In-Depth Analysis of AG73-Dental Pulp Cell Interactions Underlying Adhesion and Mineralization: Insights from Alanine Scanning.

Abstract

Background: This study aimed to explore the dental applications of AG73, a laminin-derived adhesive peptide, by examining its effects on human dental pulp cells (hDPCs), particularly in cell adhesion and mineralization. It also sought to identify the key functional residue of AG73 and the signaling pathways involved in its pro-mineralization activity.

Methods: We compared AG73's pro-mineralization activity with peptides derived from LAMA5 using alizarin red staining. Immunofluorescence staining assessed its cell adhesion properties. Alanine-scanning mutagenesis was performed to identify key residues for AG73's adhesion activity. Finally, we used p38 and JNK MAPK inhibitors to determine which signaling pathways were involved in AG73-induced mineralization.

Results: AG73 exhibited superior pro-mineralization activity compared to other peptides derived from LAMA5 in hDPCs. Immunofluorescence staining confirmed AG73's strong cell adhesive ability within 20 minutes. Alanine-scanning identified isoleucine at the tenth position as crucial for adhesion. AG73 was the most effective peptide in inducing mineralization, with no mutated versions outperforming it. p38 signaling was found to play a positive role in AG73-induced mineralization, while JNK inhibition enhanced mineralization, suggesting JNK inhibitors as potential inducers.

Conclusion: AG73 has strong pro-mineralization activity and significant cell adhesive properties in hDPCs. The isoleucine at the tenth amino acid position was identified as a critical residue for AG73's adhesive ability. Additionally, p38 was found to be involved in AG73-regulated mineralization, while JNK inhibition promoted the process. This is the first study to explore AG73's effects in hDPCs and provides a foundation for its potential use as a dental-pulp regenerative material.