Transient local depletion of Foxp3+ regulatory T cells during recovery from colitis via Fas/Fas ligand-induced death

Regulatory T cells (T_regs) play a fundamental role in regulating the immune system in health and disease. Considerable evidence exists demonstrating that transfer of T_regs can cure colitis and a variety of other inflammatory disorders. However, little is known about the effects of inflammation on resident T_regs. Mice (BALB/c or C57BL/6) treated with an intrarectal instillation of the haptenizing agent 2,4-dinitrobenzene sulfonic acid (DNBS) develop an acute inflammatory disease, the histopathology of which peaks at 3 days posttreatment and resolves spontaneously thereafter. In this study we demonstrate that DNBS (or oxazolone)-induced colitis causes a depletion of colonic Foxp3⁺ T_regs 8 days posttreatment, while the proportion of Foxp3⁺ cells in the ileum, mesenteric lymph nodes, and spleen remains unchanged. Replenishment of the colonic T _regs population was associated with the reappearance of mucosal homing (α₄β₇⁺) CD4 ⁺Foxp3⁺ T_regs. Assessing the mechanism of local T_regs depletion, we found no evidence to implicate cytokine-induced phenotypic switching in the Foxp3⁺ population or increased SMAD7 expression despite the essential role that TGF-β has in Foxp3⁺ T_regs biology. Increased Fas ligand (FasL) expression was observed in the colon of colitic mice and in vitro stimulation with a Fas cross-linking Ab resulted in apoptosis of CD4⁺Foxp3⁺ but not CD4 ⁺Foxp3^- cells. Furthermore, DNBS-induced colitis in Fas/FasL-deficient mice did not result in depletion of colonic T_regs. Finally, adoptively transferred synergic Fas^-/- but not Fas ^+/+ T_regs were protected from depletion in the colon 8 days post-DNBS treatment, thus substantiating the hypothesis that inflammation-induced local depletion of Foxp3⁺T_regs. in the colon of mice occurs via Fas/FasL-mediated death.