Biomimetic pulp scaffolds prepared from extracellular matrix derived from stem cells from human exfoliated deciduous teeth promote pulp-dentine complex regeneration

Abstract

Aim: To evaluate the role of biomimetic pulp scaffolds derived from the extracellular matrix derived of stem cells from human exfoliated deciduous teeth (SHED-ECM-PS) in promoting pulp-dentine complex regeneration.

Methodology: SHED-ECM-PS was prepared through cell aggregation and decellularization techniques. Histological and immunofluorescence analyses, scanning electron microscopy, and DNA quantification assays were used to characterize the SHED-ECM-PS. Additionally, a tooth slice implantation model was established to evaluate the effects of SHED-ECM-PS on regeneration of the pulp-dentine complex in vivo. Extraction medium for SHED-ECM-PS was prepared, and its effect on bone marrow mesenchymal stem cells (BMMSCs) was assessed in vitro. Cell counting kit-8 and Ki-67 staining assays were performed to determine cell proliferation. The rate of apoptosis was evaluated by flow cytometry. Wound healing and transwell assays were conducted to evaluate cell migration. Alizarin red S staining was performed to examine mineralized nodule formation. Western blotting was used to detect the expression of osteogenic and odontogenic markers. The results were analysed using an independent two-tailed Student's t-test. p < .05 was considered statistically significant.

Results: SHED-ECM-PS was successfully constructed, exhibiting a striped dental pulp-like shape devoid of nuclear structures or DNA components, and rich in fibronectin, collagen I, DMP1 and DSPP. Notably, SHED-ECM-PS showed no impact on the proliferation or apoptosis of BMMSCs. Histological analysis revealed that dental pulp fibroblasts formed an interwoven mesh in the root canal, and angiogenesis was observed in the SHED-ECM-PS group. Moreover, a continuous, newly formed tubular dentine layer with polarized odontoblast-like cells was observed along the inner wall of the root canal. SHED-ECM-PS promoted the migration, polar alignment and mineralized nodule formation of BMMSCs and specifically elevated the expression levels of odontogenic markers, but not osteogenic markers, compared with the control group in vitro.

Conclusion: SHED-ECM-PS exhibited no cytotoxicity and promoted pulp-dentine complex regeneration in vivo as well as cell migration and odontogenic differentiation of BMMSCs in vitro. These findings provide evidence that SHED-ECM-PS, as a novel biological scaffold, has the potential to improve the outcomes of REPs.

Keywords: biomimetic scaffold; extracellular matrix; odontogenic differentiation; pulp–dentine complex; regenerative endodontic procedures; stem cell from human exfoliated deciduous teeth.