Dexamethasone-Functionalized PLLA Membranes: Effects of Layer-by-Layer Coating and Electrospinning on Osteogenesis

The addition of dexamethasone in membranes for guided bone regeneration is promising due to its dual effect: (1) anti-inflammatory action and (2) induction of osteogenesis in host stem cells. Electrospun fiber coating with dexamethasone using the layer-by-layer (LBL) technique offers an interesting alternative for the gradual release of the drug, aiming for enhanced osteodifferentiation activity. This study aimed to develop synthetic poly-L-lactide (PLLA) membranes with dexamethasone incorporated into the fibers or coated on their surface, and to evaluate the drug release rate, as well as the material's ability to promote proliferation, osteoconduction, and osteodifferentiation of human periodontal ligament stem cells (hPDLSCs). PLLA membranes were produced by electrospinning. Dexamethasone was incorporated using three techniques: (A) electrospinning of a co-solution of PLLA with 2.5 w/w% dexamethasone; (B) deposition of four layers on the PLLA membrane using alternating solutions of chitosan and heparin/dexamethasone; (C) deposition of 10 layers on the PLLA membrane using the same solutions. hPDLSC proliferation was measured via CCK-8 at 1, 7, 14, and 21 days. Cellular differentiation was assessed by alkaline phosphatase activity (7 days) and alizarin red staining (21 days) in clonogenic and osteogenic media (ODM). Data were analyzed using one or two-way ANOVA and Tukey test. Electrospun membranes with dexamethasone and those with 4 layers showed immediate drug release within 24 h, whereas 10 layers exhibited gradual release over 14 days. Cumulative drug release was higher for electrospun membranes at 1 and 7 days, similar to 10 layers at 14 and 21 days. The 4 LBL membrane promoted lower hPDLSC proliferation compared to the 10 LBL and electrospun membranes at 21 days but showed increased extracellular matrix mineralization in osteogenic media. No significant differences in alkaline phosphatase expression were observed between materials. Therefore, the addition of dexamethasone in 10 layers, combined with heparin, enables gradual drug release. However, lower drug release in the first 24 h by four LBL membranes improved the material's osteogenesis properties. None of the materials improved the osteodifferentiation in the clonogenic medium.