Zinc Oxide Nanoparticles Induce Necrosis and Apoptosis in Macrophages in a p47phox- and Nrf2-Independent Manner

Abstract

In view of the steadily increasing use of zinc oxide nanoparticles in various industrial and consumer applications, toxicological investigations to evaluate their safety are highly justified. We have investigated mechanisms of ZnO nanoparticle-induced apoptosis and necrosis in macrophages in relation to their important role in the clearance of inhaled particulates and the regulation of immune responses during inflammation. In the murine macrophage RAW 264.7 cell line, ZnO treatment caused a rapid induction of nuclear condensation, DNA fragmentation, and the formation of hypodiploid DNA nuclei and apoptotic bodies. The involvement of the essential effector caspase-3 in ZnO-mediated apoptosis could be demonstrated by immunocytochemical detection of activated caspase-3 in RAW 264.7 cells. ZnO specifically triggered the intrinsic apoptotic pathway, because Jurkat T lymphocytes deficient in the key mediator caspase-9 were protected against ZnO-mediated toxicity whereas reconstituted cells were not. ZnO also caused DNA strand breakage and oxidative DNA damage in the RAW 264.7 cells as well as p47phox NADPH oxidase-dependent superoxide generation in bone marrow-derived macrophages. However, ZnO-induced cell death was not affected in bone marrow-derived macrophages of mice deficient in p47phox or the oxidant responsive transcription factor Nrf2. Taken together, our data demonstrate that ZnO nanoparticles trigger p47phox NADPH oxidase-mediated ROS formation in macrophages, but that this is dispensable for caspase-9/3-mediated apoptosis. Execution of apoptotic cell death by ZnO nanoparticles appears to be NADPH oxidase and Nrf2-independent but rather triggered by alternative routes.

In vitro cytotoxicity of SiO2 or ZnO nanoparticles with different sizes and surface charges on U373MG human glioblastoma cells

“…34 However, Wilhelmi et al reported that ZnO NPs induce necrosis and apoptosis in macrophages via ROS-and p53-independent pathway. 24 Consistent with the latter study, we found that both SiO 2 and ZnO NPs induced apoptosis in U373MG cells, which express mutant p53. Other authors have suggested that, in addition to the ROS-mediated p53 activation pathway, SiO 2 and ZnO NPs may activate the p38 mitogen-activated protein kinase and/or c-Jun N-terminal kinase pathways to transactivate proapoptotic genes and induce apoptosis.…”

“…[20][21][22][23] However, a recent study found that ZnO NPmediated apoptosis was not related to ROS generation or the p53 pathway. 24 Thus, the cell-death pathways mediated by SiO 2 and ZnO NPs are still the subject of some debate. In this study, we investigated the cytotoxic effects of SiO 2 and ZnO NPs on the U373MG human glioblastoma cell line.…”

Effects of zinc oxide nanoparticles on Kupffer cell phagosomal motility, bacterial clearance, and liver function

“…10,34,35 The observed toxicities have been linked to their physicochemical properties, such as dissolution into ionic Zn and their ability to generate ROS. 36 Consequently, these same properties have been exploited in the development of nanomedicines.…”

Analysis of cellular responses of macrophages to zinc ions and zinc oxide nanoparticles: a combined targeted and proteomic approach

“…RAW264 cells are one of the classical macrophage models and have been often used for studying the effects of nanoparticles in macrophages (e.g. in 26,[28][29][30][31][32][33] . They are also one of the few cell lines able to produce NO upon LPS stimulation 34 , a feature absent from the other classical human macrophage model THP-1.…”