Abstract
To identify the respective contributions of released zinc and solid particles on the cytotoxicity of zinc oxide nanoparticles (ZnO-NPs), we exposed A549 cells to ZnO-NP suspensions, their extractions collected after centrifugation, and medium containing zinc chloride (ZnCl2). We then assayed the cytotoxicity of these samples using water-soluble tetrazolium salts (WSTs) and intracellular reactive oxygen species (ROS) with 2′,7′-dichlorodihydrofluorescin diacetate (DCFH-DA) as outputs. Only the ZnO-NP suspension caused cytotoxicity and increased intracellular ROS; the extractions and ZnCl2 did not cause cytotoxicity or oxidative stress. Global gene expression analysis revealed that both ZnCl2-containing medium and the ZnO-NP suspension caused upregulation of the "cadmium binding" gene functional category. This category consisted of metallothioneins (MTs), important zinc-homeostasis proteins. To further investigate the role of MTs in ZnO-NP-dependent cytotoxicity, we inhibited the overexpression of MTs with corresponding siRNA and found that released zinc contributed to ZnO-NP-dependent cytotoxicity. We conclude that both solid particles and released zinc contributed to ZnO-NP-dependent cytotoxicity. Additionally, we propose a synergic relationship between ZnO-NPs and MTs.
