Hybrid bilayered chitosan-xanthan/PCL scaffolds as artificial periosteum substitutes for bone tissue regeneration

The periosteum, a bilayered membrane that covers bone surfaces, acts as a source of bone-forming cells and plays a pivotal role in bone homeostasis and defect healing. Tissue-engineered periosteum substitutes may be applied in different surgical modalities to treat large or complex bone fractures, as adjunctives in osteosynthesis, or even as membranes for guided bone regeneration. In this work, phosphorylated chitosan or the unmodified polysaccharide was used as osteoinductive materials to produce chitosan-xanthan complexes, which were combined with a polycaprolactone (PCL) porous membrane. This combination originated a bilayered scaffold constituted of a highly hydrated porous chitosan-xanthan layer designed to be in direct contact with bone tissue, and a slowly biodegrading PCL outer layer capable of providing long-term support for cell growth. The bilayered scaffolds presented high porosity (95%) and a gradient in pore size, with average of 261 μm for PCL and up to 900 μm for polysaccharide layers. Despite the scaffolds are thicker than the native periosteum, with thickness higher than 4.3 mm, their compression and degradation after implantation make them suitable for the application. Mechanical testing of hydrated scaffold samples showed that the major contribution to the resistance of the bilayered constructs is derived from the PCL layer, which presents Young modulus of approximately 120 kPa, about two times higher than the modulus presented by the bilayered material. Toxicity test against dental pulp stem cells revealed that the developed materials are biocompatible, as cell activity remained above 70% in comparison with negative control. Thus, these scaffolds have great potential to be applied as periosteum substitutes, as they could be simply obtained and applied, recapitulating the tissue's natural characteristics and matching the requirements for different applications.