BACKGROUND: Bone replacement is frequently needed in periodontal, orthopedic, and maxillofacial diseases. To avoid complications with autografts and allografts, artificial grafts (scaffolds) are candidates for stimulating bone regeneration after colonization with osteoblasts. Moreover, osteoblast activity can be induced by biological or physical stimulation. In this research, extracorporeal shock waves were used to improve the ability of human osteoblasts to colonize scaffolds and to induce their osteogenic properties. METHODS: Osteoblasts, treated with shock waves, were seeded on glass-ceramic macroporous scaffolds. Cells in scaffolds were counted after detachment and examined for calcium nodule formation (Alizarin staining), for differentiation markers (real time polymerase chain reaction), and for scaffold colonization (scanning electron microscope). RESULTS: Shock waves initially increased both the number and the activity of osteoblasts in the scaffold, but subsequently increased only osteoblast activity. Moreover, shock waves favored scaffold colonization even in the deeper layers. CONCLUSIONS: The calcium deposits and differentiation markers studied have demonstrated that shock waves increase osteoblast migration and penetration into scaffolds. CLINICAL RELEVANCE: This study may provide an important starting point for the introduction of shock waves to boost bone formation through osteoblast stimulation in diseases characterized by bone defects.