Impact of bacteria on the phenotype, functions, and therapeutic activities of invariant NKT cells in mice
Sungjune Kim, Saif Lalani, Vrajesh V. Parekh, Tiffaney L. Vincent, Lan Wu, Luc Van Kaer
Sungjune Kim, Saif Lalani, Vrajesh V. Parekh, Tiffaney L. Vincent, Lan Wu, Luc Van Kaer
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Research Article Immunology

Impact of bacteria on the phenotype, functions, and therapeutic activities of invariant NKT cells in mice

Abstract

Invariant NKT (iNKT) cells are innate-like lymphocytes that recognize glycolipid antigens in the context of the MHC class I–like antigen-presenting molecule CD1d. In vivo activation of mouse iNKT cells with the glycolipid α-galactosylceramide (α-GalCer) results in the acquisition of a hyporesponsive (anergic) phenotype by these cells. Because iNKT cells can become activated in the context of infectious agents, here we evaluated whether iNKT cell activation by microorganisms can influence subsequent responses of these cells to glycolipid antigen stimulation. We found that mouse iNKT cells activated in vivo by multiple bacterial microorganisms, or by bacterial LPS or flagellin, became unresponsive to subsequent activation with α-GalCer. This hyporesponsive phenotype of iNKT cells required IL-12 expression and was associated with changes in the surface phenotype of these cells, reduced severity of concanavalin A–induced hepatitis, and alterations in the therapeutic activities of α-GalCer. These findings may have important implications for the development of iNKT cell–based therapies.

Authors

Sungjune Kim, Saif Lalani, Vrajesh V. Parekh, Tiffaney L. Vincent, Lan Wu, Luc Van Kaer

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Figure 4

Live L. monocytogenes infection induces hyporesponsiveness of iNKT cells to α-GalCer rechallenge ex vivo.

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Live L. monocytogenes infection induces hyporesponsiveness of iNKT cells...
(A) In vivo dynamics of the iNKT cell population in response to L. monocytogenes infection. Mice were injected with α-GalCer (5 μg/mouse, i.p.) or infected with L. monocytogenes and sacrificed at the indicated time points, and spleen and liver mononuclear cells were prepared and stained with anti–TCR-β–FITC, anti-NK1.1–PE, anti-B220–PerCP, and tetramer-allophycocyanin. Numbers indicate the percentage of TCR-β+tetramer+ cells among B220– cells. Representative plots from 4–7 mice per group are shown. (B) Total spleen iNKT cell counts and percentage of liver iNKT cells at the indicated times, for a total of 4–7 mice per group, pooled from 2 separate experiments. *P < 0.05 compared with naive animals. (C) The in vitro α-GalCer recall response of mice at the indicated times after infection. Mice were infected with L. monocytogenes and sacrificed 3 days or 1, 2, or 4 weeks later, and splenocytes (2 × 105 per well) were cultured with graded doses of α-GalCer. After 3 days, proliferation was assessed by [3H]thymidine incorporation, and culture supernatants were evaluated for IL-4 and IFN-γ levels by ELISA. Proliferation and cytokine results represent the mean ± SEM of 4–8 mice pooled from 2 separate experiments. *P < 0.05 compared with naive splenocytes cultured with the same dose of α-GalCer. (D) iNKT cell cytokine production. Spleen cells were prepared at the indicated times, and 2 × 105 cells were cultured for 6 hours in plain medium or 100 ng/ml α-GalCer in the presence of GolgiPlug. Cells were then harvested and surface stained with tetramer-PE and anti-B220–PerCP, followed by intracellular staining with anti–IFN-γ–FITC and anti–IL-4–allophycocyanin. Data are shown for B220–tetramer+ cells. Numbers indicate the percentage of cells within each quadrant. Results shown are representative of 2 independent experiments.