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 9

Bacteria-induced iNKT cell hyporesponsiveness can be overcome by treatment with PMA plus ionomycin or with α-GalCer plus IL-2.

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Bacteria-induced iNKT cell hyporesponsiveness can be overcome by treatme...
(A) Spleen cells were prepared from mice treated 4 weeks earlier with α-GalCer or the indicated bacteria, cultured in vitro (2 × 105 per well) for 6 hours in plain medium (alone), 100 ng/ml α-GalCer, or a combination of 10 ng/ml PMA and 1 μM ionomycin (PMA + IONO), in the presence of Golgi­Plug to allow intracellular accumulation of cytokines. 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. Data shown are representative of 3 independent experiments with 2 mice in each group per experiment. (B) IL-2 overcomes the proliferative defect of hyporesponsive iNKT cells in vitro. Spleen cells from naive mice or from mice injected 1 month earlier with α-GalCer or heat-killed E. coli were labeled with CFSE. Cells (2 × 105 per well) were then cultured with α-GalCer (100 ng/ml) for 24 hours in the presence or absence of IL-2 (10 ng/ml). Cells were then washed and cultured for an additional 96 hours without α-GalCer in the presence or absence IL-2. At the end of the culture period, cells were harvested, stained with tetramer-allophycocyanin and anti-B220–PerCP, and analyzed by flow cytometry. CFSE dilution was analyzed on B220–tetramer+ cells. Representative data from 2 independent experiments are shown.