Avaliação da segurança in vitro de nanotubos de carbono de paredes múltiplas funcionalizados com polietilenoglicol em linhagem de fibroblasto de pulmão de hamster (V79-4)

Abstract

Carbon nanotubes (CNT) comprise an important class of materials with excellent physicochemical properties, which have received great attention in biological applications. Conflicting data regarding their cytotoxic effects have been reported. This discrepancy is due to the lack of methodologies standardization, which should take into account not only the dose and time of exposure to the agent, a parameter well established in classical toxicology, but also the intrinsic characteristics of the CNT and their behavior at the interface with the system biological. Thus, despite the large amount of data generated in the last 10 years, on the toxicity of these nanomaterials, nanotoxicology is still a great challenge. The objective of this study was to evaluate the cytotoxic, genotoxic and mutagenic potential of multi walled carbon nanotubes, functionalized with polyethylene glycol modified with pyrene (MWNT- PyPEG), after exposure of the hamster lung fibroblast cell line (V79-4). The research was conducted using the MTT and Cell Titer Blue methods to estimate cell viability and by means of the CBMN-Cyt with (+S9) or without (-S9) metabolic activation tests and comet assay to evaluate the mutagenicity and genotoxicity. The MTT method, in the range of concentrations tested (1.2 to 18.5 g mL-1), showed a significant decrease in the cellular viability of cells treated with MWNT- PyPEG at the concentration of 18.5 g mL-1 in relation to the control group (P <0.05) at different times (24, 48 and 72 h). Pyrene-PEG and other concentrations showed no change in cell viability compared to the control group. The concentration of 18.5 g mL-1 was cytotoxic, since cell viability was below 50% inhibitory concentration (IC50). A dose-response effect was observed when MWNT- PyPEG concentrations were used between 9.25 g mL-1 and 18.5 g mL-1. The CellTiterBlue® assay performed under the same conditions above, and only within 24 h, confirmed the MTT results. Results obtained with the CBMN-Cyt (-S9) and comet assay did not demonstrate genotoxicity and mutagenicity in the concentrations tested. However, when the MWNT- PyPEG were subjected to the CBMN-Cyt (+S9) assay, the number of micronucleus increased significantly over the negative control, and was similar to the positive control. Therefore, the biotransformation process of these NPs must be carefully investigated, since this system can potentiate their interaction with the genetic material, promoting mutations and consequently leading to the process of carcinogenesis. These results show the need and importance of more than one genotoxicity test to evaluate the safety of each compound.