Fig 2: IFN? induces upregulation of MHC-II in LECs in vitro and in vivo in the tumor periphery. (A–D) Relative mRNA expression of HLA-DRA, HLA-DM, PD-L1, LYVE-1 in cultured human dermal LECs after 24-h treatment with (A) IFN? (100 ng/mL), (B) VEGF-C (100 ng/mL), (C) IL-2 (10 ng/mL) and (D) TNF-a (10 ng/mL). (E) Representative FACS histogram plot and (F) quantification of MHC-II and PD-L1 positive human dermal LECs in culture after 24-hour treatment with IFN? (100 ng/mL) as measured by flow cytometry. (G) Representative FACS histogram plots and (H) quantification of MHC-II and PD-L1 positive peritumoral LECs in WT mice treated with IFN? neutralizing antibody or IgG isotype control at day 14 post tumor inoculation. Statistical significance assessed using unpaired student’s t test on data from at least 3 biological replicates for cell culture studies and at least 4 biological replicates for mouse experiments; statistical analysis not performed on (B–D) for which at least 2 biological replicates were used; error represented as SD. *, p < 0.05; **, p < 0.01; ***, p < 0.001. n.d, not detected mRNA in treated group.

Fig 3: Conditional knock-out of LEC-specific MHC-II expression attenuates growth of heterotopically implanted B16F10 and E0771 tumors. (A) Schematic representation of mice genotype bred to generate LECMHC II-/- mice. (B) Relative mRNA expression of MHC-II in FACS-sorted lymph node LECs (CD45-podoplanin+CD31+) and FACS-sorted lymph node CD11c+ and CD11b+ immune cells from WT and LECMHC-II-/- mice. (C) Representative FACS histogram plots of MHC-II MFI in sorted CD11c+ and CD11b+ immune cells from WT and LECMHC-II-/- mice. (D) Representative FACS histogram plots and (E) quantification of MHC-II and PD-L1 positive lymph node LECs in WT and LECMHC-II-/- mice. (F) Representative FACS histogram plots and (G) quantification of MHC-II and PD-L1 positive peritumoral LECs in WT and LECMHC-II-/- mice bearing B16F10 at 2 weeks post tumor inoculation. (H) Gross images of resected B16F10 tumors and their respective (I) mass in grams in WT and LECMHC-II-/- mice at 2 weeks post tumor inoculation. Statistical significance assessed using unpaired student’s t test on data from at least 4 biological replicates; error represented as SD. *, p < 0.05; **, p < 0.01. (J,K) Growth curves of B16F10 (G) and E0771 (H) in WT and LECMHC-II-/- mice at various time points (in weeks). Statistical significance assessed using two-way ANOVA, error represented as SD.; **, p < 0.01; ***, p < 0.001.