Figure 1.

RelB^DCko mice show an increased frequency of Langerhans cells (LCs) and Langerin⁻ dermal DCs (dDCs) among steady-state migratory DCs in skin-draining lymph nodes. (A,B) Cells from peripheral LN (PLN) of control and RelB^DCko mice were stained for B220, CD11c, and CD40 and analyzed by flow cytometry. (A) Contour plots represent CD11c⁺ B220⁻ cells within a FSC/SSC gate for live cells. Based on the expression of CD40 migratory CD40^hi dendritic cell (DC) and resident CD40^low DC are distinguished among CD11c⁺ B220⁻ cells. B220⁻ CD11c^low CD40^low cells represent RelB⁻ macrophages. Numbers indicate the percentages of cells within the gate. (B) Statistical summary of frequencies of migratory and resident DC among CD11c⁺ B220⁻ cells PLN of RelB^DCko (n = 11) or control mice (n = 7). Percentages of both DC subsets among CD11c⁺ B220⁻ cells were calculated in relation to the absolute frequency of CD11c⁺ B220⁻ cells. Each symbol represents an individual mouse and black lines indicate the mean values. Statistical analyses were performed using the Mann–Whitney Test: ns, not significant, *p < 0.05. (C) Representative flow cytometric analysis of migratory DC subpopulation in PLN of control and RelB^DCko mice after surface staining of CD11c, CD40, and CD103 and intracellular staining of Langerin. After gating specifically on CD11c⁺ B220⁻ cells (right panel), migratory CD40^hi DCs (middle panel) were further analyzed for their expression of Langerin versus CD103. Three migratory DC subsets could be defined: Langerin⁺ CD103⁻ LC, Langerin⁺ CD103⁺ dDC, and Langerin⁻ CD103^+/− dDC (left panel). Numbers indicate the percentages of cells within each gate. (D) Frequencies of Langerin⁺ CD103⁻ LC, Langerin⁺ CD103⁺ dDC, and Langerin⁻ CD103^+/− dDC among the total percentages of CD40^hi CD11c⁺ B220⁻ cells in PLN of control (n = 7) and RelB^DCko mice (n = 7) are shown. Data represent the mean values + SD from two independent experiments with 3–4 mice per genotype and each experiment. Statistical p-values were obtained by performing two-tailed unpaired Student’s t-test: ns, not significant, *p < 0.05, **p < 0.01.