Fig. 4: Lineage-specific BMAL1 deficiency abrogates rhythms in DC lymphatic trafficking.
From: Circadian clocks guide dendritic cells into skin lymphatics
a–c, Ear CD11c+ cells (green) or BMDCs (orange) after crawl-in assays with lineage-specific Bmal1–/– animals; n = 4, 4, 3 and 3 mice (left) and 5, 5, 4 and 5 mice (right) (a); n = 4, 4, 4 and 5 mice (left) and 4, 4, 3 and 3 (right) (b); n = 4, 4, 5 and 5 mice (c); data are representative of two independent experiments; two-way ANOVA with Sidak post test. b,c, Representative ear whole-mount images from control and BMAL1ΔLEC (b) or BMAL1ΔcDC (c) animals. Scale bar, 50 µm. d, Ear BMDCs after crawl-in assays with synchronized control, Per1–/–Per2–/– or Bmal1–/– BMDCs; n = 6 mice for WT cells and n = 3 for knockout (KO) cells, representative of two independent experiments; two-way ANOVA with Sidak post test. e, Velocity of synchronized WT and Per1–/–Per2–/– BMDCs in collagen gel migration assays; n = 3 mice. Data are representative of two independent experiments; Kruskal–Wallis test with Dunn’s post test. f,g, Quantitative immunofluorescence screen of LYVE-1 (f), JAM-A, CD99 and CCL21 (g) in LYVE-1+ ear LVs; n = 5 mice from two independent experiments; two-way ANOVA with Sidak post test. h,i, Ear whole-mount imaging (h) and quantification (i) of CCL21 of WT and BMAL1ΔLEC ears; n = 5 mice from two independent experiments; Kruskal–Wallis test with Dunn’s post test. Dashed lines represent LYVE-1+ capillaries. Scale bar, 50 µm. j, Flow cytometric analysis of CCR7 on CD11c+MHCII+CD103–EpCAM– DCs in control and BMAL1ΔcDC animals; n = 7, 5, 6 and 4 mice from two independent experiments; two-way ANOVA with Sidak post test. k, Representative images and quantification of the distance of CD11c+ cells to LVs after ear crawl-in assays; n = 340, 431 and 575 cells from four mice from two independent experiments; unpaired Student’s t-test. Scale bar, 10 µm. Outline represents orthogonal LV view. l, Chromatin immunoprecipitation (ChIP) of BMAL1 binding to promoter regions of Ccl21, Ccr7 and Lyve1 compared to IgG controls; n = 6 mice from two independent experiments; two-way ANOVA with Sidak post test. All data are represented as mean ± s.e.m.
