Extended Data Fig. 13: Lineage tracing of leptomeningeal CCR7+ dendritic cells and characterization metastatic cancer cells in Egfp- and Ifng-overexpressing mice.

(a) Proportion of leptomeningeal, CD11c⁺ MHC II⁺ double-positive cDC in LLC LeptoM-bearing Ifngr1-proficient and deficient mice. (b) Proportion leptomeningeal, IL12-producing CCR7+ DCs in LLC LeptoM-bearing Ifngr1-proficient and deficient mice. (c) Proportion of mCherry⁺ CCR7+ DCs across various conditions. mCherry reports the expression of Xcr1 and shows that only a minority of CCR7+ DCs expressed mCherry and originated in cDC1 subtype. (d) Classification of leptomeningeal, CD11c⁺ MHC II⁺ double-positive cDC into immature (CCR7-) and mature (CCR7+) subtypes across various conditions. (e) Proportion of Xcr1⁺ CCR7+ DCs across various conditions, confirming lineage tracing from Extended Data Fig. 12. (f) Schematic shows experimental design of strategy used to determine the cranial or extracranial origin of leptomeningeal immune cells. CD45.2 mice were irradiated with their cranium shielded and infused with CD45.1 bone marrow that colonized the depleted bones. CD45.2⁺ immune cells would after recovery predominantly originate in the shielded cranial bone marrow, while CD45.1⁺ immune cells would arrive from peripheral sites. (g) Quantification of bone marrow reconstitution. Cells in femoral bone marrow (BM) and circulation are mainly CD45.1 + , while cells isolated from cranial BM are CD45.2. (h) Determination of cDC subtype origin. Unlike neutrophils that are mostly sourced from proximal cranial BM, cDC are colonizing leptomeninges in naïve and LLC LeptoM-bearing animals from peripheral BM. (i) Quantification of cancer cells captured in the mouse single-cell atlas (n = 3,718 keratin⁺ CD63⁺ cells isolated from n = 6 mice per group); related to Fig. 4j. (j) GSEApy analysis of top 15 Reactome 2022 pathways enriched in cancer cells isolated from Ifng-overexpressing animals and subsetted as described in Fig. 4j (DEG cut-off P < 0.01). (k) GSEApy analysis of top 15 Reactome 2022 pathways enriched in cancer cells isolated from Egfp-overexpressing animals and subsetted as described in Fig. 4j (DEG cut-off P < 0.01). (l) Quantification of cleaved Caspase 3-positive cells in cancer plaques and clusters, in the leptomeninges of Egfp- or Ifng-overexpressing animals injected with LLC LeptoM cells. (m) Quantification of cleaved Caspase 3-positive cells in cancer plaques and clusters, in the leptomeninges of Egfp- or Ifng-overexpressing animals injected with E0771 LeptoM cells. (n) Quantification of cleaved Caspase 3-positive cells in cancer plaques and clusters, in the leptomeninges of Egfp- or Ifng-overexpressing animals injected with B16 LeptoM cells.

Source Data