Leishmania donovani glycosphingolipid facilitates antigen presentation by inducing relocation of CD1d into lipid rafts in infected macrophages

NKT cells respond to presentation of specific glycolipids with release of both Th1- and Th2-type cytokines. Leishmania donovani (LD)-infected splenic macrophages (sM phi(I)) and bone marrow-derived dendritic cells (BMDC(I)) failed to activate NKT cells in response to alpha-galactosyl ceramide (alpha-GalCer). The defective antigen presentation could be corrected by treating the cells with the immunostimulating glycosphingophospholipid (GSPL) of LD parasites. In vitro pulsing of BMDC(I) or sM phi(I) with GSPL, caused the activation of the V alpha 14(+) CD1d1-specific NKT cell hybridoma DN32.D3. Localization of MHC II and CD1d molecules to membrane lipid rafts has been suggested to play an important role in antigen presentation. Confocal analysis clearly demonstrated that LD infection changed the pattern of CD1d distribution to the non-lipid raft regions and this change could be reversed by GSPL treatment. Isoelectric focusing gel shift assay indicated that GSPL binds to CD1d. GSPL-treated but not untreated BMDC(I) formed immune synapses with NKT cells and this was associated with calcium mobilization. In conclusion, GSPL treatment was associated with modification of BMDC(I)/sM phi(I) lipid raft structure, which is a site for immune regulation.