The increasing risk of incidental exposure to nanomaterials has led to mounting concerns regarding nanotoxicity. Zinc oxide nanoparticles (ZnO NPs) are produced in large quantities and have come under scrutiny due to their capacity to cause cytotoxicity in vitro and potential to cause harm in vivo. Recent evidence has indicated that ZnO NPs promote autophagy in cells; however, the signaling pathways and the role of ion release inducing toxicity remain unclear. In this study, we report that ZnO NPs are immunotoxic to primary and immortalized immune cells. Importantly, such immunotoxicity is observed in mice in vivo, since death of splenocytes is seen after intranasal exposure to ZnO NPs. We determined that ZnO NPs release free Zn<sup>2+</sup> that can be taken up by immune cells, resulting in cell death. Inhibiting free Zn<sup>2+</sup> ions in solution with EDTA or their uptake with CaCl<inf>2</inf> abrogates ZnO NP-induced cell death. ZnO NP-mediated immune cell death was associated with increased levels of intracellular reactive oxygen species (ROS). ZnO NP death was not due to apoptosis, necroptosis or pyroptosis. Exposure of immune cells to ZnO NPs resulted in autophagic death and increased levels of LC3A, an essential component of autophagic vacuoles. Accordingly, ZnO NP-mediated upregulation of LC3A and induction of immune cell death were inhibited by blocking autophagy and ROS production. We conclude that release of Zn<sup>2+</sup> from ZnO NPs triggers the production of excessive intracellular ROS, resulting in autophagic death of immune cells. Our findings suggest that exposure to ZnO NPs has the potential to impact host immunity.

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Department of Immunology

Johnson, B.M, Fraietta, J.A, Gracias, D.T, Hope, J.L, Stairiker, C.J, Patel, P.R, … Katsikis, P.D. (2015). Acute exposure to ZnO nanoparticles induces autophagic immune cell death. Nanotoxicology, 9(6), 737–748. doi:10.3109/17435390.2014.974709