In vitro embryotoxicity assessment with dental restorative materials

Objectives Resin (co)monomers may be released from restorative dental materials and can diffuse into the tooth pulp or the gingiva, and can reach the saliva and the circulating blood. Genotoxic potential of some dental composite components has been clearly documented. The genotoxic effects of xenobiotics can represent a possible step in tumor initiation and/or embryotoxicity/teratogenesis. A modified fluorescent mouse embryonic stem cell test (R.E.Tox®) was used to test the embryotoxic potential of following dental restorative materials: Bisphenol A glycidylmethacrylate (BisGMA), urethanedimethacrylate (UDMA), hydroxyethylmethacrylate (HEMA), and triethyleneglycoldimethacrylate (TEGDMA), as well as some of their metabolic intermediates 2,3-epoxy-2-methyl-propionicacid-methylester (EMPME), methacrylic acid (MA), and 2,3-epoxy-2-methylpropionic acid (EMPA). Methods Mouse embryonic stem (ES) cells stably transfected with a vector containing the gene for the green fluorescent protein under control of the cardiac α-myosin heavy chain promoter were differentiated in the presence of various concentrations of the test compounds for 12 days. Fluorescence was measured using the TECAN Safire® and values were expressed as percent of control values. To distinguish between cytotoxic and embryotoxic effects, all compounds were tested in a standard MTT assay. Results HEMA, TEGDMA and EMPME did not influence the differentiation process of ES cells towards cardiac myocytes. No cytotoxic effects were observed at any of the concentration levels tested. Exposure to BisGMA resulted in a 50% decrease in cell survival and a very strong inhibition of cell differentiation at 10−5 M (p<0.01). Embryotoxic effects were also present at 10−6 and 10−7 M (p<0.05). EMPA induced a decrease in ES cell differentiation at 10−5 M (p<0.01) without cytotoxic effects. No embryotoxic effects were induced at lower concentrations. Exposure to UDMA resulted in a slight decrease of cell differentiation at 10−5 M (p<0.05). Exposure of cells to MA resulted in an increase of cardiac differentiation up to 150% (p<0.05) at 10−5 M without cytotoxic effects. Conclusions BisGMA induced a significant high embryotoxic/teratogenic effect over a large range of concentration. Therefore attention should be focused on this dental monomer, which should be investigated further by in vivo experiments.