Tumorigenic and metastatic potential of CD44/Sca1+ cells. CD44/Sca1+ and CD44/Sca1- cells were obtained by cell sorting from mammary epithelial cells (MECs), tumor cells or the lungs of mice transgenic for PyMT and MUC1 antigen (MMT) that also express green fluorescent protein (GFP MMT) at different stages of tumor development and injected into the mammary fat pads of wild type (WT) mice at the doses indicated (if not indicated in this way, 8 × 104 cells were injected). Tissue samples were obtained from WT mice injected with either sorted CD44/Sca1+ cells [(+)] or CD44/Sca1- cells [(-)]. (a) Comparison of tumorigenic potential between CD44/Sca1+ and CD44/Sca1- tumor cells. The tumor incidence and tumor size are presented for each group of mice inoculated with the indicated numbers of cells. (b) Mammary tumor from a WT mouse inoculated with CD44/Sca1+ tumor cells was viewed by H&E staining (left panel) and examined for GFP (middle panel) or CD44 (right panel) expression using immunohistochemical (IHC) staining. (c) Sections of mammary glands from WT mice inoculated with CD44/Sca1+ tumor cells (> 120 days, upper panels) or MECs (< 40 days, lower panels) were stained with H&E and examined for expression of GFP and ESA or CD44. (d) Quantification of disseminated cells. GFP-positive cells were gated from lung, lymph node cells (LNC) or blood cells of recipient mice inoculated with CD44/Sca1+ and CD44/Sca1- cells from tumor cells or MECs of GFP MMT mice, and stained with anti-ESA monoclonal antibodies and analyzed by FACS. (e) Tumors from the lungs of WT mice inoculated with CD44/Sca1+ lung cells from GFP MMT mice at more than 120 days were viewed by H&E staining and examined for GFP and estrogen receptor (ER) expression. (f) Quantification of disseminated cells. GFP-positive cells were gated from lung, LNC or blood cells of recipient mice inoculated with CD44/Sca1+ and CD44/Sca1- cells from GFP MMT mice at the indicated ages, stained with anti-ESA monoclonal antibodies and analyzed by FACS. (d and f) The average ± standard deviation of ESA/GFP-positive cells in total blood, LN or lung cells were presented.