Pulmonary exposure to zinc oxide nanoparticles (ZnONPs) could cause acute lung injury (ALI), but the underlying molecular mechanism remains unclear. Herein, we established a ZnONPs-induced ALI mouse model, characterized by the histopathological changes (edema and infiltration of inflammatory cells in lung tissues), and the elevation of total protein and cytokine interleukin-6 in bronchoalveolar lavage fluid in time- and dose-dependent manners. This model also exhibited features like the disturbance of redox-state (reduced of glutathione to glutathione disulfide ratio, elevation of heme oxygenase-1 and superoxide dismutase 2), the decrease of adenosine triphosphate synthesis and the release of zinc ions in the lung tissues. Interestingly, we found that ZnONPs exposure caused the accumulation of autophagic vacuoles and the elevation of microtubule-associated proteins 1A/1B light chain (LC)3B-II and p62, indicating the impairment of autophagic flux. Our data indicated that the above process might be regulated by the activation of AMP-activated protein kinase but not the mammalian target of rapamycin pathway. The association between ZnONPs-induced ALI and autophagy was further verified by a classical autophagy inhibitor, 3-methyladenine (3-MA). 3-MA administration reduced the accumulation of autophagic vacuoles, the expression of LC3B-II and p62, followed by a significant attenuation of histopathological changes, inflammation, and oxidative stress. More importantly, 3-MA could directly decrease the release of zinc ions in lung tissues. Taken together, our study provides the evidence that ZnONPs-induced pulmonary toxicity is autophagy-dependent, suggests that limiting the release of zinc ions by inhibiting autophagy could be a feasible strategy for the prevention of ZnONPs-associated pulmonary toxicity.

中文翻译：

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自噬依赖性释放锌离子对于氧化锌纳米颗粒触发的急性肺损伤至关重要

肺暴露于氧化锌纳米颗粒（ZnONPs）可能引起急性肺损伤（ALI），但其潜在的分子机制仍不清楚。在此，我们建立了ZnONPs诱导的ALI小鼠模型，其特征在于组织病理学改变（肺组织中的水肿和炎症细胞浸润），以及支气管肺泡灌洗液中总蛋白和细胞因子白细胞介素6在时间和剂量上的升高依赖的方式。该模型还具有氧化还原状态紊乱（谷胱甘肽与谷胱甘肽二硫化物比率降低，血红素加氧酶-1和超氧化物歧化酶2升高），三磷酸腺苷合成减少和锌离子在肺组织中释放等特征。有趣的是，我们发现ZnONPs暴露导致自噬泡的积累和微管相关蛋白1A / 1B轻链（LC）3B-II和p62的升高，表明自噬通量受损。我们的数据表明，上述过程可能受AMP激活的蛋白激酶激活的调控，但不受雷帕霉素途径的哺乳动物靶标的调控。ZnONPs诱导的ALI与自噬之间的关联已通过经典的自噬抑制剂3-甲基腺嘌呤（3-MA）进一步证实。3-MA给药减少了自噬泡的积累，LC3B-II和p62的表达，随后显着减弱了组织病理学变化，炎症和氧化应激。更重要的是，3-MA可以直接减少肺组织中锌离子的释放。在一起