The role of mTOR signal on Tertiary dentin formation.

Objectives: Rapamycin is mTOR signaling inhibitor for organ transplantation, cancer, and fibro-dysplasia ossificans progressiva. Rapamycin has been reported to inhibit dental pulp stem cells (DPSCs) differentiation in vitro. However, there is no report the effect of mTOR inhibitor on tertiary dentin formation by objective three-dimensional evaluation. Additionally, the mechanism for the inhibition of DPSCs differentiation is also unclear. Therefore, the present study investigated the influence of mTOR inhibitor on tertiary dentin formation after direct pulp capping (DPC) in rat using serial micro-CT evaluation in vivo and whole gene expression of human DPSCs using RNA sequence treated with rapamycin in vitro.
Methods: The experiment was approved by Ethical Guidelines Committee for Animal Care of Osaka University Graduate School of Dentistry (No. R-01-003-0). Rapamycin (4mg/kg, LC Laboratories) was intraperitoneally injected into Wistar rats. Then, cavity was mechanically prepared in maxillary first molar with pulp exposure. ProRoot MTA® (Dentsply Sirona) was used as a pulp-capping material and then the cavity was restored with glass-ionomer cement. Tertiary dentin volume was evaluated by micro-CT (mR_CT2, RIGAKU, Japan) and histologically 2 or 4 weeks postsurgery. For RNA sequence, hDPSCs were treated with rapamycin for 7 days. Then, total RNA was prepared using RNeasy Mini RNA isolation kit (Qiagen). RNA sequence was performed using Hiseq 2500 (illumina). Meta data were used for heatmap visualization with integrated Differential Expression and Pathway analysis (PMID:30567491) and ClustVis (PMID:25969447). Statistical significance analysis was evaluated by Student’s t-test (α=0.05).
Results: The volume of reparative dentin in rapamycin treated group showed significant decrease in both 2 and 4-week samples compared with untreated group (p<0.05). For the RNA Sequence analysis, rapamycin affected several genes expression and signal pathways related mineralized tissue formation compared with untreated group.
Conclusions: mTOR signaling plays an essential role in the healing process of pulp tissue especially dentinogenic healing.