Combined Effects of Vitamin D3 and Extracorporeal Shockwave Therapy on Osteoblast Differentiation via F-actin mediated Pdia3 Membrane Localization.

AIM: 1α,25(OH)₂D₃ (vitamin D3, VitD3) and extracorporeal shockwave therapy (ESWT) promote osteoblast differentiation and mineralization. Protein disulfide isomerase-associated 3 (Pdia3) mediates rapid membrane signaling of VitD3, but its interaction with ESWT remains unclear.

MATERIALS AND

METHODS

MC3T3-E1 cells were treated with VitD3, ESWT, or both. Alkaline phosphatase activity and Alizarin red staining assessed osteoblast differentiation and mineralization. Pdia3 expression and signaling were evaluated by Western blotting and ELISA. Pdia3 distribution was analyzed by immunofluorescence and flow cytometry.

RESULTS

The actin polymerization inhibitor latrunculin B tested the role of F-actin. Combined VitD3 and ESWT significantly increased ALP activity, mineral deposition, Pdia3 expression, and plasma membrane localization versus single treatments. These effects were mediated by Pdia3-dependent protein kinase C (PKC) activation. Disruption of F-actin reduced Pdia3 membrane localization and abolished the combined osteogenic effects. ESWT promoted F-actin repolymerization, facilitating Pdia3 trafficking to the membrane.

CONCLUSION

VitD3 and ESWT exert enhanced combined effects on osteoblast differentiation and mineralization by promoting F-actin mediated Pdia3 membrane localization and activating the PKC pathway. The integrity of the actin cytoskeleton is essential for these effects.