Applying tissue engineering strategies to modulate tertiary dentinogenesis has emerged as a promising approach to improve success rate in local dentin regeneration. Scaffolds with nanofiber architecture are capable of increasing the odontoblastic phenotype expression of dental pulp stem cells (DPSCs) due to the similarity with natural extracellular matrix. Additionally, nanofibrous mats can act as drug delivery systems, creating actively designed surfaces. The aim of the present study is to develop a functionally graded nanofibrous scaffold for dentin regeneration. A blend of artificial and natural polymers will be prepared, to provide mechanical strength and adequate cell response. The surface layer to contact the dentin substrate will be enriched with antibiotics (ABL) in order to provide local antibacterial activity. Antibiotic concentration will be selected by agar diffusion and colony-forming units assays, against Actinomyces naeslundii and Lactobacillus casei species. A bioactive layer (BL) will be designed to interact with pulp tissue and modulate dentin regeneration by resident DPSCs. This layer will be enriched with calcium oxide at a concentration capable of enhancing odontoblastic marker overexpression (pNPP assay) on DPSCs, but with no cytotoxic effect (live/dead and MTS assays). ABL and BL will be characterized by scanning electron microscopy, contact angle, atomic force microscopy, fourier transform infrared spectroscopy and thermo-gravimetrically analysis. Thereafter, a multilayer membrane (FGM) will be obtained by sequential electrospinning technique and further characterized. This innovative biomaterial will be designed to feature an antibacterial layer, a core layer and a bioactive layer. The potential of FGM to inhibit bacterial adhesion to its surface and to induce an odontoblastic phenotype on dental pulp cells as a cell-free tissue engineering system will be tested. Data obtained will be submitted to specific statistical analysis.
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