Engineered extracellular vesicles with sequential cell recruitment and osteogenic functions to effectively promote senescent bone repair

Senescent mandibular bone repair pose a formidable challenge without an entirely satisfactory strategy. Endogenous cell recruitment and osteogenic differentiation are two sequential stages for bone regeneration, and the disorder in these two processes is a significant obstacle for senescent bone repair. To tackle these problems, engineered extracellular vesicles (EV) with sequential stem cell recruitment and osteogenic functions were developed. This study demonstrated that Apt19s engineered extracellular vesicles (Apt19s-EV) recognize and recruit O-BMSCs specifically and effectively. MiR-376b-5p screen by RNA sequencing and transfection, was significantly decreased in senescent bone marrow mesenchymal stem cells (O-BMSCs), which was selected to construct miR-376b-5p engineered extracellular vesicles (miR-376b-5p-EV). miR-376b-5p-EV could promote osteogenesis and alleviate senescence of O-BMSCs by targeting Camsap1. To combine the advantages of Apt19s and miR-376b-5p, dual engineered extracellular vesicles (Apt-376b-EV) comprising Apt19s and miR-376b-5p modification simultaneously was constructed. To further validate its function, Gelatin methacryloyl (GelMA) hydrogel was used as a carrier to construct Apt-376b-EV@GelMA delivery system. The in vitro results have demonstrated that Apt-376b-EV@GelMA could recruit O-BMSCs, alleviate senescence and promote osteogenic differentiation sequentially. Notably, the in vivo study also observed that Apt-376b-EV@GelMA sequentially recruit endogenous stem cells homing and enhance new bone formation in senescent bone fracture and critical-sized defect models. In summary, the dual engineered extracellular vesicles, Apt-376b-EV, offers an appealing solution for recruiting endogenous stem cells and promoting bone repair sequentially in senescent microenvironment, which may broader clinical applications of engineered EV and provide valuable strategies for senescent bone-related diseases in the future clinical work.