Pulp ECM-derived macroporous scaffolds for stimulation of dental-pulp regeneration process

Abstract Objective Recent studies suggest xenogeneic extracellular matrices as potential regenerative tools in dental pulp regeneration. This study aimed to fabricate and characterize a novel three-dimensional macroporous pulp-derived scaffold that enables the attachment, penetration, proliferation and differentiation of mesenchymal stem cells. Method Bovine pulp was decellularized and characterized with histological and DNA content methods. This scaffold was prepared using finely milled lyophilized decellularized pulp extracellular matrix (ECM) digested with pepsin. Three different concentrations of ECM (1.50, 2.25 and 3.00 mg/ml) were freeze-dried and were tested with/without chemical crosslinking. The specimens were subjected to physicochemical characterization, cell viability and quantitative real time polymerase chain reaction assessments with human bone marrow mesenchymal stem cells (hBMMSCs). All scaffolds were subcutaneously implanted in rats for two weeks and histological and immunostaining analyses were performed. Results Histological and DNA analysis confirmed complete decellularization. All samples demonstrated more than 97% porosity and 1.50 mg/ml scaffold demonstrated highest water absorption. The highest cell viability and proliferation of hBMMSCs was observed on the 3.00 mg/ml crosslinked scaffolds. The gene expression analysis showed a significant increase of dmp-1 and collagen-I on 3.00 mg/ml crosslinked scaffolds compared to the other scaffolds. Histological examination of subcutaneous implanted scaffolds revealed low immunological response, and enhanced angiogenesis in cross-linked samples compared to non-crosslinked samples. Significance The three-dimensional macroporous pulp-derived injectable scaffold developed and characterized in this study displayed potential for regenerative therapy. While the scaffold biodegradability was decreased by crosslinking, the biocompatibility of post-crosslinked scaffold was significantly improved.