Intracellular accumulation dynamics and fate of zinc ions in alveolar epithelial cells exposed to airborne ZnO nanoparticles at the air-liquid interface.
Intracellular accumulation dynamics and fate of zinc ions in alveolar epithelial cells exposed to airborne ZnO nanoparticles at the air-liquid interface.
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DOI：
10.3109/17435390.2013.859319
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发表时间：
2015-02
期刊：
影响因子：
5
通讯作者：
Orr G
中科院分区：
文献类型：
--
作者：
Mihai C;Chrisler WB;Xie Y;Hu D;Szymanski CJ;Tolic A;Klein JA;Smith JN;Tarasevich BJ;Orr G
进入呼吸道的空气中的纳米粒子（NP）很可能到达肺泡区域。积累的观察结果支持氧化锌 (ZnO) NP 溶解在毒性中的作用，但大多数体外研究是在暴露于生长培养基中的 NP 的细胞中进行的，其中大剂量的溶解离子脱落到暴露溶液中。为了确定细胞环境中空气中纳米粒子释放的锌离子 (Zn2+) 的精确细胞内积累动态和命运，我们将肺泡上皮细胞暴露于气液界面 (ALI) 处的雾化纳米粒子。使用 Zn2+ 荧光指示剂以及细胞器特异性荧光蛋白，我们对单细胞和细胞器中的 Zn2+ 随着时间的推移进行了定量。我们发现，在 ALI 中，细胞内 Zn2+ 值在暴露后 3 小时达到峰值，并在 12 小时内衰减至正常值，而在深层培养物中，细胞内 Zn2+ 值随着时间的推移持续增加。 ALI 处的最低毒性 NP 剂量产生的细胞内 Zn2+ 峰值值比深层培养物中最低毒性剂量的 NP 产生的峰值低近三倍，比最低毒性剂量 ZnSO4 或 Zn2+ 产生的峰值低八倍。在 ALI 中，早在暴露后 1 小时就在内体和溶酶体中发现了大部分细胞内 Zn2+。相比之下，暴露于 ZnSO4 后，大部分细胞内 Zn2+ 存在于其他较大的囊泡中，内体和溶酶体中的含量不到 10%。总之，我们的观察结果表明，细胞内 Zn2+ 必须达到较低但关键的水平，特别集中在内体和溶酶体中，才会发生毒性，并指出纳米颗粒在细胞环境中的局部溶解以及离子特别在内体和溶酶体中的积累，这是空气中 ZnO 纳米颗粒潜在毒性的过程。
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 (Zn2+) 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 Zn2+, together with organelle-specific fluorescent proteins, we quantified Zn2+ in single cells and organelles over time. We found that at the ALI, intracellular Zn2+ values peaked 3 h post exposure and decayed to normal values by 12 h, while in submerged cultures, intracellular Zn2+ values continued to increase over time. The lowest toxic NP dose at the ALI generated peak intracellular Zn2+ 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 Zn2+. At the ALI, the majority of intracellular Zn2+ was found in endosomes and lysosomes as early as 1 h post exposure. In contrast, the majority of intracellular Zn2+ 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 Zn2+ 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.