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Human CD8(+) regulatory T cells, particularly the CD8(+)CD28(-) T suppressor cells, have emerged as an important modulator of alloimmunity. Understanding the conditions under which these cells are induced and/or expanded would greatly facilitate their application in future clinical trials. In the current study, we develop a novel strategy that combines common gamma chain (γc) cytokines IL-2, IL-7 and IL-15 and donor antigen presenting cells (APCs) to stimulate full HLA-mismatched allogeneic human CD8(+) T cells which results in significant expansions of donor-specific CD8(+)CD28(-) T suppressor cells in vitro. The expanded CD8(+)CD28(-) T cells exhibit increased expressions of CTLA-4, FoxP3, and CD25, while down-regulate expressions of CD56, CD57, CD127, and perforin. Furthermore, these cells suppress proliferation of CD4(+) T cells in a contact-dependent and cytokine-independent manner. Interestingly, the specificity of suppression is restricted by the donor HLA class I antigens but promiscuous to HLA class II antigens, providing a potential mechanism for linked suppression. Taken together, our results demonstrate a novel role for common γc cytokines in combination with donor APCs in the expansion of donor-specific CD8(+)CD28(-) T suppressor cells, and represent a robust strategy for in vitro generation of such cells for adoptive cellular immunotherapy in transplantation.

Immunologic risk in kidney transplantation is typically minimized by avoiding, or at least limiting, the potential of donor specific humoral responses by testing for the presence of donor-specific antibodies (DSA). Additionally, selecting donor and recipient pairs with the least number of human leukocyte antigen (HLA) mismatches has been shown to play a role in transplant outcome. However, numerous other factors may play a role in the success of transplant outcome and patient health. Specifically, the use of T-cell allospecific ELISPOT assays have helped elucidate the role of pre-formed cellular responses as additional factors in post-transplant outcome. In this review, we will evaluate numerous uses of ELISPOT assays to assess the pre- and post-transplant immunologic risk of rejection episodes, graft survival and even viral susceptibility as well as the utility of ELISPOT assays in monitoring tolerance and withdrawal of immunosuppressive medications following kidney transplantation.