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. 2019 Apr 15;10:1760. doi: 10.1038/s41467-019-09748-z

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© The Author(s) 2019

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. 2 — Lineage tracing Pdgfrα⁺ mesenchymal cells reveals GFP-labeled progeny within the mammary epithelium. a Lineage tracing model used to determine stromal cell dynamics in the mammary gland. b Whole mount confocal images of native GFP (green) and tdTomato (red) fluorescence in the mammary fat pad (left) and epithelia (right; E+P treated) of adult PdgfrαCre R26mTmG mice; scale bar = 50 µm. c GFP, tdTomato (left) and PDGFRα (magenta), DAPI (blue) (right) immunostaining spanning several epithelial structures in PdgfrαCre R26mTmG mammary tissue derived from b; scale bar = 200 µm. d High magnification image of PDGFRα, GFP and tdTomato in the fat pad and glandular regions from an E+P treated mouse (representative of n = 5 adult mice and 5 fields per tissue section); scale bar = 25 µm. e FACS profiles of GFP⁺ and tdTomato⁺ cells in mammary subpopulations: stromal, basal, luminal and luminal subsets enriched for hormone receptor (HR) positive (Sca1⁺) and negative (Sca1^-) cells of adult mammary glands isolated from untreated (n = 3) or E+P treated (n = 2) mice; f Quantification of GFP⁺ and tdTomato⁺ cells in stromal (S), basal (B) and luminal (L) subpopulations shown in e. g Droplet digital PCR analysis of Pdgfrα transcripts normalized to Gapdh in FACS-sorted mammary cell fractions from lineage tracing mice (n = 4 per group). h Immunofluorescent images of PdgfrαCre R26mTmG mammary tissue sections from pubescent and prepubescent mice (imaging of n = 3 mice per stage and 5 fields per tissue section); scale bar = 25 µm. Quantification of GFP⁺ and tdTomato⁺ cells in mammary subsets of i pubescent (n = 3) and j prepubescent (n = 3) mice. PDGFRα⁺ cells (magenta), GFP⁺ progeny (green) and epithelial EpCAM (red) in the embryonic mammary gland at embryonic day 18 k and day 13 l (n = 3 mice each, 3 fields per tissue section); scale bar = 25 µm. White arrowheads indicate GFP⁺ cells. White asterisk denotes associated Supplementary Movie 1. Data represent mean ± s.e.m. *p< 0.05, **p<0.01,***p<0.001 (Statistical analysis by one-way ANOVA in g and t-test in f, i and j). Source data are provided as a Source Data file