1 School of Marine Sciences, Ningbo University, Ningbo 315800, China; 2 Ningbo Marine Environment Monitoring Center Station, The State Oceanic Administration, Ningbo 315012, China
Abstract：The wide scale use of zinc oxide nanoparticles (ZnO NPs) makes organisms beings more prone to the exposure to ZnO NPs and its adverse effects.To investigate the toxic effect of ZnO NPs in monocytes/macrophages (MO/MΦ) from red drum (Sciaenops ocellatus), MO/MΦ from red drum were respectively exposed to the different concentrations of ZnO NPs (0, 12.5, 25, 50 μg/mL) for 6, 12, 24 h. Cell activity was determined by 3-(4, 5-dimethyl thiazol-2-yl)-2, 5-diphenyl tetrazolium bromide (MTT) assay, neutral red uptake (NRU) assay and lactate dehydrogenase (LDH) release. Cellular uptake of ZnO NPs, intracellular reactive oxygen generation and apoptosis rate were measured by flow cytometry after exposure to ZnO NPs for 6 h. The mRNA expressions of Caspase 9 in MO/MΦ were also examined using qRT-PCR assays.Compared with the control group, the experimental groups showed obvious difference in following aspects: 1) The activity of the cells detected by the MTT assay and the ability of the cells to absorb neutral red all decreased with the increase of ZnO NPs concentration and the extension of exposure time; 2) The release of LDH were increased with the extension of ZnO NPs exposure time and the increase of ZnO NPs concentration; 3) Cellular uptake of ZnO NPs was increased; 4) Intracellular reactive oxygen species (ROS) generation was increased; 5) Cell apoptosis rate showed an increasing trend; 6) mRNA expression level of gene Caspase 9 showed significant up-regulation (P＜0.05). These results demonstrated concentration and time dependent cytotoxicity after exposure to ZnO NPs on MO/MΦ from red drum. Intracellular ROS may be the major factor in cytotoxicity of ZnO NPs. When ZnO NPs entered cells, ZnO NPs increased the level of intracellular ROS which led to apoptosis. The above research provides a theoretical basis to study toxic effect of ZnO NPs to aquatic organisms.
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