Background.

Regulatory T cells (Treg) play important roles in preventing autoimmunity, graft-versus host disease and transplant rejection. In rodent transplant models, tolerance induction strategies can induce graft protective CD25+ CD4+ Treg in vivo but therapeutic exploitation of active regulation will more likely depend on protocols that allow generation or selection of regulatory cells ex vivo for use as a cellular therapy. We have used adoptive transfer skin and islet allograft models to identify, develop and evaluate ex vivo protocols that generate donor-reactive, adaptive Treg.

Methods.

Naïve CDA CD4+ T cells were stimulated with allogeneic antigen-presenting cell under neutral conditions or with cytokine modification, restimulated under identical conditions and subsequently analyzed for cytokine profile, phenotypic markers characteristic of Treg and in vivo regulatory function.

Results.

Without modification, CD4+ T cells default to a Th2 phenotype characterized by a dominant interleukin-4 response which is profoundly detrimental to allograft survival. However, addition of exogenous interferon-γ suppresses interleukin-4 production without priming for effector function, induces suppressor of cytokine signaling-1 and results in up-regulation of Foxp3 and CD62L. The generation of these populations is enhanced by, but is independent of, the presence of naturally occurring endogenous Treg. Most importantly, when tested for regulatory function in vivo, these cells prevent rejection of both skin and islet allografts mediated by effector T cells.

Conclusions.

These data reveal an unexpected role for interferon-γ in the generation of Treg ex vivo and suggest a possible route for the generation of regulatory cells for therapeutic use.