Induction of DNA double-strand breaks by monochlorophenol isomers and ChKM in human gingival fibroblasts.

Phenol has been traditionally used in dental treatment as a sedative for the pulp or as disinfectant for carious cavity and root canal. However, phenol is regarded as a mutagenic and carcinogenic agent and its use in dental practice is now therefore restricted. Monochlorophenols are derivatives of phenol, which are still used clinically as root canal disinfectants, they are even more active antiseptics/disinfectants than phenol, and the so-called Walkhoff (ChKM) solution makes use of monochlorophenol for root canal disinfection. Ingredients in the ChKM solution are the monochlorophenol compound 4-chlorophenol (4-CP), camphor, and menthol. In literature, the use of the ChKM solution is controversial because of a possible DNA toxicity of the ingredient 4-CP. However, it is unknown whether ChKM can really induce DNA damage in human oral cells. In this study, the induction of DNA double-strand breaks (DSBs) by ChKM and monochlorophenol compounds (2-chlorophenol, 2-CP; 3-chlorophenol, 3-CP; and 4-chlorophenol, 4-CP) was tested in human gingival fibroblasts (HGFs). DNA DSBs (foci) induced in HGFs unexposed and exposed to monochlorophenols or ChKM solution were investigated using the γ-H2AX DNA focus assay, which is a direct marker for DSBs. DSBs result in the ATM-dependent phosphorylation of the histone H2AX. When cells were exposed to medium or medium + DMSO (1 %) (negative controls), an average of 3 foci per cell were found. In positive control cells (H2O2 + medium, or H2O2 + medium + DMSO (1 %), an average of 35 foci each were found. About 20 DSB foci per cell were found, when HGFs were exposed to 2-CP (4 mM), 3-CP (2.3 mM), 4-CP (2.1 mM), or ChKM (corresponding to 1.5 mM 4-CP). Our results show increasing DNA toxicities in the order of 2-CP < 3-CP < 4-CP < ChKM solution. An additive DNA toxicity was found for 4-CP in combination with camphor in the ChKM solution, compared to the 4-CP alone. No significant differences regarding multi-foci cells (cells that contain more than 40 foci) were found when HGFs were exposed to the EC50 concentrations (given in parenthesis) of ChKM (1.5 mM), 4-CP (2.1 mM), or 2-CP (4 mM). Significantly fewer multi-foci cells were found when HGFs were exposed to the EC50 concentration (given in parenthesis) of 3-CP (2.3 mM), compared to the EC50 concentrations of ChKM, 4-CP, or 2-CP. Monochlorophenol compounds and/or ChKM solution can induce DSBs in primary human oral (cavity) cells, which underscores their genotoxic capacity.

Quantification of elutable substances from methacrylate based sealers and their cytotoxicity effect on with human gingival fibroblasts.

OBJECTIVES: Previous studies have shown that resin composites may cause persistent inflammation of oral or pulpal tissues as well as cell death through eluted substances. The aim of this study was to investigate the leaching of ingredients from commercial dental fissure sealers as well as their cytotoxic effects on human gingival fibroblast (HGF). METHODS: The sealers tested were: Helioseal(®) F, Helioseal(®) Refill, Fissurit(®) F, Grandio(®) Seal, Ultraseal XT(®) plus and Delton(®) FS. Ten discs of each sealer were respectively immersed in methanol or water and incubated at 37°C. The eluates were analysed by gas chromatography/mass spectrometry at day 1, 3 and 7. In the XTT-test, eight discs of each fissure sealer were immersed into medium. The eluates of the respective sealer were mixed and used undiluted and diluted with medium. HGF were incubated with the dilutions at 37°C for 24h. Then XTT-salt was added and the XTT-formazan formation was quantified. RESULTS: In eluates from polymerized sealers, comonomers (triethylene glycol dimethacrylate (TEGDMA)) and additives were found (e.g. camphorquinone (CQ), butylated hydroxytoluene, triphenylstibane). 7 d after the beginning of the experiments the highest amount of TEGDMA was found in the aqueous eluate from Grandio(®) Seal (9944.31 (2250.56) µmol/l). The most cytotoxic eluate found in the XTT-test was from Fissurit(®) F (EC50 value at 27.13 (7.04)%; (mean(SD)). SIGNIFICANCE: Because of the use of sealers in preventative dental medicine it should be taken into account that substances like TEGDMA or CQ, that are often causing allergic reactions, are elutable. Before using the sealers patients should be asked for allergic reactions to these substances.