Cytotoxic Efficiency of Human CD8+ T Cell Memory Subtypes

Immunological memory is important to protect humans against recurring diseases. Memory CD8(+) T cells are required for quick expansion into effector cells but also provide immediate cytotoxicity against their targets. Whereas many functions of the two main cytotoxic subtypes, effector memory CD8(+) T cells (T-EM) and central memory CD8(+) T cells (T-CM), are well defined, single T-EM and T-CM cell cytotoxicity has not been quantified. To quantify cytotoxic efficiency of T-EM and T-CM, we developed a FRET-based single cell fluorescent assay with NALM6 target cells which allows analysis of target cell apoptosis, secondary necrosis following apoptosis, and primary necrosis after T-EM- or T-CM-target cell contact. Both, single cell and population cytotoxicity assays reveal a higher cytotoxic efficiency of T-EM compared to T-CM, as quantified by target cell apoptosis and secondary necrosis. Perforin, granzyme B, FasL, but not TRAIL expression are higher in T-EM compared to T-CM. Higher perforin levels (likely in combination with higher granzyme levels) mediate higher cytotoxic efficiency of T-EM compared to T-CM. Both, T-EM and T-CM need the same time to find their targets, however contact time between CTL and target, time to induce apoptosis, and time to induce secondary necrosis are all shorter for T-EM. In addition, immune synapse formation in T-EM appears to be slightly more efficient than in T-CM. Defining and quantifying single T-EM and T-CM cytotoxicity and the respective mechanisms is important to optimize future subset-based immune therapies.

Automated summary

This study developed a FRET-based single cell fluorescent assay to compare the cytotoxic efficiency of effector memory CD8(+) T cells (T-EM) and central memory CD8(+) T cells (T-CM). The results showed that T-EM had higher cytotoxic efficiency compared to T-CM. This could be attributed to the higher expression levels of perforin and granzyme B in T-EM, which likely mediate the higher cytotoxic efficiency.