scholarly journals Identification of an HLA-A*0201-Restricted T-Cell Epitope on the MPT51 Protein, a Major Secreted Protein Derived from Mycobacterium tuberculosis, by MPT51 Overlapping Peptide Screening

2008 ◽  
Vol 76 (4) ◽  
pp. 1565-1571 ◽  
Author(s):  
Taiki Aoshi ◽  
Toshi Nagata ◽  
Mina Suzuki ◽  
Masato Uchijima ◽  
Dai Hashimoto ◽  
...  
Keyword(s):  
T Cells ◽  
Mycobacterium Tuberculosis ◽  
T Cell ◽  
Flow Cytometric Analysis ◽  
Cell Epitope ◽  
T Cell Epitope ◽  
Computer Algorithms ◽  
Dna Encoding ◽  
Color Flow ◽  
Ifn Γ

ABSTRACT CD8+ T cells play a pivotal role in protection against Mycobacterium tuberculosis infection. We identified a novel HLA-A*0201-restricted CD8+ T-cell epitope on a dominant secreted antigen of M. tuberculosis, MPT51, in HLA-A*0201 transgenic HHD mice. HHD mice were immunized with plasmid DNA encoding MPT51 with gene gun bombardment, and gamma interferon (IFN-γ) production by the immune splenocytes was analyzed. In response to overlapping synthetic peptides covering the mature MPT51 sequence, the splenocytes were stimulated to produce IFN-γ by only one peptide, p51-70. Three-color flow cytometric analysis of intracellular IFN-γ and cell surface CD4 and CD8 staining revealed that the MPT51 p51-70 peptide contains an immunodominant CD8+ T-cell epitope. Further analysis using computer algorithms permitted identification of a bona fide T-cell epitope, p53-62. A major histocompatibility complex class I stabilization assay using T2 cells confirmed that this epitope binds to HLA-A*0201. The T cells were capable of lysing MPT51 p53-62 peptide-pulsed T2 cells. In addition, MPT51 p53-62-specific memory CD8+ T cells were found in tuberculin skin test-positive HLA-A*0201+ healthy individuals. Use of this HLA-A*0201-restricted CD8+ T-cell epitope for analysis of the role of MPT51-specific T cells in M. tuberculosis infection and for design of vaccines against tuberculosis is feasible.

scholarly journals Identification of Murine H2-Dd- and H2-Ab-Restricted T-Cell Epitopes on a Novel Protective Antigen, MPT51, of Mycobacterium tuberculosis

2004 ◽  
Vol 72 (7) ◽  
pp. 3829-3837 ◽  
Author(s):  
Mina Suzuki ◽  
Taiki Aoshi ◽  
Toshi Nagata ◽  
Yukio Koide
Keyword(s):  
Mycobacterium Tuberculosis ◽  
T Cell ◽  
Protective Antigen ◽  
Cell Epitope ◽  
Cell Subset ◽  
Cell Phenotype ◽  
Computer Assisted ◽  
T Cell Epitope ◽  
Color Flow ◽  
Ifn Γ

ABSTRACT Both CD4+ type 1 helper T (Th1) cells and CD8+ cytotoxic T lymphocytes (CTL) play pivotal roles in protection against Mycobacterium tuberculosis infection. Here, we identified Th1 and CTL epitopes on a novel protective antigen, MPT51, in BALB/c and C57BL/6 mice. Mice were immunized with plasmid DNA encoding MPT51 by using a gene gun, and gamma interferon (IFN-γ) production from the immune spleen cells was analyzed in response to a synthetic overlapping peptide library covering the mature MPT51 sequence. In BALB/c mice, only one peptide, p21-40, appeared to stimulate the immune splenocytes to produce IFN-γ. Flow cytometric analysis with intracellular IFN-γ and the T-cell phenotype revealed that the p21-40 peptide contains an immunodominant CD8+ T-cell epitope. Further analysis with a computer-assisted algorithm permitted identification of a T-cell epitope, p24-32. In addition, a major histocompatibility complex class I stabilization assay with TAP2-deficient RMA-S cells transfected with Kd, Dd, or Ld indicated that the epitope is presented by Dd. Finally, we proved that the p24-32/Dd complex is recognized by IFN-γ-producing CTL. In C57BL/6 mice, we observed H2-Ab-restricted dominant and subdominant Th1 epitopes by using T-cell subset depletion analysis and three-color flow cytometry. The data obtained are useful for analyzing the role of MPT51-specific T cells in protective immunity and for designing a vaccine against M. tuberculosis infection.

Initial Identification of a Mouse Human Factor IX-Specific CD8+ T-Cell Epitope.

Blood ◽  
2006 ◽  
Vol 108 (11) ◽  
pp. 5490-5490
Author(s):  
Brad E. Hoffman ◽  
Roland W. Herzog
Keyword(s):  
T Cells ◽  
T Cell ◽  
Catalytic Domain ◽  
Cell Epitope ◽  
Cd8 T Cell ◽  
Factor Ix ◽  
T Cell Epitope ◽  
Cell Responses ◽  
Ifn Γ

Abstract A significant complication associated with treatment of inherited protein deficiencies, such as hemophilia B, by gene replacement therapy is the potential for the activation of transgene specific B and T cells to the therapeutic protein, coagulation factor IX (F.IX). In addition to the potential for inhibitor formation as a result of MHC class II antigen presentation (CD4+ T cell-dependent activation of B cells, which may also be observed in conventional protein-based therapy), gene expression may lead to MHC class I presentation of F.IX-derived peptides to CD8+ T cells. Upon in vivo gene transfer, such immune responses to may elicit a cytotoxic T lymphocyte (CTL) response capable of destroying target cells that express the F.IX transgene product. Therefore, to better understand the role of F.IX-specific CD8+ T-cell responses, it is essential that MHC I-restricted CD8 T-cell epitopes be identified. Here, we used a peptide library consisting of 82 individual 15-mer peptides overlapping by ten residues that spans the complete human F.IX (hF.IX) protein to preliminarily identify a specific immunodominate CD8+ T-cell epitope. The peptides were pooled into groups, each containing 8–11 peptides to create a matrix of 18 pools, with each peptide represented in two pools. C3H/HeJ were immunized with 5×1010 vector genomes of E1/E3-deleted adenovirus expressing hF.IX (Ad-hF.IX) via intramuscular injection into the quadriceps. Nine days later, the harvested spleen and popliteal lymph node cells were pooled and evaluated for CD8+ T-cell responses by intracellular cytokine staining for IFN-γ after being stimulated for 5h with peptides or controls. The frequency of IFN-γ producing hF.IX-specific CD8+ T-cells was determined by flow cytometry. While 16 pools from Ad-hF.IX immunized C3H/HeJ mice showed no response above the frequency of mock-stimulated cells, lymphocytes from two overlapping pools demonstrated a ~2.5-fold increase in frequency of CD8+ IFN-γ+ cells. From these results we can conclude that peptide 74 (SGGPHVTEVEGTSFL) contains a CD8+ T cell epitope for C3H/HeJ mice (H-2k haplotype). Furthermore, splenocytes from naive mice failed to respond to any of the peptide pools. The amino acid sequence corresponding to peptide 74 is located within the catalytic domain of hF.IX. This finding is of particular interest, in that, we previously reported a peptide containing the immunodominate CD4+ T-cell epitope in C3H/HeJ is also located within the catalytic domain of hF.IX (Blood 108:408). The definitive identification of hF.IX-specific CD8+ epitopes will facilitate the evaluation of experimental gene therapy strategies in murine models by providing a reagent for in vitro stimulation of F.IX specific CD8+ lymphocytes. For example, we can now determine the efficiency of CD8+ T cell activation as a function of vector, route, and dose following in vivo gene transfer.

scholarly journals Immunization of Volunteers with Salmonella enterica Serovar Typhi Strain Ty21a Elicits the Oligoclonal Expansion of CD8+ T Cells with Predominant Vβ Repertoires

2005 ◽  
Vol 73 (6) ◽  
pp. 3521-3530 ◽  
Author(s):  
Rosângela Salerno-Gonçalves ◽  
Rezwanul Wahid ◽  
Marcelo B. Sztein
Keyword(s):  
T Cells ◽  
T Cell ◽  
Salmonella Enterica ◽  
Flow Cytometric Analysis ◽  
Cell Receptor ◽  
Effector Memory ◽  
Color Flow ◽  
Cell Clones ◽  
Effector Memory Cells ◽  
Ifn Γ

ABSTRACT CD8+ T cells are likely to play an important role in host defense against Salmonella enterica serovar Typhi by several effector mechanisms, including lysis of infected cells (cytotoxicity) and gamma interferon (IFN-γ) secretion. In an effort to better understand these responses, we studied the T-cell receptor (TCR) repertoire of serovar Typhi-specific CD8+ T cells in humans. To this end, we determined the TCR beta chain (Vβ) usage of CD8+ T cells from three volunteers orally immunized with Ty21a typhoid vaccine by flow cytometry using a panel of monoclonal antibodies. Although TCR Vβ usage varied among volunteers, we identified oligoclonal Vβ subset expansions in individual volunteers (Vβ 2, 5.1, 8, 17, and 22 in volunteer 1; Vβ 1, 2, 5.1, 14, 17, and 22 in volunteer 2; and Vβ 3, 8, 14, and 16 in volunteer 3). These subsets were antigen specific, as shown by cytotoxicity and IFN-γ secretion assays on Vβ sorted cells and on T-cell clones derived from these volunteers. Moreover, eight-color flow cytometric analysis showed that these clones exhibited a T effector memory phenotype (i.e., CCR7− CD27− CD45RO+ CD62L−) and coexpressed gut homing molecules (e.g., high levels of integrin α4β7, intermediate levels of CCR9, and low levels of CD103). In conclusion, our results show that long-term T-cell responses to serovar Typhi in Ty21a vaccinees are oligoclonal, involving multiple TCR Vβ families. Moreover, these serovar Typhi-specific CD8+ T cells bearing defined Vβ specificities are phenotypically and functionally consistent with T effector memory cells with preferential gut homing potential.

scholarly journals Swine T-Cells and Specific Antibodies Evoked by Peptide Dendrimers Displaying Different FMDV T-Cell Epitopes

2021 ◽  
Vol 11 ◽  
Author(s):  
Patricia de León ◽  
Rodrigo Cañas-Arranz ◽  
Sira Defaus ◽  
Elisa Torres ◽  
Mar Forner ◽  
...  
Keyword(s):  
T Cells ◽  
T Cell ◽  
B Cell ◽  
Cell Epitope ◽  
T Cell Epitope ◽  
T Cell Epitopes ◽  
B Cell Epitope ◽  
Mouth Disease Virus ◽  
Ifn Γ

Dendrimeric peptide constructs based on a lysine core that comprises both B- and T-cell epitopes of foot-and-mouth disease virus (FMDV) have proven a successful strategy for the development of FMD vaccines. Specifically, B2T dendrimers displaying two copies of the major type O FMDV antigenic B-cell epitope located on the virus capsid [VP1 (140–158)], covalently linked to a heterotypic T-cell epitope from either non-structural protein 3A [3A (21–35)] or 3D [3D (56–70)], named B2T-3A and B2T-3D, respectively, elicit high levels of neutralizing antibodies (nAbs) and IFN-γ-producing cells in pigs. To assess whether the inclusion and orientation of T-3A and T-3D T-cell epitopes in a single molecule could modulate immunogenicity, dendrimers with T epitopes juxtaposed in both possible orientations, i.e., constructs B2TT-3A3D and B2TT-3D3A, were made and tested in pigs. Both dendrimers elicited high nAbs titers that broadly neutralized type O FMDVs, although B2TT-3D3A did not respond to boosting, and induced lower IgGs titers, in particular IgG2, than B2TT-3A3D. Pigs immunized with B2, a control dendrimer displaying two B-cell epitope copies and no T-cell epitope, gave no nABs, confirming T-3A and T-3D as T helper epitopes. The T-3D peptide was found to be an immunodominant, as it produced more IFN-γ expressing cells than T-3A in the in vitro recall assay. Besides, in pigs immunized with the different dendrimeric peptides, CD4+ T-cells were the major subset contributing to IFN-γ expression upon in vitro recall, and depletion of CD4+ cells from PBMCs abolished the production of this cytokine. Most CD4+IFN-γ+ cells showed a memory (CD4+2E3−) and a multifunctional phenotype, as they expressed both IFN-γ and TNF-α, suggesting that the peptides induced a potent Th1 pro-inflammatory response. Furthermore, not only the presence, but also the orientation of T-cell epitopes influenced the T-cell response, as B2TT-3D3A and B2 groups had fewer cells expressing both cytokines. These results help understand how B2T-type dendrimers triggers T-cell populations, highlighting their potential as next-generation FMD vaccines.

scholarly journals Partial Reconstitution of the CD4+-T-Cell Compartment in CD4 Gene Knockout Mice Restores Responses to Tuberculosis DNA Vaccines

2006 ◽  
Vol 74 (5) ◽  
pp. 2751-2759 ◽  
Author(s):  
Sushila D'Souza ◽  
Marta Romano ◽  
Johanna Korf ◽  
Xiao-Ming Wang ◽  
Pierre-Yves Adnet ◽  
...  
Keyword(s):  
T Cells ◽  
T Cell ◽  
Dna Vaccines ◽  
Gene Knockout ◽  
Cell Epitope ◽  
Adaptive Immune Response ◽  
Dna Encoding ◽  
Cell Compartment ◽  
Gene Knockout Mice ◽  
Ifn Γ

ABSTRACT Reactivation tuberculosis (TB) is a serious problem in immunocompromised individuals, especially those with human immunodeficiency virus (HIV) coinfection. The adaptive immune response mediated by CD4+ and CD8+ T cells is known to confer protection against TB. Hence, vaccines against TB are designed to activate these two components of the immune system. Anti-TB DNA vaccines encoding the immunodominant proteins Ag85A, Ag85B, and PstS-3 from Mycobacterium tuberculosis are ineffective in mice lacking CD4+ T cells (CD4−/− mice). In this study, we demonstrate that reconstitution of the T-cell compartment in CD4−/− mice restores vaccine-specific antibody and gamma interferon (IFN-γ) responses to these DNA vaccines. The magnitude of the immune responses correlated with the extent of reconstitution of the CD4+-T-cell compartment. Reconstituted mice vaccinated with DNA encoding PstS-3, known to encode a dominant Db-restricted CD8+-T-cell epitope, displayed CD8+-T-cell responses not observed in CD4−/− mice. M. tuberculosis challenge in reconstituted mice led to the extravasation of IFN-γ-producing CD4+ and CD8+ T cells into lungs, the primary site of bacterial replication. Importantly, a reconstitution of 12 to 15% of the CD4+-T-cell compartment resulted in Ag85B plasmid DNA-mediated protection against a challenge M. tuberculosis infection. Our findings provide evidence that anti-TB DNA vaccines could be effective in immunodeficient individuals after CD4+-T-lymphocyte reconstitution, as may occur following antiretroviral therapy in HIV+ patients.

scholarly journals CD4+ T cells induced by virus-like particles expressing a major T cell epitope down-regulate IL-5 production in an ongoing immune response to Der p 1 independently of IFN-γ production

1999 ◽  
Vol 11 (12) ◽  
pp. 1927-1934 ◽  
Author(s):  
Sandra Hirschberg ◽  
Guy T. Layton ◽  
Stephen J. Harris ◽  
Nigel Savage ◽  
Margaret J. Dallman ◽  
...  
Keyword(s):  
Immune Response ◽  
T Cells ◽  
T Cell ◽  
Cd4 T Cells ◽  
Cell Epitope ◽  
T Cell Epitope ◽  
Virus Like Particles ◽  
Der P 1 ◽  
Ifn Γ

scholarly journals Protective Vaccine Efficacy of the Complete Form of PPE39 Protein from Mycobacterium tuberculosis Beijing/K Strain in Mice

2017 ◽  
Vol 24 (11) ◽  
Author(s):  
Ahreum Kim ◽  
Yun-Gyoung Hur ◽  
Sunwha Gu ◽  
Sang-Nae Cho
Keyword(s):  
T Cells ◽  
Mycobacterium Tuberculosis ◽  
T Cell ◽  
Vaccine Development ◽  
Protective Efficacy ◽  
Interleukin 2 ◽  
T Cell Epitopes ◽  
Content Type ◽  
Complete Form ◽  
Ifn Γ

ABSTRACT The aim of this study was to evaluate the protective efficacy of MTBK_24820, a complete form of PPE39 protein derived from a predominant Beijing/K strain of Mycobacterium tuberculosis in South Korea. Mice were immunized with MTKB_24820, M. bovis Bacilli Calmette-Guérin (BCG), or adjuvant prior to a high-dosed Beijing/K strain aerosol infection. After 4 and 9 weeks, bacterial loads were determined and histopathologic and immunologic features in the lungs and spleens of the M. tuberculosis-infected mice were analyzed. Putative immunogenic T-cell epitopes were examined using synthetic overlapping peptides. Successful immunization of MTBK_24820 in mice was confirmed by increased IgG responses (P < 0.05) and recalled gamma interferon (IFN-γ), interleukin-2 (IL-2), IL-6, and IL-17 responses (P < 0.05 or P < 0.01) to MTBK_24820. After challenge with the Beijing/K strain, an approximately 0.5 to 1.0 log10 reduction in CFU in lungs and fewer lung inflammation lesions were observed in MTBK_24820-immunized mice compared to those for control mice. Moreover, MTBK_24820 immunization elicited significantly higher numbers of CD4+ T cells producing protective cytokines, such as IFN-γ and IL-17, in lungs and spleens (P < 0.01) and CD4+ multifunctional T cells producing IFN-γ, tumor necrosis factor alpha (TNF-α), and/or IL-17 (P < 0.01) than in control mice, suggesting protection comparable to that of BCG against the hypervirulent Beijing/K strain. The dominant immunogenic T-cell epitopes that induced IFN-γ production were at the N terminus (amino acids 85 to 102 and 217 to 234). Its vaccine potential, along with protective immune responses in vivo, may be informative for vaccine development, particularly in regions where the M. tuberculosis Beijing/K-strain is frequently isolated from TB patients.

scholarly journals Expression Cloning of an Immunodominant Family of Mycobacterium tuberculosis Antigens Using Human Cd4+ T Cells

2000 ◽  
Vol 191 (3) ◽  
pp. 551-560 ◽  
Author(s):  
Mark R. Alderson ◽  
Teresa Bement ◽  
Craig H. Day ◽  
Liqing Zhu ◽  
David Molesh ◽  
...  
Keyword(s):  
T Cells ◽  
Mycobacterium Tuberculosis ◽  
T Cell ◽  
Cd4 T Cells ◽  
Mononuclear Cells ◽  
Genomic Library ◽  
Cell Epitope ◽  
Single Amino Acid ◽  
Expression Cloning ◽  
Peripheral Blood Mononuclear

Development of a subunit vaccine for Mycobacterium tuberculosis (Mtb) is likely to be dependent on the identification of T cell antigens that induce strong proliferation and interferon γ production from healthy purified protein derivative (PPD)+ donors. We have developed a sensitive and rapid technique for screening an Mtb genomic library expressed in Escherichia coli using Mtb-specific CD4+ T cells. Using this technique, we identified a family of highly related Mtb antigens. The gene of one family member encodes a 9.9-kD antigen, termed Mtb9.9A. Recombinant Mtb9.9A protein, expressed and purified from E. coli, elicited strong T cell proliferation and IFN-γ production by peripheral blood mononuclear cells from PPD+ but not PPD− individuals. Southern blot analysis and examination of the Mtb genome sequence revealed a family of highly related genes. A T cell line from a PPD+ donor that failed to react with recombinant Mtb9.9A recognized one of the other family members, Mtb9.9C. Synthetic peptides were used to map the T cell epitope recognized by this line, and revealed a single amino acid substitution in this region when compared with Mtb9.9A. The direct identification of antigens using T cells from immune donors will undoubtedly be critical for the development of vaccines to several intracellular pathogens.

scholarly journals Dendritic Cell Mediated Delivery of Plasmid DNA Encoding LAMP/HIV-1 Gag Fusion Immunogen Enhances T Cell Epitope Responses in HLA DR4 Transgenic Mice

PLoS ONE ◽  
2010 ◽  
Vol 5 (1) ◽  
pp. e8574 ◽  
Author(s):  
Gregory G. Simon ◽  
Yongli Hu ◽  
Asif M. Khan ◽  
Jingshi Zhou ◽  
Jerome Salmon ◽  
...  
Keyword(s):  
T Cell ◽  
Dendritic Cell ◽  
Transgenic Mice ◽  
Plasmid Dna ◽  
Cell Epitope ◽  
T Cell Epitope ◽  
Dna Encoding ◽  
Hiv 1
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