Involvement of oxidative stress in mutagenicity and apoptosis caused by dental resin monomers in cell cultures

Abstract

Objective This investigation studied the possibility that apoptosis as well as mutagenicity induced by resin monomers are mediated by oxidative stress. Methods A range of dilutions of three resin monomers (GMA, TEGDMA, and HEMA) was added to culture medium (DMEM/10% FBS), of V79-4 fibroblasts and RPC-C2A pulp cells for 24 h. Their cytotoxic effects were measured by a colorimetric functional assay (MTT). Chromosomal aberration induced by the resin monomers was investigated by counting micronuclei in V79-4 cells. The effects of the resin monomers on DNA fragmentation were viewed by agarose gel electrophoresis of DNA, isolated from RPC-C2A pulp cells that were treated by resin compounds. Resin monomer-induced apoptosis was further confirmed by flow cytometry (staining with both annexin V-FITC and PI). Results All monomers exhibited a dose-dependent cytotoxic effect, and the ranking of the cytotoxicity based on TC 50 was GMA > TEGDMA > HEMA. The resin monomer-induced cytotoxicity was significantly decreased by co-treatment with N-acetylcystein (NAC), an antioxidant. The authors also confirmed a dose-dependent genotoxicity of the resin monomers that had induced micronucleated cells in V79-4 fibroblasts. Similar to the effects on cytotoxicity, NAC reduced the numbers of micronuclei in comparison with those generated by the resin monomers. The preventive effects of NAC were also observed in monomer-induced apoptosis in RPC-C2A cells. A DNA ladder pattern, characteristic of apoptosis, was shown at cytotoxic concentrations, but NAC blocked the resin monomer-mediated DNA fragmentation. The preventive effects of NAC on apoptosis were confirmed by Annexin V staining. Cells exposed to 300 μM GMA, 7 mM TEGDMA, or 14 mM HEMA for 24 h showed a significant increase in apoptotic cells, while NAC co-treatment caused a reduction in apoptotic cells compared to controls. Significance These findings suggest that glutathione depletion and oxidative stress are responsible for GMA, TEGDMA, and HEMA-induced mutagenicity and apoptosis.