Contribution of Regulatory T Cells and Effector T Cell Deletion in Tolerance Induction by Costimulation Blockadel

Abstract

Blocking of costimulatory signals for T cell activation leads to tolerance in several transplantation models, but the underlying mechanisms are incompletely understood. We analyzed the involvement of regulatory T cells (Treg) and deletion of alloreactive cells in the induction and maintenance of tolerance after costimulation blockade in a mouse model of graft-vs-host reaction. Injection of splenocytes from the C57BL/6 parent strain into a sublethally irradiated F1 offspring (C57BL/6 × C3H) induced a GVHR characterized by severe pancytopenia. Treatment with anti-CD40L mAb and CTLA4-Ig every 3 days during 3 wk after splenocyte injection prevented disease development and induced a long-lasting state of stable mixed chimerism (>120 days). In parallel, host-specific tolerance was achieved as demonstrated by lack of host-directed alloreactivity of donor-type T cells in vitro and in vivo. Chimerism and tolerance were also obtained after CD25+ cell-depleted splenocyte transfer, showing that CD25+ natural Treg are not essential for tolerance induction. We further show that costimulation blockade results in enhanced Treg cell activity at early time points (days 6–30) after splenocyte transfer. This was demonstrated by the presence of a high percentage of Foxp3+ cells among donor CD4+ cells in the spleen of treated animals, and our finding that isolated donor-type T cells at an early time point (day 30) after splenocyte transfer displayed suppressive capacity in vitro. At later time points (>30 days after splenocyte transfer), clonal deletion of host-reactive T cells was found to be a major mechanism responsible for tolerance.

Regulatory T Cell–Dependent and –Independent Mechanisms of Immune Suppression by CD28/B7 and CD40/CD40L Costimulation Blockade

“…Our data indeed also show a role for Tregs in the effects of costimulation blockade. It has already been suggested by several groups that Tregs are involved in tolerance induction by costimulation blockade (10,(13)(14)(15). CD40L blockade especially was found to be associated with an enhanced Treg function in previous studies (13-15, 19, 20).…”

“…We and others have shown that combined blockade of the CD40/ CD40L interaction and the CD28/B7 interaction completely prevents GVHD after allogeneic splenocyte transfer in mice and results in tolerance (8)(9)(10). Separate blockade of these costimulatory pathways is not effective.…”

“…Our previous study documented an expansion of donor Tregs when GVHD mice were treated with MR1 and CTLA-4Ig (10). Other studies have documented that the effect of MR1 is at least partially dependent on Treg activity (13-15, 19, 20), whereas data on the effect of CTLA4Ig on Tregs are controversial (10,13,(21)(22)(23). First, we analyzed the effect of MR1 and different doses of CTLA-4Ig treatment on the numbers of Foxp3 + Tregs.…”

“…Combined treatment with CTLA-4Ig and anti-CD40L mAb is clearly superior over single-agent treatment in mice and in nonhuman primates (8,9). In this context, we have previously demonstrated that treatment with CTLA-4Ig and MR1 (a blocking anti-CD40L mAb) completely prevents GVHD in a fully MHCmismatch "parent into F1" splenocyte transfer mouse model (10). Long-term survival, tolerance, and mixed chimerism were induced only if both reagents were applied together.…”

“…It is believed that TCR triggering without a costimulatory signal results in T cell hyporesponsiveness (11), followed by peripheral clonal deletion by apoptosis (10,12). It has further been suggested that regulatory T cells (Tregs) are involved (13)(14)(15).…”