Dendritic cells (DC) are professional APC that are able to modulate immune response in either a positive or negative manner depending upon their lineage and state of maturation. RelB is a NF-kappaB family member which plays a key role in the differentiation and maturation of DC. In this study, we constructed lentiviral vector expressing RelB-specific short hairpin RNAs (ShRNAs) that efficiently silenced the RelB gene in bone marrow-derived dendritic cells (BMDCs). These RelB-silenced BMDCs were maturation resistant and could functionally decrease antigen-specific T cells proliferation. We tested the therapeutic effect of RelB-silenced BMDCs in C57BL/6 mice with experimental autoimmune myasthenia gravis (EAMG). Injection i.v. with RelB-silenced BMDCs plused with Torpedo acetylcholine receptor (TAChR) dominant peptide Talpha(146-162) on days 3, 33, and 63 after first immunization decreased the incidence and severity of clinical EAMG with suppressed IFN-gamma production and increased IL-10 and IL-4 production in vitro and in vivo, and also leads to a decreased serum anti-AChR IgG, IgG1, IgG2b Ab levels. Furthermore, RelB-silenced BMDCs promoted regulatory T cell profiles as indicated by a marked increase of FoxP3 in splenocyte. Our data suggested that lentiviral-mediated RNAi targeting RelB was effective methods to inhibit the maturation of BMDCs, thus possess therapeutic potential to prevent autoimmune disorders such as EAMG or human MG.