Impact of bacteria on the phenotype, functions, and therapeutic activities of invariant NKT cells in mice
J. Clin. Invest. Sungjune Kim, et al. 118:2301 doi:10.1172/JCI33071 [
Go to this article.]
Figure 3Heat-killed
E. coli induces hyporesponsiveness of iNKT cells to α-GalCer rechallenge ex vivo.
(
A) In vivo dynamics of the iNKT cell response. Mice were injected with α-GalCer (5 μg/mouse, i.p.) or heat-killed
E. coli and sacrificed at the indicated time points, and spleen and liver mononuclear cells were prepared and stained for the identification of iNKT cells. (
B) Total spleen iNKT cell counts and percentage of liver iNKT cells at the indicated times, for a total of 5–10 mice per group, pooled from 2 separate experiments. *
P < 0.05 compared with naive animals. (
C) α-GalCer recall response of mice at the indicated times after treatment with heat-killed
E. coli. Mice were injected with α-GalCer or heat-killed
E. coli and sacrificed 3, 4, or 6 weeks later, and splenocytes were cultured with graded doses of α-GalCer. After 3 days, proliferation was assessed, and culture supernatants were evaluated for IL-4 and IFN-γ. *
P < 0.05 compared with naive splenocytes. (
D) Proliferative defect in iNKT cells from mice treated with heat-killed
E. coli. Spleen cells from naive mice or from mice injected 4 or 6 weeks earlier with α-GalCer or heat-killed
E. coli were labeled with CFSE. Cells were then cultured with α-GalCer (100 ng/ml) for 24 hours, then washed and cultured for an additional 96 hours without α-GalCer. At the end of the culture period, cells were harvested, stained, and analyzed by flow cytometry. (
E) iNKT cell cytokine production. Spleen cells were prepared at the indicated time points, and cells were cultured for 6 hours in vehicle or 100 ng/ml α-GalCer in the presence of GolgiPlug. Cells were then harvested and stained.
