Coordinated effect of simvastatin-induced osteogenesis and angiogenesis mediated by Wnt/β-catenin signaling pathway on vascularized bone tissue formation

As a typical lipid-lowering drug, simvastatin has recently demonstrated potential in modulating bone formation and neovascularization. Nevertheless, its specific role in the coupling of osteogenesis and angiogenesis during functional bone regeneration remains unclear. This study aimed to investigate the effects of simvastatin on osteogenic differentiation and angiogenic regulation of stem cells from human exfoliated deciduous teeth (SHED) and to clarify the underlying molecular mechanism. SHED were isolated from caries-free human deciduous teeth. Concentration-dependent effects of simvastatin on cell proliferation, migration, and apoptosis were evaluated using CCK-8, transwell, wound healing, and Annexin V-FITC/PI double staining assays. Osteogenic differentiation was assessed via alkaline phosphatase (ALP) activity, ALP staining, Alizarin Red S staining, qRT-PCR, and Western blotting. Angiogenic regulatory activity was examined using a SHED-human umbilical vein endothelial cell (HUVEC) co-culture system. Markers associated with Wnt/β-catenin signaling were detected, and dickkopf-related protein-1 (DKK-1) was employed to determine its role in the observed effects. Porous hydroxyapatite (HAP) scaffolds loaded with simvastatin and SHED were prepared and subcutaneously transplanted into nude mice, and the effects of simvastatin on osteogenesis and angiogenesis in vivo were evaluated by histological staining. Low-concentration simvastatin (≤ 1 µM) enhanced SHED migration without adversely affecting cell viability. Within this range, 0.1 µM simvastatin significantly upregulated osteogenic gene expression in SHED, accompanied by increased ALP activity and matrix mineralization. Simvastatin also elevated the expression of angiogenic regulatory factors in SHED, promoting more extensive reticular structure formation by HUVEC through paracrine interactions between SHED and HUVEC. Furthermore, the Wnt/β-catenin signaling pathway was activated, and the osteogenic and angiogenic effects of simvastatin were reversed by DKK-1 inhibition. Subcutaneous transplantation in nude mice confirmed that simvastatin facilitated vascularized bone formation in vivo, with enhanced expression of osteogenesis- and angiogenesis-related factors. Low-concentration simvastatin promoted osteogenesis and angiogenesis, with the activation of Wnt/β-catenin signaling pathway in SHED implicated as a contributing mechanism, highlighting the potential role of simvastatin in supporting vascularized bone-like tissue formation during functional bone regeneration.