Figure 3

Homogeneous ER ⁺PR ⁺CK5 ⁻luminal E3 and MCF-7 cells generate heterogeneous bone metastases containing ER ⁻PR ⁻CK5 ⁺luminobasal cells. (A) Representative hematoxylin and eosin (H&E) and immunohistochemistry (IHC) of control (C) bones lacking metastases despite intracardiac (IC) injection of E3 or MCF-7 cells. H&E: (B) bone, (BM) marrow. Scale bars: 50 μm. IHC: estrogen receptor (ER) or progesterone receptor (PR) (green), cytokeratin 5 (CK5) (red), and 4′,6-diamino-2-phenylindole (DAPI) (blue) by immunofluorescent (IF) staining (n = 5 per group). Scale bars: 20 μm. (B) H&E and IHC of E- or E+P-treated mice with bones containing metastases after IC injection of E3 or MCF-7 cells. H&E: bone (B) and tumor (T) cells (n = 5 per group). Scale bars: 50 μm. IHC: ER or PR (green), CK5 (red) and DAPI (blue) by IF staining (n = 5 per group). Scale bars: 20 μm. (C) Percentage of CK5⁺ cells in E- or E+P-treated E3 and MCF-7 bone metastases. Mean ± standard error of the mean (SEM) values are shown (n = 5 per group). *P <0.05, Student’s t test. (D) Proliferation rates of CK5⁺ and CK5⁻ cells measured by phosphor-histone H3 (pHH3) in E- and E+P-treated E3 and MCF-7 bone metastases. Mean ± SEM values are shown (n = 5 per group). **P <0.005, Student’s t test.