Figure 4. Lung-resident dendritic cells inhibit metastasis of B16 melanoma.

a, Maximum intensity projection of LIVM of B16ZsGreen tumour 24 h after injection into CD11c-mCherry, MacBlue host (see also Supplementary Video 9). Representative of 5 mice. b, High magnification showing tumour-loaded MacBlue⁺ cells in close interaction with B16ZsGreen cell at 24 h after injection. c, Representative image from time-lapse. Track overlays describe motion of tracked MacBlue⁺ cells over a period of ~1 h. d, Overlay of 10 tracks of MacBlue⁺ cells from c. e, Staining of ZsGreen⁺ and ZsGreen⁻ macrophages 24 h after tumour injection (n = 4), ISO represents staining with isotype control antibody. f, Gating of lung-draining mLN 72 h after injection. Contour lines, total LN; green dots, ZsGreen⁺ cells. g, Frequency of CD8⁺, CD11b⁺ and CD103⁺ DCs in total ZsGreen⁺ cells in the mLN 72 h after injection. (n = 6; *P < 0.05, one-way ANOVA with Bonferroni post-hoc test, error bars are standard deviation). h, Representative image of ZsGreen⁺CD11c-mCherry⁺ APCs interacting with GFP-labelled OT-I T cells in a mLN 72 h after injection with B16ZsGreenSL8 (see also Supplementary Video 10). Inset highlights two ZsGreen-bearing CD11c⁺ cells in close interaction with OT-I T cells (n = 3 mice). i, Representative images of metastasis-bearing lungs from wild-type (WT) or CCR2-knockout (KO) mice 2 weeks after injection with B16ZsGreen. j, Total number of lung metastases (mets) in wild-type and CCR2-knockout mice 2 weeks after injection with B16ZsGreen (n = 7 per group; *P < 0.05, unpaired t-test, horizontal bars are mean values). k, Frequency of ZsGreen⁺ macrophages in lungs of wild-type and CCR2-knockout mice 24 h after injection with B16ZsGreen (n = 4 per group; *P < 0.05 by unpaired t-test, horizontal bars are mean values). l, Frequency of ZsGreen⁺ CD103⁺ cDCs in lungs of wild-type and CCR2-knockout mice 24 h after injection with B16ZsGreen (n = 4 per group; *P < 0.05, unpaired t-test, horizontal bars are mean values). m, n, Frequency (m) and absolute number (n) of CD8⁺ T cells in lungs of wild-type or CCR20-knockout mice from k (n = 4 per group; *P < 0.05, unpaired t-test, horizontal bars are mean values). o, Representative images of lungs from Zbtb46-DTR bone marrow chimaeras treated with phosphate buffered saline (PBS) or diphtheria toxin (DT). Lungs collected 2 weeks after injection with B16ZsGreen. p, Total number of lung metastases in PBS- or DT-treated Zbtb46-DTR bone marrow chimaeras 2 weeks after injection with B16ZsGreen (n = 5 for PBS and 10 for DT groups, respectively; *P < 0.05, unpaired t-test, horizontal bars are mean values).