Autoreactive CD4+ T cells exist in normal individuals and retain the capacity to initiate autoimmune disease. The current study investigates the role of CD4+ CD25+ T-regulatory (T R) cells during autoimmune disease using the CD4+ T cell-dependent myelin oligodendrocyte glycoprotein (MOG)-specific experimental autoimmune encephalomyelitis model of multiple sclerosis. In vitro, T R cells effectively inhibited both the proliferation of and cytokine production by MOG 35–55-specific Th1 cells. In vivo, adoptive transfer of T R cells conferred significant protection from clinical experimental autoimmune encephalomyelitis which was associated with normal activation of autoreactive Th1 cells, but an increased frequency of MOG 35–55-specific Th2 cells and decreased CNS infiltration. Lastly, transferred T R cells displayed an enhanced ability to traffic to the peripheral lymph nodes and expressed increased levels of the adhesion molecules ICAM-1 and P-selectin that may promote functional interactions with target T cells. Collectively, these findings suggest that T R cells contribute notably to the endogenous mechanisms that regulate actively induced autoimmune disease.