In situ bone regeneration with sequential delivery of aptamer and BMP2 from an ECM-based scaffold fabricated by cryogenic free-form extrusion

In situ tissue engineering is a powerful strategy for the treatment of bone defects. It could overcome the limitations of traditional bone tissue engineering, which typically involves extensive cell expansion steps, low cell survival rates upon transplantation, and a risk of immuno-rejection. Here, a porous scaffold polycaprolactone (PCL)/decellularized small intestine submucosa (SIS) was fabricated via cryogenic free-form extrusion, followed by surface modification with aptamer and PlGF-2(123-144)*-fused BMP2 (pBMP2). The two bioactive molecules were delivered sequentially. The aptamer Apt19s, which exhibited binding affinity to bone marrow-derived mesenchymal stem cells (BMSCs), was quickly released, facilitating the mobilization and recruitment of host BMSCs. BMP2 fused with a PlGF-2(123-144) peptide, which showed “super-affinity” to the ECM matrix, was released in a slow and sustained manner, inducing BMSC osteogenic differentiation. In vitro results showed that the sequential release of PCL/SIS-pBMP2-Apt19s promoted cell migration, proliferation, alkaline phosphatase activity, and mRNA expression of osteogenesis-related genes. The in vivo results demonstrated that the sequential release system of PCL/SIS-pBMP2-Apt19s evidently increased bone formation in rat calvarial critical-sized defects compared to the sequential release system of PCL/SIS-BMP2-Apt19s. Thus, the novel delivery system shows potential as an ideal alternative for achieving cell-free scaffold-based bone regeneration in situ.