title: Intracellular accumulation dynamics and fate of zinc ions in alveolar epithelial cells exposed to airborne ZnO nanoparticles at the air-liquid interface.
abstract: Airborne nanoparticles (NPs) that enter the respiratory tract are likely to reach the alveolar region. Accumulating observations support a role for zinc oxide (ZnO) NP dissolution in toxicity, but the majority of in-vitro studies were conducted in cells exposed to NPs in growth media, where large doses of dissolved ions are shed into the exposure solution. To determine the precise intracellular accumulation dynamics and fate of zinc ions (Zn(2+)) shed by airborne NPs in the cellular environment, we exposed alveolar epithelial cells to aerosolized NPs at the air-liquid interface (ALI). Using a fluorescent indicator for Zn(2+), together with organelle-specific fluorescent proteins, we quantified Zn(2+) in single cells and organelles over time. We found that at the ALI, intracellular Zn(2+) values peaked 3 h post exposure and decayed to normal values by 12 h, while in submerged cultures, intracellular Zn(2+) values continued to increase over time. The lowest toxic NP dose at the ALI generated peak intracellular Zn(2+) values that were nearly three-folds lower than the peak values generated by the lowest toxic dose of NPs in submerged cultures, and eight-folds lower than the peak values generated by the lowest toxic dose of ZnSO4 or Zn(2+). At the ALI, the majority of intracellular Zn(2+) was found in endosomes and lysosomes as early as 1 h post exposure. In contrast, the majority of intracellular Zn(2+) following exposures to ZnSO4 was found in other larger vesicles, with less than 10% in endosomes and lysosomes. Together, our observations indicate that low but critical levels of intracellular Zn(2+) have to be reached, concentrated specifically in endosomes and lysosomes, for toxicity to occur, and point to the focal dissolution of the NPs in the cellular environment and the accumulation of the ions specifically in endosomes and lysosomes as the processes underlying the potent toxicity of airborne ZnO NPs.
authors.affiliation: Environmental Molecular Sciences Laboratory .
authors.forename: Cosmin
authors.initials: C
authors.lastname: Mihai
authors.forename: William B
authors.initials: WB
authors.lastname: Chrisler
authors.forename: Yumei
authors.initials: Y
authors.lastname: Xie
authors.forename: Dehong
authors.initials: D
authors.lastname: Hu
authors.forename: Craig J
authors.initials: CJ
authors.lastname: Szymanski
authors.forename: Ana
authors.initials: A
authors.lastname: Tolic
authors.forename: Jessica A
authors.initials: JA
authors.lastname: Klein
authors.forename: Jordan N
authors.initials: JN
authors.lastname: Smith
authors.forename: Barbara J
authors.initials: BJ
authors.lastname: Tarasevich
authors.forename: Galya
authors.initials: G
authors.lastname: Orr
mesh_terms: D000818:Animals; D018929:Cell Culture Techniques; D002460:Cell Line; D004305:Dose-Response Relationship, Drug; D004847:Epithelial Cells / Q000737:chemistry / Q000187:drug effects / Q000378:metabolism*; D019570:Inhalation Exposure / Q000032:analysis*; D042541:Intracellular Space / Q000737:chemistry / Q000187:drug effects / Q000378:metabolism*; D053768:Metal Nanoparticles / Q000008:administration & dosage* / Q000737:chemistry / Q000633:toxicity; D051379:Mice; D018384:Oxidative Stress / Q000187:drug effects; D011650:Pulmonary Alveoli / Q000166:cytology / Q000187:drug effects / Q000378:metabolism*; D015032:Zinc / Q000032:analysis / Q000737:chemistry / Q000493:pharmacokinetics* / Q000633:toxicity; D015034:Zinc Oxide / Q000008:administration & dosage / Q000737:chemistry / Q000493:pharmacokinetics* / Q000633:toxicity
chemical_list: D015032:Zinc; D015034:Zinc Oxide
keywords: Airborne nanoparticles; FluoZin-3; air–liquid interface; endosomes; intracellular Zn2+; lysosomes