Triptolide (TPT), a diterpenoid triepoxide, is the major component isolated from the Chinese herb Tripterygium wilfordii Hook. f. Previous studies have shown that TPT has immunosuppressive properties and is effective in prolonging graft survival and suppressing autoimmune responses. The aim of this study was to investigate the protective effect of TPT in experimental autoimmune encephalomyelitis (EAE), an animal model of multiple sclerosis. Treatment of C57BL/6 mice with TPT from the date of EAE induction significantly delayed EAE onset and suppressed disease severity, accompanied with reduced inflammation and demyelination in the central nervous system. TPT treatment lead to a significant inhibition of the mRNA expression of both Th1/Th(IL-17) and Th2 cytokines in spleen mononuclear cells (MNC) as well as in spinal cord tissues. In addition, the expression of Forkhead box p3 (Foxp3) was up-regulated in spleen MNC after TPT treatment. Furthermore, we detected apparent inhibition of nuclear factor-kappa B (NF-kappaB)-DNA binding activity, increased expression of the inhibitor of nuclear factor-kappa Balpha (IkappaBalpha) and decreased expression of pIkappaBalpha in spleen MNC in TPT-treated EAE mice. Taken together, these findings indicate that TPT has profound immunoregulatory functions and potential protective values for the treatment of autoimmune inflammatory disorders.