Recently, there has been considerable interest in developing specific cell-based immunotherapies using dendritic cells (DCs). Here the mechanisms underlying the tolerogenic properties of DCs in the suppression of experimental autoimmune uveoretinitis (EAU), the mouse model of human sight threatening autoimmune uveitis, were examined. Immature DCs have the ability to promote immune tolerance to self antigens and to prevent the development of autoimmune disorders including EAU. However there is a risk that immature DCs placed in the inflammatory environment would undergo maturation and instead of tolerance promote immunity. Therefore much effort has been directed to develop protocols that stabilize the tolerogenic DC properties which would ultimately ensure safety and effectiveness of DC-based vaccines. Previous work demonstrated that activation of DCs with lipopolysaccharide (LPS) increases the ability of these cells to prevent EAU. Here, it was demonstrated that LPS promotes the activation of both TRIF and MyD88 signalling pathways in DCs which after 24 h LPS treatment secreted a significant amount of IL-10, IFN-β, IL-1β, IL-6 and TNF-α while the level of secreted IL-12 is low. It was further shown that LPSinduced enhancement of the tolerogenic properties of DCs correlates with the state of endotoxin tolerance in the DCs, rendering them refractory to further stimulation. It was hypothesised that the LPS-induced enhancement of DC tolerogenicity is due to the reduced expression of the TLR4, which subsequently disables several signalling pathways and prevent DCs from initiating adaptive T cell immunity. In summary, the presented data provides valuable insights into the mechanisms involved in the suppression of autoimmune uveitis using a DC-based therapy.