Regulation of Osteosclerosis by Inoculated Cd133⁺ PC3 Cells in Bone-marrow Microenvironmental Niches

Bone is the most common site of prostate cancer (PC) metastasis. Studies suggest that cancer stem cells (CSCs) are associated with stemness characteristics, providing some support for the concept that CSCs act as osteosclerotic precursors in bone microenvironmental niches. Here, we asked whether ectopic overexpression of CD133 maintains stability of CSCs in human PC cell lines and induces the changes of molecular features in the bone microenvironment. Ectopic overexpression of CD133 in PC3 or DU145 cells led to increased expression of ALDHA1, OCT4, and NANOG, enhanced colony-forming ability, and increased ALDH activity. In addition, micro-CT imaging, confocal microscopy, and H&E staining of mouse tissue confirmed that CD133 overexpression in PC3 and DU145 led to marked osteolytic bone tumor. However, expression of osteoblastic markers such as collagen type I, bone sialoprotein, and osteocalcin (OC) at the tumor margin of CD133-overexpressing PC3 tumors in mouse tibiae was higher than that of CD133-overexpressing DU145 tumors with osteosclerotic molecular features. In addition, expression of osteopontin (OPN) mRNA/protein by CD133-overexpressing PC3 cells was higher than that by DU145 cells. Especially, conditioned medium (CM) from PC3^CD133+ cells increased osterix (OSX) activity in bone marrow stromal cells (BMSCs), resulting in increased expression of OC mRNA/protein resulted in increased staining of mineralized matrix by Alizarin red. However, CM from OPN silenced PC3^CD133+ cells led to a reduction of OC mRNA and protein expression through OSX activity resulted in reduced amount of mineralized matrix. In conclusion, these findings suggest that CD133 plays a functional role in regulating CSC characteristics in PCs and modulates their abilities in which induce the osteosclerosis of BMSCs. In addition, OPN from CSCs acts as a niche component that promotes osteosclerosis by supporting osteoblastic differentiation of BMSCs.