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 beta-tricalcium phosphate-doped-PLGA/PCL (PP beta). 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 (PP beta-PP-PPC) and PP beta layer facing MC cells (PPC-PP-PP beta) to evaluate surfacematerial 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 PP beta-PP-PPC compared to PP beta, PPC-PP-PP beta, and PP. PPC and PP beta-PP-PPC exhibited hydrophilic property. The three-layer membranes (PPC-PP-PP beta and PP beta-PP-PPC) demonstrated significantly higher Youngs 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-PP beta compared to the controls. Promoted osteoconductive effects of PPC-PP-PP beta were revealed by alkaline phosphatase assays and Western blot compared with the controls at 7 and 14 days. PPC, PPC-PP-PP beta and PP beta-PP-PPC exhibited a significantly wider antibacterial zone against the tested bacteria compared to PP and PP beta (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.