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Proliferation and differentiation of osteoclasts are regulated by a cytokine system that includes RANKL, which binds 2 receptors: RANK, which activates osteoclast differentiation, and osteoprotegerin (OPG), a decoy receptor that limits RANKL action. We investigated the role of the OPG/RANKL/RANK network in the pathogenesis of skeletal metastasis in neuroblastoma. Four different neuroblastoma cell lines (NB100, CHP212, SH-SY5Y, SJ-NK-P) showed a large amount of OPG and RANKL transcripts. Soluble RANKL was detectable in all cell lines, but poor release of OPG was observed. SH-SY5Y showed the lowest OPG-to-RANKL ratio and promoted osteoclastic differentiation of FLG29.1 and peripheral mononuclear cells, inducing expression of the osteoclast markers RANK, c-src, c-fos, cathepsin-K and TRAP. SJ-N-KP, which released both OPG and RANKL, did not show the same capability. OPG, neutralizing anti-RANKL antibody and antisense oligonucleotides were evaluated for their ability to inhibit RANKL activity. The neutralizing antibody hampered osteoclastic differentiation by blocking both the juxtacrine and the paracrine activity of RANKL. Our findings confirm that neuroblastoma cells induce osteoclastogenesis via RANKL and suggest that the RANKL expression associated with lack of the decoy receptor OPG could be a peculiarity of some tumors that makes them able to induce metastatic osteolysis. Moreover, our results suggest that RANKL could be a relevant target in the adjuvant therapy of bone metastatic neuroblastoma as proper neutralization revokes completely osteoclastic differentiation.