Effects of Enamel Matrix Derivative on Cell Spheroids Made of Stem Cells Obtained from the Gingiva on Osteogenic Differentiation

Abstract

Background and Objectives: A derivative of the enamel matrix was used to speed up periodontal regeneration, including the formation of new cementum, alveolar bone, and periodontal ligament. In this study, human gingiva-derived stem cell–derived cell spheroids were used to assess the effects of an enamel matrix derivative on cell viability, osteogenic differentiation, and mineralization. Materials and Methods: Human gingiva-derived stem cells were used to create spheroids, which were then coupled with unloaded control groups and an enamel matrix derivative at a final concentration of 2.7, 27, 270, and 2700 μg/mL. The morphological examination of the created stem cell spheroids took place on days 1, 3, 5, and 7. The Live/Dead Kit assay was used to determine the qualitative viability of cells on days 3 and 7. Using the Cell Counting Kit-8, the quantitative vitality of the cell spheroids was assessed on days 1, 3, and 5. On days 7 and 14, alkaline phosphatase activity assays and Alizarin Red S staining were carried out to examine the osteogenic differentiation of the cell spheroids. RUNX2 and COL1A1 expression levels on days 7 and 14 were determined using real-time polymerase chain reaction. Results: The added enamel matrix derivative at the tested concentrations did not significantly alter the morphology of the applied stem cells’ well-formed spheroids on day 1. On days 3 and 7, the majority of the spheroids’ cells fluoresced green while they were being cultivated. Alkaline phosphatase activity data revealed a substantial rise in the 2700 μg/mL group on day 7 when compared to the unloaded control (p < 0.05). On days 7 and 14, calcium deposits were distinctly seen in each group. In the 27 and 2700 μg/mL groups, the treatment with the enamel matrix derivative resulted in noticeably higher values for the Alizarin Red S staining (p < 0.05). qPCR results showed that adding an enamel matrix derivative to the culture of the 27 μg/mL group raised the level of RUNX2 mRNA expression. Conclusions: These results lead us to the conclusion that a derivative of the enamel matrix may be used to promote osteogenic differentiation in stem cell spheroids.

Regeneration of Rabbit Calvarial Defects with Combination of Stem Cells and Enamel Matrix Derivative: A Microcomputed Tomography and Histological Evaluation Comparing Two- and Three-Dimensional Cell Constructs

“…EMD has been recognized as a bone graft substitute and has demonstrated efficacy in promoting regeneration in intrabony and Class II mandibular furcation periodontal defects [ 23 ]. Further, it has been reported that the integration of EMD into 3D cultures enhances osteogenic differentiation in MSC spheroids, evidenced by significant increases in Alizarin Red S staining intensity and RUNX2 mRNA expression levels, suggesting a potential role in bone tissue engineering applications [ 13 ]. Nonetheless, earlier research has indicated that the outcomes of combining guided tissue regeneration techniques with EMD could exhibit considerable variability, leading to unpredictable regenerative results [ 24 ].…”

“…It contains amelogenin, which stimulates osteoblast activity and promotes bone formation [ 12 ]. Combining MSCs and EMDs in a tissue engineering approach may have the potential to improve bone regeneration outcomes [ 13 ]. While traditional two-dimensional cell cultures have been widely used in research, recent advances in tissue engineering have highlighted the advantages of three-dimensional cell structures [ 14 ].…”

Fibroblast Viability Through Mechanical and Chemical Root Surface Modifications in Periodontal Healing: An In Vitro Comparative Study

“…This emphasizes the potential advantages of exposure duration to specific chemical agents in facilitating fibroblast adhesion. EMD are derived from the porcine enamel matrix and promote new bone [ 22 ] and cementum [ 23 ] formation, making them a valuable component in periodontal regeneration.…”