Not all cytokine-producing CD8+ T cells suppress simian immunodeficiency virus replication

Abstract

Current assays of CD8+ T-lymphocyte function measure cytokine production rather than the ability of these lymphocytes to suppress viral replication. Here we show that CD8+ T-cell clones recognizing the same epitope vary enormously in the ability to suppress simian immunodeficiency virus SIVmac239 replication in an in vitro suppression assay. However, all Nef(165-173)IW9- and Vif(66-73)HW8-specific clones from elite controllers effectively suppressed SIV replication. Interestingly, in vitro suppression efficacy was not always associated with the ability to produce gamma interferon, tumor necrosis factor alpha, or interleukin-2.

FIG. 1.

Clonal variation of representative Mamu-A^*02-Nef_159-167YY9-specific CD8⁺ T cells in the ability to suppress SIVmac239 replication. Supernatants were collected on days 4, 6, and 8 from VSA duplicate wells with MHC class I-matched targets and six Nef_159-167YY9-specific clones from animal r00044. The clones were used at an E:T of 1:10 and analyzed by quantitative PCR to determine average viral RNA copy numbers (A). Day 8 VSA target cells were stained with an anti-Gag p27 antibody to determine the frequency of SIV-infected target cells (B). Viral suppression only occurred in an MHC class I-restricted fashion (MHC class I-mismatched target data not shown).

FIG. 2.

Effective suppressions of SIVmac239 replication by all Mamu-B^*17-Nef_165-173IW9-specific clones. Supernatants were collected on days 4, 6, and 8 from VSA duplicate wells with MHC class I-matched target cells. Seven Nef_165-173IW9 clones from animal r95061 were used at an E:T of 1:10 and analyzed by quantitative PCR to determine average viral RNA copy numbers (A). Day 8 VSA target cells were stained with an anti-Gag p27 antibody to determine the frequency of SIV-infected cells (B). Viral suppression only occurred in an MHC class I-restricted fashion (MHC class I-mismatched target data not shown).

FIG. 3.

IFN-γ, TNF-α, and IL-2 responses were not always associated with the ability to suppress SIVmac239 replication. Gag p27 staining was carried out with day 8 VSA target cells to determine the frequency of SIV-infected target cells. Clones were stimulated with autologous B-lymphoblastoid cell lines pulsed with cognate peptide and stained with anti-IFN-γ, anti-TNF-α, and/or anti-IL-2 antibodies. Nef_159-167YY9-specific clones from r95061 (A), Nef_195-203MW9- and Env_830-838FW9-specific clones from r95071 (B and C), and Vif_66-73HW8-specific clones from r98016 (D) were compared. Analyses showing the statistical significance of differences between SIV suppression and cytokine expression with log-transformed data from suppressive and nonsuppressive groups are shown in panels E, F, and G.