Fig. 2: The Scd1 deficient thymic environment imprints DN3 thymocytes with subsequent Treg differentiation propensity.

a, Schematic representation of the bone marrow chimera experiment. The bone marrow cells were isolated from age- and sex-matched CD45.1⁺CD45.2⁺Scd1^−/− mice and CD45.1⁺WT mice and mixed at a 1:1 ratio before intravenous administration to lethally irradiated (10 Gy) CD45.2⁺WT mice or CD45.2⁺Scd1^−/− mice. b, The proportions of Foxp3⁺ Treg cells among CD4⁺ T cells derived from different donors (n=9 mice for WR, n=7 mice for KR, repeated twice). c, Splenic CD4⁺CD62L⁺T cells from different donors were sorted and cultured under the Treg induction medium for 72 hours and Foxp3 levels were determined (n=4 biologically independent samples per group, repeated twice). d, Schematic representation of the adoptive transfer experiments investigating the generation of Treg cells from different subsets of WT and Scd1^−/− thymocytes. Each thymocyte population at the DN1, DN2, DN3, or DN4 stage from CD45.2⁺ Scd1^−/− mice and CD45.1⁺ WT mice was mixed at a 1:1 ratio and co-transferred to irradiated (3 Gy) CD45.1⁺CD45.2⁺WT mice via intrathymic injection. e, The Treg cells generated from different thymocyte subsets were analyzed flowcytometrically 4 weeks post transfer (n=5 mice for DN1, DN2 transfer; n=6 mice for DN3, DN4 transfer, performed once). f, Splenic CD4⁺CD62L⁺ T cells were isolated from the recipient mice to evaluate their differentiation efficiency to Treg cells in vitro (ns for DN1 and DN2 transfer, p=0.002 for DN3 transfer, p<0.0001 for DN4 transfer, n=5 biologically independent samples per group, performed once). Data are presented as Mean ± SEM. ns, no significance; by unpaired two-tailed Student’s t-test (b, c, f) or paired two-tailed Student’s t-test (e).