Extended Data Fig. 3: DNFB model of non-specific skin inflammation and regulation of CD103⁺CD8⁺ T_RM cell conditioning by IL-4.

(a) Number of transferred CD45.1⁺CD44^hiCD8⁺ OT-I T cells recovered from the flank skin of congenic wild-type (WT) CD45.1⁺CD45.2⁺ C57Bl/6 recipient mice that were injected intravenously (i.v.) with in vitro-activated (1 μg/mL CD3 and CD28 antibodies) WT CD45.1⁺CD44^hiCD8⁺ OT-I T cells and immediately painted on the flank skin with DNFB. The cells were isolated at 3, 5, 8, 11 or 14 days post-DNFB treatment. Pooled data from 4 independent experiments; day 3 (n = 4), day 5 (n = 7), day 8 (n = 6), day 11 (n = 7) and day 14 (n = 5) mice. Mean ± SEM. (b) Mean percent CD103⁺ cells of CD45.1⁺CD8⁺CD44^hi OT-I T cells isolated from the flank skin as in a. Pooled data from 4 independent experiments; day 3 (n = 4), day 5 (n = 7), day 8 (n = 6), day 11 (n = 7) and day 14 (n = 5) mice. Mean ± SEM. (c) Representative contour plots of CD103 expression in CD45.1⁺CD44^hiCD8⁺ OT-I T cells isolated from the skin as in a. (d) Representative contour plots of CD103 expression in WT CD45.1⁺CD44^hiCD8⁺ OT-I and Il4^−/− CD45.2⁺CD44^hiCD8⁺ OT-I T cells isolated from the flank skin of congenic WT CD45.1⁺CD45.2⁺ C57Bl/6 recipient mice that were injected i.v. with a mix of in vitro-activated WT CD45.1⁺CD44^hiCD8⁺ OT-I and Il4^−/− CD45.2⁺CD44^hiCD8⁺ OT-I T cells and painted with DNFB as in a, at day 5 and day 30 post-transfer. (e) CD103 gMFI in (top) and mean percent (bottom) CD103⁺ cells of WT CD45.1⁺CD44^hiCD8⁺ OT-I and Il4^−/− CD45.2⁺CD44^hiCD8⁺ OT-I T cells isolated from the flank skin as in d. (f) Representative contour plots of WT CD45.1⁺CD44^hiCD8⁺ OT-I (black gates) and Il4^−/− CD45.2⁺CD44^hiCD8⁺ OT-I (orange gates) T cells isolated from the flank skin and spleen as in d. (g) Ratios of WT compared to Il4^−/− CD44^hiCD8⁺ OT-I T cells isolated from the bLN, spleen and skin as in d. Median with IQR. (h) Numbers of transferred WT and Il4^−/− CD44^hiCD8⁺ OT-I T cells recovered from the bLN, spleen and skin as in d. *p = 0.0371; **p = 0.0039; ***p = 0.002, two-tailed Wilcoxon test. Representative (d, f) or pooled (e, g, h) data. Day 5, one experiment (n = 3); day 30, three independent experiments (n = 10).

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