Apple-Derived Vesicles Orchestrate Bone Regeneration: In Vitro Proof of Concept

The immune microenvironment critically influences bone healing, particularly in the oral cavity where inflammation and microbial biofilms can compromise regeneration. Plant-derived extracellular vesicles (PDEVs) offer a biocompatible means to modulate immune responses, and apple-derived extracellular vesicles (ADEVs) have shown antioxidant and anti-inflammatory activity, although their osteoregenerative potential remains unclear. Here, we investigate the indirect effects of ADEVs on bone regeneration by assessing how their immunomodulatory action on macrophages influences the osteogenic commitment of human dental pulp stem cells (DPSCs). ADEVs were isolated, characterized, and applied to THP-1-derived macrophages to evaluate polarization via morphology and immunofluorescence for M1 (iNOS) and M2 (ARG1) markers. Then, the extracellular vesicles (EVs) from untreated and ADEV-treated macrophages were isolated and applied to DPSCs. All EVs were efficiently internalized by both macrophages and DPSCs. Treated macrophages shifted toward an M2-like phenotype, and macrophage-derived EVs (MDEVs) promoted stem cell morphological features consistent with osteogenic activation. These findings suggest that ADEVs promote osteoregeneration indirectly by influencing macrophage polarization and modifying the osteoactive cargo of MDEVs, thereby supporting their potential in cell-free, immunomodulatory approaches for oral bone regeneration.