Mutation Research/Genetic Toxicology and Environmental Mutagenesis
Sister-chromatid exchange, chromosomal aberrations and delays in cell-cycle kinetics in human lymphocytes induced by dental composite resin eluates
Introduction
Dental composite resin-based materials have been widely studied for cytotoxicity in various cell culture systems [1], [2]. Their cytotoxic effects reported thus far have been attributed to the release of residual monomers or other substances derived either from incomplete polymerization or resin degradation [3], [4]. More than 30 different compounds have been identified as being released from the polymerized dental composites, including major monomers, co-monomers, additives and reaction products, all with different cytotoxic potential [5]. The released components may diffuse through the dentine into the pulp space or, alternatively, they may either come in contact with the gingiva, or be swallowed via the saliva, taken up by the circulating blood and excreted with the urine [6].
There are a few studies dealing with the possible mutagenicity and genotoxicity of various single components released from polymerized composite resins. TEGDMA, bisphenol A and HEMA have been found to increase the number of micronuclei at sub-toxic concentrations and promote degradation of DNA from salivary gland tissue and lymphocytes, as shown by a comet assay [7], [8], [9]. TEGDMA was also reported to induce extensive deletions of sequences in the hypoxanthine-guanine phosphoribosyl-transferase (Hprt) gene in V79 Chinese hamster lung fibroblasts [10]. Other substances released from composite resins, such as the photo-initiator camphoroquinone, have been found to promote oxidative DNA damage via the generation of reactive oxygen species (ROS) [11].
There is quite limited knowledge concerning the genotoxic and mutagenic effects of the many compounds released in eluates of commercially available, widely used composite resins and dental adhesives [12], [13]. Monitoring the genotoxicity of these eluates would provide a better understanding of their interaction with the oral tissues and the potential to cause genotoxic effects in vivo.
Recently, we have observed that eluates from specific direct composite resins show extensive time- and concentration-dependent cytotoxicity to murine WEHI 13 var fibroblasts, accompanied by induction of DNA fragmentation, as shown by agarose gel electrophoresis [14]. This prompted us to extend these observations in the present study, which was aimed at testing the effect of five widely used composite resins for direct and indirect dental restorations with respect to induction of sister-chromatid exchange (SCE), chromosomal aberrations (CAs), and effects on cell-cycle kinetics and mitotic indices of human peripheral lymphocytes. SCE have been proposed as a very sensitive indicator of DNA damage and/or subsequent DNA repair. In combination with the evaluation of chromosome aberrations, SCE can provide insight in the cytogenetic damage induced by various genotoxic agents at very low concentrations. Moreover, the determination of proliferation rates and mitotic indices in lymphocyte cultures has been proved to be a very useful and sensitive indicator of the cytostatic and cytotoxic action of various environmental hazards or therapeutic agents [15], [16]. Our test hypothesis was that laboratory-polymerized (indirect) composite resins would be less cytotoxic and genotoxic compared with conventional direct composites.
Section snippets
Materials and sample preparation
Two hybrid composites (Tetric Ceram/Ivoclar-Vivadent, Schaan, Lichtenstein and Filtek Z-250/3M ESPE, Seefeld, Germany), one nano-hybrid (Simile/Pentron, Wallingford, USA) and two second-generation laboratory composites for indirect restorations (Adoro/Ivoclar-Vivadent, Schaan, Lichtenstein and Conquest Sculpture/Pentron Wallingford, CT, USA) were tested (Table 1). Specimens of each biomaterial were prepared aseptically with a prefabricated mould as disks of 5-mm diameter and 2-mm thickness,
Results
All eluates from direct composite resins (Tetric Ceram, Simile and Filtek Z-250) induced a statistically significant increase in the frequencies of SCE, accompanied by suppression of PRI and MI in primary cultures of human lymphocytes, as shown in all three experiments, each using blood from a different donor (Table 2, Table 3, Table 4). The increase in SCE was comparable to that induced by the lower test concentrations of melphalan (i.e. 0.5 and 1.0 μM), a well-known alkylating agent that was
Discussion
According to the data presented in this study, eluates from the three direct composites (Tetric Ceram, Filtek Z-250 and Simile) were genotoxic, since they markedly increased the frequency of SCE in human lymphocyte cultures, up to levels comparable to those induced by 0.5–1.0 μM of melphalan, a well-known alkylating agent that was used as a positive control. The extracts were also clastogenic, given the significant increase in the number of CAs, up to values that were much higher than those
Acknowledgements
Dr. A.A. Bakopoulou wishes to thank “Alexandros S. Onassis” Public Benefit Foundation in Athens, Greece, for providing her with a post-graduate fellowship.
References (43)
- et al.
Responses of L929 mouse fibroblasts, primary and immortalized bovine dental papilla-derived cell lines to dental resin components
Dent. Mater.
(2002) - et al.
Cytotoxic and genotoxic effects of resin monomers in human salivary gland tissue and lymphocytes as assessed by the single cell microgel electrophoresis (Comet) assay
Biomaterials
(2006) - et al.
Interference with microtubules and induction of micronuclei in vitro by various bisphenols
Mutat. Res.
(1997) - et al.
Triethylene glycol dimethacrylate induces large deletions in the hprt gene of V 79 cells
Mutat. Res.
(1999) - et al.
The effect of camphoroquinone (CQ) and CQ-related photosensitizers on the generation of reactive oxygen species and oxidative DNA damage
Biomaterials
(2005) - et al.
Cytotoxic and mutagenic effects of dental composite materials
Biomaterials
(2005) - et al.
Post cure heat treatments for composites: properties and fractography
Dent. Mater.
(1992) - et al.
Annealing as a mechanism of increasing wear resistance of composites
Dent. Mater.
(1990) - et al.
Mass loss in urethane/TEGDMA-and-Bis-GMA/TEGDMA-based resin composites during post-cure heating
Dent. Mater.
(1997) - et al.
Quantification of organic eluates from four polymer-based dental restorative materials by use of GC/MS
J. Chromatogr. B Anal. Technol. Biomed. Life Sci.
(2007)