Accumulating studies have shown that the receptor activator of nuclear factor-κB ligand (RANKL)/RANK pathway plays an important role in tumor metastasis. However, the involvement of the RANKL/RANK signal transduction pathway in breast cancer metastasis remains unclear. The present study, therefore, investigated the role of downstream molecules of RANKL/RANK signaling in breast cancer cells using Transwell chemotaxis assays. RANKL was shown to direct the migration of MDA-MB-231 breast cancer cells. Osteoprotegerin (OPG; soluble decoy receptor of RANKL) inhibited RANKL-induced migration. RANKL activated Src kinase in MDA-MB-231 cells, as shown by Western blotting, and pretreatment with a Src inhibitor abrogated RANKL-induced cell migration, in a similar manner to OPG. Short-hairpin RNA against RANK, delivered via a lentiviral vector, significantly abolished the expression of phosphorylated Src. Stimulation by RANKL induced the phosphorylation of mitogen-activated protein kinases (MAPKs) (ERK, p38, JNK), and specific inhibitors of MAPKs blocked RANKL-induced cell migration. Furthermore, the expression of phosphorylated MAPKs could be blocked by a Src inhibitor and by small interfering RNA against Src. These findings suggest that Src and MAPK pathways may be involved in RANKL-induced MDA-MB-231 breast cancer cell migration.

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