Scaffold architecture exerts a considerable influence on the osteogenic effect through stress transmission, as the deformation of scaffolds alters the mechanical microenvironment of cells adhering to scaffold surface. Despite extensive research on bone regeneration influenced by scaffold architecture, present studies have not addressed the biological mechanism underlying scaffold architecture-induced stress stimulation (SASS) on cells yet, posing a great challenge in revealing the biomechanical cues between scaffold architecture and osteogenic progression. Therefore, graphite, fullerene, and diamond scaffolds with gradient stress stimulation to cells after deformation were prepared. The cellular biomechanical mechanisms of SASS through single-cell RNA sequencing indicated that architectures providing SASS can induce the enrichment of focal adhesion and osteogenic differentiation pathways of bone mesenchymal stem cells and balance bone resorption of osteoclasts and bone formation of osteoblasts. Besides, SASS enhances bone regeneration for repairing critical-sized defects in vivo. These results provide insights for artificial bone scaffold design and clarify the biomechanical cues between SASS and osteogenic progression.