Spleen-Resident CD4+ and CD4− CD8α− Dendritic Cell Subsets Differ in Their Ability to Prime Invariant Natural Killer T Lymphocytes

One important function of conventional dendritic cells (cDC) is their high capacity to capture, process and present Ag to T lymphocytes. Mouse splenic cDC subtypes, including CD8α+ and CD8α− cDC, are not identical in their Ag presenting and T cell priming functions. Surprisingly, few studies have reported functional differences between CD4− and CD4+ CD8α− cDC subsets. We show that, when loaded in vitro with OVA peptide or whole protein, and in steady-state conditions, splenic CD4− and CD4+ cDC are equivalent in their capacity to prime and direct CD4+ and CD8+ T cell differentiation. In contrast, in response to α-galactosylceramide (α-GalCer), CD4− and CD4+ cDC differentially activate invariant Natural Killer T (iNKT) cells, a population of lipid-reactive non-conventional T lymphocytes. Both cDC subsets equally take up α-GalCer in vitro and in vivo to stimulate the iNKT hybridoma DN32.D3, the activation of which depends solely on TCR triggering. On the other hand, and relative to their CD4+ counterparts, CD4− cDC more efficiently stimulate primary iNKT cells, a phenomenon likely due to differential production of co-factors (including IL-12) by cDC. Our data reveal a novel functional difference between splenic CD4+ and CD4− cDC subsets that may be important in immune responses

Tailored design of NKT-stimulatory glycolipids for polarization of immune responses

Natural killer T (NKT) cell is a distinct population of T lymphocytes that can rapidly release massive amount of Th1 and Th2 cytokines upon the engagement of their T cell receptor with glycolipids presented by CD1d. The secreted cytokines can promote cell-mediated immunity to kill tumor cells and intracellular pathogens, or suppress autoreactive immune cells in autoimmune diseases. Thus, NKT cell is an attractive target for developing new therapeutics to manipulate immune system. The best-known glycolipid to activate NKT cells is α-galactosylceramide (α-GalCer), which has been used as a prototype for designing new NKT stimulatory glycolipids. Many analogues have been generated by modification of the galactosyl moiety, the acyl chain or the phytosphingosine chain of α-GalCer. Some of the analogues showed greater abilities than α-GalCer in polarizing immune responses toward Th1 or Th2 dominance. Among them, several analogues containing phenyl groups in the lipid tails were more potent in inducing Th1-skewed cytokines and exhibited greater anticancer efficacy than α-GalCer. Analyses of the correlation between structure and activity of various α-GalCer analogues on the activation of iNKT cell revealed that CD1d–glycolipid complexes interacted with the same population of iNKT cell expressing similar T-cell receptor Vβ as α-GalCer. On the other hand, those phenyl glycolipids with propensity for Th1 dominant responses showed greater binding avidity and stability than α-GalCer for iNKT T-cell receptor when complexed with CD1d. Thus, it is the avidity and stability of the ternary complexes of CD1d-glycolipid-iNKT TCR that dictate the polarity and potency of immune responses. These findings provide a key to the rationale design of immune modulating glycolipids with desirable Th1/Th2 polarity for clinical application. In addition, elucidation of α-GalCer-induced anergy, liver damage and accumulation of myeloid derived suppressor cells has offered explanation for its lacklustre anti-cancer activities in clinical trials. On other hand, the lack of such drawbacks in glycolipid analogues containing phenyl groups in the lipid tails of α-GalCer coupled with the greater binding avidity and stability of CD1d-glycolipid complex for iNKT T-cell receptor, account for their superior anti-cancer efficacy in tumor bearing mice. Further clinical development of these phenyl glycolipids is warranted

Transient Foxp(3+) regulatory T-cell depletion enhances therapeutic anticancer vaccination targeting the immune-stimulatory properties of NKT cells

The natural killer T (NKT) cell ligand, alpha-galactosylceramide (-GalCer), represents a potential adjuvant to boost immunotherapeutic vaccination strategies against poorly immunogenic cancers. The objective of this study was to assess the therapeutic potential of an -GalCer-loaded tumor-cell vaccine against solid tumors in mice and to enhance the effectiveness of this approach by removing immune suppression associated with the activity of Foxp3 + regulatory T cells (Tregs). In the B16F10 melanoma model, we show that single vaccination with irradiated, -GalCer-loaded tumor cells resulted in suppression of established subcutaneous (s.c.) B16F10 tumor growth, which was mediated by NKT cell-dependent IFN-γ production and enhanced in the absence of IL-17 A. Selective depletion of Foxp3 + Tregs in transgenic DEpletion of REGulatory T cells (DEREG) mice led to significant inhibition of B16F10 tumor growth and enhanced survival of mice receiving vaccination. Short-term elimination of Foxp3 + Tregs

Interferon-γ independent formation of pulmonary granuloma in mice by injections with trehalose dimycolate (cord factor), lipoarabinomannan and phosphatidylinositol mannosides isolated from Mycobacterium tuberculosis

The mechanisms by which pulmonary granuloma formation is caused by administration of mycobacterial glycolipids such as trehalose dimycolate (TDM), lipoarabinomannan (LAM) and phosphatidylinositol mannosides (PIM) were investigated. When peritoneal and alveolar macrophages were stimulated with TDM, LAM and PIM in vitro, TDM exhibited the strongest tumour necrosis factor (TNF)-inducing activity. Responsiveness of macrophages from mice defected Toll-like receptor 4 (TLR4) was much higher than that of the wild-type mice. Although PIM and LAM also had a significant activity, LAM rather than PIM stimulated higher TNF-α production by alveolar macrophage. When mycobacterial glycolipids were injected as water-in-oil-in-water emulsion into mice via the tail vein, development of pulmonary granuloma in response to glycolipids were related closely to their TNF-inducing activity and TDM exhibited the strongest activity. Granuloma formation was observed not only in mice lacking interleukin (IL)-12 signalling but also interferon (IFN)-γ knock-out mice. Granuloma formation caused by glycolipids correlated with TNF-α levels in lungs. Administration of anti-TNF-α monoclonal antibody into TDM-injected IFN-γ knock-out mice decreased in granuloma formation, suggesting that development of pulmonary granuloma by mycobacterial glycolipids such as TDM is due to IFN-γ-independent and TNF-α-dependent pathway