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.