Biological effects of new chemical-mechanical caries removal products on human dental pulp stem cells.

Abstract

The aim of this study was to compare the biological effects of four chemical caries removal materials and to assess their cytotoxicity using human dental pulp stem cells (hDPSCs). The products evaluated are: 1 - Papain based product (BRIX 3000®); 2 - Papain/chloramine based products (NATURAL-CARE and Papacárie Duo®); and 3 - Chloramine based product (Cariesolut); The following in vitro experiments were carried out: IC50 measurement, cell metabolic activity (MTT) assay, cell migration, immunofluorescence experiment, cell apoptosis analysis, and reactive oxygen species (ROS) production analysis. Statistical analyses were performed using one-way ANOVA followed by Tukey's post hoc test (p<0.05). The IC50 values were: Brix 3000: 0.596%; Papacárie Duo: 0.052%; NATURAL CARE: 1.034%; and Cariesolut: 0.020%. The MTT assays showed non-adequate cell viability of all CMCR tested at 2% at 24, 48, and 72 h (p<0.001). The same behaviour was observed at 0.1% in the Papacárie Duo and Cariesolut groups. In contrast, 0.1% of Brix 3000 at all times and NATURAL CARE at 24h treated cells showed cell viability rates similar to the control group. At 0.01% only Brix 3000 did not show statistically significant differences at any time. Delayed cell migration was observed in all hDPSCs treated with Papacárie Duo and Cariesolut (p<0.01 and p<0.001). Phalloidin staining images showed a high confluence of cells in the presence of NATURAL CARE, similar to the control group. On the contrary, no cells were observed in Brix3000 and Cariesolut at 2% and 0.1% concentrations. Papacárie Duo showed cells at all concentrations, but hDPSCs treated at 0.01% exhibited better proliferation and spreading than those in the control group. Apoptosis assay showed that Brix 3000 at 0.1% and 0.01% had a percentage of live cells higher than 99%, with 68.4% live cells at 2%, 3.69% early apoptotic cells, and 27.9% late apoptotic cells. Conversely, the remaining materials showed abundant apoptotic cells, even at low concentrations. 0.1% and 0.01%of BRIX 3000 did not affect the ROS production levels, while 2% of BRIX 3000 counterparts significantly increased the percentage of CM-H2DCFDA positive cells. Again, all concentrations of Cariesolut showed significantly higher levels of ROS production than those observed in control cells. Our results suggest that Brix 3000 would be the most suitable material for chemical caries removal.