IADR Abstract Archives
Alternative in vitro methods to evaluate the potential of biomaterials as cell-homing therapy for dentin regeneration
Objectives: The aim of this study was to analyze the chemotactic potential of chitosan-based scaffolds as cell-free tissue engineering systems for dentin regeneration, by using realistic in vitro methods of dental pulp exposure.
Methods: Human dental pulp cells (hDPCs) were seeded on type I collagen matrix, and the 3D-culture was positioned on the pulpal surface of human dentin discs, adapted to open or sealed artificial pulp chambers (APCs). The scaffolds were placed into a central perforation prepared on dentin discs, in intimate contact with 3D-culture, simulating direct pulp capping procedure. For open APCs, the sets were incubated in 24-well plates containing 1 mL culture medium. For sealed APC, 1 mL culture medium was added to APC chamber, the system was hermetically sealed, and simulated body pressure was stablished with a 20-cm liquid column. The systems were incubated during 7 days and cells capable to migrate from 3D-culture to scaffold surface were stained with calcein AM. The number of migrating cells was obtained by means of Image J software (n = 4; t’student, p < 0.05). The expression of dentin sialoprotein (DSP) by migrating cells was analyzed by immunofluorescence.
Results: Both assays allowed observation of the improved chemotactic potential of SV-enriched scaffolds in comparison to plain CH scaffolds, with DPCs from 3D-culture being observed onto scaffold surface. However, a higher number migrating cells was observed when simulated body pressure was used (p < 0.05), with significant differences between CH and CHSV being observed. DSP-positive cells were detected only on sealed-APC assay, with CHSV featuring a higher number of positive cells compared to CH group.
Conclusions: The realistic method that simulates the in vivo conditions of pulp dentin complex allowed the observation of the chemotactic and bioactive potential of SV-enriched scaffolds for cell-homing therapy.
Methods: Human dental pulp cells (hDPCs) were seeded on type I collagen matrix, and the 3D-culture was positioned on the pulpal surface of human dentin discs, adapted to open or sealed artificial pulp chambers (APCs). The scaffolds were placed into a central perforation prepared on dentin discs, in intimate contact with 3D-culture, simulating direct pulp capping procedure. For open APCs, the sets were incubated in 24-well plates containing 1 mL culture medium. For sealed APC, 1 mL culture medium was added to APC chamber, the system was hermetically sealed, and simulated body pressure was stablished with a 20-cm liquid column. The systems were incubated during 7 days and cells capable to migrate from 3D-culture to scaffold surface were stained with calcein AM. The number of migrating cells was obtained by means of Image J software (n = 4; t’student, p < 0.05). The expression of dentin sialoprotein (DSP) by migrating cells was analyzed by immunofluorescence.
Results: Both assays allowed observation of the improved chemotactic potential of SV-enriched scaffolds in comparison to plain CH scaffolds, with DPCs from 3D-culture being observed onto scaffold surface. However, a higher number migrating cells was observed when simulated body pressure was used (p < 0.05), with significant differences between CH and CHSV being observed. DSP-positive cells were detected only on sealed-APC assay, with CHSV featuring a higher number of positive cells compared to CH group.
Conclusions: The realistic method that simulates the in vivo conditions of pulp dentin complex allowed the observation of the chemotactic and bioactive potential of SV-enriched scaffolds for cell-homing therapy.