Meakins-Christie Laboratories, Department of Medicine, McGill University, Montreal, Quebec, Canada.
Am J Respir Cell Mol Biol. 2015 Apr;52(4):513-22. doi: 10.1165/rcmb.2013-0430OC.
Chlorine gas (Cl2) inhalation causes oxidative stress, airway epithelial damage, airway hyperresponsiveness (AHR), and neutrophilia. We evaluated the effect of neutrophil depletion on Cl2-induced AHR and its effect on the endogenous antioxidant response, and if eosinophils or macrophages influence Cl2-induced AHR. We exposed male Balb/C mice to 100 ppm Cl2 for 5 minutes. We quantified inflammatory cell populations in bronchoalveolar lavage (BAL), the antioxidant response in lung tissue by quantitative PCR, and nuclear factor (erythroid-derived 2)-like 2 (NRF2) nuclear translocation by immunofluorescence. In vitro, NRF2 nuclear translocation in response to exogenous hypochlorite was assessed using a luciferase assay. Anti-granulocyte receptor-1 antibody or anti-Ly6G was used to deplete neutrophils. The effects of neutrophil depletion on IL-13 and IL-17 were measured by ELISA. Eosinophils and macrophages were depleted using TRFK5 or clodronate-loaded liposomes, respectively. AHR was evaluated with the constant-phase model in response to inhaled aerosolized methacholine. Our results show that Cl2 exposure induced neutrophilia and increased expression of NRF2 mRNA, superoxide dismutase-1, and heme-oxygenase 1. Neutrophil depletion abolished Cl2-induced AHR in large conducting airways and prevented increases in antioxidant gene expression and NRF2 nuclear translocation. Exogenous hypochlorite administration resulted in increased NRF2 nuclear translocation in vitro. After Cl2 exposure, neutrophils occupied 22 ± 7% of the luminal space in large airways. IL-17 in BAL was increased after Cl2, although this effect was not prevented by neutrophil depletion. Neither depletion of eosinophils nor macrophages prevented Cl2-induced AHR. Our data suggest the ability of neutrophils to promote Cl2-induced AHR is dependent on increases in oxidative stress and occupation of luminal space in large airways.
氯气（Cl2）吸入会导致氧化应激、气道上皮损伤、气道高反应性（AHR）和嗜中性粒细胞增多。我们评估了嗜中性粒细胞耗竭对 Cl2 诱导的 AHR 的影响，及其对内源性抗氧化反应的影响，以及嗜酸性粒细胞或巨噬细胞是否会影响 Cl2 诱导的 AHR。我们将雄性 Balb/C 小鼠暴露于 100 ppm Cl2 中 5 分钟。我们通过定量 PCR 量化了支气管肺泡灌洗液（BAL）中的炎症细胞群，通过免疫荧光量化了肺组织中的抗氧化反应，以及核因子（红细胞衍生 2）样 2（NRF2）的核易位。在体外，使用荧光素酶测定法评估了对外源次氯酸盐的 NRF2 核易位反应。使用抗粒细胞受体-1 抗体或抗 Ly6G 抗体来耗竭嗜中性粒细胞。通过 ELISA 测量嗜中性粒细胞耗竭对 IL-13 和 IL-17 的影响。使用 TRFK5 或载有氯膦酸盐的脂质体分别耗竭嗜酸性粒细胞和巨噬细胞。通过对吸入的雾化乙酰甲胆碱的恒相模型评估 AHR。我们的结果表明，Cl2 暴露诱导了嗜中性粒细胞增多，并增加了 NRF2 mRNA、超氧化物歧化酶-1 和血红素加氧酶 1 的表达。嗜中性粒细胞耗竭消除了 Cl2 诱导的大呼吸道 AHR，并防止了抗氧化基因表达和 NRF2 核易位的增加。外源性次氯酸盐给药导致体外 NRF2 核易位增加。Cl2 暴露后，嗜中性粒细胞占据了大气道管腔的 22±7%。Cl2 后 BAL 中的 IL-17 增加，尽管嗜中性粒细胞耗竭并未阻止这种增加。耗竭嗜酸性粒细胞或巨噬细胞均不能防止 Cl2 诱导的 AHR。我们的数据表明，嗜中性粒细胞促进 Cl2 诱导的 AHR 的能力取决于氧化应激的增加和大气道管腔的占据。
