Recellularisation of a Decellularised Dental Pulp Scaffold

Objectives: To assess the feasibility of a decellularised rat dental pulp as a scaffold to support human dental stem cells and promote differentiation.
Methods: Rat dental pulps were equally divided into control and decellularised groups (n=4/ group). The decellularisation protocol incorporated a low concentration sodium dodecyl sulphate and hypotonic buffers. Control and decellularised dental pulps were characterised histologically, immunohistochemistry and DNA content quantified. Biocompatibility was evaluated using a mouse fibroblast cell line (L929). Decellularised scaffolds were reseeded in-vitro with human dental pulp stem cells (5.5x10⁴cells.mL^-1) for 7 and 14-days. Cell viability, attachment and migration were monitored using Live/dead^® confocal imaging and histological analysis. Seeded stem cells were immunolabelled using makers for odontoblastic cells (alkaline phosphatase, dentine matrix protein-1, dentine sialophospoprotein) and cytoskeleton components (alpha-smooth muscle actin, nestin, vascular endothelium growth factor, vascular endothelium growth factor receptor-2, and vimentin). All histology and immunolabelled images were captured digitally and quantitative data statistically analysed (GraphPad Prism).
Results: Analysis of the decellularised scaffolds revealed an acellular matrix with preserved structural components, such as collagen, glycosaminoglycans, fibronectin and laminin. DNA content analysis indicated a significant difference between control and decellularised tissues, with DNA content loss >98% (student’s t-test, p<0.05). Decellularised scaffolds were biocompatible with normal cell growth and no difference in cellular activity compared to negative control (ANOVA, p>0.05). Recellularisation with dental pulp stem cells revealed a high majority of viable cells attached to the scaffold and distributed throughout. Antibody labelling demonstrated positive cellular expression against odontoblastic markers and cytoskeleton components following 14-days culture.
Conclusions: Decellularised pulp tissue could possibly be a suitable scaffold for regenerative endodontic treatment of non-vital immature permanent incisors. This study demonstrated the feasibility of developing a biocompatible decellularised dental pulp scaffold, which is able to support dental pulp stem cell growth and differentiation.