Characterization of a Porous BMP-6-Loaded Composite Scaffold for Bone Regeneration in Rat Calvarial Bone Defects

Abstract

Biocompatible scaffolding materials play an important role in bone tissue engineering. This study sought to assess the biocompatibility and osteoinductivity of a chitosan (CS)/collagenI(ColI)/multi-walled carbon nanotube (MWCNT) composite scaffold blended with bone morphogenetic protein-6 (BMP-6) and seeded with bone marrow mesenchymal stem cells (BMMSCs). The composite was fabricated using varying quantities of MWCNT to determine the appropriate combination, and combined with BMP-6 by blending and vacuum freeze-drying. Scanning electron microscopy (SEM) was used to observe the morphology of the CS/ColI/MWCNT (CCM) scaffolds. The porosity and mechanical properties of the composite scaffolds were measured. Scaffolds were seeded with BMMSCs in vitro and analyzed using SEM, CCK-8 assay and alkaline phosphatase (ALP) activity. To assess the in vivo utility of the composite scaffolds, critical-sized calvarial bone defects were prepared in 15 Sprague–Dawley rats and implanted with pre-seeded scaffolds (the CCM scaffold+BMMSCs group and the BMP-6/CCM scaffold+BMMSCs group) or left untreated (control). Three-dimensional computerized tomography and hematoxylin–eosin (HE) staining were used to evaluate bone formation at 8 weeks postoperatively. We found that higher concentrations of MWCNT decreased the porosity of the scaffold but increased its mechanical strength. In vitro, we found that BMP-6/CCM scaffolds provided the best conditions for BMMSCs proliferation and osteogenic differentiation. Similarly, BMP-6/CCM scaffolds seeded with BMMSCs could more effectively induce new bone formation in critical-sized calvarial bone defects compared with the other three groups. The BMP-6/CCM scaffold possesses good biocompatibility and induces bone formation in vitro and in vivo.