Fig. 1: CD301b⁺ lung cDC2s promote Treg differentiation and induce immunological tolerance.

a Timelines for mouse models of asthma. Mice were sensitized with OVA/HDE twice by o.p. aspiration and challenged once with OVA aerosol. BALF was harvested 48 h post- challenge. Tolerance assays were similar, except that mice were also exposed to OVA alone by o.p. aspiration on day -7. b Cell numbers for the indicated leukocytes in BALF, as determined by microscopy (No treatment n = 6, Sensitization n = 9, OVA pretreatment n = 12 biological replicates). c Activation of CD4⁺ T cells (left), representative cytograms of CD4⁺CD44^hi T cells (middle) and compiled data of Tregs (right) are shown. Naïve CD4⁺ T cells from Foxp3^eGFP OT-II mice were cocultured for 5 days with indicated cDC2 subsets isolated from C57BL/6 mice that received OVA by o.p. aspiration (n = 4 technical replicates). Gating strategy for purified cDC subsets and CD4⁺ T cell analysis (CD4⁺CD3ε⁺MHCII^–Live/Dead^–) is shown in Figure S1a and b. d Activation of CD4⁺ T cells (left) and Treg induction in CD44⁺ CD4⁺ T cells (right) by distinct cDC subsets isolated from mice that received OVA/HDE by o.p. aspiration (n = 5 technical replicates). Gating strategy for purified cDC subsets is shown in Fig. S1c. e Timeline for mouse model of asthma to test tolerogenic function of cDC2 subsets. CD301b⁺ or CD301b^– cDC2s were purified and incubated with OVA_323-339 peptides, then adoptively transferred by o.p. aspiration to C57BL/6 mice on day 0. After OVA/HDE sensitization and OVA challenge, cells in BALF were analyzed. f Cell numbers of the indicated leukocytes in BALF (No sensitization n = 6, No cDC transfer n = 12, CD301b⁺ cDC2 n = 16, CD301b^– cDC2 n = 8 biological replicates). b–d Data were analyzed by one-way ANOVA with Tukey’s multiple comparison test. (f) Data were analyzed by one-way ANOVA with Fisher’s LSD test. b, f Each dot represents an individual mouse. Combined results from two independent experiments are shown. (c, d) Each dot represents separately cultured CD4⁺ T cells. Representative results from 2 independent experiments are shown. Data are presented as mean values ± SEM. P values are indicated above the graphs. Source data are provided as a Source Data file. cDC conventional dendritic cells, Treg regulatory CD4⁺ T cells, OVA ovalbumin, HDE house dust extract, OP oropharyngeal, BALF bronchoalveolar lavage fluid.