To evaluate cytotoxicity of five dentin-bonding agents in terms of cytotoxicity by XTT assay, using Human Gingival Fibroblasts (HGF).
Method:
Samples of each dentin-bonding agents were prepared in black 96-well microplates under laminar air flow. Total volume of 10 µl from each material was pipetted into the wells. The ratio of sample surface area to the volume of the solution was adjusted to approximately 4.32 cm2/mL, and the bonding agents were polymerized. Total of 24 wells were prepared for each material. 200µl Dulbecco’s Modified Eagle Medium (DMEM) was added in each well and plates were incubated in highly humidified atmosphere of 5% CO2 at 37°C. Next day, medium was collected; samples were rinsed and again, topped with 200 µl DMEM. Collected medium extracts were kept in sterile eppendorf tubes at -20◦C until cytotoxicity experiments. This procedure was repeated for 24h intervals from the 1st to the 7th day, and then on the 14th, 21st and 28th day. To evaluate cytotoxicity of dentin bonding agents, three different HGF cell lines used and XTT analyze of each cell line was repeated three times. One-way ANOVA, Post-Hoc Dunnett T3 and Bonferroni Tests were used at a significance level of p < 0.05.
Result:
Cell viabilities were 7.76% for Admira, 6.60% for Adper, 24.06% Clearfil S3, 6.58% for Clearfil SE and 6.00% for Heliobond on the first day, and they were evaluated as severely cytotoxic (p<0.001). The cell viability of Admira, Clearfil SE, Clearfil S3 and Heliobond were similar on days 2 to 28 and viabilities were still under 30 per cent. For Adper single bond, viability began to increase on 14th day (18.25%) and reached to 30.78% on 28th day.
Conclusion:
Dentin-bonding agents showed severe cytotoxicity with viability results lower than 10%. Toxicity continued to 28th day except Adper Single Bond.