Figure 6. IL-4 producing iNKT cells are stimulated by self ligands.

(a) BM derived DCs pulsed with solvent (gray), βGlcCer (thick) or αGalCer (thin) were injected into B6 Nur77^GFP mice and splenic iNKT cells were analyzed 16 hours later. Data are representative of two independent analyses with a total of 4 mice per group. (b) Thymocytes from B6 and BALB/c Nur77^GFP mice were analyzed for expression of GFP in each iNKT subset. Data are representative of 3 independent experiments. (c) GFP expression in BALB/c Nur77^GFP KN2^+/− mice shows IL-4 producing NKT2 cells (hCD2⁺) express a higher level of GFP than hCD2 negative NKT2 cells. Representative data from 3 independent experiments is shown. (d) Gene expression of Nur77 was compared to IL-4 in sorted NKT2 cells (mRNA arbitrary units normalized to Gapdh). Linear regression was used to calculate goodness of fit(R²). (e) hCD2 expression is dependent on sustained TCR signaling. Thymic iNKT cells of BALB/c KN2^+/− mice were MACS enriched by depleting CD8⁺ and CD24⁺ cells, labeled with VCT and injected into WT BALB/c (N=5) or BALB/c CD1d KO (N=5) mice. Six days later, hCD2 expression in NKT2 cells was analyzed. MFI, mean fluorescence intensity. An unpaired two tailed t-test was used to compare hCD2 MFI in WT or CD1d KO hosts. ^***p=0.0004. (f) iNKT subsets have distinct TCR repertoires. TCR Vβ7⁺ usage is shown among NKT1, NKT2 and NKT17 cells in B6 mice (N=6). One-way ANOVA was used to compare the frequency of NKT1, NKT2 and NKT17 cells. ***p<0.001, *p<0.05.