL) responses. The specificities of such responses are governed by human leukocyte antigen (HLA)-restricted presentation of SARS-derived peptide epitopes......-CoV) was isolated and full-length sequenced (Marra et al., Science 2003: 300: 1399-404). Here, we have combined advanced bioinformatics and high-throughput immunology to perform an HLA supertype-, genome-wide scan for SARS-specific CTL epitopes. The scan includes all nine human HLA supertypes in total covering >99...

  6. SARS CTL vaccine candidates; HLA supertype-, genome-wide scanning and biochemical validation

    DEFF Research Database (Denmark)

    Sylvester-Hvid, C.; Nielsen, Morten; Lamberth, K.;

    2004-01-01

    of the HLA supertypes and identified almost 100 potential vaccine candidates. These should be further validated in SARS survivors and used for vaccine formulation. We suggest that immunobioinformatics may become a fast and valuable tool in rational vaccine design.......An effective Severe Acute Respiratory Syndrome (SARS) vaccine is likely to include components that can induce specific cytotoxic T-lymphocyte (CTL) responses. The specificities of such responses are governed by human leukocyte antigen (HLA)-restricted presentation of SARS-derived peptide epitopes......-CoV) was isolated and full-length sequenced (Marra et al., Science 2003: 300: 1399404). Here, we have combined advanced bioinformatics and high-throughput immunology to perform an HLA supertype-, genome-wide scan for SARS-specific CTL epitopes. The scan includes all nine human HLA supertypes in total covering >99...

  7. Therapeutic polypeptides based on HBV core 18-27 epitope can induce CD8+ CTL-mediated cytotoxicity in HLA-A2+ human PBMCs

    Institute of Scientific and Technical Information of China (English)

    Tong-Dong Shi; Yu-Zhang Wu; Zheng-Cai Jia; Li-Yun Zou; Wei Zhou

    2004-01-01

    AIM: To explore how to improve the immunogenicity of HBcAg CTL epitope based polypeptides and to trigger an HBV-specific HLA I-restricted CD8+ T cell response in vitro.METHODS: A new panel of mimetic therapeutic peptides based on the immunodominant B cell epitope of HBV PreS218-24 region, the CTL epitope of HBcAg18-27 and the universal T helper epitope of tetanus toxoid (TT) 830-843was designed using computerized molecular design method and synthesized by Merrifield's solid-phase peptide synthesis.Their immunological properties of stimulating activation and proliferation of lymphocytes, of inducing TH1 polarization,CD8+ T cell magnification and HBV-specific CD8+ CTL mediated cytotoxicity were investigatedin vitro using HLAA2+ human peripheral blood mononuclear cells (PBMCs) from healthy donors and chronic hepatitis B patients.RESULTS: Results demonstrated that the therapeutic polypeptides based on immunodominant HBcAg18-27 CTL,PreS2 B- and universal TH epitopes could stimulate the activation and proliferation of lymphocytes, induce specifically and effectively CD8+ T cell expansion and vigorous HBVspecific CTL-mediated cytotoxicity in human PBMCs.CONCLUSION: It indicated that the introduction of immunodominant T helper plus B-epitopes with short and flexible linkers could dramatically improve the immunogenicity of short CTL epitopes in vitro.

  8. Combined Transfection with EBV-Specific Epitopes and HLA-A2 genes is More Effective than Separate Transfection in Promoting CTL Lysis against Nasopharyngeal Carcinoma

    Institute of Scientific and Technical Information of China (English)

    Weijun Ding; Choylen Fong

    2004-01-01

    To augment specific cytotoxic T lymphocyte (CTL) lysis is a promising strategy for cancer therapy. In this study,we examined the boosting effect of CTLs upon autologous lymphoblastoid B cell lines (LCLs) transfected with diverse plasmids, to explore the possible CTL-based immunotherapy of nasopharyngeal carcinoma (NPC).FCM analysis displayed rather high ratio (>30%) of successfully transfected LCLs by utilizing the DMRIE-C kit. CTL assays demonstrated that substantially higher ratio of CTL specific lysis was observed upon the LCLs transfected with both expression vectors encoding EBV-specific epitopes and their presentation molecule HLA-A2, in contrast with those transfected separately. By transfecting the vector encoding HLA-A2 alone, only the LCLs of HLA-A2+ donors elicited markedly higher CTL lysis. CTL assays also showed that there existed no marked differences upon transfection by either different vectors (pcDNA3, pNGVL3 or pNGVL3-hFlex), or different EBV-derived peptides (LMP2Pep1 or LMP2Pep2), or with or without the doubled DNA sequence encoding peptides. This study indicated a promising immunotherapy strategy on NPC through boosting and eliciting the EBV-specific CTL activation by transferring vectors encoding both EBV-specific epitopes and their presentation molecule HLA-A2 into autologous LCL, the presentation cells of MHC/peptide tetrameric complex.

  9. Combined Transfection with EBV-Specific Epitopes and HLA-A2 genes is More Effective than Separate Transfection in Promoting CTL Lysis against Nasopharyngeal Carcinoma

    Institute of Scientific and Technical Information of China (English)

    WeijunDing; ChoylenFong

    2004-01-01

    To augment specific cytotoxic T lymphocyte (CTL) lysis is a promising strategy for cancer therapy. In this study,we examined the boosting effect of CTLs upon autologous lymphoblastoid B cell lines (LCLs) transfected with diverse plasmids, to explore the possible CTL-based immunotherapy of nasopharyngeal carcinoma (NPC).FCM analysis displayed rather high ratio (>30%) of successfully transfected LCLs by utilizing the DMRIE-C kit. CTL assays demonstrated that substantially higher ratio of CTL specific lysis was observed upon the LCLs transfected with both expression vectors encoding EBV-specific epitopes and their presentation molecule HLA-A2, In contrast with those transfected separately. By transfecting the vector encoding HLA-A2 alone, only the LCLs of HLA-A2+ donors elicited markedly higher CTL lysis. CTL assays also showed that there existed no marked differences upon transfection by either different vectors (pcDNA3, pNGVL3 or pNGVL3-hFIex), or different EBV-derived peptides (LMP2Pep1 or LMP2Pep2), or with or without the doubled DNA sequence encoding peptides. This study indicated a promising immunotherapy strategy on NPC through boosting and eliciting the EBV-specific CTL activation by transferring vectors encoding both EBV-specific epitopes and their presentation molecule HLA-A2 into autologous LCL, the presentation cells of MHC/peptide tetrameric complex.

  10. Chimeric virus-like particles for the delivery of an inserted conserved influenza A-specific CTL epitope.

    Science.gov (United States)

    Cheong, Wan-Shoo; Reiseger, Jessica; Turner, Stephen John; Boyd, Richard; Netter, Hans-Jürgen

    2009-02-01

    The small hepatitis B virus surface antigens (HBsAg-S) have the ability to self-assemble with host-derived lipids into empty non-infectious virus-like particles (VLPs). HBsAg-S VLPs are the sole component of the licensed hepatitis B vaccine, and they are a useful delivery platform for foreign epitopes. To develop VLPs capable of transporting foreign cytotoxic T lymphocyte (CTL) epitopes, HBsAg-S specific CTL epitopes at various sites were substituted with a conserved CTL epitope derived from the influenza matrix protein. Depending on the insertion site, the introduction of the MHC class I A2.1-restricted influenza epitope was compatible with the secretion competence of HBsAg-S indicating that chimeric VLPs were assembled. Immunizations of transgenic HHDII mice with chimeric VLPs induced anti-influenza CTL responses proving that the inserted foreign epitope can be correctly processed and cross-presented. Chimeric VLPs in the absence of adjuvant were able to induce memory T cell responses, which could be recalled by influenza virus infections in the mouse model system. The ability of chimeric HBsAg-S VLPs to induce anti-foreign CTL responses and also with the proven ability to induce humoral immune responses constitute a highly versatile platform for the delivery of selected multiple epitopes to target disease associated infectious agents.

  11. A comparative study on efficiency of adult fibroblast, putative embryonic stem cell and lymphocyte as donor cells for production of handmade cloned embryos in goat and characterization of putative ntES cells obtained from these embryos.

    Science.gov (United States)

    Dutta, Rahul; Malakar, Dhruba; Khate, Keviletsu; Sahu, Shailendra; Akshey, Yogesh; Mukesh, Manishi

    2011-09-15

    The main purpose of the experiment was to compare the efficiency of three cell types, namely adult fibroblast, putative embryonic stem (ES) cell, and lymphocyte, as donor cells for somatic cell nuclear transfer by handmade cloning in goats. The outcome clearly shows that putative embryonic stem cells, with a cleavage and blastocyst production rate of 74.69% ± 3.92 and 39.75% ± 3.86, respectively, performs better in comparison to adult fibroblast cell and lymphocyte. Between adult fibroblast cell and lymphocyte no statistically significant difference exists at P II DRB genes of cloned embryos and three donor cells were performed to verify the cloned embryos. The amplified PCR products were subjected to SSCP to confirm their genetic identity. The karyotyping of the cloned embryos showed normal chromosomal status as expected in goat. Significantly, in the second stage of the experiment, the produced cloned embryos were successfully used to derive ntES-like cells. The rate of primary colony formation rate was 62.50% ± 4.62 for fibroblast donor cell derived embryos. The same was 60.60% ± 4.62 for putative ES donor cell derived embryos and 66.66% ± 4.62 for lymphocyte donor cell derived embryos, respectively. The putative ntES colonies were positively characterized for alkaline phosphatase, Oct-4, TRA-1-60, TRA-1-81, Sox-2, and Nanog by Immunocytochemistry and Reverse Transcription PCR. To further validate the stem ness, the produced putative ntES colonies were differentiated to embryoid bodies. Immunocytochemistry revealed that embryoid bodies expressed NESTIN specific for ectodermal lineage; GATA-4 for endodermal lineage and smooth muscle actin-I, and troponin-I specific for mesodermal lineage. The study has established an efficient protocol for putative ntES cell derivation from HMC embryos. It could be of substantial significance as patient specific ntES cells have proven therapeutic significance.

  12. A three-dimensional organotypic assay to measure target cell killing by cytotoxic T lymphocytes.

    NARCIS (Netherlands)

    Weigelin, B.; Friedl, P.H.A.

    2010-01-01

    Cytotoxic T lymphocytes (CTL) mediate antigen- and cell-cell contact dependent killing of target cells, such as cancer cells and virus-infected cells. In vivo, this process requires the active migration of CTL towards and away from target cells. We here describe an organotypic 3D collagen matrix

  13. The CD11a partner in Sus scrofa lymphocyte function-associated antigen-1 (LFA-1: mRNA cloning, structure analysis and comparison with mammalian homologues

    Directory of Open Access Journals (Sweden)

    Thomas Anne VT

    2005-10-01

    Full Text Available Abstract Background Lymphocyte function-associated antigen-1 (LFA-1, CD11a/CD18, alphaLbeta2, the most abundant and widely expressed beta2-integrin, is required for many cellular adhesive interactions during the immune response. Many studies have shown that LFA-1 is centrally involved in the pathogenesis of several diseases caused by Repeats-in-toxin (RTX -producing bacteria. Results The porcine-LFA-1 CD11a (alpha subunit coding sequence was cloned, sequenced and compared with the available mammalian homologues in this study. Despite some focal differences, it shares all the main characteristics of these latter. Interestingly, as in sheep and humans, an allelic variant with a triplet insertion resulting in an additional Gln-744 was consistently identified, which suggests an allelic polymorphism that might be biologically relevant. Conclusion Together with the pig CD18-encoding cDNA, which has been available for a long time, the sequence data provided here will allow the successful expression of porcine CD11a, thus giving the first opportunity to express the Sus scrofa beta2-integrin LFA-1 in vitro as a tool to examine the specificities of inflammation in the porcine species.

  14. Therapeutic potential of CD8+ cytotoxic T lymphocytes in SLE☆

    Science.gov (United States)

    Puliaeva, I.; Puliaev, R.; Via, C.S.

    2011-01-01

    Recent evidence supports the idea that following a break in tolerance, CD8 cytotoxic T lymphocytes (CTL) may be an important but unrecognized mechanism for limiting expansion of autoreactive B cells. Failure of this mechanism could allow persistence of CD4 T cell driven polyclonal B cell activation resulting in clinical lupus. Although CD8 CTL failure may occur early in disease, work in mice supports the concept that therapeutic CTL enhancement may be both practical and beneficial in lupus. Devising such therapy for humans will first require an understanding of the in vivo mechanisms critical in CTL expansion and down regulation, particularly in the lupus setting which may differ from CTL generation in other clinical settings (e.g. tumors, infections). PMID:18725326

  15. CADM1/TSLC1 Identifies HTLV-1-Infected Cells and Determines Their Susceptibility to CTL-Mediated Lysis.

    Directory of Open Access Journals (Sweden)

    Kiruthika Manivannan

    2016-04-01

    Full Text Available Human T cell lymphotropic virus-1 (HTLV-1 primarily infects CD4+ T cells, causing inflammatory disorders or a T cell malignancy in 5% to 10% of carriers. The cytotoxic T lymphocyte (CTL response is a key factor that controls the viral load and thus the risk of disease. The ability to detect the viral protein Tax in primary cells has made it possible to estimate the rate at which Tax-expressing infected cells are eliminated by CTLs in persistently infected people. However, most HTLV-1-infected cells are Tax-at a given time, and their immunophenotype is poorly defined. Here, we aimed to identify a cell-surface molecule expressed by both Tax+ and Tax-HTLV-1-infected cells and use it to analyse the CTL response in fresh peripheral blood mononuclear cells. Cell adhesion molecule 1 (CADM1/TSLC1 was the best single marker of HTLV-1 infection, identifying HTLV-1-infected cells with greater sensitivity and specificity than CD25, CCR4 or ICAM-1. CADM1+CD4+ T cells carried a median of 65% of proviral copies in peripheral blood. In a cohort of 23 individuals, we quantified the rate of CTL-mediated killing of Tax+ and Tax-CADM1+ cells. We show that CADM1 expression is associated with enhanced susceptibility of infected cells to CTL lysis: despite the immunodominance of Tax in the CTL response, Tax+CADM1- cells were inefficiently lysed by CTLs. Upregulation of the CADM1 ligand CRTAM on CD8+ T cells correlated with efficient lysis of infected cells. Tax-CADM1+ cells were lysed at a very low rate by autologous CTLs, however, were efficiently killed when loaded with exogenous peptide antigen. High expression of CADM1 on most HTLV-1-infected cells in the face of enhanced CTL counterselection implies that CADM1 confers a strong benefit on the virus.

  16. Functional expression of choline transporter like-protein 1 (CTL1) and CTL2 in human brain microvascular endothelial cells.

    Science.gov (United States)

    Iwao, Beniko; Yara, Miki; Hara, Naomi; Kawai, Yuiko; Yamanaka, Tsuyoshi; Nishihara, Hiroshi; Inoue, Takeshi; Inazu, Masato

    2016-02-01

    In this study, we examined the molecular and functional characterization of choline transporter in human brain microvascular endothelial cells (hBMECs). Choline uptake into hBMECs was a saturable process that was mediated by a Na(+)-independent, membrane potential and pH-dependent transport system. The cells have two different [(3)H]choline transport systems with Km values of 35.0 ± 4.9 μM and 54.1 ± 8.1 μM, respectively. Choline uptake was inhibited by choline, acetylcholine (ACh) and the choline analog hemicholinium-3 (HC-3). Various organic cations also interacted with the choline transport system. Choline transporter-like protein 1 (CTL1) and CTL2 mRNA were highly expressed, while mRNA for high-affinity choline transporter 1 (CHT1) and organic cation transporters (OCTs) were not expressed in hBMECs. CTL1 and CTL2 proteins were localized to brain microvascular endothelial cells in human brain cortical sections. Both CTL1 and CTL2 proteins were expressed on the plasma membrane and mitochondria. CTL1 and CTL2 proteins are mainly expressed in plasma membrane and mitochondria, respectively. We conclude that choline is mainly transported via an intermediate-affinity choline transport system, CTL1 and CTL2, in hBMECs. These transporters are responsible for the uptake of extracellular choline and organic cations. CTL2 participate in choline transport mainly in mitochondria, and may be the major site for the control of choline oxidation. Copyright © 2015 Elsevier Ltd. All rights reserved.

  17. Distributed Maximality based CTL Model Checking

    Directory of Open Access Journals (Sweden)

    Djamel Eddine Saidouni

    2010-05-01

    Full Text Available In this paper we investigate an approach to perform a distributed CTL Model checker algorithm on a network of workstations using Kleen three value logic, the state spaces is partitioned among the network nodes, We represent the incomplete state spaces as a Maximality labeled Transition System MLTS which are able to express true concurrency. we execute in parallel the same algorithm in each node, for a certain property on an incomplete MLTS , this last compute the set of states which satisfy or which if they fail are assigned the value .The third value mean unknown whether true or false because the partial state space lacks sufficient information needed for a precise answer concerning the complete state space .To solve this problem each node exchange the information needed to conclude the result about the complete state space. The experimental version of the algorithm is currently being implemented using the functional programming language Erlang.

  18. Construction and immunogenicity prediction of Plasmodium falciparum CTL epitope minigene vaccine

    Institute of Scientific and Technical Information of China (English)

    TANG; Yuyang

    2001-01-01

    [1]Hill, A. V., Elvin, J., Willis, A. C. et al., Molecular analysis of the association of HLA-B53 and resistance to severe malaria, Nature, 1992, 360: 434.[2]Perlmann, P., Miller, L., Fogarty/WHO international conference on cellular mechanisms in malaria immunity, Immun. Letter, 1990, 25: 1.[3]Perkus, M. E., Tartaglia, J., Paoletti, E. et al., Poxvirus-based vaccine candidates for cancer, AIDS and other infectious diseases, J. Leukocyte Biol., 1995, 58(1): 1.[4]Shen, H., Slifka, M. K., Matloubian, M. et al., Recombinant Listeria monocytogenes as a live vaccine vehicle for the induction of protective anti-viral cell-mediated immunity, Proc. Natl. Acad. Sci. USA, 1995, 92: 3987.[5]Waine, G. J., McManus, D. P., Nucleic acids: vaccines of the future, Parasitol Today, 1995, 11: 113.[6]Whitton, J. L., Sheng, N., Oldstone, M. B. A. et al., A "string-of beads" vaccine, comprising linked minigenes, confers protection from lethal-dose virus challenge, J. Virol., 1993, 67: 348.[7]Lalvani, A., Aidoo, M., Allsopp, C. E. et al., An-HLA-based approach to design of a CTL-inducing vaccine against Plasmodium falciparum, Res. Immunol., 1994, 145: 461.[8]Sidney, J., Grey, H. M., Kubo, R. T. et al., Practical, biochemical and evolutionary implications of the discovery of HLA class I supermotifs, Immunology Today, 1996, 17: 261.[9]Thomson, S. A., Elliott, S. L., Sherritt, M. A. et al., Recombinant polyepitope vaccines for the delivery of multiple CD8 cytotoxic T cell epitopes, J. Immun., 1996, 157: 822.[10] Hanke, T., Schneider, J., Gilbert, S. C. et al., DNA multi-CTL epitope vaccines for HIV and Plasmodium falciparum: immunogenicity in mice, Vaccine, 1998, 16: 426.[11] Townsend, A. R. M., Rothbard, J., Gotch, F. M. et al., The epitopes of influenza nucleoprotein recognized by cytotoxic T lymphocytes can be defined with short synthetic peptides, Cell, 1986, 44: 959.[12] Ojcius, D. M., Abastado, J. P., Casrouge, A. et al., Dissociation of

  19. A new theory of cytotoxic T-lymphocyte memory: implications for HIV treatment

    DEFF Research Database (Denmark)

    Wodarz, D; Page, K M; Arnaout, R A

    2000-01-01

    reinfection is only effective in a restricted set of circumstances, we find that resolution of the primary infection requires persistence of CTL precursors (GTLp), as well as a fast rate of activation of the CTLp. Since these are commonly the defining characteristics of CTL memory, we propose that CTL memory...... may have evolved in order to clear the virus during primary challenge. We show experimental data from lymphocytic choriomeningitis virus infection in mice, supporting our theory on CTL memory. We adapt our models to HIV and find that immune impairment during the primary phase of the infection may...

  20. Preferential CTL targeting of Gag is associated with relative viral control in long-term surviving HIV-1 infected former plasma donors from China

    Institute of Scientific and Technical Information of China (English)

    Mingming Jia; Quanbi Zhao; Dan Li; Hong Peng; Marcus Altfeld; Bruce D Walker; Xu G Yu; Yiming Shao; Kunxue Hong; Jianping Chen; Yuhua Ruan; Zhe Wang; Bing Su; Guoliang Ren; Xiaoqing Zhang; Zhen Liu

    2012-01-01

    It is generally believed that CD8+ cytotoxic T lymphocytes (CTLs) play a critical role in limiting the replication of human immunodeficiency virus type 1 (HIV-1) and in determining the outcome of the infection,and this effect may partly depend on which HIV product is preferentially targeted.To address the correlation between HIV-1-specific CTL responses and virus replication in a cohort of former plasma donors (FPDs),143 antiretroviral therapy naive FPDs infected with HIV-1 clade B' strains were assessed for HIV-1-specific CTL responses with an IFN-γElispot assay at single peptide level by using overlapping peptides (OLPs) covering the whole consensus clade B proteome.By using a Spearman's rank correlation analysis,we found that the proportion of Gag-specific CTL responses among the total virus-specific CTL activity was inversely correlated with viral loads while being positively correlated to CD4 counts,as opposed to Pol- and Env-specific responses that were associated with increased viral loads and decreased CD4 counts,In addition,Vpr-specifc CTL responses showed a similar protective effect with Gag responses,but with a much lower frequency of recognition.Significantly,we also observed an association between HLA-A*30/B*13/Cw*06 haplotype and lower viral loads that was probably due to restricted Gag-specific CTL responses.Thus,our data demonstrate the prominent role of Gag-specific CTL responses in disease control.The advantage of HLA-A*30/B*13/Cw*06 haplotype in viral control may be associated with the contribution of Gag-specific CTL responses in the studied individuals.

  1. Direct Microbicidal Activity of Cytotoxic T-Lymphocytes

    Directory of Open Access Journals (Sweden)

    Paul Oykhman

    2010-01-01

    Full Text Available Cytotoxic T-lymphocytes (CTL are famous for their ability to kill tumor, allogeneic and virus-infected cells. However, an emerging literature has now demonstrated that CTL also possess the ability to directly recognize and kill bacteria, parasites, and fungi. Here, we review past and recent findings demonstrating the direct microbicidal activity of both CD4+ and CD8+ CTL against various microbial pathogens. Further, this review will outline what is known regarding the mechanisms of direct killing and their underlying signalling pathways.

  2. Modélisation aléatoire de l'activité des Lymphocytes T Cytotoxyques

    OpenAIRE

    Christophe, Claire

    2014-01-01

    This thesis studies some statistical and probability properties of the dynamic of Cytotoxic T Lymphocyte(CTL, immune cells) and a tumor nodule. This work is in close collaboration with SalvatoreValitutti team at INSERM. Considering the complexity of developing an in vitro tumor nodule, wedevelop an agent based model to describe and understand the interaction between CTL and tumor.Numerical experiments highlight two important parameters in CTL immune response against tumormass. Primarily the C...

  3. Consequences of cytotoxic T lymphocyte interaction with major histocompatibility complex class I-expressing neurons in vivo

    OpenAIRE

    1995-01-01

    Neurons have evolved strategies to evade immune surveillance that include an inability to synthesize the heavy chain of the class I major histocompatibility complex (MHC), proteins that are necessary for cytotoxic T lymphocyte (CTL) recognition of target cells. Multiple viruses have taken advantage of the lack of CTL-mediated recognition and killing of neurons by establishing persistent neuronal infections and thereby escaping attack by antiviral CTL. We have expressed a class I MHC molecule ...

  4. Specific cytotoxic T lymphocyte responses in Chinese HIV/AIDS patients

    Institute of Scientific and Technical Information of China (English)

    尚红; 韩晓旭; 王亚男; 周立平; 张子宁; 姜拥军; 张旻; 于旭

    2004-01-01

    Human immunodeficiency virus type 1 (HIV-1)-specific cytotoxic T lymphocytes (CTL) are considered to play a central role in the immune response against HIV-1. During untreated acute HIV-1 infection, virus-specific CTL activity is associated with the initial decrease in viremia.1 After HIV infection, those individuals with a slow progressive course develop stronger CTL responses than those with typical disease progression.2,3 Restoring HIV-1-specific CTL responses have been considered a key factor in immune reconstitution and vaccine development. Here we present the analysis of HIV-1 Gag-specific CTL responses in 23 Chinese HIV/AIDS cases in order to take the initial steps at identifying the epitopes that dominate the CTL response in Chinese patients.

  5. Does programmed CTL proliferation optimize virus control?

    DEFF Research Database (Denmark)

    Wodarz, Dominik; Thomsen, Allan Randrup

    2005-01-01

    CD8 T-cell or cytotoxic T-lymphocyte responses develop through an antigen-independent proliferation and differentiation program. This is in contrast to the previous thinking, which was that continuous antigenic stimulation was required. This Opinion discusses why nature has chosen the proliferati...

  6. Specific induction of anti-leukemia effects by umbilical cord cell-derived CD8~+ T cytotoxic lymphocytes

    Institute of Scientific and Technical Information of China (English)

    刘芯

    2006-01-01

    Objective To explore the specific anti-leukemia immune response of CD8+ cytotoxic T lymphocyte (CTL) derived from cord blood (CB) ex vivo and evaluate the feasibilities and values of the CTL for specific immunotherapy. Methods Dendritic cells (DC) were induced from mononuclear cells (MNC) by combination cytokines in 10 CB samples. Loading U937 cell lysate antigen on

  7. CTL escape mediated by proteasomal destruction of an HIV-1 cryptic epitope.

    Directory of Open Access Journals (Sweden)

    Sylvain Cardinaud

    2011-05-01

    Full Text Available Cytotoxic CD8+ T cells (CTLs play a critical role in controlling viral infections. HIV-infected individuals develop CTL responses against epitopes derived from viral proteins, but also against cryptic epitopes encoded by viral alternative reading frames (ARF. We studied here the mechanisms of HIV-1 escape from CTLs targeting one such cryptic epitope, Q9VF, encoded by an HIVgag ARF and presented by HLA-B*07. Using PBMCs of HIV-infected patients, we first cloned and sequenced proviral DNA encoding for Q9VF. We identified several polymorphisms with a minority of proviruses encoding at position 5 an aspartic acid (Q9VF/5D and a majority encoding an asparagine (Q9VF/5N. We compared the prevalence of each variant in PBMCs of HLA-B*07+ and HLA-B*07- patients. Proviruses encoding Q9VF/5D were significantly less represented in HLA-B*07+ than in HLA-B*07- patients, suggesting that Q9FV/5D encoding viruses might be under selective pressure in HLA-B*07+ individuals. We thus analyzed ex vivo CTL responses directed against Q9VF/5D and Q9VF/5N. Around 16% of HLA-B*07+ patients exhibited CTL responses targeting Q9VF epitopes. The frequency and the magnitude of CTL responses induced with Q9VF/5D or Q9VF/5N peptides were almost equal indicating a possible cross-reactivity of the same CTLs on the two peptides. We then dissected the cellular mechanisms involved in the presentation of Q9VF variants. As expected, cells infected with HIV strains encoding for Q9VF/5D were recognized by Q9VF/5D-specific CTLs. In contrast, Q9VF/5N-encoding strains were neither recognized by Q9VF/5N- nor by Q9VF/5D-specific CTLs. Using in vitro proteasomal digestions and MS/MS analysis, we demonstrate that the 5N variation introduces a strong proteasomal cleavage site within the epitope, leading to a dramatic reduction of Q9VF epitope production. Our results strongly suggest that HIV-1 escapes CTL surveillance by introducing mutations leading to HIV ARF-epitope destruction by proteasomes.

  8. Figuring the Context of CTL under 2013 Curiculum

    Directory of Open Access Journals (Sweden)

    Chairina Nasir

    2017-06-01

    Full Text Available The 2013 curriculum states that the purpose of teaching English for junior high school is to develop students’ communicative competence. In line with this expectation, several learning characteristics have been defined i.e. learning from model, observing, questioning, gathering information, associating, and communicating. Therefore, the teaching approaches that are used by the teacher in teaching English must suit the criteria to promote students' communicative competence. Contextual Teaching and learning (CTL seems to be compatible as an approach since it has the the characteristics of constructivism, questioning, inquiry, learning community, modelling, reflection, and authentic assessment, which are similar to the learning characteristics mentioned above, which are similar to the learning characteristics mentioned above. , which are similar to the learning characteristics mentioned above. Therefore, a qualitative research concerning the issue was conducted to see how CTL approach is implemented under the 2013 curriculum in teaching reading comprehension. From the result of observation, questionnaire, and interview as the instruments, it was found that CTL was implemented properly from phase to phase and is applicable to be implemented under the curriculum. Also, it promotes active and enjoyable learning, facilitates the students to comprehend the material and helps them to implement the knowledge in real life. The. The teacher had implemented all of the procedures of CTL under the instruction of the 2013 curriculum. Thus, applying the CTL CTL approach in the process of teaching for the 2013 curriculum for the 2013 curriculum is recommended since it gives satisfactory benefits for students.

  9. An Approach for a Synthetic CTL Vaccine Design against Zika Flavivirus Using Class I and Class II Epitopes Identified by Computer Modeling

    Directory of Open Access Journals (Sweden)

    Edecio Cunha-Neto

    2017-06-01

    Full Text Available The threat posed by severe congenital abnormalities related to Zika virus (ZKV infection during pregnancy has turned development of a ZKV vaccine into an emergency. Recent work suggests that the cytotoxic T lymphocyte (CTL response to infection is an important defense mechanism in response to ZKV. Here, we develop the rationale and strategy for a new approach to developing cytotoxic T lymphocyte (CTL vaccines for ZKV flavivirus infection. The proposed approach is based on recent studies using a protein structure computer model for HIV epitope selection designed to select epitopes for CTL attack optimized for viruses that exhibit antigenic drift. Because naturally processed and presented human ZKV T cell epitopes have not yet been described, we identified predicted class I peptide sequences on ZKV matching previously identified DNV (Dengue class I epitopes and by using a Major Histocompatibility Complex (MHC binding prediction tool. A subset of those met the criteria for optimal CD8+ attack based on physical chemistry parameters determined by analysis of the ZKV protein structure encoded in open source Protein Data File (PDB format files. We also identified candidate ZKV epitopes predicted to bind promiscuously to multiple HLA class II molecules that could provide help to the CTL responses. This work suggests that a CTL vaccine for ZKV may be possible even if ZKV exhibits significant antigenic drift. We have previously described a microsphere-based CTL vaccine platform capable of eliciting an immune response for class I epitopes in mice and are currently working toward in vivo testing of class I and class II epitope delivery directed against ZKV epitopes using the same microsphere-based vaccine.

  10. Antigenic drift in the influenza A virus (H3N2) nucleoprotein and escape from recognition by cytotoxic T lymphocytes

    NARCIS (Netherlands)

    T.M. Bestebroer (Theo); N.J. Nieuwkoop; R.A.M. Fouchier (Ron); G.F. Rimmelzwaan (Guus); J.T.M. Voeten; A.D.M.E. Osterhaus (Ab)

    2000-01-01

    textabstractViruses exploit different strategies to escape immune surveillance, including the introduction of mutations in cytotoxic T-lymphocyte (CTL) epitopes. The sequence of these epitopes is critical for their binding to major histocompatibility complex (MHC) class I molecules

  11. Antigen processing influences HIV-specific cytotoxic T lymphocyte immunodominance

    DEFF Research Database (Denmark)

    Tenzer, Stefan; Wee, Edmund; Burgevin, Anne

    2009-01-01

    Although cytotoxic T lymphocytes (CTLs) in people infected with human immunodeficiency virus type 1 can potentially target multiple virus epitopes, the same few are recognized repeatedly. We show here that CTL immunodominance in regions of the human immunodeficiency virus type 1 group-associated ......Although cytotoxic T lymphocytes (CTLs) in people infected with human immunodeficiency virus type 1 can potentially target multiple virus epitopes, the same few are recognized repeatedly. We show here that CTL immunodominance in regions of the human immunodeficiency virus type 1 group...

  12. Cytotoxic T lymphocyte activity and CD8 subpopulations in children at risk of HIV infection.

    Science.gov (United States)

    Aldhous, M C; Watret, K C; Mok, J Y; Bird, A G; Froebel, K S

    1994-07-01

    HIV-specific cytotoxic T lymphocytes (CTL) are thought to play a major role in viral control in HIV-infected adults. Changes in the relative proportions of CD8 lymphocyte subpopulations are also thought to be associated with disease progression. Less is known about the relative effectiveness of CTL against different HIV targets, or about the relationship, if any, between CTL activity and CD8 subpopulations. We have measured CTL activity against four HIV gene products (gag, tat, pol and env) and expression of CD45RO, CD45RA, HLA-DR, CD29, S6F1, and CD57 surface markers on CD8 cells from nine HIV-infected and 11 HIV-uninfected children. Of nine HIV-infected children, six showed antigen-specific CTL activity on at least one occasion: 4/6 directed against tat, 6/6 against pol, 1/6 against env, and 1/6 against gag. However, the specificity of the CTL activity varied between children and within individual children with time. Furthermore, two uninfected children showed CTL activity, one to HIV-gag, -pol and -tat, and the other to HIV-pol. All the HIV-infected and two uninfected children had abnormal proportions of CD8 subpopulations in whole blood compared with age-matched controls. There was no correlation between CTL activity and CD8 subsets in whole blood. Five children changed from CTL-positive to CTL-negative (or vice versa) during the study. In these, the occasions when CTL activity was detected coincided with an increase in CD8 cells, an expansion of HLA-DR+ CD8 cells and a loss of CD45RA+ CD8 cells.

  13. ENDOGENOUS EXPRESSION AND HLA STABILIZATION ASSAY OF PLASMODIUM FALCIPARUM CTL EPITOPE MINIGENE IN HUMAN HLA-A2.1 AND HLA-B51 CELLS

    Institute of Scientific and Technical Information of China (English)

    唐玉阳; 王恒

    2001-01-01

    Objective. To evaluate the Plasmodium falciparum CTL epitope vaccines in HLA class I allele specific human cell lines that have high frequency among Chinese population. Methods. Synthesized oligonucleotides encoding for P.f. CTL epitope genes, constructed eukaryotic expression plasmids, transfected the minigenes into HLA class I allele specific human cell lines and identified endogenous expressing of the minigenes by RT-PCR and HLA stabilization assay. Results. Two mini-genes encoding Plasmodium falciparum CTL epitopes were designed and cloned, respectively, into an eukaryotic expressing vector to form TR26 which was restricted to HLA-B51, SH6 which was restricted to HLA-A2.1, and TS, which had the two aforementioned mini-genes fused in tandem. All of these CTL epitope genes were transfected and endogenously expressed in respective cell lines containing appropriate HLA molecules. The obviously increased expressions of HLA class I molecules were detected in the transfected cell lines. It was demonstrated that the two discrete Plasmodium falciparum epitope genes were effectively processed and presented, and the close proximity of the two epitope genes in one chain as in mini-gene TS did not interfere with the processing and presenting of each epitope gene in corresponding cell line. Conclusion. A successful expression and presentation of multiple CTL epitope mini-gene in MHC class I allele specific human cell lines were demonstrated by an in vitro assay, which could be corresponding to the vaccination of CTL vaccines in people with different MHC I molecules. This work also suggested the possibility of constructing a multiple CTL epitope plasmodium falciparum DNA vaccine that could cover most of Chinese population.

  14. Cytotoxic T lymphocytes specific for the simian immunodeficiency virus.

    Science.gov (United States)

    Letvin, N L; Schmitz, J E; Jordan, H L; Seth, A; Hirsch, V M; Reimann, K A; Kuroda, M J

    1999-08-01

    A non-human primate model for acquired immunodeficiency syndrome (AIDS), the simian immunodeficiency virus (SIV)-infected rhesus monkey, was used to explore the role of the AIDS virus-specific cytotoxic T-lymphocyte (CTL) response in disease pathogenesis. This CTL response was measured using the major histocompatibility complex (MHC) class I/peptide tetramer technology. Large numbers of tetramer-binding CD8+ T lymphocytes were demonstrable not only in the peripheral blood, but in lymph nodes and even in semen of chronically SIV-infected monkeys. The central role of these effector T lymphocytes in containing SIV spread during primary infection was demonstrated by showing that early SIV clearance during primary infection correlated with the emergence of the tetramer binding CD8+ T lymphocytes and that in vivo depletion of CD8+ lymphocytes eliminated the ability of the infected monkeys to contain SIV replication. These observations suggest that an effective AIDS vaccine should elicit a potent virus-specific CTL response. In fact, a live, recombinant SIV vaccine constructed using the attenuated pox virus vector modified vaccinia Ankara (MVA) elicited a high-frequency CTL response, comparable in magnitude to that elicited by SIV infection itself. This suggests that vaccine modalities such as MVA may prove useful in creating an effective human immunodeficiency virus (HIV) vaccine. These studies also indicate the power of both the SIV/macaque model and MHC class I/peptide tetramers for assessing AIDS vaccine strategies.

  15. Peptide‐MHC class I stability is a better predictor than peptide affinity of CTL immunogenicity

    DEFF Research Database (Denmark)

    Harndahl, Mikkel Nors; Rasmussen, Michael; Roder, Gustav

    2012-01-01

    Efficient presentation of peptide‐MHC class I (pMHC‐I) complexes to immune T cells should benefit from a stable peptide‐MHC‐I interaction. However, it has been difficult to distinguish stability from other requirements for MHC‐I binding, for example, affinity. We have recently established a high......‐throughput assay for pMHC‐I stability. Here, we have generated a large database containing stability measurements of pMHC‐I complexes, and re‐examined a previously reported unbiased analysis of the relative contributions of antigen processing and presentation in defining cytotoxic T lymphocyte (CTL) immunogenicity...... [Assarsson et al., J. Immunol. 2007. 178: 7890–7901]. Using an affinity‐balanced approach, we demonstrated that immunogenic peptides tend to be more stably bound to MHC‐I molecules compared with nonimmunogenic peptides. We also developed a bioinformatics method to predict pMHC‐I stability, which suggested...

  16. Identification of a nonameric H-2Kk-restricted CD8+ cytotoxic T lymphocyte epitope on the Plasmodium falciparum circumsporozoite protein

    DEFF Research Database (Denmark)

    Malik, A; Houghten, R; Corradin, G

    1995-01-01

    Class I-restricted CD8+ cytotoxic T lymphocytes (CTL) against the circumsporozoite protein (CSP) protect mice against the rodent malaria parasite, Plasmodium yoelii, and vaccines designed to produce protective CTL against the P. falciparum CSP (PfCSP) are under development. Humans and B10.BR (H-2k...

  17. The presence of protective cytotoxic T lymphocytes does not correlate with shorter lifespans of productively infected cells in HIV-1 infection

    NARCIS (Netherlands)

    Spits, Hilde B; Mudrikova, Tania; Schellens, Ingrid M M; Wensing, Annemarie M J; Prins, Jan M; Feuth, Thijs; Spierings, Erik; Nijhuis, Monique; van Baarle, Debbie; Borghans, José A M

    2016-01-01

    OBJECTIVES AND DESIGN: CD8+ cytotoxic T lymphocytes (CTL) are important in the control of HIV infection. Although CTL are thought to reduce the lifespan of productively infected cells, CD8+ T-cell depletion in simian immunodeficiency virus-infected rhesus-macaques showed no effect on the lifespan of

  18. DAI (DLM-1/ZBP1) as a genetic adjuvant for DNA vaccines that promotes effective antitumor CTL immunity.

    Science.gov (United States)

    Lladser, Alvaro; Mougiakakos, Dimitrios; Tufvesson, Helena; Ligtenberg, Maarten A; Quest, Andrew Fg; Kiessling, Rolf; Ljungberg, Karl

    2011-03-01

    DNA vaccination is an attractive approach to induce antigen-specific cytotoxic CD8(+) T lymphocytes (CTLs), which can mediate protective antitumor immunity. The potency of DNA vaccines encoding weakly immunogenic tumor-associated antigens (TAAs) can be enhanced by codelivering gene-encoded adjuvants. Pattern recognition receptors (PRRs) that sense intracellular DNA could potentially be used to harness intrinsic immune-stimulating properties of plasmid DNA vaccines. Consequently, the cytosolic DNA sensor, DNA-dependent activator of interferon (IFN) regulatory factors (DAI), was used as a genetic adjuvant. In vivo electroporation (EP) of mice with a DAI-encoding plasmid (pDAI) promoted transcription of genes encoding type I IFNs, proinflammatory cytokines, and costimulatory molecules. Coimmunization with pDAI and antigen-encoding plasmids enhanced in vivo antigen-specific proliferation, and induction of effector and memory CTLs. Moreover, codelivery of pDAI effectively promoted CTL and CD4(+) Th1 responses to the TAA survivin. The DAI-enhanced CTL induction required nuclear factor κB (NF-κB) activation and type I IFN signaling, but did not involve the IFN regulatory factor 3 (IRF3). Codelivery of pDAI also increased CTL responses to the melanoma-associated antigen tyrosinase-related protein-2 (TRP2), enhanced tumor rejection and conferred long-term protection against B16 melanoma challenge. This study constitutes "proof-of-principle" validating the use of intracellular PRRs as genetic adjuvants to enhance DNA vaccine potency.

  19. Is There a Role for HTLV-1-Specific CTL in Adult T-Cell Leukemia/Lymphoma?

    Directory of Open Access Journals (Sweden)

    Aileen G. Rowan

    2012-01-01

    Full Text Available ATLL is an aggressive malignancy of T cells that affects about 5% of individuals infected with HTLV-1. The precise mechanism of oncogenesis is not known, but there is evidence that two regulatory viral proteins, Tax and HBZ, are involved. A high set point proviral load is associated with development of ATLL or a chronic inflammatory condition, HAM/TSP. Several lines of evidence, including HLA class 1 association studies and in vitro killing assays, indicate that cytotoxic T lymphocytes are instrumental in determining this proviral load set point. Prior studies have focused chiefly on the CTL response to the immunodominant Tax protein: efficient lysis of Tax-expressing cells inversely correlates with proviral load in nonmalignant infection. However, a recent study showed that strong binding of peptides from HBZ, but not Tax, to HLA class 1 molecules was associated with a low proviral load and a reduced risk of developing HAM/TSP, indicating an important role for HBZ-specific CTL in determining infection outcome. In comparison with nonmalignant infection, HTLV-1-specific CTLs in ATLL patients are reduced in frequency and functionally deficient. Here we discuss the nature of protective CTL responses in nonmalignant HTLV-1 infection and explore the potential of CTLs to protect against ATLL.

  20. CTL responses of high functional avidity and broad variant cross-reactivity are associated with HIV control.

    Directory of Open Access Journals (Sweden)

    Beatriz Mothe

    Full Text Available Cytotoxic T lymphocyte (CTL responses targeting specific HIV proteins, in particular Gag, have been associated with relative control of viral replication in vivo. However, Gag-specific CTL can also be detected in individuals who do not control the virus and it remains thus unclear how Gag-specific CTL may mediate the beneficial effects in some individuals but not in others. Here, we used a 10mer peptide set spanning HIV Gag-p24 to determine immunogen-specific T-cell responses and to assess functional properties including functional avidity and cross-reactivity in 25 HIV-1 controllers and 25 non-controllers without protective HLA class I alleles. Our data challenge the common belief that Gag-specific T cell responses dominate the virus-specific immunity exclusively in HIV-1 controllers as both groups mounted responses of comparable breadths and magnitudes against the p24 sequence. However, responses in controllers reacted to lower antigen concentrations and recognized more epitope variants than responses in non-controllers. These cross-sectional data, largely independent of particular HLA genetics and generated using direct ex-vivo samples thus identify T cell responses of high functional avidity and with broad variant reactivity as potential functional immune correlates of relative HIV control.

  1. A new way to generate cytolytic tumor-specific T cells: electroporation of RNA coding for a T cell receptor into T lymphocytes.

    Science.gov (United States)

    Schaft, Niels; Dörrie, Jan; Müller, Ina; Beck, Verena; Baumann, Stefanie; Schunder, Tanja; Kämpgen, Eckhart; Schuler, Gerold

    2006-09-01

    Effective T cell receptor (TCR) transfer until now required stable retroviral transduction. However, retroviral transduction poses the threat of irreversible genetic manipulation of autologous cells. We, therefore, used optimized RNA transfection for transient manipulation. The transfection efficiency, using EGFP RNA, was >90%. The electroporation of primary T cells, isolated from blood, with TCR-coding RNA resulted in functional cytotoxic T lymphocytes (CTLs) (>60% killing at an effector to target ratio of 20:1) with the same HLA-A2/gp100-specificity as the parental CTL clone. The TCR-transfected T cells specifically recognized peptide-pulsed T2 cells, or dendritic cells electroporated with gp100-coding RNA, in an IFNgamma-secretion assay and retained this ability, even after cryopreservation, over 3 days. Most importantly, we show here for the first time that the electroporated T cells also displayed cytotoxicity, and specifically lysed peptide-loaded T2 cells and HLA-A2+/gp100+ melanoma cells over a period of at least 72 h. Peptide-titration studies showed that the lytic efficiency of the RNA-transfected T cells was similar to that of retrovirally transduced T cells, and approximated that of the parental CTL clone. Functional TCR transfer by RNA electroporation is now possible without the disadvantages of retroviral transduction, and forms a new strategy for the immunotherapy of cancer.

  2. Reducing Validity in Epistemic ATL to Validity in Epistemic CTL

    Directory of Open Access Journals (Sweden)

    Dimitar P. Guelev

    2013-03-01

    Full Text Available We propose a validity preserving translation from a subset of epistemic Alternating-time Temporal Logic (ATL to epistemic Computation Tree Logic (CTL. The considered subset of epistemic ATL is known to have the finite model property and decidable model-checking. This entails the decidability of validity but the implied algorithm is unfeasible. Reducing the validity problem to that in a corresponding system of CTL makes the techniques for automated deduction for that logic available for the handling of the apparently more complex system of ATL.

  3. The role of chemokines in lymphocyte infiltration in ovarian cancer: from MRNA microarray to tissue microarray

    NARCIS (Netherlands)

    Gooden, M.J.M.; Leffers, N.; Fehrmann, R.S.N.; Ten Hoor, K.A.; Daemen, T.; Van Der Zee, A.G.J.; De Bock, G.H.; Walenkamp, A.M.E.; Nijman, H.W.

    2011-01-01

    Introduction: High numbers of CD8+ cytotoxic T lymphocytes (CTL) are associated with a survival advantage in ovarian cancer. Chemokines may play a role in T lymphocyte recruitment to the tumor, but they are also linked to metastasis and angiogenesis. Aim: To determine to what extent chemokines are

  4. Promiscuous CTL recognition of viral epitopes on multiple human leukocyte antigens: biological validation of the proposed HLA A24 supertype.

    Science.gov (United States)

    Burrows, Scott R; Elkington, Rebecca A; Miles, John J; Green, Katherine J; Walker, Susan; Haryana, Sofia M; Moss, Denis J; Dunckley, Heather; Burrows, Jacqueline M; Khanna, Rajiv

    2003-08-01

    Multiple HLA class I alleles can bind peptides with common sequence motifs due to structural similarities in the peptide binding cleft, and these groups of alleles have been classified into supertypes. Nine major HLA supertypes have been proposed, including an A24 supertype that includes A*2301, A*2402, and A*3001. Evidence for this A24 supertype is limited to HLA sequence homology and/or similarity in peptide binding motifs for the alleles. To investigate the immunological relevance of this proposed supertype, we have examined two viral epitopes (from EBV and CMV) initially defined as HLA-A*2301-binding peptides. The data clearly demonstrate that each peptide could be recognized by CTL clones in the context of A*2301 or A*2402; thus validating the inclusion of these three alleles within an A24 supertype. Furthermore, CTL responses to the EBV epitope were detectable in both A*2301(+) and A*2402(+) individuals who had been previously exposed to this virus. These data substantiate the biological relevance of the A24 supertype, and the identification of viral epitopes with the capacity to bind promiscuously across this supertype could aid efforts to develop CTL-based vaccines or immunotherapy. The degeneracy in HLA restriction displayed by some T cells in this study also suggests that the dogma of self-MHC restriction needs some refinement to accommodate foreign peptide recognition in the context of multiple supertype alleles.

  5. [Changes of T-cell clonality after induction-cultivation of peripheral T lymphocytes in adoptive immunotherapy for leukemias].

    Science.gov (United States)

    Liu, Yan; Gu, Jiang-Ying; Ou, Yuan; Li, Mian-Yang; Wang, He; Jin, Xian; Tao, Xiu-Yan; Liu, Zhao-Li; Ma, Xing-Fan; Wang, Xiu-Li; Ma, Si-Kun; Kang, Rui; Cai, Peng; Tong, Chun-Rong; Zhu, Ping

    2009-06-01

    (+) cells were 71.1 +/- 11.8%, 26.7 +/- 11.4%, 35.7 +/- 12.9%, 3.1 +/- 1.6% and 0.12 +/- 0.1% respectively before the induction and cultivation, and changed to 95.4 +/- 3.2% (p 0.01), 55.5 +/- 13.8% (p cultivation. It is concluded that the major action of this induction and cultivation method on T-lymphocytes in vitro is the promotion of CTL and NK T-cell proliferation. In leukemic patients at the remission stage, the TCRBV profile is characterized by the oligoclonal proliferation of T-lymphocytes. Several proliferated clones may have the same motif in CDR3, suggesting the recognition of the same antigen by these lymphocyte clones. Cytokine induction and co-culture with autogenous DCs can stimulate the T-lymphocytes to recover their immunocompetence as manifested by the polyclonal profile and the proliferation of CTL and NK-T cells.

  6. Dengue virus-specific cross-reactive CD8+ human cytotoxic T lymphocytes.

    OpenAIRE

    Bukowski, J F; Kurane, I; Lai, C J; Bray, M.; Falgout, B.; Ennis, F A

    1989-01-01

    Stimulation with live dengue virus of peripheral blood mononuclear cells from a dengue virus type 4-immune donor generated virus-specific, serotype-cross-reactive, CD8+, class I-restricted cytotoxic T lymphocytes (CTL) capable of lysing dengue virus-infected cells and cells pulsed with dengue virus antigens of all four serotypes. These CTL lysed autologous fibroblasts infected with vaccinia virus-dengue virus recombinant viruses containing the E gene or several nonstructural dengue virus type...

  7. T Cell Receptor Vβ Staining Identifies the Malignant Clone in Adult T cell Leukemia and Reveals Killing of Leukemia Cells by Autologous CD8+ T cells.

    Directory of Open Access Journals (Sweden)

    Aileen G Rowan

    2016-11-01

    Full Text Available There is growing evidence that CD8+ cytotoxic T lymphocyte (CTL responses can contribute to long-term remission of many malignancies. The etiological agent of adult T-cell leukemia/lymphoma (ATL, human T lymphotropic virus type-1 (HTLV-1, contains highly immunogenic CTL epitopes, but ATL patients typically have low frequencies of cytokine-producing HTLV-1-specific CD8+ cells in the circulation. It remains unclear whether patients with ATL possess CTLs that can kill the malignant HTLV-1 infected clone. Here we used flow cytometric staining of TCRVβ and cell adhesion molecule-1 (CADM1 to identify monoclonal populations of HTLV-1-infected T cells in the peripheral blood of patients with ATL. Thus, we quantified the rate of CD8+-mediated killing of the putative malignant clone in ex vivo blood samples. We observed that CD8+ cells from ATL patients were unable to lyse autologous ATL clones when tested directly ex vivo. However, short in vitro culture restored the ability of CD8+ cells to kill ex vivo ATL clones in some donors. The capacity of CD8+ cells to lyse HTLV-1 infected cells which expressed the viral sense strand gene products was significantly enhanced after in vitro culture, and donors with an ATL clone that expressed the HTLV-1 Tax gene were most likely to make a detectable lytic CD8+ response to the ATL cells. We conclude that some patients with ATL possess functional tumour-specific CTLs which could be exploited to contribute to control of the disease.

  8. T Cell Receptor Vβ Staining Identifies the Malignant Clone in Adult T cell Leukemia and Reveals Killing of Leukemia Cells by Autologous CD8+ T cells.

    Science.gov (United States)

    Rowan, Aileen G; Witkover, Aviva; Melamed, Anat; Tanaka, Yuetsu; Cook, Lucy B M; Fields, Paul; Taylor, Graham P; Bangham, Charles R M

    2016-11-01

    There is growing evidence that CD8+ cytotoxic T lymphocyte (CTL) responses can contribute to long-term remission of many malignancies. The etiological agent of adult T-cell leukemia/lymphoma (ATL), human T lymphotropic virus type-1 (HTLV-1), contains highly immunogenic CTL epitopes, but ATL patients typically have low frequencies of cytokine-producing HTLV-1-specific CD8+ cells in the circulation. It remains unclear whether patients with ATL possess CTLs that can kill the malignant HTLV-1 infected clone. Here we used flow cytometric staining of TCRVβ and cell adhesion molecule-1 (CADM1) to identify monoclonal populations of HTLV-1-infected T cells in the peripheral blood of patients with ATL. Thus, we quantified the rate of CD8+-mediated killing of the putative malignant clone in ex vivo blood samples. We observed that CD8+ cells from ATL patients were unable to lyse autologous ATL clones when tested directly ex vivo. However, short in vitro culture restored the ability of CD8+ cells to kill ex vivo ATL clones in some donors. The capacity of CD8+ cells to lyse HTLV-1 infected cells which expressed the viral sense strand gene products was significantly enhanced after in vitro culture, and donors with an ATL clone that expressed the HTLV-1 Tax gene were most likely to make a detectable lytic CD8+ response to the ATL cells. We conclude that some patients with ATL possess functional tumour-specific CTLs which could be exploited to contribute to control of the disease.

  9. CD40 agonist converting CTL exhaustion via the activation of the mTORC1 pathway enhances PD-1 antagonist action in rescuing exhausted CTLs in chronic infection.

    Science.gov (United States)

    Xu, Aizhang; Wang, Rong; Freywald, Andrew; Stewart, Kristoffor; Tikoo, Suresh; Xu, Jianqing; Zheng, Changyu; Xiang, Jim

    2017-03-11

    Expansion of PD-1-expressing CD8(+) cytotoxic T lymphocytes (CTLs) and associated CTL exhaustion are chief issues for ineffective virus-elimination in chronic infectious diseases. PD-1 blockade using antagonistic anti-PD-L1 antibodies results in a moderate conversion of CTL exhaustion. We previously demonstrated that CD40L signaling of ovalbumin (OVA)-specific vaccine, OVA-Texo, converts CTL exhaustion via the activation of the mTORC1 pathway in OVA-expressing adenovirus (AdVova)-infected B6 mice showing CTL inflation and exhaustion. Here, we developed AdVova-infected B6 and transgenic CD11c-DTR (termed AdVova-B6 and AdVova-CD11c-DTR) mice with chronic infection, and assessed a potential effect of CD40 agonist on the conversion of CTL exhaustion and on a potential enhancement of PD-1 antagonist action in rescuing exhausted CTLs in our chronic infection models. We demonstrate that a single dose of anti-CD40 alone can effectively convert CTL exhaustion by activating the mTORC1 pathway, leading to CTL proliferation, up-regulation of an effector-cytokine IFN-γ and the cytolytic effect in AdVova-B6 mice. Using anti-CD4 antibody and diphtheria toxin (DT) to deplete CD4(+) T-cells and dendritic cells (DCs), we discovered that the CD40 agonist-induced conversion in AdVova-B6 and AdVova-CD11c-DTR mice is dependent upon host CD4(+) T-cell and DC involvements. Moreover, CD40 agonist significantly enhances PD-1 antagonist effectiveness in rescuing exhausted CTLs in chronic infection. Taken together, our data demonstrate the importance of CD40 signaling in the conversion of CTL exhaustion and its ability to enhance PD-1 antagonist action in rescuing exhausted CTLs in chronic infection. Therefore, our findings may positively impact the design of new therapeutic strategies for chronic infectious diseases.

  10. Identification and cloning of a prethymic precursor T lymphocyte from a population of common acute lymphoblastic leukemia antigen (CALLA)-positive fetal bone marrow cells

    DEFF Research Database (Denmark)

    Hokland, P; Hokland, M; Daley, J

    1987-01-01

    We have cloned common acute lymphoblastic leukemia (CALLA)-positive cells from human fetal bone marrow containing less than 1 in 10,000 E-RFC in round-bottomed microtiter wells (one cell per well) using the autocloning unit of an EPICS-V cell sorter. Expansion of such cells (with IL-2 and heavily...

  11. Differential requirement for protein tyrosine kinase Fyn in the functional activation of antigen-specific T lymphocyte clones through the TCR or Thy-1.

    Science.gov (United States)

    Lancki, D W; Qian, D; Fields, P; Gajewski, T; Fitch, F W

    1995-05-01

    The protein tyrosine kinase Fyn has been shown to be involved in signal transduction through the TCR and the glycosyl-phosphatidylinositol-linked surface molecule Thy-1 expressed on T cells. In this study, we examine the requirement for Fyn expression in signaling through the TCR or Thy-1 using a panel of Ag-specific T cell clones derived from fyn-/- mutant mice. These clones do not express normal Fyn protein, as measured by immune-complex kinase reaction using anti-Fyn Ab. Stimulation through the TCR, either by APC bearing relevant Ag or by immobilized anti-CD3 mAb, resulted in comparable levels of proliferation, lymphokine production, and cytolysis by clones from both wild-type and fyn-/- mice. In contrast, stimulation through Thy-1, using soluble (or cross-linked) anti-Thy-1 mAb, was deficient, as measured by these responses. Thus, Fyn expression is selectively required for functional activation through Thy-1 in these T cell clones.

  12. Analysis of HBV-specific cytotoxic T lymphocytes in patients with hepatitis B

    Directory of Open Access Journals (Sweden)

    Ning DING

    2011-04-01

    Full Text Available Objective To investigate the expression feature of hepatitis B virus(HBV-specific cytotoxic T lymphocytes(CTL,and analyze the role of CTL in immune response.Methods Peripheral venous blood was collected from 40 acute hepatitis B(AHB patients hospitalized from Oct.2008 to Jun.2010 in 302 Hospital of PLA for the detection of human leucocyte antigen-A2(HLA-A2.Out of the 40 patients,18 were HLA-A2-positive,and their peripheral blood mononuclear cells were isolated at acute phase and convalescent phase if the patients could be contacted.Six HLA-A2-negative AHB patients,18 HLA-A2-positive chronic HB patients and 6 HLA-A2-positive normal persons served as controls.The CTL was stained by fluorescence-labeled anti-CD3 monoclonal antibody,anti-CD8 monoclonal antibody and HBV epitopic pentamers(S183-191,S335-343,and examined with flow cytometry,then analyzed by software Cell-quest.The peak value of alanine aminotransferase(ALT and HBV load was detected at acute phase of patients,and the correlation was analyzed.Results The average frequencies were 0.23%±0.18% for HBsAg183-191-specific CTL and 0.49%±0.31% for HBsAg335-343-specific CTL at the acute phase of the 18 AHB patients,and they were significantly higher than those in patients with chronic HB,in whom CTL frequencies were 0.07%±0.03% for HBsAg183-191-specific CTL and 0.08%±0.04% for HBsAg335-343-specific CTL(P < 0.01.There was no correlation between the frequency of HBsAg183-191-specific CTL and the peak value of ALT(r=0.4506,P=0.0528,while the frequency of HBsAg335-343-specific CTL was correlated with ALT level(r=0.5642,P=0.0119.Neither the two CTL frequencies showed correlation with virus load.Compared with the acute phase,the CTL frequencies obviously decreased at convalescent phase(P=0.011.Both kinds of CTL were not detectable in HLA-A2-negative AHB patients and HLA-A2-positive healthy individuals.Conclusion HBV-specific CTL may play an important role in both virus clearance and liver

  13. Engineering a CTL-Tailored Replicon RNA Vaccine against PRRSV

    DEFF Research Database (Denmark)

    Welner, Simon; Werder, Simea; Nielsen, Morten

    The development of vaccines against porcine reproductive and respiratory syndrome virus (PRRSV) has been hampered by the high mutation rate and the multiple immunoevasive strategies of the virus. With the overall aim of designing a broad coverage vaccine that induces an effective CTL response...... detection in the presence of a proteasome inhibitor. Finally, a vaccination-challenge experiment using 18 SLA-matched pigs is currently being conducted until July 2016 in which a test group and a control group are being vaccinated twice with VRPs expressing PRRSV epitopes and non-sense control epitopes...... will be available for IVIS. This study exemplifies how bioinformatics epitope prediction, recombinant SLA molecules and RNA virus replicon design can be used to engineer a replicating non-propagating vaccine tailored to deliver conserved and immunogenic CTL epitopes....

  14. Harmful Gas Recognition Exploiting a CTL Sensor Array

    Directory of Open Access Journals (Sweden)

    Yao Zheng

    2013-10-01

    Full Text Available In this paper, a novel cataluminescence (CTL-based sensor array consisting of nine types of catalytic materials is developed for the recognition of several harmful gases, namely carbon monoxide, acetone, chloroform and toluene. First, the experimental setup is constructed by using sensing nanomaterials, a heating plate, a pneumatic pump, a gas flow meter, a digital temperature device, a camera and a BPCL Ultra Weak Chemiluminescence Analyzer. Then, unique CTL patterns for the four types of harmful gas are obtained from the sensor array. The harmful gases are successful recognized by the PCA method. The optimal conditions are also investigated. Finally, experimental results show high sensitivity, long-term stability and good linearity of the sensor array, which combined with simplicity, make our system a promising application in this field.

  15. Contextual Teaching and Learning (CTL dalam Pembelajaran PAI

    Directory of Open Access Journals (Sweden)

    Muhammad Iwan Abdi

    2011-06-01

    Full Text Available Contextual teaching learning (CTL is an educational process that aims to help students see meaning in academic material they are learning by connecting academic subjects with the context of their daily, which is related to personal circumstances, social, and cultural development. In Contextual teaching and learning (CTL takes an approach that is more empowering students with the expectations of students able to construct knowledge in their minds, rather than memorizing facts. Besides, students learn through experience rather than memorize, given knowledge of the facts and not a concept readily accepted but something that must be constructed by the students. To achieve this goal include the eight components of the system, the following: making meaningful connections, doing meaningful work, conduct self-regulated learning, collaborate, think critically and creatively, helping individuals to grow and develop, achieving high standards , and using authentic assessment. In learning PAI, CTL developed by focusing on moral or affective touch protege acquired through personal experience that photographing their daily environment. This touch your heart and thoughts will inspire students to practice it.

  16. A single nine-amino acid peptide induces virus-specific, CD8+ human cytotoxic T lymphocyte clones of heterogeneous serotype specificities

    OpenAIRE

    1995-01-01

    It is generally accepted that virus-specific CD8+ cytotoxic T lymphocytes (CTLs) recognize nine-amino acid peptides in conjunction with HLA class I molecules. We recently reported that dengue virus- specific CD8+ CTLs of two different serotype specificities, which were established by stimulation with dengue virus, recognize a single nine- amino acid peptide of the nonstructural protein NS3 of dengue virus type 4 (D4V) in an HLA-B35-restricted fashion. To further analyze the relationships betw...

  17. An allospecific murine T helper clone which can help both T and B cell responses in vitro and in vivo

    DEFF Research Database (Denmark)

    Crispe, I N; Gascoigne, N R; Owens, T

    1984-01-01

    . Here we describe an in vitro and in vivo study of this problem, using a Th clone, designated MTH-1. The clone carries the cell surface markers Thy-1 and L3T4a, but lacks Lyt-2. It recognizes a minor alloantigen shared by DBA/2, B10.D2 and NZB spleen cells, and such recognition is restricted by H-2Ed....... Recognition of antigen in vitro is accompanied by secretion of IL-2. In vivo, both primary and secondary CTL responses to multiple minor alloantigens are enhanced by small numbers (less than or equal to 10(4] of MTH-1 cells. Recognition of alloantigen in a T-depleted B cell population results......Both B lymphocytes and cytotoxic T lymphocytes respond to signals from the T helper (Th) compartment, and such signals are mediated by a number of biochemically distinct factors. This raises the question whether help for B cells and T cells is a function of one or several different kinds of Th cell...

  18. Differential functional avidity of dengue virus-specific T-cell clones for variant peptides representing heterologous and previously encountered serotypes.

    Science.gov (United States)

    Imrie, Allison; Meeks, Janet; Gurary, Alexandra; Sukhbataar, Munkhzul; Kitsutani, Paul; Effler, Paul; Zhao, Zhengshan

    2007-09-01

    Proinflammatory cytokines secreted by memory CD8+ and CD4+ T cells are thought to play a direct role in the pathogenesis of dengue virus infection by increasing vascular permeability and thereby inducing the pathophysiologic events associated with dengue hemorrhagic fever and dengue shock syndrome. Severe disease is frequently observed in the setting of secondary infection with heterologous dengue virus serotypes, suggesting a role for cross-reactive memory T cells in the immunopathogenesis of severe disease. We used a large panel of well-characterized dengue virus-specific CD8+ T-cell clones isolated from Pacific Islanders previously infected with dengue virus 1 to examine effector memory function, focusing on a novel dominant HLA-B*5502-restricted NS5(329-337) epitope, and assessed T-cell responses to stimulation with variant peptides representing heterologous serotypes. Variant peptides were differentially recognized by dengue virus 1-specific effector CD8+ cytotoxic T lymphocytes (CTL) in a heterogeneous and clone-specific manner, in which cytolytic function and cytokine secretion could be enhanced, diminished, or abrogated compared with cognate peptide stimulation. Dengue virus-specific CTL stimulated with cognate and variant peptides demonstrated a cytokine response hierarchy of gamma IFN (IFN-gamma) > tumor necrosis factor alpha (TNF-alpha) > interleukin-2 (IL-2), and a subset of clones also produced IL-4 and IL-6. Individual clones demonstrated greater avidity for variant peptides representing heterologous serotypes, including serotypes previously encountered by the subject, and IFN-gamma and TNF-alpha secretion was enhanced by stimulation with these heterologous peptides. Altered antiviral T-cell responses in response to stimulation with heterologous dengue virus serotypes have implications for control of virus replication and for disease pathogenesis.

  19. A Novel CD8-Independent High-Avidity Cytotoxic T-Lymphocyte Response Directed against an Epitope in the Phosphoprotein of the Paramyxovirus Simian Virus 5

    Science.gov (United States)

    Gray, Peter M.; Parks, Griffith D.; Alexander-Miller, Martha A.

    2001-01-01

    Adoptive transfer studies have shown that cytotoxic T lymphocytes (CTL) of high avidity, capable of recognizing low levels of peptide-MHC I molecules, are more efficient at reducing viral titers than are low-avidity CTL, thus establishing CTL avidity as a critical parameter for the ability of a CTL to clear virus in vivo. It has been well documented that CTL of high avidity are relatively CD8 independent, whereas low-avidity CTL require CD8 engagement in order to become activated. In this study we have analyzed the antiviral CTL response elicited following infection with the paramyxovirus simian virus 5 (SV5). We have identified the immunodominant and subdominant CTL responses and subsequently assessed the avidity of these responses by their CD8 dependence. This is the first study in which the relationship between immunodominance and CTL avidity has been investigated. The immunodominant response was directed against an epitope present in the viral M protein, and subdominant responses were directed against epitopes present in the P, F, and HN proteins. Similarly to other CTL responses we have analyzed, the immunodominant response and the subdominant F and HN responses were comprised of both high- and low-avidity CTL. However, the subdominant response directed against the epitope present in the P protein is novel, as it is exclusively high avidity. This high-avidity response is independent of both the route of infection and expression by recombinant SV5. A further understanding of the inherent properties of P that elicit only high-avidity CTL may allow for the design of more efficacious vaccine vectors that preferentially elicit high-avidity CTL in vivo. PMID:11581375

  20. A general functional response of cytotoxic T lymphocyte-mediated killing of target cells.

    Science.gov (United States)

    Gadhamsetty, Saikrishna; Marée, Athanasius F M; Beltman, Joost B; de Boer, Rob J

    2014-04-15

    Cytotoxic T lymphocytes (CTLs) kill virus-infected cells and tumor cells, and play a critical role in immune protection. Our knowledge of how the CTL killing efficiency varies with CTL and target cell numbers is limited. Here, we simulate a region of lymphoid tissue using a cellular Potts model to characterize the functional response of CTL killing of target cells, and find that the total killing rate saturates both with the CTL and the target cell densities. The relative saturation in CTL and target cell densities is determined by whether a CTL can kill multiple target cells at the same time, and whether a target cell can be killed by many CTLs together. We find that all the studied regimes can be well described by a double-saturation (DS) function with two different saturation constants. We show that this DS model can be mechanistically derived for the cases where target cells are killed by a single CTL. For the other cases, a biological interpretation of the parameters is still possible. Our results imply that this DS function can be used as a tool to predict the cellular interactions in cytotoxicity data.

  1. Cell surface phenotype of cytolytic T lymphocyte precursors in aged nude mice.

    Science.gov (United States)

    Maryanski, J L; MacDonald, H R; Sordat, B; Cerottini, J C

    1981-12-01

    The cell surface phenotype of cytolytic T lymphocyte precursors (CTL-P) in congenitally athymic C57BL/6 nu/nu mice has been investigated. CTL-P were detected and quantitated in a limited dilution mixed leukocyte microculture assay system supplemented with interleukin 2. Minimal estimates of the frequency of CTL-P among nylon wool passed (NWP) nude spleen cells were obtained following elimination of Thy-1-bearing or Lyt-2-bearing cells with monoclonal antibodies plus complement. Alternatively, NWP spleen cells bearing Thy-1 or Lyt-2 were positively selected on a cell sorter and assayed for CTL-P frequency. Both positive and negative selection techniques demonstrated that essentially all (greater than 98%) CTL-P in NWP nude spleen expressed Thy-1 and that the majority (80-90%) expressed Lyt-2. In control NWP spleen cells from normal C57BL/6 mice, greater than 98% of CTL-P were positive for both Thy-1 and Lyt-2. These data demonstrate that most functional alloreactive CTL-P developing in the apparent absence of thymic influence already express both Thy-1 and Lyt-2 prior to exposure to antigen.

  2. A human Fab fragment specific for thyroid peroxidase generated by cloning thyroid lymphocyte-derived immunoglobulin genes in a bacteriophage lambda library.

    Science.gov (United States)

    Portolano, S; Seto, P; Chazenbalk, G D; Nagayama, Y; McLachlan, S M; Rapoport, B

    1991-08-30

    A human Fab fragment (SP2) which binds specifically to human thyroid peroxidase has been generated by expressing random combinations of heavy and light chain immunoglobulin genes (derived from Graves' thyroid cDNA) in a bacteriophage lambda library. In common with many serum TPO autoantibodies, the cloned Fab fragment is IgG1 kappa and has a high affinity for TPO (approximately 10(-9) M). On the basis of their nucleotide sequences, the heavy and light chain genes coding for SP2 belong to families VHI, (D), JH3 and VKI, JK2, respectively. These data provide the first characterization at a molecular level of a human thyroid peroxidase antibody associated with autoimmune thyroid disease.

  3. Model Checking Electronic Commerce Security Protocols Based on CTL

    Institute of Scientific and Technical Information of China (English)

    XIAO De-qin; ZHANG Huan-guo

    2005-01-01

    We present a model based on Computational Temporal Logic (CTL) methods for verifying security requirements of electronic commerce protocols. The model describes formally the authentication, confidentiality integrity,non-repudiation, denial of service and access control of the electronic commerce protocols. We illustrate as case study a variant of the Lu-Smolka protocol proposed by Lu-Smolka.Moreover, we have discovered two attacks that allow a dishonest user to purchase a good debiting the amount to another user. And also, we compared our work with relative research works and found that the formal way of this paper is more general to specify security protocols for E-Commerce.

  4. CTL Induction of Tumoricidal Nitric Oxide Production by Intratumoral Macrophages Is Critical for Tumor Elimination

    OpenAIRE

    Vicetti Miguel, Rodolfo D.; Cherpes, Thomas L.; Watson, Leah J.; McKenna, Kyle C.

    2010-01-01

    To characterize mechanisms of CTL inhibition within an ocular tumor microenvironment, tumor-specific CTLs were transferred into mice with tumors developing within the anterior chamber of the eye or skin. Ocular tumors were resistant to CTL transfer therapy whereas skin tumors were sensitive. CTLs infiltrated ocular tumors at higher CTL/tumor ratios than in skin tumors and demonstrated comparable ex vivo effector function to CTLs within skin tumors indicating that ocular tumor progression was ...

  5. The kinematics of cytotoxic lymphocytes influence their ability to kill target cells.

    Science.gov (United States)

    Bhat, Purnima; Leggatt, Graham; Matthaei, Klaus I; Frazer, Ian H

    2014-01-01

    Cytotoxic lymphocytes (CTL) have been reported to show a range of motility patterns from rapid long-range tracking to complete arrest, but how and whether these kinematics affect their ability to kill target cells is not known. Many in vitro killing assays utilize cell lines and tumour-derived cells as targets, which may be of limited relevance to the kinetics of CTL-mediated killing of somatic cells. Here, live-cell microscopy is used to examine the interactions of CTL and primary murine skin cells presenting antigens. We developed a qualitative and quantitative killing assay using extended-duration fluorescence time-lapse microscopy coupled with large-volume objective software-based data analysis to obtain population data of cell-to-cell interactions, motility and apoptosis. In vivo and ex vivo activated antigen-specific cytotoxic lymphocytes were added to primary keratinocyte targets in culture with fluorometric detection of caspase-3 activation in targets as an objective determinant of apoptosis. We found that activated CTL achieved contact-dependent apoptosis of non-tumour targets after a period of prolonged attachment - on average 21 hours - which was determined by target cell type, amount of antigen, and activation status of CTL. Activation of CTL even without engagement of the T cell receptor was sufficient to mobilise cells significantly above baseline, while the addition of cognate antigen further enhanced their motility. Highly activated CTL showed markedly increased vector displacement, and velocity, and lead to increased antigen-specific target cell death. These data show that the inherent kinematics of CTL correlate directly with their ability to kill non-tumour cells presenting cognate antigen.

  6. The kinematics of cytotoxic lymphocytes influence their ability to kill target cells.

    Directory of Open Access Journals (Sweden)

    Purnima Bhat

    Full Text Available Cytotoxic lymphocytes (CTL have been reported to show a range of motility patterns from rapid long-range tracking to complete arrest, but how and whether these kinematics affect their ability to kill target cells is not known. Many in vitro killing assays utilize cell lines and tumour-derived cells as targets, which may be of limited relevance to the kinetics of CTL-mediated killing of somatic cells. Here, live-cell microscopy is used to examine the interactions of CTL and primary murine skin cells presenting antigens. We developed a qualitative and quantitative killing assay using extended-duration fluorescence time-lapse microscopy coupled with large-volume objective software-based data analysis to obtain population data of cell-to-cell interactions, motility and apoptosis. In vivo and ex vivo activated antigen-specific cytotoxic lymphocytes were added to primary keratinocyte targets in culture with fluorometric detection of caspase-3 activation in targets as an objective determinant of apoptosis. We found that activated CTL achieved contact-dependent apoptosis of non-tumour targets after a period of prolonged attachment - on average 21 hours - which was determined by target cell type, amount of antigen, and activation status of CTL. Activation of CTL even without engagement of the T cell receptor was sufficient to mobilise cells significantly above baseline, while the addition of cognate antigen further enhanced their motility. Highly activated CTL showed markedly increased vector displacement, and velocity, and lead to increased antigen-specific target cell death. These data show that the inherent kinematics of CTL correlate directly with their ability to kill non-tumour cells presenting cognate antigen.

  7. Why Clone?

    Science.gov (United States)

    ... How might cloning be used in medicine? Cloning animal models of disease Much of what researchers learn ... issue of the genetic reshuffling that happensduring sexual reproduction and simply clone our drug-producing cow. Cloning ...

  8. Mathematical modeling of escape of HIV from cytotoxic T lymphocyte responses

    Science.gov (United States)

    Ganusov, Vitaly V.; Neher, Richard A.; Perelson, Alan S.

    2013-01-01

    Human immunodeficiency virus (HIV-1 or simply HIV) induces a persistent infection, which in the absence of treatment leads to AIDS and death in almost all infected individuals. HIV infection elicits a vigorous immune response starting about 2-3 weeks postinfection that can lower the amount of virus in the body, but which cannot eradicate the virus. How HIV establishes a chronic infection in the face of a strong immune response remains poorly understood. It has been shown that HIV is able to rapidly change its proteins via mutation to evade recognition by virus-specific cytotoxic T lymphocytes (CTLs). Typically, an HIV-infected patient will generate 4-12 CTL responses specific for parts of viral proteins called epitopes. Such CTL responses lead to strong selective pressure to change the viral sequences encoding these epitopes so as to avoid CTL recognition. Indeed, the viral population ‘escapes’ from about half of the CTL responses by mutation in the first year. Here we review experimental data on HIV evolution in response to CTL pressure, mathematical models developed to explain this evolution, and highlight problems associated with the data and previous modeling efforts. We show that estimates of the strength of the epitope-specific CTL response depend on the method used to fit models to experimental data and on the assumptions made regarding how mutants are generated during infection. We illustrate that allowing CTL responses to decay over time may improve the model fit to experimental data and provides higher estimates of the killing efficacy of HIV-specific CTLs. We also propose a novel method for simultaneously estimating the killing efficacy of multiple CTL populations specific for different epitopes of HIV using stochastic simulations. Lastly, we show that current estimates of the efficacy at which HIV-specific CTLs clear virus-infected cells can be improved by more frequent sampling of viral sequences and by combining data on sequence evolution with

  9. A comparison of parallel pyrosequencing and sanger clone-based sequencing and its impact on the characterization of the genetic diversity of HIV-1.

    Directory of Open Access Journals (Sweden)

    Binhua Liang

    Full Text Available BACKGROUND: Pyrosequencing technology has the potential to rapidly sequence HIV-1 viral quasispecies without requiring the traditional approach of cloning. In this study, we investigated the utility of ultra-deep pyrosequencing to characterize genetic diversity of the HIV-1 gag quasispecies and assessed the possible contribution of pyrosequencing technology in studying HIV-1 biology and evolution. METHODOLOGY/PRINCIPAL FINDINGS: HIV-1 gag gene was amplified from 96 patients using nested PCR. The PCR products were cloned and sequenced using capillary based Sanger fluorescent dideoxy termination sequencing. The same PCR products were also directly sequenced using the 454 pyrosequencing technology. The two sequencing methods were evaluated for their ability to characterize quasispecies variation, and to reveal sites under host immune pressure for their putative functional significance. A total of 14,034 variations were identified by 454 pyrosequencing versus 3,632 variations by Sanger clone-based (SCB sequencing. 11,050 of these variations were detected only by pyrosequencing. These undetected variations were located in the HIV-1 Gag region which is known to contain putative cytotoxic T lymphocyte (CTL and neutralizing antibody epitopes, and sites related to virus assembly and packaging. Analysis of the positively selected sites derived by the two sequencing methods identified several differences. All of them were located within the CTL epitope regions. CONCLUSIONS/SIGNIFICANCE: Ultra-deep pyrosequencing has proven to be a powerful tool for characterization of HIV-1 genetic diversity with enhanced sensitivity, efficiency, and accuracy. It also improved reliability of downstream evolutionary and functional analysis of HIV-1 quasispecies.

  10. Response of ELA-A1 horses immunized with lipopeptide containing an equine infectious anemia virus ELA-A1-restricted CTL epitope to virus challenge.

    Science.gov (United States)

    Ridgely, Sherritta L; Zhang, Baoshan; McGuire, Travis C

    2003-01-17

    Lipopeptide containing an ELA-A1-restricted cytotoxic T lymphocyte (CTL) epitope from the envelope surface unit (SU) protein of the EIAV(WSU5) strain was used to immunize three horses having the ELA-A1 haplotype. Peptide-specific ELA-A1-restricted CTL were induced in all three horses, although these were present transiently in PBMC. These horses were further immunized with lipopeptide containing the corresponding CTL epitope from the EIAV(PV) strain. Then, the three immunized horses and three non-immunized horses were challenged by intravenous inoculation with 300 TCID(50) EIAV(PV). All horses developed cell free viremia, fever and thrombocytopenia. However, there was a statistically lower fever and thrombocytopenia severity score in the immunized group. Shorter duration of plasma viral load in two of the three immunized horses likely explains the less severe clinical disease in this group. Results indicate that lipopeptide immunization had a protective effect against development of clinical disease following virus challenge.

  11. Effect of insulin on functional status of cord blood-derived dendritic cells and on dendritic cell-induced CTL cytotoxicity against pancreatic cancer cell lines

    Institute of Scientific and Technical Information of China (English)

    Qiu-Liang Liu; Yi-Sheng Wang; Jia-Xiang Wang

    2009-01-01

    BACKGROUND: Dendritic cells (DCs) are the most important antigen-presenting cells in the human body, and DCs with different mature status possess different or even opposite functions. This study was designed to explore the influence of insulin on the functional status of cord blood-derived DCs and on DC-induced cytotoxic T lymphocyte (CTL) activity against pancreatic cancer cell lines. METHODS: Mononuclear cells were isolated from fresh cord blood. Interleukin-4 (IL-4) and granulocyte-macrophage colony-stimulating factor (GM-CSF) were used to induce or stimulate the mononuclear cells. Insulin at different concentrations served to modify DCs, and then DC morphology, number, and growth status were assessed. The DC immunophenotype was detected with a flow cytometer. The IL-12 in DC supernatant was determined by ELISA. DC functional status was evaluated by the autologous mixed lymphocyte reaction. T lymphocytes were induced by insulin-modified DCs to become CTLs. The CTL cytotoxicity against pancreatic cancer cell lines was determined. RESULTS:  Mononuclear cells from cord blood can be differentiated into DCs by cytokine induction and insulin modification. With the increase in insulin concentration (2.5-25 mg/L), the expression of DC HLA-DR, CD1α, CD80, and CD83 was significantly increased, the DC ability to secrete IL-12 was significantly improved, DC function to activate autologous lymphocytes was significantly enhanced, and the cytotoxicity of CTLs induced by insulin-modified DCs against pancreatic cancer cell lines was significantly strengthened. CONCLUSIONS: Insulin may facilitate DC induction and maturation, and improve the reproductive activity of autologous lymphocytes. The cytotoxicity of CTLs induced by insulin-modified DCs against pancreatic cancer cell lines was significantly enhanced. Insulin may serve as a factor modifying DCs and inducing CTLs in vitro in insulin biotherapy.

  12. Th cells promote CTL survival and memory via acquired pMHC-I and endogenous IL-2 and CD40L signaling and by modulating apoptosis-controlling pathways.

    Directory of Open Access Journals (Sweden)

    Channakeshava Sokke Umeshappa

    Full Text Available Involvement of CD4(+ helper T (Th cells is crucial for CD8(+ cytotoxic T lymphocyte (CTL-mediated immunity. However, CD4(+ Th's signals that govern CTL survival and functional memory are still not completely understood. In this study, we assessed the role of CD4(+ Th cells with acquired antigen-presenting machineries in determining CTL fates. We utilized an adoptive co-transfer into CD4(+ T cell-sufficient or -deficient mice of OTI CTLs and OTII Th cells or Th cells with various gene deficiencies pre-stimulated in vitro by ovalbumin (OVA-pulsed dendritic cell (DCova. CTL survival was kinetically assessed in these mice using FITC-anti-CD8 and PE-H-2K(b/OVA257-264 tetramer staining by flow cytometry. We show that by acting via endogenous CD40L and IL-2, and acquired peptide-MHC-I (pMHC-I complex signaling, CD4(+ Th cells enhance survival of transferred effector CTLs and their differentiation into the functional memory CTLs capable of protecting against highly-metastasizing tumor challenge. Moreover, RT-PCR, flow cytometry and Western blot analysis demonstrate that increased survival of CD4(+ Th cell-helped CTLs is matched with enhanced Akt1/NF-κB activation, down-regulation of TRAIL, and altered expression profiles with up-regulation of prosurvival (Bcl-2 and down-regulation of proapoptotic (Bcl-10, Casp-3, Casp-4, Casp-7 molecules. Taken together, our results reveal a previously unexplored mechanistic role for CD4(+ Th cells in programming CTL survival and memory recall responses. This knowledge could also aid in the development of efficient adoptive CTL cancer therapy.

  13. The cytotoxic T lymphocyte immune synapse at a glance.

    Science.gov (United States)

    Dieckmann, Nele M G; Frazer, Gordon L; Asano, Yukako; Stinchcombe, Jane C; Griffiths, Gillian M

    2016-08-01

    The immune synapse provides an important structure for communication with immune cells. Studies on immune synapses formed by cytotoxic T lymphocytes (CTLs) highlight the dynamic changes and specialised mechanisms required to facilitate focal signalling and polarised secretion in immune cells. In this Cell Science at a Glance article and the accompanying poster, we illustrate the different steps that reveal the specialised mechanisms used to focus secretion at the CTL immune synapse and allow CTLs to be such efficient and precise serial killers.

  14. Identification of a cyclin B1-derived CTL epitope eliciting spontaneous responses in both cancer patients and healthy donors

    DEFF Research Database (Denmark)

    Andersen, Rikke Sick; Sørensen, Rikke Bæk; Ritter, Cathrin

    2011-01-01

    was verified in an HLA stabilization assay. By stimulation with peptide-loaded dendritic cells a CTL clone was established, which was able to kill two breast cancer cell lines in an HLA-A2-dependent and peptide-specific manner, demonstrating presentation of the peptide on the surface of cancer cells....... Furthermore, blood from cancer patients and healthy donors was screened for spontaneous T-cell reactivity against the peptide in IFN-γ ELISPOT assays. Patients with breast cancer, malignant melanoma, or renal cell carcinoma hosted powerful and high-frequency T-cell responses against the peptide. In addition......, when blood from healthy donors was tested, similar responses were observed. Ultimately, serum from cancer patients and healthy donors was analyzed for anti-cyclin B1 antibodies. Humoral responses against cyclin B1 were frequently detected in both cancer patients and healthy donors. In conclusion...

  15. Identification of a cyclin B1-derived CTL epitope eliciting spontaneous responses in both cancer patients and healthy donors

    DEFF Research Database (Denmark)

    Andersen, Rikke Sick; Sørensen, Rikke Bæk; Ritter, Cathrin

    2011-01-01

    was verified in an HLA stabilization assay. By stimulation with peptide-loaded dendritic cells a CTL clone was established, which was able to kill two breast cancer cell lines in an HLA-A2-dependent and peptide-specific manner, demonstrating presentation of the peptide on the surface of cancer cells....... Furthermore, blood from cancer patients and healthy donors was screened for spontaneous T-cell reactivity against the peptide in IFN-¿ ELISPOT assays. Patients with breast cancer, malignant melanoma, or renal cell carcinoma hosted powerful and high-frequency T-cell responses against the peptide. In addition......, when blood from healthy donors was tested, similar responses were observed. Ultimately, serum from cancer patients and healthy donors was analyzed for anti-cyclin B1 antibodies. Humoral responses against cyclin B1 were frequently detected in both cancer patients and healthy donors. In conclusion...

  16. Cationic influenza virosomes as an adjuvanted delivery system for CTL induction by DNA vaccination

    NARCIS (Netherlands)

    Jamali, Abbas; Holtrop, Marijke; de Haan, Aalzen; Hashemi, Hamidreza; Shenagari, Mohammad; Memarnejadian, Arash; Roohvand, Farzin; Sabahi, Farzaneh; Kheiri, Masumeh Tavassoti; Huckriede, Anke

    2012-01-01

    DNA vaccines have emerged as an attractive approach to induce CTL responses against cancer and infectious agents in recent years. Although CTL induction by DNA vaccination would be a valuable strategy for controlling viral infections, increasing the potency of DNA vaccines is mandatory before DNA

  17. CD3 directed bispecific antibodies induce increased lymphocyte-endothelial cell interactions in vitro

    NARCIS (Netherlands)

    Molema, G; Tervaert, JWC; Kroesen, BJ; Helfrich, W; Meijer, DKF; de Leij, LFMH

    2000-01-01

    Bispecific antibody (BsMAb) BIS-1 has been developed to redirect the cytolytic activity of cytotoxic T lymphocytes (CTL) to epithelial glycoprotein-2 (EGP-2) expressing tumour cells; intravenous administration of BIS-1 F(ab')(2) to carcinoma patients in a phase I/II clinical trial, caused immunomodu

  18. Strategi Bisnis pada PT CTL Dengan Pendekatan Metode Tows

    Directory of Open Access Journals (Sweden)

    Tjia Fie Tjoe

    2010-11-01

    Full Text Available The purpose of this research is determining direction of the correct business strategy to be applied by PT CTL, a garment company producing cloth for men. Research method used by the author is descriptive analysis with a case study research method. Research is conducted by collecting data obtained through survey by interview and giving questionnaire to all staff and head and also observation by evaluating directly the research object and also through literature study. Data analysis is conducted through input phase by using IFAS and EFAS matrix, adaptation phase with TOWS diagram, TOWS matrix and Internal-External matrix, and also uses SPACE matrix and also BCG matrix to analyse company's finance situation. Based on the conducted analysist the recommended corporation level strategy to be used by the company is diversification strategy direct to growth and stability. 

  19. Chemokines, lymphocytes, and HIV

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    Farber J.M.

    1998-01-01

    Full Text Available Chemokines are members of a family of more than 30 human cytokines whose best-described activities are as chemotactic factors for leukocytes and that are presumed to be important in leukocyte recruitment and trafficking. While many chemokines can act on lymphocytes, the roles of chemokines and their receptors in lymphocyte biology are poorly understood. The recent discoveries that chemokines can suppress infection by HIV-1 and that chemokine receptors serve, along with CD4, as obligate co-receptors for HIV-1 entry have lent urgency to studies on the relationships between chemokines and lymphocytes. My laboratory has characterized Mig and Crg-2/IP-10, chemokines that are induced by IFN-g and that specifically target lymphocytes, particularly activated T cells. We have demonstrated that the genes for these chemokines are widely expressed during experimental infections in mice with protozoan and viral pathogens, but that the patterns of mig and crg-2 expression differed, suggesting non-redundant roles in vivo. Our related studies to identify new chemokine receptors from activated lymphocytes resulted in the cloning of STRL22 and STRL33. We and others have shown that STRL22 is a receptor for the CC chemokine MIP-3a, and STRL22 has been re-named CCR6. Although STRL33 remains an orphan receptor, we have shown that it can function as a co-receptor for HIV-1 envelope glycoproteins, and that it is active with a broader range of HIV-1 envelope glycoproteins than the major co-receptors described to date. The ability of STRL33 to function with a wide variety of envelope glycoproteins may become particularly important if therapies are instituted to block other specific co-receptors. We presume that investigations into the roles of chemokines and their receptors in lymphocyte biology will provide information important for understanding the pathogenesis of AIDS and for manipulating immune and inflammatory responses for clinical benefit

  20. CD30 Is Dispensable for T-Cell Responses to Influenza Virus and Lymphocytic Choriomeningitis Virus Clone 13 but Contributes to Age-Associated T-Cell Expansion in Mice

    Directory of Open Access Journals (Sweden)

    Angela C. Zhou

    2017-09-01

    Full Text Available CD30 is a tumor necrosis factor receptor (TNFR family member whose expression is associated with Hodgkin’s disease, anaplastic large cell lymphomas, and other T and B lymphoproliferative disorders in humans. A limited number of studies have assessed the physiological role of CD30/CD30 ligand interactions in control of infection in mice. Here, we assess the role of CD30 in T-cell immunity to acute influenza and chronic lymphocytic choriomeningitis virus (LCMV clone 13 infection, two viral infections in which other members of the TNFR superfamily are important for T-cell responses. We show that CD30 is expressed on activated but not resting CD4 and CD8 T cells in vitro, as well as on regulatory T cells and marginally on T helper 1 cells in vivo during influenza infection. Despite this, CD4 and CD8 T-cell expansion in response to influenza virus was comparable in CD30+/+ and CD30−/− littermates, with no discernable role for the pathway in the outcome of influenza infection. Similarly, during persistent infection with LCMV clone 13, CD30 plays no obvious role in CD4 or CD8 T-cell responses, the level of T-cell exhaustion or viral control. In contrast, in the steady state, we observed increased numbers of total CD4 and CD8 T cells as well as increased numbers of regulatory T cells in unimmunized older (~8 months CD30+/+ but not in CD30−/− age-matched littermates. Naive T-cell numbers were unchanged in the aged CD30+/+ mice compared to their CD30−/− littermate controls, rather the T-cell expansions were explained by an increase in CD4+ and CD8+ CD44mid-hiCD62L− effector memory cells, with a similar trend in the central memory T-cell compartment. In contrast, CD30 did not impact the numbers of T cells in young mice. These data suggest a role for CD30 in the homeostatic regulation of T cells during aging, contributing to memory T-cell expansions, which may have relevance for CD30 expression in human T-cell lymphoproliferative diseases.

  1. Major Histocompatibility Complex-Dependent Cytotoxic T Lymphocyte Repertoire and Functional Avidity Contribute to Strain-Specific Disease Susceptibility after Murine Respiratory Syncytial Virus Infection ▿

    Science.gov (United States)

    Jessen, Birthe; Faller, Simone; Krempl, Christine D.; Ehl, Stephan

    2011-01-01

    Susceptibility to respiratory syncytial virus (RSV) infection in mice is genetically determined. While RSV causes little pathology in C57BL/6 mice, pulmonary inflammation and weight loss occur in BALB/c mice. Using major histocompatibility complex (MHC)-congenic mice, we observed that the H-2d allele can partially transfer disease susceptibility to C57BL/6 mice. This was not explained by altered viral elimination or differences in the magnitude of the overall virus-specific cytotoxic T lymphocyte (CTL) response. However, H-2d mice showed a more focused response, with 70% of virus-specific CTL representing Vβ8.2+ CTL directed against the immunodominant epitope M2-1 82, while in H-2b mice only 20% of antiviral CTL were Vβ9+ CTL specific for the immunodominant epitope M187. The immunodominant H-2d-restricted CTL lysed target cells less efficiently than the immunodominant H-2b CTL, probably contributing to prolonged CTL stimulation and cytokine-mediated immunopathology. Accordingly, reduction of dominance of the M2-1 82-specific CTL population by introduction of an M187 response in the F1 generation of a C57BL/6N × C57BL/6-H-2d mating (C57BL/6-H-2dxb mice) attenuated disease. Moreover, disease in H-2d mice was less pronounced after infection with an RSV mutant failing to activate M2-1 82-specific CTL or after depletion of Vβ8.2+ cells. These data illustrate how the MHC-determined diversity and functional avidity of CTL responses contribute to disease susceptibility after viral infection. PMID:21795345

  2. Human infection with Trypanosoma cruzi induces parasite antigen-specific cytotoxic T lymphocyte responses.

    Science.gov (United States)

    Wizel, B; Palmieri, M; Mendoza, C; Arana, B; Sidney, J; Sette, A; Tarleton, R

    1998-09-01

    Experimental models of Chagas' disease, an infection caused by the intracellular protozoan Trypanosoma cruzi, have demonstrated the crucial immunoprotective role played by CD8(+) T lymphocytes. These cells dominate inflammatory foci in parasitized tissues and their elimination from mice leads to uncontrolled parasite replication and subsequent death of the infected host. A trypomastigote surface antigen, TSA-1, and two amastigote surface molecules, ASP-1 and ASP-2, were recently identified as targets of CD8(+) cytotoxic T lymphocytes (CTL) in T. cruzi-infected mice. Until now, however, there was no evidence for the development of parasite-specific CTL in T. cruzi-infected humans. In this study, human CTL specific for TSA-1-, ASP-1-, and ASP-2-derived peptides were detected in the peripheral blood mononuclear cells from 21 of 24 HLA-A2(+) T. cruzi-infected patients. CTL recognition was antigen specific, A2-restricted, and CD8(+) T cell-dependent. Demonstration of human CTL against T. cruzi and against target molecules identified using the murine model provides important information for the optimal design and evaluation of vaccines to prevent or ameliorate Chagas' disease.

  3. Monitoring of WT1-specific cytotoxic T lymphocytes after allogeneic hematopoietic stem cell transplantation.

    Science.gov (United States)

    Morita, Yuriko; Heike, Yuji; Kawakami, Mami; Miura, Osamu; Nakatsuka, Shin-Ichi; Ebisawa, Michiko; Mori, Shin-Ichiro; Tanosaki, Ryuji; Fukuda, Takahiro; Kim, Sung-Won; Tobinai, Kensei; Takaue, Yoichi

    2006-09-15

    Donor-derived cytotoxic T lymphocytes (CTL) that respond to tumor antigens emerge after hematopoietic stem cell transplantation (HSCT), particularly in association with the status of immune recovery. To analyze the frequency of CTL against PR1, PRAME and WT1 after HSCT, a tetramer-based analysis was performed in 97 samples taken from 35 patients (9 AML, 11 MDS, 2 CML, 4 ALL, 7 lymphoma and 2 renal cell carcinoma [RCC]) with the HLA-A02 phenotype. Regarding PR1, only 1 sample showed the presence of tetramer-positive cells (0.04%/lymphocyte). Similarly, in PRAME, only 10 of 97 samples were sporadically positive with low titers. For WT1, positive results were detected in 39 of 97 samples and 7 (2 CML, 1 ALL, 2 lymphoma and 2 RCC) patients clearly showed positive results more than once. On the basis of these results, we performed serial analyses of WT1-specific CTL during the clinical course in 2 patients with RCC, who underwent HSCT with a reduced-intensity regimen, to examine the precise correlation between the kinetics of CTL, the occurrence of GVHD and the observed clinical response. A higher positive rate for WT1-specific CTL and a correlation with the clinical response suggest that WT1 may be a useful antigen for a wider monitoring application.

  4. Identification of the epitopes on HCV core protein recognized by HLA-A2 restricted cytotoxic T lymphocytes

    Institute of Scientific and Technical Information of China (English)

    Hong-Chao Zhou; De-Zhong Xu; Xue-Ping Wang; Jing-Xia Zhang; Ying-Huang; Yong-Ping Yan; Yong Zhu; Bo-Quan Jin

    2001-01-01

    AIM To identify hepatitis C virus (HCV) core protein epitopes recognized by HLA-A2 restricted cytotoxic T lymphocyte (CTL). METHODS Utilizing the method of computer prediction followed by a 4 h 51 Cr-release assay confirmation. RESULTS The results showed that peripheral blood mononuclear cells (PBMC) obtained from two HLA-A2 positive donors who were infected with HCV could lyse autologous target cells labeled with peptide "ALAHGVFAL (core TS0-158)".The rates of specific lysis of the cells from the two donors were 37.5% and 15.8%,respectively. Blocking of the CTL response with anti-CD4 mAb caused no significant decrease of the specific lysis.But blocking of CTL response with anti-CD8 mAb could abolish the Iysis. CONCLUSION The peptide (core 150 - 158 ) is the candidate epitope recognized by HLA-A2 restricted CTL.

  5. Generation of cytotoxic T lymphocytes specific for B-cell acute lymphoblastic leukemia family-shared peptides derived from immunoglobulin heavy chain framework region

    Institute of Scientific and Technical Information of China (English)

    LIU Ying; ZHU Ping; HU Ya-mei

    2007-01-01

    Background Immunoglobulin heavy chain variable region (IgHV) is a well-characterized tumor antigen for B-cell malignancies. It can function as a target for T cell-mediated immune response. Clinical trials of IgHV protein vaccines against lymphoma have demonstrated induction of tumor-specific cytotoxic T lymphocyte (CTL) responses. However,complementary determining regions-based individual vaccines have disadvantages for wide clinical application. Although a recent study demonstrated that immunogenic peptides are derived from framework regions (FR) shared among patients with B-cell lymphoma, how to choose the appropriate peptides for each patient is still unsolved. The aim of this study was to investigate whether immunoglobulin heavy chain FR-derived peptides shared in each IgHV family are potential CTL epitopes presented by B-cell acute lymphoblastic leukemia (B-ALL). Such CTL epitopes might be beneficial to shifting vaccination strategies against B-ALL from individual specificity to family specificity.Methods Seven IgHV gene families were amplified respectively by PCR and sequenced directly from 71 childhood B-ALL cases. Bioinformatics was applied in analyzing characteristics of sequences available and predicting HLA-A*0201-restricted CTL epitopes for each IgHV family. An antigen-specific T cell expansion system was used to generate peptide-specific CTLs. The cytotoxicity of CTLs against B-ALL cells was assessed in the lactate dehydrogenase release assay.Results Complete IgHV rearrangements were identified in all of the 71 B-ALL cases. All of 40 sequences available showed ≥98% homology with the nearest germline IgHV genes, indicating IgHV genes in B-ALL of germline nature.Twelve nonapeptides of high HLA-A*0201-binding scores were obtained from 26 productive IgHV protein sequences. Ten (83%) of the peptides were located in FR1 and FR3 shared among the corresponding IgHV family. CTLs specific for the peptide QLVQSGAEV located in FR1 (3-11) shared among the IgHV1

  6. Symbolic Model Checking Algorithm for Temporal Epistemic Logic CTL * K%时态认知逻辑CTL*K的符号化模型检查算法

    Institute of Scientific and Technical Information of China (English)

    陈彬; 王智学

    2009-01-01

    时序认知逻辑是由时序逻辑和认知逻辑组合而成的逻辑,主要应用于多主体系统的规范定义.大多数时序认知逻辑是基于CTL的,表达能力有限.并且已知的一些模型检查算法存在内存不足和状态爆炸等问题.讨论了基于CTL*的时态认知逻辑cTL*K的语法、语义和模型,它能够在表达力很强的时态逻辑CTL*基础上描述智能体的知识、目标等意向特征.并给出了CTL*K的模型检查算法,其核心思想就是将CTL*K公式的检查问题转化为CTL*公式的模型检查问题,可以使检查的系统规模得以大幅度提高.并且将算法编码后容易集成到NuSMV模型检查器.

  7. Cytotoxic T lymphocyte activity in children infected with HIV.

    Science.gov (United States)

    Froebel, K S; Aldhous, M C; Mok, J Y; Hayley, J; Arnott, M; Peutherer, J F

    1994-01-01

    Of the Edinburgh cohort of approximately 130 children born to HIV-infected women, 9 are infected and alive. This article describes results from the first 18 months of a natural history study of seven of these, and two adopted children, studying the CD8 T cell-mediated cytotoxicity against HIV proteins (Gag, Tat, Pol, and Env), over time, and relating it to clinical progression and viral activity. Autologous EBV cell lines infected with vaccinia-HIV constructs were used as target cells, and bulk-cultured peripheral blood mononuclear cells as effector cells. The children ranged in age from 0 to 93 months, with six of the nine showing CTL activity to one or more HIV proteins. The specificity of the response was directed against Tat in the younger children, switching to Pol, then Gag or Env. Preliminary analysis of virological data showed no association between CTL and virus activity. The children with CTLs tended to be well clinically, but the cohort needs to be studied longer before conclusions can be made about CTL activity and HIV disease progression. Cytotoxic T lymphocyte activity has also been observed in two children diagnosed as HIV uninfected. These results show the importance of looking at CTL specificity, and may have implications in vaccine design.

  8. Why do cytotoxic T lymphocytes fail to eliminate hepatitis C virus? Lessons from studies using major histocompatibility complex class I peptide tetramers.

    Science.gov (United States)

    Lechner, F; Sullivan, J; Spiegel, H; Nixon, D F; Ferrari, B; Davis, A; Borkowsky, B; Pollack, H; Barnes, E; Dusheiko, G; Klenerman, P

    2000-08-29

    Hepatitis C virus (HCV) infection is a major public health problem, affecting an estimated 3% of the world's population, and over 10% in some countries. Infection in most cases becomes persistent, and can lead to hepatic inflammation, fibrosis and liver failure. The T lymphocyte reponse, in particular that mediated by cytotoxic T lymphocytes (CTLs), is likely to be involved in determining the outcome of infection, although its overall role is not clear. The use of major histocompatibility complex (MHC) class I peptide tetrameric complexes (tetramers) to study antiviral CTL responses has revolutionized our approach to the study of human infection. We have used a panel of MHC class I tetramers to analyse immune responses in HCV-infected individuals at various stages of disease. We find that the CTL response against HCV is vigorous in its early phases but dwindles over time both in terms of lymphocyte number and function. A number of potential explanations for this 'CTL failure' are discussed.

  9. Building blocks for institutional preparation of CTL019 delivery.

    Science.gov (United States)

    McGuirk, Joseph; Waller, Edmund K; Qayed, Muna; Abhyankar, Sunil; Ericson, Solveig; Holman, Peter; Keir, Christopher; Myers, G Douglas

    2017-09-01

    Chimeric antigen receptor (CAR) T-cell therapy is an investigational immunocellular therapy that reprograms a patient's cytotoxic T cells to engage and eliminate malignant cells. CAR T-cell therapies targeting the CD19 antigen have demonstrated high efficacy in clinical trials for patients with B-cell malignancies and may potentially be available on a broader scale in the future. CAR T-cell therapy begins with the collection of a sufficient number of T cells from a patient's peripheral blood through leukapheresis. Several factors must be considered when patients undergo leukapheresis for CAR T-cell therapy, including age and prior therapies. The leukapheresis material is shipped to a manufacturing facility, followed by return of the CAR T cells to the treatment center. Careful coordination of a multidisciplinary team composed of physicians, nurses, pharmacists and other hospital personnel is critical for the proper care of the patient before, during and after CAR T-cell therapy. CAR T-cell therapy has been associated with adverse events (AEs) such as cytokine release syndrome, which requires rapid attention by the emergency department, intensive care unit and hospital pharmacy. In this review, we discuss several aspects of institutional preparation for leukapheresis, CAR T-cell infusion and AE management based on our experience with clinical trials of the CD19 CAR T-cell therapy CTL019. Copyright © 2017 International Society for Cellular Therapy. Published by Elsevier Inc. All rights reserved.

  10. Immune-mediated bone marrow failure syndromes of progenitor and stem cells: molecular analysis of cytotoxic T cell clones.

    Directory of Open Access Journals (Sweden)

    Ramon Tiu

    2007-03-01

    Full Text Available The unique structure of the T cell receptor (TCR enables molecular identification of individual T cell clones and provides an unique opportunity for the design of molecular diagnostic tests based on the structure of the rearranged TCR chain e.g., using the TCR CDR3 region. Initially, clonal T cell malignancies, including T cell large granular lymphocyte leukemia (T-LGL, mucosis fungoides and peripheral T cell lymphoma were targets for the TCR-based analytic assays such as detection of clonality by T-gamma rearrangement using y-chain-specific PCR or Southern Blotting. Study of these disorders facilitated further analytic concepts and application of rational methods of TCR analysis to investigations of polyclonal T cell-mediated diseases. In hematology, such conditions include graft versus host disease (GvHD and immune-mediated bone marrow failure syndromes. In aplastic anemia (AA, myelodysplastic syndrome (MDS or paroxysmal nocturnal hemoglobinuria (PNH, cytotoxic T cell responses may be directed against certain antigens located on stem or more lineage-restricted progenitor cells in single lineage cytopenias. The nature of the antigenic targets driving polyclonal CTL responses remains unclear. Novel methods of TCR repertoire analysis, include VB flow cytometry, peptide-specific tetramer staining, in vitro stimulation assays and TCR CDR3-specific PCR. Such PCR assay can be either VB family-specific or multiplexed for all VB families. Amplified products can be characterized and quantitated to facilitate detection of the most immunodominant clonotypes. Such clonotypes may serve as markers for the global polyclonal T cell response. Identification of these clonotypes can be performed in blood and tissue biopsy material by various methods. Once immunodominant clonotypes corresponding to pathogenic CTL clones are identified they can serve as surrogate markers for the activity of the pathophysiologic process or even indicate the presence of specific

  11. Efficient lysis of epithelial ovarian cancer cells by MAGE-A3-induced cytotoxic T lymphocytes using rAAV-6 capsid mutant vector.

    Science.gov (United States)

    Batchu, Ramesh B; Gruzdyn, Oksana V; Moreno-Bost, Amberly M; Szmania, Susann; Jayandharan, Giridhararao; Srivastava, Arun; Kolli, Bala K; Weaver, Donald W; van Rhee, Frits; Gruber, Scott A

    2014-02-12

    MAGE-A3 is highly expressed in epithelial ovarian cancer (EOC), making it a promising candidate for immunotherapy. We investigated whether dendritic cells (DCs) transduced with a rAAV-6 capsid mutant vector Y445F could elicit effective MAGE-A3-specific anti-tumor cytotoxic T lymphocyte (CTL) responses in vitro. MAGE-A3 was cloned and rAAV-6-MAGE-A3 purified, followed by proviral genome detection using real-time PCR. Immunofluorescence detection of rAAV-6-Y445F-MAGE-A3-transduced DCs demonstrated 60% transduction efficiency. Fluorescent in situ hybridization analysis confirmed chromosomal integration of rAAV vectors. Flow cytometric analysis of transduced DCs showed unaltered expression of critical monocyte-derived surface molecules with retention of allo-stimulatory activity. Co-culture of autologous T lymphocytes with MAGE-A3-expressing DCs produced CTLs that secreted IFN-γ, and efficiently killed MAGE-A3+ EOC cells. This form of rAAV-based DC immunotherapy, either alone or more likely in combination with other immune-enhancing protocols, may prove useful in the clinical setting for management of EOC.

  12. What is Cloning?

    Science.gov (United States)

    Donate Home Cloning What is Cloning What is Cloning Clones are organisms that are exact genetic copies. ... clones made through modern cloning technologies. How Is Cloning Done? Many people first heard of cloning when ...

  13. Prior population immunity reduces the expected impact of CTL-inducing vaccines for pandemic influenza control.

    Directory of Open Access Journals (Sweden)

    Kirsty J Bolton

    Full Text Available Vaccines that trigger an influenza-specific cytotoxic T cell (CTL response may aid pandemic control by limiting the transmission of novel influenza A viruses (IAV. We consider interventions with hypothetical CTL-inducing vaccines in a range of epidemiologically plausible pandemic scenarios. We estimate the achievable reduction in the attack rate, and, by adopting a model linking epidemic progression to the emergence of IAV variants, the opportunity for antigenic drift. We demonstrate that CTL-inducing vaccines have limited utility for modifying population-level outcomes if influenza-specific T cells found widely in adults already suppress transmission and prove difficult to enhance. Administration of CTL-inducing vaccines that are efficacious in "influenza-experienced" and "influenza-naive" hosts can likely slow transmission sufficiently to mitigate a moderate IAV pandemic. However if neutralising cross-reactive antibody to an emerging IAV are common in influenza-experienced hosts, as for the swine-variant H3N2v, boosting CTL immunity may be ineffective at reducing population spread, indicating that CTL-inducing vaccines are best used against novel subtypes such as H7N9. Unless vaccines cannot readily suppress transmission from infected hosts with naive T cell pools, targeting influenza-naive hosts is preferable. Such strategies are of enhanced benefit if naive hosts are typically intensively mixing children and when a subset of experienced hosts have pre-existing neutralising cross-reactive antibody. We show that CTL-inducing vaccination campaigns may have greater power to suppress antigenic drift than previously suggested, and targeting adults may be the optimal strategy to achieve this when the vaccination campaign does not have the power to curtail the attack rate. Our results highlight the need to design interventions based on pre-existing cellular immunity and knowledge of the host determinants of vaccine efficacy, and provide a framework

  14. Prior Population Immunity Reduces the Expected Impact of CTL-Inducing Vaccines for Pandemic Influenza Control

    Science.gov (United States)

    Bolton, Kirsty J.; McCaw, James M.; Brown, Lorena; Jackson, David; Kedzierska, Katherine; McVernon, Jodie

    2015-01-01

    Vaccines that trigger an influenza-specific cytotoxic T cell (CTL) response may aid pandemic control by limiting the transmission of novel influenza A viruses (IAV). We consider interventions with hypothetical CTL-inducing vaccines in a range of epidemiologically plausible pandemic scenarios. We estimate the achievable reduction in the attack rate, and, by adopting a model linking epidemic progression to the emergence of IAV variants, the opportunity for antigenic drift. We demonstrate that CTL-inducing vaccines have limited utility for modifying population-level outcomes if influenza-specific T cells found widely in adults already suppress transmission and prove difficult to enhance. Administration of CTL-inducing vaccines that are efficacious in "influenza-experienced" and "influenza-naive" hosts can likely slow transmission sufficiently to mitigate a moderate IAV pandemic. However if neutralising cross-reactive antibody to an emerging IAV are common in influenza-experienced hosts, as for the swine-variant H3N2v, boosting CTL immunity may be ineffective at reducing population spread, indicating that CTL-inducing vaccines are best used against novel subtypes such as H7N9. Unless vaccines cannot readily suppress transmission from infected hosts with naive T cell pools, targeting influenza-naive hosts is preferable. Such strategies are of enhanced benefit if naive hosts are typically intensively mixing children and when a subset of experienced hosts have pre-existing neutralising cross-reactive antibody. We show that CTL-inducing vaccination campaigns may have greater power to suppress antigenic drift than previously suggested, and targeting adults may be the optimal strategy to achieve this when the vaccination campaign does not have the power to curtail the attack rate. Our results highlight the need to design interventions based on pre-existing cellular immunity and knowledge of the host determinants of vaccine efficacy, and provide a framework for assessing the

  15. Consequences of cytotoxic T lymphocyte interaction with major histocompatibility complex class I-expressing neurons in vivo.

    Science.gov (United States)

    Rall, G F; Mucke, L; Oldstone, M B

    1995-11-01

    Neurons have evolved strategies to evade immune surveillance that include an inability to synthesize the heavy chain of the class I major histocompatibility complex (MHC), proteins that are necessary for cytotoxic T lymphocyte (CTL) recognition of target cells. Multiple viruses have taken advantage of the lack of CTL-mediated recognition and killing of neurons by establishing persistent neuronal infections and thereby escaping attack by antiviral CTL. We have expressed a class I MHC molecule (Db) in neurons of transgenic mice using the neuron-specific enolase (NSE) promoter to determine the pathogenic consequences of CTL recognition of virally infected, MHC-expressing central nervous system (CNS) neurons. The NSE-Db transgene was expressed in H-2b founder mice, and transgene-derived messenger RNA was detected by reverse transcriptase-polymerase chain reaction in transgenic brains from several lines. Purified primary neurons from transgenic but not from nontransgenic mice adhered to coverslips coated with a conformation-dependent monoclonal antibody directed against the Dv molecule and presented viral peptide to CTL in an MHC-restricted manner, indicating that the Db molecule was expressed on transgenic neurons in a functional form. Transgenic mice infected with the neurotropic lymphocytic choriomeningitis virus (LCMV) and given anti-LCMV, MHC-restricted CTL displayed a high morbidity and mortality when compared with controls receiving MHC-mismatched CTL or expressing alternative transgenes. After CTL transfer, transgenic brains showed an increased number of CD8+ cells compared with nontransgenic controls as well as an increased rate of clearance of infectious virus from the CNS. Additionally, an increase in blood-brain barrier permeability was detected during viral clearance in NSE-Db transgenic mice and lasted several months after clearance of virus from neurons. In contrast, LCMV-infected, nontransgenic littermates and mice expressing other gene products from the

  16. Function of Helper T Cells in the Memory CTL-mediated Anti-tumor Immunity

    Institute of Scientific and Technical Information of China (English)

    高丰光; GermainJ.P.Fernendo; 刘文军

    2004-01-01

    Abstract To investigate the role of CD4+ helper T (Th) cells in the memory CTL-mediated anti-tumor immunity, the RAG-1 gene knock out mice were adoptively transferred with OT-1 cells to generate the memory CTL, the C57B1/6 mice immunized with the epitope peptide of OVA specific Th cells and with different adjuvants were adopfively transferred with these memory-CTLs, and then the animals were challenged with tumor cells EGT. It was found that although the simple immunization of mice with the epitope peptide of the OVA specific Th cells could generate more effect CTL, but this effect was not so strong enough to resist completely the challenges with tumor cells. Nevertheless, the memory CTL-mediated anti-tumor immune effect required the helps of Th1 and Th2 cells. The cross-regulation between Thl and Th2 cells seemed to be beneficial for the host to generate more effector CTL for mounting an efficient anti-tumor response. It concluded that the interaction between Thl and Th2 cells might be more important than the single subset of Th cells in the memory CTL-mediated anti-tumor immune response. More attention should be paid in this regard for the future studies.

  17. Recombinant poxvirus boosting of DNA-primed rhesus monkeys augments peak but not memory T lymphocyte responses

    OpenAIRE

    Santra, Sampa; Barouch, Dan H.; Korioth-Schmitz, Birgit; Lord, Carol I.; Krivulka, Georgia R.; Yu, Faye; Beddall, Margaret H; Gorgone, Darci A.; Lifton, Michelle A.; Miura, Ayako; Philippon, Valerie; Manson, Kelledy; Markham, Phillip D.; Parrish, John; Kuroda, Marcelo J.

    2004-01-01

    Although a consensus has emerged that an HIV vaccine should elicit a cytotoxic T lymphocyte (CTL) response, the characteristics of an effective vaccine-induced T lymphocyte response remain unclear. We explored this issue in the simian human immunodeficiency virus/rhesus monkey model in the course of assessing the relative immunogenicity of vaccine regimens that included a cytokine-augmented plasmid DNA prime and a boost with DNA or recombinant pox vectors. Recombinant vaccinia virus, recombin...

  18. Mathematical Modeling of Cytotoxic Lymphocyte-Mediated Immune Response to Hepatitis B Virus Infection

    Directory of Open Access Journals (Sweden)

    Changjiang Long

    2008-01-01

    Full Text Available Nowak's model of the human immunodeficiency virus (HIV infection has been extensively and successfully used to simulate the interaction between HIV and cytotoxic lymphocyte- (CTL- mediated immune response. However, this model is not available for hepatitis B virus (HBV infection. As the enhanced recruitment of virus-specific CTLs into the liver has been an important novel concept in the pathogenesis of hepatitis B, we develop a specific mathematical model analyzing the relationship between HBV and the CTL-mediated immune response, and the indicator of the liver cell damage, alanine aminotransferase (ALT. The stability condition of the complete recovery equilibrium point at which HBV will be eliminated entirely from the body is discussed. A different set of parameters is used in the simulation, and the results show that the model can interpret the wide variety of clinical manifestations of HBV infection. The model suggests that a rapid and vigorous CTL response is required for resolution of HBV infection.

  19. Posttranscriptional clearance of hepatitis B virus RNA by cytotoxic T lymphocyte-activated hepatocytes.

    Science.gov (United States)

    Tsui, L V; Guidotti, L G; Ishikawa, T; Chisari, F V

    1995-01-01

    Using transgenic mice that replicate the hepatitis B virus (HBV) genome, we recently demonstrated that class I-restricted, hepatitis B surface antigen-specific cytotoxic T lymphocytes (CTLs) can noncytolytically eliminate HBV pregenomic and envelope RNA transcripts from the hepatocyte. We now demonstrate that the steady-state content of these viral transcripts is profoundly reduced in the nucleus and cytoplasm of CTL-activated hepatocytes, but their transcription rates are only slightly reduced. Additionally, we demonstrate that transcripts covering the HBV X coding region are resistant to downregulation by the CTL. These results imply the existence of CTL-inducible hepatocellular factors that interact with a discrete element(s) between nucleotides 3157 and 1239 within the viral pregenomic and envelope transcripts and mediate their degradation, thus converting the hepatocyte from a passive victim to an active participant in the host response to HBV infection. Images Fig. 2 Fig. 3 Fig. 4 Fig. 5 PMID:8618909

  20. Induction of systemic CTL responses in melanoma patients by dendritic cell vaccination: Cessation of CTL responses is associated with disease progression

    DEFF Research Database (Denmark)

    Andersen, M.H.; Keikavoussi, P.; Brocker, E.B.

    2001-01-01

    by Western blotting was decreased in PBL at this time. In summary, our data confirm that DC-based vaccinations induce peptide-specific T cells in the peripheral blood of advanced-stage melanoma patients. Although successful induction of systemic tumor antigen-specific CTL may not lead to objective clinical...

  1. Chimeric flagellin expressed by Salmonella typhimurium induces an ESAT-6-specific Th 1-type immune response and CTL effects following intranasal immunization

    Institute of Scientific and Technical Information of China (English)

    Hui Zhang; Liu Liu; Ke Wen; Jinlin Huang; Shizhong Geng; Junsong Shen; Zhiming Pan; Xinan Jiao

    2011-01-01

    The flagellin component FliC of Salmonella typhimurium is capable of activating the innate immune system via specific interactions with TLR5 and can also act as a carrier of foreign antigen to elicit antigen-specific immune responses.Thus,we constructed an attenuated Salmonella strain SL5928(fliC/esat) expressing chimeric flagellin that contained the ESAT-6 antigen coding sequence of Mycobacterium tuberculosis inserted into the highly variable region of the Salmonella flagellin coding gene fliCi.The chimeric flagellin functioned normally,as demonstrated using a flagella swarming assay and electron microscopy.To analyze the effects of chimeric flagellin,the cell-mediated immune response and cytotoxic T lymphocyte (CTL) effects specific for ESAT-6antigen were tested after intranasal immunization of mice with flagellated Salmonella SL5928(fliC/esat).The results showed that SL5928(fliC/esat) intranasal immunization can strongly elicit an ESAT-6-specific T helper (Th) 1-type immune response in mucosal lymphoid tissues,such as nasopharynx-associated lymph nodes,lung and Peyer's patches,and a Th 1/Th2 response was elicited in spleen and mesenteric lymph nodes.Furthermore,intranasal immunization of SL5928(fliC/esat) produced efficient CTL effects,as demonstrated using a 5-and 6-carboxyfluorescein diacetate succinimidyl ester (CFSE) assay.Thus,our study revealed that Salmonella flagellin acts as a carrier for foreign antigen and triggers strong Th 1 and CTL responses during intranasal immunization.Chimeric flagellin is potentially an effective strategy for the development of novel vaccines against tuberculosis in humans and animals.

  2. Quantum cloning

    OpenAIRE

    Scarani, Valerio; Iblisdir, Sofyan; Gisin, Nicolas; Acin, Antonio

    2005-01-01

    The impossibility of perfectly copying (or cloning) an arbitrary quantum state is one of the basic rules governing the physics of quantum systems. The processes that perform the optimal approximate cloning have been found in many cases. These "quantum cloning machines" are important tools for studying a wide variety of tasks, e.g. state estimation and eavesdropping on quantum cryptography. This paper provides a comprehensive review of quantum cloning machines (both for discrete-dimensional an...

  3. Construction and immunogenicity prediction of Plasmodium falciparum CTL epitope minigene vaccine

    Institute of Scientific and Technical Information of China (English)

    2001-01-01

    The minigenes encoding Plasmodium falciparum CTL epitopesrestricted to human MHC class I molecular HLA-A2 and HLA-B51, which were both at high frequency among Chinese population, were constructed as mono-epitope CTL vaccines named pcDNA3.1/tr and pcDNA3.1/ sh. The minigenes of the two epitopes were then tandem linked to form a dimeric CTL epitope minigene recombinant vaccine. After DNA transfection, the epitope minigenes were expressed respectively in two human cell lines, each bearing one MHC class I molecule named CIR/HLA-A2.1 and K562/HLA-B51. The intracellular expression of the CTL epitope minigenes not only enhanced the stability of HLA-A2.1 and HLA-B51 molecules but also increased the assemblage of MHC class I molecules on cell surfaces, which testified the specific process and presentation of those endogenous expressed epitopes. For the cells transfected with the dimeric minigene encoding two tandem linked epitopes, the expression and presentation of each epitope were also detected on cell membranes that bore different MHC class I molecules. It meant that the adjacency of the two CTL epitopes did not interfere with the specific process and presentation of each epitope. Compared with the ordinary CTL studies that inoculated synthesized epitope peptides with peripheral blood cells, this work aimed to process the epitopes directly inside HLA class I allele specific human cells, and thus theoretically imitated the same procedure in vivo. It was also an economical way to predict the immunogenicity of CTL epitopes at an early stage especially in laboratories with limited financial resource.

  4. A strategy for efficient cross-presentation of CTL-epitope peptides leading to enhanced induction of in vivo tumor immunity.

    Science.gov (United States)

    Hayashi, Akira; Wakita, Hisashi; Yoshikawa, Tomoaki; Nakanishi, Tsuyoshi; Tsutsumi, Yasuo; Mayumi, Tadanori; Mukai, Yohei; Yoshioka, Yasuo; Okada, Naoki; Nakagawa, Shinsaku

    2007-01-22

    The activation of antitumor cytotoxic T-lymphocytes (CTLs) depends on how efficiently the relevant tumor antigen peptides are delivered into the major histocompatibility complex (MHC) class I presentation pathway in antigen presenting cells (APCs). An elegant approach to promote the peptide-MHC class I association has been described for enhanced peptide transportation into the endoplasmic reticulum (ER) by adding an ER insertion signal sequence (Eriss). Nevertheless, this approach does not appear potent enough to induce in vivo tumor protective immunity. Herein, we present a novel peptide-vaccine strategy based on the combined utilization of Eriss and fusogenic liposomes (FLs) capable of directly introducing encapsulated CTL-epitope peptides into the MHC class I pathway of APCs. APCs pulsed with free peptides, FL-encapsulated peptides, or FL-encapsulated Eriss-conjugated peptides exhibited comparable levels of antigen-presenting activity at early phases after pulsing. Interestingly, whereas in the first two methods the APC ability began to decline 40 to 60 h after pulsing, FL-encapsulated Eriss(+) peptides allowed APCs to retain peptide-presentation activity for at least 140 h. This advantage of FL-encapsulated Eriss(+) peptides correlated with the induction of more potent antitumor immunity compared with soluble Eriss(+) or Eriss(-) peptides or FL-encapsulated Eriss(-) peptides when they were administered in vivo. Thus, Eriss-conjugated CTL-epitope peptides encapsulated in FLs provide a highly efficient tumor-vaccine to enhance the induction of in vivo tumor immunity.

  5. Tissue Dimensionality Influences the Functional Response of Cytotoxic T Lymphocyte-Mediated Killing of Targets.

    Science.gov (United States)

    Gadhamsetty, Saikrishna; Marée, Athanasius F M; de Boer, Rob J; Beltman, Joost B

    2016-01-01

    Cytotoxic T lymphocyte (CTL)-mediated killing of virus infections and tumors occurs over a wide range of conditions. The spatial environments in which CTLs encounter target cells vary from narrow vessels, to two-dimensional epithelial tissues, to densely populated 3-dimensional (3D) T cell areas within lymphoid tissues. How such spatial environments alter the functional response of CTL-mediated killing, i.e., how the killing efficiency depends on cell densities, is unclear. In this study, we perform cellular Potts model simulations in different spatial configurations to investigate how the dimensionality of the space affects the functional response of CTL-mediated killing. Irrespective of the spatial configuration, the function with separate saturation constants for CTL and for target cell densities that we previously proposed can in all cases describe the response, demonstrating its generality. However, the tissue dimensionality determines at which cell densities the killing rate starts to saturate. We show that saturation in a fully 3D environment is stronger than in a "flat" 3D environment, which is largely due to accompanying differences in the CTL-target encounter rates.

  6. Syntaxin 8 is required for efficient lytic granule trafficking in cytotoxic T lymphocytes.

    Science.gov (United States)

    Bhat, Shruthi S; Friedmann, Kim S; Knörck, Arne; Hoxha, Cora; Leidinger, Petra; Backes, Christina; Meese, Eckart; Keller, Andreas; Rettig, Jens; Hoth, Markus; Qu, Bin; Schwarz, Eva C

    2016-07-01

    Cytotoxic T lymphocytes (CTL) eliminate pathogen-infected and cancerous cells mainly by polarized secretion of lytic granules (LG, containing cytotoxic molecules like perforin and granzymes) at the immunological synapse (IS). Members of the SNARE (soluble N-ethylmaleimide-sensitive factor attachment protein receptor) family are involved in trafficking (generation, transport and fusion) of vesicles at the IS. Syntaxin 8 (Stx8) is expressed in LG and colocalizes with the T cell receptor (TCR) upon IS formation. Here, we report the significance of Stx8 for human CTL cytotoxicity. We found that Stx8 mostly localized in late, recycling endosomal and lysosomal compartments with little expression in early endosomal compartments. Down-regulation of Stx8 by siRNA resulted in reduced cytotoxicity. We found that following perforin release of the pre-existing pool upon target cell contact, Stx8 down-regulated CTL regenerate perforin pools less efficiently and thus release less perforin compared to control CTL. CD107a degranulation, real-time and end-point population cytotoxicity assays, and high resolution microscopy support our conclusion that Stx8 is required for proper and timely sorting and trafficking of cytotoxic molecules to functional LG through the endosomal pathway in human CTL.

  7. DR3 regulation of apoptosis of naive T-lymphocytes in children with acute infectious mononucleosis.

    Science.gov (United States)

    Filatova, Elena Nikolaevna; Anisenkova, Elena Viktorovna; Presnyakova, Nataliya Borisovna; Utkin, Oleg Vladimirovich

    2016-09-01

    Acute infectious mononucleosis (AIM) is a widespread viral disease that mostly affects children. Development of AIM is accompanied by a change in the ratio of immune cells. This is provided by means of different biological processes including the regulation of apoptosis of naive T-cells. One of the potential regulators of apoptosis of T-lymphocytes is a death receptor 3 (DR3). We have studied the role of DR3 in the regulation of apoptosis of naive CD4(+) (nTh) and CD8(+) (nCTL) T-cells in healthy children and children with AIM. In healthy children as well as in children with AIM, the activation of DR3 is accompanied by inhibition of apoptosis of nTh. In healthy children, the stimulation of DR3 resulted in the increase in apoptosis of nCTL. On the contrary, in children with AIM, the level of apoptosis of nCTL decreased after DR3 activation, which is a positive contribution to the antiviral immune response. In children with AIM, nCTL are characterized by reduced level of apoptosis as compared with healthy children. These results indicate that DR3 can be involved in the reduction of sensitivity of nCTL to apoptosis in children with AIM.

  8. The progenitor cell inhibition assay to measure the anti-leukemic reactivity of T cell clones against acute and chronic myeloid leukemia.

    Science.gov (United States)

    van der Hoorn, M A W G; van Luxemburg-Heijs, S A P; van Bergen, C A M; Bongaerts, R; Willemze, R; Falkenburg, J H F

    2003-10-01

    Adoptive immunotherapy with donor T lymphocytes may be used as a treatment for relapsed leukemia after allogeneic hematopoietic stem cell transplantation (SCT). In vitro selected and expanded anti-leukemic T cells may be more effective in inducing a response in vivo. To identify the anti-leukemic reactivity of in vitro generated T cells, standard target cell read-out assays like the 51Cr-release assay are not always appropriate. We developed an assay in which the ability of T cells to antigen specifically inhibit the in vitro growth of leukemic progenitor cells in the presence of cytokines can be measured. This assay allows the evaluation of the cytolytic or suppressive potential of leukemia reactive T cells for prolonged periods of time. The assay is based on inhibition of [3H]thymidine incorporation by the leukemic progenitor cells induced by multiple hematopoietic growth factors. T cell clones with a known specificity were used to compare the analytic potential of the new assay with those of other cytotoxicity assays. Based on the results of the T cell clones, a modification of a limiting dilution assay was developed to identify anti-leukemic allogeneic T cells in HLA identical donor-recipient combinations selected on their ability to inhibit the in vitro growth of CML or AML progenitor cells, to be used for the generation of leukemia-reactive CTL lines for clinical use.

  9. Induction of cytotoxic T-cell responses by gene gun DNA vaccination with minigenes encoding influenza A virus HA and NP CTL-epitopes

    DEFF Research Database (Denmark)

    Fomsgaard, A; Nielsen, H V; Kirkby, N

    1999-01-01

    degree of controllability. We have examined the induction of murine CTL's by this approach using DNA plasmid minigene vaccines encoding known mouse K(k) minimal CTL epitopes (8 amino acids) from the influenza A virus hemagglutinin and nucleoprotein. We here report that such an approach is feasible...... and that wild type influenza virus flanking amino acid sequences can influence the CTL response but are not essential for optimal CTL induction. We also examined the effect of different new amino acid sequences flanking the CTL epitopes. In one version, two CTL epitopes were linked together as 'string of beads......-induced CTL responses and tested for their protective effect against a lethal influenza A virus infection in mice and no effect was found. We conclude that a specific and highly directed CTL induction is possible by unlinked minigene DNA immunisation, but that CTL induction solely is not always sufficient...

  10. Adoptive transfer of PR1 cytotoxic T lymphocytes associated with reduced leukemia burden in a mouse acute myeloid leukemia xenograft model.

    Science.gov (United States)

    Ma, Qing; Wang, Changqing; Jones, Dan; Quintanilla, Kathryn E; Li, Dan; Wang, Yang; Wieder, Eric D; Clise-Dwyer, Karen; Alatrash, Gheath; Mj, You; Munsell, Mark F; Lu, Sijie; Qazilbash, Muzaffar H; Molldrem, Jeffrey J

    2010-12-01

    Tumor antigen-specific cytotoxic T lymphocytes (CTL) have been used in the treatment of human cancer, including leukemia. Several studies have established PR1 peptide, an HLA-A2.1-restricted peptide derived from proteinase 3 (P3), as a human leukemia-associated antigen. PR1-specific CTL elicited in vitro from healthy donors have been shown to lyse P3-expressing AML cells from patients. We investigated whether PR1-CTL can be adoptively transferred into NOD/SCID mice to eliminate human leukemia cells. PR1-CTL were generated in bulk culture from peripheral blood mononuclear cells (PBMC) stimulated with autologous dendritic cells. Human acute myeloid leukemia (AML) patient samples were injected and engrafted in murine bone marrow at 2 weeks post-transfer. Following adoptive transfer, bone marrow aspirate from mice that received AML alone had 72-88% blasts in a hypercellular marrow, whereas mice that received AML plus PR1-CTL co-infusion had normal hematopoietic elements and only 3-18% blasts in a hypocellular marrow. The PR1-CTL persisted in the bone marrow and liver and maintained a CD45RA⁻CD28+ effector phenotype. We found that adoptive transfer of PR1-CTL generated in vitro is associated with reduced AML cells in NOD/SCID mice. PR1-CTL can migrate to the sites of disease and maintain their capacity to kill the AML cells. The surface phenotype of PR1-CTL was consistent with their trafficking pattern in both vascular and end-organ tissues.

  11. Gag Protein Epitopes Recognized by ELA-A-Restricted Cytotoxic T Lymphocytes from Horses with Long-Term Equine Infectious Anemia Virus Infection

    Science.gov (United States)

    Zhang, Wei; Lonning, Scott M.; McGuire, Travis C.

    1998-01-01

    Most equine infectious anemia virus (EIAV)-infected horses have acute clinical disease, but they eventually control the disease and become lifelong carriers. Cytotoxic T lymphocytes (CTL) are considered an important immune component in the control of infections with lentiviruses including EIAV, but definitive evidence for CTL in the control of disease in carrier horses is lacking. By using retroviral vector-transduced target cells expressing different Gag proteins and overlapping synthetic peptides of 16 to 25 amino acids, peptides containing at least 12 Gag CTL epitopes recognized by virus-stimulated PBMC from six long-term EIAV-infected horses were identified. All identified peptides were located within Gag matrix (p15) and capsid (p26) proteins, as no killing of target cells expressing p11 and p9 occurred. Each of the six horses had CTL recognizing at least one Gag epitope, while CTL from one horse recognized at least eight different Gag epitopes. None of the identified peptides were recognized by CTL from all six horses. Two nonamer peptide epitopes were defined from Gag p26; one (18a) was likely restricted by class I equine leukocyte alloantigen A5.1 (ELA-A5.1) molecules, and the other (28b-1) was likely restricted by ELA-A9 molecules. Sensitization of equine kidney target cells for CTLm killing required 10 nM peptide 18a and 1 nM 28b-1. The results demonstrated that diverse CTL responses against Gag epitopes were generated in long-term EIAV-infected horses and indicated that ELA-A class I molecules were responsible for the diversity of CTL epitopes recognized. This information indicates that multiple epitopes or whole proteins will be needed to induce CTL in horses with different ELA-A alleles in order to evaluate their role in controlling EIAV. PMID:9811694

  12. Computer-based design of an HLA-haplotype and HIV-clade independent cytotoxic T-lymphocyte assay for monitoring HIV-specific immunity.

    Science.gov (United States)

    Amicosante, Massimo; Gioia, Cristiana; Montesano, Carla; Casetti, Rita; Topino, Simone; D'Offizi, Gianpiero; Cappelli, Giulia; Ippolito, Giuseppe; Colizzi, Vittorio; Poccia, Fabrizio; Pucillo, Leopoldo P.

    2002-01-01

    BACKGROUND: Human immunodeficiency virus (HIV)- specific CD8-positive cytotoxic T-lymphocytes (CTL) play a key role in controlling HIV infection. Monitoring CTL response could be clinically relevant during structured therapy interruption (STI), HIV exposure, and vaccine trials. However, HLA patients' restriction and HIV variability limited the development of a CTL assay with broad specificity. MATERIALS AND METHODS: We designed an HLA-class I/HIV-1 clade independent assay for assessing HIV- specific CTL by using a computer-assisted selection ofthe CTL epitopes. Twenty-eight 15-mers were selected by peptide-binding motifs analysis using different databases (HIV-Immunology Database, SYFPEITHI, BIMAS). Altogether they putatively bind to more than 90% of HLA haplotypes in different populations, with an overall HIV-1 variability below 9%. The peptide pool was used as an antigen in an intracellular cytokine staining (ICS) assay for quantifying HIV-specific CTL response. RESULTS: The test can be performed using both fresh and cryopreserved peripheral blood mononuclear cells (PBMC), whereas GAG protein as antigen works only on fresh PBMC. A significantly higher CTL response with respect to HIV-negative controls was detected in all HIV-1 infected subjects of two groups of patients with different ethnicities (Caucasians and Africans) and coming from areas with different HIV-1 clade prevalences (clade B and A/G, respectively). In Caucasian patients, after month of STI, the number of HIV-1 specific CTL (2,896 +/- 2,780 IFN-gamma specific CD8 cells/ml) was significantly higher than that found at enrolment (2,125 +/- 4,426 IFN-gamma specific CD8 cells/ml, p< 0.05). CONCLUSIONS: These data indicate that this CTL assay is broadly specific and could represent a useful clinical tool for HIV immunodiagnostic independent of HLA-haplotype and HIV-clade variabilities. PMID:12606814

  13. PEMBELAJARAN CONTEXTUAL TEACHING AND LEARNING (CTL) DAN HASIL BELAJAR MATEMATIKA SISWA SEKOLAH DASAR

    OpenAIRE

    Sunandar Sunandar

    2016-01-01

    The Influences of CTL Teaching-learning on the Learning Achievement of Primary School Students. The main purpose of this study was knowing whether using Contextual Teaching and Learn­ing (CTL) approach was more effective than Textual Teaching and Learning (TTL) approach in mathematics study. The samples of this study were 85 students from class VA and VB of Ngesrep 01/02 Elementary School at Semarang. The instruments in this study were (1) learning achievement test (2) observation sheet of st...

  14. The loss of immunodominant epitopes affects interferon-γ production and lytic activity of the human influenza virus-specific cytotoxic T lymphocyte response in vitro

    NARCIS (Netherlands)

    E.G.M. Berkhoff (Eufemia); M.M. Geelhoed-Mieras (Martina); E.J. Verschuren (Esther); C.A. van Baalen (Carel); R.A. Gruters (Rob); R.A.M. Fouchier (Ron); A.D.M.E. Osterhaus (Albert); G.F. Rimmelzwaan (Guus)

    2007-01-01

    textabstractIn the present study, we examined the effect of the loss of the human leucocyte antigen (HLA)-B*3501-restricted nucleoprotein (NP)418-426epitope on interferon (IFN)-γ-production and lytic activity of the human cytotoxic T lymphocyte (CTL) response in vitro. Extensive amino acid variation

  15. Protective Role of Cytotoxic T Lymphocytes in Filovirus Hemorrhagic Fever

    Directory of Open Access Journals (Sweden)

    Kelly Lyn Warfield

    2011-01-01

    Full Text Available Infection with many emerging viruses, such as the hemorrhagic fever disease caused by the filoviruses, Marburg (MARV, and Ebola virus (EBOV, leaves the host with a short timeframe in which to mouse a protective immune response. In lethal cases, uncontrolled viral replication and virus-induced immune dysregulation are too severe to overcome, and mortality is generally associated with a lack of notable immune responses. Vaccination studies in animals have demonstrated an association of IgG and neutralizing antibody responses against the protective glycoprotein antigen with survival from lethal challenge. More recently, studies in animal models of filovirus hemorrhagic fever have established that induction of a strong filovirus-specific cytotoxic T lymphocyte (CTL response can facilitate complete viral clearance. In this review, we describe assays used to discover CTL responses after vaccination or live filovirus infection in both animal models and human clinical trials. Unfortunately, little data regarding CTL responses have been collected from infected human survivors, primarily due to the low frequency of disease and the inability to perform these studies in the field. Advancements in assays and technologies may allow these studies to occur during future outbreaks.

  16. Academic Cloning.

    Science.gov (United States)

    Sikula, John P.; Sikula, Andrew F.

    1980-01-01

    The authors define "cloning" as an integral feature of all educational systems, citing teaching practices which reward students for closely reproducing the teacher's thoughts and/or behaviors and administrative systems which tend to promote like-minded subordinates. They insist, however, that "academic cloning" is not a totally…

  17. Activation of secretion and surface alteration of cytolytic T-lymphocytes interacting with target cells.

    Science.gov (United States)

    Bykovskaya, S N; Shevelev, A A; Kupriyanova, T A

    1988-01-01

    Cells obtained in mixed lymphocyte culture (MLC) and memory cells adsorbed on the surface of target cells (TC) were examined using scanning and transmission electron microscopy depending on the time of interaction with TC. Three types of lymphocytes were revealed: type I - cells of spherical shape with a smooth surface or an insignificant amount of microvilli; predominantly small and medium-sized lymphocytes contacting TC with non significant involvement of their surface or by several microvilli; type II - oval or round-shaped lymphocytes evenly covered with microvilli with considerably enlarged region of contact; type III cells - predominantly large lymphocytes and lymphoblasts flattened (spread) on TC, with multiple microvilli, ridge-like projections, and ruffles on their surface. TEM revealed activation of the secretory apparatus in the cytoplasm of such lymphocytes. With increased time of interaction, type III cells increase in number (from 8.6% after 10 min to 90.2% after 60 min of incubation). Memory cells show no morphologic signs of secretion in correlation with the absence of lysis of TC on which they are adsorbed. The surface of the lymphocytes adsorbed on the substrate with poly-L-lysin is not noticeably altered. It is suggested that 3 morphological types of lymphocytes correspond to 3 stages of secretion activation. Lymphocyte contact with TC surface is evidently a specific stimulus for activating secretory apparatus of CTL. SEM can be used for quantitation of activated lymphocytes.

  18. Peptide-MHC class I stability is a stronger predictor of CTL immunogenicity than peptide affinity

    DEFF Research Database (Denmark)

    Harndahl, Mikkel Nors; Rasmussen, Michael; Nielsen, Morten

    2012-01-01

    Peptide-MHC class I stability is a stronger predictor of CTL immunogenicity than peptide affinity Mikkel Harndahla, Michael Rasmussena, Morten Nielsenb, Soren Buusa,∗ a Laboratory of Experimental Immunology, Faculty of Health Sciences, University of Copenhagen, Denmark b Center for Biological Seq...... al., 2007. J. Immunol. 178, 7890–7901. doi:10.1016/j.molimm.2012.02.025...

  19. MODEL PEMBELAJARAN EKONOMI SYARI’AH MELALUI CTL PADA JURUSAN PENDIDIKAN EKONOMI UNIVERSITAS NEGERI SURABAYA

    Directory of Open Access Journals (Sweden)

    Yoyok Soesatyo

    2015-09-01

    material given by using direct instructional model is still less than the standard of competencespecified, therefore it is necessary efforts to increase the understanding material through appropriate learning model in this study is a CTL (Contectual Teaching Learning

  20. EFEKTIVITAS PENDEKATAN OPEN-ENDED DAN CTL DITINJAU DARI BERPIKIR KREATIF SISWA KELAS VII

    Directory of Open Access Journals (Sweden)

    Yeni Rahmawati ES

    2016-06-01

    Full Text Available This research aimed to describe the effectiveness of the open-ended and CTL approach in terms of creative thinking. This research was quasi-experimental study design with nonequivalent pretest-posttest design goup. The research population was all students in class VII SMP Negeri 1 Sekampung which consisted of five classes. The research sample consisted of two classes is determined randomly. First experimental group was treated the open-ended and experimental group II were treated CTL approach. The one sample t-test carried out to investigate the effectiveness of the open-ended and CTL approach. Data collection techniques used were tests. Engineering tests used to measure student creative thinking on basic competencies: 1 identify the properties of a rectangle, square, trapezoid, parallelogram, rhombus and a kite; 2 calculate the circumference and area of a rectangle up and use it in problem solving both before and after treatment. The results of the research show that the open-ended and CTL approach is effective in terms of creative thinking. Keywords: open-ended, contextual teaching and learning, creative thinking

  1. A Sigmoid Functional Response Emerges When Cytotoxic T Lymphocytes Start Killing Fresh Target Cells.

    Science.gov (United States)

    Gadhamsetty, Saikrishna; Marée, Athanasius F M; Beltman, Joost B; de Boer, Rob J

    2017-03-28

    Cytotoxic T lymphocyte (CTL)-mediated killing involves the formation of a synapse with a target cell, followed by delivery of perforin and granzymes. Previously, we derived a general functional response for CTL killing while considering that CTLs form stable synapses (i.e., single-stage) and that the number of conjugates remains at steady state. However, the killing of target cells sometimes requires multiple engagements (i.e., multistage). To study how multistage killing and a lack of steady state influence the functional response, we here analyze a set of differential equations as well as simulations employing the cellular Potts model, in both cases describing CTLs that kill target cells. We find that at steady state the total killing rate (i.e., the number of target cells killed by all CTLs) is well described by the previously derived double saturation function. Compared to single-stage killing, the total killing rate during multistage killing saturates at higher CTL and target cell densities. Importantly, when the killing is measured before the steady state is approached, a qualitatively different functional response emerges for two reasons: First, the killing signal of each CTL gets diluted over several targets and because this dilution effect is strongest at high target cell densities; this can result in a peak in the dependence of the total killing rate on the target cell density. Second, the total killing rate exhibits a sigmoid dependence on the CTL density when killing is a multistage process, because it takes typically more than one CTL to kill a target. In conclusion, a sigmoid dependence of the killing rate on the CTLs during initial phases of killing may be indicative of a multistage killing process. Observation of a sigmoid functional response may thus arise from a dilution effect and is not necessarily due to cooperative behavior of the CTLs. Copyright © 2017 Biophysical Society. Published by Elsevier Inc. All rights reserved.

  2. Regulatory effect of Ganoderma lucidum polysaccharides on cytotoxic T-lymphocytes induced by dendritic cells in vitro

    Institute of Scientific and Technical Information of China (English)

    CAOLi-Zhen; LINZhi-Bin

    2003-01-01

    AIM:To study the regulatory effects of Ganoderma lucidum polysaccharides(Gl-PS)on cytotoxicity and mechanism of specific cytotoxic T-lymphocytes(CTL)induced by dendritic cells(DC)in vitro during the stage of antigen presentation.METHODS:Cultured murine bone marrow-derived DC were pulsed with P815 tumor cell lysates and co-incubated with or without various concentrations of Gl-PS(0.8,3.2,or 12.8mg/L)at the same time.P815 specific CTL were induced by spleen lymphocytes stimulated with mature DC.Non-adherent cells and culture supernatants were harvested on d 5 for analysis of specific cytotoxicity with lactate dehydrogenase(LDH)activity assay,mRNA expression of IFNγ,granzyme B with RT-PCR assay,and protein expression of IFNγ,granzyme B with ELISA or Western blot assay,respectively,RESULTS:Three concentrations of Gl-PS promoted LDH activities released into culture supernatants(P<0.01).It also increased mRNA expression of IFNγin CTL(Gl-PS 12.8mg/L vs RPMI medium 1640,P<0.05)and granzyme B in CTL(P<0.01).Protein production of IFNγin culture supernatants(P<0.05)and protein expression of granzyme B in CTL(Gl-PS 12.8mg/L vs RPMI medium 1640,P<0.05)were also augmented by Gl-PS,CONCLUSION:Gl-PS is shown to promote the cytotoxicity of specific CTL induced by DC which were pulsed with P815 tumor antigen during the stage of antigen presentation,and the mechanism of cytotoxicity is believed to be going through IFNγ and granzyme B pathways.

  3. Combinatorial peptide libraries as an alternative approach to the identification of ligands for tumor-reactive cytolytic T lymphocytes.

    Science.gov (United States)

    Pinilla, C; Rubio-Godoy, V; Dutoit, V; Guillaume, P; Simon, R; Zhao, Y; Houghten, R A; Cerottini, J C; Romero, P; Valmori, D

    2001-07-01

    The recent identification of molecularly defined human tumor antigens recognized by autologous CTLs has opened new opportunities for the development of antigen-specific cancer vaccines. Despite extensive work, however, the number of CTL-defined tumor antigens that are suitable targets for generic vaccination of cancer patients is still limited, mostly because of the painstaking and lengthy nature of the procedures currently used for their identification. A novel approach is based on the combined use of combinatorial peptide libraries in positional scanning format (positional scanning synthetic combinatorial peptide libraries, PS-SCLs) and tumor-reactive CTL clones. To validate this approach, we herein analyzed in detail the recognition of PS-SCLs by Melan-A-specific CTL clones. Our results indicate that, at least for some clones, most of the amino acids composing the native antigenic peptide can be identified through the use of PS-SCLs. Interestingly, this analysis also allowed the identification of peptide analogues with increased antigenic activity as well as agonist peptides containing multiple amino-acid substitutions. In addition, biometrical analysis of the data generated by PS-SCL screening allowed the identification of the native ligand in a public database. Overall, these data demonstrate the successful use of PS-SCLs for the identification and optimization of tumor-associated CTL epitopes.

  4. Recombinant poxvirus boosting of DNA-primed rhesus monkeys augments peak but not memory T lymphocyte responses.

    Science.gov (United States)

    Santra, Sampa; Barouch, Dan H; Korioth-Schmitz, Birgit; Lord, Carol I; Krivulka, Georgia R; Yu, Faye; Beddall, Margaret H; Gorgone, Darci A; Lifton, Michelle A; Miura, Ayako; Philippon, Valerie; Manson, Kelledy; Markham, Phillip D; Parrish, John; Kuroda, Marcelo J; Schmitz, Jörn E; Gelman, Rebecca S; Shiver, John W; Montefiori, David C; Panicali, Dennis; Letvin, Norman L

    2004-07-27

    Although a consensus has emerged that an HIV vaccine should elicit a cytotoxic T lymphocyte (CTL) response, the characteristics of an effective vaccine-induced T lymphocyte response remain unclear. We explored this issue in the simian human immunodeficiency virus/rhesus monkey model in the course of assessing the relative immunogenicity of vaccine regimens that included a cytokine-augmented plasmid DNA prime and a boost with DNA or recombinant pox vectors. Recombinant vaccinia virus, recombinant modified vaccinia Ankara (MVA), and recombinant fowlpox were comparable in their immunogenicity. Moreover, whereas the magnitude of the peak vaccine-elicited T lymphocyte responses in the recombinant pox virus-boosted monkeys was substantially greater than that seen in the monkeys immunized with plasmid DNA alone, the magnitudes of recombinant pox boosted CTL responses decayed rapidly and were comparable to those of the DNA-alone-vaccinated monkeys by the time of viral challenge. Consistent with these comparable memory T cell responses, the clinical protection seen in all groups of experimentally vaccinated monkeys was similar. This study, therefore, indicates that the steady-state memory, rather than the peak effector vaccine-elicited T lymphocyte responses, may be the critical immune correlate of protection for a CTL-based HIV vaccine.

  5. Recognition of HIV-1 peptides by host CTL is related to HIV-1 similarity to human proteins.

    Directory of Open Access Journals (Sweden)

    Morgane Rolland

    Full Text Available BACKGROUND: While human immunodeficiency virus type 1 (HIV-1-specific cytotoxic T lymphocytes preferentially target specific regions of the viral proteome, HIV-1 features that contribute to immune recognition are not well understood. One hypothesis is that similarities between HIV and human proteins influence the host immune response, i.e., resemblance between viral and host peptides could preclude reactivity against certain HIV epitopes. METHODOLOGY/PRINCIPAL FINDINGS: We analyzed the extent of similarity between HIV-1 and the human proteome. Proteins from the HIV-1 B consensus sequence from 2001 were dissected into overlapping k-mers, which were then probed against a non-redundant database of the human proteome in order to identify segments of high similarity. We tested the relationship between HIV-1 similarity to host encoded peptides and immune recognition in HIV-infected individuals, and found that HIV immunogenicity could be partially modulated by the sequence similarity to the host proteome. ELISpot responses to peptides spanning the entire viral proteome evaluated in 314 individuals showed a trend indicating an inverse relationship between the similarity to the host proteome and the frequency of recognition. In addition, analysis of responses by a group of 30 HIV-infected individuals against 944 overlapping peptides representing a broad range of individual HIV-1B Nef variants, affirmed that the degree of similarity to the host was significantly lower for peptides with reactive epitopes than for those that were not recognized. CONCLUSIONS/SIGNIFICANCE: Our results suggest that antigenic motifs that are scarcely represented in human proteins might represent more immunogenic CTL targets not selected against in the host. This observation could provide guidance in the design of more effective HIV immunogens, as sequences devoid of host-like features might afford superior immune reactivity.

  6. Cytotoxic T lymphocytes promote cytarabine-induced acute myeloid leukemia cell apoptosis via inhibiting Bcl-2 expression.

    Science.gov (United States)

    Deng, Rui; Fan, Fang-Yi; Yi, Hai; Fu, Li; Zeng, Yan; Wang, Yi; Miao, Xiao-Juan; Shuai, Yan-Rong; He, Guang-Cui; Su, Yi

    2017-08-01

    Acute myeloid leukemia (AML) remains difficult to cure due to its drug tolerance and refractoriness. Immunotherapy is a growing area of cancer research, which has been applied for the treatment of numerous types of cancer, including leukemia. The present study generated AML cell-specific cytotoxic T lymphocytes (CTLs) in vitro and investigated the effect of combining CTL treatment with one of the most commonly used drugs for the treatment of hematological malignancies, cytarabine, on AML cell apoptosis. Firstly, it was observed that monocyte-depleted peripheral blood lymphocytes from healthy donors could be used to generate large numbers of CD3(+)CD8(+) CTLs through immune stimulation. These CD3(+)CD8(+) CTLs could effectively recognize and induce the apoptosis of human Kasumi-3 AML cells. In addition, cytarabine-induced AML cell apoptosis was enhanced by CTL treatment. Western blotting revealed that Bcl-2 expression was downregulated in AML cells following cytarabine and CTL treatment, indicating that the synergistic effect of this treatment on AML cell apoptosis is due to the downregulation of Bcl-2. These results highlight the potential application of CTL immunotherapy for the treatment of AML. Further studies optimizing the specificity and potency of CTLs, and identifying favorable combinations with other chemotherapeutic drug are required.

  7. Quantifying factors determining the rate of CTL escape and reversion during acute and chronic phases of HIV infection

    Energy Technology Data Exchange (ETDEWEB)

    Ganusov, Vitaly V [Los Alamos National Laboratory; Korber, Bette M [Los Alamos National Laboratory; Perelson, Alan S [Los Alamos National Laboratory

    2009-01-01

    Human immunodeficiency virus (HIV) often evades cytotoxic T cell (CTL) responses by generating variants that are not recognized by CTLs. However, the importance and quantitative details of CTL escape in humans are poorly understood. In part, this is because most studies looking at escape of HIV from CTL responses are cross-sectional and are limited to early or chronic phases of the infection. We use a novel technique of single genome amplification (SGA) to identify longitudinal changes in the transmitted/founder virus from the establishment of infection to the viral set point at 1 year after the infection. We find that HIV escapes from virus-specific CTL responses as early as 30-50 days since the infection, and the rates of viral escapes during acute phase of the infection are much higher than was estimated in previous studies. However, even though with time virus acquires additional escape mutations, these late mutations accumulate at a slower rate. A poor correlation between the rate of CTL escape in a particular epitope and the magnitude of the epitope-specific CTL response suggests that the lower rate of late escapes is unlikely due to a low efficacy of the HIV-specific CTL responses in the chronic phase of the infection. Instead, our results suggest that late and slow escapes are likely to arise because of high fitness cost to the viral replication associated with such CTL escapes. Targeting epitopes in which virus escapes slowly or does not escape at all by CTL responses may, therefore, be a promising direction for the development of T cell based HIV vaccines.

  8. Keefektifan CTL Berbantuan Macromedia Flash Terhadap Kemampuan Berpikir Kritis pada Materi Segiempat

    Directory of Open Access Journals (Sweden)

    O.N. Kusumadewi

    2013-06-01

    Full Text Available Tujuan pada penelitian ini adalah untuk mengetahui nilai rata-rata dan tingkat ketuntasan klasikal pada kemampuan berpikir kritis peserta didik dalam materi segiempat setelah diterapkan model Contextual Teaching and Learning/CTL berbantuan Macromedia Flash 8, dan membandingkan model CTL berbantuan Macromedia Flash 8 dengan model direct instruction pada kemampuan berpikir kritis materi segiempat kelas VII. Populasi pada penelitian ini adalah peserta didik kelas VII SMP Negeri 1 Semarang tahun pelajaran 2012/ 2013. Sampel dipilih dengan menggunakan teknik cluster sampling. Hasil penelitian menunjukkan menunjukkan bahwa rata-rata kemampuan berpikir kritis kelas eksperimen mencapai kriteria ketuntasan minimum (KKM. Persentase banyaknya peserta didik yang tuntas melebihi ketuntasan klasikal minimal. Kemampuan berpikir kritis pada kelas eksperimen lebih baik dibandingkan pada kelas kontrol.  Hal tersebut terlihat dari rata-rata nilai dan persentase peserta didik yang tuntas pada kelas eksperimen lebih tinggi daripada kelas kontrol. Berdasarkan hasil penelitian, dapat disimpulkan bahwa pembelajaran dengan model CTL berbantuan Macromedia Flash 8 efektif terhadap kemampuan berpikir kritis materi segiempat kelas VII.   The purpose of the experiment is to find out the students’ completeness mean score and percentage in critical thinking ability about quadrilateral in CTL model with Macromedia Flash 8, and to compare the CTL model with direct instruction model in students’ critical thinking ability at 7th grade. The popula­tion of this study was 7th grade students of SMP N 1 Semarang academic year 2012/2013. Study learning showed that means score of critical thinking ability in experiment class achieves minimum completeness criteria (KKM. Percentage of students that achieve KKM was also more than minimum classical completeness score. Critical thinking ability of the experiment class is better than control class. It is proven from mean score and

  9. T-lymphocyte and B-lymphocyte dichotomy in anuran amphibians: III. Assessment and identification of inducible killer T-lymphocytes (IKTL) and spontaneous killer T-lymphocytes (SKTL).

    Science.gov (United States)

    Klempau, A E; Cooper, E L

    1984-01-01

    We have established the existence of alloreactive inducible killer (IK) T-lymphocytes in Rana pipiens by injecting immunogenic concentrations of allogeneic frog erythrocytes (RBC). Assessment of specific IK activity was determined microscopically, observing effector-target conjugate formation, and spectrophotometrically as released hemoglobin (Hb) from lysed targets (RBC). The presence of spontaneous killer (SK) T-lymphocyte activity was also determined using unimmunized frogs and similar assay conditions. Assays using rabbit anti-frog Thy-1.1 antiserum inhibition, but not E-rosetted T-lymphocyte depletion, confirmed the T-lymphocyte category of both effector cell populations in Rana pipiens. For IK activity, we determined the 1) best priming doses, 2) best effector cell source (peripheral blood), 3) best priming route (intraperitoneal), 4) kinetics of immunity development, and 5) kinetics of lysis. Kinetics of lysis and organ distribution for spontaneous killer cells were also determined. Our results may assist 1) in establishing the evolutionary origin of cytotoxic T-lymphocytes (CTL) and natural killer (NK) cells, and 2) in predicting where the capacity of immuno-surveillance against modified-self appeared in phylogeny. The implications are important for understanding origins of mechanisms of resistance against neoplastic conditions.

  10. Optimization and immune recognition of multiple novel conserved HLA-A2, human immunodeficiency virus type 1-specific CTL epitopes

    DEFF Research Database (Denmark)

    Corbet, S; Nielsen, HV; Vinner, L;

    2003-01-01

    MHC-I-restricted cytotoxic responses are considered a critical component of protective immunity against viruses, including human immunodeficiency virus type 1 (HIV-1). CTLs directed against accessory and early regulatory HIV-1 proteins might be particularly effective; however, CTL epitopes....... This study demonstrates the potency of ANNs for identifying putative virus CTL epitopes, and the new HIV-1 CTL epitopes identified should have significant implications for HIV-1 vaccine development. As a novel vaccine approach, it is proposed to increase the coverage of HIV variants by including multiple...

  11. Optimization and immune recognition of multiple novel conserved HLA-A2, human immunodeficiency virus type 1-specific CTL epitopes

    DEFF Research Database (Denmark)

    Corbet, S.; Nielsen, H.V.; Vinner, L.;

    2003-01-01

    MHC-I-restricted cytotoxic responses are considered a critical component of protective immunity against viruses, including human immunodeficiency virus type 1 (HIV-1). CTLs directed against accessory and early regulatory HIV-1 proteins might be particularly effective; however, CTL epitopes...... demonstrates the potency of ANNs for identifying putative virus CTL epitopes, and the new HIV-1 CTL epitopes identified should have significant implications for HIV-1 vaccine development. As a novel vaccine approach, it is proposed to increase the coverage of HIV variants by including multiple anchor...

  12. Identification and enhancement of HLA-A2.1-restricted CTL epitopes in a new human cancer antigen-POTE.

    Directory of Open Access Journals (Sweden)

    Yi-Hsiang Huang

    Full Text Available Identification of CD8(+ T cell epitopes that can induce T cells to kill tumor cells is a fundamental step for development of a peptide cancer vaccine. POTE protein is a newly identified cancer antigen that was found to be expressed in a wide variety of human cancers, including prostate, colon, lung, breast, ovary and pancreas. Here, we determined HLA-A2.1-restricted cytotoxic T lymphocyte (CTL epitopes in the POTE protein, and also designed enhanced epitopes by amino acid (AA substitutions. Five 9-mer peptides were first selected and their binding affinity to HLA-A2 molecules was measured by the T2 binding assay. POTE 272-280 and POTE 323-331 showed the strongest HLA-A2 binding affinity. AA substituted peptides POTE 252-9V (with valine at position 9, POTE 553-1Y (with tyrosine at position 1 and POTE 323-3F (with phenylalanine at position 3 conferred higher affinity for HLA-A2, and induced CTL responses cross-reactive with wild type antigens. While POTE 252-9V was the strongest in this respect, POTE 323-3F had the greatest increase in immunogenicity compared to wild type. Importantly, two modified epitopes (POTE-553-1Y and POTE-323-3F induced CTLs that killed NCI-H522, a POTE-expressing HLA-A2(+ human non-small cell lung cancer cell line, indicating natural endogenous processing of these epitopes. In conclusion, the immunogenicity of POTE epitopes can be enhanced by peptide modification to induce T cells that kill human cancer cells. A combination of POTE 553-1Y and POTE 323-3F epitopes might be an attractive vaccine strategy for HLA-A2 cancer patients to overcome tolerance induced by tumors and prevent escape.

  13. Inhibition of serine-peptidase activity enhances the generation of a survivin-derived HLA-A2-presented CTL epitope in colon-carcinoma cells.

    Science.gov (United States)

    Preta, G; Marescotti, D; Fortini, C; Carcoforo, P; Castelli, C; Masucci, M; Gavioli, R

    2008-12-01

    Cytotoxic T lymphocytes eliminate tumor cells expressing antigenic peptides in the context of MHC-I molecules. Peptides are generated during protein degradation by the proteasome and resulting products, surviving cytosolic amino-peptidases activity, may be presented by MHC-I molecules. The MHC-I processing pathway is altered in a large number of malignancies and modulation of antigen generation is one strategy employed by cells to evade immune control. In this study we analyzed the generation and presentation of a survivin-derived CTL epitope in HLA-A2-positive colon-carcinoma cells. Although all cell lines expressed the anti-apoptotic protein survivin, some tumors were poorly recognized by ELTLGEFLKL (ELT)-specific CTL cultures. The expression of MHC-I or TAP molecules was similar in all cell lines suggesting that tumors not recognized by CTLs may present defects in the generation of the ELT-epitope which could be due either to lack of generation or to subsequent degradation of the epitope. The cells were analyzed for the expression and the activity of extra-proteasomal peptidases. A significant overexpression and higher activity of TPPII was observed in colon-carcinoma cells which are not killed by ELT-specific CTLs, suggesting a possible role of TPPII in the degradation of the ELT-epitope. To confirm the role of TPPII in the degradation of the ELT-peptide, we showed that treatment of colon-carcinoma cells with a TPPII inhibitor resulted in a dose-dependent increased sensitivity to ELT-specific CTLs. These results suggest that TPPII is involved in degradation of the ELT-peptide, and its overexpression may contribute to the immune escape of colon-carcinoma cells.

  14. Applying generalized hydrophobicity scale of amino acids to quantitative prediction of human leukocyte antigen-A*0201-restricted cytotoxic T lymphocyte epitope

    Institute of Scientific and Technical Information of China (English)

    ZHOU Peng; TIAN Feifei; ZHANG Mengjun; LI Zhiliang

    2006-01-01

    Derived from 149 hydrophobic factors of 20 natural amino acids, a novel amino acid descriptor termed as generalized hydrophobicity scale (GH-scale) was proposed by principal component analysis (PCA). Via genetic algorithm-partial least square (GPLS) method, quantitative structure-activity relationship (QSAR) model was constructed by GH-scale for 152 human leukocyte antigen (HLA)-A*0201-restricted cytotoxic T lymphocyte (CTL) epitopes with the model estimated and cross-validated correlative coefficients of R2 = 0.813 and Q2 = 0.725, respectively. It was indicated that hydrophobic interaction played an important role in HLA-A*0201-CTL interaction, prominently at anchor residues.

  15. Gag Protein Epitopes Recognized by CD4+ T-Helper Lymphocytes from Equine Infectious Anemia Virus-Infected Carrier Horses

    Science.gov (United States)

    Lonning, S. M.; Zhang, W.; McGuire, T. C.

    1999-01-01

    Antigen-specific T-helper (Th) lymphocytes are critical for the development of antiviral humoral responses and the expansion of cytotoxic T lymphocytes (CTL). Identification of relevant Th lymphocyte epitopes remains an important step in the development of an efficacious subunit peptide vaccine against equine infectious anemia virus (EIAV), a naturally occurring lentivirus of horses. This study describes Th lymphocyte reactivity in EIAV carrier horses to two proteins, p26 and p15, encoded by the relatively conserved EIAV gag gene. Using partially overlapping peptides, multideterminant and possibly promiscuous epitopes were identified within p26. One peptide was identified which reacted with peripheral blood mononuclear cells (PBMC) from all five EIAV-infected horses, and three other peptides were identified which reacted with PBMC from four of five EIAV-infected horses. Four additional peptides containing both CTL and Th lymphocyte epitopes were also identified. Multiple epitopes were recognized in a region corresponding to the major homology region of the human immunodeficiency virus, a region with significant sequence similarity to other lentiviruses including simian immunodeficiency virus, puma lentivirus, feline immunodeficiency virus, Jembrana disease virus, visna virus, and caprine arthritis encephalitis virus. PBMC reactivity to p15 peptides from EIAV carrier horses also occurred. Multiple p15 peptides were shown to be reactive, but not all infected horses had Th lymphocytes recognizing p15 epitopes. The identification of peptides reactive with PBMC from outbred horses, some of which encoded both CTL and Th lymphocyte epitopes, should contribute to the design of synthetic peptide or recombinant vector vaccines for EIAV. PMID:10196322

  16. Global stability for delay-dependent HTLV-I model with CTL immune response

    Science.gov (United States)

    Wang, Yan; Liu, Jun

    2016-06-01

    We present a delay-dependent HTLV-I model with CTL immune response. The basic reproduction number is obtained for the existence of positive steady state. By constructing suitable Lyapunov functions, when the basic reproduction number is less than one, the infection-free steady state is globally asymptotically stable; when the basic reproduction number is greater than one, the infected steady state is globally asymptotically stable.

  17. PENGARUH PENDEKATAN CONTEXTUAL TEACHING AND LEARNING (CTL BERBANTUAN MEDIA POWERPOINT TERHADAP PENINGKATAN HASIL BELAJAR IPA FISIKA

    Directory of Open Access Journals (Sweden)

    Suprianto Suprianto

    2016-12-01

    Full Text Available Berdasarkan hasil wawancara terbatas dengan sebagian guru IPA fisika MTs di kecamatan Camplong Kabupaten Sampang didapatkan informasi bahwa hasil belajar siswa masih rendah dibawah KKM yang sudah ditentukan pihak sekolah. Hal ini disebabkan karena kurangnya penggunaan media pembelajaran, proses pembelajaran masih bersifat teacher centered sehingga sebagian besar siswa tidak mampu menghubungkan antara apa yang mereka pelajari dengan bagaimana pemanfaatannya dalam kehidupan nyata. Adapun tujuan penelitian ini adalah untuk mengidentifikasi pengaruh pendekatan CTL berbantuan media powerpoint terhadap peningkatan hasil belajar IPA fisika siswa. Metode penelitian yang digunakan adalah metode Quasi Experimen dengan menggunakan Nonrandomized Control Group Pretest-Postest Design dan dibagi menjadi dua kelompok, yaitu kelompok eksperimen dan kelompok kontrol. Instrumen penelitian yang digunakan berupa tes objektif bentuk pilihan ganda. Tes ini terdiri dari empat pilihan (opsi dan hasilnya diuji melalui statistik uji “t”. Dari hasil perhitungan diperoleh nilai thitung sebesar 10,81 sedangkan ttabel sebesar 2,021 pada taraf signifikansi 0,05 atau dapat diketahui thitung > ttabel. Dari perhitungan N-gain, dapat dinyatakan bahwa peningkatan hasil belajar fisika yang diterapkan pendekatan CTL berbantuan media powerpoint lebih tinggi dari pada pembelajaran konvensional yaitu 0,71 > 0,52. Berdasarkan hasil analisis statistika dan deskriptif maka dapat di simpulkan bahwa terdapat pengaruh yang signifikan pendekatan CTL berbantuan media powerpoint terhadap peningkatan hasil belajar fisika siswa di kelas VIII MTs.

  18. Role of vitamin D in cytotoxic T lymphocyte immunity to pathogens and cancer.

    Science.gov (United States)

    Sarkar, Surojit; Hewison, Martin; Studzinski, George P; Li, Yan Chun; Kalia, Vandana

    2016-01-01

    The discovery of vitamin D receptor (VDR) expression in immune cells has opened up a new area of research into immunoregulation by vitamin D, a niche that is distinct from its classical role in skeletal health. Today, about three decades since this discovery, numerous cellular and molecular targets of vitamin D in the immune system have been delineated. Moreover, strong clinical associations between vitamin D status and the incidence/severity of many immune-regulated disorders (e.g. infectious diseases, cancers and autoimmunity) have prompted the idea of using vitamin D supplementation to manipulate disease outcome. While much is known about the effects of vitamin D on innate immune responses and helper T (T(H)) cell immunity, there has been relatively limited progress on the frontier of cytotoxic T lymphocyte (CTL) immunity--an arm of host cellular adaptive immunity that is crucial for the control of such intracellular pathogens as human immunodeficiency virus (HIV), tuberculosis (TB), malaria, and hepatitis C virus (HCV). In this review, we discuss the strong historical and clinical link between vitamin D and infectious diseases that involves cytotoxic T lymphocyte (CTL) immunity, present our current understanding as well as critical knowledge gaps in the realm of vitamin D regulation of host CTL responses, and highlight potential regulatory connections between vitamin D and effector and memory CD8 T cell differentiation events during infections.

  19. Vaccination with a recombinant protein encoding the tumor-specific antigen NY-ESO-1 elicits an A2/157-165-specific CTL repertoire structurally distinct and of reduced tumor reactivity than that elicited by spontaneous immune responses to NY-ESO-1-expressing Tumors.

    Science.gov (United States)

    Bioley, Gilles; Guillaume, Philippe; Luescher, Immanuel; Bhardwaj, Nina; Mears, Gregory; Old, Lloyd; Valmori, Danila; Ayyoub, Maha

    2009-01-01

    In a recent vaccination trial assessing the immunogenicity of an NY-ESO-1 (ESO) recombinant protein administered with Montanide and CpG, we have obtained evidence that this vaccine induces specific cytolytic T lymphocytes (CTL) in half of the patients. Most vaccine-induced CTLs were directed against epitopes located in the central part of the protein, between amino acids 81 and 110. This immunodominant region, however, is distinct from another ESO CTL region, 157-165, that is a frequent target of spontaneous CTL responses in A2+ patients bearing ESO tumors. In this study, we have investigated the CTL responses to ESO 157-165 in A2+ patients vaccinated with the recombinant protein. Our data indicate that after vaccination with the protein, CTL responses to ESO 157-165 are induced in some, but not all, A2+ patients. ESO 157-165-specific CTLs induced by vaccination with the ESO protein were functionally heterogeneous in terms of tumor recognition and often displayed decreased tumor reactivity as compared with ESO 157-165-specific CTLs isolated from patients with spontaneous immune responses to ESO. Remarkably, protein-induced CTLs used T-cell receptors similar to those previously isolated from patients vaccinated with synthetic ESO peptides (Vbeta4.1) and distinct from those used by highly tumor-reactive CTLs isolated from patients with spontaneous immune responses (Vbeta1.1, Vbeta8.1, and Vbeta13.1). Together, these results demonstrate that vaccination with the ESO protein elicits a repertoire of ESO 157-165-specific CTLs bearing T-cell receptors that are structurally distinct from those of CTLs found in spontaneous immune responses to the antigen and that are heterogeneous in terms of tumor reactivity, being often poorly tumor reactive.

  20. Molecular cloning.

    Science.gov (United States)

    Lessard, Juliane C

    2013-01-01

    This protocol describes the basic steps involved in conventional plasmid-based cloning. The goals are to insert a DNA fragment of interest into a receiving vector plasmid, transform the plasmid into E. coli, recover the plasmid DNA, and check for correct insertion events.

  1. Clinical-Grade Human Multipotent Adult Progenitor Cells Block CD8+ Cytotoxic T Lymphocytes.

    Science.gov (United States)

    Plessers, Jeroen; Dekimpe, Emily; Van Woensel, Matthias; Roobrouck, Valerie D; Bullens, Dominique M; Pinxteren, Jef; Verfaillie, Catherine M; Van Gool, Stefaan W

    2016-12-01

    : MultiStem cells are clinical-grade multipotent adult bone marrow-derived progenitor cells (MAPCs), with extensive replication potential and broader differentiation capacity compared with mesenchymal stem cells. Human MAPCs suppress T-cell proliferation induced by alloantigens and mutually interact with allogeneic natural killer cells. In this study, the interaction between MultiStem and CD8(+) cytotoxic T lymphocytes (CTLs) was addressed for the first time. In an in vitro setting, the immunogenicity of MultiStem, the susceptibility of MultiStem toward CTL-mediated lysis, and its effects on CTL function were investigated. MultiStem was nonimmunogenic for alloreactive CTL induction and was-even after major histocompatibility complex class I upregulation-insensitive to alloantigen-specific CTL-mediated lysis. Furthermore, MultiStem reduced CTL proliferation and significantly decreased perforin expression during the T-cell activation phase. As a consequence, MultiStem dose-dependently impaired the induction of CTL function. These effects of MultiStem were mediated predominantly through contact-dependent mechanisms. Moreover, MultiStem cells considerably influenced the expression of T-cell activation markers CD25, CD69, and human leukocyte antigen-DR. The MultiStem-induced CD8(-)CD69(+) T-cell population displayed a suppressive effect on the induction of CTL function during a subsequent mixed-lymphocyte culture. Finally, the killer activity of activated antigen-specific CTLs during their cytolytic effector phase was also diminished in the presence of MultiStem. This study confirms that these clinical-grade MAPCs are an immune-modulating population that inhibits CTL activation and effector responses and are, consequently, a highly valuable cell population for adoptive immunosuppressive therapy in diseases where damage is induced by CTLs. Because multipotent adult progenitor cells (MAPCs) are among the noteworthy adult mesenchymal stem cell populations for immune

  2. Suppressor T cells, distinct from "veto cells," are induced by alloantigen priming and mediate transferable suppression of cytotoxic T lymphocyte responses in vivo

    DEFF Research Database (Denmark)

    Owens, T; Crispe, I N

    1985-01-01

    Primary and secondary cytotoxic T lymphocyte responses to minor alloantigens can be suppressed by priming host mice with a high dose (10(8) cells) of alloantigenic donor spleen cells (SC). Such suppression is antigen specific and transferable into secondary hosts with T cells. One interpretation...... of this is that antigen-specific host suppressor T cells (Ts) are activated. Alternatively, donor Lyt-2+ T cells, introduced in the priming inoculum, may inactivate host CTL precursors (CTLp) that recognize the priming (donor) alloantigens. Donor cells that act in this way are termed veto T cells. The experiments...... for the transfer of suppression of a secondary CTL response to B10 minors was of the host Thy-1 allotype, and so originated in the host spleen and was not introduced in the priming inoculum. Secondly, antigen-specific Ts generated in CBA female mice against B10 minors could act on CTL responses to an unequivocally...

  3. Identification of putative human T cell receptor delta complementary DNA clones

    Energy Technology Data Exchange (ETDEWEB)

    Hata, S.; Brenner, M.B.; Krangel, M.S.

    1987-10-30

    A novel T cell receptor (TCR) subunit termed TCR delta, associated with TCY ..gamma.. and CD3 polypeptides, were recently found on a subpopulation of human T lymphocytes. T cell-specific complementary DNA clones present in a human TCR..gamma..delta T cell complementary DNA library were obtained and characterized in order to identify candidate clones encoding TCR delta. One cross-hybridizing group of clones detected transcripts that are expressed in lymphocytes bearing TCR ..gamma..delta but not in other T lymphocytes and are encoded by genes that are rearranged in TCR ..gamma..delta lymphocytes but deleted in other T lymphocytes. Their sequences indicate homology to the variable, joining, and constant elements of other TCR and immunoglobulin genes. These characteristics are strong evidence that the complementary DNA clones encode TCR delta.

  4. Control of mucosal virus infection by influenza nucleoprotein-specific CD8+ cytotoxic T lymphocytes

    Directory of Open Access Journals (Sweden)

    Couch Robert B

    2007-06-01

    Full Text Available Abstract Background MHC class I-restricted CD8+ cytotoxic T lymphocytes (CTL are thought to play a major role in clearing virus and promoting recovery from influenza infection and disease. This has been demonstrated for clearance of influenza virus from the lungs of infected mice. However, human influenza infection is primarily a respiratory mucosal infection involving the nasopharynx and tracheobronchial tree. The role of CD8+ CTL directed toward the influenza nucleoprotein (NP in defense against influenza virus infection at the respiratory mucosa was evaluated in two separate adoptive transfer experiments. Methods Influenza nucleoprotein (NP-specific CD8+ CTL were generated from splenocytes obtained from Balb/c mice previously primed with influenza A/Taiwan/1/86 (H1N1 infection or with influenza A/PR/8/34 (H1N1-derived NP plasmid DNA vaccine followed by infection with A/Hong Kong/68 (H3N2 virus. After in vitro expansion by exposure to an influenza NP-vaccinia recombinant, highly purified CD8+ T cells exhibited significant lysis in vitro of P815 target cells infected with A/Hong Kong/68 (H3N2 virus while the CD8- fraction (CD4+ T cells, B cells and macrophages had no CTL activity. Purified CD8+ and CD8- T cells (1 × 107 were injected intravenously or interperitoneally into naive mice four hours prior to intranasal challenge with A/HK/68 (H3N2 virus. Results The adoptively transferred NP-vaccinia-induced CD8+ T cells caused significant reduction of virus titers in both the lungs and nasal passages when compared to CD8- cells. Neither CD8+ nor CD8- T cells from cultures stimulated with HIV gp120-vaccinia recombinant reduced virus titers. Conclusion The present data demonstrate that influenza NP-specific CD8+ CTL can play a direct role in clearance of influenza virus from the upper respiratory mucosal surfaces.

  5. Serglycin determines secretory granule repertoire and regulates natural killer cell and cytotoxic T lymphocyte cytotoxicity.

    Science.gov (United States)

    Sutton, Vivien R; Brennan, Amelia J; Ellis, Sarah; Danne, Jill; Thia, Kevin; Jenkins, Misty R; Voskoboinik, Ilia; Pejler, Gunnar; Johnstone, Ricky W; Andrews, Daniel M; Trapani, Joseph A

    2016-03-01

    The anionic proteoglycan serglycin is a major constituent of secretory granules in cytotoxic T lymphocyte (CTL)/natural killer (NK) cells, and is proposed to promote the safe storage of the mostly cationic granule toxins, granzymes and perforin. Despite the extensive defects of mast cell function reported in serglycin gene-disrupted mice, no comprehensive study of physiologically relevant CTL/NK cell populations has been reported. We show that the cytotoxicity of serglycin-deficient CTL and NK cells is severely compromised but can be partly compensated in both cell types when they become activated. Reduced intracellular granzyme B levels were noted, particularly in CD27(+) CD11b(+) mature NK cells, whereas serglycin(-/-) TCR-transgenic (OTI) CD8 T cells also had reduced perforin stores. Culture supernatants from serglycin(-/-) OTI T cells and interleukin-2-activated NK contained increased granzyme B, linking reduced storage with heightened export. By contrast, granzyme A was not significantly reduced in cells lacking serglycin, indicating differentially regulated trafficking and/or storage for the two granzymes. A quantitative analysis of different granule classes by transmission electronmicroscopy showed a selective loss of dense-core granules in serglycin(-/-) CD8(+) CTLs, although other granule types were maintained quantitatively. The findings of the present study show that serglycin plays a critical role in the maturation of dense-core cytotoxic granules in cytotoxic lymphocytes and the trafficking and storage of perforin and granzyme B, whereas granzyme A is unaffected. The skewed retention of cytotoxic effector molecules markedly reduces CTL/NK cell cytotoxicity, although this is partly compensated for as a result of activating the cells by physiological means.

  6. Human Cloning

    Science.gov (United States)

    2006-07-20

    genes , for example, has led to new treatments developed by the biotechnology industry for diseases such as diabetes and hemophilia . In the context of...stem cells should be permitted because of the potential for developing new therapies and advancing biomedical knowledge. On May 24, 2005, the House...to describe many different processes that involve making copies of biological material, such as a gene , a cell, a plant or an animal. The cloning of

  7. Peripheral blood and intrathyroidal T cell clones from patients with thyroid autoimmune diseases.

    Science.gov (United States)

    Massart, C; Caroff, G; Maugendre, D; Genetet, N; Gibassier, J

    1999-01-01

    For a better understanding of the pathogenesis of thyroid autoimmune diseases, we have studied morphological and functional properties of T clones from peripheral blood lymphocytes (PBL) and from intrathyroidal lymphocytes (ITL) obtained from 3 patients with Graves' disease or 1 Hashimoto's thyroiditis. Investigations were carried out on clones cultured alone or cocultured with autologous thyrocytes. Clonage efficiency ranged from 30% to 33% for PBL and 10% to 36% for ITL. A predominance of CD4-positive clones was observed whatever the origin of the lymphocytes or the autoimmune pathology. Gamma interferon (IFN-gamma) was detected in the majority (17/19) of the clones tested. Intracytoplasmic interleukin (IL-4) was secreted in 7/19 clones and both cytokines were produced in 5/19 clones. In coculture a proliferative response and tumour necrosis factor (TNF-alpha) production were observed with 6 clones (4 from Graves thyrocytes and 2 from thyroiditis). No cytotoxic clone was derived from Graves or thyroiditis tissues. These data demonstrate that the large majority of T clones are principally CD4-T cells; all the clones secreted TNF-alpha and a large majority produced IFN-gamma. Only a few clones produced IL-4 alone or associated with IFN-gamma. Six T clones induced proliferative response and of TNF-alpha secretion in coculture. Further investigations must be performed on these antigen-reactive T clones to analyse their role in the pathogenesis of the human thyroid autoimmune diseases.

  8. FLT3-regulated antigens as targets for leukemia-reactive cytotoxic T lymphocytes.

    Science.gov (United States)

    Brackertz, B; Conrad, H; Daniel, J; Kast, B; Krönig, H; Busch, D H; Adamski, J; Peschel, C; Bernhard, H

    2011-03-01

    The FMS-like tyrosine kinase 3 (FLT3) is highly expressed in acute myeloid leukemia (AML). Internal tandem duplications (ITD) of the juxtamembrane domain lead to the constitutive activation of the FLT3 kinase inducing the activation of multiple genes, which may result in the expression of leukemia-associated antigens (LAAs). We analyzed the regulation of LAA in FLT3-wild-type (WT)- and FLT3-ITD(+) myeloid cells to identify potential targets for antigen-specific immunotherapy for AML patients. Antigens, such as PR-3, RHAMM, Survivin, WT-1 and PRAME, were upregulated by constitutively active FLT3-ITD as well as FLT3-WT activated by FLT3 ligand (FL). Cytotoxic T-cell (CTL) clones against PR-3, RHAMM, Survivin and an AML-directed CTL clone recognized AML cell lines and primary AML blasts expressing FLT3-ITD, as well as FLT3-WT(+) myeloid dendritic cells in the presence of FL. Downregulation of FLT3 led to the abolishment of CTL recognition. Comparing our findings concerning LAA upregulation by the FLT3 kinase with those already made for the Bcr-Abl kinase, we found analogies in the LAA expression pattern. Antigens upregulated by both FLT3 and Bcr-Abl may be promising targets for the development of immunotherapeutical approaches against myeloid leukemia of different origin.

  9. CTL Responses to Regulatory Proteins Tat and Rev in HIV-1 B'/C Virus-Infected Individuals

    Institute of Scientific and Technical Information of China (English)

    MING-MING JIA; KUN-XUE HONG; JIAN-PING CHEN; HONG-WEI LIU; SHA LIU; XIAO-QING ZHANG; HONG-JING ZHAO; YI-MING SHAO

    2008-01-01

    To characterize HIV-1 specific CTL responses to regulatory proteins Tat and Rev in HIV-B'/C vires-infected ART-naive individuals. Methods HIV-1-specific CTL responses were analyzed by IFN-γ ELISPOT assay using overlapping peptides spanning the consensus sequences of HIV-1 clade C Tat and Rev proteins. Statistical analysis and graphical presentation were performed using SIGMAPLOT 10.0 and SIGMASTAT 3.5. For samples with a positive response, the magnitude of CTL responses was compared between HIV-1 C proteins by Wilcoxon rank sum test, and the significance threshold was P<0.05. Results Tat and Rev were frequently recognized, with 23% and 52% of the tested individuals having detectable responses to these proteins, respectively. Several immunodominant regions were detected in Rev. No significant correlation was observed between the magnitude and breadth of CTL responses to regulatory proteins and the control of virus replication in this study. Conclusion Tat and Rev can serve as targets for HIV-1-specific CTL, and several immunodominant regions are detectable in Rev. Further characterization of epitopes and their role in virus control may shed light on pathogenesis of HIV-1 natural infection and also be useful for the design and testing of candidate vaccines.

  10. In vivo CD8+ T-cell suppression of siv viremia is not mediated by CTL clearance of productively infected cells.

    Directory of Open Access Journals (Sweden)

    Joseph K Wong

    2010-01-01

    Full Text Available The CD8+ T-cell is a key mediator of antiviral immunity, potentially contributing to control of pathogenic lentiviral infection through both innate and adaptive mechanisms. We studied viral dynamics during antiretroviral treatment of simian immunodeficiency virus (SIV infected rhesus macaques following CD8+ T-cell depletion to test the importance of adaptive cytotoxic effects in clearance of cells productively infected with SIV. As previously described, plasma viral load (VL increased following CD8+ T-cell depletion and was proportional to the magnitude of CD8+ T-cell depletion in the GALT, confirming a direct relationship between CD8+ T-cell loss and viral replication. Surprisingly, first phase plasma virus decay following administration of antiretroviral drugs was not slower in CD8+ T-cell depleted animals compared with controls indicating that the short lifespan of the average productively infected cell is not a reflection of cytotoxic T-lymphocyte (CTL killing. Our findings support a dominant role for non-cytotoxic effects of CD8+ T-cells on control of pathogenic lentiviral infection and suggest that cytotoxic effects, if present, are limited to early, pre-productive stages of the viral life cycle. These observations have important implications for future strategies to augment immune control of HIV.

  11. Combinatorial peptide library-based identification of peptide ligands for tumor-reactive cytolytic T lymphocytes of unknown specificity.

    Science.gov (United States)

    Rubio-Godoy, Verena; Ayyoub, Maha; Dutoit, Valerie; Servis, Catherine; Schink, Amy; Rimoldi, Donata; Romero, Pedro; Cerottini, Jean-Charles; Simon, Richard; Zhao, Yindong; Houghten, Richard A; Pinilla, Clemencia; Valmori, Danila

    2002-08-01

    A novel approach for the identification of tumor antigen-derived sequences recognized by CD8(+) cytolytic T lymphocytes (CTL) consists in using synthetic combinatorial peptide libraries. Here we have screened a library composed of 3.1 x 10(11) nonapeptides arranged in a positional scanning format, in a cytotoxicity assay, to search the antigen recognized by melanoma-reactive CTL of unknown specificity. The results of this analysis enabled the identification of several optimal peptide ligands, as most of the individual nonapeptides deduced from the primary screening were efficiently recognized by the CTL. The results of the library screening were also analyzed with a mathematical approach based on a model of independent and additive contribution of individual amino acids to antigen recognition. This biometrical data analysis enabled the retrieval, in public databases, of the native antigenic peptide SSX-2(41-49), whose sequence is highly homologous to the ones deduced from the library screening, among the ones with the highest stimulatory score. These results underline the high predictive value of positional scanning synthetic combinatorial peptide library analysis and encourage its use for the identification of CTL ligands.

  12. Ofatumumab, Pentostatin, and Cyclophosphamide in Treating Patients With Untreated Chronic Lymphocytic Leukemia or Small Lymphocytic Lymphoma

    Science.gov (United States)

    2014-10-30

    Hematopoietic/Lymphoid Cancer; B-cell Chronic Lymphocytic Leukemia; Contiguous Stage II Small Lymphocytic Lymphoma; Noncontiguous Stage II Small Lymphocytic Lymphoma; Stage 0 Chronic Lymphocytic Leukemia; Stage I Chronic Lymphocytic Leukemia; Stage I Small Lymphocytic Lymphoma; Stage II Chronic Lymphocytic Leukemia; Stage III Chronic Lymphocytic Leukemia; Stage III Small Lymphocytic Lymphoma; Stage IV Chronic Lymphocytic Leukemia; Stage IV Small Lymphocytic Lymphoma

  13. Induction of protective CTL immunity against peptide transporter TAP-deficient tumors through dendritic cell vaccination.

    Science.gov (United States)

    Chambers, Benedict; Grufman, Per; Fredriksson, Vanoohi; Andersson, Kenth; Roseboom, Marjet; Laban, Sandra; Camps, Marcel; Wolpert, Elisabeth Z; Wiertz, Emmanuel J H J; Offringa, Rienk; Ljunggren, Hans-Gustaf; van Hall, Thorbald

    2007-09-15

    A large proportion of human cancers show deficiencies in the MHC class I antigen-processing machinery. Such defects render tumors resistant to immune eradication by tumoricidal CTLs. We recently identified a unique population of CTL that selectively targets tumor immune-escape variants through recognition of MHC-presented peptides, termed TEIPP (T cell epitopes associated with impaired peptide processing), expressed on cells lacking functional TAP-peptide transporters. Previously, we showed that vaccination with TEIPP peptides mediates protection against TAP-deficient tumors. Here, we further explored the concept of TEIPP-targeted therapy using a dendritic cell (DC)-based cellular vaccine. Impairment of TAP function in DC induced the presentation of endogenous TEIPP antigens by MHC class I molecules, and immunization with these DCs protected mice against the outgrowth of TAP-deficient lymphomas and fibrosarcomas. Immune analysis of vaccinated mice revealed strong TEIPP-specific CTL responses, and a crucial role for CD8(+) cells in tumor resistance. Finally, we show that TEIPP antigens could be successfully induced in wild-type DC by introducing the viral TAP inhibitor UL49.5. Our results imply that immune intervention strategies with TAP-inhibited DC could be developed for the treatment of antigen processing-deficient cancers in humans.

  14. A scallop C-type lectin from Argopecten irradians (AiCTL5) with activities of lipopolysaccharide binding and Gram-negative bacteria agglutination.

    Science.gov (United States)

    Mu, Changkao; Song, Xiaoyan; Zhao, Jianmin; Wang, Lingling; Qiu, Limei; Zhang, Huan; Zhou, Zhi; Wang, Mengqiang; Song, Linsheng; Wang, Chunlin

    2012-05-01

    C-type lectins are a family of calcium-dependent carbohydrate-binding proteins. In the present study, a C-type lectin (designated as AiCTL5) was identified and characterized from Argopecten irradians. The full-length cDNA of AiCTL5 was of 673 bp, containing a 5' untranslated region (UTR) of 24 bp, a 3' UTR of 130 bp with a poly (A) tail, and an open reading frame (ORF) of 519 bp encoding a polypeptide of 172 amino acids with a putative signal peptide of 17 amino acids. A C-type lectin-like domain (CRD) containing 6 conserved cysteines and a putative glycosylation sites were identified in the deduced amino acid sequence of AiCTL5. AiCTL5 shared 11%-27.5% identity with the previous reported C-type lectin from A. irradians. The cDNA fragment encoding the mature peptide of AiCTL5 was recombined into pET-21a (+) with a C-terminal hexa-histidine tag fused in-frame, and expressed in Escherichia coli Origami (DE3). The recombinant AiCTL5 (rAiCTL5) agglutinated Gram-negative E. coli TOP10F' and Listonella anguillarum, but did not agglutinate Gram-positive bacteria Bacillus thuringiensis and Micrococcus luteus, and the agglutination could be inhibited by EDTA, indicating that AiCTL5 was a Ca(2+)-dependent lectin. rAiCTL5 exhibited a significantly strong activity to bind LPS from E. coli, which conformed to the agglutinating activity toward Gram-negative bacteria. Moreover, rAiCTL5 also agglutinated rabbit erythrocytes. These results indicated that AiCTL5 could function as a pattern recognition receptor to protect bay scallop from Gram-negative bacterial infection, and also provide evidence to understand the structural and functional diverse of lectin.

  15. Characterization of the cysK2-ctl1-cysE2 gene cluster involved in sulfur metabolism in Lactobacillus casei.

    Science.gov (United States)

    Bogicevic, Biljana; Irmler, Stefan; Portmann, Reto; Meile, Leo; Berthoud, Hélène

    2012-01-16

    The up- and downstream regions of ctl1 and ctl2 that encode a cystathionine lyase were analyzed in various Lactobacillus casei strains. ctl1 and ctl2 were found to be part of a gene cluster encoding two other open reading frames. One of the two open reading frames precedes ctl1 and encodes a putative cysteine synthase. The other open reading frame lies downstream of ctl1 and encodes a putative serine acetyltransferase. The gene cluster is not present in the publicly available genome sequences of L. casei ATCC 334, BL23 and Zhang. Apparently, the gene cluster was acquired by a horizontal gene transfer event and can also be found in other lactic acid bacteria such as Lactobacillus helveticus, Lactobacillus delbrueckii subsp. bulgaricus and Streptococcus thermophilus. RT-PCR was used to analyze the expression of the gene cluster. Additionally, an mass spectrometry-based selected reaction monitoring method was developed for quantifying Ctl1 in a cell-free extract of lactic acid bacteria. The gene cluster cysK2-ctl1-cysE2 was expressed as single transcript, and expression was down-regulated by cysteine. In addition, cystathionine lyase activity present in cell-free extracts disappeared when L. casei was grown in the presence of cysteine. Whereas the transcript and the gene product of ctl1 protein were found in all studied ctl1(+)L. casei strains, only the transcript but not the protein or cystathionine lyase activity was detected in L. helveticus FAM2888, L. delbrueckii subsp. bulgaricus ATCC 11842 and S. thermophilus FAM17014, which actually possess a homolog of the cysK2-ctl1-cysE2 gene cluster.

  16. Optimization and immune recognition of multiple novel conserved HLA-A2, human immunodeficiency virus type 1-specific CTL epitopes

    DEFF Research Database (Denmark)

    Corbet, Sylvie; Nielsen, Henrik Vedel; Vinner, Lasse

    2003-01-01

    and more conserved. Such epitope peptides were anchor-optimized to improve immunogenicity and further increase the number of potential vaccine epitopes. About 67 % of anchor-optimized vaccine epitopes induced immune responses against the corresponding non-immunogenic naturally occurring epitopes....... This study demonstrates the potency of ANNs for identifying putative virus CTL epitopes, and the new HIV-1 CTL epitopes identified should have significant implications for HIV-1 vaccine development. As a novel vaccine approach, it is proposed to increase the coverage of HIV variants by including multiple...

  17. Cytotoxic-T-Lymphocyte-Mediated Elimination of Target Cells Transduced with Engineered Adeno-Associated Virus Type 2 Vector In Vivo▿

    OpenAIRE

    Li, Chengwen; Hirsch, Matt; DiPrimio, Nina; Asokan, Aravind; Goudy, Kevin; Tisch, Roland; Samulski, R. Jude

    2009-01-01

    A recent clinical trial in patients with hemophilia B has suggested that adeno-associated virus (AAV) capsid-specific cytotoxic T lymphocytes (CTLs) eliminated AAV-transduced hepatocytes and resulted in therapeutic failure. AAV capsids elicit a CTL response in animal models; however, these capsid-specific CTLs fail to kill AAV-transduced target cells in mice. To better model the human clinical trial data in mice, we introduced an immunodominant epitope derived from ovalbumin (OVA; SIINFEKL) i...

  18. Measurement of background translocation frequencies in individuals with clones

    Energy Technology Data Exchange (ETDEWEB)

    Wade, M.J.

    1996-08-01

    In the leukemia case the unseparated B and T lymphocytes had a high translocation frequency even after 0.0014, respectively. After purging all clones from the data, the translocation frequencies for Bio 8 and Bio 23 were 0.00750.0014 and 0.0073 metaphases were scored for chromosomal aberrations,, specifically reciprocal translocations, using fluorescence in situ hybridization (FISH). Metaphase spreads were used from two healthy, unexposed individuals (not exposed to radiation, chemotherapy or radiotherapy) and one early B- precursor acute lymphocytic leukemia (ALL) patient (metaphase spreads from both separated T lymphocytes and unseparated B and T lymphocytes were scored). All three individuals had an abnormally high translocation frequency. The high translocation frequencies resulted from clonal expansion of specific translocated chromosomes. I show in this thesis that by purging (discounting or removing) clones from the data of unexposed individuals, one can obtain true background translocation frequencies. In two cases, Bio 8 and Bio 23, the measured translocation frequency for chromosomes 1, 2 and 4 was 0.0124 purging all of the clones from the data. This high translocation frequency may be due to a low frequency of some clones and may not be recognized. The separated T lymphocytes had a higher translocation frequency than expected.

  19. Induction of lytic pathways in T cell clones derived from wild-type or protein tyrosine kinase Fyn mutant mice.

    Science.gov (United States)

    Lancki, D W; Fields, P; Qian, D; Fitch, F W

    1995-08-01

    The OVA-reactive CD4+ Th1 clones and alloreactive CD8+ clones derived from wild-type or fyn-/- mice serve as model systems which have allowed us to investigate several aspects of the molecular events associated with T cell-mediated cytotoxicity, including 1) the differential utilization of two distinct cytolytic pathways by CD4+ Th1 clones and CD8+ CTL, 2) a comparison of the pathways of lysis induced by stimulation of the TCR or by alternative stimuli, 3) the requirement of Fyn for derivation of antigen-specific T-cell clones having properties of CD4+ Th1 and CD8+ CTL cells 4) the differential requirement of Fyn in the induction of responses by TCR and the alternative stimuli. Stimulation through the TCR, either by APC bearing relevant antigen or by immobilized anti-CD3 mAb, resulted in comparable levels of target cell lysis by clones from both wild-type and fyn-/- mice. These clones also utilize the Fas pathway to lyse target cells. Thus, Fyn does not appear to be required for expression of the Fas pathway when triggered through the TCR. In contrast, lysis of target cells by T-cell clones lacking Fyn was deficient when stimulated through Thy-1 or Ly-6C (using mAb) or with Con A or phorbol ester as compared to clones derived from wild-type mice. The basis for the defect in response to stimulation through the GPI-linked molecules appears to be a signaling defect which affects all of the functional responses we measured, while the defect in response to Con A stimulation appears to affect lysis but not lymphokine production. Thus, Fyn expression is selectively required for efficient activation of the Fas pathway of lysis through Thy-1, Ly-6C, and by Con A or phorbol ester in these T-cell clones. CD8+ clones derived from fyn-/- mutant mice, like clones derived from wild-type mice, display antigen-specific lysis, and appear to express perforin message and perforin protein. A Ca(++)-dependent (presumably perforin/exocytosis) component and Fas component of lysis was

  20. Clusters versus affinity-based approaches in F. tularensis whole genome search of CTL epitopes.

    Directory of Open Access Journals (Sweden)

    Anat Zvi

    Full Text Available Deciphering the cellular immunome of a bacterial pathogen is challenging due to the enormous number of putative peptidic determinants. State-of-the-art prediction methods developed in recent years enable to significantly reduce the number of peptides to be screened, yet the number of remaining candidates for experimental evaluation is still in the range of ten-thousands, even for a limited coverage of MHC alleles. We have recently established a resource-efficient approach for down selection of candidates and enrichment of true positives, based on selection of predicted MHC binders located in high density "hotspots" of putative epitopes. This cluster-based approach was applied to an unbiased, whole genome search of Francisella tularensis CTL epitopes and was shown to yield a 17-25 fold higher level of responders as compared to randomly selected predicted epitopes tested in Kb/Db C57BL/6 mice. In the present study, we further evaluate the cluster-based approach (down to a lower density range and compare this approach to the classical affinity-based approach by testing putative CTL epitopes with predicted IC(50 values of <10 nM. We demonstrate that while the percent of responders achieved by both approaches is similar, the profile of responders is different, and the predicted binding affinity of most responders in the cluster-based approach is relatively low (geometric mean of 170 nM, rendering the two approaches complimentary. The cluster-based approach is further validated in BALB/c F. tularensis immunized mice belonging to another allelic restriction (Kd/Dd group. To date, the cluster-based approach yielded over 200 novel F. tularensis peptides eliciting a cellular response, all were verified as MHC class I binders, thereby substantially increasing the F. tularensis dataset of known CTL epitopes. The generality and power of the high density cluster-based approach suggest that it can be a valuable tool for identification of novel CTLs in

  1. Bendamustine Plus Alemtuzumab for Refractory Chronic Lymphocytic Leukemia (CLL)

    Science.gov (United States)

    2013-08-20

    Recurrent Small Lymphocytic Lymphoma; Refractory Chronic Lymphocytic Leukemia; Stage III Chronic Lymphocytic Leukemia; Stage III Small Lymphocytic Lymphoma; Stage IV Chronic Lymphocytic Leukemia; Stage IV Small Lymphocytic Lymphoma

  2. Human immunodeficiency virus type 1 infection of antigen-specific CD4 cytotoxic T lymphocytes.

    Science.gov (United States)

    Robbins, P A; Roderiquez, G L; Peden, K W; Norcross, M A

    1998-11-01

    The effect of macrophage (M)-tropic and T cell line (T)-tropic human immunodeficiency virus type 1 (HIV-1) infection on antigen-specific CD4 cytotoxic T lymphocytes (CTLs) has been studied using a CD4 CTL line specific for a peptide from influenza B virus hemagglutinin. In the absence of antigen presentation, the production of CC chemokines was low. Both the M-tropic HIV-1 strain (HIV-1AD) and the T-tropic HIV-1 strain (HIV-1LAI) established productive infections in the CD4 CTLs, decreasing antigen-specific cytotoxicity. Peptide presented to the CD4 CTLs increased their secretion of RANTES and MIP-1beta, suppressed M-tropic HIV-1 replication, downmodulated CCR5 expression, and preserved CTL recognition. The suppression of M-tropic HIV-1 replication and downmodulation of the CCR5 receptor likely resulted from CC chemokine secretion since antibodies to CC chemokines restored M-tropic HIV-1 replication. Antigen presentation did not protect CD4 CTLs from T-tropic HIV-1 infection or preserve their CTL recognition. Thus, these CD4 CTLs do not make suppressor factors that inhibit the T-tropic HIV-1LAI isolate. The results indicate that these CD4 CTLs can either harbor or suppress M-tropic HIV-1 infection, depending on whether antigen is present. CD4 CTLs might therefore provide some protection in the early stages of HIV-1 infection when M-tropic isolates are present.

  3. What Is Chronic Lymphocytic Leukemia?

    Science.gov (United States)

    ... Chronic Lymphocytic Leukemia (CLL) About Chronic Lymphocytic Leukemia What Is Chronic Lymphocytic Leukemia? Cancer starts when cells ... body, including the lymph nodes, liver, and spleen. What is leukemia? Leukemia is a cancer that starts ...

  4. Specific killing of P53 mutated tumor cell lines by a cross-reactive human HLA-A2-restricted P53-specific CTL line

    DEFF Research Database (Denmark)

    Würtzen, P A; Pedersen, L O; Poulsen, H S

    2001-01-01

    , 4 HLA-A2(+) p53-mutated tumor cell lines were lysed by the CTL line, indicating that these peptides are endogenously processed and presented on HLA-A2 molecule. Thus, monocyte-derived DC pulsed with a pool of peptides are able to induce CTL reactivity to wild-type p53 peptides presented by several...

  5. Alvocidib in Treating Patients With B-Cell Chronic Lymphocytic Leukemia or Small Lymphocytic Lymphoma

    Science.gov (United States)

    2013-07-01

    B-cell Chronic Lymphocytic Leukemia; Contiguous Stage II Small Lymphocytic Lymphoma; Noncontiguous Stage II Small Lymphocytic Lymphoma; Stage I Chronic Lymphocytic Leukemia; Stage I Small Lymphocytic Lymphoma; Stage II Chronic Lymphocytic Leukemia; Stage III Chronic Lymphocytic Leukemia; Stage III Small Lymphocytic Lymphoma; Stage IV Chronic Lymphocytic Leukemia; Stage IV Small Lymphocytic Lymphoma

  6. Local Model Checking of Weighted CTL with Upper-Bound Constraints

    DEFF Research Database (Denmark)

    Jensen, Jonas Finnemann; Larsen, Kim Guldstrand; Srba, Jiri

    2013-01-01

    graphs. We implement all algorithms in a publicly available tool prototype and evaluate them on several experiments. The principal conclusion is that our local algorithm is the most efficient one with an order of magnitude improvement for model checking problems with a high number of “witnesses”.......We present a symbolic extension of dependency graphs by Liu and Smolka in order to model-check weighted Kripke structures against the logic CTL with upper-bound weight constraints. Our extension introduces a new type of edges into dependency graphs and lifts the computation of fixed-points from...... boolean domain to nonnegative integers in order to cope with the weights. We present both global and local algorithms for the fixed-point computation on symbolic dependency graphs and argue for the advantages of our approach compared to the direct encoding of the model checking problem into dependency...

  7. Universal cytotoxic activity of a HTLV-1 Tax-specific T cell clone from an HLA-A*24:02⁺ patient with adult T-cell leukemia against a variety of HTLV-I-infected T-cells.

    Science.gov (United States)

    Tanaka, Yukie; Yamazaki, Rie; Terasako-Saito, Kiriko; Nakasone, Hideki; Akahoshi, Yu; Nakano, Hirofumi; Ugai, Tomotaka; Wada, Hidenori; Yamasaki, Ryoko; Ishihara, Yuko; Kawamura, Koji; Sakamoto, Kana; Ashizawa, Masahiro; Sato, Miki; Kimura, Shun-ichi; Kikuchi, Misato; Kako, Shinichi; Kanda, Junya; Tanihara, Aki; Nishida, Junji; Kanda, Yoshinobu

    2014-01-01

    Adult T cell leukemia/lymphoma (ATL) is an aggressive mature T cell malignancy that is causally associated with human T cell lymphotropic virus type 1 (HTLV-1) infection. The HTLV-1 regulatory protein Tax aggressively accelerates the proliferation of host cells and is also an important target antigen for CD8(+) cytotoxic T cells (CTLs). We previously reported that several predominant HLA-A*24:02-restricted HTLV-1 Tax301-309-specific CTL clones commonly expressed a particular amino acid sequence motif (P-D-R) in complementarity-determining region 3 of T-cell receptor (TCR)-β chain among unrelated ATL patients who underwent allogeneic stem cell transplantation (allo-HSCT). Furthermore, a PDR-motif(+) CTL clone persistently existed in a long-term survivor as a central CTL clone with strong CTL activities after HSCT. Although a larger analysis of the relationship between PDR-motif(+) CTLs and the clinical course is required, the expression of PDR-motif(+) TCR on CD8(+) T cells may play a critical role in the management of anti-HTLV-1 activities for HLA-A24:02(+) ATL patients. Therefore, in this study, we prepared an HTLV-1 Tax301-309 peptide-specific CTL clone (HT-9) expressing PDR-motif(+) TCR isolated from a long-term survivor after HSCT, and evaluated its CTL activity against a variety of HTLV-1-infected T-cells from HLA-A*24:02(+) ATL patients. Before the assay of CTL function, we confirmed that HT-9 expressed less-differentiated effector-memory phenotypes (CD45RA(-)CCR7(-)CD27(+)CD28(+/-)CD57(+/-)) and T-cell exhaustion marker PD-1(+). In assays of CTL function, HT-9 recognized HTLV-1 Tax in an HLA-restricted fashion and demonstrated strong CTL activities against a variety of HTLV-1-infected T-cells from HLA-A*24:02(+) ATL patients regardless of whether the sources were autologous or allogeneic, but not normal cells. These data indicate that PDR-motif(+) TCR could be an important TCR candidate for TCR-gene immunotherapy for HLA-A24:02(+) ATL patients, provided

  8. Enhancement of lytic activity of leukemic cells by CD8+ cytotoxic T lymphocytes generated against a WT1 peptide analogue.

    Science.gov (United States)

    Al Qudaihi, Ghofran; Lehe, Cynthia; Negash, Muna; Al-Alwan, Monther; Ghebeh, Hazem; Mohamed, Said Yousuf; Saleh, Abu-Jafar Mohammed; Al-Humaidan, Hind; Tbakhi, Abdelghani; Dickinson, Anne; Aljurf, Mahmoud; Dermime, Said

    2009-02-01

    The Wilms tumor antigen 1 (WT1) antigen is over-expressed in human leukemias, making it an attractive target for immunotherapy. Most WT1-specific Cytotoxic T Lymphocytes (CTLs) described so far displayed low avidity, limiting its function. To improve the immunogenicity of CTL epitopes, we replaced the first-amino-acid of two known immunogenic WT1-peptides (126 and 187) with a tyrosine. This modification enhances 126Y analogue-binding ability, triggers significant number of IFN-gamma-producing T cells (P = 0.0003), induces CTL that cross-react with the wild-type peptide, exerts a significant lytic activity against peptide-loaded-targets (P = 0.0006) and HLA-A0201-matched-leukemic cells (P = 0.0014). These data support peptide modification as a feasible approach for the development of a leukemia-vaccine.

  9. A HLA-A2 restricted human CTL line recognizes a novel tumor cell expressed p53 epitope

    DEFF Research Database (Denmark)

    Würtzen, Peter A; Claesson, Mogens H

    2002-01-01

    line, indicating that this novel p53 peptide epitope is endogenously processed and presented by the HLA-A2 molecules of the tumor cells. In conclusion, CTL reactivity towards a wild-type p53 peptide was revealed through induction with DC pulsed with a pool of HLA-A2 binding p53 peptides. In addition...

  10. Development of oral CTL vaccine using a CTP-integrated Sabin 1 poliovirus-based vector system.

    Science.gov (United States)

    Han, Seung-Soo; Lee, Jinjoo; Jung, Yideul; Kang, Myeong-Ho; Hong, Jung-Hyub; Cha, Min-Suk; Park, Yu-Jin; Lee, Ezra; Yoon, Cheol-Hee; Bae, Yong-Soo

    2015-09-11

    We developed a CTL vaccine vector by modification of the RPS-Vax system, a mucosal vaccine vector derived from a poliovirus Sabin 1 strain, and generated an oral CTL vaccine against HIV-1. A DNA fragment encoding a cytoplasmic transduction peptide (CTP) was integrated into the RPS-Vax system to generate RPS-CTP, a CTL vaccine vector. An HIV-1 p24 cDNA fragment was introduced into the RPS-CTP vector system and a recombinant poliovirus (rec-PV) named vRPS-CTP/p24 was produced. vRPS-CTP/p24 was genetically stable and efficiently induced Th1 immunity and p24-specific CTLs in immunized poliovirus receptor-transgenic (PVR-Tg) mice. In challenge experiments, PVR-Tg mice that were pre-immunized orally with vRPS-CTP/p24 were resistant to challenge with a lethal dose of p24-expressing recombinant vaccinia virus (rMVA-p24). These results suggested that the RPS-CTP vector system had potential for developing oral CTL vaccines against infectious diseases.

  11. TCR gamma delta cytotoxic T lymphocytes expressing the killer cell-inhibitory receptor p58.2 (CD158b) selectively lyse acute myeloid leukemia cells.

    Science.gov (United States)

    Dolstra, H; Fredrix, H; van der Meer, A; de Witte, T; Figdor, C; van de Wiel-van Kemenade, E

    2001-05-01

    Cytotoxic T lymphocytes (CTL) are thought to play an important role in the graft-versus-leukemia (GVL) response. Unfortunately, GVL reactivity is often associated with life-threatening graft-versus-host disease (GVHD). Characterization of CTL that selectively attack leukemic cells but not normal cells may lead to the development of adjuvant immunotherapy that separates GVL from GVHD. Here, we describe TCR gamma delta (V gamma 9/V delta 1) CTL, isolated from the peripheral blood of an AML patient after stem cell transplantation (SCT), that very efficiently lysed freshly isolated acute myeloid leukemia (AML) cells and AML cell lines. Interestingly, HLA-matched non-malignant hematopoietic cells were not killed. We revealed that the killer cell-inhibitory receptor (KIR) p58.2 (CD158b) specific for group 2 HLA-C molecules negatively regulates the cytotoxic effector function displayed by these TCR gamma delta CTL. First, an antibody against HLA-C enhances lysis of non-malignant cells. Secondly, stable transfection of HLA-Cw*0304 into the class I-negative cell line 721.221 inhibited lysis. Finally, engagement of p58.2 by antibodies immobilized on Fc gamma R-expressing murine P815 cells inhibits CD3- and TCR gamma delta-directed lysis. Compared to non-malignant hematopoietic cells, AML cells express much lower levels of MHC class I molecules making them susceptible to lysis by p58.2(+) TCR gamma delta CTL. Such KIR-regulated CTL reactivity may have a role in the GVL response without affecting normal tissues of the host and leading to GVHD.

  12. Cytotoxic-T-lymphocyte-mediated elimination of target cells transduced with engineered adeno-associated virus type 2 vector in vivo.

    Science.gov (United States)

    Li, Chengwen; Hirsch, Matt; DiPrimio, Nina; Asokan, Aravind; Goudy, Kevin; Tisch, Roland; Samulski, R Jude

    2009-07-01

    A recent clinical trial in patients with hemophilia B has suggested that adeno-associated virus (AAV) capsid-specific cytotoxic T lymphocytes (CTLs) eliminated AAV-transduced hepatocytes and resulted in therapeutic failure. AAV capsids elicit a CTL response in animal models; however, these capsid-specific CTLs fail to kill AAV-transduced target cells in mice. To better model the human clinical trial data in mice, we introduced an immunodominant epitope derived from ovalbumin (OVA; SIINFEKL) into the AAV capsid and tested CTL-mediated killing of AAV2-transduced target tissues in vivo. Initially, in vitro experiments demonstrated both classical class I and cross-presentation of the OVA antigen, following endogenous expression or AAV2-OVA vector transduction, respectively. Furthermore, an OVA-specific CTL response was elicited after muscular or systemic injection of the AAV2-OVA vector. Finally, CTL reactivity was enhanced in mice with established SIINFEKL-specific immunity after AAV2-OVA/alpha1 anti-trypsin (AAT) administration. Most importantly, these OVA-specific CTLs decreased AAT expression in mice treated with AAV2-OVA/AAT vector that followed a time course mimicking uncoating kinetics of AAV2 transduction in OVA-immunized mice. These results demonstrate that AAV capsid-derived antigens elicit CD8(+) CTL reactivity, and these CTLs eliminated AAV-transduced target cells in mice. Notably, this model system can be exploited to study the kinetics of capsid presentation from different serotypes of AAV and permit the design of novel strategies to block CTL-mediated killing of AAV-transduced cells.

  13. JAK-STAT-mediated chronic inflammation impairs cytotoxic T lymphocyte activation to decrease anti-PD-1 immunotherapy efficacy in pancreatic cancer.

    Science.gov (United States)

    Lu, Chunwan; Talukder, Asif; Savage, Natasha M; Singh, Nagendra; Liu, Kebin

    2017-01-01

    Human pancreatic cancer does not respond to immune check point blockade immunotherapy. One key feature of pancreatic cancer is the association between its progression and chronic inflammation. Emerging evidence supports a key role for the JAK-STAT pathway in pancreatic cancer inflammation. We aimed at testing the hypothesis that sustained JAK-STAT signaling suppresses cytotoxic T lymphocyte (CTL) activation to counteract anti-PD-1 immunotherapy-induced CTL activity in pancreatic cancer. We show that human pancreatic carcinomas express high level of PD-L1 and exhibit low level of CTL infiltration. JAK-STAT inhibitor Ruxolitinib selectively inhibits STAT1 and STAT3 activation and increases CTL infiltration to induce a Tc1/Th1 immune response in the tumor microenvironment in an orthotopic pancreatic cancer mouse model. Ruxilitinib-mediated tumor suppressive efficacy diminishes in T-cell-deficient mice. Pancreatic tumor grows significantly faster in IFNγ-deficient mice. However, neutralizing IFNγ does not alter tumor growth but diminishes Ruxolitinib-induced tumor suppression in vivo, indicating that lymphocytes and IFNγ are essential for Ruxolitinib-induced host antitumor immune response. Both type I and type II interferons upregulate PD-L1 expression through the JAK-STAT signaling pathway in mouse pancreatic tumor cells. Tumor cells respond to activated T cells by activating STAT3. The inhibition of STAT3 downregulates immune suppressive cytokines production by tumor cells, resulting in increased T cell activation and effector function. Consequently, Ruxolitinib significantly improves the efficacy of anti-PD-1 immunotherapy. Our data demonstrate that Ruxolitinib is effective in the inhibition of systemic inflammation in the tumor microenvironment and therefore upregulates CTL infiltration and activation to overcome pancreatic cancer resistance to anti-PD-1 immunotherapy.

  14. Histone deacetylase inhibitors impair the elimination of HIV-infected cells by cytotoxic T-lymphocytes.

    Directory of Open Access Journals (Sweden)

    Richard Brad Jones

    2014-08-01

    Full Text Available Resting memory CD4+ T-cells harboring latent HIV proviruses represent a critical barrier to viral eradication. Histone deacetylase inhibitors (HDACis, such as suberanilohydroxamic acid (SAHA, romidepsin, and panobinostat have been shown to induce HIV expression in these resting cells. Recently, it has been demonstrated that the low levels of viral gene expression induced by a candidate HDACi may be insufficient to cause the death of infected cells by viral cytopathic effects, necessitating their elimination by immune effectors, such as cytotoxic T-lymphocytes (CTL. Here, we study the impact of three HDACis in clinical development on T-cell effector functions. We report two modes of HDACi-induced functional impairment: i the rapid suppression of cytokine production from viable T-cells induced by all three HDACis ii the selective death of activated T-cells occurring at later time-points following transient exposures to romidepsin or, to a lesser extent, panobinostat. As a net result of these factors, HDACis impaired CTL-mediated IFN-γ production, as well as the elimination of HIV-infected or peptide-pulsed target cells, both in liquid culture and in collagen matrices. Romidepsin exerted greater inhibition of antiviral function than SAHA or panobinostat over the dose ranges tested. These data suggest that treatment with HDACis to mobilize the latent reservoir could have unintended negative impacts on the effector functions of CTL. This could influence the effectiveness of HDACi-based eradication strategies, by impairing elimination of infected cells, and is a critical consideration for trials where therapeutic interruptions are being contemplated, given the importance of CTL in containing rebound viremia.

  15. MOLECULAR CLONING OF HUMAN NEUROTROPHIN-4 GENE

    Institute of Scientific and Technical Information of China (English)

    2001-01-01

    Objective Cloning and sequencing of the human neurotrophin-4(hNT-4) gene.Methods With the chromosomal DNA of human blood lymphocytes as template,hNT-4 coding genes were amplified by polymerase chain reaction(PCR) and recombinated into phage vector pGEM-T Easy,which were sequenced by using Sanger's single stranded DNA terminal termination method.Results The sequence of the cloned gene is completely the same as that reported in the literature(GenBank data base,M86528).Conclusion This study successfully cloning and sequenced the gene of mhNT-4,and it would be convenient for us to study the expression of mhNT-4 in eukaryote,and to continue the research on the gene therapy of Alzheimer's disease intensively.This study indicate that the hNT-4 is conservative in different races and individuals.

  16. The Clone Factory

    Science.gov (United States)

    Stoddard, Beryl

    2005-01-01

    Have humans been cloned? Is it possible? Immediate interest is sparked when students are asked these questions. In response to their curiosity, the clone factory activity was developed to help them understand the process of cloning. In this activity, students reenact the cloning process, in a very simplified simulation. After completing the…

  17. Eliciting cytotoxic T lymphocytes against acute myeloid leukemia-derived antigens: evaluation of dendritic cell-leukemia cell hybrids and other antigen-loading strategies for dendritic cell-based vaccination.

    Science.gov (United States)

    Galea-Lauri, Joanna; Darling, David; Mufti, Ghulam; Harrison, Phillip; Farzaneh, Farzin

    2002-08-01

    Dendritic cells (DC) have been successfully used in clinical pilot studies to induce tumor-specific immunity as well as clinical response in selected patients. However, DC-based immunotherapy remains a challenge and several parameters need to be examined in order to optimize the induction of anti-tumor immune responses. This study focuses on DC vaccination for leukemia and evaluates the in vitro efficacy of three different strategies for generating antigen-loaded DC-based vaccines for the induction of major histocompatibility complex (MHC) class I-restricted anti-leukemia cytotoxic T lymphocyte (CTL) responses. These included direct fusion of DC with leukemia cells to generate DC-leukemia cell hybrids, and DC pulsed with either apoptotic leukemia cell fragments or whole tumor cell lysates. Using either the U937 cell line or primary human acute myeloid leukemia blasts (AML), DC-leukemia cell hybrids were found to be the most potent in vitro inducers of CTL activity. DC pulsed with apoptotic tumor cell fragments were less efficient, but induced a more potent CTL response compared to tumor lysate-pulsed DC. The CTL responses were both MHC class I-restricted and antigen-specific, as shown by the inability of the CTL to lyse other control targets. The data presented here suggest that the method of antigen loading onto DC may be critical in the design of tumor vaccines.

  18. Maintenance or Emergence of Chronic Phase Secondary Cytotoxic T Lymphocyte Responses after Loss of Acute Phase Immunodominant Responses Does Not Protect SIV-Infected Rhesus Macaques from Disease Progression

    Directory of Open Access Journals (Sweden)

    M. Shannon Keckler

    2010-01-01

    Full Text Available The simian immunodeficiency virus- (SIV- infected rhesus macaque is the preferred animal model for vaccine development, but the correlates of protection in this model are not completely understood. In this paper, we document the cytotoxic T lymphocyte (CTL response to SIV and its effects on viral evolution in an effort to identify events associated with disease progression regardless of MHC allele expression. We observed the evolution of epitopes targeted by CTLs in a group of macaques that included long-term nonprogressing (LTNP, slowly progressing (SP, normally progressing (NP, and rapidly progressing (RP animals. Collectively, our data (1 identify novel CTL epitopes from an SP animal that are not restricted by known protective alleles, (2 illustrate that, in this small study, RP and NP animals accrue more mutations in CTL epitopes than in SP or LTNP macaques, and (3 demonstrate that the loss of CTL responses to immunodominant epitopes is associated with viral replication increases, which are not controlled by secondary CTL responses. These findings provide further evidence for the critical role of the primary cell-mediated immune responses in the control of retroviral infections.

  19. Cloning of observables

    OpenAIRE

    Ferraro, Alessandro; Galbiati, Matteo; Paris, Matteo G. A.

    2005-01-01

    We introduce the concept of cloning for classes of observables and classify cloning machines for qubit systems according to the number of parameters needed to describe the class under investigation. A no-cloning theorem for observables is derived and the connections between cloning of observables and joint measurements of noncommuting observables are elucidated. Relationships with cloning of states and non-demolition measurements are also analyzed.

  20. Acute Lymphocytic Leukemia

    Science.gov (United States)

    ... for information in your local library and on the Internet. Good sources include the National Cancer Institute, the ... mayoclinic.org/diseases-conditions/acute-lymphocytic-leukemia/basics/definition/CON-20042915 . Mayo Clinic Footer Legal Conditions and ...

  1. The clinical implications of mixed lymphocyte reaction with leukemic cells.

    Science.gov (United States)

    Kim, Hee-Je; Kim, Tai-Gyu; Cho, Hyun Il; Han, Hoon; Min, Woo-Sung; Kim, Chun-Choo

    2002-11-01

    To evaluate the clinical implications of a mixed lymphocyte reaction between leukemic cells and lymphocytes from HLA-matched sibling donors, we attempted to generate donor-derived, graft-versus-leukemia-effective cells and to define their characteristics. We studied 8 patients with chronic myelogenous leukemia (CML), including 5 patients in the chronic phase (CP), 3 patients in the accelerated phase (AP), and 2 patients with acute myelogenous leukemia (AML) in their first complete remission. Cells from these patients were used as stimulators in a mixed lymphocyte reaction.The effects of natural killer (NK) cells and cytotoxic T-lymphocytes (CTLs) were separated by observing tests for cytotoxicity to target cells, including K562 cells, the patient's leukemic cells, and phytohemagglutinin (PHA) blasts. Donor-derived antileukemic CTLs againstthe patient's own leukemic cells are productive in vitro. The efficacy of generating CTLs against leukemic target cells was (in decreasing order) AML, CML-CP, and CML-AP. Cytotoxic activity against leukemic targets was prominent in 4 cases--2 CML-CP and the 2 AML cases. On the contrary, the 3 cases of CML-AP showed low CTL activity. In cases showing 1 positive result among 3 targets (K562 cells, the patient's leukemic cells, and PHA blasts), the relapse rate was significantly lower (P = .022) on follow-up (median, 33 months; 7-40 months) after hematopoietic stem cell transplantation. By a combined analysis of the cytotoxicity effects for all 3 target cells, we were able to demonstrate a correlation between leukemic relapse and the variable degree of the cytotoxicity test results. Although the total sample numbers for this study were low, we speculate that these results may come from differences in the individual characteristics of the leukemic cells that are in line with their clinical disease status.

  2. Molecular cloning and characterization of duck interleukin-17

    Science.gov (United States)

    Interleukin-17 (IL-17) belonging to the Th17 family is a proinflammatory cytokine produced by activated T cells. A 1034-bp cDNA encoding duck IL-17 (duIL-17) was cloned from ConA-activated splenic lymphocytes of ducks. The encoded protein, predicted to consisted of 169 amino acids, displayed a molec...

  3. Chitinase-like (CTL) and cellulose synthase (CESA) gene expression in gelatinous-type cellulosic walls of flax (Linum usitatissimum L.) bast fibers.

    Science.gov (United States)

    Mokshina, Natalia; Gorshkova, Tatyana; Deyholos, Michael K

    2014-01-01

    Plant chitinases (EC 3.2.1.14) and chitinase-like (CTL) proteins have diverse functions including cell wall biosynthesis and disease resistance. We analyzed the expression of 34 chitinase and chitinase-like genes of flax (collectively referred to as LusCTLs), belonging to glycoside hydrolase family 19 (GH19). Analysis of the transcript expression patterns of LusCTLs in the stem and other tissues identified three transcripts (LusCTL19, LusCTL20, LusCTL21) that were highly enriched in developing bast fibers, which form cellulose-rich gelatinous-type cell walls. The same three genes had low relative expression in tissues with primary cell walls and in xylem, which forms a xylan type of secondary cell wall. Phylogenetic analysis of the LusCTLs identified a flax-specific sub-group that was not represented in any of other genomes queried. To provide further context for the gene expression analysis, we also conducted phylogenetic and expression analysis of the cellulose synthase (CESA) family genes of flax, and found that expression of secondary wall-type LusCESAs (LusCESA4, LusCESA7 and LusCESA8) was correlated with the expression of two LusCTLs (LusCTL1, LusCTL2) that were the most highly enriched in xylem. The expression of LusCTL19, LusCTL20, and LusCTL21 was not correlated with that of any CESA subgroup. These results defined a distinct type of CTLs that may have novel functions specific to the development of the gelatinous (G-type) cellulosic walls.

  4. Chitinase-like (CTL and cellulose synthase (CESA gene expression in gelatinous-type cellulosic walls of flax (Linum usitatissimum L. bast fibers.

    Directory of Open Access Journals (Sweden)

    Natalia Mokshina

    Full Text Available Plant chitinases (EC 3.2.1.14 and chitinase-like (CTL proteins have diverse functions including cell wall biosynthesis and disease resistance. We analyzed the expression of 34 chitinase and chitinase-like genes of flax (collectively referred to as LusCTLs, belonging to glycoside hydrolase family 19 (GH19. Analysis of the transcript expression patterns of LusCTLs in the stem and other tissues identified three transcripts (LusCTL19, LusCTL20, LusCTL21 that were highly enriched in developing bast fibers, which form cellulose-rich gelatinous-type cell walls. The same three genes had low relative expression in tissues with primary cell walls and in xylem, which forms a xylan type of secondary cell wall. Phylogenetic analysis of the LusCTLs identified a flax-specific sub-group that was not represented in any of other genomes queried. To provide further context for the gene expression analysis, we also conducted phylogenetic and expression analysis of the cellulose synthase (CESA family genes of flax, and found that expression of secondary wall-type LusCESAs (LusCESA4, LusCESA7 and LusCESA8 was correlated with the expression of two LusCTLs (LusCTL1, LusCTL2 that were the most highly enriched in xylem. The expression of LusCTL19, LusCTL20, and LusCTL21 was not correlated with that of any CESA subgroup. These results defined a distinct type of CTLs that may have novel functions specific to the development of the gelatinous (G-type cellulosic walls.

  5. In vitro methods for generating CD8+ T-cell clones for immunotherapy from the naïve repertoire.

    Science.gov (United States)

    Ho, William Y; Nguyen, Hieu N; Wolfl, Matthias; Kuball, Juergen; Greenberg, Philip D

    2006-03-20

    Innovations in gene discovery and the analysis of gene expression are facilitating the identification of a growing number of antigens that could potentially be targeted for immunotherapy of tumors. Methods to reliably generate antigen-specific T-cell responses in vitro would be useful not only to screen candidate antigens for immunogenicity prior to embarking on in vivo vaccination trials, but also to generate T-cell lines or clones that could be used directly for adoptive immunotherapy approaches. Although many techniques have proven successful for expanding ex vivo effector cells from antigen-specific memory CD8(+) cells that have been primed in vivo, methods to reliably generate high-avidity CTL clones from the naïve repertoire have not been well described. Various methods for the induction and expansion of antigen-specific CD8(+) CTL clones from healthy A2(+) donors were compared, using WT1 as a model tumor-associated antigen for which there is a low frequency of precursor T cells in naïve individuals. In contrast to the well-studied Melan-A/MART-1 (Melan-A) A2-restricted response, for which the CD8(+) T-cell precursor frequency in the naïve repertoire is unusually high, successful expansion of WT1-specific CD8(+) T cells appeared to be more dependent upon cell culture conditions. In particular, primary stimulation with autologous peptide-loaded monocyte-derived DC generated in 48 h (DC2d) was more effective in expanding WT1-reactive populations of CTL than stimulation with DC generated using the more standard week-long protocol (DC7d). Adding supplemental IL-7 2 to 3 days after initiation of a stimulation cycle expanded antigen-specific cells within CTL lines more efficiently than including the cytokine from the beginning of the cycle. Following primary stimulation with peptide-loaded mature DC, subsequent restimulation with peptide-loaded PBMC as the stimulators was more effective at expanding antigen-specific cells than repeated stimulation with mature DC

  6. Dendritic cells pulsed with alpha-fetoprotein and mutant P53 fused gene induce bi-targeted cytotoxic T lymphocyte response against hepatic carcinoma.

    Science.gov (United States)

    Ren, Jun; Jia, Jun; Zhang, Hongmei; Zhang, Liwang; Ma, Bo; Jiang, Hanfang; Di, Lijun; Song, Guohong; Yu, Jing

    2008-07-01

    Dendritic cell (DC)-based immunotherapy is rapidly emerging as a promising treatment in cancer therapy. We had previously shown that DC pulsed with either defined mRNA of tumor antigen (Ag) such as alpha-fetoprotein (AFP), or total RNA of hepatocellular carcinoma (HCC) could elicit Ag-specific cytotoxic T lymphocyte (CTL) response. Therefore, we suggested a novel DC-based therapeutic method, in which DCs derived from CD34(+) cells enriched peripheral blood mononuclear cells were pulsed with liposome-coated AFP and mutant P53 (mtP53) fused gene pEGFP-C3/AFP-mtP53 to induce bi-targeted specific CTL responses against HCC. Three different genotype HCC cell lines, HepG2 (human histocompatibility leukocyte antigens (HLA) A2 positive, AFP expressing positive, P53 expressing negative), SMMC7721 (HLA A2 positive, neither AFP nor P53 expressing positive), and HMCC97 (HLA A2 positive, both AFP and P53 expressing positive) were selected as targets for CTL responses. An important finding was that DCs pulsed with the liposome-coated fused gene could evoke more intensive bi-targeted Ag-specific CTL responses against HMCC97 than DCs pulsed with either AFP or P53 single gene (P gene of different Ags might induce more extensive multitargeted antitumor immunity.

  7. CD8-dependent CTL require co-engagement of CD8 and the TCR for phosphatidylinositol hydrolysis, but CD8-independent CTL do not and can kill in the absence of phosphatidylinositol hydrolysis.

    Science.gov (United States)

    Knall, C; Smith, P A; Potter, T A

    1995-06-01

    Most instances of MHC class I recognition and target cell killing by CD8+ CTL require the involvement of CD8. The role of CD8 in these events may be both for adhesion of the CTL with the APC, as well as for signal transduction through the TCR. The precise mechanism by which CD8 mediates signal transduction remains enigmatic. Similarly, it is unclear whether only the CD8 molecules which bind to the same class I molecule as the TCR contribute to signaling in the T cell responding to antigen. We have investigated the requirement for co-engagement of CD8 and the TCR in the induction of the hydrolysis of phosphatidylinositol-4,5-bisphosphate (PIP2) during the interaction of CTL and APC transfected with either wild-type or mutant (CD8 non-binding) class I molecules. Our results show that for conventional CD8-dependent killing co-engagement of both CD8 and the TCR is required to initiate PIP2 hydrolysis. This requirement for co-engagement, however, can be overcome by a high density of ligand, such as that provided by high concentrations of exogenous peptide. In such situations, the binding of CD8 to non-antigenic class I molecules can elicit PIP2 hydrolysis. Therefore, during interactions between CTL and APC, which generally occur at low concentrations of antigenic peptide, triggering of PIP2 hydrolysis requires TCR and CD8 co-engagement, and the binding of CD8 to non-antigenic class I molecules does not contribute significantly to signaling within the T cell.(ABSTRACT TRUNCATED AT 250 WORDS)

  8. Dynamics of a Delayed HIV-1 Infection Model with Saturation Incidence Rate and CTL Immune Response

    Science.gov (United States)

    Guo, Ting; Liu, Haihong; Xu, Chenglin; Yan, Fang

    2016-12-01

    In this paper, we investigate the dynamics of a five-dimensional virus model incorporating saturation incidence rate, CTL immune response and three time delays which represent the latent period, virus production period and immune response delay, respectively. We begin this model by proving the positivity and boundedness of the solutions. Our model admits three possible equilibrium solutions, namely the infection-free equilibrium E0, the infectious equilibrium without immune response E1 and the infectious equilibrium with immune response E2. Moreover, by analyzing corresponding characteristic equations, the local stability of each of the feasible equilibria and the existence of Hopf bifurcation at the equilibrium point E2 are established, respectively. Further, by using fluctuation lemma and suitable Lyapunov functionals, it is shown that E0 is globally asymptotically stable when the basic reproductive numbers for viral infection R0 is less than unity. When the basic reproductive numbers for immune response R1 is less than unity and R0 is greater than unity, the equilibrium point E1 is globally asymptotically stable. Finally, some numerical simulations are carried out for illustrating the theoretical results.

  9. HIV-1 Gag-specific exosome-targeted T cell-based vaccine stimulates effector CTL responses leading to therapeutic and long-term immunity against Gag/HLA-A2-expressing B16 melanoma in transgenic HLA-A2 mice

    Directory of Open Access Journals (Sweden)

    Rong Wang

    2014-01-01

    Full Text Available Human immunodeficiency virus type-1 (HIV-1-specific dendritic cell (DC vaccines have been applied to clinical trials that show only induction of some degree of immune responses, warranting the search of other more efficient vaccine strategies. Since HIV-1-specific CD8+ cytotoxic T lymphocytes (CTLs have been found to recognize some HIV-1 structural protein Gag conserved and cross-strain epitopes, Gag has become one of the most attractive target candidates for HIV-1 vaccine development. In this study, we generated HIV-1 Gag-specific Gag-Texo vaccine by using ConA-stimulated polyclonal CD8+ T-cells with uptake of Gag-expressing adenoviral vector AdVGag-transfected DC (DCGag-released exosomes (EXOs, and assessed its stimulation of Gag-specific CD8+ CTL responses and antitumor immunity. We demonstrate that Gag-Texo and DCGag vaccines comparably stimulate Gag-specific effector CD8+ CTL responses. Gag-Texo-stimulated CTL responses result in protective immunity against Gag-expressing BL6-10Gag melanoma in 8/8 wild-type C57BL/6 mice. In addition, we show that Gag-Texo vaccine also induces CTL responses leading to protective and long-term immunity against Gag/HLA-A2-expressing BL6-10Gag/A2 melanoma in 8/8 and 2/8 transgenic HLA-A2 mice, respectively. The average number of lung tumor colonies in mice with 30-days post-immunization is only 23, which is significantly less than that (>300 in control ConA-T-immunized HLA-A2 mice. Furthermore, Gag-Texo vaccine also induces some degree of therapeutic immunity. The average number (50 and size (0.23 mm in diameter of lung tumor colonies in Gag-Texo-immunized HLA-A2 mice bearing 6-day-established lung BL6-10Gag/A2 melanoma metastasis are significantly less than the average number (>300 and size (1.02 mm in diameter in control ConA-T-immunized HLA-A2 mice. Taken together, HIV-1 Gag-Texo vaccine capable of stimulating Gag-specific CTL responses and therapeutic immunity may be useful as a new immunotherapeutic

  10. Candidate mosaic proteins for a pan-filoviral cytotoxic T-Cell lymphocyte vaccine

    Energy Technology Data Exchange (ETDEWEB)

    Fenimore, Paul W [Los Alamos National Laboratory; Fischer, William M [Los Alamos National Laboratory; Kuiken, Carla [Los Alamos National Laboratory; Foley, Brian T [Los Alamos National Laboratory; Thurmond, J R [Los Alamos National Laboratory; Yusim, K [Los Alamos National Laboratory; Korber, B T [Los Alamos National Laboratory

    2008-01-01

    The extremely high fatality rates of many filovirus (FILV) strains the recurrent but rarely identified origin of human epidemics, the only partly identified viral reservoirs and the continuing non-human primate epizootics in Africa make a broadly-protective filovirus vaccine highly desirable. Cytotoxic T-cells (CTL) have been shown to be protective in mice, guinea pigs and non-human primates. In murine models the cytotoxic T-cell epitopes that are protective against Ebola virus have been mapped and in non-human primates CTL-mediated protection between viral strains (John Dye: specify) has been demonstrated using two filoviral proteins, nucleoprotein (NP) and glycoprotein (GP). These immunological results suggest that the CTL avenue of immunity deserves consideration for a vaccine. The poorly-understood viral reservoirs means that it is difficult to predict what strains are likely to cause epidemics. Thus, there is a premium on developing a pan-filoviral vaccine. The genetic diversity of FILV is large, roughly the same scale as human immunodeficiency virus (HIV). This presents a serious challenge for the vaccine designer because a traditional vaccine aspiring to pan-filoviral coverage is likely to require the inclusion of many antigenic reagents. A recent method for optimizing cytotoxic T-cell lymphocyte epitope coverage with mosaic antigens was successful in improving potential CTL epitope coverage against HIV and may be useful in the context of very different viruses, such as the filoviruses discussed here. Mosaic proteins are recombinants composed of fragments of wild-type proteins joined at locations resulting in exclusively natural k-mers, 9 {le} k {le} 15, and having approximately the same length as the wild-type proteins. The use of mosaic antigens is motivated by three conjectures: (1) optimizing a mosaic protein to maximize coverage of k-mers found in a set of reference proteins will give better odds of including broadly-protective CTL epitopes in a vaccine

  11. Prediction of HLA-A 2.1-restricted CTL epitopes from IGFBP7 antigen of lung carcinoma

    Institute of Scientific and Technical Information of China (English)

    Zhao Weipeng; Long Haixia; Zhu Bo; Duan Yuzhong; Chen Zhengtang

    2009-01-01

    Objective: With the development of peptide-based cancer specific immunotherapy, the prediction of CTL epitopes from insulin-like growth factor-binding protein 7 (IGFBP7) is very important for some research about tumor metastasis. Because HLA-A2.1-expressing individuals cover >50% in the population of China, we aimed at identifying IGFBP7-encoded peptide presented by HLA-A2.1. Methods: In our study, a HLA-A2.1 restricted CTL epitope was identified by using the following two-step procedure: (a) computer-based epitope prediction from the amino acid sequence of IGFBP7 antigen; (b) Validation with epitope molecular modeling. Results: We obtained four epitopes with high immunogenicity scores by all of the three algorithms, i.e., BIMAS, SYFPEITHI and IMTECH. Each of the four candidates satisfied the criteria of the HLA-A2.1-restricted CTL epitopes in molecular modeling analysis. Conclusion: The combination of BIMAS, SYFPEITHI and IMTECH method can improve the prediction efficiency and accuracy. Due to this research herein, this four epitopes have potential value for further studied, also have potential application in peptide-mediated immunotherapy. These epitopes may be useful in the design of therapeutic peptide vaccine for lung carcinoma and as immunotherapeutic strategies against lung carcinoma after identified by immunology experiment.

  12. CTL epitopes for influenza A including the H5N1 bird flu; genome-, pathogen-, and HLA-wide screening.

    Science.gov (United States)

    Wang, Mingjun; Lamberth, Kasper; Harndahl, Mikkel; Røder, Gustav; Stryhn, Anette; Larsen, Mette V; Nielsen, Morten; Lundegaard, Claus; Tang, Sheila T; Dziegiel, Morten H; Rosenkvist, Jørgen; Pedersen, Anders E; Buus, Søren; Claesson, Mogens H; Lund, Ole

    2007-04-12

    The purpose of the present study is to perform a global screening for new immunogenic HLA class I (HLA-I) restricted cytotoxic T cell (CTL) epitopes of potential utility as candidates of influenza A-virus diagnostics and vaccines. We used predictions of antigen processing and presentation, the latter encompassing 12 different HLA class I supertypes with >99% population coverage, and searched for conserved epitopes from available influenza A viral protein sequences. Peptides corresponding to 167 predicted peptide-HLA-I interactions were synthesized, tested for peptide-HLA-I interactions in a biochemical assay and for influenza-specific, HLA-I-restricted CTL responses in an IFN-gamma ELISPOT assay. Eighty-nine peptides could be confirmed as HLA-I binders, and 13 could be confirmed as CTL targets. The 13 epitopes, are highly conserved among human influenza A pathogens, and all of these epitopes are present in the emerging bird flu isolates. Our study demonstrates that present technology enables a fast global screening for T cell immune epitopes of potential diagnostics and vaccine interest. This technology includes immuno-bioinformatics predictors with the capacity to perform fast genome-, pathogen-, and HLA-wide searches for immune targets. To exploit this new potential, a coordinated international effort to analyze the precious source of information represented by rare patients, such as the current victims of bird flu, would be essential.

  13. Induction of protective anti-CTL epitope responses against HER-2-positive breast cancer based on multivalent T7 phage nanoparticles.

    Directory of Open Access Journals (Sweden)

    Somayeh Pouyanfard

    Full Text Available We report here the development of multivalent T7 bacteriophage nanoparticles displaying an immunodominant H-2k(d-restricted CTL epitope derived from the rat HER2/neu oncoprotein. The immunotherapeutic potential of the chimeric T7 nanoparticles as anti-cancer vaccine was investigated in BALB/c mice in an implantable breast tumor model. The results showed that T7 phage nanoparticles confer a high immunogenicity to the HER-2-derived minimal CTL epitope, as shown by inducing robust CTL responses. Furthermore, the chimeric nanoparticles protected mice against HER-2-positive tumor challenge in both prophylactic and therapeutic setting. In conclusion, these results suggest that CTL epitope-carrying T7 phage nanoparticles might be a promising approach for development of T cell epitope-based cancer vaccines.

  14. Statement on Human Cloning

    Science.gov (United States)

    ... ban on efforts to implant a human cloned embryo for the purpose of reproduction. The scientific evidence ... stem cell research, including the use of nuclear transplantation techniques (also known as research or therapeutic cloning), ...

  15. Ethical issues in cloning.

    Science.gov (United States)

    Satris, S

    2000-01-01

    There is great public concern with the ethics of human cloning. This paper briefly examines some of what I identify as pseudo-problems or myths associated with cloning, and some of the more substantial ethical concerns.

  16. Do HIV-specific CTL continue to have an antiviral function during antiretroviral therapy? If not, why not, and what can be done about it?

    OpenAIRE

    Dorian eMcIlroy

    2013-01-01

    Pharmacological re-activation of HIV expression from latent proviruses coupled with fully suppressive anti-retroviral therapy (ART) has been suggested as a strategy to eradicate HIV infection. In order for this strategy to be effective, latently infected cells must be killed either by the cytopathic effect of reactivated HIV gene expression, or by HIV-specific CTL. However, a review of current data reveals little evidence that CTL retain an anti-viral effector capacity in patients on fully su...

  17. An integrative approach to CTL epitope prediction: A combined algorithm integrating MHC class I binding, TAP transport efficiency, and proteasomal cleavage predictions

    DEFF Research Database (Denmark)

    Larsen, Mette Voldby; Lundegaard, Claus; Lamberth, K;

    2005-01-01

    Reverse immunogenetic approaches attempt to optimize the selection of candidate epitopes, and thus minimize the experimental effort needed to identify new epitopes. When predicting cytotoxic T cell epitopes, the main focus has been on the highly specific MHC class I binding event. Methods have al.......The method is available at http://www.cbs.dtu.dk/services/NetCTL. Supplementary material is available at http://www.cbs.dtu.dk/suppl/immunology/CTL.php....

  18. Blister fluid T lymphocytes during toxic epidermal necrolysis are functional cytotoxic cells which express human natural killer (NK) inhibitory receptors

    Science.gov (United States)

    Le Cleach, L; Delaire, S; Boumsell, L; Bagot, M; Bourgault-Villada, I; Bensussan, A; Roujeau, J C

    2000-01-01

    Toxic epidermal necrolysis (TEN) is a rare life-threatening adverse drug reaction characterized by a massive destruction of the epidermis. Immunohistological studies of skin biopsies of TEN showed infiltrates of predominantly CD8+ T lymphocytes even though other authors reported a prominent involvement of cells of the monocyte-macrophage lineage. The aim of this study was to characterize phenotypically and functionally the cells present in the cutaneous blister fluid of four patients with TEN. We first determined that lymphocytes were predominant in blister fluid obtained early, while monocytes/macrophages later became the most important population. We then showed that this lymphocyte population, mainly CD3+CD8+, corresponded to a peculiar cell subset as they expressed cutaneous leucocyte antigen, killer inhibitory receptors KIR/KAR and failed to express CD28 molecule. Functionally, we determined that blister T lymphocytes had a cytotoxic T lymphocyte (CTL)- and NK-like cytotoxicity. The role of this cytotoxic lymphocyte population present at the site of lesions during TEN remains to be understood. PMID:10606987

  19. Global Gene Expression Profiling in Interleukin-12-Induced Activation of CD8+ Cytotoxic T Lymphocytes against Mouse Mammary Carcinoma

    Institute of Scientific and Technical Information of China (English)

    Shanjin Cao; Zhaoying Xiang; Xiaojing Ma

    2004-01-01

    Interleukin-12 (IL-12) is a critical cytokine representing the link between the cellular and humoral branches of host immune defense apparatus. IL-12-induced cytotoxic lymphocyte (CTL) development is a central mechanism in immune responses against intracellular infectious agents as well as malignant growth. However,the molecular basis of tumor-specific CTL responses mediated by IL-12 remains poorly defined. In this study,we addressed this issue in a comprehensive manner to probe into IL-12-induced anti-tumor responses by global gene expression profiling of mRNA expression in CD8+T cells in a transplantable syngeneic mouse mammary carcinoma model treated or not with recombinant IL-12. A strong tumor regression was induced by the IL-12 treatment. An introspection of differential gene expression at an early stage of the IL-12-initiated CTL activation reveals interesting genes and molecular pathways that may account for the marked tumor regression,and is likely to provide a rich source of potential targets for further research and development of effective therapeutic modalities. Cellular & Molecular Immunology. 2004;1(5):357-366.

  20. Do HIV-specific CTL continue to have an antiviral function during antiretroviral therapy? If not, why not, and what can be done about it?

    Directory of Open Access Journals (Sweden)

    Dorian eMcIlroy

    2013-03-01

    Full Text Available Pharmacological re-activation of HIV expression from latent proviruses coupled with fully suppressive anti-retroviral therapy (ART has been suggested as a strategy to eradicate HIV infection. In order for this strategy to be effective, latently infected cells must be killed either by the cytopathic effect of reactivated HIV gene expression, or by HIV-specific CTL. However, a review of current data reveals little evidence that CTL retain an anti-viral effector capacity in patients on fully suppressive ART, implying that the HIV-specific CTL present in these patients will not be able to eliminate HIV-infected CD4+ T-cells effectively. If this is due to functional impairment or a quantitative deficit of HIV-specific CTL during ART, then therapeutic vaccination may improve the prospects for eradicating latent reservoirs. However, data from the macaque SIV model indicate that in vivo, SIV-specific CTL are only effective during the early stages of the viral replication cycle, and this constitutes an alternative explanation why HIV-specific CTL do not appear to have an impact on HIV reservoirs during ART. In that case, immunotoxins that target HIV-expressing cells may be a more promising approach for HIV eradication.

  1. Enhancing the cellulose-degrading activity of cellulolytic bacteria CTL-6 (Clostridium thermocellum) by co-culture with non-cellulolytic bacteria W2-10 (Geobacillus sp.).

    Science.gov (United States)

    Lü, Yucai; Li, Ning; Yuan, Xufeng; Hua, Binbin; Wang, Jungang; Ishii, Masaharu; Igarashi, Yasuo; Cui, Zongjun

    2013-12-01

    The effect of a non-cellulolytic bacterium W2-10 (Geobacillus sp.) on the cellulose-degrading activity of a cellulolytic bacterium CTL-6 (Clostridium thermocellum) was determined using cellulose materials (paper and straw) in peptone cellulose solution (PCS) medium under aerobic conditions. The results indicated that in the co-culture, addition of W2-10 resulted in a balanced medium pH, and may provide the required anaerobic environment for CTL-6. Overall, addition of W2-10 was beneficial to CTL-6 growth in the adverse environment of the PCS medium. In co-culture with W2-10, the CTL-6 cellulose degradation efficiency of filter paper and alkaline-treated wheat straw significantly increased up to 72.45 and 37.79 %, respectively. The CMCase activity and biomass of CTL-6 also increased from 0.23 U ml(-1) and 45.1 μg ml(-1) (DNA content) up to 0.47 U ml(-1) and 112.2 μg ml(-1), respectively. In addition, co-culture resulted in accumulation of acetate and propionate up to 4.26 and 2.76 mg ml(-1). This was a respective increase of 2.58 and 4.45 times, in comparison to the monoculture with CTL-6.

  2. T-cell receptor (TCR) phenotype of nodal Epstein-Barr virus (EBV)-positive cytotoxic T-cell lymphoma (CTL): a clinicopathologic study of 39 cases.

    Science.gov (United States)

    Kato, Seiichi; Asano, Naoko; Miyata-Takata, Tomoko; Takata, Katsuyoshi; Elsayed, Ahmed Ali; Satou, Akira; Takahashi, Emiko; Kinoshita, Tomohiro; Nakamura, Shigeo

    2015-04-01

    Among Epstein-Barr virus (EBV)-positive cytotoxic T/NK-cell lymphoma, there are only a few reports on the clinicopathologic features of patients with primary nodal presentation (nodal EBV cytotoxic T-cell lymphoma [CTL]). Here, we compared the clinicopathologic profiles of 39 patients with nodal EBV CTL with those of 27 cases of "extranasal" NK/T-cell lymphoma of nasal type (ENKTL), especially addressing their T-cell receptor (TCR) phenotype. Histologically, 22 of 39 nodal EBV CTL cases (56%) were unique in having centroblastoid appearance, which was contrasted with the lower incidence of this feature in ENKTL (15%, P=0.001). In contrast, pleomorphic appearance was more frequently seen in ENKTL than in nodal EBV CTL (67% vs. 23%, P=0.001). Thirty-three of 39 nodal EBV CTL cases (85%) were of T-cell lineage on the basis of TCR expression and/or TCRγ gene rearrangement; in detail, 18 cases (46%) were TCRβ positive (αβ T), 5 (13%) were TCRγ and/or δ positive (γδ T), and 10 (26%) were TCR-silent type with clonal TCRγ gene rearrangement but no expression of TCRβ, γ, or δ. These results were clearly contrasted by a lower incidence of T-cell lineage in ENKTL (7 cases, 26%, PEBV CTL is distinct from ENKTL.

  3. Cyclophosphamide, Alvocidib, and Rituximab in Treating Patients With High Risk B-Cell Chronic Lymphocytic Leukemia or Small Lymphocytic Lymphoma

    Science.gov (United States)

    2015-11-10

    Chronic Lymphocytic Leukemia; Prolymphocytic Leukemia; Recurrent Small Lymphocytic Lymphoma; Refractory Chronic Lymphocytic Leukemia; Stage I Chronic Lymphocytic Leukemia; Stage I Small Lymphocytic Lymphoma; Stage II Chronic Lymphocytic Leukemia; Stage II Small Lymphocytic Lymphoma; Stage III Chronic Lymphocytic Leukemia; Stage III Small Lymphocytic Lymphoma; Stage IV Chronic Lymphocytic Leukemia; Stage IV Small Lymphocytic Lymphoma

  4. Kinetics of circulating B lymphocytes in human myeloma

    Energy Technology Data Exchange (ETDEWEB)

    Boccadoro, M.; Gavarotti, P.; Fossati, G.; Massaia, M.; Pileri, A.; Durie, B.G.

    1983-04-01

    The tritiated thymidine labeling index (LI%) of peripheral B lymphocytes was studied in eight myeloma patients using simultaneous immunofluorescence and autoradiography. The LI% values were low (0.3%-5.1%), but significantly increased as compared to normal controls. In addition, there was excellent correlation between the LI% values and myeloma disease activity: lowest LI% values were observed in remission patients and the highest at the time of relapse. Simultaneous LI% evaluation of bone marrow myeloma cells in five patients gave concordant results, indicating the same kinetic behavior in both these compartments, particularly in the relapse phase. These data indicate both that circulating B lymphocytes include the neoplastic clone and that these B lymphocytes and bone marrow myeloma cells have similar kinetics.

  5. Quick and clean cloning.

    Science.gov (United States)

    Thieme, Frank; Marillonnet, Sylvestre

    2014-01-01

    Identification of unknown sequences that flank known sequences of interest requires PCR amplification of DNA fragments that contain the junction between the known and unknown flanking sequences. Since amplified products often contain a mixture of specific and nonspecific products, the quick and clean (QC) cloning procedure was developed to clone specific products only. QC cloning is a ligation-independent cloning procedure that relies on the exonuclease activity of T4 DNA polymerase to generate single-stranded extensions at the ends of the vector and insert. A specific feature of QC cloning is the use of vectors that contain a sequence called catching sequence that allows cloning specific products only. QC cloning is performed by a one-pot incubation of insert and vector in the presence of T4 DNA polymerase at room temperature for 10 min followed by direct transformation of the incubation mix in chemo-competent Escherichia coli cells.

  6. LYMPHOCYTE APOPTOSIS IN PSORIASIS

    Directory of Open Access Journals (Sweden)

    О. M. Kapuler

    2006-01-01

    Full Text Available Abstract. Forty-two patients with progressive vulgar psoriasis (PASI = 19.7 ± 1.5 and 40 healthy volunteers were under investigation. Psoriatic patients were characterized by increased number of CD4+ CD95+ peripheral blood T lymphocytes, which correlates with clinical psoriatic score, and by increased levels of soluble Fas (sFas in serum, as compared to controls (resp., 1868.1 ± 186.8 pg/ml vs. 1281.4 ± 142.5 pg/ml, PLSD = 0.019. The levels of spontaneous lymphocyte apoptosis and anti-Fas (Mab-induced apoptosis in psoriatic patients did not differ from the controls. However, apoptosis induced by “oxidative stress” (50 M Н202, 4 hrs was depressed in the patients. Moreover, a simultaneous assessment of cell cycle structure (metachromatic staining with Acridine Orange, apoptosis and Fas receptor expression (AnnV-FITC/antiFas mAbs-PE staining following a short-term mitogenic stimulation (PHA-P, 5 µg/ml, 24 hrs were performed. We found no marked differences in mitogenic reactivity, activation-induced apoptosis, and activation-induced Fas receptor expression when studying lymphocytes from healthy donors and psoriatic patients. However, PHA-activated lymphocytes from psoriatic patients displayed a significantly decreased ratio of AnnV+CD95+ to the total AnnV+ subpopulation, thus suggesting a decreased role of Fas-dependent mechanisms of apoptosis during the cell activation. The data obtained confirm a view, that an abnormal lymphocyte “apoptotic reactivity”, which plays a crucial role in the mechanisms of autoimmunity, may also of importance in the pathogenesis of psoriasis.

  7. Discovery of a Good Responder Subtype of Esophageal Squamous Cell Carcinoma with Cytotoxic T-Lymphocyte Signatures Activated by Chemoradiotherapy.

    Directory of Open Access Journals (Sweden)

    Yosuke Tanaka

    Full Text Available Definitive chemoradiotherapy (CRT is a less invasive therapy for esophageal squamous cell carcinoma (ESCC. Five-year survival rate of locally advanced ESCC patients by definitive CRT were 37%. We previously reported that tumor-specific cytotoxic T-lymphocyte (CTL activation signatures were preferentially found in long-term survivors. However, it is unknown whether the CTL activation is actually driven by CRT. We compared gene expression profiles among pre- and post-treatment biopsy specimens of 30 ESCC patients and 121 pre-treatment ESCC biopsy specimens. In the complete response (CR cases, 999 overexpressed genes including at least 234 tumor-specific CTL-activation associated genes such as IFNG, PRF1, and GZMB, were found in post-treatment biopsy specimens. Clustering analysis using expression profiles of these 234 genes allowed us to distinguish the immune-activated cases, designating them as I-type, from other cases. However, despite the better CR rate in the I-type, overall survival was not significantly better in both these 30 cases and another 121 cases. Further comparative study identified a series of epithelial to mesenchymal transition-related genes overexpressed in the early relapse cases. Importantly, the clinical outcome of CDH2-negative cases in the I-type was significantly better than that of the CDH2-positive cases in the I-type. Furthermore, NK cells, which were activated by neutrophils-producing S100A8/S100A9, and CTLs were suggested to cooperatively enhance the effect of CRT in the CDH2-negative I-type. These results suggested that CTL gene activation may provide a prognostic advantage in ESCCs with epithelial characteristics.

  8. Increased breadth and depth of cytotoxic T lymphocytes responses against HIV-1-B Nef by inclusion of epitope variant sequences.

    Directory of Open Access Journals (Sweden)

    Morgane Rolland

    Full Text Available Different vaccine approaches cope with HIV-1 diversity, ranging from centralized(1-4 to variability-encompassing(5-7 antigens. For all these strategies, a concern remains: how does HIV-1 diversity impact epitope recognition by the immune system? We studied the relationship between HIV-1 diversity and CD8(+ T Lymphocytes (CTL targeting of HIV-1 subtype B Nef using 944 peptides (10-mers overlapping by nine amino acids (AA that corresponded to consensus peptides and their most common variants in the HIV-1-B virus population. IFN-γ ELISpot assays were performed using freshly isolated PBMC from 26 HIV-1-infected persons. Three hundred and fifty peptides elicited a response in at least one individual. Individuals targeted a median of 7 discrete regions. Overall, 33% of responses were directed against viral variants but not elicited against consensus-based test peptides. However, there was no significant relationship between the frequency of a 10-mer in the viral population and either its frequency of recognition (Spearman's correlation coefficient ρ = 0.24 or the magnitude of the responses (ρ = 0.16. We found that peptides with a single mutation compared to the consensus were likely to be recognized (especially if the change was conservative and to elicit responses of similar magnitude as the consensus peptide. Our results indicate that cross-reactivity between rare and frequent variants is likely to play a role in the expansion of CTL responses, and that maximizing antigenic diversity in a vaccine may increase the breadth and depth of CTL responses. However, since there are few obvious preferred pathways to virologic escape, the diversity that may be required to block all potential escape pathways may be too large for a realistic vaccine to accommodate. Furthermore, since peptides were not recognized based on their frequency in the population, it remains unclear by which mechanisms variability-inclusive antigens (i.e., constructs enriched with

  9. Statistical mechanics of clonal expansion in lymphocyte networks modelled with slow and fast variables

    Science.gov (United States)

    Mozeika, Alexander; Coolen, Anthony C. C.

    2017-01-01

    We use statistical mechanical techniques to model the adaptive immune system, represented by lymphocyte networks in which B cells interact with T cells and antigen. We assume that B- and T-clones evolve in different thermal noise environments and on different timescales, and derive stationary distributions and study expansion of B clones for the case where these timescales are adiabatically separated. We compute characteristics of B-clone sizes, such as average concentrations, in parameter regimes where T-clone sizes are modelled as binary variables. This analysis is independent of the network topology, and its results are qualitatively consistent with experimental observations. To obtain the full distributions of B-clone sizes we assume further that the network topologies are random and locally equivalent to trees. This allows us to compete these distributions via the Bethe-Peierls approach. As an example we calculate B-clone distributions for immune models defined on random regular networks.

  10. Identification of Novel HLA-A*0201-Restricted CTL Epitopes in Chinese Vitiligo Patients

    Science.gov (United States)

    Cui, Tingting; Yi, Xiuli; Guo, Sen; Zhou, Fubo; Liu, Ling; Li, Chunying; Li, Kai; Gao, Tianwen

    2016-01-01

    Generalized vitiligo is an autoimmune disease characterized by melanocyte loss, which results in patchy depigmentation of skin and hair. Recent studies suggested the key role of CD8+T lymphocytes for mediating immune response in vitiligo through melanocyte differentiation antigens, including tyrosinase, gp100 and MelanA/Mart-1. However, the specific epitopes of these auto-antigens are still unknown. In our study, we predicted the possible HLA-A*0201-restricted nonapeptides overlaying the full-length amino acid sequences of these three known antigens and investigated the lymphocytes reactivity to these nonapeptides by Elispot assay. In addition, we evaluated the abilities of these nonapeptides to activate CD8+T cells. We screened out 5 possible epitopes originated from tyrosinase and gp100, numbered P28, P41, P112, P118 and P119. Among these 5 epitopes, notably, P28 and P119 played the dominant role in activating CTLs, with a significant increase in proliferation rate and Interferon-γ (IFN-γ) production of CD8+T cells. Nevertheless, antigen-specific T cell reactivity was not detected in MelanA/Mart-1 peptides. Our studies identified two novel epitopes originated from proteins of gp100 and tyrosinase, which may have implications for the development of immunotherapies for vitiligo. PMID:27821860

  11. Quantifying the impact of human immunodeficiency virus-1 escape from cytotoxic T-lymphocytes.

    Directory of Open Access Journals (Sweden)

    Ulrich D Kadolsky

    Full Text Available HIV-1 escape from the cytotoxic T-lymphocyte (CTL response leads to a weakening of viral control and is likely to be detrimental to the patient. To date, the impact of escape on viral load and CD4(+ T cell count has not been quantified, primarily because of sparse longitudinal data and the difficulty of separating cause and effect in cross-sectional studies. We use two independent methods to quantify the impact of HIV-1 escape from CTLs in chronic infection: mathematical modelling of escape and statistical analysis of a cross-sectional cohort. Mathematical modelling revealed a modest increase in log viral load of 0.051 copies ml(-1 per escape event. Analysis of the cross-sectional cohort revealed a significant positive association between viral load and the number of "escape events", after correcting for length of infection and rate of replication. We estimate that a single CTL escape event leads to a viral load increase of 0.11 log copies ml(-1 (95% confidence interval: 0.040-0.18, consistent with the predictions from the mathematical modelling. Overall, the number of escape events could only account for approximately 6% of the viral load variation in the cohort. Our findings indicate that although the loss of the CTL response for a single epitope results in a highly statistically significant increase in viral load, the biological impact is modest. We suggest that this small increase in viral load is explained by the small growth advantage of the variant relative to the wildtype virus. Escape from CTLs had a measurable, but unexpectedly low, impact on viral load in chronic infection.

  12. Low dose decitabine treatment induces CD80 expression in cancer cells and stimulates tumor specific cytotoxic T lymphocyte responses.

    Science.gov (United States)

    Wang, Li-Xin; Mei, Zhen-Yang; Zhou, Ji-Hao; Yao, Yu-Shi; Li, Yong-Hui; Xu, Yi-Han; Li, Jing-Xin; Gao, Xiao-Ning; Zhou, Min-Hang; Jiang, Meng-Meng; Gao, Li; Ding, Yi; Lu, Xue-Chun; Shi, Jin-Long; Luo, Xu-Feng; Wang, Jia; Wang, Li-Li; Qu, Chunfeng; Bai, Xue-Feng; Yu, Li

    2013-01-01

    Lack of immunogenicity of cancer cells has been considered a major reason for their failure in induction of a tumor specific T cell response. In this paper, we present evidence that decitabine (DAC), a DNA methylation inhibitor that is currently used for the treatment of myelodysplastic syndrome (MDS), acute myeloid leukemia (AML) and other malignant neoplasms, is capable of eliciting an anti-tumor cytotoxic T lymphocyte (CTL) response in mouse EL4 tumor model. C57BL/6 mice with established EL4 tumors were treated with DAC (1.0 mg/kg body weight) once daily for 5 days. We found that DAC treatment resulted in infiltration of IFN-γ producing T lymphocytes into tumors and caused tumor rejection. Depletion of CD8(+), but not CD4(+) T cells resumed tumor growth. DAC-induced CTL response appeared to be elicited by the induction of CD80 expression on tumor cells. Epigenetic evidence suggests that DAC induces CD80 expression in EL4 cells via demethylation of CpG dinucleotide sites in the promoter of CD80 gene. In addition, we also showed that a transient, low-dose DAC treatment can induce CD80 gene expression in a variety of human cancer cells. This study provides the first evidence that epigenetic modulation can induce the expression of a major T cell co-stimulatory molecule on cancer cells, which can overcome immune tolerance, and induce an efficient anti-tumor CTL response. The results have important implications in designing DAC-based cancer immunotherapy.

  13. Novel plant virus-based vaccine induces protective cytotoxic T-lymphocyte-mediated antiviral immunity through dendritic cell maturation.

    Science.gov (United States)

    Lacasse, Patrick; Denis, Jérôme; Lapointe, Réjean; Leclerc, Denis; Lamarre, Alain

    2008-01-01

    Currently used vaccines protect mainly through the production of neutralizing antibodies. However, antibodies confer little or no protection for a majority of chronic viral infections that require active involvement of cytotoxic T lymphocytes (CTLs). Virus-like particles (VLPs) have been shown to be efficient inducers of cell-mediated immune responses, but administration of an adjuvant is generally required. We recently reported the generation of a novel VLP system exploiting the self-assembly property of the papaya mosaic virus (PapMV) coat protein. We show here that uptake of PapMV-like particles by murine splenic dendritic cells (DCs) in vivo leads to their maturation, suggesting that they possess intrinsic adjuvant-like properties. DCs pulsed with PapMV-like particles displaying the lymphocytic choriomeningitis virus (LCMV) p33 immunodominant CTL epitope (PapMV-p33) efficiently process and cross-present the viral epitope to p33-specific transgenic T cells. Importantly, the CTL epitope is also properly processed and presented in vivo, since immunization of p33-specific T-cell receptor transgenic mice with PapMV-p33 induces the activation of large numbers of specific CTLs. C57BL/6 mice immunized with PapMV-p33 VLPs in the absence of adjuvant develop p33-specific effector CTLs that rapidly expand following LCMV challenge and protect vaccinated mice against LCMV infection in a dose-dependent manner. These results demonstrate the efficiency of this novel plant virus-based vaccination platform in inducing DC maturation leading to protective CTL responses.

  14. Low dose decitabine treatment induces CD80 expression in cancer cells and stimulates tumor specific cytotoxic T lymphocyte responses.

    Directory of Open Access Journals (Sweden)

    Li-Xin Wang

    Full Text Available Lack of immunogenicity of cancer cells has been considered a major reason for their failure in induction of a tumor specific T cell response. In this paper, we present evidence that decitabine (DAC, a DNA methylation inhibitor that is currently used for the treatment of myelodysplastic syndrome (MDS, acute myeloid leukemia (AML and other malignant neoplasms, is capable of eliciting an anti-tumor cytotoxic T lymphocyte (CTL response in mouse EL4 tumor model. C57BL/6 mice with established EL4 tumors were treated with DAC (1.0 mg/kg body weight once daily for 5 days. We found that DAC treatment resulted in infiltration of IFN-γ producing T lymphocytes into tumors and caused tumor rejection. Depletion of CD8(+, but not CD4(+ T cells resumed tumor growth. DAC-induced CTL response appeared to be elicited by the induction of CD80 expression on tumor cells. Epigenetic evidence suggests that DAC induces CD80 expression in EL4 cells via demethylation of CpG dinucleotide sites in the promoter of CD80 gene. In addition, we also showed that a transient, low-dose DAC treatment can induce CD80 gene expression in a variety of human cancer cells. This study provides the first evidence that epigenetic modulation can induce the expression of a major T cell co-stimulatory molecule on cancer cells, which can overcome immune tolerance, and induce an efficient anti-tumor CTL response. The results have important implications in designing DAC-based cancer immunotherapy.

  15. Statement on Human Cloning

    Science.gov (United States)

    ... as our understanding of this technology advances. Support Stem Cell Research (including Research Cloning) AAAS supports stem cell research, including the use of nuclear transplantation techniques (also ...

  16. 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.

  17. Effects of interval and continuous exercise training on CD4 lymphocyte apoptotic and autophagic responses to hypoxic stress in sedentary men.

    Directory of Open Access Journals (Sweden)

    Tzu-Pin Weng

    Full Text Available Exercise is linked with the type/intensity-dependent adaptive immune responses, whereas hypoxic stress facilitates the programmed death of CD4 lymphocytes. This study investigated how high intensity-interval (HIT and moderate intensity-continuous (MCT exercise training influence hypoxia-induced apoptosis and autophagy of CD4 lymphocytes in sedentary men. Thirty healthy sedentary males were randomized to engage either HIT (3-minute intervals at 40% and 80%VO2max, n=10 or MCT (sustained 60%VO2max, n=10 for 30 minutes/day, 5 days/week for 5 weeks, or to a control group that did not received exercise intervention (CTL, n=10. CD4 lymphocyte apoptotic and autophagic responses to hypoxic exercise (HE, 100 W under 12%O2 for 30 minutes were determined before and after various regimens. The results demonstrated that HIT exhibited higher enhancements of pulmonary ventilation, cardiac output, and VO2 at ventilatory threshold and peak performance than MCT did. Before the intervention, HE significantly down-regulated autophagy by decreased beclin-1, Atg-1, LC3-II, Atg-12, and LAMP-2 expressions and acridine orange staining, and simultaneously enhanced apoptosis by increased phospho-Bcl-2 and active caspase-9/-3 levels and phosphotidylserine exposure in CD4 lymphocytes. However, five weeks of HIT and MCT, but not CTL, reduced the extents of declined autophagy and potentiated apoptosis in CD4 lymphocytes caused by HE. Furthermore, both HIT and MCT regimens manifestly lowered plasma myeloperoxidase and interleukin-4 levels and elevated the ratio of interleukin-4 to interferon-γ at rest and following HE. Therefore, we conclude that HIT is superior to MCT for enhancing aerobic fitness. Moreover, either HIT or MCT effectively depresses apoptosis and promotes autophagy in CD4 lymphocytes and is accompanied by increased interleukin-4/interferon-γ ratio and decreased peroxide production during HE.

  18. Dendritic cells transduced with Rsf-1/HBXAP gene generate specific cytotoxic T lymphocytes against ovarian cancer in vitro

    Energy Technology Data Exchange (ETDEWEB)

    Sun, Li [Department of Gynecology Oncology, Shan Dong Tumor Hospital, Jinan, Shandong (China); Department of Obstetrics and Gynecology, Qilu Hospital, Shandong University, Jinan, Shandong (China); Kong, Beihua, E-mail: kongbeihua@sdu.edu.cn [Department of Obstetrics and Gynecology, Qilu Hospital, Shandong University, Jinan, Shandong (China); Sheng, Xiugui [Department of Gynecology Oncology, Shan Dong Tumor Hospital, Jinan, Shandong (China); Sheu, Jim Jinn-Chyuan [Human Genetic Center, China Medical University Hospital and Graduate Institute of Chinese Medical Science, China Medical University, Taichung City, Taiwan (China); Shih, Ie-Ming [Departments of Pathology, Oncology, and Gynecology and Obstetrics, Johns Hopkins Medical Institutions, Baltimore, MD 21231 (United States)

    2010-04-09

    Recently, some studies have indicated that Rsf-1/HBXAP plays a role in chromatin remodeling and transcriptional regulation that may contribute to tumorigenesis in ovarian cancer. The present study demonstrates that using dendritic cells (DCs) from human cord blood CD34{sup +} cells transduced with Rsf-1/HBXAP DNA plasmids by nucleofection generate specific cytotoxic T lymphocytes (CTL) against ovarian cancer in vitro. After transfection, DCs were analyzed for Rsf-1/HBXAP mRNA expression by RT-PCR and protein expression by Western blot. Then the DC phenotypes, T-cell stimulatory capacity, endocytic activity and migration capacity were explored by flow cytometry analysis, allogeneic mixed lymphocyte reaction, endocytosis and transwell chemotaxis assay, respectively. After transfection, Rsf-1/HBXAP expression was detected at mRNA and protein levels. Allogeneic T-cell proliferation induced by transfected DCs was obviously higher than non-transfected DCs, but the endocytosis capacity and migratory ability were not different. Rsf-1/HBXAP gene-transduced DCs could induce antigen-specific CTL and generate a very potent cytotoxicity to OVCAR3 cells. These data suggest that Rsf-1/HBXAP gene-transduced DCs may be a potential adjuvant immunotherapy for ovarian cancer in clinical applications.

  19. Curcumin and Cholecalciferol in Treating Patients With Previously Untreated Stage 0-II Chronic Lymphocytic Leukemia or Small Lymphocytic Lymphoma

    Science.gov (United States)

    2016-10-04

    Contiguous Stage II Small Lymphocytic Lymphoma; Noncontiguous Stage II Small Lymphocytic Lymphoma; Stage 0 Chronic Lymphocytic Leukemia; Stage I Chronic Lymphocytic Leukemia; Stage I Small Lymphocytic Lymphoma; Stage II Chronic Lymphocytic Leukemia

  20. Cloning-free CRISPR

    NARCIS (Netherlands)

    Arbab, Mandana; Srinivasan, Sharanya; Hashimoto, Tatsunori; Geijsen, Niels; Sherwood, Richard I

    2015-01-01

    We present self-cloning CRISPR/Cas9 (scCRISPR), a technology that allows for CRISPR/Cas9-mediated genomic mutation and site-specific knockin transgene creation within several hours by circumventing the need to clone a site-specific single-guide RNA (sgRNA) or knockin homology construct for each targ

  1. Cloning-free CRISPR

    NARCIS (Netherlands)

    Arbab, Mandana; Srinivasan, Sharanya; Hashimoto, Tatsunori; Geijsen, Niels; Sherwood, Richard I

    2015-01-01

    We present self-cloning CRISPR/Cas9 (scCRISPR), a technology that allows for CRISPR/Cas9-mediated genomic mutation and site-specific knockin transgene creation within several hours by circumventing the need to clone a site-specific single-guide RNA (sgRNA) or knockin homology construct for each

  2. Cloning-free CRISPR

    NARCIS (Netherlands)

    Arbab, Mandana; Srinivasan, Sharanya; Hashimoto, Tatsunori; Geijsen, Niels|info:eu-repo/dai/nl/194303403; Sherwood, Richard I

    2015-01-01

    We present self-cloning CRISPR/Cas9 (scCRISPR), a technology that allows for CRISPR/Cas9-mediated genomic mutation and site-specific knockin transgene creation within several hours by circumventing the need to clone a site-specific single-guide RNA (sgRNA) or knockin homology construct for each targ

  3. Estimating the number of hematopoietic or lymphoid stem cells giving rise to clonal chromosome aberrations in blood T lymphocytes.

    Science.gov (United States)

    Nakano, M; Kodama, Y; Ohtaki, K; Itoh, M; Awa, A A; Cologne, J; Kusunoki, Y; Nakamura, N

    2004-03-01

    Quantifying the proliferative capacity of long-term hematopoietic stem cells in humans is important for bone marrow transplantation and gene therapy. Obtaining appropriate data is difficult, however, because the experimental tools are limited. We hypothesized that tracking clonal descendants originating from hematopoietic stem cells would be possible if we used clonal chromosome aberrations as unique tags of individual hematopoietic stem cells in vivo. Using FISH, we screened 500 blood T lymphocytes from each of 513 atomic bomb survivors and detected 96 clones composed of at least three cells with identical aberrations. The number of clones was inversely related to their population size, which we interpreted to mean that the progenitor cells were heterogeneous in the number of progeny that they could produce. The absolute number of progenitor cells contributing to the formation of the observed clones was estimated as about two in an unexposed individual. Further, scrutiny of ten clones revealed that lymphocyte clones could originate roughly equally from hematopoietic stem cells or from mature T lymphocytes, thereby suggesting that the estimated two progenitor cells are shared as one hematopoietic stem cell and one mature T cell. Our model predicts that one out of ten people bears a non- aberrant clone comprising >10% of the total lymphocytes, which indicates that clonal expansions are common and probably are not health-threatening.

  4. In vitro culture of skin-homing T lymphocytes from inflammatory skin diseases

    DEFF Research Database (Denmark)

    Bang, Karen; C Mogensen, Søren; Thestrup-Pedersen, Kristian

    2005-01-01

    -chain studies by using reverse transcription-polymerase chain reaction showed that all T lymphocytes had access to the skin compartment. Single-stranded conformational analysis showed clonally expanded T cells numbering between 40 and 60 clones. After approximately 2 months of growth, the mean CD4+ : CD8+ ratio...

  5. Lymphocyte 'homing' and chronic inflammation.

    Science.gov (United States)

    Sakai, Yasuhiro; Kobayashi, Motohiro

    2015-07-01

    Chronic inflammation is a response to prolonged exposure to injurious stimuli that harm and destroy tissues and promote lymphocyte infiltration into inflamed sites. Following progressive accumulation of lymphocytes, the histology of inflamed tissue begins to resemble that of peripheral lymphoid organs, which can be referred to as lymphoid neogenesis or formation of tertiary lymphoid tissues. Lymphocyte recruitment to inflamed tissues is also reminiscent of lymphocyte homing to peripheral lymphoid organs. In the latter, under physiological conditions, homing receptors expressed on lymphocytes adhere to vascular addressin expressed on high endothelial venules (HEVs), initiating a lymphocyte migration process composed of sequential adhesive interactions. Intriguingly, in chronic inflammation, HEV-like vessels are induced de novo, despite the fact that the inflamed site is not originally lymphoid tissue, and these vessels contribute to lymphocyte recruitment in a manner similar to physiological lymphocyte homing. In this review, we first describe physiological lymphocyte homing mechanisms focusing on vascular addressins. We then describe HEV-like vessel-mediated pathogenesis seen in various chronic inflammatory disorders such as Helicobacter pylori gastritis, inflammatory bowel disease (IBD), autoimmune pancreatitis and sclerosing sialadenitis, as well as chronic inflammatory cell neoplasm MALT lymphoma, with reference to our work and that of others.

  6. Lymphocyte Trafficking to Mucosal Tissues

    DEFF Research Database (Denmark)

    Mikhak, Zamaneh; Agace, William Winston; Luster, Andrew D.

    2015-01-01

    Lymphocytes are the key cells of the adaptive immune system that provide antigen-specific responses tailored to the context of antigen exposure. Through cytokine release and antibody production, lymphocytes orchestrate and amplify the recruitment and function of other immune cells and contribute...... to host defense against invading pathogens and the pathogenesis of many inflammatory diseases. Lymphocyte function is critically dependent on their ability to traffic into the correct anatomic locations at the appropriate times. This process is highly regulated and requires that lymphocytes interact...

  7. Lymphocytic Interstitial Pneumonia.

    Science.gov (United States)

    Panchabhai, Tanmay S; Farver, Carol; Highland, Kristin B

    2016-09-01

    Lymphocytic interstitial pneumonia (LIP) is a rare lung disease on the spectrum of benign pulmonary lymphoproliferative disorders. LIP is frequently associated with connective tissue diseases or infections. Idiopathic LIP is rare; every attempt must be made to diagnose underlying conditions when LIP is diagnosed. Computed tomography of the chest in patients with LIP may reveal ground-glass opacities, centrilobular and subpleural nodules, and randomly distributed thin-walled cysts. Demonstrating polyclonality with immunohistochemistry is the key to differentiating LIP from lymphoma. The 5-year mortality remains between 33% and 50% and is likely to vary based on the underlying disease process.

  8. Unsuccessful CTL transfusion in a case of post-BMT Epstein-Barr virus-associated lymphoproliferative disorder (EBV-LPD).

    Science.gov (United States)

    Imashuku, S; Goto, T; Matsumura, T; Naya, M; Yamori, M; Hojo, M; Hibi, S; Todo, S

    1997-08-01

    A patient with AML (FAB M4Eo) developed EBV-LPD 1.5 months after allogeneic BMT from his one locus-mismatched mother, the diagnosis being confirmed on day +82. Attempts to eradicate the monoclonally proliferating LPD using chemotherapy (VP16/dexamethasone) followed by two doses of EBV-specific CTL and one dose of unstimulated donor leukocytes were not successful. We assume delay of infusions (day +100, +107) and insufficient CTL cell doses (total 9.2 x 10(6)) may have been responsible for the poor outcome in this case.

  9. Frequent occurrence of highly expanded but unrelated B-cell clones in patients with multiple myeloma.

    Science.gov (United States)

    Kriangkum, Jitra; Motz, Sarah N; Debes Marun, Carina S; Lafarge, Sandrine T; Gibson, Spencer B; Venner, Christopher P; Johnston, James B; Belch, Andrew R; Pilarski, Linda M

    2013-01-01

    Clonal diversity in multiple myeloma (MM) includes both MM-related and MM-unrelated clonal expansions which are subject to dominance exerted by the MM clone. Here we show evidence for the existence of minor but highly expanded unrelated B-cell clones in patients with MM defined by their complementary determining region 3 (CDR3) peak. We further characterize these clones over the disease and subsequent treatment. Second clones were identified by their specific IgH-VDJ sequences that are distinct from those of dominant MM clones. Clonal frequencies were determined through semi-quantitative PCR, quantitative PCR and single-cell polymerase chain reaction of the clone-specific sequence. In 13/74 MM patients, more than one dominant CDR3 peak was identified with 12 patients (16%) being truly biclonal. Second clones had different frequencies, were found in different locations and were found in different cell types from the dominant MM clone. Where analysis was possible, they were shown to have chromosomal characteristic distinct from those of the MM clone. The frequency of the second clone also changed over the course of the disease and often persisted despite treatment. Molecularly-defined second clones are infrequent in monoclonal gammopathy of undetermined significance (MGUS, 1/43 individuals or 2%), suggesting that they may arise at relatively late stages of myelomagenesis. In further support of our findings, biclonal gammopathy and concomitant MM and CLL (chronic lymphocytic leukemia) were confirmed to originate from two unrelated clones. Our data supports the idea that the clone giving rise to symptomatic myeloma exerts clonal dominance to prevent expansion of other clones. MM and second clones may arise from an underlying niche permissive of clonal expansion. The clinical significance of these highly expanded but unrelated clones remains to be confirmed. Overall, our findings add new dimensions to evaluating related and unrelated clonal expansions in MM and the

  10. Frequent occurrence of highly expanded but unrelated B-cell clones in patients with multiple myeloma.

    Directory of Open Access Journals (Sweden)

    Jitra Kriangkum

    Full Text Available Clonal diversity in multiple myeloma (MM includes both MM-related and MM-unrelated clonal expansions which are subject to dominance exerted by the MM clone. Here we show evidence for the existence of minor but highly expanded unrelated B-cell clones in patients with MM defined by their complementary determining region 3 (CDR3 peak. We further characterize these clones over the disease and subsequent treatment. Second clones were identified by their specific IgH-VDJ sequences that are distinct from those of dominant MM clones. Clonal frequencies were determined through semi-quantitative PCR, quantitative PCR and single-cell polymerase chain reaction of the clone-specific sequence. In 13/74 MM patients, more than one dominant CDR3 peak was identified with 12 patients (16% being truly biclonal. Second clones had different frequencies, were found in different locations and were found in different cell types from the dominant MM clone. Where analysis was possible, they were shown to have chromosomal characteristic distinct from those of the MM clone. The frequency of the second clone also changed over the course of the disease and often persisted despite treatment. Molecularly-defined second clones are infrequent in monoclonal gammopathy of undetermined significance (MGUS, 1/43 individuals or 2%, suggesting that they may arise at relatively late stages of myelomagenesis. In further support of our findings, biclonal gammopathy and concomitant MM and CLL (chronic lymphocytic leukemia were confirmed to originate from two unrelated clones. Our data supports the idea that the clone giving rise to symptomatic myeloma exerts clonal dominance to prevent expansion of other clones. MM and second clones may arise from an underlying niche permissive of clonal expansion. The clinical significance of these highly expanded but unrelated clones remains to be confirmed. Overall, our findings add new dimensions to evaluating related and unrelated clonal expansions in

  11. CRTAM determines the CD4+ cytotoxic T lymphocyte lineage.

    Science.gov (United States)

    Takeuchi, Arata; Badr, Mohamed El Sherif Gadelhaq; Miyauchi, Kosuke; Ishihara, Chitose; Onishi, Reiko; Guo, Zijin; Sasaki, Yoshiteru; Ike, Hiroshi; Takumi, Akiko; Tsuji, Noriko M; Murakami, Yoshinori; Katakai, Tomoya; Kubo, Masato; Saito, Takashi

    2016-01-11

    Naive T cells differentiate into various effector T cells, including CD4(+) helper T cell subsets and CD8(+) cytotoxic T cells (CTL). Although cytotoxic CD4(+) T cells (CD4 +: CTL) also develop from naive T cells, the mechanism of development is elusive. We found that a small fraction of CD4(+) T cells that express class I-restricted T cell-associated molecule (CRTAM) upon activation possesses the characteristics of both CD4(+) and CD8(+) T cells. CRTAM(+) CD4(+) T cells secrete IFN-γ, express CTL-related genes, such as eomesodermin (Eomes), Granzyme B, and perforin, after cultivation, and exhibit cytotoxic function, suggesting that CRTAM(+) T cells are the precursor of CD4(+)CTL. Indeed, ectopic expression of CRTAM in T cells induced the production of IFN-γ, expression of CTL-related genes, and cytotoxic activity. The induction of CD4(+)CTL and IFN-γ production requires CRTAM-mediated intracellular signaling. CRTAM(+) T cells traffic to mucosal tissues and inflammatory sites and developed into CD4(+)CTL, which are involved in mediating protection against infection as well as inducing inflammatory response, depending on the circumstances, through IFN-γ secretion and cytotoxic activity. These results reveal that CRTAM is critical to instruct the differentiation of CD4(+)CTL through the induction of Eomes and CTL-related gene.

  12. Chronic lymphocytic leukaemia

    Science.gov (United States)

    Kipps, Thomas J.; Stevenson, Freda K.; Wu, Catherine J.; Croce, Carlo M.; Packham, Graham; Wierda, William G.; O’Brien, Susan; Gribben, John; Rai, Kanti

    2017-01-01

    Chronic lymphocytic leukaemia (CLL) is a malignancy of CD5+ B cells that is characterized by the accumulation of small, mature-appearing lymphocytes in the blood, marrow and lymphoid tissues. Signalling via surface immunoglobulin, which constitutes the major part of the B cell receptor, and several genetic alterations play a part in CLL pathogenesis, in addition to interactions between CLL cells and other cell types, such as stromal cells, T cells and nurse-like cells in the lymph nodes. The clinical progression of CLL is heterogeneous and ranges from patients who require treatment soon after diagnosis to others who do not require therapy for many years, if at all. Several factors, including the immunoglobulin heavy-chain variable region gene (IGHV) mutational status, genomic changes, patient age and the presence of comorbidities, should be considered when defining the optimal management strategies, which include chemotherapy, chemoimmunotherapy and/or drugs targeting B cell receptor signalling or inhibitors of apoptosis, such as BCL-2. Research on the biology of CLL has profoundly enhanced our ability to identify patients who are at higher risk for disease progression and our capacity to treat patients with drugs that selectively target distinctive phenotypic or physiological features of CLL. How these and other advances have shaped our current understanding and treatment of patients with CLL is the subject of this Primer. PMID:28102226

  13. Decreased deformability of lymphocytes in chronic lymphocytic leukemia

    Science.gov (United States)

    Zheng, Yi; Wen, Jun; Nguyen, John; Cachia, Mark A.; Wang, Chen; Sun, Yu

    2015-01-01

    This paper reports the first study of stiffness/deformability changes of lymphocytes in chronic lymphocytic leukemia (CLL) patients, demonstrating that at the single cell level, leukemic metastasis progresses are accompanied by biophysical property alterations. A microfluidic device was utilized to electrically measure cell volume and transit time of single lymphocytes from healthy and CLL patients. The results from testing thousands of cells reveal that lymphocytes from CLL patients have higher stiffness (i.e., lower deformability), as compared to lymphocytes in healthy samples, which was also confirmed by AFM indentation tests. This observation is in sharp contrast to the known knowledge on other types of metastatic cells (e.g., breast and lung cancer cells) whose stiffness becomes lower as metastasis progresses.

  14. Clonal dominance among T-lymphocyte infiltrates in arthritis

    Energy Technology Data Exchange (ETDEWEB)

    Stamenkovic, I.; Stegagno, M.; Wright, K.A.; Krane, S.M.; Amento, E.P.; Colvin, R.B.; Duquesnoy, R.J.; Kurnick, J.T.

    1988-02-01

    Synovial membranes in patients with rheumatoid arthritis as well as other types of chronic destructive inflammatory arthritis contain infiltrates of activated T lymphocytes that probably contribute to the pathogenesis of the disease. In an effort to elucidate the nature of these infiltrates, interleukin 2 (IL-2)-responsive T lymphocytes were grown out of synovial fragments from 14 patients undergoing surgery for advanced destructive inflammatory joint disease. Eleven of the samples examined were from patients with classical rheumatoid arthritis, while three others were obtained from individuals with clinical osteoarthritis. Southern blot analysis of T-cell receptor (TCR) ..beta..-chain genes in 13 of 14 cultures showed distinct rearrangements, indicating that each culture was characterized by the predominance of a limited number of clones. T-cell populations from peripheral blood stimulated with a variety of activators and expanded with IL-2 did not demonstrate evidence of similar clonality in long-term culture. These results suggest that a limited number of activated T-cell clones predominate at the site of tissue injury in rheumatoid synovial membranes as well as in other types of destructive inflammatory joint disease. Further characterization of these T-cell clones may aid our understanding of the pathogenesis of these rheumatic disorders.

  15. Use of "one-pot, mix-and-read" peptide-MHC class I tetramers and predictive algorithms to improve detection of cytotoxic T lymphocyte responses in cattle

    DEFF Research Database (Denmark)

    Svitek, Nicholas; Hansen, Andreas Martin; Steinaa, Lucilla

    2014-01-01

    Peptide-major histocompatibility complex (p-MHC) class I tetramer complexes have facilitated the early detection and functional characterisation of epitope specific CD8(+) cytotoxic T lymphocytes (CTL). Here, we report on the generation of seven recombinant bovine leukocyte antigens (BoLA....... parva CTL epitopes. Using an ELISA assay to measure peptide-BoLA monomer formation and p-MHC class I tetramers of new specificity, we demonstrate that a predicted alternative epitope Tp2(29-37) rather than the previously reported Tp2(27-37) epitope is the correct Tp2 epitope presented by BoLA-6......*04101. We also verified the prediction by NetMHCpan that the Tp5(87-95) epitope reported as BoLA-T5 restricted can also be presented by BoLA-1*02301, a molecule similar in sequence to BoLA-T5. In addition, Tp5(87-95) specific bovine CTL were simultaneously stained by Tp5-BoLA-1*02301 and Tp5-BoLA-T5...

  16. Global Gene Expression Profiling in Interleukin-12-Induced Activation of CD8+ Cytotoxic T Lymphocytes against MouseMammary Carcinoma

    Institute of Scientific and Technical Information of China (English)

    ShanjinCao; ZhaoyingXiang; XiaojingMa

    2004-01-01

    Interleukin-12 (IL-12) is a critical cytokine representing the link between the cellular and humoral branches of host immune defense apparatus. IL-12-induced cytotoxic lymphocyte (CTL) development is a central mechanism in immune responses against intracellular infectious agents as well as malignant growth. However,the molecular basis of tumor-specific CTL responses mediated by IL-12 remains poorly defined. In this study,we addressed this issue in a comprehensive manner to probe into IL-12-induced anti-tumor responses by global gene expression profiling of mRNA expression in CD8+T cells in a transplantable syngeneic mouse mammarycarcinoma model treated or not with recombinant IL-12. A strong tumor regression was induced by the IL-12 treatment. An introspection of differential gene expression at an early stage of the IL-12-initiated CTL activation reveals interesting genes and molecular pathways that may account for the marked tumor regression,and is likely to provide a rich source of potential targets for further research and development of effective therapeutic modalities. Cellular & Molecular Immunology. 2004;1(5):357-366.

  17. Cytotoxic T lymphocytes and CD4 epitope mutations in the pre-core/core region of hepatitis B virus in chronic hepatitis B carriers in Northeast Iran.

    Science.gov (United States)

    Zhand, Sareh; Tabarraei, Alijan; Nazari, Amineh; Moradi, Abdolvahab

    2017-07-25

    Hepatitis B virus (HBV) is vulnerable to many various mutations. Those within epitopes recognized by sensitized T cells may influence the re-emergence of the virus. This study was designed to investigate the mutation in immune epitope regions of HBV pre-core/core among chronic HBV patients of Golestan province, Northeast Iran. In 120 chronic HBV carriers, HBV DNA was extracted from blood plasma samples and PCR was done using specific primers. Direct sequencing and alignment of the pre-core/core region were applied using reference sequence from Gene Bank database (Accession Number AB033559). The study showed 27 inferred amino acid substitutions, 9 of which (33.3%) were in CD4 and 2 (7.4%) in cytotoxic T lymphocytes' (CTL) epitopes and 16 other mutations (59.2%) were observed in other regions. CTL escape mutations were not commonly observed in pre-core/core sequences of chronic HBV carriers in the locale of study. It can be concluded that most of the inferred amino acid substitutions occur in different immune epitopes other than CTL and CD4.

  18. NY-ESO-1 致敏树突状细胞诱导的细胞毒性 T 淋巴细胞对视网膜母细胞瘤细胞的特异性杀伤作用%The killing effect of NY-ESO-1-sensitzed dendritic cells-induced cellular toxic lymphocyte on human retinoblastoma cell

    Institute of Scientific and Technical Information of China (English)

    周学智; 周希瑗

    2012-01-01

    Background Cell immunologic therapy for retinoblastoma(RB)is becoming a hot research topic.Cancer-testis antigen is a human immunogenic protein and is used to treat some tumors.However,its effect on RB has not been investigated.Objective The present study was to discuss the antigen specific anti-tumor effect of cytotoxic T lymphocytes(CTL)induced by the cancer-testis antigen,NY-ESO-1-sensitized dendritic cells(DCs),on human RB.Methods PCR was performed to amplify target gene fragments from the NY-ESO-1 plasmids,and then the target gene fragments were digested with the restriction enzymes SalI and EcoRI.Harvested fragments were inserted into the pDC316 plasmid to construct the recombinant plasmid pDC316/NY-ESO-1.The expression of NY-ESO-1 protein in human RB cells strain,HXO-RB44,was detected by immunofluorescence and Western blot.Monocytes were isolated from 60 ml of peripheral blood from a healthy donor using Ficoll density-gradient centrifugation with a cell density of 1 × 107/ml.DCs isolated from blood were stimulated with recombinant human granulocyte-macrophage colony stimulating factor(rhGM-CSF)and recombinant human interleukin-4(rhIL-4).The recombinant plasmid pDC316/NY-ESO-1 was transfected into DCs and the DCs were co-cultured with T lymphocytes.The resultant CTL were used as effector cells.The growth of the CTL was detected by MTT assay.The CTL were then added into the growth medium used for culturing HXO-RB44 cells and the vitality of the HXO-RB44 cells was assayed by MTT assay.Results The sequence of the cloned DNA fragment of the recombinant plasmid pDC316/NY-ESO-1 was conforms with the sequence of the NY-ESO-1 gene.The expression of the NY-ESO-1 protein in HXO-RB44 cells was tested by immunofluorescence and Western blot.DCs were successfully induced with rhIL-4 and rhGM-CSF from PBMC.The recombinant expression plasmid pDC316/NY-ESO-1 was successfully transferred into DCs.These DCs had high expression of surface molecules such as HLA-DR(42.1%),CD80

  19. Obinutuzumab in chronic lymphocytic leukemia.

    Science.gov (United States)

    Dupuis, Jehan

    2015-09-01

    Obinutuzumab is the second next-generation monoclonal anti-CD20 antibody (after ofatumumab) to enter clinical practice in chronic lymphocytic leukemia. Its superiority in association with chlorambucil as compared with chlorambucil alone has led to its approval as a first-line treatment for chronic lymphocytic leukemia, for patients who are not candidates for a more intensive treatment.

  20. Enzyme free cloning for high throughput gene cloning and expression

    NARCIS (Netherlands)

    de Jong, R.N.; Daniëls, M.; Kaptein, R.; Folkers, G.E.

    2006-01-01

    Structural and functional genomics initiatives significantly improved cloning methods over the past few years. Although recombinational cloning is highly efficient, its costs urged us to search for an alternative high throughput (HTP) cloning method. We implemented a modified Enzyme Free Cloning (EF

  1. Enzyme free cloning for high throughput gene cloning and expression

    NARCIS (Netherlands)

    de Jong, R.N.; Daniëls, M.; Kaptein, R.; Folkers, G.E.

    2006-01-01

    Structural and functional genomics initiatives significantly improved cloning methods over the past few years. Although recombinational cloning is highly efficient, its costs urged us to search for an alternative high throughput (HTP) cloning method. We implemented a modified Enzyme Free Cloning

  2. Unified Approach to Universal Cloning and Phase-Covariant Cloning

    OpenAIRE

    Hu, Jia-Zhong; Yu, Zong-Wen; Wang, Xiang-Bin

    2008-01-01

    We analyze the problem of approximate quantum cloning when the quantum state is between two latitudes on the Bloch's sphere. We present an analytical formula for the optimized 1-to-2 cloning. The formula unifies the universal quantum cloning (UQCM) and the phase covariant quantum cloning.

  3. Enzyme free cloning for high throughput gene cloning and expression

    NARCIS (Netherlands)

    de Jong, R.N.; Daniëls, M.; Kaptein, R.; Folkers, G.E.

    2006-01-01

    Structural and functional genomics initiatives significantly improved cloning methods over the past few years. Although recombinational cloning is highly efficient, its costs urged us to search for an alternative high throughput (HTP) cloning method. We implemented a modified Enzyme Free Cloning (EF

  4. Functional balance between T cell chimeric receptor density and tumor associated antigen density: CTL mediated cytolysis and lymphokine production.

    Science.gov (United States)

    Weijtens, M E; Hart, E H; Bolhuis, R L

    2000-01-01

    Genetically engineered expression of tumor-specific single chain antibody chimeric receptors (ch-Rec) on human T lymphocytes endow these cells with the parental monoclonal antibody (mAb) dictated tumor specificity and may be useful for clinical immuno-genetherapy. Therefore it was of importance to assess how the densities of tumor-specific receptors and tumor associated antigens (TAA), respectively, affect primary human T lymphocyte functions in relation to target cell susceptibilities to lysis. We therefore studied the functional balance between ch-Rec densities on human T lymphocytes and TAA on tumor cells. The gene construct encoding a ch-Rec derived from (1) a renal carcinoma cell (RCC) specific mouse mAb (G250), and (2) the human signal transducing Fc(epsilon)RI gamma-chain was used. To obtain ch-RecHIGH-POS and ch-RecLOW-POS T lymphocytes, two distinct retroviral vectors were used to introduce the gene constructs into primary human T lymphocytes. Levels of ch-Rec-redirected T lymphocyte mediated tumor cell lysis, as well as lymphokine production were determined using RCC lines as target/stimulator cells, which express either no or increasing densities of the TAA. A functional and dynamic balance between ch-Rec densities on cytotoxic T lymphocytes (CTLs) on the one hand and TAA densities on RCCs on the other, was found. In short, ch-RecHIGH-POS CTLs are triggered by TAAHIGH-POS as well as TAALOW-POS RCCs to lyse tumor cells and produce (IFN-gamma and TNF-alpha) lymphokine. In contrast, ch-RecLOW-POS T lymphocytes are only triggered for cytolysis and lymphokine production by relatively TAAHIGH-POS RCCs. In conclusion, (1) the activation of T lymphocyte responses is co-determined by the expression levels of the ch-Rec on T lymphocytes and the TAA on tumor cells and (2) at relatively high T lymphocyte ch-Rec expression levels the CTLs lyse tumor cells with a wide range of TAA densities. Gene Therapy (2000) 7, 35-42.

  5. Main: Clone Detail [KOME

    Lifescience Database Archive (English)

    Full Text Available Clone Detail Mapping Pseudomolecule data detail Detail information Mapping to the TIGR japonica Pseudomolecu...les kome_mapping_pseudomolecule_data_detail.zip kome_mapping_pseudomolecule_data_detail ...

  6. BIOETHICS AND HUMAN CLONING

    Directory of Open Access Journals (Sweden)

    Željko Kaluđerović

    2011-12-01

    Full Text Available In this paper the authors analyze the process of negotiating and beginning of the United Nations Declaration on Human Cloning as well as the paragraphs of the very Declaration. The negotiation was originally conceived as a clear bioethical debate that should have led to a general agreement to ban human cloning. However, more often it had been discussed about human rights, cultural, civil and religious differences between people and about priorities in case of eventual conflicts between different value systems. In the end, a non-binding Declaration on Human Cloning had been adopted, full of numerous compromises and ambiguous formulations, that relativized the original intention of proposer states. According to authors, it would have been better if bioethical discussion and eventual regulations on cloning mentioned in the following text had been left over to certain professional bodies, and only after the public had been fully informed about it should relevant supranational organizations have taken that into consideration.

  7. Do Managers Clone Themselves?

    Science.gov (United States)

    Baron, Alma S.

    1981-01-01

    A recent questionnaire survey provides statistics on male managers' views of female managers. The author recommends that male managers break out of their cloning behavior and that the goal ought to be a plurality in management. (Author/WD)

  8. Murine T cell clones specific for Hymenolepis nana: generation and functional analysis in vivo and in vitro.

    Science.gov (United States)

    Asano, K; Okamoto, K

    1991-12-01

    To examine the role of the T cell in protective immunity to Hymenolepis nana, H. nana-specific clonal lymphocytes were generated from mesenteric lymph nodes of BALB/c mice infected with H. nana, and some of their functions were analyzed in vitro and in vivo. Following limiting dilution techniques, five clones were generated from mesenteric lymph node cell populations. All of these clones expressed the L3T4+, Lyt-2.2- phenotype and proliferated in vitro in response to soluble egg antigen of H. nana. Of five clones, three secreted interleukin 2 (IL-2) and interferon-gamma (IFN-gamma) after stimulation with egg antigen. Furthermore, these three clones conferred local delayed-type hypersensitivity to egg antigen. The remaining two clones produced interleukin 4 (IL-4) in response to egg antigen, and could not mediate local delayed-type hypersensitivity. Adoptive transfer experiments using clonal lymphocytes were also undertaken in an attempt to define cell types involved in protective immunity. Clonal lymphocytes secreting both IL-2 and IFN-gamma transferred protective immunity, equivalent to that obtained by non-cultured-sensitized mesenteric lymph node cells. They were effective in very small numbers. However, clonal lymphocytes that secreted IL-4 did not transfer protective immunity. These results suggest that helper T lymphocytes, especially the Th1 subtype, are involved in protective immunity against H. nana.

  9. Model Pembelajaran CTL (Contextual Teaching and Learning dalam Meningkatkan Hasil Belajar Mahasiswa PGSD Pada MataKuliah Konsep IPS Dasar

    Directory of Open Access Journals (Sweden)

    Wahyu Susiloningsih

    2016-03-01

    Full Text Available Hasil belajar adalah suatu proses perubahan tingkah laku dalam pengetahuan, sikap, dan ketrampilan yang diperoleh dalam jangka waktu yang lama.Mata kuliah Konsep IPS Dasar merupakan mata kuliah yang memberikan pemahaman kepada mahasiswa PGSD tentang konsep dasar IPS sebagai landasan kajian yang bahannya bersumber dari kehidupan manusia di masyarakat, yang aspek-aspeknya meliputi social science (ilmu sosial, social studies (studi sosial, Ilmu Pengetahuan Sosial (IPS. IPS merupakan pembelajaran pada tingkat sekolah yang berperan mengfungsionalkan ilmu-ilmu sosial yang bersifat teoritik dalam kehidupan nyata dalam masyarakat. Model pembelajaran CTL adalah model pembelajaran yang menuntut kreatifitas guru dalam mengaitkan subject matter dengan kehidupan nyata mahasiswa guna membantu mahasiswa untuk lebih mudah memaknai materi tersebut.

  10. PEMBELAJARAN PENJUMLAHAN BILANGAN PECAHAN DENGAN METODE CONTEXTUAL TEACHING AND LEARNING (CTL DI SD MUHAMMADIYAH PROGRAM KHUSUS, KOTA BARAT, SURAKARTA

    Directory of Open Access Journals (Sweden)

    Yulia Maftuhah Hidayati

    2015-07-01

    Full Text Available This study is aimed to describe the lesson plan, teaching learning process of sum of fractions based of Contextual Teaching and Learning (CTL, and the motivation of learners attend classes in learning. This study used a qualitative approach. The type of research is a case study. Validation of data is done through triangulation. Data were collected through interviews, observation, documentation, and testing. The technique of data analysis is descriptive, entrepretative. The results of this study indicate that (1 the development of lesson plan has been implemented routinely in every new school year, (2 the process of learning mathematics goes through three stages, namely preinstructional phase (preliminary / initial activity, instructional phase (core activities, and appraisal, (3 during the learning process, the students have a high motivation to participate in activities  because of the method used by teachers is fun and enjoyable

  11. Site-directed mutagenesis of an HLA-A3 gene identifies amino acid 152 as crucial for major-histocompatibility-complex-restricted and alloreactive cytotoxic-T-lymphocyte recognition.

    OpenAIRE

    Cowan, E P; Jelachich, M L; Coligan, J E; Biddison, W E

    1987-01-01

    Major histocompatibility complex-restricted and alloreactive cytotoxic T lymphocytes (CTL) can discriminate between the HLA-A3.1 and HLA-A3.2 antigens. HLA-A3.1 and the rare variant HLA-A3.2 have been shown to differ by two amino acids in the alpha 2 domain at positions 152 (A3.1, glutamic acid; A3.2, valine) and 156 (A3.1, leucine; A3.2, glutamine). To determine the structural basis for the ability of CTL to differentiate A3.1 from A3.2, two site-directed mutants of the HLA-A3.2 gene were pr...

  12. Suppression induction in vivo by a T helper clone?

    DEFF Research Database (Denmark)

    Crispe, I N; Owens, T

    1985-01-01

    We have previously described a helper T cell clone which augments in vivo cytotoxic T cell responses when injected at 10(4) cells per mouse, but not at 10(5) per mouse (Crispe, I. N. et al., Immunology 1984. 52:55). To test whether this dose-response relationship was due to the induction...... of suppression, naive syngeneic mice were injected with 10(5) cloned T helper cells, and their spleen cells were subsequently assayed for suppressive activity in adoptive transfer experiments. Lymphocytes from such mice indeed suppressed an antigen-specific cytotoxic response, but only in the presence...... of the same T helper cell clone freshly added at the time of adoptive transfer. On this basis we argue that the distinction between T helper cell activity and T suppressor-inducer activity corresponds to differences in cell numbers, rather than to two separate cell lineages....

  13. A replicating cytomegalovirus-based vaccine encoding a single Ebola virus nucleoprotein CTL epitope confers protection against Ebola virus.

    Directory of Open Access Journals (Sweden)

    Yoshimi Tsuda

    2011-08-01

    Full Text Available BACKGROUND: Human outbreaks of Ebola virus (EBOV are a serious human health concern in Central Africa. Great apes (gorillas/chimpanzees are an important source of EBOV transmission to humans due to increased hunting of wildlife including the 'bush-meat' trade. Cytomegalovirus (CMV is an highly immunogenic virus that has shown recent utility as a vaccine platform. CMV-based vaccines also have the unique potential to re-infect and disseminate through target populations regardless of prior CMV immunity, which may be ideal for achieving high vaccine coverage in inaccessible populations such as great apes. METHODOLOGY/PRINCIPAL FINDINGS: We hypothesize that a vaccine strategy using CMV-based vectors expressing EBOV antigens may be ideally suited for use in inaccessible wildlife populations. To establish a 'proof-of-concept' for CMV-based vaccines against EBOV, we constructed a mouse CMV (MCMV vector expressing a CD8+ T cell epitope from the nucleoprotein (NP of Zaire ebolavirus (ZEBOV (MCMV/ZEBOV-NP(CTL. MCMV/ZEBOV-NP(CTL induced high levels of long-lasting (>8 months CD8+ T cells against ZEBOV NP in mice. Importantly, all vaccinated animals were protected against lethal ZEBOV challenge. Low levels of anti-ZEBOV antibodies were only sporadically detected in vaccinated animals prior to ZEBOV challenge suggesting a role, at least in part, for T cells in protection. CONCLUSIONS/SIGNIFICANCE: This study demonstrates the ability of a CMV-based vaccine approach to protect against an highly virulent human pathogen, and supports the potential for 'disseminating' CMV-based EBOV vaccines to prevent EBOV transmission in wildlife populations.

  14. Rapid analysis of rearranged kappa light chain genes of circulating polysaccharide-specific B lymphocytes by means of immunomagnetic beads and the polymerase chain reaction

    DEFF Research Database (Denmark)

    Hougs, L; Barington, T; Madsen, HO

    1993-01-01

    of the B lymphocytes activated in vivo. Here, we present a method for rapid analysis of the rearranged kappa light chain genes used by human circulating antigen-specific B lymphocytes. After vaccination with Haemophilus influenzae type b capsular polysaccharide (HibCP) conjugated with protein, the Hib......CP-specific B lymphocytes were isolated by antigen-coated immunomagnetic beads. After the purification, at least 98% of the immunoglobulin-secreting recovered cells were HibCP specific. The RNA was isolated and amplified by cDNA synthesis using a kappa constant region primer followed by polymerase chain...... reaction (PCR) using in addition a degenerate kappa light chain signal peptide region primer. The PCR product was cloned into the M13mp18 phage. The cloning efficiency was 100-600 clones/ml of blood. Of the 86 clones sequenced, 90% represented rearranged kappa light chain genes from different antibody...

  15. Rapid analysis of rearranged kappa light chain genes of circulating polysaccharide-specific B lymphocytes by means of immunomagnetic beads and the polymerase chain reaction

    DEFF Research Database (Denmark)

    Hougs, L; Barington, T; Madsen, HO

    1993-01-01

    reaction (PCR) using in addition a degenerate kappa light chain signal peptide region primer. The PCR product was cloned into the M13mp18 phage. The cloning efficiency was 100-600 clones/ml of blood. Of the 86 clones sequenced, 90% represented rearranged kappa light chain genes from different antibody......CP-specific B lymphocytes were isolated by antigen-coated immunomagnetic beads. After the purification, at least 98% of the immunoglobulin-secreting recovered cells were HibCP specific. The RNA was isolated and amplified by cDNA synthesis using a kappa constant region primer followed by polymerase chain...... of the B lymphocytes activated in vivo. Here, we present a method for rapid analysis of the rearranged kappa light chain genes used by human circulating antigen-specific B lymphocytes. After vaccination with Haemophilus influenzae type b capsular polysaccharide (HibCP) conjugated with protein, the Hib...

  16. The fourth dimension in immunological space: how the struggle for nutrients selects high-affinity lymphocytes.

    Science.gov (United States)

    Wensveen, Felix M; van Gisbergen, Klaas P J M; Eldering, Eric

    2012-09-01

    Lymphocyte activation via the antigen receptor is associated with radical shifts in metabolism and changes in requirements for nutrients and cytokines. Concomitantly, drastic changes occur in the expression of pro-and anti-apoptotic proteins that alter the sensitivity of lymphocytes to limiting concentrations of key survival factors. Antigen affinity is a primary determinant for the capacity of activated lymphocytes to access these vital resources. The shift in metabolic needs and the variable access to key survival factors is used by the immune system to eliminate activated low-affinity cells and to generate an optimal high-affinity response. In this review, we focus on the control of apoptosis regulators in activated lymphocytes by nutrients, cytokines, and costimulation. We propose that the struggle among individual clones that leads to the formation of high-affinity effector cell populations is in effect an 'invisible' fourth signal required for effective immune responses.

  17. Broadly Immunogenic HLA Class I Supertype-Restricted Elite CTL Epitopes Recognized in a Diverse Population Infected with Different HIV-1 Subtypes

    DEFF Research Database (Denmark)

    Pérez, Carina L; Larsen, Mette Voldby; Gustafsson, Rasmus

    2008-01-01

    The genetic variations of the HIV-1 virus and its human host constitute major obstacles for obtaining potent HIV-1-specific CTL responses in individuals of diverse ethnic backgrounds infected with different HIV-1 variants. In this study, we developed and used a novel algorithm to select 184 predi...

  18. Resistance mutations and CTL epitopes in archived HIV-1 DNA of patients on antiviral treatment: toward a new concept of vaccine.

    Directory of Open Access Journals (Sweden)

    Jennifer Papuchon

    Full Text Available Eleven patients responding successfully to first-line antiretroviral therapy (ART were investigated for proviral drug resistance mutations (DRMs in RT by ultra-deep pyrosequencing (UDPS. After molecular typing of the class I alleles A and B, the CTL epitopes in the Gag, Nef and Pol regions of the provirus were sequenced and compared to the reference HXB2 HIV-1 epitopes. They were then matched with the HLA alleles with determination of theoretical affinity (TA. For 3 patients, the results could be compared with an RNA sample of the circulating virus at initiation of therapy. Five out of 11 patients exhibited DRMs by UDPS. The issue is whether a therapeutic switch is relevant in these patients by taking into account the identity of the archived resistance mutations. When the archived CTL epitopes were determined on the basis of the HLA alleles, different patterns were observed. Some epitopes were identical to those reported for the reference with the same TA, while others were mutated with a decrease in TA. In 2 cases, an epitope was observed as a combination of subpopulations at entry and was retrieved as a single population with lower TA at success. With regard to immunological stimulation and given the variability of the archived CTL epitopes, we propose a new concept of curative vaccine based on identification of HIV-1 CTL epitopes after prior sequencing of proviral DNA and matching with HLA class I alleles.

  19. [原著]Prethymic Nylon Wool-Passed Bone Marrow Cells Can Make Distinction between Self and Non-Self X-Chromosome-Linked Gene Products (Xir Antigens) on the Stimulator Cells, Resulting in Regulation of the Generation of Cytotoxic T Lymphocytes in Mixed Lymphocyte Cultures

    OpenAIRE

    Higa, Moritake; Tanabe, MasaoJ; Department of Bacteriology, Faculty of, University of the Ryukyus, Okinawa, Japan; Research Institute of Comprehensive Medicine, School of Medicine, University of the Ryukyus, Okinawa, Japan

    1994-01-01

    We have previously reported that nylon wool-passed bone marrow cells treated with anti-Thy.1 antibody and complement (Thy.1 NW-BM cells) had helper-like activity which could augment the generation of cytotoxic T lymphocytes (CTL). In this study, we determined the antigens to which these NW-BM cells responded and recognized. When a few responder lymph node (LN) cells and an excess of NW-BM responder cells from BIOBR (H-2^k B10 background) mice were cultured with stimulator spleen cells from ei...

  20. 负载HER-2/neu多肽的树突状细胞激发特异性CTL反应%Specific CTL response induced by dendritic cells pulsed with HER-2/neu peptide

    Institute of Scientific and Technical Information of China (English)

    孟东; 时伟锋; 孙春雷; 时宏珍; 史央; 朱晨瑶; 唐金海

    2012-01-01

    目的:探讨以HER-2/neu为靶抗原、树突状细胞(dendritic cell,DC)为抗原载体激发特异性细胞毒性T淋巴细胞(cytotoxic T lymphocytes,CTL)反应的能力及研制治疗型乳腺癌疫苗的可行性.方法:采集17例HLA-A201+ HER-2/neu+乳腺癌患者外周血,分离单个核细胞与外周血淋巴细胞(peripheral blood lymphocyte,PBL),并诱导为成熟DC(mature dendritic cell,mDC);人工合成HER-2/neu多肽[E75 (KIFGSLAFL)和GP2(IISAVVGIL)2条]负载mDC后体外反复致敏PBL(3次,每周1次),检测其激发HER-2/neu特异性CTL的能力与CTL的杀伤活性.同时于患者腹股沟淋巴结富集区皮内注射负载HER-2/neu多肽的DC,每周1次,共接种4次,检测接种前后患者外周血细胞因子和特异性的CTL水平变化,并进行DTH试验.结果:患者外周PBL经过负载HER-2/neu多肽DC共3轮致敏后,HER-2/neu多肽特异的CTL平均比例比对照组(未负载HER-2/neu多肽DC组)明显增高[(5.41±1.44)%vs(0.41±0.12)%,P< 0.05];致敏后PBL对负载HER-2/neu多肽T2靶细胞的杀伤率明显高于对照组(未负载DC诱导的CTL)[效靶比为30∶1时,(35.5±4.7)%vs(11.2±1.4)%,P< 0.05].接种负载HER-2/neu多肽的DC后,患者体内血清中细胞因子IL-2、IL-12、IFN-γ水平较治疗前显著升高[(409.09±89.39)vs(148.79±28.32) ng/ml,(56.23±14.08) vs(24.49±56.23) ng/ml,(146.57±25.97) vs(67.77±39.35) ng/ml;均P<0.05],TNF-α和IL-10水平较治疗前变化不大(P>0.05).患者DTH试验阳性率为47%(8/17),DTH阳性患者外周血中特异性CTL比例明显上升.结论:负载HER-2/neu多肽的DC体内、外均具有激发特异性CTL反应能力,可诱导Th1型细胞因子的分泌,未发生临床不良反应.%Objective; To explore the potential of autologous dendritic cells (DCs) pulsed with HER-2/neu peptide in inducing specific cytotoxic T lymphocyte (CTL) response and feasibility of breast cancer vaccines. Methods; Seventeen breast cancer patients with positive HLA-A201

  1. Whole-genome immunoinformatic analysis of F. tularensis: predicted CTL epitopes clustered in hotspots are prone to elicit a T-cell response.

    Directory of Open Access Journals (Sweden)

    Anat Zvi

    Full Text Available The cellular arm of the immune response plays a central role in the defense against intracellular pathogens, such as F. tularensis. To date, whole genome immunoinformatic analyses were limited either to relatively small genomes (e.g. viral or to preselected subsets of proteins in complex pathogens. Here we present, for the first time, an unbiased bacterial global immunoinformatic screen of the 1740 proteins of F. tularensis subs. holarctica (LVS, aiming at identification of immunogenic peptides eliciting a CTL response. The very large number of predicted MHC class I binders (about 100,000, IC(50 of 1000 nM or less required the design of a strategy for further down selection of CTL candidates. The approach developed focused on mapping clusters rich in overlapping predicted epitopes, and ranking these "hotspot" regions according to the density of putative binding epitopes. Limited by the experimental load, we selected to screen a library of 1240 putative MHC binders derived from 104 top-ranking highly dense clusters. Peptides were tested for their ability to stimulate IFNγ secretion from splenocytes isolated from LVS vaccinated C57BL/6 mice. The majority of the clusters contained one or more CTL responder peptides and altogether 127 novel epitopes were identified, of which 82 are non-redundant. Accordingly, the level of success in identification of positive CTL responders was 17-25 fold higher than that found for a randomly selected library of 500 predicted MHC binders (IC(50 of 500 nM or less. Most proteins (ca. 2/3 harboring the highly dense hotspots are membrane-associated. The approach for enrichment of true positive CTL epitopes described in this study, which allowed for over 50% increase in the dataset of known T-cell epitopes of F. tularensis, could be applied in immunoinformatic analyses of many other complex pathogen genomes.

  2. Laboratorial diagnosis of lymphocytic meningitis

    Directory of Open Access Journals (Sweden)

    Sérgio Monteiro de Almeida

    Full Text Available Meningitis is the main infectious central nervous system (CNS syndrome. Viruses or bacteria can cause acute meningitis of infectious etiology. The term "Aseptic Meningitis" denotes a clinical syndrome with a predominance of lymphocytes in the cerebrospinal fluid (CSF, with no common bacterial agents identified in the CSF. Viral meningitis is considered the main cause of lymphocyte meningitis. There are other etiologies of an infectious nature. CSF examination is essential to establish the diagnosis and to identify the etiological agent of lymphocytic meningitis. We examined CSF characteristics and the differential diagnosis of the main types of meningitis.

  3. Laboratorial diagnosis of lymphocytic meningitis

    Directory of Open Access Journals (Sweden)

    Sérgio Monteiro de Almeida

    2007-10-01

    Full Text Available Meningitis is the main infectious central nervous system (CNS syndrome. Viruses or bacteria can cause acute meningitis of infectious etiology. The term "Aseptic Meningitis" denotes a clinical syndrome with a predominance of lymphocytes in the cerebrospinal fluid (CSF, with no common bacterial agents identified in the CSF. Viral meningitis is considered the main cause of lymphocyte meningitis. There are other etiologies of an infectious nature. CSF examination is essential to establish the diagnosis and to identify the etiological agent of lymphocytic meningitis. We examined CSF characteristics and the differential diagnosis of the main types of meningitis.

  4. Early detection of dominant Env-specific and subdominant Gag-specific CD8+ lymphocytes in equine infectious anemia virus-infected horses using major histocompatibility complex class I/peptide tetrameric complexes.

    Science.gov (United States)

    Mealey, Robert H; Sharif, Amin; Ellis, Shirley A; Littke, Matt H; Leib, Steven R; McGuire, Travis C

    2005-08-15

    Cytotoxic T lymphocytes (CTL) are critical for control of lentiviruses, including equine infectious anemia virus (EIAV). Measurement of equine CTL responses has relied on chromium-release assays, which do not allow accurate quantitation. Recently, the equine MHC class I molecule 7-6, associated with the ELA-A1 haplotype, was shown to present both the Gag-GW12 and Env-RW12 EIAV CTL epitopes. In this study, 7-6/Gag-GW12 and 7-6/Env-RW12 MHC class I/peptide tetrameric complexes were constructed and used to analyze Gag-GW12- and Env-RW12-specific CTL responses in two EIAV-infected horses (A2164 and A2171). Gag-GW12 and Env-RW12 tetramer-positive CD8+ cells were identified in nonstimulated peripheral blood mononuclear cells as early as 14 days post-EIAV inoculation, and frequencies of tetramer-positive cells ranged from 0.4% to 6.7% of nonstimulated peripheral blood CD8+ cells during the 127-day study period. Although both horses terminated the initial viremic peak, only horse A2171 effectively controlled viral load. Neutralizing antibody was present during the initial control of viral load in both horses, but the ability to maintain control correlated with Gag-GW12-specific CD8+ cells in A2171. Despite Env-RW12 dominance, Env-RW12 escape viral variants were identified in both horses and there was no correlation between Env-RW12-specific CD8+ cells and control of viral load. Although Gag-GW12 CTL escape did not occur, a Gag-GW12 epitope variant arose in A2164 that was recognized less efficiently than the original epitope. These data indicate that tetramers are useful for identification and quantitation of CTL responses in horses, and suggest that the observed control of EIAV replication and clinical disease was associated with sustained CTL recognition of Gag-specific epitopes.

  5. BURDEN OF ABNORMAL HEMATOPOIETIC CLONE IN PATIENTS WITH MYELODYSPLASTIC SYNDROMES

    Institute of Scientific and Technical Information of China (English)

    2006-01-01

    Objective To investigate the role of the burden of abnormal hematopoietic clone in the development of myelodys plastic syndromes (MDS).Methods The ratio of the bone marrow cells with abnormal chromosomes to the total counted bone marrow cells was regarded as the index of MDS clone burden. The disease severity related parameters including white blood cell count, hemoglobin, platelet count, lactate dehydrogenase level, bone marrow blast, myeloid differentiation index, micromegakaryocyte, transfusion, interleukin-2, tumor necrosis factor ( TNF), CD4 + and CD8 + T cells of MDS patients were assayed, and the correlations between those parameters and MDS clone burden were also analyzed.Results The clone burden of MDS patients was 67.4% ± 36. 2%. MDS clone burden positively correlated with bone marrow blasts (r=0.483, P<0.05), negatively with hemoglobin level (r=-0.445, P<0.05). The number of blasts, hemoglobin, and erythrocytes in high clone burden (>50%) and low clone burden (≤50%) groups were 7.78%±5.51% and 3.45%±3.34%, 56.06±14. 28 g/L and 76.40±24.44 g/L, (1.82±0.48)×1012/L and (2. 32±0.66)×1012/L, respectively (all P <0.05). CD4 + T lymphocytes of MDS patients and normal controls were (0. 274±0.719)×109/L and (0.455±0.206)×109/L, respectively (P<0.05). CD8 ± T lymphocytes of MDS patients and normal controls were (0.240±0.150)×109/L and (0.305 ±0.145)×109/L, respectively. The serum level of interleukin-2 of MDS patients (6.29±3.58 ng/mL) was significantly higher than normal control (3.11±1.40ng/mL, P<0.05). The serum level of TNF of MDS patients and normal control group were 2.42±1.79 ng/mL and 1.68 ±0.69 ng/mL, respectively. The ratio of CD4 to CD8 was higher in high clone burden MDS patients (1.90 ±0.52) than that in low clone burden patients (0.97±0.44, P<0.05).Conclusion The quantitive clonal karyotype abnormalities and deficient T cell immunity are important parameters for evaluating MDS severity and predicting its

  6. Identification of Melanoma-specific Peptide Epitopes by HLA-A2.1-restricted Cytotoxic T Lymphocytes

    Institute of Scientific and Technical Information of China (English)

    Hai-Liang GE; Ying WANG; Shu-Jun WANG; Yong ZHANG

    2006-01-01

    HLA-A2.1-associated peptides, extracted from human melanoma cells, were used to study epitopes for melanoma-specific HLA-A2.1-restricted cytotoxic T lymphocytes (CTLs) by epitope reconstitution, active peptide sequence characterization and synthetic peptide verification. CTL were generated from tumor-involved nodes by in vitro stimulation, initially with autologous melanoma cells and subsequently with allogeneic HLA-A2.1 positive melanoma cells. The CTLs could lyse autologous and allogeneic HLA-A2.1 positive melanomas, but not HLA-A2.1 negative melanomas or HLA-A2.1 positive non-melanomas. The lysis of melanomas could be inhibited by anti-CD3, anti-HLA class Ⅰ and anti-HLA-A2.1 monoclonal antibodies.HLA-A2.1 molecules were purified from detergent-solubilized human melanoma cells by immunoaffinity column chromatography and further fractionated by reversed phase high performance liquid chromatography.The fractions were assessed for their ability to reconstitute melanoma-specific epitopes with HLA-A2.1 positive antigen-processing mutant T2 cells. Three reconstitution peaks were observed in lactate dehydrogenase release assay. Mass spectrometry and ion-exchange high performance liquid chromatography analysis were used to identify peptide epitopes. Peptides with a mass-to-charge ratio of 948 usually consist of nine amino acid residues. The data from reconstitution experiments confirmed that the synthetic peptides contained epitopes and that the peptides associated with HLA-A2.1 and recognized by melanoma-specific CTL were present in these different melanoma cells. These peptides could be potentially exploited in novel peptide-based antitumor vaccines in immunotherapy for CTL.

  7. Polymer nanoparticles for cross-presentation of exogenous antigens and enhanced cytotoxic T-lymphocyte immune response

    Directory of Open Access Journals (Sweden)

    Song C

    2016-08-01

    Full Text Available Chanyoung Song,* Young-Woock Noh,* Yong Taik Lim SKKU Advanced Institute of Nanotechnology (SAINT, School of Chemical Engineering, Sungkyunkwan University, Suwon, South Korea *These authors contributed equally to this work Abstract: Effective induction of an antigen-specific cytotoxic T lymphocyte (CTL immune response is one of the key goals of cancer immunotherapy. We report the design and fabrication of polyethylenimine (PEI-coated polymer nanoparticles (NPs as efficient antigen-delivery carriers that can induce antigen cross-presentation and a strong CTL response. After synthesis of poly(d,l-lactide-co-glycolide (PLGA NPs containing ovalbumin (OVA by the double-emulsion solvent-evaporation method, cationic-charged PLGA NPs were generated by coating them with PEI. In a methyl tetrazolium salt assay, no discernible cytotoxic effect of PEI-coated PLGA (OVA NPs was observed. The capacity and mechanism of PEI-coated PLGA (OVA NPs for antigen delivery and cross-presentation on dendritic cells (DCs were determined by fluorescence microscopy and flow cytometry. PEI-coated PLGA (OVA NPs were internalized efficiently via phagocytosis or macropinocytosis in DCs and induced efficient cross-presentation of the antigen on MHC class I molecules via both endosome escape and a lysosomal processing mechanism. The DCs treated with PEI-coated PLGA (OVA NPs induced a release of IL-2 cytokine from OVA-specific CD8-OVA1.3 T cells more efficiently than DCs treated with PLGA (OVA NPs. Therefore, the PEI-coated PLGA (OVA NPs can induce antigen cross-presentation and are expected to be used for induction of a strong CTL immune response and for efficient anticancer immunotherapy. Keywords: antigen delivery, dendritic cells, polymer NPs, vaccine, cross-presentation

  8. HLA-A2–Restricted Cytotoxic T Lymphocyte Epitopes from Human Heparanase as Novel Targets for Broad-Spectrum Tumor Immunotherapy

    Directory of Open Access Journals (Sweden)

    Ting Chen

    2008-09-01

    Full Text Available Peptide vaccination for cancer immunotherapy requires identification of peptide epitopes derived from antigenic proteins associated with tumors. Heparanase (Hpa is broadly expressed in various advanced tumors and seems to be an attractive new tumor-associated antigen. The present study was designed to predict and identify HLA-A2– restricted cytotoxic T lymphocyte (CTL epitopes in the protein of human Hpa. For this purpose, HLA-A2–restricted CTL epitopes were identified using the following four-step procedure: 1 a computer-based epitope prediction from the amino acid sequence of human Hpa, 2 a peptide-binding assay to determine the affinity of the predicted protein with the HLA-A2 molecule, 3 stimulation of the primary T-cell response against the predicted peptides in vitro, and 4 testing of the induced CTLs toward different kinds of carcinoma cells expressing Hpa antigens and/or HLA-A2. The results demonstrated that, of the tested peptides, effectors induced by peptides of human Hpa containing residues 525-533 (PAFSYSFFV, Hpa525, 277-285 (KMLKSFLKA, Hpa277, and 405-413 (WLSLLFKKL, Hpa405 could effectively lyse various tumor cell lines that were Hpa-positive and HLA-A2-matched. We also found that these peptide-specific CTLs could not lyse autologous lymphocytes with low Hpa activity. Further study revealed that Hpa525, Hpa277, and Hpa405 peptides increased the frequency of IFN-γ–producing T cells compared to a negative peptide. Our results suggest that Hpa525, Hpa277, and Hpa405 peptides are new HLA-A2–restricted CTL epitopes capable of inducing Hpa-specific CTLs in vitro. Because Hpa is expressed in most advanced malignant tumors, Hpa525, Hpa277, and Hpa405 peptide–based vaccines may be useful for the immunotherapy for patients with advanced tumors.

  9. Improved cytotoxic T-lymphocyte immune responses to a tumor antigen by vaccines co-expressing the SLAM-associated adaptor EAT-2.

    Science.gov (United States)

    Aldhamen, Y A; Seregin, S S; Kousa, Y A; Rastall, D P W; Appledorn, D M; Godbehere, S; Schutte, B C; Amalfitano, A

    2013-10-01

    The signaling lymphocytic activation molecule-associated adaptor Ewing's sarcoma's-activated transcript 2 (EAT-2) is primarily expressed in dendritic cells, macrophages and natural killer cells. Including EAT-2 in a vaccination regimen enhanced innate and adaptive immune responses toward pathogen-derived antigens, even in the face of pre-existing vaccine immunity. Herein, we investigate whether co-vaccinations with two recombinant Ad5 (rAd5) vectors, one expressing the carcinoembryonic antigen (CEA) and one expressing EAT-2, can induce more potent CEA-specific cytotoxic T lymphocyte (CTL) and antitumor activity in the therapeutic CEA-expressing MC-38 tumor model. Our results suggest that inclusion of EAT-2 significantly alters the kinetics of Th1-biasing proinflammatory cytokine and chemokine responses, and enhances anti-CEA-specific CTL responses. As a result, rAd5-EAT2-augmented rAd5-CEA vaccinations are more efficient in eliminating CEA-expressing target cells as measured by an in vivo CTL assay. Administration of rAd5-EAT2 vaccines also reduced the rate of growth of MC-38 tumor growth in vivo. Also, an increase in MC-38 tumor cell apoptosis (as measured by hematoxylin and eosin staining, active caspase-3 and granzyme B levels within the tumors) was observed. These data provide evidence that more efficient, CEA-specific effector T cells are generated by rAd5 vaccines expressing CEA, when augmented by rAd5 vaccines expressing EAT-2, and this regimen may be a promising approach for cancer immunotherapy in general.

  10. Initiation of lymphocyte DNA synthesis.

    Science.gov (United States)

    Coffman, F D; Fresa, K L; Cohen, S

    1991-01-01

    The initiation of DNA replication in T lymphocytes appears to be regulated by two distinct activities: one associated with proliferation which mediates initiation, and another associated with quiescence which blocks initiation. Activated lymphocytes and proliferating lymphoid cell lines produce an activity, termed ADR, which can initiate DNA replication in isolated, quiescent nuclei. ADR is heat-labile, has protease activity or interacts closely with a protease, and is distinct from the DNA polymerases. ADR activity is absent in quiescent lymphocytes and appears in mitogen-stimulated lymphocytes after IL-2 binding. The generation of active ADR appears to be mediated by phosphorylation of a precursor which is present in resting cells. Nuclei from mitogen-unresponsive lymphocytes fail to initiate DNA replication in response to ADR, of potential importance in the age-related decline of immunity. Quiescent lymphocytes lack ADR and synthesize an ADR-inhibitory activity. The ADR inhibitor is a heat-stable protein which suppresses the initiation of DNA synthesis, but is ineffective at suppressing elongation once DNA strand replication has begun. Nuclei from several neoplastic cell lines fail to respond to the ADR inhibitor, which may play a role in the continuous proliferation of these cells. At least one of these neoplastic cell lines produces both ADR and an inhibitory factor. These findings suggest that the regulation of proliferation is dependent on the balance between activating and inhibitory pathways.

  11. Placentation in cloned cattle

    DEFF Research Database (Denmark)

    Miglino, M A; Pereira, F T V; Visintin, J A

    2007-01-01

    To elucidate the morphological differences between placentas from normal and cloned cattle pregnancies reaching term, the umbilical cord, placentomes and interplacentomal region of the fetal membranes were examined macroscopically as well as by light and scanning electron microscopy. In pregnancies...... than one primary villus, as opposed to a single villus in non-cloned placentae. Scanning electron microscopy of blood vessel casts revealed that there was also more than one stem artery per villous tree and that the ramification of the vessels failed to form dense complexes of capillary loops...

  12. In vitro co-stimulation with anti-CD28 synergizes with IL-12 in the generation of T cell immune responses to leukaemic cells; a strategy for ex-vivo generation of CTL for immunotherapy.

    Science.gov (United States)

    Orleans-Lindsay, J K; Deru, A; Craig, J I O; Prentice, H G; Lowdell, M W

    2003-09-01

    The existence of an immune based graft-versus-leukaemia (GvL) effect highlighted the prospect of managing relapsed leukaemias with T cell-based adoptive immunotherapy. Thus, various strategies have been explored for the in vitro expansion of acute myeloid leukaemia (AML)-specific T cells. In a popular approach, AML blasts have been genetically modified to express co-stimulatory molecules essential for effective T cell priming. One such tactic has been the modification of AML cells to express the B7/CD80 co-stimulatory molecule that binds to CD28 on T cells initiating events that culminate in enhanced cytokine production, proliferation and development of effector functions by T cells. The success of these strategies has been limited by difficulties in attaining sufficient transduction efficiencies and associated high levels of CD80 expression. We demonstrate that these problems can be circumvented by using anti-CD28 monoclonal antibody. Furthermore, we show that the synergistic relationship between CD80/CD28 pathway and interleukin 12 cytokine (IL-12), documented in the generation of cytotoxic T lymphocytes (CTL) for solid tumours, also applies to AML. CD28/IL-12 synergy facilitated the proliferation of allogeneic T cells in response to stimulation with primary AML blasts. The synergy also favoured generation of a Th1-type immune response, evidenced by gamma interferon (IFN-gamma) secretion and facilitated naive and memory T cell proliferation. Unlike some methods of in vitro T cell expansion, use of CD28/IL-12 synergy left T cells in the physiologically appropriate CD45RA-/CCR7- subsets known to be associated with immediate cytotoxic functions.

  13. Why clone flies? Using cloned Drosophila to monitor epigenetic defects.

    Science.gov (United States)

    Haigh, Andrew J; Lloyd, Vett K

    2007-01-01

    Since the birth of the first cloned sheep in 1996, advances in nuclear transplantation have led to both the creation of genetically tailored stem cells and the generation of a number of cloned organisms. The list of cloned animals reared to adulthood currently includes the frog, sheep, mouse, cow, goat, pig, rabbit, cat, zebrafish, mule, horse, rat and dog. The addition of Drosophila to this elite bestiary of cloned animals has prompted the question - why clone flies? Organisms generated by nuclear transplantation suffer from a high rate of associated defects, and many of these defects appear to be related to aberrant genomic imprinting. Imprinted gene expression also appears to be compromised in Drosophila clones. Proper imprinted gene regulation relies on a suite of highly conserved chromatin-modifying genes first identified in Drosophila. Thus, Drosophila can potentially be used to study epigenetic dysfunction in cloned animals and to screen for genetic and epigenetic conditions that promote the production of healthy clones.

  14. A Single Amino Acid Difference within the α-2 Domain of Two Naturally Occurring Equine MHC Class I Molecules Alters the Recognition of Gag and Rev Epitopes by Equine Infectious Anemia Virus-Specific CTL1

    Science.gov (United States)

    Mealey, Robert H.; Lee, Jae-Hyung; Leib, Steven R.; Littke, Matt H.; McGuire, Travis C.

    2012-01-01

    Although CTL are critical for control of lentiviruses, including equine infectious anemia virus, relatively little is known regarding the MHC class I molecules that present important epitopes to equine infectious anemia virus-specific CTL. The equine class I molecule 7-6 is associated with the equine leukocyte Ag (ELA)-A1 haplotype and presents the Env-RW12 and Gag-GW12 CTL epitopes. Some ELA-A1 target cells present both epitopes, whereas others are not recognized by Gag-GW12-specific CTL, suggesting that the ELA-A1 haplotype comprises functionally distinct alleles. The Rev-QW11 CTL epitope is also ELA-A1-restricted, but the molecule that presents Rev-QW11 is unknown. To determine whether functionally distinct class I molecules present ELA-A1-restricted CTL epitopes, we sequenced and expressed MHC class I genes from three ELA-A1 horses. Two horses had the 7-6 allele, which when expressed, presented Env-RW12, Gag-GW12, and Rev-QW11 to CTL. The other horse had a distinct allele, designated 141, encoding a molecule that differed from 7-6 by a single amino acid within the α-2 domain. This substitution did not affect recognition of Env-RW12, but resulted in more efficient recognition of Rev-QW11. Significantly, CTL recognition of Gag-GW12 was abrogated, despite Gag-GW12 binding to 141. Molecular modeling suggested that conformational changes in the 141/Gag-GW12 complex led to a loss of TCR recognition. These results confirmed that the ELA-A1 haplotype is comprised of functionally distinct alleles, and demonstrated for the first time that naturally occurring MHC class I molecules that vary by only a single amino acid can result in significantly different patterns of epitope recognition by lentivirus-specific CTL. PMID:17082657

  15. The immune system as a self-centered network of lymphocytes.

    Science.gov (United States)

    Santori, Fabio R

    2015-08-01

    This essay makes a brief historical and comparative review of selective and network theories of the immune system which is presented as a chemical sensory system with immune and non-immune functions. The ontogeny of immune networks is the result of both positive and negative selection of lymphocytes to self-epitopes that serve as a "template" for the recognition of foreign antigens. The development of immune networks progresses from single individual clones in early ontogeny into complex "information processing networks" in which lymphocytes are linked to inhibitory and stimulatory immune cells. The results of these regulatory interactions modulate immune responses and tolerance.

  16. Measuring melanoma-specific cytotoxic T lymphocytes elicited by dendritic cell vaccines with a tumor inhibition assay in vitro.

    Science.gov (United States)

    Paczesny, Sophie; Shi, Honhgzhen; Saito, Hiroaki; Mannoni, Patrice; Fay, Joseph; Banchereau, Jacques; Palucka, A Karolina

    2005-01-01

    Improving cancer vaccines depends on assays measuring elicited tumor-specific T-cell immunity. Cytotoxic effector cells are essential for tumor clearance and are commonly evaluated using 51Cr release from labeled target cells after a short (4 hours) incubation with T cells. The authors used a tumor inhibition assay (TIA) that assesses the capacity of cytotoxic T lymphocytes (CTLs) to control the survival/growth of EGFP-labeled tumor cell lines. TIA was validated using CD8+ T cells primed in vitro against melanoma and breast cancer cells. TIA was then used to assess the CTL function of cultured CD8+ T cells isolated from patients with metastatic melanoma who underwent vaccination with peptide-pulsed CD34+ HPCs-derived DCs. After the DC vaccination, T cells from six of eight patients yielded CTLs that could inhibit the survival/growth of melanoma cells. The results of TIA correlated with killing of tumor cells in a standard 4-hour 51Cr release assay, yet TIA allowed detection of CTL activities that appeared marginal in the 51Cr release assay. Thus, TIA might prove valuable for measuring spontaneous and induced antigen-specific cytotoxic T cells.

  17. Capacity of toxitoxic T lymphocytes to control the reproduction of human immunodeficiency virus

    NARCIS (Netherlands)

    C.A. van Baalen (Carel)

    2002-01-01

    textabstractThe focus of the work presented in this thesis is on CTL directed against HIV-1. Chapter 2 addresses frequencies of circulating CTL, their specificity at the protein and epitope level, and associations of CTL responses with rapid or slow disease progression in HIV -1 infection. Studies o

  18. Clip, connect, clone

    DEFF Research Database (Denmark)

    Fujima, Jun; Lunzer, Aran; Hornbæk, Kasper

    2010-01-01

    using three mechanisms: clipping of input and result elements from existing applications to form cells on a spreadsheet; connecting these cells using formulas, thus enabling result transfer between applications; and cloning cells so that multiple requests can be handled side by side. We demonstrate...

  19. The Cloning of America.

    Science.gov (United States)

    Dobson, Judith E.; Dobson, Russell L.

    1981-01-01

    Proposes that the U.S. school system purports to prize human variability, but many educators are engaged in activities that seek to homogenize students. Describes these activities, including diagnosis, labeling, ability grouping, and positive reinforcement. Presents suggestions for counselors to combat sources of cloning and self-validation. (RC)

  20. Asian Yellow Goat Cloned

    Institute of Scientific and Technical Information of China (English)

    2005-01-01

    @@ It was released on August 24,2005 by Prof. CHEN Dayuan (Da-Yuan Chen) from the CAS Institute of Zoology that the first success in cloning the Asian Yellow Goat by nuclear transfer had recently been achieved in east China's Shandong Province.

  1. Sequential cloning of chromosomes

    Energy Technology Data Exchange (ETDEWEB)

    Lacks, S.A.

    1991-12-31

    A method for sequential cloning of chromosomal DNA and chromosomal DNA cloned by this method are disclosed. The method includes the selection of a target organism having a segment of chromosomal DNA to be sequentially cloned. A first DNA segment, having a first restriction enzyme site on either side. homologous to the chromosomal DNA to be sequentially cloned is isolated. A first vector product is formed by ligating the homologous segment into a suitably designed vector. The first vector product is circularly integrated into the target organism`s chromosomal DNA. The resulting integrated chromosomal DNA segment includes the homologous DNA segment at either end of the integrated vector segment. The integrated chromosomal DNA is cleaved with a second restriction enzyme and ligated to form a vector-containing plasmid, which is replicated in a host organism. The replicated plasmid is then cleaved with the first restriction enzyme. Next, a DNA segment containing the vector and a segment of DNA homologous to a distal portion of the previously isolated DNA segment is isolated. This segment is then ligated to form a plasmid which is replicated within a suitable host. This plasmid is then circularly integrated into the target chromosomal DNA. The chromosomal DNA containing the circularly integrated vector is treated with a third, retrorestriction enzyme. The cleaved DNA is ligated to give a plasmid that is used to transform a host permissive for replication of its vector. The sequential cloning process continues by repeated cycles of circular integration and excision. The excision is carried out alternately with the second and third enzymes.

  2. [Nuclear transfer and therapeutic cloning].

    Science.gov (United States)

    Xu, Xiao-Ming; Lei, An-Min; Hua, Jin-Lian; Dou, Zhong-Ying

    2005-03-01

    Nuclear transfer and therapeutic cloning have widespread and attractive prospects in animal agriculture and biomedical applications. We reviewed that the quality of oocytes and nuclear reprogramming of somatic donor cells were the main reasons of the common abnormalities in cloned animals and the low efficiency of cloning and showed the problems and outlets in therapeutic cloning, such as some basic problems in nuclear transfer affected clinical applications of therapeutic cloning. Study on isolation and culture of nuclear transfer embryonic stem (ntES) cells and specific differentiation of ntES cells into important functional cells should be emphasized and could enhance the efficiency. Adult stem cells could help to cure some great diseases, but could not replace therapeutic cloning. Ethics also impeded the development of therapeutic cloning. It is necessary to improve many techniques and reinforce the research of some basic theories, then somatic nuclear transfer and therapeutic cloning may apply to agriculture reproduction and benefit to human life better.

  3. The First Human Cloned Embryo.

    Science.gov (United States)

    Cibelli, Jose B.; Lanza, Robert P.; West, Michael D.; Ezzell, Carol

    2002-01-01

    Describes a process known as parthenogenesis which produces cloned, early-stage embryos and human embryos generated only from eggs. Speculates that this technology puts therapeutic cloning within reach. (DDR)

  4. Animal Cloning and Food Safety

    Science.gov (United States)

    ... For Consumers Home For Consumers Consumer Updates Animal Cloning and Food Safety Share Tweet Linkedin Pin it ... evaluate the issue. back to top FDA Studies Cloning For more than five years, CVM scientists studied ...

  5. Ex vivo expansion of tumor-infiltrating lymphocytes from nasopharyngeal carcinoma patients for adoptive immunotherapy

    Institute of Scientific and Technical Information of China (English)

    Jia He; Xiao-Feng Tang; Qiu-Yan Chen; Hai-Qiang Mai; Zhou-Feng Huang; Jiang Li; Yi-Xin Zeng

    2012-01-01

    Establishing Epstein-Barr virus (EBV)-specific cytolytic T lymphocytes (EBV-CTLs) from peripheral blood mononuclear cells (PBMCs) for adoptive immunotherapy has been reported in EBV-associated malignancies including Hodgkin's lymphoma and nasopharyngeal carcinoma (NPC).In the current study,we performed ex vivo expansion of tumor-infiltrating lymphocytes (TILs) obtained from NPC biopsy specimens with a rapid expansion protocol using anti-CD3 monoclonal antibody (OKT3),recombinanthuman interleukin (IL)-2,and irradiated PBMCs from healthy donors to initiate the growth of TILs.Young TIL cultures comprised of more than 90% of CD3+ T cells,a variable percentage of CD3+CD8+ and CD3+CD4+ T cells,and less than 10% of CD3-CD16+ natural killer cells,a similar phenotype of EBV-CTL cultures from PBMCs.Interestingly,TIL cultures secreted high levels of the Th1 cytokines,interferon gamma (IFNγ) and tumor necrosis factor-alpha (TNF-α),and low levels of the Th2 cytokines,IL-4 and IL10.Moreover,young TILs could recognize autologous EBV-transformed B lymphoblast cell lines,but not autologous EBV-negative blast cells or allogeneic EBV-negative tumor cells.Taken together,these data suggest that ex vivo expansion of TILs from NPC biopsy tissue is an appealing alternative method to establish T cell-based immunotherapy for NPC.

  6. Engineered exosomes boost the HCV NS3-specific CD8+ T lymphocyte immunity in humans

    Directory of Open Access Journals (Sweden)

    Simona Anticoli

    2016-01-01

    Full Text Available At the present, no anti-Hepatitis C virus (HCV HCV vaccine is available, and many patients failed the treatment with new class of HCV inhibitors. In HCV infection, both experimental and clinic evidences indicate that a strong CTL-immune response could have significant therapeutic effects. We developed an innovative anti-HCV CD8+ T immunogen based on the uploading in engineered exosomes of full-length HCV-NS3 protein. HCV NS3 exosomes appeared immunogenic when injected in mice, as proven by the detection of a memory CD8+ T lymphocyte pool two weeks after the last of three immunizations. On the other hand, dendritic cells isolated from PBMCs of HCV infected patients activate autologous HCV NS3-specific CD8+ T lymphocytes upon challenge with HCV NS3 exosomes. These results provide the proof-of-principle that engineered exosomes can boost the CD8+ T cell immunity in HCV-infected patients, thus representing a suitable option for patients resisting the therapies with recently discovered HCV inhibitors.

  7. Probabilistic Cloning and Quantum Computation

    Institute of Scientific and Technical Information of China (English)

    GAO Ting; YAN Feng-Li; WANG Zhi-Xi

    2004-01-01

    @@ We discuss the usefulness of quantum cloning and present examples of quantum computation tasks for which the cloning offers an advantage which cannot be matched by any approach that does not resort to quantum cloning.In these quantum computations, we need to distribute quantum information contained in the states about which we have some partial information. To perform quantum computations, we use a state-dependent probabilistic quantum cloning procedure to distribute quantum information in the middle of a quantum computation.

  8. 乙型肝炎病毒包膜区和核心区CTL抗原表位变异对感染慢性化及疾病转归的影响%Impact of CTL epitope mutations of hepatitis B virus envelope protein and core protein on chronicity and disease prognosis in patients with hepatitis B

    Institute of Scientific and Technical Information of China (English)

    哈明昊; 佘会元; 吴建秋; 孙莉萍; 沈文娟; 黄钟鸣; 陈晓兰; 单文艳; 何慧芳

    2015-01-01

    Objective To investigate the impact of cytotoxic T lymphocyte (CTL) epitope mutation of HBV envelope and core protein on disease progress in patients with hepatitis B. Methods 38 individuals with asymptomatic HBsAg carrier,35 patients with chronic hepatitis B and 23 with liver cirrhosis were recruited in this study. Gene sequencing analysis of HBV core region and envelope region was carried out in all enrolled patients to unveil any possible CTL epitope mutation. Results The mutation rates of locus 41~49,88~96,97~108,172~180 and 185~194 of CTL epitope in HBV envelope region were 48(55.2%),13(14.9%),11(12.6%),9(10.3%) and 6 (6.9%),respectively;60% of mutation at locus 41~49 in patients with chronic hepatitis B and 65.2% in with liver cirrhosis were significantly higher than 31.6% in asymptomatic HBsAg carriers(P<0.05);the mutation rates at 18~27 and 91~95 locus of CTL epitope in HBV core region were 56.2% and 43.8%,respectively;the mutation rates at locus 18~27 were also higher in patients with CHB (20.0%) and LC (30.4%) than that in asymptomatic HBsAg carriers (10.5%,P<0.05);Multivariate analysis showed that locus 41~49 mutation was an independent risk factor for the progress of CHB to LC. Conclusion CTL epitope mutation of HBV is a significant cause for the chronicity of hepatitis B virus infection.%目的:探讨HBV包膜区和核心区细胞毒性T淋巴细胞(CTL)抗原表位变异的规律及对疾病发展转归的影响。方法本研究选择无症状HBsAg携带者(ASC)38例,慢性乙型肝炎(CHB)35例,乙型肝炎肝硬化(LC)23例。采用DNA测序法检测HBV包膜区和核心区病毒序列及表位变异情况,并分析其与临床转归的关系。结果在入组患者中,HBV包膜区CTL抗原表位41~49、88~96、97~108、172~180和185~194变异率分别为48个(55.2%)、13个(14.9%)、11个(12.6%)、9个(10.3%)和6个(6.9%);HBV CTL 41~49位变异率在CHB和肝硬化患者中的变异率分别为60.0%

  9. THE PROFILE OF STUDENT ACTIVITIES IN LEARNING BASIC NATURAL SCIENCE CONCEPTS THROUGH THE CONTEXTUAL TEACHING AND LEARNING (CTL APPROACH WITH GROUP INVESTIGATION (GI MODEL

    Directory of Open Access Journals (Sweden)

    Muhammad Minan Chusni

    2017-05-01

    Full Text Available Science learning essentially requires students to cultivate curiosity so that it triggers to conduct investigations by doing science activities. The purpose of this research is to know the profile of student activity in learning basic concept of IPA through contextual teaching and learning approach (CTL with group investigation (GI model.The subject of this research is Program of primary teacher education UMMGL students consisting of two classes with 83 people. The research method is descriptive. Data collection techniques were conducted by setting the focus of research, selecting informants as data sources, collecting data, assessing data quality, analyzing data, interpreting data, and drawing conclusions on the findings.From the research results can be concluded that the profile of student activity in learning basic concept of science through CTL approach with the average GI model is in very good category.

  10. Comparison of CTL reactivity in the spleen and draining lymph nodes after immunization with peptides pulsed on dendritic cells or mixed with Freund's incomplete adjuvant

    DEFF Research Database (Denmark)

    Wang, Ming-Jun; Nissen, Mogens Holst; Buus, Søren

    2003-01-01

    OBJECTIVE: To compare CTL reactivity in the spleen and the draining lymph nodes (LN) from C57BL/6 mice after immunization with self and non-self peptides pulsed on autologous dendritic cells (DC) or mixed with Freund's incomplete adjuvant (FIA). METHODS: Peptides showing high to low binding...... in the draining LN, whereas non-self peptides mixed with FIA generated the strongest response in the spleen. CONCLUSIONS: DC-based immunization with non-self and self peptides is more efficient than immunization based on peptides mixed with FIA. DC-based immunization focuses the CTL response towards the spleen....... Immunization based on FIA focuses the response against self peptides towards the draining LN and non-self peptides towards the spleen....

  11. HTLV-1 Tax-Specific CTL Epitope-Pulsed Dendritic Cell Therapy Reduces Proviral Load in Infected Rats with Immune Tolerance against Tax.

    Science.gov (United States)

    Ando, Satomi; Hasegawa, Atsuhiko; Murakami, Yuji; Zeng, Na; Takatsuka, Natsuko; Maeda, Yasuhiro; Masuda, Takao; Suehiro, Youko; Kannagi, Mari

    2017-02-01

    Adult T cell leukemia/lymphoma (ATL), a CD4(+) T cell malignancy with a poor prognosis, is caused by human T cell leukemia virus type 1 (HTLV-1) infection. High proviral load (PVL) is a risk factor for the progression to ATL. We previously reported that some asymptomatic carriers had severely reduced functions of CTLs against HTLV-1 Tax, the major target Ag. Furthermore, the CTL responses tended to be inversely correlated with PVL, suggesting that weak HTLV-1-specific CTL responses may be involved in the elevation of PVL. Our previous animal studies indicated that oral HTLV-1 infection, the major route of infection, caused persistent infection with higher PVL in rats compared with other routes. In this study, we found that Tax-specific CD8(+) T cells were present, but not functional, in orally infected rats as observed in some human asymptomatic carriers. Even in the infected rats with immune unresponsiveness against Tax, Tax-specific CTL epitope-pulsed dendritic cell (DC) therapy reduced the PVL and induced Tax-specific CD8(+) T cells capable of proliferating and producing IFN-γ. Furthermore, we found that monocyte-derived DCs from most infected individuals still had the capacity to stimulate CMV-specific autologous CTLs in vitro, indicating that DC therapy may be applicable to most infected individuals. These data suggest that peptide-pulsed DC immunotherapy will be useful to induce functional HTLV-1-specific CTLs and decrease PVL in infected individuals with high PVL and impaired HTLV-1-specific CTL responses, thereby reducing the risk of the development of ATL. Copyright © 2017 by The American Association of Immunologists, Inc.

  12. Limited T-cell receptor diversity in liver-infiltrating lymphocytes from patients with primary biliary cirrhosis.

    Science.gov (United States)

    Diu, A; Moebius, U; Ferradini, L; Genevée, C; Roman-Roman, S; Claudon, M; Delorme, D; Meuer, S; Hercend, T; Praz, F

    1993-10-01

    Primary biliary cirrhosis is associated with the presence of high-titer anti-mitochondrial autoantibodies as well as T-cell infiltration of the liver, suggesting the involvement of autoimmune mechanisms. We have studied here the sequences of T-cell receptor alpha and beta chains expressed by T-cell clones derived from liver-infiltrating lymphocytes of two patients with primary biliary cirrhosis. Among the eight clones studied from the first patient, four expressed the same member of the V beta 6 subfamily, associated with either V alpha 4 (three clones) or V alpha 21 (one clone) gene segment. Two other clones expressed an identical V beta 12 transcript, and two in-frame alpha chain transcripts, involving V alpha 2 and V alpha 7 gene segments. From the second patient, eight out of the nine clones were found to rearrange V beta 17-J beta 2.1 and V alpha 3 gene segments. The remaining clone expressed distinct T-cell receptor chains, involving V beta 9 and V alpha 11 gene segments. As deduced from the analysis of their junctional regions, the eight T-cell clones expressing V beta 17/V alpha 3 gene segments derived from only three different T cells. Furthermore, conserved amino acid motifs were found to be encoded in both the alpha and the beta-chain junctional regions. Together, these data show a local amplification of unique T lymphocytes in both patients. The use of identical V beta J beta and V alpha gene segments with similar junctional sequences by three different cells, evidenced in one of the two cases, strengthens the view that liver-infiltrating T lymphocytes are selected locally by autoantigens in PBC.

  13. Entering the Clone Age

    Institute of Scientific and Technical Information of China (English)

    1995-01-01

    Suppose you make your parents so happy,they decide to have another baby just like you.It might be flattering,but how would you feel about having a little brother or sister who is also your twin? A laboratory experiment conducted last fall suggests it may someday be possible.For the first time ever,scientists made exact copies, or clones, of a human embryo.

  14. Analysis of cDNAs and Genomic DNA of Snake Venom CTL-like Proteins Revealed An Extraordinary Post-transcriptional Processing Event%蛇毒C型凝集素类蛋白cDNA与基因组DNA序列分析显示特别的转录后加工

    Institute of Scientific and Technical Information of China (English)

    查向东; 周立志; 黄河胜; 刘兢; 徐康森

    2004-01-01

    为研究蛇毒C型凝集素类蛋白的快速进化机制和结构功能关系,使用PCR技术扩增了若干编码C型凝集素类蛋白β链的cDNA分子以及agkisasin β的基因组DNA,并将这些扩增产物进行克隆和测序.对测序结果与试验过程中的具体条件进行了因果关系分析,并且进行点阵图比较和多序列比对.结果表明,可能存在"转录后同源重组"等转录后的事件,在蛇毒C型凝集素类蛋白的多样性上起着重要的作用.对于解释基因数目与蛋白质数目的差异这一后基因组时代的重要问题,具有一定的参考价值.首次报告蛇毒C型凝集素类蛋白的基因组DNA序列,其中未发现有内含子.%To better understand the accelerated evolution of snake venom C-type lectin-like proteins (CTL-like proteins) and to investigate the structure-function relationships, PCR was conducted to amplify cDNAs coding for the β chains of snake venom CTL-like proteins and the genomic DNA of agkisasin β. The reaction products were cloned and sequenced. The causal relationships between the sequences and the experimental conditions were established. Dot plot analysis and multiple alignments were also performed. The results suggested the existence of a post-transcriptional processing event that was essentially homologous recombination at the RNA level, which might play an important role in the diversity of snake venom CTL-like proteins. This inference would provide a novel perspective for explaining a challenging problem of the post-genomic era: the discrepancy between the limited number of genes and the large collection of cDNAs, which was most prominent with regard to certain snake venom proteins. The genomic DNA of a snake venom C-type lectin-like protein was elucidated and no introns were found in the coding region.

  15. Astrocyte cytolysis by MHC class II-specific mouse T cell clones.

    Science.gov (United States)

    Reder, A T; Lascola, C D; Flanders, S A; Maimone, D; Jensen, M A; Skias, D D; Lancki, D W

    1993-08-01

    The brain is "immunologically privileged," in part because class I and II MHC antigens are not normally present on glia or neurons. However, under certain conditions such as transplantation, glial cells express MHC proteins at levels sufficient for glia to become targets of immune responses. Cultured astrocytes expressing very low levels of MHC class I protein are killed efficiently by MHC class I antigen-specific CTL. Mouse brain allografts, however, are rejected by CD4+ T cells that are likely to be class II MHC-specific. The level of expression of MHC class II antigen needed to trigger specific killing of astrocytes by CD4+ T cells, independent of exogenous antigen, has not been studied. We examined the role of glial class II MHC in the lysis of cultured neonatal mouse astrocytes by an alloreactive murine CD4+ CTL alone. IFN-gamma induced functionally relevant increases in MHC class II antigen on target cells. Astrocytes were lysed by the CD4+ clone only when class II MHC antigens reached levels readily detectable by flow cytometry. MHC class II expression and lysis increased when astrocytes were coincubated with IFN-gamma and TNF-alpha. Conversely, lysis decreased when class II expression was downregulated by IFN-alpha/beta or dbcAMP. Cytolysis by CD4+ clones was blocked by antibodies to CD4 and LFA-1 on T cells, and to ICAM-1 and class II molecules on astrocytes. The role of LFA-1 in CD4+ cell-mediated lysis was greater than that of LFA-1/ICAM-1 in CD8+ T cell-mediated lysis. CD4+ cells may lyse activated astrocytes when the immune privilege of the brain is compromised as in transplantation, tumors, and inflammatory diseases.

  16. Sequential cloning of chromosomes

    Science.gov (United States)

    Lacks, S.A.

    1995-07-18

    A method for sequential cloning of chromosomal DNA of a target organism is disclosed. A first DNA segment homologous to the chromosomal DNA to be sequentially cloned is isolated. The first segment has a first restriction enzyme site on either side. A first vector product is formed by ligating the homologous segment into a suitably designed vector. The first vector product is circularly integrated into the target organism`s chromosomal DNA. The resulting integrated chromosomal DNA segment includes the homologous DNA segment at either end of the integrated vector segment. The integrated chromosomal DNA is cleaved with a second restriction enzyme and ligated to form a vector-containing plasmid, which is replicated in a host organism. The replicated plasmid is then cleaved with the first restriction enzyme. Next, a DNA segment containing the vector and a segment of DNA homologous to a distal portion of the previously isolated DNA segment is isolated. This segment is then ligated to form a plasmid which is replicated within a suitable host. This plasmid is then circularly integrated into the target chromosomal DNA. The chromosomal DNA containing the circularly integrated vector is treated with a third, retrorestriction (class IIS) enzyme. The cleaved DNA is ligated to give a plasmid that is used to transform a host permissive for replication of its vector. The sequential cloning process continues by repeated cycles of circular integration and excision. The excision is carried out alternately with the second and third enzymes. 9 figs.

  17. Cloning-free CRISPR

    Directory of Open Access Journals (Sweden)

    Mandana Arbab

    2015-11-01

    Full Text Available We present self-cloning CRISPR/Cas9 (scCRISPR, a technology that allows for CRISPR/Cas9-mediated genomic mutation and site-specific knockin transgene creation within several hours by circumventing the need to clone a site-specific single-guide RNA (sgRNA or knockin homology construct for each target locus. We introduce a self-cleaving palindromic sgRNA plasmid and a short double-stranded DNA sequence encoding the desired locus-specific sgRNA into target cells, allowing them to produce a locus-specific sgRNA plasmid through homologous recombination. scCRISPR enables efficient generation of gene knockouts (∼88% mutation rate at approximately one-sixth the cost of plasmid-based sgRNA construction with only 2 hr of preparation for each targeted site. Additionally, we demonstrate efficient site-specific knockin of GFP transgenes without any plasmid cloning or genome-integrated selection cassette in mouse and human embryonic stem cells (2%–4% knockin rate through PCR-based addition of short homology arms. scCRISPR substantially lowers the bar on mouse and human transgenesis.

  18. Secure the Clones

    CERN Document Server

    Jensen, Thomas; Pichardie, David

    2012-01-01

    Exchanging mutable data objects with untrusted code is a delicate matter because of the risk of creating a data space that is accessible by an attacker. Consequently, secure programming guidelines for Java stress the importance of using defensive copying before accepting or handing out references to an internal mutable object. However, implementation of a copy method (like clone()) is entirely left to the programmer. It may not provide a sufficiently deep copy of an object and is subject to overriding by a malicious sub-class. Currently no language-based mechanism supports secure object cloning. This paper proposes a type-based annotation system for defining modular copy policies for class-based object-oriented programs. A copy policy specifies the maximally allowed sharing between an object and its clone. We present a static enforcement mechanism that will guarantee that all classes fulfil their copy policy, even in the presence of overriding of copy methods, and establish the semantic correctness of the ove...

  19. Ethical issues in livestock cloning.

    Science.gov (United States)

    Thompson, P B

    1999-01-01

    Although cloning may eventually become an important technology for livestock production, four ethical issues must be addressed before the practice becomes widespread. First, researchers must establish that the procedure is not detrimental to the health or well-being of affected animals. Second, animal research institutions should evaluate the net social benefits to livestock producers by weighing the benefits to producers against the opportunity cost of research capacity lost to biomedical projects. Third, scientists should consider the indirect effects of cloning research on the larger ethical issues surrounding human cloning. Finally, the market structure for products of cloned animals should protect individual choice, and should recognize that many individuals find the prospect of cloning (or consuming cloned animals) repugnant. Analysis of these four issues is complicated by spurious arguments alleging that cloning will have a negative impact on environment and genetic diversity.

  20. Prognostic Factors Related to Clinical Response in Patients with Metastatic MelanomaTreated by CTL-Associated Antigen-4 Blockade

    Science.gov (United States)

    Downey, Stephanie G.; Klapper, Jacob A.; Smith, Franz O.; C.Yang, James; Sherry, Richard M.; Royal, Richard E.; Kammula, Udai S.; Hughes, Marybeth S.; Allen, Tamika E.; Levy, Catherine L.; Michael, Yellin; Nichol, Geoffrey; E.White, Donald; Steinberg, Seth M.; Rosenberg, Steven A.

    2007-01-01

    Purpose CTL-associated antigen 4 (CTLA-4) can inhibit T-cell activation and helps maintain peripheral self-tolerance. Previously, we showed immune-related adverse events (IRAE) and objective, durable clinical responses in patients with metastatic melanoma treated with CTLA-4 blockade.We have now treated139 patients in two trials and have sufficient follow-up to examine factors associated with clinical response. Experimental Design A total of 139 patients with metastatic melanoma were treated: 54 patients received ipilimumab in conjunction with peptide vaccinations and 85 patients were treated with intrapatient dose escalation of ipilimumab and randomized to receive peptides in accordance with HLA-A*0201status. Results Three patients achieved complete responses (CR; ongoing at 29+, 52+, and 53+ months); an additional 20 patients achieved partial responses (PR) for an overall objective response rate of 17%. The majority of patients (62%, 86 of 139) developed some form of IRAE, which was associated with a greater probability of objective antitumor response (P = 0.0004); all patients with CR had more severe IRAEs. Prior therapy with IFNα-2b was a negative prognostic factor, whereas prior high-dose interleukin-2 did not significantly affect the probability of response. There were no significant differences in the rate of clinical response or development of IRAEs between the two trials. The duration of tumor response was not affected by the use of high-dose steroids for abrogation of treatment-related toxicities (P = 0.23). There were no treatment-related deaths. Conclusion In patients with metastatic melanoma, ipilimumab can induce durable objective clinical responses, which are related to the induction of IRAEs. PMID:17982122

  1. Anergy in self-directed B lymphocytes: A statistical mechanics perspective.

    Science.gov (United States)

    Agliari, Elena; Barra, Adriano; Del Ferraro, Gino; Guerra, Francesco; Tantari, Daniele

    2015-06-21

    Self-directed lymphocytes may evade clonal deletion at ontogenesis but still remain harmless due to a mechanism called clonal anergy. For B-lymphocytes, two major explanations for anergy developed over the last decades: according to Varela theory, anergy stems from a proper orchestration of the whole B-repertoire, such that self-reactive clones, due to intensive feed-back from other clones, display strong inertia when mounting a response. Conversely, according to the model of cognate response, self-reacting cells are not stimulated by helper lymphocytes and the absence of such signaling yields anergy. Through statistical mechanics we show that helpers do not prompt activation of a sub-group of B-cells: remarkably, the latter are just those broadly interacting in the idiotypic network. Hence Varela theory can finally be reabsorbed into the prevailing framework of the cognate response model. Further, we show how the B-repertoire architecture may emerge, where highly connected clones are self-directed as a natural consequence of ontogenetic learning.

  2. Studies on rabbit lymphocytes in vitro

    Science.gov (United States)

    Sell, S.; Gell, P. G. H.

    1969-01-01

    Anti-allotypic sera that have no known allotypic determinants other than those also present in the genotype of the lymphocyte donor are as able to induce lymphocyte `blast' transformation in vitro as are anti-allotypic sera that do have allotypic determinants that are not present in the lymphocyte donor. Therefore, anti-allotypic sera do not appear to function in the stimulation of blast transformation by providing access for any of the known allotypic determinants into lymphocytes. PMID:5769980

  3. Rapid exacerbation of lymphocytic infundibuloneurohypophysitis

    Science.gov (United States)

    Shibue, Kimitaka; Fujii, Toshihito; Goto, Hisanori; Yamashita, Yui; Sugimura, Yoshihisa; Tanji, Masahiro; Yasoda, Akihiro; Inagaki, Nobuya

    2017-01-01

    Abstract Rationale: Lymphocytic hypophysitis is a relatively rare autoimmune disease defined by lymphocytic infiltration to the pituitary. Its rarity and wide spectrum of clinical manifestations make clarification of the pathology difficult. Here, we describe a case we examined from the primary diagnosis to final discharge, showing the serial progression of lymphocytic infundibuloneurohypophysitis (LINH) to panhypopituitarism with extrapituitary inflammatory invasion in a short period, and responding favorably to high-dose glucocorticoid treatment. Patient concerns: Polyuria, General fatigue and Nausea/Vomiting. Diagnoses: Central diabetes insipidus (CDI), Lymphocytic infundibuloneurohypophysitis (LINH). Interventions: Desmopressin acetate, High-dose glucocorticoid (GC) treatment. Outcomes: He was prescribed desmopressin acetate and subsequently discharged. A month later, he revisited our hospital with general fatigue and nausea/vomiting. A screening test disclosed hypopituitarism with adrenal insufficiency. MRI revealed expanded contrast enhancement to the peripheral extrapituitary lesion. He received high-dose GC treatment and the affected lesion exhibited marked improvement on MRI, along with the recovery of the anterior pituitary function. Lessons: This case demonstrates the potential for classical LINH to develop into panhypopituitarsim. We consider this is the first documentation of approaching the cause of atypical LINH with progressive clinical course from the pathological viewpoint. PMID:28248860

  4. Treatment of chronic lymphocytic leukemia.

    Science.gov (United States)

    Ferrajoli, Alessandra; O'Brien, Susan M

    2004-04-01

    Treatment options for patients with chronic lymphocytic leukemia have changed over the past two decades. This article reviews the experience accumulated with the use of alkylating agents alone and in combination; purine analogues alone and in combination and monoclonal antibodies such as rituximab, and alemtuzumab alone and in combination. The results obtained with different treatment strategies are summarized, compared, and reviewed.

  5. Anergy in self-directed B lymphocytes from a statistical mechanics perspective

    CERN Document Server

    Agliari, Elena; Del Ferraro, Gino; Guerra, Francesco; Tantari, Daniele

    2012-01-01

    The ability of the adaptive immune system to discriminate between self and non-self mainly stems from the ontogenic clonal-deletion of lymphocytes expressing strong binding affinity with self-peptides. However, some self-directed lymphocytes may evade selection and still be harmless due to a mechanism called clonal anergy. As for B lymphocytes, two major explanations for anergy developed over three decades: according to "Varela theory", it stems from a proper orchestration of the whole B-repertoire, in such a way that self-reactive clones, due to intensive interactions and feed-back from other clones, display more inertia to mount a response. On the other hand, according to the `two-signal model", which has prevailed nowadays, self-reacting cells are not stimulated by helper lymphocytes and the absence of such signaling yields anergy. The first result we present, achieved through disordered statistical mechanics, shows that helper cells do not prompt the activation and proliferation of a certain sub-group of ...

  6. Long term presence of a single predominant tyrosinase-specific T-cell clone associated with disease control in a patient with metastatic melanoma.

    Science.gov (United States)

    Ochsenreither, Sebastian; Fusi, Alberto; Busse, Antonia; Letsch, Anne; Haase, Doreen; Thiel, Eckhard; Scheibenbogen, Carmen; Keilholz, Ulrich

    2010-05-15

    In an earlier study, we described a patient who developed an anti-tyrosinase T-cell response leading to long-term tumor control. Here we analyzed this response with regard to T-cell receptor (TCR) Vbeta family usage and clonality in order to further elucidate the nature of the T cell response in this patient. For identification of expanded specific cytotoxic T-cell (CTL) clones, tetramer enrichment of tyrosinase reactive T-cells was followed by comparative quantitative reverse transcribed PCR (qRT PCR) quantification of all TCR Vbeta-families and sequencing of family Vbeta4 elevated in the enriched fraction. The predominant specific clone was quantified by clonotypic qRT PCR in multiple samples from blood, bone marrow, and tumor tissue. FACS analyses with staining of TYR.A2 and TCR Vbeta4 were performed. Epitope specific enrichment revealed an isolated increase of Vbeta-family 4. FACS analysis showed a shift of specific CTLs to Vbeta-family 4 during tumor regression with a maximum of 80% of all TYR.A2 specific cells belonging to this family. Sequencing revealed a single predominant clone against polyclonal background coding for identical CDR3 loops. The predominant clone was highly expressed in bone marrow and tumor tissue, and was detectable in blood over a period of ten years. Considering the results of previous studies showing a specific effector phenotype in blood and a specific memory compartment in bone marrow of this patient, this data implicate the predominant clone featured all attributes of a sufficient CTL response including homing capacity and memory formation resulting in long term clonal persistence and tumor control.

  7. Ethical issues in animal cloning.

    Science.gov (United States)

    Fiester, Autumn

    2005-01-01

    The issue of human reproductive cloning has recently received a great deal attention in public discourse. Bioethicists, policy makers, and the media have been quick to identify the key ethical issues involved in human reproductive cloning and to argue, almost unanimously, for an international ban on such attempts. Meanwhile, scientists have proceeded with extensive research agendas in the cloning of animals. Despite this research, there has been little public discussion of the ethical issues raised by animal cloning projects. Polling data show that the public is decidedly against the cloning of animals. To understand the public's reaction and fill the void of reasoned debate about the issue, we need to review the possible objections to animal cloning and assess the merits of the anti-animal cloning stance. Some objections to animal cloning (e.g., the impact of cloning on the population of unwanted animals) can be easily addressed, while others (e.g., the health of cloned animals) require more serious attention by the public and policy makers.

  8. Exosomes derived form bladder transitional cell carcinoma cells induce CTL cytotoxicity in vitro%膀胱移行细胞癌来源的exosome诱导体外细胞毒性T细胞杀伤效应

    Institute of Scientific and Technical Information of China (English)

    张家模; 吴小候; 张尧; 夏雨果; 罗春丽

    2009-01-01

    目的 观察膀胱移行细胞癌T24细胞来源的exosome体外诱导细胞毒性特异性T淋巴细胞(CTL)对肿瘤细胞的杀伤效应.方法 采用超滤和蔗糖密度梯度离心法分离T24细胞释放的exosome,电镜、Western blot观察exosome的特征.将exosome和肿瘤细胞负载到人外周血分离培养的树突状细胞(Dc)上,并与T细胞体外共同培养,分为exosome致敏DC组、未致敏DC组和对照组,Alamar blue检测CTL对T24细胞的细胞毒活性.结果 T24细胞分泌的exosome为直径约30~90nm的类圆碟形小囊泡.Western blot证实,exosome表达热休克蛋白70(HSP70)、细胞间黏附分子1(ICAM-1)和人细胞角蛋白20(CK20)分子.与未致敏DC组和对照组比较,exosome致敏DC组活化的T细胞对T24细胞有更强的细胞毒活性(P<0.01).结论 T24细胞来源的exosome负载了HSP70、ICAM-1等免疫相关蛋白;exosome经DC负载后活化CTL产生抗肿瘤活性.%Objective To isolate and purify exosomes derived from human bladder transitional cell carcinoma T24 cells,analyze the morphology and protein composition,and investigate the antitumor effect of specific cytotoxic T lymphocytes induced by exosomes.Methods Exosomes were isolated and purified by ultrafihration and sucrose gradient centrifugation,and characterized by electron microscopy and Western blot.Dendritic cells were amplified and purified from peripheral blood and pulsed with exosomes.Then they were co-cultured with T cells,and divided into 3 groups:exosome-pulsed DC group,unplused DC group and control group.Alamar-Blue assay was used to evaluate the specific cytolytic activity.Results The exosomes were in size about 30~90 nm saucer-shaped membranous vesicles.HSP70,ICAM-1 and CK20 were detected by Western blot.The CTL induced by DC pulsed with exosomes had significant cytolytic activity (P<0.01).Conclusion The exosomes derived from T24 cells are loaded with immunoprotein HSP70 and ICAM-1,and DC pulsed with exosomes can promote the anti

  9. To clone or not to clone--whither the law?

    Science.gov (United States)

    Lupton, M L

    1999-01-01

    The cloning of Dolly the lamb from adult cells by scientists at the Roslin Laboratories near Edinburgh in February 1997 has startled the world because it now opens the way to clone adult human beings. The reaction to Ian Wilmut's breakthrough has been instant and largely negative. Bills were rushed into both the US Senate and House of Representatives aimed at banning the cloning of human beings. Human cloning is premature at this stage, but there are many positive spin-offs of cloning in the field of genetic engineering, such as the production of human proteins such as blood clotting factors which aid in healing wounds. Progress by means of cloning can also be made into devising a cure for Parkinson's Disease amongst others. No lesser ethicist than John C. Fletcher of the University of Virginia foresees circumstances in which human cloning is acceptable e.g. to enable a couple to replace a dying child, to enable a couple, one of whom is infertile, to clone a child from either partner. Extensive regulation of cloning by the law is inevitable but, in doing so, the legislation should be careful not to outlaw research in this area which could be beneficial to mankind.

  10. Crystal structure of swine major histocompatibility complex class I SLA-1 0401 and identification of 2009 pandemic swine-origin influenza A H1N1 virus cytotoxic T lymphocyte epitope peptides.

    Science.gov (United States)

    Zhang, Nianzhi; Qi, Jianxun; Feng, Sijia; Gao, Feng; Liu, Jun; Pan, Xiaocheng; Chen, Rong; Li, Qirun; Chen, Zhaosan; Li, Xiaoying; Xia, Chun; Gao, George F

    2011-11-01

    The presentation of viral epitopes to cytotoxic T lymphocytes (CTLs) by swine leukocyte antigen class I (SLA I) is crucial for swine immunity. To illustrate the structural basis of swine CTL epitope presentation, the first SLA crystal structures, SLA-1 0401, complexed with peptides derived from either 2009 pandemic H1N1 (pH1N1) swine-origin influenza A virus (S-OIV(NW9); NSDTVGWSW) or Ebola virus (Ebola(AY9); ATAAATEAY) were determined in this study. The overall peptide-SLA-1 0401 structures resemble, as expected, the general conformations of other structure-solved peptide major histocompatibility complexes (pMHC). The major distinction of SLA-1 0401 is that Arg(156) has a "one-ballot veto" function in peptide binding, due to its flexible side chain. S-OIV(NW9) and Ebola(AY9) bind SLA-1 0401 with similar conformations but employ different water molecules to stabilize their binding. The side chain of P7 residues in both peptides is exposed, indicating that the epitopes are "featured" peptides presented by this SLA. Further analyses showed that SLA-1 0401 and human leukocyte antigen (HLA) class I HLA-A 0101 can present the same peptides, but in different conformations, demonstrating cross-species epitope presentation. CTL epitope peptides derived from 2009 pandemic S-OIV were screened and evaluated by the in vitro refolding method. Three peptides were identified as potential cross-species influenza virus (IV) CTL epitopes. The binding motif of SLA-1 0401 was proposed, and thermostabilities of key peptide-SLA-1 0401 complexes were analyzed by circular dichroism spectra. Our results not only provide the structural basis of peptide presentation by SLA I but also identify some IV CTL epitope peptides. These results will benefit both vaccine development and swine organ-based xenotransplantation.

  11. Crystal Structure of Swine Major Histocompatibility Complex Class I SLA-1*0401 and Identification of 2009 Pandemic Swine-Origin Influenza A H1N1 Virus Cytotoxic T Lymphocyte Epitope Peptides ▿

    Science.gov (United States)

    Zhang, Nianzhi; Qi, Jianxun; Feng, Sijia; Gao, Feng; Liu, Jun; Pan, Xiaocheng; Chen, Rong; Li, Qirun; Chen, Zhaosan; Li, Xiaoying; Xia, Chun; Gao, George F.

    2011-01-01

    The presentation of viral epitopes to cytotoxic T lymphocytes (CTLs) by swine leukocyte antigen class I (SLA I) is crucial for swine immunity. To illustrate the structural basis of swine CTL epitope presentation, the first SLA crystal structures, SLA-1*0401, complexed with peptides derived from either 2009 pandemic H1N1 (pH1N1) swine-origin influenza A virus (S-OIVNW9; NSDTVGWSW) or Ebola virus (EbolaAY9; ATAAATEAY) were determined in this study. The overall peptide–SLA-1*0401 structures resemble, as expected, the general conformations of other structure-solved peptide major histocompatibility complexes (pMHC). The major distinction of SLA-1*0401 is that Arg156 has a “one-ballot veto” function in peptide binding, due to its flexible side chain. S-OIVNW9 and EbolaAY9 bind SLA-1*0401 with similar conformations but employ different water molecules to stabilize their binding. The side chain of P7 residues in both peptides is exposed, indicating that the epitopes are “featured” peptides presented by this SLA. Further analyses showed that SLA-1*0401 and human leukocyte antigen (HLA) class I HLA-A*0101 can present the same peptides, but in different conformations, demonstrating cross-species epitope presentation. CTL epitope peptides derived from 2009 pandemic S-OIV were screened and evaluated by the in vitro refolding method. Three peptides were identified as potential cross-species influenza virus (IV) CTL epitopes. The binding motif of SLA-1*0401 was proposed, and thermostabilities of key peptide–SLA-1*0401 complexes were analyzed by circular dichroism spectra. Our results not only provide the structural basis of peptide presentation by SLA I but also identify some IV CTL epitope peptides. These results will benefit both vaccine development and swine organ-based xenotransplantation. PMID:21900158

  12. Simultaneous assessment of cytotoxic T lymphocyte responses against multiple viral infections by combined usage of optimal epitope matrices, anti- CD3 mAb T-cell expansion and "RecycleSpot"

    Directory of Open Access Journals (Sweden)

    Wong Johnson T

    2005-05-01

    Full Text Available Abstract The assessment of cellular anti-viral immunity is often hampered by the limited availability of adequate samples, especially when attempting simultaneous, high-resolution determination of T cell responses against multiple viral infections. Thus, the development of assay systems, which optimize cell usage, while still allowing for the detailed determination of breadth and magnitude of virus-specific cytotoxic T lymphocyte (CTL responses, is urgently needed. This study provides an up-to-date listing of currently known, well-defined viral CTL epitopes for HIV, EBV, CMV, HCV and HBV and describes an approach that overcomes some of the above limitations through the use of peptide matrices of optimally defined viral CTL epitopes in combination with anti-CD3 in vitro T cell expansion and re-use of cells from negative ELISpot wells. The data show that, when compared to direct ex vivo cell preparations, antigen-unspecific in vitro T cell expansion maintains the breadth of detectable T cell responses and demonstrates that harvesting cells from negative ELISpot wells for re-use in subsequent ELISpot assays (RecycleSpot, further maximized the use of available cells. Furthermore when combining T cell expansion and RecycleSpot with the use of rationally designed peptide matrices, antiviral immunity against more than 400 different CTL epitopes from five different viruses can be reproducibly assessed from samples of less than 10 milliliters of blood without compromising information on the breadth and magnitude of these responses. Together, these data support an approach that facilitates the assessment of cellular immunity against multiple viral co-infections in settings where sample availability is severely limited.

  13. Fludarabine Phosphate, Radiation Therapy, and Rituximab in Treating Patients Who Are Undergoing Donor Stem Cell Transplant Followed by Rituximab for High-Risk Chronic Lymphocytic Leukemia or Small Lymphocytic Lymphoma

    Science.gov (United States)

    2017-03-27

    Chronic Lymphocytic Leukemia; Prolymphocytic Leukemia; Recurrent Small Lymphocytic Lymphoma; Refractory Chronic Lymphocytic Leukemia; Stage III Chronic Lymphocytic Leukemia; Stage III Small Lymphocytic Lymphoma; Stage IV Chronic Lymphocytic Leukemia; Stage IV Small Lymphocytic Lymphoma; T-Cell Large Granular Lymphocyte Leukemia

  14. Fludarabine Phosphate and Total-Body Irradiation Before Donor Peripheral Blood Stem Cell Transplant in Treating Patients With Chronic Lymphocytic Leukemia or Small Lymphocytic Leukemia

    Science.gov (United States)

    2016-07-18

    B-Cell Prolymphocytic Leukemia; Chronic Lymphocytic Leukemia; Prolymphocytic Leukemia; Recurrent Chronic Lymphocytic Leukemia; Recurrent Small Lymphocytic Lymphoma; Refractory Chronic Lymphocytic Leukemia; T-Cell Prolymphocytic Leukemia

  15. Lessons learned from cloning dogs.

    Science.gov (United States)

    Kim, M J; Oh, H J; Kim, G A; Park, J E; Park, E J; Jang, G; Ra, J C; Kang, S K; Lee, B C

    2012-08-01

    The aim of this article is to review dog cloning research and to suggest its applications based on a discussion about the normality of cloned dogs. Somatic cell nuclear transfer was successfully used for production of viable cloned puppies despite limited understanding of in vitro dog embryo production. Cloned dogs have similar growth characteristics to those born from natural fertilization, with no evidence of serious adverse effects. The offspring of cloned dogs also have similar growth performance and health to those of naturally bred puppies. Therefore, cloning in domestic dogs can be applied as an assisted reproductive technique to conserve endangered species, to treat sterile canids or aged dogs, to improve reproductive performance of valuable individuals and to generate disease model animals.

  16. Molecular Cloning of Adenosinediphosphoribosyl Transferase.

    Science.gov (United States)

    1987-09-08

    ACCESSION NO.D,. 03261102F 2312 A~5 11. TITLE (include Securqt Classification) 0 Molecular Cloning of Adenosinediphosphoribosyl Transferase 12. PERSONAL...I’:- AFOSR.Tlt. 8 7 - 0 9 8,2 0IL * pi AFOSR- 85 -0377 PROGRESS REPORT Molecular Cloning of Adenosinediphosphoribosyl Transferase 5." Period of...Pharmacology and the Cardiovascular Research Institute September 8, 1987 .’, 5.’- "’S ". -f, AFOSR - 85 -0377 PROGRESS REPORT Molecular Cloning of

  17. Human cloning and child welfare.

    Science.gov (United States)

    Burley, J; Harris, J

    1999-01-01

    In this paper we discuss an objection to human cloning which appeals to the welfare of the child. This objection varies according to the sort of harm it is expected the clone will suffer. The three formulations of it that we will consider are: 1. Clones will be harmed by the fearful or prejudicial attitudes people may have about or towards them (H1); 2. Clones will be harmed by the demands and expectations of parents or genotype donors (H2); 3. Clones will be harmed by their own awareness of their origins, for example the knowledge that the genetic donor is a stranger (H3). We will show why these three versions of the child welfare objection do not necessarily supply compelling reasons to ban human reproductive cloning. The claim that we will develop and defend in the course of our discussion is that even if it is the case that a cloned child will suffer harms of the type H1-H3, it is none the less permissible to conceive by cloning so long as these cloning-induced welfare deficits are not such as to blight the existence of the resultant child, whoever this may be. PMID:10226914

  18. Molecular cloning of a new immunomodulatory protein from Anoectochilus formosanus which induces B cell IgM secretion through a T-independent mechanism.

    Directory of Open Access Journals (Sweden)

    Yen-Chou Kuan

    Full Text Available An immunomodulatory protein (IPAF was purified and cloned from Anoectochilus formosanus, an Orchidaceae herbal plant in Asia. The major targeting immune cells of IPAF and its modulating effects toward B lymphocytes were investigated. Rapid amplification of cDNA ends (RACE was conducted to clone the IPAF gene, and the obtained sequence was BLAST compared on the NCBI database. MACS-purified mouse T and B lymphocytes were stimulated with IPAF and the cell proliferation, activation, and Igs production were examined. IPAF comprised a 25 amino acids signal peptide and a 138 amino acids protein which was homologous to the lectins from Orchidaceae plant. IPAF selectively induced the cell proliferation in mouse splenic B lymphocytes but not T lymphocytes. The IPAF-induced B cells exhibited increased CD69 and MHC class II expression, and a dose- and time-dependent enhancement in IgM production. These results suggested potential benefits of IPAF to strengthen the humoral immunity.

  19. Development of a Luminex Bead Based Assay for Diagnosis of Toxocariasis Using Recombinant Antigens Tc-CTL-1 and Tc-TES-26.

    Science.gov (United States)

    Anderson, John P; Rascoe, Lisa N; Levert, Keith; Chastain, Holly M; Reed, Matthew S; Rivera, Hilda N; McAuliffe, Isabel; Zhan, Bin; Wiegand, Ryan E; Hotez, Peter J; Wilkins, Patricia P; Pohl, Jan; Handali, Sukwan

    2015-01-01

    The clinical spectrum of human disease caused by the roundworms Toxocara canis and Toxocara cati ranges from visceral and ocular larva migrans to covert toxocariasis. The parasite is not typically recovered in affected tissues, so detection of parasite-specific antibodies is usually necessary for establishing a diagnosis. The most reliable immunodiagnostic methods use the Toxocara excretory-secretory antigens (TES-Ag) in ELISA formats to detect Toxocara-specific antibodies. To eliminate the need for native parasite materials, we identified and purified immunodiagnostic antigens using 2D gel electrophoresis followed by electrospray ionization mass spectrometry. Three predominant immunoreactive proteins were found in the TES; all three had been previously described in the literature: Tc-CTL-1, Tc-TES-26, and Tc-MUC-3. We generated Escherichia coli expressed recombinant proteins for evaluation in Luminex based immunoassays. We were unable to produce a functional assay with the Tc-MUC-3 recombinant protein. Tc-CTL-1 and Tc-TES-26 were successfully coupled and tested using defined serum batteries. The use of both proteins together generated better results than if the proteins were used individually. The sensitivity and specificity of the assay for detecting visceral larval migrans using Tc-CTL-1 plus Tc-TES-26 was 99% and 94%, respectively; the sensitivity for detecting ocular larval migrans was 64%. The combined performance of the new assay was superior to the currently available EIA and could potentially be employed to replace current assays that rely on native TES-Ag.

  20. PD1 blockade reverses the suppression of melanoma antigen-specific CTL by CD4+ CD25(Hi) regulatory T cells.

    Science.gov (United States)

    Wang, Wenshi; Lau, Roy; Yu, Daohai; Zhu, Weiwei; Korman, Alan; Weber, Jeffrey

    2009-09-01

    Regulatory CD4(+)CD25(Hi) T cells (Treg) and programmed death-1 (PD-1) molecule have emerged as pivotal players in immune regulation. However, the underlying mechanisms by which they impact antigen-specific CD8(+) immune responses in cancer patients and how they interact with each other under physiologic conditions remain unclear. Herein, we examined the relationship of PD-1 and its abrogation to the function of Treg in patients with melanoma using short-term in vitro assays to generate melanoma-specific T cells. We identified Treg in the circulation of vaccinated melanoma patients and detected PD-1 expression on vaccine-induced melanoma antigen-specific CTLs, as well as on and within Treg from patients' peripheral blood. Programmed death ligand (PD-L) 1 expression was also detected on patients' Treg. PD-1 blockade promoted the generation of melanoma antigen-specific CTLs and masked their inhibition by Treg. The mechanisms by which PD-1 blockade mediated immune enhancement included direct augmentation of melanoma antigen-specific CTL proliferation, heightening their resistance to inhibition by Treg and direct limitation of the inhibitory ability of Treg. PD-1 blockade reversed the increased expression of PD-1 and PD-L1 on melanoma antigen-specific CTL by Treg, rescued INF-gamma and IL-2 or INF-gamma and tumor necrosis factor-alpha co-expression and expression of IL-7 receptor by melanoma antigen-specific CTL which were diminished by Treg. PD-1 blockade also resulted in down-regulation of intracellular FoxP3 expression by Treg. These data suggest that PD-1 is importantly implicated in the regulation of Treg function in melanoma patients.

  1. PD1 blockade reverses the suppression of melanoma antigen-specific CTL by CD4+CD25Hi regulatory T cells

    Science.gov (United States)

    Lau, Roy; Yu, Daohai; Zhu, Weiwei; Korman, Alan; Weber, Jeffrey

    2009-01-01

    Regulatory CD4+CD25Hi T cells (Treg) and programmed death-1 (PD-1) molecule have emerged as pivotal players in immune regulation. However, the underlying mechanisms by which they impact antigen-specific CD8+ immune responses in cancer patients and how they interact with each other under physiologic conditions remain unclear. Herein, we examined the relationship of PD-1 and its abrogation to the function of Treg in patients with melanoma using short-term in vitro assays to generate melanoma-specific T cells. We identified Treg in the circulation of vaccinated melanoma patients and detected PD-1 expression on vaccine-induced melanoma antigen-specific CTLs, as well as on and within Treg from patients’ peripheral blood. Programmed death ligand (PD-L) 1 expression was also detected on patients’ Treg. PD-1 blockade promoted the generation of melanoma antigen-specific CTLs and masked their inhibition by Treg. The mechanisms by which PD-1 blockade mediated immune enhancement included direct augmentation of melanoma antigen-specific CTL proliferation, heightening their resistance to inhibition by Treg and direct limitation of the inhibitory ability of Treg. PD-1 blockade reversed the increased expression of PD-1 and PD-L1 on melanoma antigen-specific CTL by Treg, rescued INF-γ and IL-2 or INF-γ and tumor necrosis factor-α co-expression and expression of IL-7 receptor by melanoma antigen-specific CTL which were diminished by Treg. PD-1 blockade also resulted in down-regulation of intracellular FoxP3 expression by Treg. These data suggest that PD-1 is importantly implicated in the regulation of Treg function in melanoma patients. PMID:19651643

  2. Lamprey buccal gland secretory protein-2 (BGSP-2 inhibits human T lymphocyte proliferation

    Directory of Open Access Journals (Sweden)

    Jing SUN, Shuiyan YU, Zhuang XUE, Cenjie LIU, Yu WU, Xin LIU, Qingwei LI

    2010-04-01

    Full Text Available Lamprey is a representative of the agnathans, the most ancient class of vertebrates. Parasitic lampreys secrete anticoagulant from their buccal glands and prevent blood coagulation of host fishes. We identified a buccal gland secretory protein-2 (BGSP-2 from a buccal gland cDNA library of Lampetra japonica. The full-length BGSP-2 gene was cloned and the recombinant BGSP-2 protein was generated. The role of BGSP-2 on lymphocyte proliferation was studied by examining its effects on human T lymphocytes. We found that lamprey BGSP-2 was able to effectively block the proliferation of T cells in vitro by inducing G1/S cell cycle arrest. Furthermore, it inhibited the proliferation of human T lymphocytes stimulated by phytohemagglutinin (PHA at a minimum concentration of 0.1μg/ml. Our data suggest that lamprey BGSP-2 is able to block the mitosis of human T lymphocytes at the G1/S point, and has the potential of anti-proliferative effect on PHA-activated T lymphocytes [Current Zoology 56 (2: 252–258, 2010].

  3. Potential elucidation of a novel CTL epitope in HIV-1 protease by the protease inhibitor resistance mutation L90M.

    Directory of Open Access Journals (Sweden)

    Werner Smidt

    Full Text Available The combination of host immune responses and use of antiretrovirals facilitate partial control of human immunodeficiency virus type 1 (HIV-1 infection and result in delayed progression to Acquired Immunodeficiency Syndrome (AIDS. Both treatment and host immunity impose selection pressures on the highly mutable HIV-1 genome resulting in antiretroviral resistance and immune escape. Researchers have shown that antiretroviral resistance mutations can shape cytotoxic T-lymphocyte immunity by altering the epitope repertoire of HIV infected cells. Here it was discovered that an important antiretroviral resistance mutation, L90M in HIV protease, occurs at lower frequencies in hosts that harbor the B*15, B*48 or A*32 human leukocyte antigen subtypes. A likely reason is the elucidation of novel epitopes by L90M. NetMHCPan predictions reveal increased affinity of the peptide spanning the HIV protease region, PR 89-97 and PR 90-99 to HLA-B*15/B*48 and HLA-A*32 respectively due to the L90M substitution. The higher affinity could increase the chance of the epitope being presented and recognized by Cytotoxic T-lymphocytes and perhaps provide additional immunological pressures in the presence of antiretroviral attenuating mutations. This evidence supports the notion that knowledge of HLA allotypes in HIV infected individuals could augment antiretroviral treatment by the elucidation of epitopes due to antiretroviral resistance mutations in HIV protease.

  4. [The discrete horror of cloning].

    Science.gov (United States)

    Guibourg, Ricardo A

    2009-01-01

    The author raises the topic of cloning after the decision of the Argentine government, which concerned for the "dignity of the human person", passed a decree of need and urgency, No. 200/97 (Annex), prohibiting cloning experiments with human beings. Therefore, considering that the topic is so terribly urgent and necessary, the author feels it is timely to consider it.

  5. CATO: The Clone Alignment Tool.

    Directory of Open Access Journals (Sweden)

    Peter V Henstock

    Full Text Available High-throughput cloning efforts produce large numbers of sequences that need to be aligned, edited, compared with reference sequences, and organized as files and selected clones. Different pieces of software are typically required to perform each of these tasks. We have designed a single piece of software, CATO, the Clone Alignment Tool, that allows a user to align, evaluate, edit, and select clone sequences based on comparisons to reference sequences. The input and output are designed to be compatible with standard data formats, and thus suitable for integration into a clone processing pipeline. CATO provides both sequence alignment and visualizations to facilitate the analysis of cloning experiments. The alignment algorithm matches each of the relevant candidate sequences against each reference sequence. The visualization portion displays three levels of matching: 1 a top-level summary of the top candidate sequences aligned to each reference sequence, 2 a focused alignment view with the nucleotides of matched sequences displayed against one reference sequence, and 3 a pair-wise alignment of a single reference and candidate sequence pair. Users can select the minimum matching criteria for valid clones, edit or swap reference sequences, and export the results to a summary file as part of the high-throughput cloning workflow.

  6. CATO: The Clone Alignment Tool.

    Science.gov (United States)

    Henstock, Peter V; LaPan, Peter

    2016-01-01

    High-throughput cloning efforts produce large numbers of sequences that need to be aligned, edited, compared with reference sequences, and organized as files and selected clones. Different pieces of software are typically required to perform each of these tasks. We have designed a single piece of software, CATO, the Clone Alignment Tool, that allows a user to align, evaluate, edit, and select clone sequences based on comparisons to reference sequences. The input and output are designed to be compatible with standard data formats, and thus suitable for integration into a clone processing pipeline. CATO provides both sequence alignment and visualizations to facilitate the analysis of cloning experiments. The alignment algorithm matches each of the relevant candidate sequences against each reference sequence. The visualization portion displays three levels of matching: 1) a top-level summary of the top candidate sequences aligned to each reference sequence, 2) a focused alignment view with the nucleotides of matched sequences displayed against one reference sequence, and 3) a pair-wise alignment of a single reference and candidate sequence pair. Users can select the minimum matching criteria for valid clones, edit or swap reference sequences, and export the results to a summary file as part of the high-throughput cloning workflow.

  7. Role of signaling lymphocytic activation molecule in T helper cell responses

    Directory of Open Access Journals (Sweden)

    Jan E. de Vries

    1998-01-01

    Full Text Available Signaling lymphocytic activation molecule (SLAM; CDw150 is a 70 kDa glycoprotein. Signaling lymphocytic activation molecule is constitutively expressed on memory T cells, CD56+ T cells, a subset of T cell receptor γδ+ cells, immature thymocytes and, at low levels, on a proportion of peripheral blood B cells. Signaling lymphocytic activation molecule is rapidly upregulated on all T and B cells after activation. Engagement of SLAM by F(ab’2 fragments of an anti-SLAM monoclonal antibody (mAb A12 enhances antigen-specific T cell proliferation. In addition, mAb A12 was directly mitogenic for T cell clones and activated T cells. T cell proliferation induced by mAb A12 is independent of interleukin (IL-2, IL-4, IL-12 and IL-15, but is cyclosporin A sensitive. Ligation of SLAM during antigen-specific T cell proliferation resulted in upregulation of interferon (IFN-γ production, even by allergen-specific T helper cell (Th 2 clones, whereas the levels of IL-4 and IL-5 production were only marginally affected. The mAb A12 was unable to induce IL-4 and IL-5 production by Th1 clones. Co-stimulation of skin-derived Der P1-specific Th2 cells from patients with atopic dermatitis via SLAM resulted in the generation of a population of IFN-γ-producing cells, thereby reverting their phenotype to a Th0 pattern. Signaling lymphocytic activation molecule is a high-affinity self ligand mediating homophilic cell interaction. In addition, soluble SLAM enhances both T and B cell proliferation. Collectively, these data indicate that SLAM molecules act both as receptors and ligands that are not only involved in T cell expansion but also drive the expanding T cells during immune responses into the Th0/Th1 pathway. This suggests that signaling through SLAM plays a role in directing Th0/Th1 development.

  8. [Scientific ethics of human cloning].

    Science.gov (United States)

    Valenzuela, Carlos Y

    2005-01-01

    True cloning is fission, budding or other types of asexual reproduction. In humans it occurs in monozygote twinning. This type of cloning is ethically and religiously good. Human cloning can be performed by twinning (TWClo) or nuclear transfer (NTClo). Both methods need a zygote or a nuclear transferred cell, obtained in vitro (IVTec). They are under the IVTec ethics. IVTecs use humans (zygotes, embryos) as drugs or things; increase the risk of malformations; increase development and size of abnormalities and may cause long-term changes. Cloning for preserving extinct (or almost extinct) animals or humans when sexual reproduction is not possible is ethically valid. The previous selection of a phenotype in human cloning violates some ethical principles. NTClo for reproductive or therapeutic purposes is dangerous since it increases the risk for nucleotide or chromosome mutations, de-programming or re-programming errors, aging or malignancy of the embryo cells thus obtained.

  9. Quantum probabilistically cloning and computation

    Institute of Scientific and Technical Information of China (English)

    2008-01-01

    In this article we make a review on the usefulness of probabilistically cloning and present examples of quantum computation tasks for which quantum cloning offers an advantage which cannot be matched by any approach that does not resort to it.In these quantum computations,one needs to distribute quantum information contained in states about which we have some partial information.To perform quantum computations,one uses state-dependent probabilistic quantum cloning procedure to distribute quantum information in the middle of a quantum computation.And we discuss the achievable efficiencies and the efficient quantum logic network for probabilistic cloning the quantum states used in implementing quantum computation tasks for which cloning provides enhancement in performance.

  10. Artificial antigen-presenting cells plus IL-15 and IL-21 efficiently induce melanoma-specific cytotoxic CD8+CD28+ T lymphocyte responses

    Institute of Scientific and Technical Information of China (English)

    Xia Yu; Yuan Fang; Xi Li; Nuo Zhou; Yong-Xiang Zhao; Xiao-Ling Lu; Jian He; Sodaly Mongkhoune; Yi Peng; Yuan Xie; Jing Su; Su-Fang Zhou; Xiao-Xun Xie; Guo-Rong Luo

    2013-01-01

    To develop a novel artificial antigen-presenting system for efficiently inducing melanoma-specific CD8+CD28+ cytotoxic T lymphocyte (CTL) responses. Methods: Cell-sized Dynabeads® M-450 Epoxy beads coated with H-2Kb:Ig-TRP2180-188 and anti-CD28 antibody were used as artificial antigen-presenting cells (aAPCs) to induce melanoma-specific CD8+CD28+CTL responses with the help of IL-21 and IL-15. Dimer staining, proliferation, ELISPOT, and cytotoxicity experiments were conducted to evaluate the frequency and activity of induced CTLs. Results: Dimer staining demonstrated that the new artificial antigen-presenting system efficiently induced melanoma TRP2-specific CD8+CD28+ CTLs. Proliferation and ELISPOT assays indicated that the induced CTLs rapidly proliferate and produce increased IFN-γ under the stimulation of H-2Kb:Ig-TRP2-aAPCs, IL-15, and IL-21. In addition, cytotoxicity experiments showed that induced CTLs have specific killing activity of target cells. Conclusions: The new artificial antigen-presenting system including aAPCs plus IL-21 and IL-15 can induce a large number of antigen-specific CD8+CD28+ CTLs against the melanoma. Our study provides evidence for a novel adoptive immunotherapy against tumors.

  11. Identification and characterization of a membrane-bound cytotoxin of murine cytolytic lymphocytes that is related to tumor necrosis factor/cachectin

    Energy Technology Data Exchange (ETDEWEB)

    Liu, Chauching; Detmers, P.A.; Jiang, Shibo; Young, J.D.E (Rockefeller Univ., New York, NY (USA))

    1989-05-01

    Cytotoxic T lymphocytes (CTLs) kill their targets by a contact-dependent mechanism. The authors investigated the possibility that the CTL membranes themselves could exert direct cytotoxic activity. Murine CTLs that had been fixed with paraformaldehyde retained a slow cytotoxic activity toward various target cells that are also sensitive to another cytokine, tumor necrosis factor (TNF)/cachectin. This cytotoxic activity was neutralized by antibodies specific for TNF. Membrane fractions obtained from CTLs were cytotoxic to TNF-sensitive targets but not to several TNF-resistant cell lines. Immunoblot analysis revealed a membrane protein band of 50-60 kDa from CTLs that reacts with anti-TNF antibodies. The surface localization of this cytokine was further ascertained by flow cytometry, indirect immunofluorescence, and immunoelectron microscopy studies using TNF-specific antibodies. Radioiodination of CTL surface proteins followed by immunoprecipitation with anti-TNF antibodies confirmed the presence of a TNF-related cytokine in the plasma membranes of CTLs that migrated with an apparent molecular mass of 50-60 kDa under disulfide-reducing conditions. This cytokine can be removed from membranes by treatment with detergents but not with high-salt buffers, suggesting that it may be an integral membrane protein.

  12. Clone Networks, Clone Extensions and Biregularizations of Varieties of Algebras

    Institute of Scientific and Technical Information of China (English)

    J. Plonka

    2001-01-01

    We consider algebras of type τ- without nullary operations. An identity ψ≈ψ of type τ is clone compatible if ψ and ψ are the same variable or the sets of fundamental operation symbols in ψ and ψ are non-empty and identical. For a variety V, we denote by Vc the variety defined by all clone compatible identities from Id(V). In this paper, we give a construction of algebras called a clone network. Under some assumptions, we describe algebras from Vc by means of this construction. We find some properties of Vc and applications.

  13. Opinion: Interactions of innate and adaptive lymphocytes

    Science.gov (United States)

    Gasteiger, Georg; Rudensky, Alexander Y.

    2015-01-01

    Innate lymphocytes, including natural killer (NK) cells and the recently discovered innate lymphoid cells (ILCs) have crucial roles during infection, tissue injury and inflammation. Innate signals regulate the activation and homeostasis of innate lymphocytes. Less well understood is the contribution of the adaptive immune system to the orchestration of innate lymphocyte responses. We review our current understanding of the interactions between adaptive and innate lymphocytes, and propose a model in which adaptive T cells function as antigen-specific sensors for the activation of innate lymphocytes to amplify and instruct local immune responses. We highlight the potential role of regulatory and helper T cells in these processes and discuss major questions in the emerging area of crosstalk between adaptive and innate lymphocytes. PMID:25132095

  14. Sequence conservation, HLA-E-Restricted peptide, and best-defined CTL/CD8+ epitopes in gag P24 (capsid) of HIV-1 subtype B

    Science.gov (United States)

    Prasetyo, Afiono Agung; Dharmawan, Ruben; Sari, Yulia; Sariyatun, Ratna

    2017-02-01

    Human immunodeficiency virus type 1 (HIV-1) remains a cause of global health problem. Continuous studies of HIV-1 genetic and immunological profiles are important to find strategies against the virus. This study aimed to conduct analysis of sequence conservation, HLA-E-restricted peptide, and best-defined CTL/CD8+ epitopes in p24 (capsid) of HIV-1 subtype B worldwide. The p24-coding sequences from 3,557 HIV subtype B isolates were aligned using MUSCLE and analysed. Some highly conserved regions (sequence conservation ≥95%) were observed. Two considerably long series of sequences with conservation of 100% was observed at base 349-356 and 550-557 of p24 (HXB2 numbering). The consensus from all aligned isolates was precisely the same as consensus B in the Los Alamos HIV Database. The HLA-E-restricted peptide in amino acid (aa) 14-22 of HIV-1 p24 (AISPRTLNA) was found in 55.9% (1,987/3,557) of HIV-1 subtype B worldwide. Forty-four best-defined CTL/CD8+ epitopes were observed, in which VKNWMTETL epitope (aa 181-189 of p24) restricted by B*4801 was the most frequent, as found in 94.9% of isolates. The results of this study would contribute information about HIV-1 subtype B and benefits for further works willing to develop diagnostic and therapeutic strategies against the virus.

  15. Feasibility of Telomerase-Specific Adoptive T-cell Therapy for B-cell Chronic Lymphocytic Leukemia and Solid Malignancies.

    Science.gov (United States)

    Sandri, Sara; Bobisse, Sara; Moxley, Kelly; Lamolinara, Alessia; De Sanctis, Francesco; Boschi, Federico; Sbarbati, Andrea; Fracasso, Giulio; Ferrarini, Giovanna; Hendriks, Rudi W; Cavallini, Chiara; Scupoli, Maria Teresa; Sartoris, Silvia; Iezzi, Manuela; Nishimura, Michael I; Bronte, Vincenzo; Ugel, Stefano

    2016-05-01

    Telomerase (TERT) is overexpressed in 80% to 90% of primary tumors and contributes to sustaining the transformed phenotype. The identification of several TERT epitopes in tumor cells has elevated the status of TERT as a potential universal target for selective and broad adoptive immunotherapy. TERT-specific cytotoxic T lymphocytes (CTL) have been detected in the peripheral blood of B-cell chronic lymphocytic leukemia (B-CLL) patients, but display low functional avidity, which limits their clinical utility in adoptive cell transfer approaches. To overcome this key obstacle hindering effective immunotherapy, we isolated an HLA-A2-restricted T-cell receptor (TCR) with high avidity for human TERT from vaccinated HLA-A*0201 transgenic mice. Using several relevant humanized mouse models, we demonstrate that TCR-transduced T cells were able to control human B-CLL progression in vivo and limited tumor growth in several human, solid transplantable cancers. TERT-based adoptive immunotherapy selectively eliminated tumor cells, failed to trigger a self-MHC-restricted fratricide of T cells, and was associated with toxicity against mature granulocytes, but not toward human hematopoietic progenitors in humanized immune reconstituted mice. These data support the feasibility of TERT-based adoptive immunotherapy in clinical oncology, highlighting, for the first time, the possibility of utilizing a high-avidity TCR specific for human TERT. Cancer Res; 76(9); 2540-51. ©2016 AACR.

  16. Treatment of solid organ transplant recipients with autologous Epstein Barr virus–specific cytotoxic T lymphocytes (CTLs)

    Science.gov (United States)

    Savoldo, Barbara; Goss, John A.; Hammer, Markus M.; Zhang, Lan; Lopez, Teresita; Gee, Adrian P.; Lin, Yu-Feng; Quiros-Tejeira, Ruben E.; Reinke, Petra; Schubert, Stephan; Gottschalk, Stephen; Finegold, Milton J.; Brenner, Malcolm K.; Rooney, Cliona M.; Heslop, Helen E.

    2006-01-01

    We have investigated the in vivo safety, efficacy, and persistence of autologous Epstein Barr virus (EBV)–specific cytotoxic T lymphocytes (CTLs) for the treatment of solid organ transplant (SOT) recipients at high risk for EBV-associated posttransplantation lymphoproliferative disease (PTLD). EBV-CTLs generated from 35 patients expanded with normal kinetics contained both CD8 and CD4 lymphocytes and produced significant specific killing of autologous EBV-transformed B lymphoblastoid cell lines (LCLs). Twelve SOT recipients at high risk for PTLD, or with active disease, received autologous CTL infusions without toxicity. Real-time polymerase chain reaction (PCR) monitoring of EBV-DNA showed a transient increase in plasma EBV-DNA suggestive of lysis of EBV-infected cells, although there was no consistent decrease in virus load in peripheral-blood mononuclear cells. Interferon-γ enzyme-linked immunospot (ELISPOT) assay and tetramer analysis showed an increase in the frequency of EBV-responsive T cells, which returned to preinfusion levels after 2 to 6 months. None of the treated patients developed PTLD. One patient with liver PTLD showed a complete response, and one with ocular disease has had a partial response stable for over one year. These data are consistent with an expansion and persistence of adoptively transferred EBV-CTLs that is limited in the presence of continued immunosuppression but that nonetheless produces clinically useful antiviral activity. PMID:16835376

  17. Limitations on Cloning in Classical Mechanics

    OpenAIRE

    Fenyes, Aaron

    2010-01-01

    In this paper, we show that a result precisely analogous to the traditional quantum no-cloning theorem holds in classical mechanics. This classical no-cloning theorem does not prohibit classical cloning, we argue, because it is based on a too-restrictive definition of cloning. Using a less popular, more inclusive definition of cloning, we give examples of classical cloning processes. We also prove that a cloning machine must be at least as complicated as the object it is supposed to clone.

  18. [Adoptive transfer of T lymphocytes].

    Science.gov (United States)

    Vié, H; Clémenceau, B

    2017-09-01

    Within a few years, the success of treatments based on the use of T-cells armed with a chimeric T-receptor for the CD19 molecule (CAR-T CD19) has revolutionized the perception of adoptive transfer approaches. The levels of responses observed in acute leukemias, of the order of 70-90 % are indeed unprecedented. The medical and financial enthusiasm aroused by these results has led to the current situation where more than 300 clinical trials are under way, against some thirty different antigens. This enthusiasm, well justified by the first successes, must however be tempered by the difficulties associated with the use of these cells. Indeed, the management of patients is made very complex both for medical reasons, because the toxicities associated with these treatments are important, and for technical reasons, because the preparation of T lymphocytes for therapeutic use requires dedicated structures. During this same period, knowledge of the mechanisms of regulation of T lymphocytes and the possibilities offered by synthetic biology and techniques of genome engineering have progressed considerably. Combined, they allow envisaging a true "programming" of the T lymphocytes, intended to improve the efficiency of the treatments and the safety of the patients. Medical and industrial perspectives and the role of these approaches in the arsenal of cancer therapies will depend largely on two conditions: the emergence of a robust demonstration of their effectiveness in solid tumors, and the establishment of an acceptable production and distribution model 1. Copyright © 2017 Elsevier Masson SAS. All rights reserved.

  19. Lymphocyte subpopulations in Sheehan's syndrome.

    Science.gov (United States)

    Atmaca, Hulusi; Araslı, Mehmet; Yazıcı, Zihni Acar; Armutçu, Ferah; Tekin, Ishak Özel

    2013-06-01

    The role of autoimmunity in the development of Sheehan's syndrome is obscure. There are a limited number of studies investigating the immunological alterations accompanying Sheehan's Syndrome. Our objective was to evaluate lymphocyte subsets in these patients. We conducted a cross-sectional clinical study. Cytofluorometry was used for the immunophenotyping of peripheral blood leukocytes from patients with Sheehan's syndrome followed up in the endocrine clinic during 2005-2009. Fifteen consecutive patients (mean age 61.6 ± 11.3, range 34-75 years) and 25 healthy controls (mean age 56.7 ± 10.6, range 34-80 years) were included. There was no statistically significant difference between the groups in terms of mean age. The percentages of CD19(+), CD16(+)/56(+), CD8(+)28(-), γδTCR(+), CD8(+); the total lymphocyte counts; and the ratio of CD8(+)28(-)/CD8(+)28(+) were similar (p > 0.05) between patients and controls. Whereas the leucocyte counts (p = 0.003), the percentage of CD3 (+) DR (+) (p Sheehan's syndrome compared to healthy controls. There was a positive correlation between the duration of illness and the percentage of CD3(+)DR(+) (r = 0.53, p = 0.03) expression. Some peripheral lymphocyte cell subsets show marked variation in patients with Sheehan's syndrome in comparison to matched healthy subjects, which may have implications for altered immune regulation in these patients. High CD3 (+) DR (+) expression that correlates with the duration of illness in Sheehan's patients is suggestive of an ongoing inflammation accompanying the slow progression of pituitary dysfunction in Sheehan's syndrome. It is not clear if these cellular alterations contribute to the cause or consequence of pituitary deficiency in Sheehan's syndrome.

  20. Adipose tissue lymphocytes: types and roles.

    Science.gov (United States)

    Caspar-Bauguil, S; Cousin, B; Bour, S; Casteilla, L; Castiella, L; Penicaud, L; Carpéné, C

    2009-12-01

    Besides adipocytes, specialized in lipid handling and involved in energy balance regulation, white adipose tissue (WAT) is mainly composed of other cell types among which lymphocytes represent a non-negligible proportion. Different types of lymphocytes (B, alphabetaT, gammadeltaT, NK and NKT) have been detected in WAT of rodents or humans, and vary in their relative proportion according to the fat pad anatomical location. The lymphocytes found in intra-abdominal, visceral fat pads seem representative of innate immunity, while those present in subcutaneous fat depots are part of adaptive immunity, at least in mice. Both the number and the activity of the different lymphocyte classes, except B lymphocytes, are modified in obesity. Several of these modifications in the relative proportions of the lymphocyte classes depend on the degree of obesity, or on leptin concentration, or even fat depot anatomical location. Recent studies suggest that alterations of lymphocyte number and composition precede the macrophage increase and the enhanced inflammatory state of WAT found in obesity. Lymphocytes express receptors to adipokines while several proinflammatory chemokines are produced in WAT, rendering intricate crosstalk between fat and immune cells. However, the evidences and controversies available so far are in favour of an involvement of lymphocytes in the control of the number of other cells in WAT, either adipocytes or immune cells and of their secretory and metabolic activities. Therefore, immunotherapy deserves to be considered as a promising approach to treat the endocrino-metabolic disorders associated to excessive fat mass development.

  1. Autoimmune hepatitis in association with lymphocytic colitis.

    LENUS (Irish Health Repository)

    Cronin, Edmond M

    2012-02-03

    Autoimmune hepatitis is a rare, chronic inflammatory disorder which has been associated with a number of other auto-immune conditions. However, there are no reports in the medical literature of an association with microscopic (lymphocytic) colitis. We report the case of a 53-year-old woman with several autoimmune conditions, including lymphocytic colitis, who presented with an acute hepatitis. On the basis of the clinical features, serology, and histopathology, we diagnosed autoimmune hepatitis. To our knowledge, this is the first report of autoimmune hepatitis in association with lymphocytic colitis, and lends support to the theory of an autoimmune etiology for lymphocytic colitis.

  2. Lymphocyte-platelet crosstalk in Graves' disease.

    Science.gov (United States)

    Kuznik, Boris I; Vitkovsky, Yuri A; Gvozdeva, Olga V; Solpov, Alexey V; Magen, Eli

    2014-03-01

    Platelets can modulate lymphocytes' role in the pathophysiology of thyroid autoimmune diseases. The present study was performed to clarify the status of platelet-lymphocyte subpopulations aggregation in circulating blood in patients with Graves' disease (GD). One hundred and fifty patients with GD (GD group) and 45 hyperthyroid patients with toxic multinodular goiter (TMG group) were recruited in the study. Control group consisted 150 healthy subjects. Immunophenotyping of lymphocytes was performed by flow cytometry. Detection of lymphocyte-platelet aggregates (LPAs) was done using light microscope after Ficoll-gradient centrifugation. The group of GD patients exhibited reduced CD8 lymphocyte and higher CD19 cell counts compared with TMG group and healthy controls. A greater number of activated CD3, HLA-DR+ lymphocytes were observed in GD than in TMG group and control group. GD group was characterized by lower blood platelet count (232 ± 89 × 10 cells/µL) than TMG group (251 ± 97 × 10 cells/µL; P < 0.05) and control group (262 ± 95 × 10 cells/µL; P < 0.05). In GD group, more platelet-bound lymphocytes (332 ± 91 /µL) were found than that in TMG group (116 ± 67/µL, P < 0.005) and control group (104 ± 58 /µL; P < 0.001). GD is associated with higher levels of activated lymphocytes and lymphocyte-platelet aggregates.

  3. Composite mantle cell lymphoma and chronic lymphocytic leukemia/small lymphocytic lymphoma: a clinicopathologic and molecular study.

    Science.gov (United States)

    Hoeller, Sylvia; Zhou, Yi; Kanagal-Shamanna, Rashmi; Xu-Monette, Zijun Y; Hoehn, Daniela; Bihl, Michel; Swerdlow, Steven H; Rosenwald, Andreas; Ott, German; Said, Jonathan; Dunphy, Cherie H; Bueso-Ramos, Carlos E; Lin, Pei; Wang, Michael; Miranda, Roberto N; Tzankov, Alexander; Medeiros, L Jeffrey; Young, Ken H

    2013-01-01

    Mantle cell lymphoma (MCL) and chronic lymphocytic leukemia/small lymphocytic lymphoma (CLL/SLL) share many features and both arise from CD5+ B-cells; their distinction is critical as MCL is a more aggressive neoplasm. Rarely, cases of composite MCL and CLL/SLL have been reported. Little is known about the nature of these cases and, in particular, the clonal relationship of the 2 lymphomas. Eleven composite MCL and CLL/SLL cases were identified. The clinical, morphologic and immunophenotypic features of the MCL and CLL/SLL were characterized. IGH (immunoglobulin heavy chain) gene analysis was performed on microdissected MCL and CLL/SLL components to assess their clonal relationship. Ten patients had lymphadenopathy, and 7 patients had bone marrow involvement. The MCL component had the following growth patterns: in situ (n = 1), mantle zone (n = 3), nodular and diffuse (n = 3), diffuse (n = 3), and interstitial in the bone marrow (the only patient without lymphadenopathy) (n = 1); 6 MCLs had blastoid or pleomorphic and 5 small lymphocytic features. The CLL/SLL component was nodular (n = 9) or diffuse (n = 2). All MCL were CD5(+) and cyclin D1(+) with t(11;14) translocation. All CLL/SLL were CD5(+), CD23(+) and negative for cyclin D1 or t(11;14). IGH gene analysis showed that the MCL and CLL/SLL components displayed different sized fragments, indicating that the MCL and CLL/SLL are likely derived from different neoplastic B-cell clones. The lack of a clonal relationship between the MCL and CLL/SLL components suggests that MCL and CLL/SLL components represent distinct disease processes and do not share a common progenitor B-cell.

  4. Seamless Ligation Cloning Extract (SLiCE) cloning method.

    Science.gov (United States)

    Zhang, Yongwei; Werling, Uwe; Edelmann, Winfried

    2014-01-01

    SLiCE (Seamless Ligation Cloning Extract) is a novel cloning method that utilizes easy to generate bacterial cell extracts to assemble multiple DNA fragments into recombinant DNA molecules in a single in vitro recombination reaction. SLiCE overcomes the sequence limitations of traditional cloning methods, facilitates seamless cloning by recombining short end homologies (15-52 bp) with or without flanking heterologous sequences and provides an effective strategy for directional subcloning of DNA fragments from bacterial artificial chromosomes or other sources. SLiCE is highly cost-effective and demonstrates the versatility as a number of standard laboratory bacterial strains can serve as sources for SLiCE extract. We established a DH10B-derived E. coli strain expressing an optimized λ prophage Red recombination system, termed PPY, which facilitates SLiCE with very high efficiencies.

  5. Viral load and clinical disease enhancement associated with a lentivirus cytotoxic T lymphocyte vaccine regimen

    Science.gov (United States)

    Mealey, Robert H.; Leib, Steven R.; Littke, Matt H.; Wagner, Bettina; Horohov, David W.; McGuire, Travis C.

    2009-01-01

    Effective DNA-based vaccines against lentiviruses will likely induce CTL against conserved viral proteins. Equine infectious anemia virus (EIAV) infects horses worldwide, and serves as a useful model for lentiviral immune control. Although attenuated live EIAV vaccines have induced protective immune responses, DNA-based vaccines have not. In particular, DNA-based vaccines have had limited success in inducing CTL responses against intracellular pathogens in the horse. We hypothesized that priming with a codon-optimized plasmid encoding EIAV Gag p15/p26 with co-administration of a plasmid encoding an equine IL-2/IgG fusion protein as a molecular adjuvant, followed by boosting with a vaccinia vector expressing Gag p15/p26, would induce protective Gag-specific CTL responses. Although the regimen induced Gag-specific CTL in four of seven vaccinated horses, CTL were not detected until after the vaccinia boost, and protective effects were not observed in EIAV challenged vaccinates. Unexpectedly, vaccinates had significantly higher viral loads and more severe clinical disease, associated with the presence of vaccine-induced CTL. It was concluded that 1.) further optimization of the timing and route of DNA immunization was needed for efficient CTL priming in vivo, 2.) co-administration of the IL-2/IgG plasmid did not enhance CTL priming by the Gag p15/p26 plasmid, 3.) vaccinia vectors are useful for lentivirus-specific CTL induction in the horse, 4.) Gag-specific CTL alone are either insufficient or a more robust Gag-specific CTL response is needed to limit EIAV viremia and clinical disease, and 5.) CTL-inducing vaccines lacking envelope immunogens can result in lentiviral disease enhancement. Although the mechanisms for enhancement associated with this vaccine regimen remain to be elucidated, these results have important implications for development of lentivirus T cell vaccines. PMID:19368787

  6. Therapeutic cloning in the mouse

    Science.gov (United States)

    Mombaerts, Peter

    2003-01-01

    Nuclear transfer technology can be applied to produce autologous differentiated cells for therapeutic purposes, a concept termed therapeutic cloning. Countless articles have been published on the ethics and politics of human therapeutic cloning, reflecting the high expectations from this new opportunity for rejuvenation of the aging or diseased body. Yet the research literature on therapeutic cloning, strictly speaking, is comprised of only four articles, all in the mouse. The efficiency of derivation of embryonic stem cell lines via nuclear transfer is remarkably consistent among these reports. However, the efficiency is so low that, in its present form, the concept is unlikely to become widespread in clinical practice. PMID:12949262

  7. Biomimetic Cloning of Quantum Observables

    CERN Document Server

    Alvarez-Rodriguez, U; Lamata, L; Solano, E

    2013-01-01

    We propose a bio-inspired sequential quantum protocol for the cloning and preservation of the statistics associated to quantum observables of a given system. It combines the cloning of a set of commuting observables, permitted by the no-cloning and no-broadcasting theorems, with a controllable propagation of the initial state coherences to the subsequent generations. The protocol mimics the scenario in which an individual in an unknown quantum state copies and propagates its quantum information into an environment of blank qubits. Finally, we propose a realistic experimental implementation of this protocol in trapped ions.

  8. Cloning: revisiting an old debate.

    Science.gov (United States)

    Verhey, Allen D

    1994-09-01

    The debate about cloning that took place 25 years ago, although directed toward a different sort of cloning, elucidates fundamental issues currently at stake in reproductive technologies and research. Paul Ramsey and Joseph Fletcher were participants in this early debate. The differences between Ramsey and Fletcher about the meaning and sufficiency of freedom, the understanding and weighing of good and evil, the connection between embodiment and personhood, the relationship of humans with nature, and the meaning of parenthood suggest both a broader agenda for the debate about cloning and a cautious move forward in the development of embryo-splitting.

  9. Anti-tumor Effects of T Lymphocytes Induced by Dendritic Cells Pulsed with ATC-IC%ATC-IC致敏的树突状细胞诱导T淋巴细胞抗肿瘤效应的研究

    Institute of Scientific and Technical Information of China (English)

    孙海燕; 刘晓辉; 杨明花; 马楠; 刘云鹏; 姜又红

    2012-01-01

    Objective To investigate the activation and proliferation of cytotoric T lymphocytes (CTL) induced by dendritic cells pulsed with the complexes ATC-IC and their anti-tumor effects on renal carcinoma cells. Methods The complexes ATC-1C were prepared with apoptotic renal carcinoma cells and G250mAb. Monocytes and T lymphocytes were isolated from the peripheral blood in healthy subjects. DCs were cultivated in monocytes by granulocyte-macrophage colony and interleukin. The DCs stimulated by ATC-IC were experimental group, while the DCs loaded with ATC and without any stimulation were control group. The effects of the DCs in each group on CTL were tested. The effects of CTL on renal carcinoma cells (786-0) and lung cancer cells (.4549) were tested by CCK-8 assays. Results Compared with the control group,the proliferation of CTL induced by DC pulsed with the complexes ATC-IC was statistically significant (P < 0.01). Compared with the control group,proliferated CTL had higher cytotnxicity against 786-0 cells(P<0.05). Two groups had no significant killing activity on A549 cells. Conclusion DCs pulsed by ATC-IC could induce a significant T lymphocyte proliferation and the proliferated CTL could induce specific anti-kidney tumor effects in vitro.%目的 探讨凋亡的肾癌细胞与G250单克隆抗体(mAb)形成的复合物(ATC-IC)致敏树突状细胞(DC)后,DC对T淋巴细胞的激活、增殖作用及对肾透明细胞癌细胞的抑癌效应.方法 制备凋亡的肾癌细胞与G250 mAb形成的复合物ATC-IC;分离健康供血者外周血单个核细胞及T淋巴细胞,联合应用粒—巨细胞集落刺激因子及白细胞介素从单个核细胞中培养出DC,以制备的ATC-IC刺激DC为实验组,以凋亡的肾癌细胞刺激DC和未经任何因素刺激的DC为对照组,检测各组DC对T淋巴细胞的细胞增殖动力学的影响,并用CCK-8测定诱导的细胞毒性T淋巴细胞对肾癌细胞株786-0和肺癌细胞株A549的杀伤活性.结果 ATC-IC

  10. Methylotroph cloning vehicle

    Science.gov (United States)

    Hanson, Richard S.; Allen, Larry N.

    1989-04-25

    A cloning vehicle comprising: a replication determinant effective for replicating the vehicle in a non-C.sub.1 -utilizing host and in a C.sub.1 -utilizing host; DNA effective to allow the vehicle to be mobilized from the non-C.sub.1 -utilizing host to the C.sub.1 -utilizing host; DNA providing resistance to two antibiotics to which the wild-type C.sub.1 -utilizing host is susceptible, each of the antibiotic resistance markers having a recognition site for a restriction endonuclease; a cos site; and a means for preventing replication in the C.sub.1 -utilizing host. The vehicle is used for complementation mapping as follows. DNA comprising a gene from the C.sub.1 -utilizing organism is inserted at the restriction nuclease recognition site, inactivating the antibiotic resistance marker at that site. The vehicle can then be used to form a cosmid structure to infect the non-C.sub.1 -utilizing (e.g., E. coli) host, and then conjugated with a selected C.sub.1 -utilizing mutant. Resistance to the other antibiotic by the mutant is a marker of the conjugation. Other phenotypical changes in the mutant, e.g., loss of an auxotrophic trait, is attributed to the C.sub.1 gene. The vector is also used to inactivate genes whose protein products catalyze side reactions that divert compounds from a biosynthetic pathway to a desired product, thereby producing an organism that makes the desired product in higher yields.

  11. Changes of lymphocyte kinetics in the normal rat, induced by the lymphocyte mobilizing agent polymethacrylic acid

    NARCIS (Netherlands)

    Ormai, S.; Hagenbeek, A.; Palkovits, M.; Bekkum, D.W. van

    1973-01-01

    The changes in lymphocyte kinetics induced by the lymphocyte mobilizing agent polymethacrylic acid (PMAA) were studied in the normal rat. Quantitative data are presented concerning the degree of lymphocyte mobilization in the spleen and in various lymph nodes at different times after PMAA administra

  12. Allorestricted T lymphocytes with a high avidity T-cell receptor towards NY-ESO-1 have potent anti-tumor activity.

    Science.gov (United States)

    Krönig, Holger; Hofer, Kathrin; Conrad, Heinke; Guilaume, Philippe; Müller, Julia; Schiemann, Matthias; Lennerz, Volker; Cosma, Antonio; Peschel, Christian; Busch, Dirk H; Romero, Pedro; Bernhard, Helga

    2009-08-01

    The cancer-testis antigen NY-ESO-1 has been targeted as a tumor-associated antigen by immunotherapeutical strategies, such as cancer vaccines. The prerequisite for a T-cell-based therapy is the induction of T cells capable of recognizing the NY-ESO-1-expressing tumor cells. In this study, we generated human T lymphocytes directed against the immunodominant NY-ESO-1(157-165) epitope known to be naturally presented with HLA-A*0201. We succeeded to isolate autorestricted and allorestricted T lymphocytes with low, intermediate or high avidity TCRs against the NY-ESO-1 peptide. The avidity of the established CTL populations correlated with their capacity of lysing HLA-A2-positive, NY-ESO-1-expressing tumor cell lines derived from different origins, e.g. melanoma and myeloma. The allorestricted NY-ESO-1-specific T lymphocytes displayed TCRs with the highest avidity and best anti-tumor recognition activity. TCRs derived from allorestricted, NY-ESO-1-specific T cells may be useful reagents for redirecting primary T cells by TCR gene transfer and, therefore, may facilitate the development of adoptive transfer regimens based on TCR-transduced T cells for the treatment of NY-ESO-1-expressing hematological malignancies and solid tumors.

  13. Paraflagellar rod protein-specific CD8+ cytotoxic T lymphocytes target Trypanosoma cruzi-infected host cells.

    Science.gov (United States)

    Wrightsman, Ruth A; Luhrs, Keith A; Fouts, David; Manning, Jerry E

    2002-08-01

    Our previous studies show that in mice immunized with the paraflagellar rod (PFR) proteins of Trypanosoma cruzi protective immunity against this protozoan parasite requires MHC class I-restricted T cell function. To determine whether PFR-specific CD8+ T cell subsets are generated during T. cruzi infection, potential CTL targets in the PFR proteins were identified by scanning the amino acid sequences of the four PFR proteins for regions of 8-10 amino acids that conform to predicted MHC class I H-2b binding motifs. A subset of the peptide sequences identified were synthesized and tested as target antigen in 51Cr-release assays with effector cells from chronically infected T. cruzi mice. Short-term cytotoxic T lymphocyte (CTL) lines specific for two of the peptides, PFR-1(164-171) and PFR-3(123-130), showed high levels of lytic activity against peptide-pulsed target cells, secreted interferon (IFN)-gamma in response to parasite-infected target cells, and were found to be CD8+, CD4-, CD3+, TCRalphabeta+ cells of the Tc1 subset. Challenge of PFR immunized CD8-/- and perforin-deficient (PKO) mice confirmed that while CD8+ cells are required for survival of T. cruzi challenge infection, perforin activity is not required. Furthermore, while lytic activity of PFR-specific CD8+ T cell lines derived from PKO mice was severely impaired, the IFN-gamma levels secreted by CTLs from PKO mice were equivalent to that of normal mice, suggesting that the critical role played by CD8+ T cells in immunity to the parasite may be secretion of type 1 cytokines rather than lysis of parasite infected host cells.

  14. Effective Cytotoxic T Lymphocyte Targeting of Persistent HIV-1 during Antiretroviral Therapy Requires Priming of Naive CD8+ T Cells

    Directory of Open Access Journals (Sweden)

    Kellie N. Smith

    2016-05-01

    Full Text Available Curing HIV-1 infection will require elimination of persistent cellular reservoirs that harbor latent virus in the face of combination antiretroviral therapy (cART. Proposed immunotherapeutic strategies to cure HIV-1 infection include enhancing lysis of these infected cells by cytotoxic T lymphocytes (CTL. A major challenge in this strategy is overcoming viral immune escape variants that have evaded host immune control. Here we report that naive CD8+ T cells from chronic HIV-1-infected participants on long-term cART can be primed by dendritic cells (DC. These DC must be mature, produce high levels of interleukin 12p70 (IL-12p70, be responsive to CD40 ligand (CD40L, and be loaded with inactivated, autologous HIV-1. These DC-primed CD8+ T cell responders produced high levels of gamma interferon (IFN-γ in response to a broad range of both conserved and variable regions of Gag and effectively killed CD4+ T cell targets that were either infected with the autologous latent reservoir-associated virus or loaded with autologous Gag peptides. In contrast, HIV-1-specific memory CD8+ T cells stimulated with autologous HIV-1-loaded DC produced IFN-γ in response to a narrow range of conserved and variable Gag peptides compared to the primed T cells and most notably, displayed significantly lower cytolytic function. Our findings highlight the need to selectively induce new HIV-1-specific CTL from naive precursors while avoiding activation of existing, dysfunctional memory T cells in potential curative immunotherapeutic strategies for HIV-1 infection.

  15. Human Cloning: Let's Discuss It.

    Science.gov (United States)

    Taras, Loretta; Stavroulakis, Anthea M.; Ortiz, Mary T.

    1999-01-01

    Describes experiences with holding discussions on cloning at a variety of levels in undergraduate biology courses. Discusses teaching methods used and student reactions to the discussions. Contains 12 references. (WRM)

  16. A Clone of Your Own.

    Science.gov (United States)

    Bilodeau, Kirsten

    1997-01-01

    Describes an activity used at the Washington Park Arboretum that helps students understand cloning through plant propagation. Students also learn how to make a pot from recycled newspapers and how to make soil that is appropriate for the plants. (DDR)

  17. Human cloning and 'posthuman' society.

    Science.gov (United States)

    Blackford, Russell

    2005-01-01

    Since early 1997, when the creation of Dolly the sheep by somatic cell nuclear transfer was announced in Nature, numerous government reports, essays, articles and books have considered the ethical problems and policy issues surrounding human reproductive cloning. In this article, I consider what response a modern liberal society should give to the prospect of human cloning, if it became safe and practical. Some opponents of human cloning have argued that permitting it would place us on a slippery slope to a repugnant future society, comparable to that portrayed in Aldous Huxley's novel, Brave New World. I conclude that, leaving aside concerns about safety, none of the psychological or social considerations discussed in this article provides an adequate policy justification for invoking the state's coercive powers to prevent human cloning.

  18. Use of “one-pot, mix-and-read” peptide-MHC class I tetramers and predictive algorithms to improve detection of cytotoxic T lymphocyte responses in cattle

    Science.gov (United States)

    2014-01-01

    Peptide-major histocompatibility complex (p-MHC) class I tetramer complexes have facilitated the early detection and functional characterisation of epitope specific CD8+ cytotoxic T lymphocytes (CTL). Here, we report on the generation of seven recombinant bovine leukocyte antigens (BoLA) and recombinant bovine β2-microglobulin from which p-MHC class I tetramers can be derived in ~48 h. We validated a set of p-MHC class I tetramers against a panel of CTL lines specific to seven epitopes on five different antigens of Theileria parva, a protozoan pathogen causing the lethal bovine disease East Coast fever. One of the p-MHC class I tetramers was tested in ex vivo assays and we detected T. parva specific CTL in peripheral blood of cattle at day 15-17 post-immunization with a live parasite vaccine. The algorithm NetMHCpan predicted alternative epitope sequences for some of the T. parva CTL epitopes. Using an ELISA assay to measure peptide-BoLA monomer formation and p-MHC class I tetramers of new specificity, we demonstrate that a predicted alternative epitope Tp229-37 rather than the previously reported Tp227-37 epitope is the correct Tp2 epitope presented by BoLA-6*04101. We also verified the prediction by NetMHCpan that the Tp587-95 epitope reported as BoLA-T5 restricted can also be presented by BoLA-1*02301, a molecule similar in sequence to BoLA-T5. In addition, Tp587-95 specific bovine CTL were simultaneously stained by Tp5-BoLA-1*02301 and Tp5-BoLA-T5 tetramers suggesting that one T cell receptor can bind to two different BoLA MHC class I molecules presenting the Tp587-95 epitope and that these BoLA molecules fall into a single functional supertype. PMID:24775445

  19. Clonal heterogeneity of synovial fluid T lymphocytes from patients with rheumatoid arthritis

    Energy Technology Data Exchange (ETDEWEB)

    Duby, A.D.; Sinclair, A.K.; Osborne-Lawrence, S.L. (Univ. of Texas Southwestern Medical Center, Dallas (USA)); Zeldes, W.; Kan, Li; Fox, D.A. (Univ. of Michigan, Ann Arbor (USA))

    1989-08-01

    Although substantial evidence suggests that synovial T lymphocytes are critical in the pathogenesis of rheumatoid arthritis (RA), little is known regarding their antigenic specificities, antigen receptor gene rearrangements, and mechanisms of activation. To assess the extend of expansion of specific clones among RA synovial fluid T cells, Southern blot analyses of T-cell receptor (TCR) gene rearrangements were performed on 40 RA synovial fluid T-cell clones, as well as on fresh and polyclonally activated T cells from RA synovial fluid, RA peripheral blood, and normal peripheral blood. Two of the clones had identical TCR rearrangement patterns, but the remainder were unique. The nonclonal RA T-cell samples showed the same pattern of TCR {beta}-chain rearrangement that was observed among normal peripheral blood T cells, indicating no dominant clonal T-cell population in these samples. It was noted that with sufficient exposure of autoradiograms of the Southern blots, discrete TCR gene rearrangements, representing in some cases common D{sub {beta}}J{sub {beta}} (D, diversity; J, joining) rearrangements, were evident in T cells from peripheral blood of normal individuals and patients with RA, as well as T cells from RA synovial fluid. Taken together, the findings indicate that only a minor degree of oligoclonality can be demonstrated among T lymphocytes from RA synovial fluid.

  20. Sonographic characteristics of chronic lymphocytic thyroiditis with or without nodules%慢性淋巴细胞性甲状腺炎声像图表现及其结节的鉴别

    Institute of Scientific and Technical Information of China (English)

    原韶玲; 王寅; 张洪英; 卢伟; 赵培鹏

    2009-01-01

    Objective To study and distinguish the sonographic characteristics of chronic lymphocytic thyroiditis (CTL) and its nodules.Methods Ultrasonographic characteristics of 107 patients with histologically confirmed CTL was observed with a 7.5-12 MHz transducer and confirmed with the operation and pathology.Seventy four cases of CTL were examined by color Doppler flow image (CDFI),serum thyroid hormone levels were determined in 38 cases of CTL.Results In 57 cases of CTL,the pattern of diffuse low echoes was found in 14 cases (24.6 %),and nodulose pattern in 43 cases (75.4 %) in which diffuse numerous hypoecoie microdules in 16 (37.2%),and sporadic small nodules in 27 (62.8 %).All cases showed degeneration and disappearance of thyroid follicles,lymph cells invasion in varying degrees on pathology.Nodular goiter was commonly detected in 40 cases CTL (37.4 %),adenoma in 4 cases (3.7 %),hyperthyroidism in one case,non-Hodgkin lymphoma in one case,thyroid carcinoma in 4 cases (3.7 %).Color dopple flow imaging showed in 74 cases,0-Ⅰ 34 cases,Ⅱ 38 cases,Ⅲ 2 cases.The increased flow were observed in 38 cases in both Anti-Tpo and Anti-TG,and in 26 cases in TSH.Conclusion Ultrasonography is complex in CTL,with more complications.Ultrasonography are distinguished between the nodule of CTL and the nodule in the other thyroid diseases.Ultrasonography could help to avoid unnecessary surgical intervention,and provide strong evidence for CTL and its complication in clinic treatment.%目的 探讨慢性淋巴细胞性甲状腺炎(CTL)声像图表现及其结节的鉴别.方法 对107例CTL患者应用常规超声二维高频探头检测甲状腺大小、回声、内部结节及其钙化、液化情况,部分病例进行彩色多普勒血流及血清甲状腺内分泌水平测定,并与手术病理结果对照.结果 单纯CTL57例中,回声弥漫减低型CTL 14例(24.6%),结节型CTL 43例(75.4%),其中弥漫小结节型16例(37.2%),散在结节型27例(62.8%).病理表现以间

  1. Interpretation of NCCN Guideline: Chronic Lymphocytic Leukemia/Small Lymphocytic Lymphoma (Version 1, 2017

    Directory of Open Access Journals (Sweden)

    Lei XIA

    2016-12-01

    Full Text Available Chronic lymphocytic leukemia (CLL is a kind of chronic lymphocyte proliferative disease with corresponding clinical symptoms caused by the accumulation of mature B lymphocytes in peripheral blood, bone marrow and lymphatic tissues. In recent years, great achievements have been reached on the basic research, new prognostic markers, diagnostic criteria and therapeutic methods in CLL. This study mainly interpreted the corresponding diagnosis and treatment of CLL in NCCN Clinical Practice Guidelines in Oncology (NCCN Guidelines®: Chronic Lymphocytic Leukemia/Small Lymphocytic Lymphoma (Version 1, 2017.

  2. Islamic perspectives on human cloning.

    Science.gov (United States)

    Sadeghi, Mahmoud

    2007-01-01

    The present paper seeks to assess various views from Islamic jurists relating to human cloning, which is one of the controversial topics in the recent past. Taking Islamic jurisprudence principles, such as the rule of necessity for self preservation and respect for human beings, the rule of la darar wa la dirar ('the necessity to refrain from causing harm to oneself and others') and the rule of usr wa haraj, one may indicate that if human cloning could not be prohibited, as such, it could still be opposed because it gives way to various harmful consequences, which include family disorder, chaos in the clone's family relationships, physical and mental diseases for clones and suffering of egg donors and surrogate mothers. However with due attention to the fact that the reasons behind the prohibition of abortion only restrict the destruction of human embryos in their post-implantation stages, human cloning for biomedical research and exploitation of stem cells from cloned embryos at the blastocyst stage for therapeutic purposes would be acceptable.

  3. Cloning goes to the movies.

    Science.gov (United States)

    Cormick, Craig

    2006-10-01

    Public attitude research conducted by Biotechnology Australia shows that one of the major sources of information on human reproductive cloning is movies. Traditionally, understanding of new and emerging technologies has come through the mass media but human cloning, being so widely addressed through the popular culture of movies, is more effectively defined by Hollywood than the news media or science media. But how well are the science and social issues of cloning portrayed in box office hits such as The Island, Multiplicity, Star Wars: Attack of the Clones and Jurassic Park? These movies have enormous reach and undoubted influence, and are therefore worth analyzing in some detail. This study looks at 33 movies made between 1971 and 2005 that address human reproductive cloning, and it categorizes the films based on their genre and potential influence. Yet rather than simply rating the quality of the science portrayed, the study compares the key messages in these movies with public attitudes towards cloning, to examine the correlations.

  4. Artificial cloning of domestic animals.

    Science.gov (United States)

    Keefer, Carol L

    2015-07-21

    Domestic animals can be cloned using techniques such as embryo splitting and nuclear transfer to produce genetically identical individuals. Although embryo splitting is limited to the production of only a few identical individuals, nuclear transfer of donor nuclei into recipient oocytes, whose own nuclear DNA has been removed, can result in large numbers of identical individuals. Moreover, clones can be produced using donor cells from sterile animals, such as steers and geldings, and, unlike their genetic source, these clones are fertile. In reality, due to low efficiencies and the high costs of cloning domestic species, only a limited number of identical individuals are generally produced, and these clones are primarily used as breed stock. In addition to providing a means of rescuing and propagating valuable genetics, somatic cell nuclear transfer (SCNT) research has contributed knowledge that has led to the direct reprogramming of cells (e.g., to induce pluripotent stem cells) and a better understanding of epigenetic regulation during embryonic development. In this review, I provide a broad overview of the historical development of cloning in domestic animals, of its application to the propagation of livestock and transgenic animal production, and of its scientific promise for advancing basic research.

  5. Genetics Home Reference: bare lymphocyte syndrome type I

    Science.gov (United States)

    ... Home Health Conditions bare lymphocyte syndrome type I bare lymphocyte syndrome type I Enable Javascript to view ... boxes. Download PDF Open All Close All Description Bare lymphocyte syndrome type I (BLS I) is an ...

  6. A novel method for producing target cells and assessing cytotoxic T lymphocyte activity in outbred hosts

    Directory of Open Access Journals (Sweden)

    Bendinelli Mauro

    2009-03-01

    Full Text Available Abstract Background Cytotoxic T lymphocytes play a crucial role in the immunological control of microbial infections and in the design of vaccines and immunotherapies. Measurement of cytotoxic T lymphocyte activity requires that the test antigen is presented by target cells having the same or compatible class I major hystocompatibility complex antigens as the effector cells. Conventional assays use target cells labeled with 51chromium and infer cytotoxic T lymphocyte activity by measuring the isotope released by the target cells lysed following incubation with antigen-specific cytotoxic T lymphocytes. This assay is sensitive but needs manipulation and disposal of hazardous radioactive reagents and provides a bulk estimate of the reporter released, which may be influenced by spontaneous release of the label and other poorly controllable variables. Here we describe a novel method for producing target in outbred hosts and assessing cytotoxic T lymphocyte activity by flow cytometry. Results The method consists of culturing skin fibroblasts, immortalizing them with a replication defective clone of simian virus 40, and finally transducing them with a bicistronic vector encoding the target antigen and the reporter green fluorescent protein. When used in a flow cytometry-based assay, the target cells obtained with this method proved valuable for assessing the viral envelope protein specific cytotoxic T lymphocyte activity in domestic cats acutely or chronically infected with feline immunodeficiency virus, a lentivirus similar to human immunodeficiency virus and used as animal model for AIDS studies. Conclusion Given the versatility of the bicistronic vector used, its ability to deliver multiple and large transgenes in target cells, and its extremely wide cell specificity when pseudotyped with the vesicular stomatitis virus envelope protein, the method is potentially exploitable in many animal species.

  7. The Danish National Chronic Lymphocytic Leukemia Registry

    DEFF Research Database (Denmark)

    da Cunha-Bang, Caspar; Geisler, Christian Hartmann; Enggaard, Lisbeth

    2016-01-01

    AIM: In 2008, the Danish National Chronic Lymphocytic Leukemia Registry was founded within the Danish National Hematology Database. The primary aim of the registry is to assure quality of diagnosis and care of patients with chronic lymphocytic leukemia (CLL) in Denmark. Secondarily, to evaluate...

  8. Structured Review of Code Clone Literature

    NARCIS (Netherlands)

    Hordijk, Wiebe; Ponisio, María Laura; Wieringa, Roel

    2008-01-01

    This report presents the results of a structured review of code clone literature. The aim of the review is to assemble a conceptual model of clone-related concepts which helps us to reason about clones. This conceptual model unifies clone concepts from a wide range of literature, so that findings ab

  9. Structured Review of Code Clone Literature

    NARCIS (Netherlands)

    Hordijk, W.T.B.; Ponisio, Laura; Wieringa, Roelf J.

    2008-01-01

    This report presents the results of a structured review of code clone literature. The aim of the review is to assemble a conceptual model of clone-related concepts which helps us to reason about clones. This conceptual model unifies clone concepts from a wide range of literature, so that findings ab

  10. The anti-tumor activity of dendritic cell/cytotoxic T lymphocyte induced by cholangiocarcinoma-derived exosome and its ultrafiltered lysate%基于exosome裂解超滤液的树突状细胞/细胞毒性T淋巴细胞诱导及其抗胆管癌研究

    Institute of Scientific and Technical Information of China (English)

    陈炯煌; 丁国平; 陈文超; 曹利平

    2013-01-01

    Objective To observe the impact of cholangiocarcinoma-derived exosome and ultrafiltered exosome lysate on the anti-tumor activity of dendritic celL/cytotoxic T lymphocyte (DC/CTL) cells and discuss the mechanism involved.Methods Exosomes derived from RBE cells (human cholangiocarcinoma line) were collected by ultracentrifugation,ow-osmotic splitting followed by ultrafiltration was performed to remove exosomal microRNAs and purify the ultrafiltered exosome lysate.ImDCs (immature dendritic cells) were induced from peripheral blood and were co-cultured with CTL(cytotoxic lymphocyte),which were impulsed by exosome and ultrafiltered exosome lysate,respectively.The concentrations of tumor necrosis factor (TNF)-αt and perforin in culture medium supernatant were examined by enzyme linked immunosorbent assay (ELISA).Cell counting kit-8 (CCK-8) was used to evaluate the cytotoxic activity of DC/CTL to RBE cells.Results The concentrations of TNF-αand perforin in E-DC/CTL group were 138.61 ng/L and 2.41 μg/L respectively,lower than those of DC/CTL (194.08 ng/L and 3.39 μg/L) and EL-DC/CTL group (210.87 ng/L and 3.79 μg/L).The killing rate of E-DC/CTL was 33.35%,lower than that of DC/CTL (47.35%) and EL-DC/CTL (66.23%) significantly.The killing rate of EL-DC/CTL was significantly higher than that of DC/CTL (P < 0.01).Conclusion RBE cell-derived exosome inhibits the anti-tumor activity of DC/CTL by down-regulating TNF-α and perforin,exosomal miRNA may play important roles in the immune escape of cholangiocarcinoma.We built a new model based on ultrafiltered exosome lysate derived from cholangiocarcinoma to enhance the anti-tumor activity of DC/CTL.%目的 观察人胆管癌细胞来源的exosome及exosome裂解超滤液(UEL)对树突状细胞/细胞毒性T淋巴细胞诱导(DC/CTL)混合细胞抗肿瘤活性的影响及机制.方法 采用超速离心法提取人胆管癌细胞(RBE细胞)释放的exosome,低渗法裂解exosome,超

  11. Dendritic Cell-Derived Exosomes Stimulate Stronger CD8+ CTL Responses and Antitumor Immunity than Tumor Cell-Derived Exosomes

    Institute of Scientific and Technical Information of China (English)

    Siguo Hao; Ou Bai; Jinying Yuan; Mabood Qureshi; Jim Xiang

    2006-01-01

    Exosomes (EXO) derived from dendritic cells (DC) and tumor cells have been used to stimulate antitumor immune responses in animal models and in clinical trials. However, there has been no side-by-side comparison of the stimulatory efficiency of the antitumor immune responses induced by these two commonly used EXO vaccines. In this study, we selected to study the phenotype characteristics of EXO derived from a transfected EG7 tumor cells expressing ovalbumin (OVA) and OVA-pulsed DC by flow cytometry. We compared the stimulatory effect in induction of OVA-specific immune responses between these two types of EXO. We found that OVA protein-pulsed DCovA-derived EXO (EXODC) can more efficiently stimulate naive OVA-specific CD8+ T cell proliferation and differentiation into cytotoxic T lymphocytes in vivo, and induce more efficient antitumor immunity than EG7 tumor cell-derived EXO (EXOEG7). In addition, we elucidated the important role of the host DC in EXO vaccines that the stimulatory effect of EXO is delivered to T cell responses by the host DC. Therefore, DC-derived EXO may represent a more effective EXO-based vaccine in induction of antitumor immunity.

  12. Role of the ABO, Secretor, and Lewis determinants in the primed lymphocyte test.

    Science.gov (United States)

    Singal, D P; Blajchman, M A; Naipaul, N; Joseph, S

    1981-05-01

    The antigenic determinants of the combined ABO, Lewis, and Secretor genes have been detected on the surface of lymphocytes by the lymphocytotoxicity test. We have studied the role of these determinants in the primed lymphocyte test (PLT), and the data demonstrate that Lewis incompatibility causes proliferative responses in PLT. On the other hand, no effects of ABO and Secretor incompatibilities were observed in this test. The frequency of the alloantigen-reactive cells (ARC) responding to Lewis and HLA-DR antigens in PLT was estimated by the limiting dilution analysis. The frequency of ARC to allogeneic Lewis-negative donors, who are positive for the sensitizing HLA-DR antigens ranged between 1:58 and 1:97. The incidence of ARC to Lewis-positive allogeneic donors who did not carry the sensitizing HLA-DR specificity was 1:94 to 1:142. These results demonstrate the presence of lymphocyte clones that are able to respond to antigens of the Lewis system. This study suggests that non-HLA antigens belonging to the Lewis system can cause stimulation of lymphocytes in the PLT test.

  13. Tracking in vivo migration and distribution of antigen-specific cytotoxic T lymphocytes by 5,6-carboxyfluorescein diacetate succinimidyl ester staining during cancer immunotherapy

    Institute of Scientific and Technical Information of China (English)

    XU Wei-li; LI Suo-lin; WEN Ming; WEN Jun-ye; HAN Jie; ZHANG Hong-zhen; GAO Fei

    2013-01-01

    Background Killing of targeted tumors during adoptive cell transfer therapy is associated with cytotoxic T lymphocyte (CTL) numbers,immunophenotype,tumor-specificity,and in vivo residence time,migration,and distribution.Therefore,tracing in vivo persistence,migration,and distribution of CTLs is important for cancer immunotherapy.Methods Optimal staining concentration for CTL proliferation was determined by cell counting kit-8 (CCK-8) assay and killing efficiencies of CTLs or carboxyfluorescein diacetate succinimidyl ester (CFSE)-labeled melanoma antigen-specific cytotoxic T lymphocytes (CFSE-CTLs) for malignant melanoma cells in vitro were compared.Additionally,CFSE-CTLs were intravenously transfused to mice receiving B16 melanoma,and their residence time,migration,and distribution in vivo were observed by measuring fluorescence intensities of CFSE-CTLs per gram of tissue (%FI/g) in various tissues and analyzing tumor/non-tumor (T/NT) values.Anti-tumor effects of transferred CTLs and correlation between %FI/g and D-value of tumor size were analyzed.Results Five-micromolar CFSE was optimal for labeling CTLs with minimal cytotoxicity.No significant difference occurred between CTLs and CFSE-CTLs for tumor cell killing (P=0.849) or interleukin-2 (P=0.318) and interferon-y (P=0.201)levels.Distribution of CTLs in vivo varied with time.A negative correlation between %FI/g in tumors and D-value of tumor sizes by Spearman correlation analysis was observed.CTLs were recruited to and killed tumors from 6 hours to 3 days after cell infusion.CTLs were observed up to three weeks later in the tumor,liver,kidneys,and spleen; this was related to the abundant blood supply or the nature of immune organs.Conclusions CCK-8 assay is a novel method to select optimal CFSE staining concentrations.Fluorescence intensity of transferred CTLs reflects their killing efficiency of tumors.CFSE fluorescent markers can trace in vivo CTL persistence,migration,and distribution because of its

  14. Lenalidomide and Chronic Lymphocytic Leukemia

    Directory of Open Access Journals (Sweden)

    Ana Pilar González-Rodríguez

    2013-01-01

    Full Text Available Lenalidomide is an oral immunomodulatory drug used in multiple myeloma and myelodysplastic syndrome and most recently it has shown to be effective in the treatment of various lymphoproliferative disorders such as chronic lymphocytic leukemia (CLL and non-Hodgkin lymphoma. The mechanism of action of lenalidomide varies depending on the pathology, and in the case of CLL, it appears to primarily act by restoring the damaged mechanisms of tumour immunosurveillance. This review discusses the potential mechanism of action and efficacy of lenalidomide, alone or in combination, in treatment of CLL and its toxic effects such as tumor lysis syndrome (TLS and tumor flare reaction (TFR, that make its management different from other hematologic malignancies.

  15. 煤制油项目饱和蒸汽余热利用研究%Study on Saturated Steam Waste Heat Utilization of CTL Project

    Institute of Scientific and Technical Information of China (English)

    周翔

    2016-01-01

    Take a CTL project as an example, the paper mainly introduces the installed capacity and type of saturated steam turbogenerator by calculating plantwide steam balance, and presents the issues needing attention in type selection of saturated steam turbogenerator by analyzing variables like exhaust pressure, exhaust dryness, relative internal efficiency and so on of saturated steam turbogenerator.%以某干粉气化间接合成油项目为例,通过全厂蒸汽平衡的计算,确定了饱和余热发电机组的装机容量和形式;分析了饱和余热发电汽轮机的排汽压力、排汽干度、相对内效率等变量,提出了饱和余热汽轮机选型中应注意的问题。

  16. Rapid progressing allele HLA-B35 Px restricted anti-HIV-1 CD8+ T cells recognize vestigial CTL epitopes.

    Directory of Open Access Journals (Sweden)

    Christian B Willberg

    Full Text Available The HLA-B*35-Px allele has been associated with rapid disease progression in HIV-1 infection, in contrast to the HLA-B*35-Py allele.Immune responses to two HLA-B*35 restricted HIV-1 specific CTL epitopes and their variants were followed longitudinally during early HIV-1 infection in 16 HLA-B*35+ individuals. Subjects expressing HLA-B*35-Px alleles showed no difference in response to the consensus epitopes compared to individuals with HLA-B*35-Py alleles. Surprisingly, all the HLA-B*35-Px+ individuals responded to epitope-variants even in the absence of a consensus response. Sequencing of the viral population revealed no evidence of variant virus in any of the individuals.This demonstrates a novel phenomenon that distinguishes individuals with the HLA-B*35-Px rapid progressing allele and those with the HLA-B*35-Py slower progressing allele.

  17. Recognition of melanoma-derived antigens by CTL: possible mechanisms involved in down-regulating anti-tumor T-cell reactivity

    DEFF Research Database (Denmark)

    Rivoltini, L; Loftus, D J; Squarcina, P

    1998-01-01

    Several T cell-recognized epitopes presented by melanoma cells have been identified recently. Despite the large array of epitopes potentially available for clinical use, it is still unclear which of these antigens could be effective in mediating anti-tumor responses when used as a vaccine...... (detected as increased antigen-specific CTL activity in peripheral blood) was obtained by vaccinating HLA-A2.1+ melanoma patients with the immunodominant epitope (residues 27-35) of the differentiation antigen MART-1, but this immunization was not accompanied by a significant clinical response. To implement...... immunotherapeuties capable of significantly impacting disease outcome, it is necessary to identify the potential mechanisms responsible for the failure of some antigens to mediate significant anti-tumor responses in vivo. In the case of the MART-1(27-35) epitope, we hypothesize that one of these mechanisms may...

  18. Therapy of established B16-F10 melanoma tumors by a single vaccination of CTL/T helper peptides in VacciMax®

    Directory of Open Access Journals (Sweden)

    Korets-Smith Ella

    2007-04-01

    Full Text Available Abstract Background Melanoma tumors are known to express antigens that usually induce weak immune responses of short duration. Expression of both tumor-associated antigens p53 and TRP2 by melanoma cells raises the possibility of simultaneously targeting more than one antigen in a therapeutic vaccine. In this report, we show that VacciMax® (VM, a novel liposome-based vaccine delivery platform, can increase the immunogenicity of melanoma associated antigens, resulting in tumor elimination. Methods C57BL/6 mice bearing B16-F10 melanoma tumors were vaccinated subcutaneously 6 days post tumor implantation with a mixture of synthetic peptides (modified p53: 232–240, TRP-2: 181–188 and PADRE and CpG. Tumor growth was monitored and antigen-specific splenocyte responses were assayed by ELISPOT. Results Vaccine formulated in VM increased the number of both TRP2- and p53-specific IFN-γ producing splenocytes following a single vaccination. Vaccine formulated without VM resulted only in enhanced IFN-γ producing splenocytes to one CTL epitopes (TRP2:180–188, suggesting that VM overcomes antigen dominance and enhances immunogenicity of multiple epitopes. Vaccination of mice bearing 6-day old B16-F10 tumors with both TRP2 and p53-peptides formulated in VM successfully eradicated tumors in all mice. A control vaccine which contained all ingredients except liposomes resulted in eradication of tumors in no more than 20% of mice. Conclusion A single administration of VM is capable of inducing an effective CTL response to multiple tumor-associated antigens. The responses generated were able to reject 6-day old B16-F10 tumors.

  19. Increased PRAME-specific CTL killing of acute myeloid leukemia cells by either a novel histone deacetylase inhibitor chidamide alone or combined treatment with decitabine.

    Directory of Open Access Journals (Sweden)

    Yushi Yao

    Full Text Available As one of the best known cancer testis antigens, PRAME is overexpressed exclusively in germ line tissues such as the testis as well as in a variety of solid and hematological malignant cells including acute myeloid leukemia. Therefore, PRAME has been recognized as a promising target for both active and adoptive anti-leukemia immunotherapy. However, in most patients with PRAME-expressing acute myeloid leukemia, PRAME antigen-specific CD8(+ CTL response are either undetectable or too weak to exert immune surveillance presumably due to the inadequate PRAME antigen expression and PRAME-specific antigen presentation by leukemia cells. In this study, we observed remarkably increased PRAME mRNA expression in human acute myeloid leukemia cell lines and primary acute myeloid leukemia cells after treatment with a novel subtype-selective histone deacetylase inhibitor chidamide in vitro. PRAME expression was further enhanced in acute myeloid leukemia cell lines after combined treatment with chidamide and DNA demethylating agent decitabine. Pre-treatment of an HLA-A0201(+ acute myeloid leukemia cell line THP-1 with chidamide and/or decitabine increased sensitivity to purified CTLs that recognize PRAME(100-108 or PRAME(300-309 peptide presented by HLA-A0201. Chidamide-induced epigenetic upregulation of CD86 also contributed to increased cytotoxicity of PRAME antigen-specific CTLs. Our data thus provide a new line of evidence that epigenetic upregulation of cancer testis antigens by a subtype-selective HDAC inhibitor or in combination with hypomethylating agent increases CTL cytotoxicity and may represent a new opportunity in future design of treatment strategy targeting specifically PRAME-expressing acute myeloid leukemia.

  20. HIV-specific cvtotoxic T-lymphocyte responses in patients co-infected with HIV and HCV%艾滋病病毒/丙型肝炎病毒双重感染者的艾滋病病毒特异性CTL应答

    Institute of Scientific and Technical Information of China (English)

    康文臻; 黄德东; 李媛; 庄严; 翟嵩; 赵柯; 谢玉梅; 孙永涛

    2011-01-01

    Objective To investigate the features of HIV-spocific cytotoxic T-lymphocyte (CTL) responses in patients co-infected with HIV and HCV in China. Methods The patients were divided into 2 groups, HIV/HCV co-infected group (n=22) and simply HIV infected group (n=20). The HIV-l-specific CTL responses were analyzed in an ELISPOT assay by using peptides spanning the entire HIV-1 Clade B consensus sequence. Comparison was made between the difference of HIV specific CTL of HIV/HCV coinfectied group and simply HIV infected group. Results In comparison of the response between HIV/HCV co-infected and simply HIV infected group, the magnitude and frequency of HIV specific CTL was roughly identical with some difference found at singlepeptide level. In HIV/HCV co-infected group, the magnitude and frequency of HIV specific CTL in Gag was higher than those in HIV group. There was positive linear correlation between the magnitude and the frequency of HIV specific CTL in the two groups.Gag and Nef were the most frequently targeted proteins. There was no significant correlation between the total responses of HIV specific CTL and CD4+T cell count. Conclusion HIV-l-specifie CTL may play an important role in aggravating HIV in patients with HIV/HCV co-infection.%目的 探讨中国艾滋病病毒/丙型肝炎病毒(HIV/HCV)双重感染患者HIV特异性细胞毒性T淋巴细胞(CTL)应答的特征.方法 观察对象为HIV/HCV双重患者、单纯HIV感染者.以HIV-1 B亚型构建的全基因组肽库作为抗原,通过酶联免疫斑点(ELISPOT)法检测HIV/AIDS患者HIV特异性CTL应答.并对HIV/HCV双重感染患者的HIV特异性CTL应答进行相关性分析.结果 HIV/HCV双重感染组和单纯HIV感染组的HIV特异性CTL应答强度与频率在肽段水平存在一定差异,与单纯HIV感染组相比,合并HCV感染组反应增强的肽段主要集中在Gag.两组特异性CTL应答强度与频率均呈明显正相关,且合并HCV感染组较单纯感染组正相关性增强

  1. [Mystery and problems of cloning].

    Science.gov (United States)

    Nikitin, V A

    2010-01-01

    The attention of investigators is attracted to the fact that, in spite of great efforts in mammalian cloning, advances that have been made in this area of research are not great, and cloned animals have developmental pathologies often incompatible with life and/or reproduction ability. It is yet not clear what technical or biological factors underlie this, and how they are connected or interact with each other, which is more realistic strategically. There is a great number of articles dealing with the influence of cloning with the nuclear transfer on genetic and epigenetic reprogramming of donor cells. At the same time we can see the practical absence of analytical investigations concerning the technology of cloning as such, its weak points, and possible sources of cellular trauma in the course of microsurgery of nuclear transfer or twinning. This article discusses step by step several nuclear transfer techniques and the methods of dividing early preimplanted embryos for twinning with the aim to reveal possible sources of cell damage during micromanipulation that may have negative influence on the development of cloned organisms. Several new author's technologies based on the study of cell biophysical characteristics are described, which allow one to avoid cellular trauma during manipulation and minimize the possibility of cell damage at any rate.

  2. [Cloning and law in Hungary].

    Science.gov (United States)

    Julesz, Máté

    2015-03-01

    Reproductive human cloning is prohibited in Hungary, as in many other countries. Therapeutic human cloning is not prohibited, just like in many other countries. Stem cell therapy is also allowed. Article III, paragraph (3) of the Hungarian basic law (constitution) strictly forbids total human cloning. Article 1 of the Additional Protocol to the Oviedo Convention, on the Prohibition of Cloning Human Beings (1998) stipulates that any intervention seeking to create a human being genetically identical to another human being, whether living or dead, is prohibited. In Hungary, according to Article 174 of the Criminal Code, total human cloning constitutes a crime. Article 180, paragraph (3) of the Hungarian Act on Health declares that embryos shall not be brought about for research purposes; research shall be conducted only on embryos brought about for reproductive purposes when this is authorized by the persons entitled to decide upon its disposal, or when the embryo is damaged. Article 180, paragraph (5) of the Hungarian Act on Health stipulates that multiple individuals who genetically conform to one another shall not be brought about. According to Article 181, paragraph (1) of the Hungarian Act on Health, an embryo used for research shall be kept alive for not longer than 14 days, not counting the time it was frozen for storage and the time period of research.

  3. [Evolution and phylogeny of B lymphocytes].

    Science.gov (United States)

    Claudio-Piedras, Fabiola; Lanz-Mendoza, Humberto

    2016-01-01

    B lymphocytes are one of the most important cell types involved in the immune response of mammals. The origin and evolution of this cellular type is unknown, but the B lymphocyte bona fide appeared first in fish. In this review we analize the principal components of the immune response of invertebrates, their phylogenetic distribution and the permancence of some properties that allowed the emergence of the B lymphocyte. We started from the idea that many of the components that characterize the B lymphocyte are found distributed among the invertebrates, however, it is in the B lymphocyte, where all these components that give this type of cell its identity, converged. The actual knowledge we have in regards of the lymphocytes comes, in the most part, from physiological studies in mammals, being the mice the more representative. The origin of the B lymphocyte, its alternative mechanisms for generating receptor diversity, its immune effector response, and the generation of memory, require an evolutionary and multidisiplinary approach for its study.

  4. Vaccination against lymphocytic choriomeningitis virus infection in MHC class II-deficient mice

    DEFF Research Database (Denmark)

    Holst, Peter Johannes; Christensen, Jan Pravsgaard; Thomsen, Allan Randrup

    2011-01-01

    response could be elicited in MHC class II-deficient mice by vaccination with adenovirus encoding lymphocytic choriomeningitis virus (LCMV) glycoprotein tethered to MHC class II-associated invariant chain. Moreover, the response induced conferred significant cytolytic CD8(+) T cell-mediated protection...... against challenge with a high dose of the invasive clone 13 strain of LCMV. In contrast, vaccination with adenovirus encoding unlinked LCMV glycoprotein induced weak virus control in the absence of CD4(+) T cells, and mice may die of increased immunopathology associated with incomplete protection. Acute...

  5. Activation of human T lymphocytes by Leishmania lipophosphoglycan

    DEFF Research Database (Denmark)

    Kemp, M; Theander, T G; Handman, E

    1991-01-01

    This study describes Leishmania antigen-induced activation of lymphocytes isolated from Kenyan donors, previously treated for visceral leishmaniasis, and from Danish and Kenyan controls. Peripheral blood mononuclear cells (PBMC) from cured Kala-Azar patients proliferated and produced Interferon...... 63 failed to activate PBMC from any of the donors tested. These results show that the individuals cured from visceral leishmaniasis had expanded T-cell clones recognizing LPG, conceivably as a result of Leishmania infection. The LPG preparation was without detectable protein contamination. Thus...

  6. Cloning and characterization of exodus, a novel beta-chemokine.

    Science.gov (United States)

    Hromas, R; Gray, P W; Chantry, D; Godiska, R; Krathwohl, M; Fife, K; Bell, G I; Takeda, J; Aronica, S; Gordon, M; Cooper, S; Broxmeyer, H E; Klemsz, M J

    1997-05-01

    Chemokines are a family of related proteins that regulate leukocyte infiltration into inflamed tissue. Some chemokines such as MIP-1 alpha also inhibit hematopoietic progenitor cell proliferation. Recently, three chemokines, MIP-1 alpha, MIP-1 beta, and RANTES, have been found to significantly decrease human immunodeficiency virus production from infected T cells. We report here the cloning and characterization of a novel human chemokine termed Exodus for its chemotactic properties. This novel chemokine is distantly related to other chemokines (28% homology with MIP-1 alpha) and shares several biological activities. Exodus is expressed preferentially in lymphocytes and monocytes, and its expression is markedly upregulated by mediators of inflammation such as tumor necrosis factor or lipopolysaccharide. Purified synthetic Exodus was found to inhibit proliferation of myeloid progenitors in colony formation assays. Exodus also stimulated chemotaxis of peripheral blood mononuclear cells. The sequence homology, expression, and biological activity indicate that Exodus represents a novel divergent beta-chemokine.

  7. The topsy-turvy cloning law.

    Science.gov (United States)

    Brassington, Iain; Oultram, Stuart

    2011-03-01

    In debates about human cloning, a distinction is frequently drawn between therapeutic and reproductive uses of the technology. Naturally enough, this distinction influences the way that the law is framed. The general consensus is that therapeutic cloning is less morally problematic than reproductive cloning--one can hold this position while holding that both are morally unacceptable--and the law frequently leaves the way open for some cloning for the sake of research into new therapeutic techniques while banning it for reproductive purposes. We claim that the position adopted by the law has things the wrong way around: if we accept a moral distinction between therapeutic and reproductive cloning, there are actually more reasons to be morally worried about therapeutic cloning than about reproductive cloning. If cloning is the proper object of legal scrutiny, then, we ought to make sure that we are scrutinising the right kind of clone.

  8. Metabolism of peripheral lymphocytes, interleukin-2-activated lymphocytes and tumor-infiltrating lymphocytes from sup 31 P NMR studies

    Energy Technology Data Exchange (ETDEWEB)

    Kaplan, O.; Cohen, J.S.; Aebersold, P.

    1989-11-20

    {sup 31}O NMR spectra of tumor-infiltrating lymphocytes (TILs) were found to be significantly diefferent form those of normal peripheral lymphocytes. The greatest difference was in the phosphodiester (PDE) region, mainly in the glycerophosphocholine (GPC) signal. Short-term activation of peripheral lymphocytes with interleukin-2 induced a small increase in ATP levels. In all lumphocytes the phosphomonoester (PME) region is dominated by phosphoethanolamine (PE), while there is an unusual absence of phosphocholine (PC). Perfusion of these cells with high concentrations of choline caused only a minimal increase in PC, indicating that choline kinase is not the rate limiting step of lecithin synthesis in lymphocytes. (author). 13 refs.; 3 figs.; 1 tab.

  9. Animal cloning: advances and prospects

    Directory of Open Access Journals (Sweden)

    Chuaire Lilian

    2004-05-01

    Full Text Available Few recent advances have revolutionized the developmental biology as the animal cloning has. Since the birth of Dolly, the sheep, in 1996, which was the first derived clone of a mature animal, a new scientific era began. It has been characterized by growing demystification that differentiated cells are unalterable entities in its nuclear organization and chromatin structure, and by a better understanding of the mechanisms that regulate the development. Throughout this paper, we will review some of the achievements and limitations of the techniques used, both in therapeutic and in the reproductive cloning, as well as the perspectives that its application allows to glimpse within a close future. At the same time, we will point out some considerations regarding the ethical debate that surrounds such a controversial issue.

  10. Public perceptions of animal cloning

    DEFF Research Database (Denmark)

    Jelsøe, Erling; Vincentsen, Ulla; Andersen, Ida-Elisabeth

    What was from the outset meant to be a survey testing predefined categories of ethical positions related to new biotechnologies with animal cloning as an example was subsequently developed into a process of broader involvement of groups of citizens in the issue. The survey was conducted at meetings...... in four different cities in Denmark. The participants were introduced to animal cloning and after that they filled out the questionnaire. Finally, the issue was discussed in focus groups. The process as a whole was run in a dialogue oriented way. Through the information they received in combination...... with reflecting on the survey questions the participants were well prepared for discussions in the focus groups. This approach made it possible, on the one hand to get a measure of the citizen's perceptions of the ethical aspects of animal cloning, but also to go deeper into their own thoughts of the issue...

  11. 树突状细胞诱导的细胞毒性T细胞对黑色素瘤的抑制作用%Inhibitory effects of cytotoxic T lymphocyte induced by dendritic cells on melanoma

    Institute of Scientific and Technical Information of China (English)

    张燕; 谭延伟; 蔡建辉; 胡玲玲

    2014-01-01

    Objective To investigate the role of CD4 + CD25 + regulatory T cell (Treg) in cytotoxic T lymphocyte (CTL) inhibiting B16 melanoma and the mechanism.Methods B16 melanoma cells were injected into subcutaneous tissue under the left shoulder of the C57BL/6 mouse to make the melanoma animal models.Then the animal models were irradiated by 60Co-ray on the total body in order to get a no-marrow lymphocyte deletion (NMLD) state.Eighty C57BL/6 mice were divided into 4 groups:3 experimental groups and a control group,with 20 mice each.In vitro T cells were induced into CTL with tumor specificity.CTL were injected back into the mouse body through the vein in the tail.The changes of tumor volume and tumor weight were observed.Results The tumor inhibition rate in the three experiment groups was 90.89%,92.52% and 60.02% respectively,which was significantly higher than that in the control group (P < 0.05).A significant tumor inhibition was observed in groups A and B with a deletion of CD4 + CD25 + Tregs,in comparison with that in group C with no deletion of Tregs (P < 0.05).But no significant difference was found between groups A and B with deletion of Tregs at different time points (P > 0.05).Conclusion The deletion of CD4 + CD25 + Treg could significantly increase the tumor inhibition effect of antigen-specific CTL in melanoma.%目的 探讨CD4+ CD25+ Treg细胞对树突状细胞(DC)诱导的细胞毒性T细胞(CTL)对B16黑色素瘤抑瘤作用的影响及其机制.方法 制备C57BL/6小鼠B16黑色素瘤动物模型,行全身放疗达非髓性淋巴细胞删除(NMLD)状态.随机将80只小鼠分为4组.A组:在DC与T细胞共同培养前应用免疫磁珠法(MACS)删除CD4+ CD25+ Treg细胞;B组:DC与T细胞共同培养后删除CD4+ CD25+ Treg细胞;C组:DC与T细胞共同培养时不删除CD4+ CD25+ Treg细胞.D组:对照组.体外应用DC诱导T细胞产生效应细胞CTL,将CTL经鼠尾静脉回输小鼠体内,观察对黑色素瘤的抑瘤效果.结果 A

  12. HLA class I-restricted cytotoxic T-cell epitopes of the respiratory syncytial virus fusion protein

    NARCIS (Netherlands)

    A.H. Brandenburg (Afke); L. de Waal (Leon); H.H. Timmerman (Helga); P. Hoogerhout (Peter); R.L. de Swart (Rik); A.D.M.E. Osterhaus (Albert)

    2000-01-01

    textabstractVirus-specific cytotoxic T lymphocytes (CTL) play a major role in the clearance of respiratory syncytial virus (RSV) infection. We have generated cytotoxic T-cell clones (TCC) from two infants who had just recovered from severe RSV infection. These TCC were

  13. Monoclonal antibodies in chronic lymphocytic leukemia.

    Science.gov (United States)

    Ferrajoli, Alessandra; Faderl, Stefan; Keating, Michael J

    2006-09-01

    Multiple options are now available for the treatment of chronic lymphocytic leukemia. Over the last 10 years, monoclonal antibodies have become an integral part of the management of this disease. Alemtuzumab has received approval for use in patients with fludarabine-refractory chronic lymphocytic leukemia. Rituximab has been investigated extensively in chronic lymphocytic leukemia both as a single agent and in combination with chemotherapy and other monoclonal antibodies. Epratuzumab and lumiliximab are newer monoclonal antibodies in the early phase of clinical development. This article will review the monoclonal antibodies more commonly used to treat chronic lymphocytic leukemia, the results obtained with monoclonal antibodies as single agents and in combination with chemotherapy, and other biological agents and newer compounds undergoing clinical trials.

  14. Chronic lymphocytic leukemia: present status.

    Science.gov (United States)

    Montserrat, E; Rozman, C

    1995-03-01

    Chronic lymphocytic leukemia (CLL) is the form of leukemia which occurs most frequently in Western countries. Its etiology is unknown, and no relationship with viruses or genes has been demonstrated. Epidemiological data suggest that genetic and ambiental factors might be of some significance. Clinical features of CLL are due to the accumulation of leukemic cells in bone marrow and lymphoid organs as well as the immune disturbances that accompany the disease. The prognosis of patients with CLL varies. Treatment is usually indicated by the risk of the individual patient, which is clearly reflected by the stage of the disease. In the early stage (Binet A, Rai O) it is reasonable to defer therapy until disease progression is observed. By contrast, because their median survival is less than five years, patients with more advanced stages require therapy. For almost 50 years, no major advances in the management of CLL, which has revolved around the use of alkylating agents, have been made. In recent years, the therapeutic approach in patients with CLL has changed as a result of the introduction of combination chemotherapy regimens and, in particular, purine analogues. The latter are already the treatment of choice for patients not responding to standard therapies, and their role as front-line therapy is being investigated. Bone marrow transplants are also being increasingly used. It is to be hoped that in years to come the outcome of patients with CLL will be improved by these advances.

  15. Lymphocyte subsets in pediatric migraine.

    Science.gov (United States)

    Cseh, Aron; Farkas, Kristof Mark; Derzbach, Laszlo; Muller, Katalin; Vasarhelyi, Barna; Szalay, Balazs; Treszl, Andras; Farkas, Viktor

    2013-07-01

    Aseptic inflammation due to activated immune cells has been implicated in the pathomechanism of migraine. We measured the prevalence of regulatory T cells (Tregs), along with that of CD4(+)/CD8(+) lymphocytes and their Th1/Th2 commitment in pediatric migraine. Children and adolescents suffering from migraine without aura, migraine with aura and hemiplegic migraine ictally (n = 53, 27, and 20, respectively), also interictally (n = 33) were recruited and compared to 24 healthy children. Our results indicated comparable prevalence of Tregs, CD4(+) and Th1/Th2 committed cells. CD8(+) prevalence was lower, and CD4(+)/CD8(+) ratio was higher in ictal phase irrespective of the subtype of migraine. No association between CD8(+) prevalence and gender, body weight, disease onset and attack duration in migraine subtypes was found. CD8(+) prevalence was normal in patients in interictal phase. These results suggest the absence of major systemic alteration of adaptive immunity in children and adolescents suffering from migraine; however, a transient decrease of CD8(+) prevalence during the ictal phase was detected irrespective of the subtype of migraine.

  16. Obinutuzumab for chronic lymphocytic leukemia.

    Science.gov (United States)

    Rioufol, Catherine; Salles, Gilles

    2014-10-01

    Chronic lymphocytic leukemia (CLL) is a frequent hematological malignancy that is incurable using standard approaches. Two anti-CD20 monoclonal antibodies (mAb), rituximab and ofatumumab, have been approved for CLL treatment. A new glycoengineered type II humanized anti-CD20 mAb, obinutuzumab (GA101), has been developed and demonstrates increased activity against B-cell malignancies by inducing direct cell death and better antibody-dependent cellular cytotoxicity. In a recent randomized Phase III study in patients with newly diagnosed CLL and coexisting conditions, obinutuzumab plus chlorambucil demonstrated significant improvement in progression-free survival and several other outcome parameters, in contrast to rituximab plus chlorambucil. Grade 3-4 infusion-related reactions and neutropenia occurred more frequently in patients who received obinutuzumab compared with those who received rituximab; however, the rate of serious infections was similar. Obinutuzumab represents a promising new option for patients with CLL and must be investigated with other chemotherapy regimens or with new targeted agents.

  17. How T lymphocytes see antigen

    Science.gov (United States)

    Chakraborty, Arup K.

    2009-03-01

    Complex organisms, like humans, have an adaptive immune system that enables us to do battle with diverse pathogens. This flexible system can also go awry, and many diseases are the direct consequence of the adaptive immune system failing to discriminate between markers of self and non-self. The orchestrators of adaptive immunity are a class of cells called T lymphocytes (T cells). T cells recognize minute numbers of molecular signatures of pathogens, and T cell recognition of these molecular markers of non-self is both specific and degenerate. The specific (yet, cross-reactive), diverse, and self-tolerant T cell repertoire is designed in the thymus. I will describe how an approach that brings together theoretical and computational studies (rooted in statistical physics) with experiments (carried out by key collaborators) has allowed us to shed light on the mechanistic principles underlying how T cells respond to pathogens in a digital fashion (``on'' or ``off''), and how this molecular machinery coupled with frustration (a la spin glasses) plays a key role in designing the special properties of the T cell repertoire during development in the thymus.

  18. Bilateral dacryoadenitis complicated by lymphocytic hypophysitis.

    Science.gov (United States)

    Baoke, Hou; Shihui, Wei; Maonian, Zhang; Zhaohui, Li; Zhitong, Zou; Zhigang, Song; Yan, Hei

    2009-09-01

    Three patients developed dacryoadenitis (DA) or lymphocytic pneumonitis before the diagnosis of lymphocytic hypophysitis (LyH). There were two previous reports of concurrence of DA and LyH. Our patients add support to the idea that DA and LyH are manifestations of a systemic autoimmune disease. We suggest that the discovery of DA should prompt imaging and endocrine investigation of LyH.

  19. Expression of tenascin in lymphocytic autoimmune thyroiditis.

    OpenAIRE

    Back, W; Heubner, C; Winter, J.; Bleyl, U

    1997-01-01

    AIMS: To study the distribution of tenascin by immunocytochemistry in autoimmune diseases of the thyroid. METHODS: Thyroids from patients with inflammatory lesions of the thyroid (lymphocytic thyroiditis Hashimoto, Grave's disease, thyroiditis DeQuervain) were studied by immunocytochemistry using antibodies against tenascin, collagen III, and collagen IV. RESULTS: In autoimmune lymphocytic thyroiditis Hashimoto there was a characteristic corona-like staining pattern of tenascin around all act...

  20. Quantum cloning machines and the applications

    Energy Technology Data Exchange (ETDEWEB)

    Fan, Heng, E-mail: hfan@iphy.ac.cn [Beijing National Laboratory for Condensed Matter Physics, Institute of Physics, Chinese Academy of Sciences, Beijing 100190 (China); Collaborative Innovation Center of Quantum Matter, Beijing 100190 (China); Wang, Yi-Nan; Jing, Li [School of Physics, Peking University, Beijing 100871 (China); Yue, Jie-Dong [Beijing National Laboratory for Condensed Matter Physics, Institute of Physics, Chinese Academy of Sciences, Beijing 100190 (China); Shi, Han-Duo; Zhang, Yong-Liang; Mu, Liang-Zhu [School of Physics, Peking University, Beijing 100871 (China)

    2014-11-20

    No-cloning theorem is fundamental for quantum mechanics and for quantum information science that states an unknown quantum state cannot be cloned perfectly. However, we can try to clone a quantum state approximately with the optimal fidelity, or instead, we can try to clone it perfectly with the largest probability. Thus various quantum cloning machines have been designed for different quantum information protocols. Specifically, quantum cloning machines can be designed to analyze the security of quantum key distribution protocols such as BB84 protocol, six-state protocol, B92 protocol and their generalizations. Some well-known quantum cloning machines include universal quantum cloning machine, phase-covariant cloning machine, the asymmetric quantum cloning machine and the probabilistic quantum cloning machine. In the past years, much progress has been made in studying quantum cloning machines and their applications and implementations, both theoretically and experimentally. In this review, we will give a complete description of those important developments about quantum cloning and some related topics. On the other hand, this review is self-consistent, and in particular, we try to present some detailed formulations so that further study can be taken based on those results.

  1. T-cell receptor Vβ repertoire of CD8+ T-lymphocyte subpopulations in cutaneous leishmaniasis patients from the state of Rio de Janeiro, Brazil

    Directory of Open Access Journals (Sweden)

    Raquel Ferraz

    2015-08-01

    Full Text Available In human cutaneous leishmaniasis (CL, the immune response is mainly mediated by T-cells. The role of CD8+ T-lymphocytes, which are related to healing or deleterious functions, in affecting clinical outcome is controversial. The aim of this study was to evaluate T-cell receptor diversity in late-differentiated effector (LDE and memory CD8+ T-cell subsets in order to create a profile of specific clones engaged in deleterious or protective CL immune responses. Healthy subjects, patients with active disease (PAD and clinically cured patients were enrolled in the study. Total CD8+ T-lymphocytes showed a disturbance in the expression of the Vβ2, Vβ9, Vβ13.2, Vβ18 and Vβ23 families. The analyses of CD8+T-lymphocyte subsets showed high frequencies of LDE CD8+T-lymphocytes expressing Vβ12 and Vβ22 in PAD, as well as effector-memory CD8+ T-cells expressing Vβ22. We also observed low frequencies of effector and central-memory CD8+ T-cells expressing Vβ2 in PAD, which correlated with a greater lesion size. Particular Vβ expansions point to CD8+ T-cell clones that are selected during CL immune responses, suggesting that CD8+ T-lymphocytes expressing Vβ12 or Vβ22 are involved in a LDE response and that Vβ2 contractions in memory CD8+T-cells are associated with larger lesions.

  2. T-cell receptor Vβ repertoire of CD8+ T-lymphocyte subpopulations in cutaneous leishmaniasis patients from the state of Rio de Janeiro, Brazil

    Science.gov (United States)

    Ferraz, Raquel; Cunha, Clarissa Ferreira; Pimentel, Maria Inês; Lyra, Marcelo Rosandiski; Schubach, Armando Oliveira; de Mendonça, Sérgio Coutinho Furtado; Da-Cruz, Alda Maria; Bertho, Alvaro Luiz

    2015-01-01

    In human cutaneous leishmaniasis (CL), the immune response is mainly mediated by T-cells. The role of CD8+ T-lymphocytes, which are related to healing or deleterious functions, in affecting clinical outcome is controversial. The aim of this study was to evaluate T-cell receptor diversity in late-differentiated effector (LDE) and memory CD8+ T-cell subsets in order to create a profile of specific clones engaged in deleterious or protective CL immune responses. Healthy subjects, patients with active disease (PAD) and clinically cured patients were enrolled in the study. Total CD8+ T-lymphocytes showed a disturbance in the expression of the Vβ2, Vβ9, Vβ13.2, Vβ18 and Vβ23 families. The analyses of CD8+T-lymphocyte subsets showed high frequencies of LDE CD8+T-lymphocytes expressing Vβ12 and Vβ22 in PAD, as well as effector-memory CD8+ T-cells expressing Vβ22. We also observed low frequencies of effector and central-memory CD8+ T-cells expressing Vβ2 in PAD, which correlated with a greater lesion size. Particular Vβ expansions point to CD8+ T-cell clones that are selected during CL immune responses, suggesting that CD8+ T-lymphocytes expressing Vβ12 or Vβ22 are involved in a LDE response and that Vβ2 contractions in memory CD8+T-cells are associated with larger lesions. PMID:26107186

  3. Sequence conservation of subdominant HLA-A2-binding CTL epitopes in HIV-1 clinical isolates and CD8+ T-lymphocyte cross-recognition may explain the immune reaction in infected individuals

    DEFF Research Database (Denmark)

    Thorn, Mette; Tang, Sheila; Therrien, Dominic;

    2007-01-01

    genome sequenced HIV-1 from 11 HLA-A2(+) patients to examine the sequence variation of these natural epitopes and compared them with the patient's CD8(+) T-cell recall response. Often the epitope was conserved but only a few patients showed a CD8(+) T-cell recall response. This infrequent targeting may...... be explained by immune subdominance. CD8(+) T-cell recall response to a natural epitope could be measured despite sequence differences in the patient's virus. T-cell cross-reaction between such variants could be demonstrated in HLA-A2 transgenic mice. Nine infrequently targeted but conserved or cross...

  4. Lymphocyte migration into syngeneic implanted lymph nodes

    Energy Technology Data Exchange (ETDEWEB)

    Gordeeva, M.S.

    1986-03-01

    To judge the functional activity of lymphocytes of an implanted lymph node (LN), the proliferative response of lymphocytes of the implanted organ in the blast-transformation reaction in vitro and their ability to induce a local graft versus host reaction (GVHR) were determined. The lymphocyte suspension for labeling with /sup 51/Cr was obtained from peripheral LN in different situations from syngeneic mice. The resulting lymphocyte suspension was labeled with a solution of sodium chromate-/sup 51/Cr in a concentration of 20-40 microCi/100.10/sup 6/ cells in 1 ml for 40 min at 37/sup 0/C. The proliferative activity of a suspension of lymphocytes was estimated as incorporation of /sup 3/H-thymidine into DNA during incubation of the cell suspension for 3 days. Data on migration of /sup 51/Cr-labeled cells and the results of the morphological observations revealed marked ability of lymphocytes of the peripheral pool to colonize the regenerating stroma.

  5. Prenatal ontogeny of lymphocyte subpopulations in pigs.

    Science.gov (United States)

    Sinkora, M; Sinkora, J; Reháková, Z; Splíchal, I; Yang, H; Parkhouse, R M; Trebichavsk, I

    1998-12-01

    Although porcine lymphocytes have been classified into numerous subpopulations in postnatal animals, little is known about the ontogeny of these complex cell subsets. Using double- and triple-colour flow cytometry (FCM), we investigated the surface phenotype of fetal lymphoid cells in the thymus, cord blood, spleen and mesenteric lymph nodes at different stages of gestation. It was found that the major lymphocyte subpopulations started to appear at the beginning of the second third of the gestation period, with B cells being the earliest lymphocyte subpopulation to appear in the periphery. The T-cell receptor (TCR) gamma delta+ cells were the earliest detectable T-cell subset, developing first in the thymus and subsequently arriving in the periphery. Later in ontogeny, however, the number of TCRalpha beta+ lymphocytes rapidly increased, becoming the predominant T cells both in the thymus and in the periphery. Cells with the phenotype of adult natural killer cells were also identified in pig fetuses, though their nature and functional roles remain to be investigated. In addition, CD2 was expressed on most B cells whilst very few CD4+ TCRalpha beta+ cells or CD2+ TCRgamma delta+ cells expressed CD8, suggesting that the expression of CD2 and CD8 may reflect the functional status of the cells in postnatal animals. Taken together, this study has provided a systematic analysis of fetal porcine lymphocyte subpopulations and may provide the base for studies to establish the physiological roles of these lymphocyte subsets.

  6. SHARPIN Regulates Uropod Detachment in Migrating Lymphocytes

    Directory of Open Access Journals (Sweden)

    Jeroen Pouwels

    2013-11-01

    Full Text Available SHARPIN-deficient mice display a multiorgan chronic inflammatory phenotype suggestive of altered leukocyte migration. We therefore studied the role of SHARPIN in lymphocyte adhesion, polarization, and migration. We found that SHARPIN localizes to the trailing edges (uropods of both mouse and human chemokine-activated lymphocytes migrating on intercellular adhesion molecule-1 (ICAM-1, which is one of the major endothelial ligands for migrating leukocytes. SHARPIN-deficient cells adhere better to ICAM-1 and show highly elongated tails when migrating. The increased tail lifetime in SHARPIN-deficient lymphocytes decreases the migration velocity. The adhesion, migration, and uropod defects in SHARPIN-deficient lymphocytes were rescued by reintroducing SHARPIN into the cells. Mechanistically, we show that SHARPIN interacts directly with lymphocyte-function-associated antigen-1 (LFA-1, a leukocyte counterreceptor for ICAM-1, and inhibits the expression of intermediate and high-affinity forms of LFA-1. Thus, SHARPIN controls lymphocyte migration by endogenously maintaining LFA-1 inactive to allow adjustable detachment of the uropods in polarized cells.

  7. Mammalian cloning: possibilities and threats.

    Science.gov (United States)

    Mitalipov, S M; Wolf, D P

    2000-10-01

    The cloning of mammals originated with the production of limited numbers of genetically identical offspring by blastomere separation or embryo splitting. In the past few years, remarkable progress has been reported in cloning by nuclear transfer (NT) with donor nuclei recovered from embryonic, fetal or adult cells. Factors that contribute to the successful reprogramming of the transferred nucleus and the normal term development of the newly reconstructed embryo include the cell cycle stage of both the donor nucleus and recipient cytoplast, the timing of fusion and cytoplast activation, and the source of donor nuclei. The possibility of producing live offspring by somatic cell NT carries potential applications in animal husbandry, biotechnology, transgenic and pharmaceutical production, biomedical research, and the preservation of endangered species. However, the low efficiencies of cloning by NT coupled with high embryonic, fetal and neonatal losses may restrict immediate commercial applications in agriculture. These limitations notwithstanding, the greatest benefits and practical implications of this new technology could be in transplantation medicine and therapeutic cloning.

  8. Operads, clones, and distributive laws

    CERN Document Server

    Curien, Pierre-Louis

    2012-01-01

    We show how non-symmetric operads (or multicategories), symmetric operads, and clones, arise from three suitable monads on Cat, each extending to a (pseudo-)monad on the bicategory of categories and profunctors. We also explain how other previous categorical analyses of operads (via Day's tensor products, or via analytical functors) fit with the profunctor approach.

  9. Positional cloning of deafness genes

    NARCIS (Netherlands)

    Kremer, H.; Cremers, F.P.M.

    2009-01-01

    The identification of the majority of the known causative genes involved in nonsyndromic sensorineural hearing loss (NSHL) started with linkage analysis as part of a positional cloning procedure. The human and mouse genome projects in combination with technical developments on genotyping, transcript

  10. EasyClone-MarkerFree

    DEFF Research Database (Denmark)

    Fabre, Mathew Malcolm Jessop; Jakociunas, Tadas; Stovicek, Vratislav

    2016-01-01

    Clone-MarkerFree. The integration of linearized expression cassettes into defined genomic loci is facilitated by CRISPR/Cas9. Cas9 is recruited to the chromosomal location by specific guide RNAs (gRNAs) expressed from a set of gRNA helper vectors. Using our genome engineering vector suite, single and triple insertions are obtained...

  11. Clone Poems and the Microcomputer.

    Science.gov (United States)

    Irizarry, Estelle

    1989-01-01

    Describes how students can use the computer to study and create clone poems (altering original Spanish-language poems by substituting words and expressions), and how students can gain a deeper appreciation of the original poem's poetic structure and semantics. (CB)

  12. Graph rewriting with polarized cloning

    CERN Document Server

    Duval, Dominique; Prost, Frédéric

    2009-01-01

    We tackle the problem of graph transformation with a particular focus on node cloning. We propose a graph rewriting framework where nodes can be cloned zero, one or more times. A node can be cloned together with all its incident edges, with only the outgoing edges, with only the incoming edges or without any of the incident edges. We thus subsume previous works such as the sesqui-pushout, the heterogeneous pushout and the adaptive star grammars approaches. A rule is defined as a span $\\spa{\\grpol{L}}{l}{\\grpol{K}}{r}{R}$ where the right-hand side $R$ is a multigraph, the left-hand side $\\grpol{L}$ and the interface $\\grpol{K}$ are polarized multigraphs. A polarized multigraph is a multigraph endowed with some cloning annotations on nodes and edges. We introduce the notion of polarized multigraphs and define a rewriting step as pushback followed by a pushout in the same way as in the sesqui-pushout approach.

  13. Entospletinib and Obinutuzumab in Treating Patients With Relapsed Chronic Lymphocytic Leukemia, Small Lymphocytic Lymphoma, or Non-Hodgkin Lymphoma

    Science.gov (United States)

    2017-07-24

    Anemia; B-Cell Prolymphocytic Leukemia; Fatigue; Fever; Grade 1 Follicular Lymphoma; Grade 2 Follicular Lymphoma; Grade 3a Follicular Lymphoma; Hairy Cell Leukemia; Lymphadenopathy; Lymphocytosis; Lymphoplasmacytic Lymphoma; Mantle Cell Lymphoma; Marginal Zone Lymphoma; Night Sweats; Recurrent Chronic Lymphocytic Leukemia; Recurrent Small Lymphocytic Lymphoma; Refractory Chronic Lymphocytic Leukemia; Refractory Small Lymphocytic Lymphoma; Richter Syndrome; Splenomegaly; Thrombocytopenia; Weight Loss

  14. Human reproductive cloning: a conflict of liberties.

    Science.gov (United States)

    Havstad, Joyce C

    2010-02-01

    Proponents of human reproductive cloning do not dispute that cloning may lead to violations of clones' right to self-determination, or that these violations could cause psychological harms. But they proceed with their endorsement of human reproductive cloning by dismissing these psychological harms, mainly in two ways. The first tactic is to point out that to commit the genetic fallacy is indeed a mistake; the second is to invoke Parfit's non-identity problem. The argument of this paper is that neither approach succeeds in removing our moral responsibility to consider and to prevent psychological harms to cloned individuals. In fact, the same commitment to personal liberty that generates the right to reproduce by means of cloning also creates the need to limit that right appropriately. Discussion of human reproductive cloning ought to involve a careful and balanced consideration of both the relevant aspects of personal liberty - the parents' right to reproductive freedom and the cloned child's right to self-determination.

  15. Economical Phase-Covariant Cloning of Qudits

    CERN Document Server

    Buscemi, F; Macchiavello, C; Buscemi, Francesco; Ariano, Giacomo Mauro D'; Macchiavello, Chiara

    2004-01-01

    We derive the optimal $N\\to M$ phase-covariant quantum cloning for equatorial states in dimension $d$ with $M=kd+N$, $k$ integer. The cloning maps are optimal for both global and single-qudit fidelity. The map is achieved by an ``economical'' cloning machine, which works without ancilla. The connection between optimal phase-covariant cloning and optimal multi-phase estimation is finally established.

  16. Dealing with clones in the tracking

    CERN Document Server

    Rodrigues, E

    2006-01-01

    The note describes the way clone tracks are found and eliminated in the LHCb tracking. Both the "clone killer" algorithm and the related "clone finder" tool are presented. The performance of the algorithm as it is used at present in Brunel is also discussed.

  17. Monoclonal Antibody Therapy in Treating Patients With Chronic Lymphocytic Leukemia, Lymphocytic Lymphoma, Acute Lymphoblastic Leukemia, or Acute Myeloid Leukemia

    Science.gov (United States)

    2013-06-03

    Extranodal Marginal Zone B-cell Lymphoma of Mucosa-associated Lymphoid Tissue; Nodal Marginal Zone B-cell Lymphoma; Noncontiguous Stage II Marginal Zone Lymphoma; Noncontiguous Stage II Small Lymphocytic Lymphoma; Recurrent Adult Acute Lymphoblastic Leukemia; Recurrent Adult Acute Myeloid Leukemia; Recurrent Marginal Zone Lymphoma; Recurrent Small Lymphocytic Lymphoma; Refractory Chronic Lymphocytic Leukemia; Splenic Marginal Zone Lymphoma; Stage III Marginal Zone Lymphoma; Stage III Small Lymphocytic Lymphoma; Stage IV Marginal Zone Lymphoma; Stage IV Small Lymphocytic Lymphoma

  18. A single-copy galK promoter cloning vector suitable for cloning strong promoters

    DEFF Research Database (Denmark)

    Dandanell, Gert; Court, Donald L.; Hammer, Karin

    1986-01-01

    We report the construction of lambda galK promoter cloning vectors for cloning and characterization of strong promoters. This phage, which contains a unique HindIII cloning site, was applied to the cloning and analysis of transcription initiations of the regulatory region of the deo-operon of...

  19. A single-copy galK promoter cloning vector suitable for cloning strong promoters

    DEFF Research Database (Denmark)

    Dandanell, Gert; Court, Donald L.; Hammer, Karin

    1986-01-01

    We report the construction of lambda galK promoter cloning vectors for cloning and characterization of strong promoters. This phage, which contains a unique HindIII cloning site, was applied to the cloning and analysis of transcription initiations of the regulatory region of the deo-operon of...

  20. Promiscuous prediction and conservancy analysis of CTL binding epitopes of HCV 3a viral proteome from Punjab Pakistan: an In Silico Approach

    Directory of Open Access Journals (Sweden)

    Idrees Muhammad

    2011-02-01

    Full Text Available Abstract Background HCV is a positive sense RNA virus affecting approximately 180 million people world wide and about 10 million Pakistani populations. HCV genotype 3a is the major cause of infection in Pakistani population. One of the major problems of HCV infection especially in the developing countries that limits the limits the antiviral therapy is the long term treatment, high dosage and side effects. Studies of antigenic epitopes of viral sequences of a specific origin can provide an effective way to overcome the mutation rate and to determine the promiscuous binders to be used for epitope based subunit vaccine design. An in silico approach was applied for the analysis of entire HCV proteome of Pakistani origin, aimed to identify the viral epitopes and their conservancy in HCV genotypes 1, 2 and 3 of diverse origin. Results Immunoinformatic tools were applied for the predictive analysis of HCV 3a antigenic epitopes of Pakistani origin. All the predicted epitopes were then subjected for their conservancy analysis in HCV genotypes 1, 2 and 3 of diverse origin (worldwide. Using freely available web servers, 150 MHC II epitopes were predicted as promiscuous binders against 51 subjected alleles. E2 protein represented the 20% of all the predicted MHC II epitopes. 75.33% of the predicted MHC II epitopes were (77-100% conserve in genotype 3; 47.33% and 40.66% in genotype 1 and 2 respectively. 69 MHC I epitopes were predicted as promiscuous binders against 47 subjected alleles. NS4b represented 26% of all the MHC I predicted epitopes. Significantly higher epitope conservancy was represented by genotype 3 i.e. 78.26% and 21.05% for genotype 1 and 2. Conclusions The study revealed comprehensive catalogue of potential HCV derived CTL epitopes from viral proteome of Pakistan origin. A considerable number of predicted epitopes were found to be conserved in different HCV genotype. However, the number of conserved epitopes in HCV genotype 3 was

  1. Novel immunodominant peptide presentation strategy: a featured HLA-A*2402-restricted cytotoxic T-lymphocyte epitope stabilized by intrachain hydrogen bonds from severe acute respiratory syndrome coronavirus nucleocapsid protein.

    Science.gov (United States)

    Liu, Jun; Wu, Peng; Gao, Feng; Qi, Jianxun; Kawana-Tachikawa, Ai; Xie, Jing; Vavricka, Christopher J; Iwamoto, Aikichi; Li, Taisheng; Gao, George F

    2010-11-01

    Antigenic peptides recognized by virus-specific cytotoxic T lymphocytes (CTLs) are presented by major histocompatibility complex (MHC; or human leukocyte antigen [HLA] in humans) molecules, and the peptide selection and presentation strategy of the host has been studied to guide our understanding of cellular immunity and vaccine development. Here, a severe acute respiratory syndrome coronavirus (SARS-CoV) nucleocapsid (N) protein-derived CTL epitope, N1 (QFKDNVILL), restricted by HLA-A*2402 was identified by a series of in vitro studies, including a computer-assisted algorithm for prediction, stabilization of the peptide by co-refolding with HLA-A*2402 heavy chain and β(2)-microglobulin (β(2)m), and T2-A24 cell binding. Consequently, the antigenicity of the peptide was confirmed by enzyme-linked immunospot (ELISPOT), proliferation assays, and HLA-peptide complex tetramer staining using peripheral blood mononuclear cells (PBMCs) from donors who had recovered from SARS donors. Furthermore, the crystal structure of HLA-A*2402 complexed with peptide N1 was determined, and the featured peptide was characterized with two unexpected intrachain hydrogen bonds which augment the central residues to bulge out of the binding groove. This may contribute to the T-cell receptor (TCR) interaction, showing a host immunodominant peptide presentation strategy. Meanwhile, a rapid and efficient strategy is presented for the determination of naturally presented CTL epitopes in the context of given HLA alleles of interest from long immunogenic overlapping peptides.

  2. The immune system strikes back: cellular immune responses against indoleamine 2,3-dioxygenase

    DEFF Research Database (Denmark)

    Sørensen, Rikke Baek; Berge-Hansen, Linda; Junker, Niels

    2009-01-01

    lymphocytes (CTL) from the peripheral blood of cancer patients were cloned and expanded. The functional capacity of the established CTL clones was examined by chrome release assays. The study unveiled spontaneous cytotoxic T-cell reactivity against IDO in peripheral blood as well as in the tumor...... microenvironment of different cancer patients. We demonstrate that these IDO reactive T cells are indeed peptide specific, cytotoxic effector cells. Hence, IDO reactive T cells are able to recognize and kill tumor cells including directly isolated AML blasts as well as IDO-expressing dendritic cells, i.e. one...

  3. Selective effects of alpha interferon on human T-lymphocyte subsets during mixed lymphocyte cultures

    DEFF Research Database (Denmark)

    Hokland, M; Hokland, P; Heron, I

    1983-01-01

    Mixed lymphocyte reaction (MLR) cultures of human lymphocyte subsets with or without the addition of physiological doses of human alpha interferon (IFN-alpha) were compared with respect to surface marker phenotypes and proliferative capacities of the responder cells. A selective depression on the T...

  4. Cytotoxic T cell response against the chimeric ETV6-AML1 protein in childhood acute lymphoblastic leukemia.

    Science.gov (United States)

    Yotnda, P; Garcia, F; Peuchmaur, M; Grandchamp, B; Duval, M; Lemonnier, F; Vilmer, E; Langlade-Demoyen, P

    1998-07-15

    Cytotoxic T lymphocytes (CTL) are potent effector cells that could provide long term antitumor immunity if induced by appropriate vaccines. CTL recognize 8-14 amino acid-long peptides processed intracellularly and presented by MHC class I molecules. A well-characterized example of a potential tumor antigen in childhood pre-B Acute Lymphoblastic Leukemia (ALL) results from the chromosomal translocation 12;21 leading to the fusion of the ETV6 and AML1 genes. This translocation is observed in > 25% of ALL-patients. In this study, we have examined whether the chimeric ETV6-AML1 protein could serve as a tumor specific antigen for CTL in HLA-A2.1 individuals. We have identified a nonapeptide (RIAECILGM), encoded by the fusion region of the ETV6-AML1 protein, that binds to HLA-A2.1 molecules and induces specific primary CTL in peripheral blood lymphocytes from healthy donors. These CTL specifically lysed HLA-A2.1 tumor cells endogeneously expressing the ETV6-AML fusion protein. CTL with similar functional capacities were found with high frequencies and cloned from one patient's bone marrow indicating that ETV6-AML1-specific anti-ALL CTL are, at least in some patients, spontaneously stimulated and might participate to host antileukemia defense.

  5. Cloning of aminopeptidase Npromoter and its activity in hematopoietic cell and different tumor cell lines

    Institute of Scientific and Technical Information of China (English)

    2001-01-01

    Aminopeptidase N (APN) promoter region was cloned and sequenced from peripheral blood mononuclear cells. The recombinant reporter construct containing the promoter and luciferase gene, designated pXP1-APNLuc, was introduced into myeloblastic cell line, T lymphocyte cell line and various tumor cell lines. Luciferase assay showed that APN upstream promoter is myeloid-specific for high expression in myeloblastic cell line and much lower expres sion in T lymphocyte cell line. The promoter activity was relatively high in lung adenoma cell line compared with other tumor cell lines including hepatoma cell line, tong cancer cell line and esophageal cancer cell line in which the promoter activity significantly diminished or was almost undetectable. The characteristics of APN promoter may pro vide a new strategy for specific myeloprotection while tumor patients are being treated with chemotherapy and/or radio therapy.

  6. Characterization of the atypical lymphocytes in African swine fever

    Directory of Open Access Journals (Sweden)

    Z. A. Karalyan

    2016-07-01

    Full Text Available Aim: Atypical lymphocytes usually described as lymphocytes with altered shape, increased DNA amount, and larger size. For analysis of cause of genesis and source of atypical lymphocytes during African swine fever virus (ASFV infection, bone marrow, peripheral blood, and in vitro model were investigated. Materials and Methods: Atypical lymphocytes under the influence of ASFV were studied for morphologic, cytophotometric, and membrane surface marker characteristics and were used in vivo and in vitro models. Results: This study indicated the increased size, high metabolic activity, and the presence of additional DNA amount in atypical lymphocytes caused by ASFV infection. Furthermore, in atypical lymphocytes, nuclear-cytoplasmic ratio usually decreased, compared to normal lymphocytes. In morphology, they looking like lymphocytes transformed into blasts by exposure to mitogens or antigens in vitro. They vary in morphologic detail, but most of them are CD2 positive. Conclusions: Our data suggest that atypical lymphocytes may represent an unusual and specific cellular response to ASFV infection.

  7. Characterization of the atypical lymphocytes in African swine fever

    Science.gov (United States)

    Karalyan, Z. A.; Ter-Pogossyan, Z. R.; Abroyan, L. O.; Hakobyan, L. H.; Avetisyan, A. S.; Karalyan, N. Yu; Karalova, E. M.

    2016-01-01

    Aim: Atypical lymphocytes usually described as lymphocytes with altered shape, increased DNA amount, and larger size. For analysis of cause of genesis and source of atypical lymphocytes during African swine fever virus (ASFV) infection, bone marrow, peripheral blood, and in vitro model were investigated. Materials and Methods: Atypical lymphocytes under the influence of ASFV were studied for morphologic, cytophotometric, and membrane surface marker characteristics and were used in vivo and in vitro models. Results: This study indicated the increased size, high metabolic activity, and the presence of additional DNA amount in atypical lymphocytes caused by ASFV infection. Furthermore, in atypical lymphocytes, nuclear-cytoplasmic ratio usually decreased, compared to normal lymphocytes. In morphology, they looking like lymphocytes transformed into blasts by exposure to mitogens or antigens in vitro. They vary in morphologic detail, but most of them are CD2 positive. Conclusions: Our data suggest that atypical lymphocytes may represent an unusual and specific cellular response to ASFV infection. PMID:27536044

  8. Flow cytometric analysis of lymphocytes and lymphocyte subpopulations in induced sputum from patients with asthma

    Directory of Open Access Journals (Sweden)

    Yutaro Shiota

    2000-01-01

    Full Text Available Study objectives were to compare the numbers of lymphocytes and lymphocyte subpopulations in induced sputum from asthmatic patients and from healthy subjects, and to determine the effect of inhaled anti-asthmatic steroid therapy on these cell numbers. Hypertonic saline inhalation was used to non-invasively induce sputum samples in 34 patients with bronchial asthma and 21 healthy subjects. The sputum samples were reduced with dithioerythritol and absolute numbers of lymphocytes and lymphocyte subpopulations were assessed by direct immunofluorescence and flow cytometry. To assess the effect of beclomethasone dipropionate (BDP on induced sputum, numbers of lymphocytes and lymphocyte subpopulations in sputum also were evaluated after 4 weeks of BDP inhalation treatment in seven asthmatic patients. An adequate sample was obtained in 85.3% of patients with asthma and in 79.2% of the healthy subjects. Induced sputum from patients with asthma had increased numbers of lymphocytes (P = 0.009; CD4+ cells (P = 0.044; CD4+ cells-bearing interleukin-2 receptor (CD25; P = 0.016; and CD4+ cells bearing human histocompatibility leukocyte antigen (HLA-DR (P = 0.033. CD8+ cells were not increased in asthmatic patients. In patients treated with inhaled steroids, numbers of lymphocytes, CD4+ cells, CD25-bearing CD4+ cells and HLA-DR-bearing CD4+ cells in sputum decreased from pretreatment numbers (P = 0.016, 0.002, 0.003 and 0.002, respectively. Analysis of lymphocytes in induced sputum by flow cytometry is useful in assessing bronchial inflammation, and activated CD4+ lymphocytes may play a key role in the pathogenesis of airway inflammation in bronchial asthma.

  9. Antiviral Effect of Interferon Lambda Against Lymphocytic Choriomeningitis Virus.

    Science.gov (United States)

    Lukacikova, Lubomira; Oveckova, Ingrid; Betakova, Tatiana; Laposova, Katarina; Polcicova, Katarina; Pastorekova, Silvia; Pastorek, Jaromir; Tomaskova, Jana

    2015-07-01

    Lambda interferons inhibit replication of many viruses, but their role in the inhibition of lymphocytic choriomeningitis virus (LCMV) infection remains unclear. In this study, we examined the antiviral effects of interferon (IFN)-λ2 and IFN-λ3 against LCMV in A549 cells. We found that IFN-λ2 is a more potent inhibitor of LCMV strain MX compared with IFN-λ3, whereas both cytokines have similar antiviral effects against an immunosuppressive variant of LCMV, clone-13. We also demonstrated that the antiviral activity of IFN-λ2 is more effective if it is delivered early rather than after establishment of a long-term infection, suggesting that virus replication is only partially responsive to the cytokine. In agreement with this observation, we showed that LCMV infection significantly reduces IFNLR1 mRNA expression in infected cells. In addition, LCMV infection, to some extent, compromises the signal transduction pathway of IFN-λ2. This implies that IFN receptors as well as their downstream signaling components could be selectively targeted either directly by LCMV proteins or indirectly by cellular factor(s) that are induced or activated by LCMV infection.

  10. Therapeutic and reproductive cloning: a critique.

    Science.gov (United States)

    Bowring, Finn

    2004-01-01

    This article is a critical examination of the science and ethics of human cloning. It summarises the key scientific milestones in the development of nuclear transplantation, explains the importance of cloning to research into the medical potential of embryonic stem cells, and discusses the well-worn distinction between 'therapeutic' and 'reproductive' cloning. Suggesting that this distinction will be impossible to police, it goes on to consider the ethics of full human cloning. It is concluded that it represents an unacceptable form of parental despotism, and that the genetic engineering and cloning of future human beings will fracture the foundations of modern humanism.

  11. Clone DB: an integrated NCBI resource for clone-associated data.

    Science.gov (United States)

    Schneider, Valerie A; Chen, Hsiu-Chuan; Clausen, Cliff; Meric, Peter A; Zhou, Zhigang; Bouk, Nathan; Husain, Nora; Maglott, Donna R; Church, Deanna M

    2013-01-01

    The National Center for Biotechnology Information (NCBI) Clone DB (http://www.ncbi.nlm.nih.gov/clone/) is an integrated resource providing information about and facilitating access to clones, which serve as valuable research reagents in many fields, including genome sequencing and variation analysis. Clone DB represents an expansion and replacement of the former NCBI Clone Registry and has records for genomic and cell-based libraries and clones representing more than 100 different eukaryotic taxa. Records provide details of library construction, associated sequences, map positions and information about resource distribution. Clone DB is indexed in the NCBI Entrez system and can be queried by fields that include organism, clone name, gene name and sequence identifier. Whenever possible, genomic clones are mapped to reference assemblies and their map positions provided in clone records. Clones mapping to specific genomic regions can also be searched for using the NCBI Clone Finder tool, which accepts queries based on sequence coordinates or features such as gene or transcript names. Clone DB makes reports of library, clone and placement data on its FTP site available for download. With Clone DB, users now have available to them a centralized resource that provides them with the tools they will need to make use of these important research reagents.

  12. Rice's Salt Tolerance Gene Cloned

    Institute of Scientific and Technical Information of China (English)

    2005-01-01

    @@ In cooperation with US colleagues, CAS researchers have made significant progress in their studies into functional genes for key agronomic traits by cloning SKC1, a salt-tolerant functional gene of rice and making clear its biological functions and mechanisms. This pioneering work,which was reported in the Oct. issue of Nature Genetics (37:1141-1146), is believed to hold promise to increase the output of the crop plant in this country.

  13. El envejecimiento de los clones

    OpenAIRE

    Trippi, Victorio S.

    2007-01-01

    El envejecimiento de los clones se observa en plantas que muestran crecimiento definido por un determinismo genético, cuando se multiplican con tejidos que evolucionan hacia el crecimiento reproductivo. Las plantas fuertemente influenciadas por el ambiente, pueden mostrar fenómenos de senescencia cuando la condición de ambiente determina el crecimiento reproductivo. Los cambios asociados con la edad resultan de alteraciones del citoplasma como un tipo de diferenciación cel...

  14. Conotoxins Are Purified and Cloned

    Institute of Scientific and Technical Information of China (English)

    2006-01-01

    @@ A group of CAS scientists have succeeded in purifying many conotoxins and cloning more than 100 new genes from six species of cone snails living in waters off the coast of the South China Sea, paving the way for the development of new drugs to relieve neuropathic pains. The work has been honored with a first prize from the 2005 Awards for S&T Progress in Shanghai.

  15. Cloning expeditions: risky but rewarding.

    Science.gov (United States)

    Lodish, Harvey

    2013-12-01

    In the 1980s, a good part of my laboratory was using the then-new recombinant DNA techniques to clone and characterize many important cell surface membrane proteins: GLUT1 (the red cell glucose transporter) and then GLUT2 and GLUT4, the red cell anion exchange protein (Band 3), asialoglycoprotein receptor subunits, sucrase-isomaltase, the erythropoietin receptor, and two of the subunits of the transforming growth factor β (TGF-β) receptor. These cloned genes opened many new fields of basic research, including membrane insertion and trafficking of transmembrane proteins, signal transduction by many members of the cytokine and TGF-β families of receptors, and the cellular physiology of glucose and anion transport. They also led to many insights into the molecular biology of several cancers, hematopoietic disorders, and diabetes. This work was done by an exceptional group of postdocs and students who took exceptionally large risks in developing and using novel cloning technologies. Unsurprisingly, all have gone on to become leaders in the fields of molecular cell biology and molecular medicine.

  16. Demethylating agent decitabine induces autologous cancer testis antigen specific cytotoxic T lymphocytes in vivo

    Institute of Scientific and Technical Information of China (English)

    ZHOU Ji-hao; YAO Yu-shi; WANG Li-xin; WANG Jia; LI Yong-hui; JIANG Meng-meng; ZHOU Min-hang

    2013-01-01

    Background Cancer testis antigens (CTAs) are a novel group of tumor associated antigens.Demethylating agent decitabine was reported to be able to up-regulate CTAs through its hypomethylation mechanism,thus enhance the immunogenicity of leukemia cells.However,few researches have ever focused on the questions that whether this immunostimulatory effect of decitabine could induce autologous CTA specific cytotoxic T lymphocytes (CTLs) in vivo,and if so,whether this effect contributes to disease control.In this study,we aimed to show that decitabine could induce specific autologous CTLs against some mouse CTAs in leukemia cells in vitro and in vivo.Methods Several mouse CTAs were screened by RT-PCR.CTL specific to one of the CTAs named P1A was detected and sorted by P1A specific dimer by flow cytometry.The activity of specific CTLs was measured by real time RT-PCR.Results We firstly screened expression of some CTAs in mouse leukemia cells before and after decitabine treatment and found that decitabine treatment did up-regulate expression of many CTAs.Then we measured the CTLs' activity specific to a mouse CTA P1A in vivo and showed that this activity increased after decitabine treatment.Finally,we sorted these in vivo induced P1A specific CTLs by flow cytometry and demonstrated their cytotoxicity against decitabine treated leukemia cells.Conclusions Our study showed the autologous immune response induced by decitabine in vivo.And more importantly,we firstly proved that this response may contribute to disease control.We believe that this immunostimulatory effect is another anti-cancer mechanism of decitabine,and this special effect would inspire new applications of decitabine in the field of leukemia treatment in the future.

  17. Structural Basis for Degenerate Recognition of Natural HIV Peptide Variants by Cytotoxic Lymphocytes

    Energy Technology Data Exchange (ETDEWEB)

    Martinez-Hackert,E.; Anikeeva, N.; Kalams, S.; Walker, B.; Hendrickson, W.; Sykulev, Y.

    2006-01-01

    It is well established that even small changes in amino acid side chains of antigenic peptide bound to MHC protein may completely abrogate recognition of the peptide-MHC (pMHC) complex by the T-cell receptor (TCR). Often, however, several non-conservative substitutions in the peptide antigen are accommodated and do not impair its recognition by TCR. For example, a preponderance of natural sequence variants of the HIV p17 Gag-derived peptide SLYNTVATL (SL9) are recognized by cytotoxic T lymphocytes (CTL), which implies that interactions with SL9 variants are degenerate both with respect to the class I MHC molecule and with respect to TCR. Here we study the molecular basis for this degenerate recognition of SL9 variants. We show that several SL9 variants bind comparably well to soluble HLA-A2 and to a particular soluble TCR and that these variants are active in the cognate cytotoxicity assay. Natural SL9 variation is restricted by its context in the HIV p17 matrix protein, and we have used synthetic variants to explore the wider spectrum of recognition. High-resolution crystal structures of seven selected SL9 variants bound to HLA-A2 all have remarkably similar peptide conformations and side-chain dispositions outside sites of substitution. This preservation of the peptide conformation despite epitope variations suggests a mechanism for the observed degeneracy in pMHC recognition by TCR, and may contribute to the persistence of SL9-mediated immune responses in chronically infected individuals.

  18. Dynamics of immune escape during HIV/SIV infection.

    Directory of Open Access Journals (Sweden)

    Christian L Althaus

    Full Text Available Several studies have shown that cytotoxic T lymphocytes (CTLs play an important role in controlling HIV/SIV infection. Notably, the observation of escape mutants suggests a selective pressure induced by the CTL response. However, it remains difficult to assess the definite role of the cellular immune response. We devise a computational model of HIV/SIV infection having a broad cellular immune response targeting different viral epitopes. The CTL clones are stimulated by viral antigen and interact with the virus population through cytotoxic killing of infected cells. Consequently, the virus population reacts through the acquisition of CTL escape mutations. Our model provides realistic virus dynamics and describes several experimental observations. We postulate that inter-clonal competition and immunodominance may be critical factors determining the sequential emergence of escapes. We show that even though the total killing induced by the CTL response can be high, escape rates against a single CTL clone are often slow and difficult to estimate from infrequent sequence measurements. Finally, our simulations show that a higher degree of immunodominance leads to more frequent escape with a reduced control of viral replication but a substantially impaired replicative capacity of the virus. This result suggests two strategies for vaccine design: Vaccines inducing a broad CTL response should decrease the viral load, whereas vaccines stimulating a narrow but dominant CTL response are likely to induce escape but may dramatically reduce the replicative capacity of the virus.

  19. Genetic vaccination against the melanocyte lineage-specific antigen gp100 induces cytotoxic T lymphocyte-mediated tumor protection.

    Science.gov (United States)

    Schreurs, M W; de Boer, A J; Figdor, C G; Adema, G J

    1998-06-15

    Melanocyte lineage-specific antigens, such as gp100, have been shown to induce both cellular and humoral immune responses against melanoma. Therefore, these antigens are potential targets for specific antimelanoma immunotherapy. A novel approach to induce both cellular and humoral immunity is genetic vaccination, the injection of antigen-encoding naked plasmid DNA. In a mouse model, we investigated whether genetic vaccination against the human gp100 antigen results in specific antitumor immunity. The results demonstrate that vaccinated mice were protected against a lethal challenge with syngeneic B16 melanoma-expressing human gp100, but not control-transfected B16. Both cytotoxic T cells and IgG specific for human gp100 could be detected in human gp100-vaccinated mice. However, only adoptive transfer of spleen-derived lymphocytes, not of the serum, isolated from protected mice was able to transfer antitumor immunity to nonvaccinated recipients, indicating that CTLs are the predominant effector cells. CTI, lines generated from human gp100-vaccinated mice specifically recognized human gp100. Interestingly, one of the CTL lines cross-reacted between human and mouse gp100, indicating the recognition of a conserved epitope. However, these CTLs did not appear to be involved in the observed tumor protection. Collectively, our results indicate that genetic vaccination can result in a potent antitumor response in vivo and constitutes a potential immunotherapeutic strategy to fight cancer.

  20. Antigen-specific cytotoxic T lymphocytes target airway CD103+ and CD11b+ dendritic cells to suppress allergic inflammation.

    Science.gov (United States)

    Daniels, N J; Hyde, E; Ghosh, S; Seo, K; Price, K M; Hoshino, K; Kaisho, T; Okada, T; Ronchese, F

    2016-01-01

    Allergic airway inflammation is driven by the recognition of inhaled allergen by T helper type 2 (Th2) cells in the airway and lung. Allergen-specific cytotoxic T lymphocytes (CTLs) can strongly reduce airway inflammation, however, the mechanism of their inhibitory activity is not fully defined. We used mouse models to show that allergen-specific CTLs reduced early cytokine production by Th2 cells in lung, and their subsequent accumulation and production of interleukin (IL)-4 and IL-13. In addition, treatment with specific CTLs also increased the proportion of caspase(+) dendritic cells (DCs) in mediastinal lymph node (MLN), and decreased the numbers of CD103(+) and CD11b(+) DCs in the lung. This decrease required expression of the cytotoxic mediator perforin in CTLs and of the appropriate MHC-antigen ligand on DCs, suggesting that direct CTL-DC contact was necessary. Lastly, lung imaging experiments revealed that in airway-challenged mice XCR1-GFP(+) DCs, corresponding to the CD103(+) DC subset, and XCR1-GFP(-) CD11c(+) cells, which include CD11b(+) DCs and alveolar macrophages, both clustered in the areas surrounding the small airways and were closely associated with allergen-specific CTLs. Thus, allergen-specific CTLs reduce allergic airway inflammation by depleting CD103(+) and CD11b(+) DC populations in the lung, and may constitute a mechanism through which allergic immune responses are regulated.