Biomimetic Domain-Active Electrospun Scaffolds Facilitating Bone Regeneration Synergistically with Antibacterial Efficacy for Bone Defects.

To improve bone regeneration in oral microenvironment, we generated a novel biodegradable, antibacterial, and osteoconductive electrospun PLGA/PCL membrane as an ideal osteogenic scaffold. The novel three-layer membranes were structured with serial layers of electrospun chlorhexidine-doped-PLGA/PCL (PPC), PLGA/PCL (PP), and β-tricalcium phosphate-doped-PLGA/PCL (PPβ). To characterize osteoconductive properties of these membranes, MC3T3-E1 (MC) cultures were seeded onto the membranes for 14 days for evaluation of cell proliferation, morphology and gene/protein expression. In addition, MC cells were cultured onto different surfaces of the three-layer membranes, PPC layer facing MC cells (PPβ-PP-PPC) and PPβ layer facing MC cells (PPC-PP-PPβ) to evaluate surface-material effects. Membrane properties and structures were evaluated. Antibacterial properties against Streptococcus mutans and Staphylococcus aureus were determined. Scanning electron microscope demonstrated smaller interfiber spaces of PPC and PPβ-PP-PPC compared to PPβ, PPC-PP-PPβ, and PP. PPC and PPβ-PP-PPC exhibited hydrophilic property. The three-layer membranes (PPC-PP-PPβ and PPβ-PP-PPC) demonstrated significantly higher Young's modulus (94.99 ± 4.03 MPa and 92.88 ± 4.03 MPa) compared to PP (48.76 ± 18.15 MPa) or PPC (7.92 ± 3.97 MPa) (p < 0.05). No significant difference of cell proliferation was found among any groups at any time point (p > 0.05). Higher expression of integrins were detected at 12 h of cultures on PPC-PP-PPβ compared to the controls. Promoted osteoconductive effects of PPC-PP-PPβ were revealed by alkaline phosphatase assays and Western blot compared with the controls at 7 and 14 days. PPC, PPC-PP-PPβ and PPβ-PP-PPC exhibited a significantly wider antibacterial zone against the tested bacteria compared to PP and PPβ (p < 0.05). These results suggested that the three-layer electrospun membranes demonstrated superior properties: higher strength, better cell adhesion, and promoted osteoconductive properties compared to single-layer membrane: however, antibacterial properties were exhibited in three-layer electrospun membranes and chlorhexidine-doped single-layer membrane. We concluded that the novel three-layer membranes could be used as a biocompatible scaffold for intraoral bone regeneration due to its enhanced osteoconductive activity and antibacterial effect.