LAUSR

AIM In order to obtain a 3-dimentional scaffold with predictable clinical results for pulp regeneration, this study aims to fabricate and characterize a porous decellularized human amniotic membrane (HAM) extracellular matrix (ECM) scaffold, and evaluate its potential to promote pulp regeneration in vitro and in vivo. METHODOLOGY The HAM was decellularized, and its histology and DNA content were analyzed to confirm decellularization. The scaffolds were synthesized with 15, 22.5, and 30 mg/mL concentrations. The porosity, pore size, phosphate buffered saline (PBS) absorption, and degradation rate of the scaffolds were assessed. In vitro experiments were performed on human dental pulp stem cells (hDPSCs) to assess their viability, proliferation, adhesion, and migration on the scaffolds. The optimal group was selected for in vivo immunogenicity assessment and was also used as the cell-free or cell-loaded scaffold in root segment models to evaluate pulp regeneration. All nonparametric data were analyzed with the Kruskal-Wallis test followed by Dunn's post hoc test, while quantitative data were analyzed with one-way ANOVA. RESULTS Decellularization of HAM was confirmed (P<0.05). The porosity of all scaffolds was more than 95%, and the pore size decreased with an increase in ECM concentration (P<0.01). PBS absorption was not significantly different among the groups, while 30 mg/mL ECM scaffold had the highest degradation rate (P<0.01). The hDPSCs adhered to the scaffold, while their proliferation rate increased over time in all groups (P<0.001). Cell migration was higher in 30 mg/mL ECM scaffold (P<0.05). In vivo investigation with 30 mg/mL ECM scaffold revealed mild to moderate inflammatory response. In root segments, both cell-free and cell-loaded 30 mg/mL scaffolds were replaced with newly formed, pulp-like tissue with no significant difference between groups. Immunohistochemical (IHC) assessments revealed high revascularization and collagen content with no significant difference among the groups. CONCLUSION The 30 mg/mL HAM ECM scaffold had optimal physical properties and better supported hDPSC migration. The HAM ECM scaffold did not interfere with formation of pulp-like tissue and revascularization within the root canal when employed as both cell-free and cell-loaded scaffold. These results highlight the potential of HAM ECM membrane for further investigations in regenerative endodontics.