Interação de nanotubos de carbono de paredes múltiplas com cádmio e seus efeitos em células de peixe-zebra in vitro

Abstract

Carbon nanotubes presence in the environment increases every year as a result of exponential production around the world. In aquatic environments, carbon nanotubes can interact with other pollutants based on their nanoadsorbent characteristics. Heavy metals ions represent one of the biggest concerns in water resources nowadays, as a consequence of anthropogenic actions, in which cadmium (Cd) is one of the most harmful metal for aquatic organisms. This study investigates the effects of oxidized multiwalled carbon nanotubes (ox-MWCNT), of Cd ions and two interaction protocols in zebrafish liver cell line (ZFL): Incubation Protocol A, the ox-MWCNT were incubated in RPMI/L-15 (Roswell Park Memorial Institute and Leibovitz's-15) medium + 10% FBS for 30 min (step 1) and, subsequently, the CdCl2 were added and incubated for more 30 min (step 2); and (B) Incubation Protocol B, the ox-MWCNT were incubated with CdCl2 for 30 min (step 1) and, thereafter, the RPMI/L-15 medium + 10% FBS were added and incubated for more 30 min (step 2). Ox-MWCNT were physical and chemical characterized and its Cd adsorption capacity and colloidal stability in cell culture medium were determined. Cytotoxicity was investigated by four assays and the necrosis and apoptosis events and cell cycle were determined using flow cytometer. Cd content and uptake in cells was analyzed and ten biochemical biomarkers and the effects on DNA damage were evaluated. In this study, Cd presence in medium did not interfere in the protein corona composition of ox-MWCNT but the order of interaction of FBS and Cd can change its colloidal stability and metal adsorption rate. This study demonstrates that ox-MWCNT can increase the ZFL content of cadmium. The treatment with only oxidized ox-MWCNT promotes depletion of catalase, glutathione peroxidase, and glutathione S-transferase and led to alterations in cell cycle with a reduction of cells in G2/M phase. The ox-MWCNT affects Cd toxicity probably due to a “Trojan horse” and synergistic effects of both contaminants and induce apoptosis and necrosis in ZFL cells. We verify that the two co-exposure treatments performed increases the biological cadmium effect in ROS production and led to more genotoxicity damage in DNA. Thus, we show that an alteration in the order protocol method used for ox-MWCNT and Cd interaction in in vitro assays can modify the MWCNT-metal complexes formed and led to different biological effects. This study evidence the modulation of ox-MWCNT on Cd biological effects and contributes to future co-exposure investigations and legislation about with nanomaterials interactions with aquatic pollutants.