Sequential Delivery of BMP2-Derived Peptide P24 by Thiolated Chitosan/Calcium Carbonate Composite Microspheres Scaffolds for Bone Regeneration

The combination of tissue-engineered bone scaffolds with osteogenic induction molecules is an important strategy for critical-sized bone defects repair. We synthesized a novel thiolated chitosan/calcium carbonate composite microsphere (TCS-P24/CA) scaffold as a carrier for bone morphogenetic protein 2- (BMP2-) derived peptide P24 and evaluated the release kinetics of P24. The effect of TCS-P24/CA scaffolds on the proliferation and differentiation of bone marrow mesenchymal stem cells (BMSCs) was evaluated by scanning electron microscope (SEM), CCK-8, ALP assay, alizarin red staining, and PCR. A 5 mm diameter calvarial defect was created, then new bone formation was evaluated by Micro-CT and histological examination at 4 and 8 weeks after operation. We found the sequential release of P24 could last for 29 days. Meanwhile, BMSCs revealed spindle-shaped surface morphology, indicating the TCS-P24/CA scaffolds could support cell adhesion and mRNA levels for ALP, Runx2, and COL1a1 were significantly upregulated on TCS-10%P24/CA scaffold compared with other groups in vitro (p<0.05). Similarly, the BMSCs exhibited a higher ALP activity as well as calcium deposition level on TCS-10%P24/CA scaffolds compared with other groups (p<0.05). Analysis of in vivo bone formation showed that the TCS-10%P24/CA scaffold induced more bone formation than TCS-5%P24/CA, TCS/CA, and control groups. This study demonstrates that the novel TCS-P24/CA scaffolds might contribute to the delivery of BMP2-derived Peptide P24 and is considered to be a potential candidate for repairing bone defects.