Extended Data Fig. 6: Extended analysis of OT-II CD45.1 cell seeding and fate in gLNs upon intestinal OVA injection or infection.

a–h, Frequency of FOXP3⁺RORγT^– (a), FOXP3⁺RORγT⁺ (b), FOXP3^–RORγT⁺ (c), FOXP3⁺CD25⁺ (d), or FOXP3^–CD25⁺ (e) cells among CD45.1⁺ OT-II cells, frequency of CD45.1⁺ OT-II cells among TCRβ⁺CD4⁺ cells (f), division index (g) and total TCRβ⁺CD4⁺ cell counts (h) in indicated gLNs of mice 48 h after duodenal or ileal injection with OVA or OVA–CT, performed 16 h after adoptive transfer of 1 × 10⁶ naive OT-II cells into SPF CD45.2 C57BL/6 host mice (n = 3). i–l, Survival curve (sham n = 5, duo n = 6, ile n = 8; i), and frequency in ileal lamina propria of FOXP3^–RORγT⁺CD45.1⁺ (j), FOXP3^–RORγT⁺CD45.1⁺ (k) and FOXP3⁺RORγt^–CD45.1⁺ (l) cells among TCRβ⁺CD4⁺ cells (n = 4 per group) of mice infected with Stm-OVA 9 days after duodenal or ileal injection with OVA–CT versus sham operation, and 10 days after adoptive transfer of 1 × 10⁵ naive OT-II cells into SPF CD45.2 C57BL/6 host mice. j–l, Intestines removed 48 h after infection. *P < 0.05 (Mantel–Cox and Gehan–Breslow–Wilcoxon, i), ^##P < 0.005 ⁽logrank, i), *P < 0.05 (ANOVA, j–l). m–p, Flow cytometry plot of CD25⁺ and CFSE (m), frequency of CD451⁺ OT-II cells among TCRβ⁺CD4⁺ cells (n), CD45.1⁺ division index (o) and total CD45.1⁺ cell counts (p) in indicated gLNs 64 h after adoptive transfer of 1 × 10⁶ naive CD45.1⁺ OT-II cells into SPF CD45.2 C57BL/6 mice infected with Citro-OVA 9 days earlier, and after gavage with OVA or PBS 48 h and 24 h before analysis (n = 5). Data representative of two independent experiments. q–u, Frequency of total FOXP3⁺ (q), FOXP3^–CD25⁺ (r), and FOXP3^–RORγT⁺CD451⁺ (s) cells among CD45.1⁺ cells, frequency of CD45.1⁺ cells among TCRβ⁺CD4⁺ cells (t), and frequency of FOXP3^–RORγT⁺CD45.2⁺ cells among TCRβ⁺CD4⁺ cells (u) in indicated gLNs 64 h after adoptive transfer of 1 × 10⁶ naive CD45.1⁺ OT-II cells into SPF CD45.2 C57BL/6 host mice infected or not with Citrobacter 9 days earlier, and after gavage with OVA 48 h and 24 h before analysis (n = 5). ns, not significant in ANOVA or one-tailed t-test comparing gLNs of infected versus non-infected mice. *P < 0.05, **P < 0.01, ***P < 0.005 (ANOVA); ^#P < 0.05 (t-test).