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. 2022 Nov 22;13:7149. doi: 10.1038/s41467-022-34791-8

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© The Author(s) 2022

Open Access This article is licensed under a Creative Commons Attribution 4.0 International License, which permits use, sharing, adaptation, distribution and reproduction in any medium or format, as long as you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons license, and indicate if changes were made. The images or other third party material in this article are included in the article’s Creative Commons license, unless indicated otherwise in a credit line to the material. If material is not included in the article’s Creative Commons license and your intended use is not permitted by statutory regulation or exceeds the permitted use, you will need to obtain permission directly from the copyright holder. To view a copy of this license, visit http://creativecommons.org/licenses/by/4.0/.

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Fig. 7 — a Flt3L+NDV-treated GFP⁺ A20-tumor-bearing Wt or Batf3^−/− mice were followed for tumor growth (mean ± s.e.m) and survival (untreated, n = 12; Flt3L+NDV Wt, n = 11; Flt3L+NDV Batf3^−/−, n = 11). Two-way ANOVA with Tukey’s multiple comparisons test (tumor size) and Log-rank (Mantel-Cox) test (survival). b, c Tumors/TdLNs from treated GFP⁺ A20-tumor-bearing mice were analyzed by spectral flow cytometry. Stacked bar graphs and representative contour plots of cDC1, cDC2, DN cDCs or pDCs (b) and stacked bar graphs of CD86/CD40 expression (vs Untr) in all cDCs (c). b, c One-way ANOVA with Tukey’s multiple comparisons test; n = 5 mice per group. d, e GFP⁺ A20-tumor-bearing Wt or Batf3^−/− mice were treated with Flt3L+NDV. Anti-GFP CD45.1⁺CD8⁺ (JEDI) T cells were adoptively transferred and tumoral/TdLN T cells were analyzed after 5 days. IFN-γ, TNF and Tbet expression in intratumoral CD8⁺ T cells (Untr; n = 5, Flt3L+NDV Wt, n = 4, Flt3L+NDV Batf3^−/−, n = 5) (d) and representative contour plots and bar graphs showing IFN-γ in JEDI T cells from TdLNs (after completed treatment) upon GFP-peptide stimulation ex vivo (e) (Untr; n = 4, Flt3L+NDV Wt, n = 6, Flt3L+NDV Batf3^−/−, n = 3). f Mice in (a) were analyzed for blood anti-GFP tetramer⁺CD8⁺ T cells 7 days after completed treatment. Kruskal-Wallis with Dunn’s multiple comparisons test; n = 12 (untr, Flt3L+NDV Wt) and n = 6 (Flt3L + NDV Batf3^−/−). g TdLN cells from tumor-bearing Flt3L+NDV-treated mice were co-cultured with Lrrk1_mut peptide-pulsed DCs; T cells were analyzed after 24 h. IFN-γ in CD44⁺PD1⁺CD8⁺ T cells, representative from two A20- and three GFP⁺ A20-bearing mice. h GFP⁺ A20-tumor-bearing Wt mice treated with anti-CD8, anti-CD4, anti-IFNAR or isotype-control antibodies were treated with Flt3L+NDV and followed for tumor growth (mean ± s.e.m) and survival. Two-way ANOVA with Sidak’s multiple comparisons test (tumor size) and Log-rank (Mantel-Cox) test (survival); n = 8 (IgG2b), n = 9 (IgG1, anti-CD8), n = 10 (anti-CD4, anti-IFNAR). i TdLNs from mice treated as in (g) with anti-CD4 or isotype-control antibodies and Flt3L+NDV were co-cultured with unstimulated (No pept) or GFP-peptide pulsed (GFP-pept) DCs and analyzed after 24 h. Representative contour plots and quantification of IFN-γ in CD44⁺PD1⁺CD8⁺ T cells. One-way ANOVA with Tukey’s multiple comparisons test, n = 4 mice per group. Data show mean ± SD.