Figure 4

YY1^lo NKT cells produce less IL-4 and IFN-γ as compared to YY1^hi NKT cells. (a–d) C57BL/6J (WT) mice were injected I.V. with either α-GalCer (10ug) suspended in saline or saline only (control). 90 minutes after injection, splenic NKT cells were harvested and stained for IL-4 and IFN-γ. (a) Representative FACs plot for IL-4 and IFN-γ expression in each WT activated NKT cell (MHCII⁻, CD3⁺ CD1dtet⁺ CD24⁻) subset: YY1^lo PLZF^lo, YY1^hi PLZF^lo, and YY1^hi PLZF^hi. (b) Cumulative data from 6 mice, depicting the % of IL-4⁺ and IFN-γ ⁺ double positive cells. The IL-4 MFI of IL-4⁺ cells (d) and IFN-γ MFI of IFN-γ⁺ cells (d) from each NKT cell subset is plotted. The IL-4 MFI and IFN-γ MFI of each experimental sample was normalized to the IL-4 MFI and IFN-γ MFI of control, unactivated NKT cells stained in the same experiment. (e–f) NKT cells were isolated from the spleens of WT mice and activated in vitro for 5 hours by culture with PMA (50 ng/mL) and Ionomycin (500n/mL). Activated NKT cells were stained for IL-4, IFN-γ, and YY1. (e) Representative histograms and (f) and aggregate data compare the production of IFN-γ (left) and IL-4 (right) between YY1^lo and YY1^hi NKT cells following in vitro activation. Data are representative of 4 (a–d) or 3 (e–f) independent experiments. Each dot represents one mouse, N = 6 for (b–d), N = 3 for (f). The horizontal lines indicate the mean (±s.e.m.). *P < 0.05 **P < 0.01, ***P < 0.001 determined by one-way ANOVA (b–d).