Background: HIV-1-specific CD8⁺ T cells play a crucial role for the spontaneous control of HIV-1 replication, yet, in numerous studies, no correlation was found between HIV-specific CD8⁺ T-cell responses in interferon-g ELISpot and clinical or surrogate markers of HIV disease progression, even when comprehensive screening approaches were applied. Thus, it is currently unclear if interferon-g secretion by HIV-specific CD8⁺ T cells adequately reflects their ability to eliminate HIV-infected cells by MHC-class I restricted cytolysis.

Methods: Here, we adapted a novel cytotoxicity assay based on caspase-3 substrates as indicators of target cell death, allowing for the direct ex vivo measurement of the cytolytic capacity of HIV-specific CD8⁺ T cells. Using this method, we directly compared interferon-g secretion, as determined by intracellular cytokine staining, with the cytotoxic capacity of freshly-isolated HIV-specific CD8⁺ T cells.

Results: In a total of 48 different HIV-specific CD8⁺ T-cell populations from 8 different study subjects, we found considerable intra-individual and inter-individual differences between cytotoxic- and IFN-g-secreting responses to individuals epitopes, indicating that the execution of these effector functions is only weakly correlated (R=0.46, p=0.0015). A substantially stronger relationship (R=0.65, p<0.0001) was found when the cytotoxic potential of CD8⁺ T cells was plotted against the proportion of HIV-specific CD8⁺ T cells that produced both TNF-a and interferon-g following viral stimulation. In contrast, no correlation was found between the cytotoxic potential of HIV-specific CD8⁺ T cells and the proportion of cells that produce interferon-g but no TNF-a following viral stimulation (R=0.3, p=0.4).  

Conclusion: These results suggest that the direct cytolytic effects of HIV-specific CD8⁺ T cells are preferentially mediated by the subset of cells capable of producing both interferon-g and TNF-a in response to viral stimulation. The characterization of this CD8⁺ T cell subset will be thus relevant for a more precise evaluation of CD8⁺ T cell mediated HIV-specific immune responses generated naturally or following immunization with HIV vaccine candidates.  

Keywords: HIV-specific CD8+ T cells; cytotoxicity; effector functions