Effect of 3-dimensional Collagen Fibrous Scaffolds with Different Pore Sizes on Pulp Regeneration

In this study, we generated a 3-dimensional (3D) collagen fibrous scaffold for potential pulp regeneration and investigated the influence of various pore sizes of these scaffolds on proliferation, odontoblastic differentiation of human dental pulp cells (hDPCs), and subsequent tissue formation during pulp regeneration.Electrospinning followed by freeze-drying was used to fabricate 3D fibrous collagen scaffolds. hDPCs were cultured on these scaffolds. Cell growth was detected by a Cell Counting Kit-8 assay and observed via scanning electron microscopy. Odontogenic genes and protein expression were analyzed by real-time reverse transcription polymerase chain reaction and immunofluorescence staining. The formation of mineralized nodules was tested by von Kossa staining, scanning electron microscopy, and energy-dispersive X-ray microanalysis. Subcutaneous transplantation of the seeded scaffold/tooth fragments into nude mice was performed to observe tissue formation for pulp regeneration.Collagen 3D fibrous scaffolds with 3 distinct mean pore sizes (approximately 20 μm, 65 μm, and 145 μm) were fabricated, which showed good biocompatibility and bioactivity. Scaffolds with larger mean pore sizes of 65 and 145 μm improved hDPC ingrowth and proliferation, with the 65-μm scaffold group presenting the highest level of odontogenic gene expression (DSPP and DMP-1), protein expression (DMP-1), mineralized area ratio, and vascular pulplike tissue formation after 6 weeks of subcutaneous implantation.The pore size of collagen 3D fibrous scaffolds significantly affected cell adhesion, proliferation, odontoblastic differentiation, and tissue rehabilitation. Scaffolds with a mean pore size of 65 μm presented superior results and could be an alternative for pulp regeneration.