Activated iNKT Cells Promote Memory CD8+ T Cell Differentiation during Viral Infection

α-galactosylceramide (α-GalCer) is the prototypical lipid ligand for invariant NKT cells. Recent studies have proposed that α-GalCer is an effective adjuvant in vaccination against a range of immune challenges, however its mechanism of action has not been completely elucidated. A variety of delivery methods have been examined including pulsing dendritic cells with α-GalCer to optimize the potential of α-GalCer. These methods are currently being used in a variety of clinical trials in patients with advanced cancer but cannot be used in the context of vaccine development against pathogens due to their complexity. Using a simple delivery method, we evaluated α-GalCer adjuvant properties, using the mouse model for cytomegalovirus (MCMV). We measured several key parameters of the immune response to MCMV, including inflammation, effector, and central memory CD8+ T cell responses. We found that α-GalCer injection at the time of the infection decreases viral titers, alters the kinetics of the inflammatory response, and promotes both increased frequencies and numbers of virus-specific memory CD8+ T cells. Overall, our data suggest that iNKT cell activation by α-GalCer promotes the development of long-term protective immunity through increased fitness of central memory CD8+ T cells, as a consequence of reduced inflammation

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

Combining Carbon ion Radiotherapy and Local Injection of Alpha-galactosylceramide-pulsed Dendritic Cells Inhibits Lung Metastases in an in vivo Murine Model

Purpose: Our previous report indicated that carbon ion beam irradiation upregulatedimmunogenic, underlining the potential clinical application of radioimmunotherapy. The antimetastatic efficacy of a local combination therapy of carbon-ion radiotherapy and immunotherapy was examined using in vivo murine model.Methods and Materials: Tumors of the mouse squamous cell carcinoma (NR-S1) cells inoculated in the leg of C3H/HeMsNrs mice were locally irradiated with a single 6 Gy dosage of carbon ions (290 MeV/nucleon, 6 cm spread-out Bragg peak). Thirty-six hours after irradiation, alpha-galactosylceramide (alpha-GalCer)-pulsed dendritic cells (DCs) were injected into the leg tumor. The effects on distant lung metastases were investigated using pathological observations and immunohistochemistry.Results: The mice in the control group, which were not irradiated and not injected with alpha-GalCerpulsed DCs, had 168 +/- 53.8 metastatic nodules in their lungs at 2 weeks. The lung metastases were significantly decreased in the mouse which received the combination therapy, and the mean number of lung metastatic nodules was 2.6 +/- 1.9 (P = 0.009). In addition, the size of each lung metastasis was significantly decreased in compared to the control group (P = 0.009). Immunohistochemistry showed that ICAM-1, which activates DCs, was highly expressed on the local tumors of the carbon ionirradiated group. The expression of S100A8 in lung tissue, a marker of the lung pre-metastatic phase, was decreased in the combination therapy group.Conclusion: The combination of carbon ion radiotherapy with the injection of alpha-GalCer-pulsed DCs into the primary tumors significantly decreased the number of lung metastases

Establishment of an Improved Mouse Model for Infantile Neuroaxonal Dystrophy That Shows Early Disease Onset and Bears a Point Mutation in Pla2g6

Calcium-independent group VIA phospholipase A2 (iPLA2β), encoded by PLA2G6, has been shown to be involved in various physiological and pathological processes, including immunity, cell death, and cell membrane homeostasis. Mutations in the PLA2G6 gene have been recently identified in patients with infantile neuroaxonal dystrophy (INAD). Subsequently, it was reported that similar neurological impairment occurs in gene-targeted mice with a null mutation of iPLA2β, whose disease onset became apparent approximately 1 to 2 years after birth. Here, we report the establishment of an improved mouse model for INAD that bears a point mutation in the ankyrin repeat domain of Pla2g6 generated by N-ethyl-N-nitrosourea mutagenesis. These mutant mice developed severe motor dysfunction, including abnormal gait and poor performance in the hanging grip test, as early as 7 to 8 weeks of age, in a manner following Mendelian law. Neuropathological examination revealed widespread formation of spheroids containing tubulovesicular membranes similar to human INAD. Molecular and biochemical analysis revealed that the mutant mice expressed Pla2g6 mRNA and protein, but the mutated Pla2g6 protein had no glycerophospholipid-catalyzing enzyme activity. Because of the significantly early onset of the disease, this mouse mutant (Pla2g6-inad) could be highly useful for further studies of pathogenesis and experimental interventions in INAD and neurodegeneration