(A) human PC-3 and RWPE-1 cells were intracardiac (i.c.) or intra-tail vein (i.v.) injected in mice bearing heterotopic humanized bone/BM ossicles. (B) Western blot analysis of PC3 and RWPE-1 cell lysates. β-actin was used as control (a). Fluorescent Activated Cell Sorting (FACS) analysis of PC3 cells stained with anti-human CD146 (b). (C) GFP expression of PC-3 cells performed before i.c. injection of SCID/beige mice. (D) I.c. injected PC3 cells not survive and growth in murine BM. Representative FACS analysis for human CD146 and GFP in murine BM three days and five weeks after i.c. injection of 1 × 106 PC3 cells show that human PC3 cells reach murine BM, but do not survive or generate histologically detectable metastatic deposits. PC-3 cells are refractory to colonize murine bone/BM. Scale bars represent 120, 300, 500 μm as indicated. (E) X-ray analysis did not detect any bone metastasis in mouse skeleton following prostate cancer PC3 or RPWE-1 injection. H&E, Hematoxylin/Eosin; hem, hematopoiesis.