scholarly journals Identification of Dominant Optimal HLA-B60- and HLA-B61-Restricted Cytotoxic T-Lymphocyte (CTL) Epitopes: Rapid Characterization of CTL Responses by Enzyme-Linked Immunospot Assay

2000 ◽  
Vol 74 (18) ◽  
pp. 8541-8549 ◽  
Author(s):  
Marcus A. Altfeld ◽  
Alicja Trocha ◽  
Robert L. Eldridge ◽  
Eric S. Rosenberg ◽  
Mary N. Phillips ◽  
...  
Keyword(s):  
T Lymphocyte ◽  
Mononuclear Cells ◽  
Cytotoxic T Lymphocyte ◽  
Hla Class I ◽  
Peripheral Blood Mononuclear ◽  
Antiviral Immune Response ◽  
Ctl Epitopes ◽  
Hla Class I Molecules ◽  
Hiv 1 ◽  
Ctl Responses

ABSTRACT Human immunodeficiency virus type 1 (HIV-1)-specific cytotoxic T-lymphocyte (CTL) responses play a major role in the antiviral immune response, but the relative contribution of CTL responses restricted by different HLA class I molecules is less well defined. HLA-B60 or the related allele B61 is expressed in 10 to 20% of Caucasoid populations and is even more highly prevalent in Asian populations, but yet no CTL epitopes restricted by these alleles have been defined. Here we report the definition of five novel HLA-B60-restricted HIV-1-specific CTL epitopes, using peripheral blood mononuclear cells in enzyme-linked immunospot (Elispot) assays and using CTL clones and lines in cytolytic assays. The dominant HLA-B60-restricted epitope, Nef peptide KEKGGLEGL, was targeted by all eight subjects with B60 and also by both subjects with B61 studied. This study additionally establishes the utility of the Elispot assay as a more rapid and efficient method of defining novel CTL epitopes. This approach will help to define new CTL epitopes that may play an important role in the immune control of HIV-1.

HIV-1-Specific Cytotoxic T Lymphocyte (CTL) Responses Against Immunodominant Optimal Epitopes Slow the Progression of AIDS in China

2008 ◽  
Vol 6 (4) ◽  
pp. 335-350 ◽  
Author(s):  
Song Zhai ◽  
Yan Zhuang ◽  
Yang Song ◽  
Shu Li ◽  
Dedong Huang ◽  
...  
Keyword(s):  
T Lymphocyte ◽  
Cytotoxic T Lymphocyte ◽  
Hiv 1 ◽  
Ctl Responses

scholarly journals Rapid Definition of Five Novel HLA-A∗3002-Restricted Human Immunodeficiency Virus-Specific Cytotoxic T-Lymphocyte Epitopes by Elispot and Intracellular Cytokine Staining Assays

2001 ◽  
Vol 75 (3) ◽  
pp. 1339-1347 ◽  
Author(s):  
Philip J. R. Goulder ◽  
, Marylyn M. Addo ◽  
Marcus A. Altfeld ◽  
Eric S. Rosenberg ◽  
Yanhua Tang ◽  
...  
Keyword(s):  
Human Immunodeficiency Virus ◽  
Mononuclear Cells ◽  
Cytotoxic T Lymphocyte ◽  
Intracellular Cytokine Staining ◽  
Initial Step ◽  
Peripheral Blood Mononuclear ◽  
Hla Restriction ◽  
Immunodeficiency Virus ◽  
Definition Of ◽  
Ctl Responses

ABSTRACT Human immunodeficiency virus (HIV)-specific cytotoxic T lymphocytes (CTL) play a major role in control of viral replication. To understand the contribution of this antiviral response, an initial step is to fully define the specific epitopes targeted by CTL. These studies focused on CTL responses restricted by HLA-A∗3002, one of the HLA-A molecules most prominent in African populations. To avoid the time-consuming effort and expense involved in culturing CTL prior to defining epitopes and restricting alleles, we developed a method combining Elispot assays with intracellular gamma interferon staining of peripheral blood mononuclear cells to first map the optimal epitopes targeted and then define the HLA restriction of novel epitopes. In two A∗3002-positive subjects whose CTL responses were characterized in detail, the strongest response in both cases was to an epitope in p17 Gag, RSLYNTVATLY (residues 76 to 86). Using this method, CTL epitopes for which there were no motif predictions were optimized and the HLA restriction was established within 48 to 72 h of receipt of blood. This simple and convenient approach should prove useful especially in the characterization of CTL responses specific to HIV and other viruses, particularly in localities where performing cytotoxicity assays would be problematic.

scholarly journals HIV-1 Variation Diminishes CD4 T Lymphocyte Recognition

1998 ◽  
Vol 188 (10) ◽  
pp. 1785-1793 ◽  
Author(s):  
Gillian C. Harcourt ◽  
Sarah Garrard ◽  
Miles P. Davenport ◽  
Anne Edwards ◽  
Rodney E. Phillips
Keyword(s):  
T Lymphocyte ◽  
Mononuclear Cells ◽  
Antigenic Variation ◽  
Class Ii ◽  
Hla Class Ii ◽  
Peripheral Blood Mononuclear ◽  
Altered Peptide Ligands ◽  
Peptide Epitopes ◽  
Naturally Occurring ◽  
Hiv 1

Effective long-term antiviral immunity requires specific cytotoxic T lymphocytes and CD4+ T lymphocyte help. Failure of these helper responses can be a principle cause of viral persistence. We sought evidence that variation in HIV-1 CD4+ T helper epitopes might contribute to this phenomenon. To determine this, we assayed fresh peripheral blood mononuclear cells from 43 asymptomatic HIV-1+ patients for proliferative responses to HIV-1 antigens. 12 (28%) showed a positive response, and we went on to map dominant epitopes in two individuals, to p24 Gag restricted by human histocompatibility leukocyte antigen (HLA)-DR1 and to p17 Gag restricted by HLA-DRB52c. Nine naturally occurring variants of the p24 Gag epitope were found in the proviral DNA of the individual in whom this response was detected. All variants bound to HLA-DR1, but three of these peptides failed to stimulate a CD4+ T lymphocyte line which recognized the index sequence. Antigenic variation was also detected in the p17 Gag epitope; a dominant viral variant present in the patient was well recognized by a specific CD4+ T lymphocyte line, whereas several natural mutants were not. Importantly, variants detected at both epitopes also failed to stimulate fresh uncultured cells while index peptide stimulated successfully. These results demonstrate that variant antigens arise in HIV-1+ patients which fail to stimulate the T cell antigen receptor of HLA class II–restricted lymphocytes, although the peptide epitopes are capable of being presented on the cell surface. In HIV-1 infection, naturally occurring HLA class II–restricted altered peptide ligands that fail to stimulate the circulating T lymphocyte repertoire may curtail helper responses at sites where variant viruses predominate.

Cytolytic activity and regulatory functions of inhibitory NK cell receptor–expressing T cells expanded from granulocyte colony-stimulating factor–mobilized peripheral blood mononuclear cells

Blood ◽  
2004 ◽  
Vol 104 (3) ◽  
pp. 768-774 ◽  
Author(s):  
Junji Tanaka ◽  
Tomomi Toubai ◽  
Yutaka Tsutsumi ◽  
Yoko Miura ◽  
Naoko Kato ◽  
...  
Keyword(s):  
T Cells ◽  
Mononuclear Cells ◽  
Hla Class I ◽  
Cell Receptor ◽  
Cytolytic Activity ◽  
Leukemic Cells ◽  
Peripheral Blood Mononuclear ◽  
Hla Class I Molecules ◽  
Blood Mononuclear Cells ◽  
Mobilized Peripheral Blood

AbstractInhibitory natural killer cell receptor (NKR)–expressing cells may induce a graft-versus-leukemia/tumor (GVL/T) effect against leukemic cells and tumor cells that have mismatched or decreased expression of HLA class I molecules and may not cause graft-versus-host disease (GVHD) against host cells that have normal expression of HLA class I molecules. In our study, we were able to expand inhibitory NKR (CD94/NKG2A)–expressing CD8+ T cells from granulocyte colonystimulating factor (G-CSF)–mobilized peripheral blood mononuclear cells (G-PBMCs) by more than 500-fold using stimulation by an anti-CD3 monoclonal antibody with interleukin 15 (IL-15). These expanded and purified CD94-expressing cells attacked various malignant cell lines, including solid cancer cell lines, as well as the patients' leukemic cells but not autologous and allogeneic phytohemagglutinin (PHA) blasts in vitro. Also, these CD94-expressing cells prevented the growth of K562 leukemic cells and CW2 colon cancer cells in NOD/SCID mice in vivo. On the other hand, the CD94-expressing cells have low responsiveness to alloantigen in mixed lymphocyte culture (MLC) and have high transforming growth factor (TGF)–β1– but low IL-2– producing capacity. Therefore, CD94-expressing cells with cytolytic activity against the recipient's leukemic and tumor cells without enhancement of alloresponse might be able to be expanded from donor G-PBMCs.

scholarly journals Detection of equine arteritis virus (EAV)-specific cytotoxic CD8+ T lymphocyte precursors from EAV-infected ponies

2003 ◽  
Vol 84 (10) ◽  
pp. 2745-2753 ◽  
Author(s):  
J. Castillo-Olivares ◽  
J. P. Tearle ◽  
F. Montesso ◽  
D. Westcott ◽  
J. H. Kydd ◽  
...  
Keyword(s):  
T Lymphocyte ◽  
Peripheral Blood ◽  
Mononuclear Cells ◽  
Cytotoxic T Lymphocyte ◽  
Severe Disease ◽  
Systemic Infection ◽  
Neutralizing Antibody ◽  
Equine Arteritis Virus ◽  
Peripheral Blood Mononuclear ◽  
Virus Clearance

Equine arteritis virus (EAV) causes a systemic infection in equids with variable outcome, ranging from subclinical infections to severe disease, and also has the capacity to induce abortion in pregnant mares and persistent infections in stallions. The serum virus-neutralizing antibody response that invariably develops in the infected animal lasts for many months or years and is believed to play an important role in virus clearance. However, very little is known about cellular immunity against EAV because of a lack of methods for evaluating these immune responses. In the present study, we describe methods for detecting cytotoxic T lymphocyte (CTL) precursors in the peripheral blood of EAV-convalescent ponies using a 51Cr release cytolysis assay. Primary equine dermal cells, used as CTL targets, were shown to express MHC I but not MHC II and to retain 51Cr efficiently and support EAV replication. Peripheral blood mononuclear cells (PBMC) collected from EAV-convalescent ponies that had been incubated with or without live EAV were used as effectors. EAV-induced PBMC cultures showed evidence of expansion and activation of lymphoblasts, with an increase in the CD8+/CD4+ ratio in comparison with mock-induced PBMC. The cytotoxicity induced by EAV-stimulated PBMC was virus specific, showed genetic restriction, was mediated by CD8+ T lymphocytes and could be detected for periods of 4 months to more than 1 year post-infection. These findings and methods will hopefully contribute to an understanding of virus–host interactions in horses, in particular the mechanisms of virus clearance occurring during EAV infection.

scholarly journals Predominance of HLA-Restricted Cytotoxic T-Lymphocyte Responses to Serotype-Cross-Reactive Epitopes on Nonstructural Proteins following Natural Secondary Dengue Virus Infection

1998 ◽  
Vol 72 (5) ◽  
pp. 3999-4004 ◽  
Author(s):  
Anuja Mathew ◽  
Ichiro Kurane ◽  
Sharone Green ◽  
Henry A. F. Stephens ◽  
David W. Vaughn ◽  
...  
Keyword(s):  
Virus Infection ◽  
Dengue Virus ◽  
T Lymphocyte ◽  
Mononuclear Cells ◽  
Cytotoxic T Lymphocyte ◽  
Nonstructural Proteins ◽  
Dengue Virus Infection ◽  
Peripheral Blood Mononuclear ◽  
Blood Mononuclear Cells ◽  
Lymphocyte Responses

ABSTRACT We examined the memory cytotoxic T-lymphocytic (CTL) responses of peripheral blood mononuclear cells (PBMC) obtained from patients in Thailand 12 months after natural symptomatic secondary dengue virus infection. In all four patients analyzed, CTLs were detected in bulk culture PBMC against nonstructural dengue virus proteins. Numerous CD4+ and CD8+ CTL lines were generated from the bulk cultures of two patients, KPP94-037 and KPP94-024, which were specific for NS1.2a (NS1 and NS2a collectively) and NS3 proteins, respectively. All CTL lines derived from both patients were cross-reactive with other serotypes of dengue virus. The CD8+ NS1.2a-specific lines from patient KPP94-037 were HLA B57 restricted, and the CD8+ NS3-specific lines from patient KPP94-024 were HLA B7 restricted. The CD4+ CTL lines from patient KPP94-037 were HLA DR7 restricted. A majority of the CD8+ CTLs isolated from patient KPP94-024 were found to recognize amino acids 221 to 232 on NS3. These results demonstrate that in Thai patients after symptomatic secondary natural dengue infections, CTLs are mainly directed against nonstructural proteins and are broadly cross-reactive.

scholarly journals Deep sequencing of the HIV-1 polymerase gene for characterisation of cytotoxic T-lymphocyte epitopes during early and chronic disease stages

2021 ◽  
Author(s):  
Paballo Nkone ◽  
Shayne Loubser ◽  
Thomas C. Quinn ◽  
Andrew D. Redd ◽  
Arshad Ismail ◽  
...  
Keyword(s):  
T Lymphocyte ◽  
Deep Sequencing ◽  
Sanger Sequencing ◽  
Cytotoxic T Lymphocyte ◽  
Sequencing Data ◽  
Subtype C ◽  
Ctl Epitopes ◽  
Significant Difference ◽  
Minority Variant ◽  
Hiv 1

Abstract Background Despite multiple attempts, there is still no effective HIV-1 vaccine available. The HIV-1 polymerase (pol) gene is highly conserved and encodes cytotoxic T-lymphocyte (CTL) epitopes. In this study, deep sequencing was employed for characterisation of HIV-1 Pol CTL epitopes in mostly paired samples obtained during early and chronic stages of infection. Deep sequencing data was then compared to Sanger sequencing data only in samples obtained at baseline. Results Fifty-two participants were enrolled in the study. Their median age was 28 years (interquartile range: 24–32 years) and the majority of participants (92.3%) were female. Illumina minority variant analysis identified a significantly higher number of CTL epitopes (n = 65) compared to epitopes (n = 8) identified through Sanger sequencing. Most of the identified epitopes mapped to reverse transcriptase (RT) and integrase (IN) regardless of the method of sequencing. There was a significantly higher proportion of minority variant epitopes in RT (n = 39, 60.0%) compared to IN (n = 17, 26.2%) and PR (n = 9, 13.8%), p = 0.002 and < 0.0001, respectively. However, no significant difference was observed between the proportion of minority variant epitopes in IN versus PR, p = 0.06. Some epitopes were detected in either early or chronic HIV-1 infection whereas others were detected in both stages. Different distribution patterns of minority variant epitopes were observed in sample pairs; with some increasing or decreasing over time, while others remained constant. Some of the identified epitopes have not been previously reported for HIV-1 subtype C. There were also variants that could not be mapped to reported CTL epitopes in the Los Alamos HIV database. Conclusion Deep sequencing revealed many Pol CTL epitopes, including some not previously reported for HIV-1 subtype C. The findings of this study support the inclusion of RT and IN epitopes in HIV-1 vaccine candidates as these proteins harbour many CTL epitopes. Variants that were not mapped within CTL epitopes could represent new epitopes.

scholarly journals Between-Host Evolution of Cytotoxic T-Lymphocyte Epitopes in Human Immunodeficiency Virus Type 1: an Approach Based on Phylogenetically Independent Comparisons

2004 ◽  
Vol 78 (21) ◽  
pp. 11758-11765 ◽  
Author(s):  
Helen Piontkivska ◽  
Austin L. Hughes
Keyword(s):  
Natural Selection ◽  
T Lymphocyte ◽  
Cytotoxic T Lymphocyte ◽  
Ctl Epitopes ◽  
Term Evolution ◽  
Immunodeficiency Virus ◽  
Host Evolution ◽  
Hiv 1

ABSTRACT In human immunodeficiency virus type 1 (HIV-1), mutations that escape from cytotoxic T-lymphocyte (CTL) recognition have been documented, and sequence analyses have provided indirect support for the hypothesis that natural selection has favored CTL escape mutants within an infected host. In spite of such evidence for within-host selection by CTL, it has been more difficult to determine how natural selection by host CTL has influenced long-term evolution of HIV-1. We used statistical analysis of published HIV-1 genomic sequences to examine the role of natural selection in between-host evolution of CTL epitopes. Based on a phylogenetic analysis, we identified 21 pairs of closely related genomes isolated from different hosts and examined the pattern of nucleotide substitution in genomic regions encoding well-characterized CTL epitopes. The results revealed that certain CTL epitopes have been subject to repeated positive selection across the population, while others are generally conserved. Furthermore, evidence of positive selection was associated with divergence from the canonical epitope sequence and with an enhanced frequency of convergent amino acid sequence changes in CTL epitopes. The results support the hypothesis that CTL-driven selection has been a major factor in the long-term evolution of HIV-1.

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