Class I major histocompatibility complex-restricted cytotoxic T lymphocytes specific for Epstein-Barr Virus (EBV) nuclear antigens fail to lyse the EBV-transformed B lymphoblastoid cell lines against which they were raised

Ann Hill, Steven P. Lee, John S. Haurum, Nick Murray, Qing Yen Yao, Martin Rowe, Nathalie Signoret, Alan B. Rickinson, Andrew J. McMichael

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Abstract

We have raised CD8+ cytotoxic T lymphocytes (CTL) from three Epstein-Barr virus-seropositive donors by incubating peripheral blood lymphocytes with irradiated autologous B95.8-strain EBV-transformed B lymphoblastoid cells (LCL). However, to detect lysis in a standard 51Cr release assay of the LCL against which these CTL were raised, superinfection with recombinant vaccinia expressing the appropriate EBV protein or incubation with the peptide epitope was necessary. The untreated LCL were not lysed, even though Western blotting demonstrated that they expressed the EBV antigens containing the CTL epitopes. We have found CTL of this phenotype that are restricted by human leukocyte antigen-A2, -A3, -B7, or -B39, and which recognize the EBV latent proteins, EBV nuclear antigen (EBNA)-3A, EBNA-3C, or terminal protein. During these experiments, we identified a new human leukocyte antigen-A3-restricted EBNA-3A epitope, residues 603-611, RLRAEAGVK. We raised a spontaneous LCL, transformed by endogenous EBV, from one donor, but this was also not lysed. For at least one of the epitopes, CTL from another donor lysed the LCL without superinfection or addition of peptides. We conclude that the CTL were unable to achieve a high enough avidity of interaction with untreated LCL to trigger effector function, although the LCL were able to stimulate them to grow in vitro for up to 4 mo. To assess whether a small percentage of the LCL might possess a higher antigen density, we used an assay of tumor necrosis factor release from a CTL clone, which was able to detect antigen-bearing cells representing only 1% of a stimulating LCL population. Nevertheless, the untreated autologous LCL line failed to stimulate tumor necrosis factor release.

Original languageEnglish (US)
Pages (from-to)2221-2228
Number of pages8
JournalJournal of Experimental Medicine
Volume181
Issue number6
StatePublished - Jun 1 1995
Externally publishedYes

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Epstein-Barr Virus Nuclear Antigens
Cytotoxic T-Lymphocytes
Major Histocompatibility Complex
Human Herpesvirus 4
Cell Line
Epitopes
Superinfection
HLA Antigens
Antigens
Tumor Necrosis Factor-alpha
Vaccinia
Peptides
Proteins
T-Lymphocyte Epitopes
varespladib methyl
Clone Cells
Western Blotting

ASJC Scopus subject areas

  • Immunology

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Class I major histocompatibility complex-restricted cytotoxic T lymphocytes specific for Epstein-Barr Virus (EBV) nuclear antigens fail to lyse the EBV-transformed B lymphoblastoid cell lines against which they were raised. / Hill, Ann; Lee, Steven P.; Haurum, John S.; Murray, Nick; Yao, Qing Yen; Rowe, Martin; Signoret, Nathalie; Rickinson, Alan B.; McMichael, Andrew J.

In: Journal of Experimental Medicine, Vol. 181, No. 6, 01.06.1995, p. 2221-2228.

Research output: Contribution to journalArticle

Hill, Ann ; Lee, Steven P. ; Haurum, John S. ; Murray, Nick ; Yao, Qing Yen ; Rowe, Martin ; Signoret, Nathalie ; Rickinson, Alan B. ; McMichael, Andrew J. / Class I major histocompatibility complex-restricted cytotoxic T lymphocytes specific for Epstein-Barr Virus (EBV) nuclear antigens fail to lyse the EBV-transformed B lymphoblastoid cell lines against which they were raised. In: Journal of Experimental Medicine. 1995 ; Vol. 181, No. 6. pp. 2221-2228.
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abstract = "We have raised CD8+ cytotoxic T lymphocytes (CTL) from three Epstein-Barr virus-seropositive donors by incubating peripheral blood lymphocytes with irradiated autologous B95.8-strain EBV-transformed B lymphoblastoid cells (LCL). However, to detect lysis in a standard 51Cr release assay of the LCL against which these CTL were raised, superinfection with recombinant vaccinia expressing the appropriate EBV protein or incubation with the peptide epitope was necessary. The untreated LCL were not lysed, even though Western blotting demonstrated that they expressed the EBV antigens containing the CTL epitopes. We have found CTL of this phenotype that are restricted by human leukocyte antigen-A2, -A3, -B7, or -B39, and which recognize the EBV latent proteins, EBV nuclear antigen (EBNA)-3A, EBNA-3C, or terminal protein. During these experiments, we identified a new human leukocyte antigen-A3-restricted EBNA-3A epitope, residues 603-611, RLRAEAGVK. We raised a spontaneous LCL, transformed by endogenous EBV, from one donor, but this was also not lysed. For at least one of the epitopes, CTL from another donor lysed the LCL without superinfection or addition of peptides. We conclude that the CTL were unable to achieve a high enough avidity of interaction with untreated LCL to trigger effector function, although the LCL were able to stimulate them to grow in vitro for up to 4 mo. To assess whether a small percentage of the LCL might possess a higher antigen density, we used an assay of tumor necrosis factor release from a CTL clone, which was able to detect antigen-bearing cells representing only 1{\%} of a stimulating LCL population. Nevertheless, the untreated autologous LCL line failed to stimulate tumor necrosis factor release.",
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AU - Hill, Ann

AU - Lee, Steven P.

AU - Haurum, John S.

AU - Murray, Nick

AU - Yao, Qing Yen

AU - Rowe, Martin

AU - Signoret, Nathalie

AU - Rickinson, Alan B.

AU - McMichael, Andrew J.

PY - 1995/6/1

Y1 - 1995/6/1

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