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. 2018 Dec 11;9:5279. doi: 10.1038/s41467-018-07685-x

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© The Author(s) 2018

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. 3 — Embryo-derived spleen CD11b^lo F4/80^hi macrophages depend on CSF1R-mediated signals for their generation. a Photograph of E10.5 embryo with yolk sac (YS). Scale bar corresponds to 1 mm. b, c Dot plots show yolk sac (b) or body cells excluding head tissues (c) analyzed for the expression of CD45 and KIT (left). CD45-expressing cells are further resolved for the expression of CD11b and CSF1R (second left). CSF1R⁺ cells co-express F4/80 (third left). Plots show the frequencies of F4/80-expressing cells within the CSF1R-positive population (second right). Each dot represents one mouse. CD45-negative cells are resolved for the expression of F4/80 and CSF1R (right). d Schematic outline of TAM-induced depletion of CSF1R expression in the embryo (top). Contour plots resolve CD45⁺ fetal liver cells from E14.5 R26-CreER^T2+;Csf1r^F/⁻ and control embryos for the expression of CD11b and F4/80 (bottom). e Plots show the frequencies of CD11b⁺ F4/80^lo (left) or CD11b^lo F4/80^hi (middle) cells in the fetal liver of R26-CreER^T2+;Csf1r^F/⁻ and control embryos. Right plot shows fold-change differences of leukocyte and macrophage populations in the fetal livers. Data are summarized from three independent experiments. Two-sided t test (left) and Mann–Whitney U test (right) were performed. SD is shown. f Schematic outline of TAM-induced depletion of CSF1R expression in R26-CreER^T2+;Flt3^−/−;Csf1r^F/⁻ and control embryos. TAM was administered at E10.5. g Frequencies (left) and fold-change differences (right) of leukocytes and RP-Mps in the spleens of R26-CreER^T2+;Flt3^−/−;Csf1r^F/⁻ and control pups at E19.5. Data are summarized from four independent experiments. Two-sided t test (left) and Mann–Whitney U test (right) were performed. SD is shown. h Dot plots show DCs in R26-CreER^T2+;Flt3^−/−;Csf1r^F/⁻ and control mice at E19.5. Numbers indicate frequencies of dendritic cells within Dapi⁻ cells. i, j Plots show DC frequencies and fold-change differences of leukocytes and DCs in the spleen of E19.5 pups (i) or at 3 weeks of age (j) in R26-CreER^T2+;Flt3^−/−;Csf1r^F/⁻ and control mice that were TAM-induced at E10.5. Data are summarized from three independent experiments. Two-sided t tests (left) and Mann–Whitney U tests (right) were performed. SD are shown