Ultrasound-induced activation of Wnt signaling in human MG-63 osteoblastic cells

Abstract

The benefit from an ultrasound (US) exposure for fracture healing has been clearly shown. However, the molecular mechanisms behind this effect are not fully known. Recently, the canonical Wnt signaling pathway has been recognized as one of the essential regulators of osteoblastogenesis and bone mass, and thereby considered crucial for bone health. Mechanical loading and fluid shear stress have been reported to activate the canonical Wnt signaling pathway in bone cells, but previous reports on the effects of therapeutic US on Wnt signaling in general or in bone, in particular, have not been published yet. Therefore, activation of Wnt signaling pathway was assayed in human osteoblastic cells, and indeed, this pathway was found to be activated in MG-63 cells through the phosphoinositol 3-kinase/Akt (PI3K/Akt) and mTOR cascades following a single 10 min US exposure (2 W, 1.035 MHz). In addition to the reporter assay results, the Wnt pathway activation was also observed as nuclear localization of beta-catenin. Wnt activation showed also temperature dependence at elevated temperatures, and the expression of canonical Wnt ligands was induced under the thermal exposures. However, existence of a specific, non-thermal US component was evident as well, perhaps evidence of a potential dual action of therapeutic US on bone. Neither US nor heat exposures affected cell viability in our experiments. In summary, this is the first study to report that Wnt signaling cascade, important for osteoblast function and bone health, is one of the pathways activated by therapeutic US as well as by hyperthermia in human osteoblastic cells. Our results provide evidence for the potential molecular mechanisms behind the beneficial effects of US on fracture healing. Combinations of US, heat, and possible pharmacological treatment could provide useful flexibility for clinical cases in treating various bone disorders.