Chitosan-based endodontic irrigation solutions and TGF-β1 treatment: Creating the most favourable environment for the survival and proliferation of stem cells of the apical papilla in vitro

Abstract

Background: The dental pulp's environment is essential for the regulation of mesenchymal stem cells' homeostasis and thus, it is of great importance to evaluate the materials used in regenerative procedures.

Aim: To assess in vitro (i) the effect of chitosan nanoparticles, 0.2% chitosan irrigation solution, Dual Rinse®, 17% EDTA, 10% citric acid and 2.5% NaOCl on DSCS viability; (ii) the effect of different concentrations of TGF-β1 on DCSC proliferation; and (iii) whether treatment with TGF-β1 following exposure to the different irrigation solutions could compensate for their negative effects.

Methodology: (i) DSCS were treated with three dilutions (1:10, 1:100 and 1:1000) of the six irrigation solutions prepared in DMEM for 10 and 60 min to assess the effect on viability. (ii) The effect of different concentrations (0, 1, 5 and 10 ng/mL) of TGF-β1 on DCSC proliferation was assessed at 1, 3 and 7 days. (iii) The proliferative effect of TGF-β1 following 10-min exposure to 1:10 dilution of each irrigation solution was also tested. We used MTT assay to assess viability and proliferation. We performed statistical analysis using Prism software.

Results: (i) The different endodontic irrigation solutions tested showed a significant effect on cell viability (p ≤ .0001). Significant interactions between the endodontic irrigation solutions and their dilutions were also found for all parameters (p ≤ .0001). Chitosan nanoparticles and 0.2% chitosan irrigation solution were the least cytotoxic to DSCS whilst 2.5% NaOCl was the most cytotoxic followed by 17% EDTA. (ii) TGF-β1 at concentrations of 1 and 5 ng/mL resulted in significantly higher proliferation compared to the control group. (iii) Exposure to 17% EDTA or 2.5% NaOCl for 10 min was sufficient to make DSCS cells refractory to the proliferative effects of TGF-β1. DSCS groups treated with TGF-β1 following exposure to chitosan nanoparticles, 0.2% chitosan irrigation solution, Dual Rinse® and 10% CA demonstrated significantly higher proliferation compared to non-TGF-β1-treated groups (p ≤ .0001, p ≤ .0001, p ≤ .0001 and p = .01 respectively).

Conclusions: The current study offers data that can be implemented to improve the outcome of regenerative endodontic procedures by using less toxic irrigation solutions and adding TGF-β1 to the treatment protocol.

Keywords: EDTA; chitosan; citric acid; nanoparticles; regenerative endodontic procedures; stem cells.