krn-7000 and Leishmaniasis--Visceral

NKT cells respond to presentation of specific glycolipids with release of both Th1- and Th2-type cytokines. Leishmania donovani (LD)-infected splenic macrophages (sMϕ(I)) and bone marrow-derived dendritic cells (BMDC(I)) failed to activate NKT cells in response to α-galactosyl ceramide (α-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 sMϕ(I) with GSPL, caused the activation of the Vα14(+) 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)/sMϕ(I) lipid raft structure, which is a site for immune regulation.

Topics: Animals; Antigen Presentation; Antigens, CD1d; Blotting, Western; Calcium; Cell Membrane; Cells, Cultured; Cricetinae; Electrophoresis, Polyacrylamide Gel; Flow Cytometry; Fluorescent Antibody Technique; Galactosylceramides; Glycosphingolipids; Immunological Synapses; Isoelectric Focusing; Leishmania donovani; Leishmaniasis, Visceral; Lymphocyte Activation; Macrophages; Membrane Microdomains; Mice; Natural Killer T-Cells

We report that natural killer T (NKT) cells play only a minor physiological role in protection from Leishmania donovani infection in C57BL/6 mice. Furthermore, attempts at therapeutic activation of invariant NKT (iNKT) cells with alpha-galactosylceramide (alpha-GalCer) during L. donovani infection exacerbated, rather than ameliorated, experimental visceral leishmaniasis. The inability of alpha-GalCer to promote anti-parasitic immunity did not result from inefficient antigen presentation caused by infection because alpha-GalCer-loaded bone marrow-derived dendritic cells were also unable to improve disease resolution. The immune-dampening affect of alpha-GalCer correlated with a bias towards increased IL-4 production by iNKT cells following alpha-GalCer stimulation in infected mice compared to naïve controls. However, studies in IL-4-deficient mice, and IL-4 neutralisation in cytokine-sufficient mice revealed that alpha-GalCer-induced IL-4 production during infection had only a minor role in impaired parasite control. Analysis of liver cell composition following alpha-GalCer stimulation during an established L. donovani infection revealed important differences, predominantly a decrease in IFNgamma+ CD8+ T cells, compared with control-treated mice. Our data clearly illustrate the double-edged sword of NKT cell-based therapy, showing that in some circumstances, such as when sub-clinical or chronic infections exist, iNKT cell activation can have adverse outcomes.

Topics: Animals; Antibodies, Blocking; Biomarkers; CD8-Positive T-Lymphocytes; Disease Models, Animal; Female; Galactosylceramides; Gene Silencing; Host-Parasite Interactions; Immunologic Factors; Interferon-gamma; Interleukin-4; Killer Cells, Natural; Leishmania donovani; Leishmaniasis, Visceral; Liver; Lymphocyte Activation; Lymphocyte Subsets; Mice; Mice, Inbred BALB C; Mice, Inbred C57BL; Mice, Knockout; RNA, Messenger