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.