The Toxicity of Universal Dental Adhesives: An In Vitro Study

Abstract

There is no consensus in the literature regarding the potential toxicity of universal dental adhesives (UDA). Being used in close proximity to the pulp, their biocompatibility should be an important factor in dental research. The aim of the present study was to evaluate the biocompatibility of UDA in an in vitro model. The study was performed using a monocyte/macrophage peripheral blood SC cell line (ATCC CRL-9855) on four specific UDA, namely: All-Bond Universal (Bisco); CLEARFIL Universal Bond Quick (Kuraray); G-Premio BOND (GC); Single Bond Universal (3M ESPE). The cytotoxicity of the investigated UDA was measured using the XTT colorimetric assay. The genotoxicity of the analyzed compounds was evaluated using an alkaline version of the comet assay. Furthermore, flow cytometry (FC) apoptosis detection was performed using the FITC Annexin V Apoptosis Detection Kit I. FC cell-cycle arrest assessment was performed using propidium iodide staining. The study observed significant differences in the toxicity of the UDA that were tested, as G-Premio BOND showed significant in vitro toxicity in all of the tests performed, while All-Bond Universal, CLEARFIL Universal Bond Quick and Single Bond Universal did not present any significant toxic effects toward SC cell line. The in vitro toxicity of UDA should be taken into consideration prior to in vivo and clinical studies. The flow cytometry could improve the accuracy of dental materials research and should be incorporated into the standardization criteria.

Keywords: biocompatibility; cytotoxicity; dental materials; flow cytometry; genotoxicity; universal dental adhesives.

Figure 1

Cytotoxicity of the investigated adhesives. Test performed using the Cell Proliferation Kit II (XTT) assay after 24 h incubation of cells with the tested compounds. The positive control constituted cells were suspended in 96% isopropyl alcohol, while the negative control cells were cultured in a complete medium. The differences were statistically significant on the graphs as follows: *** p < 0.001 versus negative control.

Figure 2

Genotoxicity of the investigated adhesives. Analysis was performed using an alkaline version of the comet assay after 24 h incubation of cells with the tested compounds. Cells suspended in 10% DMSO constituted a positive control. Cells suspended in 1 mL of complete culture medium constituted a negative control. The differences were statistically significant on the graphs as follows: *** p < 0.001 versus negative control.

Figure 3

Flow cytometric FITC annexin V/propidium iodide (PI) double staining analysis of apoptosis after 24 h incubation of cells with the tested compounds. Dot plot graphs indicate the percentage of viable (FITC annexin V negative, PI negative), early apoptotic (FITC annexin V positive, PI negative) late apoptotic (FITC annexin V positive, PI positive) and necrotic (FITC annexin V negative, PI positive) cells. Cells treated with staurosporine at a concentration of 1 µM constituted a positive control. Cells suspended in the complete culture medium constituted a negative control.

Figure 4

Flow cytometry (FC) analysis of cell cycle progression using the propidium iodide (PI) staining after 24 h incubation of cells with the tested compounds. Cells treated with 1 µM nocodazole constituted a positive control. Cells cultured in the complete medium constituted a negative control. The differences were statistically significant on the graphs as follows: * p < 0.01, *** p < 0.001 versus negative control.