Identification of New Alloantigen-Reactive CD8⁺ Cytotoxic and Suppressor T Cell Subpopulations.

We sought to develop a better understanding of the T cells involved in the human allogeneic immune response, in order to eventually engineer a donor graft with reduced GVHD-mediating potential, without ablating general immune competence. Prior studies reported that all the activated CD4⁺ T cells responding to a specific antigen challenge reside within the CD4^high population expressing high levels of membrane CD4. We identified a new population of activated CD8⁺ T cells that developed during an in vitro allogeneic immune response, along with the allo-activated CD4^high T cell population. Analogous to activated CD4⁺ T cells, this new T cell population was distinguished by up-regulated CD8 (and CD38) expression (CD8^highCD38⁺). In accordance with Martins et al. (Blood 2004, 104:3429), we found that the depletion of the CD4^highCD38⁺ population resulted in reduced 2^o response to the original 2^nd party stimulators. In contrast, depletion of the CD8^highCD38⁺ population resulted in an increased 2^o response to 2^nd party cells, with no change in the response to 3^rd party or CMV antigens. Elevated numbers of CD8^highCD38⁺ T cells potently reduced the 1^o and 2^o responses to 2^nd party, but not to 3^rd party cells or CMV antigens. The complementary, non-activated CD8^normalCD38⁻ T cell population had no inhibitory effect. Importantly, we found that

• CD8^highCD38⁺ T cells mediated both a specific cytotoxic response (that could be inhibited by the pan-caspase inhibitor, Z-VAD), and a specific suppressive response toward the original 2^nd party stimulators (that was not affected by Z-VAD), and

• within this CD8^highCD38⁺ population, there was a subpopulation of cytotoxic T cells (perforin⁺LAMP1⁺CD56⁺CD11b⁺CD11c⁺) and a subpopulation of non-cytotoxic T cells.

Furthermore, we found that although CD8^highCD38⁺ T cells differentially expressed CD28, both CD8^highCD38⁺CD28⁻ and CD8^highCD38⁺CD28⁻ T cells mediated a cytotoxic as well as a suppressor T cell response toward the original 2^nd party cells (different from the published suppressive function of CD8⁺CD28⁻ T cells observed by Liu et al, Int Immunol 1998, 10:775). Upon separation of cytotoxic CD8^highCD38⁺ T cells from suppressor CD8^highCD38⁺ T cells, we will explore the GVHD potential of these 2 novel activated CD8^high T cell subpopulations, in a sensitive in vivo xenograft model for GVHD using NOD/SCID/IL2Rγ^null immunodeficient mice.