Figure 4. NKT cells exhibit a glutamine-addicted phenotype.

(A) The graph shows hexokinase 2 (HK2) expression in NKT and CD4 T cells with and without stimulation (n = 3).

(B) Heatmap shows relative levels of the indicated PPP metabolites in resting NKT cells compared with resting CD4 T cells analyzed after LC-MS/MS analysis (n = 3).

(C) Heatmap shows PPP metabolites in NKT cells with and without stimulation as analyzed by LC-MS/MS (n = 3).

(D and E) Cell survival and proliferation of sorted NKT cells stimulated in the presence or absence of EGCG (20 μM) with or without sodium pyruvate (1 mM) or DMαKG (1.5 mM) (n = 3).

(F and G) WT and GLS1 KO mice were injected with αGalCer (5 μg/mouse), and splenic NKT cells were sorted on day 3 of activation. The representative graphs show ECAR using the glycolytic stress test (F) and OCR using the Mito Stress test (G) in Seahorse assay (n = 6 replicates per group pooled from two mice). The graph in (G) shows basal respiration (BR), maximum respiration capacity (MRC), and reserve capacity (RC) (n = 3).

(H and I) NKT from WT and GLS1 KO mice were stimulated in the presence or absence of DMαKG (1.5 mM). (H) Cell survival and proliferation of NKT cells are shown (n = 3). (I) Representative histograms and summary graphs show mitochondrial mass and membrane potential (n = 3). All relative levels were calculated using the average control value as a reference point. All data are representative of or combined from at least two or three different experiments. Seahorse data are shown as mean ± SD. All other data are shown as mean ± SEM. *p < 0.05, **p < 0.01, ****p < 0.0001.