NOX4过度表达与中性粒细胞哮喘患者上皮纤毛功能障碍相关
2017/04/28
摘要
背景：哮喘的一个重要特征在于支气管上皮纤毛功能障碍。我们旨在探索中性粒细胞炎症和烟酰胺腺嘌呤二核苷酸磷酸（nicotinamide adenine dinucleotide phosphate，NADPH）氧化酶在哮喘患者纤毛功能障碍中的作用。
方法：通过视频显微镜对哮喘患者新鲜离体上皮细胞检测支气管上皮纤毛功能，受试者包括健康受试者与哮喘患者，根据痰细胞分类对哮喘进行分层。通过8-羟基脱氧鸟苷（8-oxo-dG）表达测定支气管上皮氧化损伤。支气管上皮细胞的NADPH氧化酶（NOX）/双氧化酶（DUOX）表达通过使用基因芯片进行评估，氧化酶4（NOX4）和双氧化酶1/2(DOUX1/2)表达情况通过支气管活检标本进行评估。对哮喘患者以及采用卵蛋白致敏和激发后的小鼠模型，获得其新鲜支气管上皮细胞，使用NADPH氧化酶抑制剂（GKT137831）后，评估纤毛功能障碍。
结果：与非中性粒细胞性哮喘患者（n=10）比较，中性粒细胞性哮喘患者(n=11)的纤毛摆动频率受损(5.8 [0.6] Hz vs 8.8 [0.5] Hz; P = .003)，且与痰中性粒细胞计数呈负相关(r = –0.70; P < .001)。原代支气管上皮细胞表达DUOX1/2和NOX4。与非中性粒细胞性哮喘患者比较，8-oxo-dG及NOX4在中性粒细胞性哮喘患者中升高，DUOX1在两种表型中均升高，，DUOX2仅在非中性粒细胞性哮喘中升高。虽然中性粒细胞性哮喘患者NOX4的表达与活性氧（ROS）的产生增加，但在培养的原代上皮细胞中，纤毛功能障碍并未持续存在。GKT137831使体外上皮细胞（n=13）的纤毛功能得到改善，且与中性粒细胞炎症强度相关；它亦消除了小鼠哮喘模型的纤毛功能障碍，且炎症无明显减轻。
结论：中性粒细胞性哮喘其纤毛功能障碍增高，它与NOX4表达增加相关，且能被NADPH氧化酶抑制剂减轻。
NADPH Oxidase-4 Overexpression Is Associated With Epithelial Ciliary Dysfunction in Neutrophilic Asthma
Wing-Yan Heidi Wan; Fay Hollins; Louise Haste; Lucy Woodman; Robert A. Hirst; Sarah Bolton; Edith Gomez; Amanda Sutcliffe; Dhananjay Desai; Latifa Chachi; Vijay Mistry; Cédric Szyndralewiez; Andrew Wardlaw; Ruth Saunders; Christopher O’Callaghan; Peter W. Andrew; and Christopher E. Brightling,
Chest; 2016; 149(6):1445-1459
Abstract
Background: Bronchial epithelial ciliary dysfunction is an important feature of asthma. We sought to determine the role in asthma of neutrophilic inflammation and nicotinamide adenine dinucleotide phosphate (NADPH) oxidases in ciliary dysfunction.
Methods: Bronchial epithelial ciliary function was assessed by using video microscopy in fresh ex vivo epithelial strips from patients with asthma stratified according to their sputum cell differentials and in culture specimens from healthy control subjects and patients with asthma. Bronchial epithelial oxidative damage was determined by 8-oxo-dG expression. Nicotinamide adenine dinucleotide phosphate oxidase (NOX)/dual oxidase (DUOX) expression was assessed in bronchial epithelial cells by using microarrays, with NOX4 and DUOX1/2 expression assessed in bronchial biopsy specimens. Ciliary dysfunction following NADPH oxidase inhibition, using GKT137831, was evaluated in fresh epithelial strips from patients with asthma and a murine model of ovalbumin sensitization and challenge.
Results: Ciliary beat frequency was impaired in patients with asthma with sputum neutrophilia (n =11) vs those without (n = 10) (5.8 [0.6] Hz vs 8.8 [0.5] Hz; P = .003) and was correlated with sputum neutrophil count (r = –0.70; P < .001). Primary bronchial epithelial cells expressed DUOX1/2 and NOX4. Levels of 8-oxo-dG and NOX4 were elevated in patients with neutrophilic vs nonneutrophilic asthma, DUOX1 was elevated in both, and DUOX2 was elevated in nonneutrophilic asthma in vivo. In primary epithelial cultures, ciliary dysfunction did not persist, although NOX4 expression and reactive oxygen species generation was increased from patients with neutrophilic asthma. GKT137831 both improved ciliary function in ex vivo epithelial strips (n = 13), relative to the intensity of neutrophilic inflammation, and abolished ciliary dysfunction in the murine asthma model with no reduction in inflammation.
Conclusions: Ciliary dysfunction is increased in neutrophilic asthma associated with increased NOX4 expression and is attenuated by NADPH oxidase inhibition.
背景：哮喘的一个重要特征在于支气管上皮纤毛功能障碍。我们旨在探索中性粒细胞炎症和烟酰胺腺嘌呤二核苷酸磷酸（nicotinamide adenine dinucleotide phosphate，NADPH）氧化酶在哮喘患者纤毛功能障碍中的作用。
方法：通过视频显微镜对哮喘患者新鲜离体上皮细胞检测支气管上皮纤毛功能，受试者包括健康受试者与哮喘患者，根据痰细胞分类对哮喘进行分层。通过8-羟基脱氧鸟苷（8-oxo-dG）表达测定支气管上皮氧化损伤。支气管上皮细胞的NADPH氧化酶（NOX）/双氧化酶（DUOX）表达通过使用基因芯片进行评估，氧化酶4（NOX4）和双氧化酶1/2(DOUX1/2)表达情况通过支气管活检标本进行评估。对哮喘患者以及采用卵蛋白致敏和激发后的小鼠模型，获得其新鲜支气管上皮细胞，使用NADPH氧化酶抑制剂（GKT137831）后，评估纤毛功能障碍。
结果：与非中性粒细胞性哮喘患者（n=10）比较，中性粒细胞性哮喘患者(n=11)的纤毛摆动频率受损(5.8 [0.6] Hz vs 8.8 [0.5] Hz; P = .003)，且与痰中性粒细胞计数呈负相关(r = –0.70; P < .001)。原代支气管上皮细胞表达DUOX1/2和NOX4。与非中性粒细胞性哮喘患者比较，8-oxo-dG及NOX4在中性粒细胞性哮喘患者中升高，DUOX1在两种表型中均升高，，DUOX2仅在非中性粒细胞性哮喘中升高。虽然中性粒细胞性哮喘患者NOX4的表达与活性氧（ROS）的产生增加，但在培养的原代上皮细胞中，纤毛功能障碍并未持续存在。GKT137831使体外上皮细胞（n=13）的纤毛功能得到改善，且与中性粒细胞炎症强度相关；它亦消除了小鼠哮喘模型的纤毛功能障碍，且炎症无明显减轻。
结论：中性粒细胞性哮喘其纤毛功能障碍增高，它与NOX4表达增加相关，且能被NADPH氧化酶抑制剂减轻。
(吕燕1 张红萍1 王刚 四川大学华西医院中西医结合科呼吸病组 610041 摘译)
（Chest; 2016; 149(6):1445-1459）
（Chest; 2016; 149(6):1445-1459）
NADPH Oxidase-4 Overexpression Is Associated With Epithelial Ciliary Dysfunction in Neutrophilic Asthma
Wing-Yan Heidi Wan; Fay Hollins; Louise Haste; Lucy Woodman; Robert A. Hirst; Sarah Bolton; Edith Gomez; Amanda Sutcliffe; Dhananjay Desai; Latifa Chachi; Vijay Mistry; Cédric Szyndralewiez; Andrew Wardlaw; Ruth Saunders; Christopher O’Callaghan; Peter W. Andrew; and Christopher E. Brightling,
Chest; 2016; 149(6):1445-1459
Abstract
Background: Bronchial epithelial ciliary dysfunction is an important feature of asthma. We sought to determine the role in asthma of neutrophilic inflammation and nicotinamide adenine dinucleotide phosphate (NADPH) oxidases in ciliary dysfunction.
Methods: Bronchial epithelial ciliary function was assessed by using video microscopy in fresh ex vivo epithelial strips from patients with asthma stratified according to their sputum cell differentials and in culture specimens from healthy control subjects and patients with asthma. Bronchial epithelial oxidative damage was determined by 8-oxo-dG expression. Nicotinamide adenine dinucleotide phosphate oxidase (NOX)/dual oxidase (DUOX) expression was assessed in bronchial epithelial cells by using microarrays, with NOX4 and DUOX1/2 expression assessed in bronchial biopsy specimens. Ciliary dysfunction following NADPH oxidase inhibition, using GKT137831, was evaluated in fresh epithelial strips from patients with asthma and a murine model of ovalbumin sensitization and challenge.
Results: Ciliary beat frequency was impaired in patients with asthma with sputum neutrophilia (n =11) vs those without (n = 10) (5.8 [0.6] Hz vs 8.8 [0.5] Hz; P = .003) and was correlated with sputum neutrophil count (r = –0.70; P < .001). Primary bronchial epithelial cells expressed DUOX1/2 and NOX4. Levels of 8-oxo-dG and NOX4 were elevated in patients with neutrophilic vs nonneutrophilic asthma, DUOX1 was elevated in both, and DUOX2 was elevated in nonneutrophilic asthma in vivo. In primary epithelial cultures, ciliary dysfunction did not persist, although NOX4 expression and reactive oxygen species generation was increased from patients with neutrophilic asthma. GKT137831 both improved ciliary function in ex vivo epithelial strips (n = 13), relative to the intensity of neutrophilic inflammation, and abolished ciliary dysfunction in the murine asthma model with no reduction in inflammation.
Conclusions: Ciliary dysfunction is increased in neutrophilic asthma associated with increased NOX4 expression and is attenuated by NADPH oxidase inhibition.
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