Figure 4.

Increased expression of RBP4 and PLAC8 in prostate cancer cells remaining in bone of nude mice following osteolytic tumor growth inhibition by OPG and the effect of RBP4 and PLAC8 knockdown on the growth of prostate cancer cells. (A) Bone tumors were minced with a sharp-edged knife and cultured for 1 week. PC3-GFP cells were then separated from growing cells by a cell sorter. Total RNA from PC3-OPG cells was reverse-transcribed and the obtained cDNA was subjected to PCR for RBP4, PLAC8 and β-actin. (B) Formalin-fixed paraffin-embedded sections of bone lesions were subjected to immunohistochemical staining for GFP, RBP4 and PLAC8. Scale bar, 100 μm. (C) PC-3 cells were transfected with negative control siRNA, RBP4 siRNA and PLAC8 siRNA. Specific knockdown of RBP4 and PLAC8 was confirmed. (D) Transfectants were plated onto 96-well plates (5×10³ cells/well) and pre-incubated overnight. Subsequently, cells were further incubated for 24 h and the growth rates were evaluated using the MTT method (n=6).