Wnt5a Promotes Migration Of Human Osteosarcoma Cells By Triggering A Phosphatidylinositol-3 Kinase/Akt Signals

Objective : In this study, we hypothesized that the PI3K/Akt signaling pathway might mediate Wnt5a-induced osteosarcoma cell migration.Methods : Human osteosarcoma cell line MG-63 were cultured in Dulbecco-modified Eagle’s medium(DMEM) supplemented with 10% fetal bovine serum(FBS). As for wound healing assay, MG-63 cells were plated onto 96-well cell culture clusters and grown to confluence, and then serum-starved for 24 h. The monolayer cells were scratched manually with a plastic pipette tip, and after two washes with PBS, the wounded cellular monolayer was allowed to heal for 10 h in DMEM containing 100 ng/ml recombinant Wnt5a(r Wnt5a)(R&D Systems). Photographs of central wound edges per condition were taken at time 0 and at the indicated time points using digital camera. For gene knockdown,si RNA duplexes specific for Akt(Cell Signaling) were transfected into MG-63 cells by using Lipofectamine 2000 reagent(Invitrogen, Carlsbad, CA) in serum-free OPTI-MEM according to the manufacturer’s instructions. Knockdown efficiency was evaluated 48 h after transfection by measuring protein levels in cell lysates through using immunoblotting.Result: we found that Wnt5 a stimulated the migration of human osteosarcoma cells(MG-63), with the maximal effect at 100 ng/ml, via enhancing phosphorylation of phosphatidylinositol-3 kinase(PI3K)/Akt. PI3 K and Akt showed visible signs of basal phosphorylation and elevated phosphorylation at 15 min after stimulation with Wnt5 a.Pharmaceutical inhibition of PI3 K with LY294002 significantly blocked the Wnt5a-induced activation of Akt(p-Ser473) and decreased Wnt5a-induced cell migration.Additionally, Akt si RNA remarkably inhibited Wnt5a-induced cell migration.Conclusion : Our study show that Wnt5 a promotes osteosarcoma cell migration via PI3K/Akt signaling. These findings elucidate a molecular pathway linking Wnt5 a signalingto PI3K/Akt in cell motility. This result will contribute to further understanding of biological roles of Wnt5a/PI3K/Akt in cell migration of osteosarcoma and other cancers.