Glycolipid antigen induces long-term natural killer T cell anergy in mice
Vrajesh V. Parekh, Michael T. Wilson, Danyvid Olivares-Villagómez, Avneesh K. Singh, Lan Wu, Chyung-Ru Wang, Sebastian Joyce, Luc Van Kaer
Vrajesh V. Parekh, Michael T. Wilson, Danyvid Olivares-Villagómez, Avneesh K. Singh, Lan Wu, Chyung-Ru Wang, Sebastian Joyce, Luc Van Kaer
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Research Article Immunology

Glycolipid antigen induces long-term natural killer T cell anergy in mice

Abstract

Natural killer T (NKT) cells recognize glycolipid antigens presented by the MHC class I–related glycoprotein CD1d. The in vivo dynamics of the NKT cell population in response to glycolipid activation remain poorly understood. Here, we show that a single administration of the synthetic glycolipid α-galactosylceramide (α-GalCer) induces long-term NKT cell unresponsiveness in mice. NKT cells failed to proliferate and produce IFN-γ upon α-GalCer restimulation but retained the capacity to produce IL-4. Consequently, we found that activation of anergic NKT cells with α-GalCer exacerbated, rather than prevented, B16 metastasis formation, but that these cells retained their capacity to protect mice against experimental autoimmune encephalomyelitis. NKT cell anergy was induced in a thymus-independent manner and maintained in an NKT cell–autonomous manner. The anergic state could be broken by IL-2 and by stimuli that bypass proximal TCR signaling events. Collectively, the kinetics of initial NKT cell activation, expansion, and induction of anergy in response to α-GalCer administration resemble the responses of conventional T cells to strong stimuli such as superantigens. Our findings have important implications for the development of NKT cell–based vaccines and immunotherapies.

Authors

Vrajesh V. Parekh, Michael T. Wilson, Danyvid Olivares-Villagómez, Avneesh K. Singh, Lan Wu, Chyung-Ru Wang, Sebastian Joyce, Luc Van Kaer

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α-GalCer induces NKT cell anergy. Naive mice or mice injected with α-Gal...
α-GalCer induces NKT cell anergy. Naive mice or mice injected with α-GalCer (5 μg/mouse, i.p.) 1 month earlier were reinjected with α-GalCer or vehicle. (A) NKT cell population dynamics. Single-cell suspensions of the spleen were prepared, stained with anti–TCR-β–FITC, tetramer-PE, and anti-B220–PerCP, and analyzed by flow cytometry. Numbers indicate the percentage of TCR-β+tetramer+ cells among B220– cells. (B) Graphical representation of the data from A. (C) Expression of activation markers by NKT cells. Expression of CD69 and CD40L by NKT cells was evaluated at 24 hours and 4 hours, respectively. (D) NKT cell cytokine production. Spleen cells were prepared at the indicated time points, and 2 × 105 cells were cultured for 6 hours in plain medium (alone), 100 ng/ml α-GalCer, or a combination of PMA and ionomycin (PMA+IONO), in the presence of GolgiPlug (BD Biosciences) 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–IL-4–allophycocyanin and anti–IFN-γ–FITC. Data are shown for B220–tetramer+ cells. Numbers indicate the percentage of cells within each quadrant. Results shown are representative of 3 individual experiments.