Donor-specific transplantation tolerance promises to free recipients from complications associated with lifelong immunosuppression. Anergy, a CD4+ conventional T cell (Tconv cell)-intrinsic state of long-term hypofunction, is thought to arise when T cells encounter cognate antigen in the absence of costimulation. Anti-CD154 (αCD154), an antibody that blocks T cell costimulation and is currently in clinical trials, can induce donor-specific transplantation tolerance in mice that is associated with anergy of Tconv cells recognizing persistently expressed alloantigens. Using murine models of fully mismatched cardiac allograft tolerance or repeated alloantigen exposure, we demonstrate that αCD154 is not sufficient and regulatory T cells (Treg cells) are necessary to induce and sustain αCD154-mediated alloreactive Tconv cell anergy. Transient depletion of Treg cells disrupted the induction of Tconv cell anergy, and in mice with established tolerance, Treg cell depletion transcriptionally and functionally reinvigorated previously anergic Tconv cells. Mechanistically, Treg cell depletion increased IL-2 availability and IL-2R signaling in Tconv cells, and exogenous IL-2 could bypass Treg cell-mediated control to partially restore Tconv cell function. Thus, the state of Tconv cell anergy is less cell-intrinsic than previously thought and requires Treg cells to limit IL-2 availability for both its induction and its continuous enforcement. Selecting immunosuppressive drugs that preserve this Treg cell function may be critical for achieving stable clinical transplantation tolerance.
