scholarly journals Functionally Inert HIV-Specific Cytotoxic T Lymphocytes Do Not Play a Major Role in Chronically Infected Adults and Children

2000 ◽  
Vol 192 (12) ◽  
pp. 1819-1832 ◽  
Author(s):  
Philip J.R. Goulder ◽  
Yanhua Tang ◽  
Christian Brander ◽  
Michael R. Betts ◽  
Marcus Altfeld ◽  
...  
Keyword(s):  
T Lymphocytes ◽  
Hiv Infection ◽  
Cytotoxic T Lymphocytes ◽  
Intracellular Cytokine Staining ◽  
Cell Number ◽  
Lymphocytic Choriomeningitis ◽  
Intracellular Cytokine ◽  
Virus Infections ◽  
Tetramer Binding ◽  
Ifn Γ

The highly sensitive quantitation of virus-specific CD8+ T cells using major histocompatibility complex–peptide tetramer assays has revealed higher levels of cytotoxic T lymphocytes (CTLs) in acute and chronic virus infections than were recognized previously. However, studies in lymphocytic choriomeningitis virus infection have shown that tetramer assays may include measurement of a substantial number of tetramer-binding cells that are functionally inert. Such phenotypically silent CTLs, which lack cytolytic function and do not produce interferon (IFN)-γ, have been hypothesized to explain the persistence of virus in the face of a quantitatively large immune response, particularly when CD4 help is impaired. In this study, we examined the role of functionally inert CTLs in chronic HIV infection. Subjects studied included children and adults (n = 42) whose viral loads ranged from <50 to >100,000 RNA copies/ml plasma. Tetramer assays were compared with three functional assays: enzyme-linked immunospot (Elispot), intracellular cytokine staining, and precursor frequency (limiting dilution assay [LDA]) cytotoxicity assays. Strong positive associations were observed between cell numbers derived by the Elispot and the tetramer assay (r = 0.90). An even stronger association between tetramer-derived numbers and intracellular cytokine staining for IFN-γ was present (r = 0.97). The majority (median 76%) of tetramer-binding cells were consistently detectable via intracellular IFN-γ cytokine staining. Furthermore, modifications to the LDA, using a low input cell number into each well, enabled LDAs to reach equivalence with the other methods of CTL enumeration. These data together show that functionally inert CTLs do not play a significant role in chronic pediatric or adult HIV infection.

scholarly journals CD40 Ligand-Mediated Interactions Are Involved in the Generation of Memory CD8+ Cytotoxic T Lymphocytes (CTL) but Are Not Required for the Maintenance of CTL Memory following Virus Infection

1998 ◽  
Vol 72 (9) ◽  
pp. 7440-7449 ◽  
Author(s):  
Persephone Borrow ◽  
David F. Tough ◽  
Danelle Eto ◽  
Antoinette Tishon ◽  
Iqbal S. Grewal ◽  
...  
Keyword(s):  
T Cell ◽  
T Lymphocytes ◽  
Cytotoxic T Lymphocytes ◽  
Cd40 Ligand ◽  
Lymphocytic Choriomeningitis ◽  
T Cell Response ◽  
Primary Immune Response ◽  
Virus Infections ◽  
Ctl Response ◽  
Deficient Mice

ABSTRACT CD8+ cytotoxic T lymphocytes (CTL) play a key role in the control of many virus infections, and the need for vaccines to elicit strong CD8+ T-cell responses in order to provide optimal protection in such infections is increasingly apparent. However, the mechanisms involved in the induction and maintenance of CD8+ CTL memory are currently poorly understood. In this study, we investigated the involvement of CD40 ligand (CD40L)-mediated interactions in these processes by analyzing the memory CTL response of CD40L-deficient mice following infection with lymphocytic choriomeningitis virus (LCMV). The maintenance of memory CD8+ CTL precursors (CTLp) at stable frequencies over time was not impaired in CD40L-deficient mice. By contrast, the initial generation of memory CTLp was affected. CD40L-deficient mice produced lower levels of CD8+ CTLp during the primary immune response to LCMV than did wild-type controls, despite the fact that the LCMV-specific effector CTL response of CD40L-deficient mice was indistinguishable from that of control animals. The differentiation of naı̈ve CD8+ T cells into effector and memory CTL thus involves pathways that can be discriminated from each other by their requirement for CD40L-mediated interactions. Expression of CD40L by CTLp themselves was not an essential step during their expansion and differentiation from naı̈ve CD8+ cells into memory CTLp; instead, the reduction in memory CTLp generation in CD40L-deficient mice was likely a consequence of defects in the CD4+ T-cell response mounted by these animals. These results thus suggest a previously unappreciated role for CD40L in the generation of CD8+ memory CTLp, the probable nature of which is discussed.

Production of interferon-γ by lung lymphocytes in HIV-infected individuals

1999 ◽  
Vol 276 (2) ◽  
pp. L256-L262 ◽  
Author(s):  
Homer L. Twigg ◽  
Blake A. Spain ◽  
Diaa M. Soliman ◽  
Kenneth Knox ◽  
Richard A. Sidner ◽  
...  
Keyword(s):  
T Lymphocytes ◽  
Hiv Infection ◽  
Cytotoxic T Lymphocytes ◽  
Late Stage ◽  
Opportunistic Infections ◽  
Suppressor Cells ◽  
Interferon Γ ◽  
Lymphocyte Function ◽  
Hiv Disease ◽  
Ifn Γ

A CD8+lymphocytic alveolitis occurs in up to 60% of asymptomatic human immunodeficiency virus (HIV)-infected individuals. Early in HIV infection, lymphocytes consist predominantly of cytotoxic T lymphocytes directed against HIV-infected targets. As HIV disease progresses, they are replaced by CD8+CD57+suppressor cells. Virus-specific cytotoxic T lymphocytes secrete interferon-γ (IFN-γ), an important cytokine in upregulating immune responses, primarily through macrophage activation. We examined the ability of lung and blood lymphocytes from HIV-positive patients at various stages of HIV infection to secrete IFN-γ spontaneously and in response to phytohemagglutinin A. IFN-γ production and secretion were determined with ELISA, Western blot, immunoprecipitation, and Northern blot techniques. Lung lymphocytes from HIV-infected individuals secreted large amounts of IFN-γ. However, this ability was lost in patients with late-stage disease. Correlation between blood and lung lymphocyte IFN-γ secretion was poor, suggesting regional differences in lymphocyte function. These data suggest that lung levels of IFN-γ are high until late in HIV disease. These findings support the concept of administering exogenous IFN-γ to patients with late-stage HIV disease and opportunistic infections.

scholarly journals Identification of HLA-A*02:01-restricted candidate epitopes derived from the non-structural polyprotein 1a of SARS-CoV-2 that may be natural targets of CD8+ T cell recognition in vivo

2020 ◽  
Author(s):  
Akira Takagi ◽  
Masanori Matsui
Keyword(s):  
T Lymphocytes ◽  
Transgenic Mice ◽  
Cytotoxic T Lymphocytes ◽  
Intracellular Cytokine Staining ◽  
Binding Affinities ◽  
Intracellular Cytokine ◽  
Peptide Vaccination ◽  
Ctl Epitopes ◽  
Selection For ◽  
Epitope Selection

COVID-19 vaccines are being rapidly developed and human trials are underway. Almost all of these vaccines have been designed to induce antibodies targeting spike protein of SARS-CoV-2 in expectation of neutralizing activities. However, non-neutralizing antibodies are at risk of causing antibody-dependent enhancement. Further, the longevity of SARS-CoV-2-specific antibodies is very short. Therefore, in addition to antibody-induced vaccines, novel vaccines on the basis of SARS-CoV-2-specific cytotoxic T lymphocytes (CTLs) should be considered in the vaccine development. Here, we attempted to identify HLA-A*02:01-restricted CTL epitopes derived from the non-structural polyprotein 1a of SARS-CoV-2. Eighty-two peptides were firstly predicted as epitope candidates on bioinformatics. Fifty-four in 82 peptides showed high or medium binding affinities to HLA-A*02:01. HLA-A*02:01 transgenic mice were then immunized with each of the 54 peptides encapsulated into liposomes. The intracellular cytokine staining assay revealed that 18 out of 54 peptides were CTL epitopes because of the induction of IFN-γ-producing CD8+ T cells. In the 18 peptides, 10 peptides were chosen for the following analyses because of their high responses. To identify dominant CTL epitopes, mice were immunized with liposomes containing the mixture of the 10 peptides. Some peptides were shown to be statistically predominant over the other peptides. Surprisingly, all mice immunized with the liposomal 10 peptide mixture did not show the same reaction pattern to the 10 peptides. There were three response patterns, suggesting the existence of an immunodominance hierarchy following peptide vaccination, which may provide us more variations in the epitope selection for designing CTL-based COVID-19 vaccines. IMPORTANCE For the development of vaccines based on SARS-CoV-2-specific cytotoxic T lymphocytes (CTLs), we attempted to identify HLA-A*02:01-restricted CTL epitopes derived from the non-structural polyprotein 1a of SARS-CoV-2. Out of 82 peptides predicted on bioinformatics, 54 peptides showed good binding affinities to HLA-A*02:01. Using HLA-A*02:01 transgenic mice, 18 in 54 peptides were found to be CTL epitopes in the intracellular cytokine staining assay. Out of 18 peptides, 10 peptides were chosen for the following analyses because of their high responses. To identify dominant epitopes, mice were immunized with liposomes containing the mixture of the 10 peptides. Some peptides were shown to be statistically predominant. Surprisingly, all immunized mice did not show the same reaction pattern to the 10 peptides. There were three reaction patterns, suggesting the existence of an immunodominance hierarchy following peptide vaccination, which may provide us more variations in the epitope selection for designing CTL-based COVID-19 vaccines.

scholarly journals An immunodominance hierarchy exists in CD8+ T cell responses to HLA-A*02:01-restricted epitopes identified from the non-structural polyprotein 1a of SARS-CoV-2

2020 ◽  
Author(s):  
Akira Takagi ◽  
Masanori Matsui
Keyword(s):  
T Lymphocytes ◽  
Transgenic Mice ◽  
Cytotoxic T Lymphocytes ◽  
Intracellular Cytokine Staining ◽  
Reaction Pattern ◽  
Binding Affinities ◽  
Intracellular Cytokine ◽  
Ctl Epitopes ◽  
Selection For ◽  
Epitope Selection

AbstractCOVID-19 vaccines are being rapidly developed and human trials are underway. Almost all of these vaccines have been designed to induce antibodies targeting spike protein of SARS-CoV-2 in expectation of neutralizing activities. However, non-neutralizing antibodies are at risk of causing antibody-dependent enhancement. Further, the longevity of SARS-CoV-2-specific antibodies is very short. Therefore, in addition to antibody-induced vaccines, novel vaccines on the basis of SARS-CoV-2-specific cytotoxic T lymphocytes (CTLs) should be considered in the vaccine development. Here, we attempted to identify HLA-A*02:01-restricted CTL epitopes derived from the non-structural polyprotein 1a of SARS-CoV-2. Eighty-two peptides were firstly predicted as epitope candidates on bioinformatics. Fifty-four in 82 peptides showed high or medium binding affinities to HLA-A*02:01. HLA-A*02:01 transgenic mice were then immunized with each of the 54 peptides encapsulated into liposomes. The intracellular cytokine staining assay revealed that 18 out of 54 peptides were CTL epitopes because of the induction of IFN-γ-producing CD8+ T cells. In the 18 peptides, 10 peptides were chosen for the following analyses because of their high responses. To identify dominant CTL epitopes, mice were immunized with liposomes containing the mixture of the 10 peptides. Some peptides were shown to be statistically predominant over the other peptides. Surprisingly, all mice immunized with the liposomal 10 peptide mixture did not show the same reaction pattern to the 10 peptides. There were three pattern types that varied sequentially, suggesting the existence of an immunodominance hierarchy, which may provide us more variations in the epitope selection for designing CTL-based COVID-19 vaccines.ImportanceFor the development of vaccines based on SARS-CoV-2-specific cytotoxic T lymphocytes (CTLs), we attempted to identify HLA-A*02:01-restricted CTL epitopes derived from the non-structural polyprotein 1a of SARS-CoV-2. Out of 82 peptides predicted on bioinformatics, 54 peptides showed good binding affinities to HLA-A*02:01. Using HLA-A*02:01 transgenic mice, 18 in 54 peptides were found to be CTL epitopes in the intracellular cytokine staining assay. Out of 18 peptides, 10 peptides were chosen for the following analyses because of their high responses. To identify dominant epitopes, mice were immunized with liposomes containing the mixture of the 10 peptides. Some peptides were shown to be statistically predominant. Surprisingly, all immunized mice did not show the same reaction pattern to the 10 peptides. There were three pattern types that varied sequentially, suggesting the existence of an immunodominance hierarchy, which may provide us more variations in the epitope selection for designing CTL-based COVID-19 vaccines.

scholarly journals Gamma Interferon Expression in CD8+ T Cells Is a Marker for Circulating Cytotoxic T Lymphocytes That Recognize an HLA A2-Restricted Epitope of Human Cytomegalovirus Phosphoprotein pp65

2001 ◽  
Vol 8 (3) ◽  
pp. 628-631 ◽  
Author(s):  
Smita A. Ghanekar ◽  
Laurel E. Nomura ◽  
Maria A. Suni ◽  
Louis J. Picker ◽  
Holden T. Maecker ◽  
...  
Keyword(s):  
T Cells ◽  
T Lymphocytes ◽  
Cytotoxic T Lymphocytes ◽  
Human Cytomegalovirus ◽  
Surrogate Marker ◽  
Gamma Interferon ◽  
Strong Positive Correlation ◽  
Specific Lysis ◽  
Cytokine Flow Cytometry ◽  
Ifn Γ

ABSTRACT Antigen-specific CD8+ T cells with cytotoxic activity are often critical in immune responses to infectious pathogens. To determine whether gamma interferon (IFN-γ) expression is a surrogate marker for cytotoxic T lymphocytes (CTL), human cytomegalovirus-specific CTL responses were correlated with CD8+ T-cell IFN-γ expression determined by cytokine flow cytometry. A strong positive correlation was observed between specific lysis of peptide-pulsed targets in a 51Cr release assay and frequencies of peptide-activated CD8+ T cells expressing IFN-γ at 6 h (r 2 = 0.72) or 7 days (r 2 = 0.91). Enumeration of responding cells expressing perforin, another marker associated with CTL, did not improve this correlation. These results demonstrate that IFN-γ expression can be a functional surrogate for identification of CTL precursor cells.

Induction of Multiple Myeloma-Specific Cytotoxic T Lymphocytes Using HLA-A2.1-Specific CD19 and CD20 Peptides.

Blood ◽  
2004 ◽  
Vol 104 (11) ◽  
pp. 2477-2477
Author(s):  
Jooeun Bae ◽  
Jeff A. Martinson ◽  
Hans G. Klingemann ◽  
Steven Treon ◽  
Kenneth C. Anderson ◽  
...  
Keyword(s):  
Multiple Myeloma ◽  
T Lymphocytes ◽  
Cytotoxic Activity ◽  
Cell Lines ◽  
Cytotoxic T Lymphocytes ◽  
Myeloma Cell ◽  
Myeloma Cells ◽  
Specific Ctls ◽  
Ifn Γ ◽  
Hla A2.1

Abstract We have identified novel CD19 and CD20 antigen-derived HLA-A2.1-specific immunogenic peptides, CD19150–158 (KLMSPKLYV) and CD20188–196 (SLFLGILSV), for generating cytotoxic T lymphocytes (CTLs) against malignant B-cell diseases. Initial testing showed that the CTLs displayed antigen-specific and HLA-A2.1-restriced cytotoxic activity against both Burkitt’s lymphoma and chronic lymphoid leukemia cell lines. The observed cytotoxic activity of the CTLs was shown to be specific to the CD19150–158 or the CD20188–196 peptides. Additionally, the CTLs displayed a distinct phenotype (majority CD69+/CD45RO+) along with a significant (p<0.05) increase in cell proliferation and IFN-γ release following re-stimulation with HLA-A2.1+/CD19+/CD20+ tumor cell lines. Based on emerging information that clonogenic myeloma cells express CD19 and/or CD20, we evaluated the activity of the CD19 and CD20 peptide specific-CTLs against several multiple myeloma cell lines. Five of 10 myeloma cell lines evaluated were HLA-A2.1-positive and expressed both CD19 and CD20 antigens. CD19 peptide specific-CTLs generated from normal donors were able to specifically lyse CD19+/HLA-A2.1+ MM cell lines (30% lysis; 10:1 E:T ratio) but did not lyse CD19−/HLA-A2.1+ or CD19+/HLA-A2.1− cell lines. Similarly, the CD20-specific CTLs generated from normal donors lysed CD20+/HLA-A2.1+ MM cell lines (25% lysis; 10:1 E:T ratio), in a manner restricted to HLA-A2.1 and specific to antigens. We next showed IFN-γ production by the CTLs after exposure to CD19+/HLA-A2.1+ or CD20+/HLA-A2.1+ MM cells. Moreover, we have demonstrated the ability to expand CD20-CTLs under serum-free culture conditions while maintaining their cytotoxic activity (28–49%). In ongoing studies, we are evaluating the ability of CD19- and CD20-specific CTLs to eliminate clonogenic myeloma cells in vitro and in vivo in a SCID mouse model of myeloma. These preclinical studies strongly suggest that immunogenic CD19 and CD20 peptide-based vaccines represent a promising immunotherapeutic approach in myeloma.

The Roles of Wilms’ Tumor Gene Peptide-Specific Cytotoxic T Lymphocytes in Immunologic Pathophysiology of Paroxysmal Nocturnal Hemoglobinuria.

Blood ◽  
2007 ◽  
Vol 110 (11) ◽  
pp. 3676-3676
Author(s):  
Kazuhiko Ikeda ◽  
Hideyoshi Noji ◽  
Masaki Yasukawa ◽  
Akiko Shichishima ◽  
Kazuko Akutsu ◽  
...  
Keyword(s):  
Flow Cytometry ◽  
T Lymphocytes ◽  
Cytotoxic T Lymphocytes ◽  
Colony Formation ◽  
Paroxysmal Nocturnal Hemoglobinuria ◽  
Wilms Tumor ◽  
Wilms Tumor Gene ◽  
Ifn Γ ◽  
Tumor Gene ◽  
Pnh Clone

Abstract It is unclear how a paroxysmal nocturnal hemoglobinuria (PNH) clone expands and bone marrow failure (BMF) occurs in PNH patients, although an immunologic mechanism by human leukocyte antigen (HLA)-restricted cytotoxic T lymphocytes (CTLs) has been suggested. It has been also reported that immunization with HLA-binding peptides of Wilms’ tumor gene (WT1) in hematopoietic cells induces a WT1 peptide-specific CTL response, and WT1 RNA is highly expressed in BM mononuclear cells (MNCs) in PNH patients (Shichishima T et al., Blood, 2002). In this study, to clarify some roles of WT1 peptide-specific and HLA-restricted CTLs, the frequencies of peripheral blood (PB) WT1 peptide-specific and HLA-A*2402-restricted CTLs by flow cytometric tetramer analysis and WT1 peptide-stimulated interferon (IFN)-γ-producing MNCs by enzyme-linked immunospot assay in 5 PNH patients with the HLA-A*2402 allele were examined. We also investigated cytotoxicity of WT1 peptide-specific and HLA-A*2402-restricted CTL clone (TAK-1) cells on BM MNCs by 51Cr-releasing assay, colony forming-unit granulocyte-macrophage colony formation of CD34+CD59+ and CD34+CD59− cells, and CD59 expression in viable 7AAD−CD34+ cells by flow cytometry in those patients, and expression of IFN-γ in TAK-1 cells by flow cytometry, after co-incubation of BM cells from them with TAK-1 cells. As controls, 8 healthy volunteers (HV) with the HLA-A*2402 allele and 2 PNH patients and HV without the allele were examined. We found that the frequencies of PB WT1 peptide-specific and HLA-A*2402-restricted CD8+ cells (p<0.005) and WT1 peptide-stimulated IFN-γ-producing MNCs (p<0.02) were significantly higher in PNH patients with the HLA-A*2402 allele (0.255 ± 0.164% and 25.2 ± 15.4 / 5 x 105 cells, respectively) than HV with the allele (0.052 ± 0.025% and 6.6 ± 6.8 / 5 x 105 cells, respectively). In PNH patients or HV, TAK-1 cells significantly killed BM MNCs, suppressed colony formations of CD34+CD59+ and CD34+CD59− cells, and expressed IFN-γ in the absence and presence of a WT1 peptide or only in the presence of the peptide, respectively, in an HLA-A*2402-restricted manner. Reduction rates of colony formation of CD34+CD59− cells from the patients with the HLA-A*2402 allele by TAK-1 cells were significantly less than those of CD34+CD59+ cells in PNH patients, in the absence (38.3 ± 23.0% and 59.0 ± 28.0%, respectively, p<0.01) and presence (74.7 ± 12.8% and 90.6 ± 11.1%, respectively, p<0.002) of a WT1 peptide. After co-incubation of BM MNCs from the patients with TAK-1 cells, proportions of viable CD34+CD59− cells from PNH patients significantly increased in the absence (62.87 ± 27.29%; p<0.01) and presence (62.32 ± 25.73%; p<0.01) of a WT1 peptide compared with those of the controls incubated without TAK-1 cells (52.40 ± 24.58%) in an HLA-A*2402-restricted manner. In conclusion, WT1 peptide-specific and HLA-restricted CTLs may play important roles in the expansion of a PNH clone during immunologic selection and in the occurrence of BMF via IFN-γ in PNH.

scholarly journals Lesional CD4+ IFN-γ+ cytotoxic T lymphocytes in IgG4-related dacryoadenitis and sialoadenitis

2016 ◽  
Vol 76 (2) ◽  
pp. 377-385 ◽  
Author(s):  
Takashi Maehara ◽  
Hamid Mattoo ◽  
Miho Ohta ◽  
Vinay S Mahajan ◽  
Masafumi Moriyama ◽  
...  
Keyword(s):  
T Lymphocytes ◽  
Cytotoxic T Lymphocytes ◽  
Transforming Growth Factor ◽  
Large Fraction ◽  
Quantitative Imaging ◽  
Healthy Controls ◽  
Tissue Samples ◽  
Study Gene Expression ◽  
Serum Igg4 ◽  
Ifn Γ

ObjectivesIgG4-related disease (IgG4-RD) is a chronic, systemic, inflammatory condition of unknown aetiology. We have recently described clonally expanded circulating CD4+ cytotoxic T lymphocytes (CTLs) in IgG4-RD that infiltrate affected tissues where they secrete interleukin (IL)-1β and transforming growth factor -β1 (TGF-β1). In this study, we sought to examine the role of CD4+ CTLs in the pathogenesis of IgG4-related dacryoadenitis and sialoadenitis (IgG4-DS) and to determine whether these cells secrete interferon-gamma (IFN-γ) at lesional sites.MethodsSalivary glands of 25 patients with IgG4-DS, 22 patients with Sjögren's syndrome (SS), 12 patients with chronic sialoadenitis (CS) and 12 healthy controls were analysed in this study. Gene expression analysis was performed on submandibular glands (SMGs) from five patients with IgG4-DS, three with CS and three healthy controls. Infiltrating CD4+ CTLs were examined by quantitative multicolour imaging in tissue samples from 20 patients with IgG4-DS, 22 patients with SS, 9 patients with CS and 9 healthy controls.ResultsIn IgG4-DS tissues, nine genes associated with CD4+ CTLs were overexpressed. The expression of granzyme A (GZMA) mRNA was significantly higher in samples from patients with IgG4-RD compared with corresponding tissues from SS and healthy controls. Quantitative imaging showed that infiltrating CD4+ GZMA+ CTLs were more abundant in patients with IgG4-DS than in the other groups. The ratio of CD4+GZMA+ CTLs in SMGs from patients with IgG4-DS correlated with serum IgG4 concentrations and the number of affected organs. A large fraction of CD4+GZMA+ CTLs in SMGs from patients with IgG4-DS secreted IFN-γ.ConclusionsThe pathogenesis of IgG4-DS is associated with tissue infiltration by CD4+GZMA+ CTLs that secrete IFN-γ.

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