title: 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 p47(phox) 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 p47(phox) or the oxidant responsive transcription factor Nrf2. Taken together, our data demonstrate that ZnO nanoparticles trigger p47(phox) 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.
authors.affiliation: IUF - Leibniz Research Institute for Environmental Medicine, Düsseldorf, Germany.
authors.forename: Verena
authors.initials: V
authors.lastname: Wilhelmi
authors.forename: Ute
authors.initials: U
authors.lastname: Fischer
authors.forename: Heike
authors.initials: H
authors.lastname: Weighardt
authors.forename: Klaus
authors.initials: K
authors.lastname: Schulze-Osthoff
authors.forename: Carmen
authors.initials: C
authors.lastname: Nickel
authors.forename: Burkhard
authors.initials: B
authors.lastname: Stahlmecke
authors.forename: Thomas A J
authors.initials: TA
authors.lastname: Kuhlbusch
authors.forename: Agnes M
authors.initials: AM
authors.lastname: Scherbart
authors.forename: Charlotte
authors.initials: C
authors.lastname: Esser
authors.forename: Roel P F
authors.initials: RP
authors.lastname: Schins
authors.forename: Catrin
authors.initials: C
authors.lastname: Albrecht
mesh_terms: D000818:Animals; D017209:Apoptosis / Q000187:drug effects*; D001854:Bone Marrow Cells / Q000166:cytology / Q000187:drug effects* / Q000378:metabolism; D053148:Caspase 3 / Q000235:genetics* / Q000378:metabolism; D053453:Caspase 9 / Q000172:deficiency / Q000235:genetics; D002460:Cell Line; D053938:DNA Fragmentation / Q000187:drug effects; D005786:Gene Expression Regulation; D006801:Humans; D019169:Jurkat Cells; D008264:Macrophages / Q000166:cytology / Q000187:drug effects* / Q000378:metabolism; D051379:Mice; D008810:Mice, Inbred C57BL; D019255:NADPH Oxidases / Q000235:genetics / Q000378:metabolism; D051267:NF-E2-Related Factor 2 / Q000235:genetics / Q000378:metabolism; D053758:Nanoparticles / Q000633:toxicity*; D009336:Necrosis / Q000139:chemically induced / Q000378:metabolism; D018384:Oxidative Stress; D017382:Reactive Oxygen Species / Q000819:agonists / Q000378:metabolism; D015398:Signal Transduction; D015034:Zinc Oxide / Q000633:toxicity*
chemical_list: D051267:NF-E2-Related Factor 2; C495636:Nfe2l2 protein, mouse; D017382:Reactive Oxygen Species; D019255:NADPH Oxidases; C060505:neutrophil cytosolic factor 1; C505409:Casp3 protein, mouse; D053148:Caspase 3; D053453:Caspase 9; D015034:Zinc Oxide