Department of Clinical and Experimental Medicine - University of Catania, Catania, Italy.
Acclarogen Ltd, St John's Innovation Centre, Cambridge, United Kingdom.
PLoS One. 2018 Sep 21;13(9):e0203874. doi: 10.1371/journal.pone.0203874. eCollection 2018.
Oxidative stress is believed to be a major driver of inflammation in smoking asthmatics. The U-BIOPRED project recruited a cohort of Severe Asthma smokers/ex-smokers (SAs/ex) and non-smokers (SAn) with extensive clinical and biomarker information enabling characterization of these subjects. We investigated oxidative stress in severe asthma subjects by analysing urinary 8-iso-PGF2α and the mRNA-expression of the main pro-oxidant (NOX2; NOSs) and anti-oxidant (SODs; CAT; GPX1) enzymes in the airways of SAs/ex and SAn. All the severe asthma U-BIOPRED subjects were further divided into current smokers with severe asthma (CSA), ex-smokers with severe asthma (ESA) and non-smokers with severe asthma (NSA) to deepen the effect of active smoking. Clinical data, urine and sputum were obtained from severe asthma subjects. A bronchoscopy to obtain bronchial biopsy and brushing was performed in a subset of subjects. The main clinical data were analysed for each subset of subjects (urine-8-iso-PGF2α; IS-transcriptomics; BB-transcriptomics; BBr-transcriptomics). Urinary 8-iso-PGF2α was quantified using mass spectrometry. Sputum, bronchial biopsy and bronchial brushing were processed for mRNA expression microarray analysis. Urinary 8-iso-PGF2α was increased in SAs/ex, median (IQR) = 31.7 (24.5-44.7) ng/mmol creatinine, compared to SAn, median (IQR) = 26.6 (19.6-36.6) ng/mmol creatinine (p< 0.001), and in CSA, median (IQR) = 34.25 (24.4-47.7), vs. ESA, median (IQR) = 29.4 (22.3-40.5), and NSA, median (IQR) = 26.5 (19.6-16.6) ng/mmol creatinine (p = 0.004). Sputum mRNA expression of NOX2 was increased in SAs/ex compared to SAn (probe sets 203922_PM_s_at fold-change = 1.05 p = 0.006; 203923_PM_s_at fold-change = 1.06, p = 0.003; 233538_PM_s_at fold-change = 1.06, p = 0.014). The mRNA expression of antioxidant enzymes were similar between the two severe asthma cohorts in all airway samples. NOS2 mRNA expression was decreased in bronchial brushing of SAs/ex compared to SAn (fold-change = -1.10; p = 0.029). NOS2 mRNA expression in bronchial brushing correlated with FeNO (Kendal's Tau = 0.535; p< 0.001). From clinical and inflammatory analysis, FeNO was lower in CSA than in ESA in all the analysed subject subsets (p< 0.01) indicating an effect of active smoking. Results about FeNO suggest its clinical limitation, as inflammation biomarker, in severe asthma active smokers. These data provide evidence of greater systemic oxidative stress in severe asthma smokers as reflected by a significant changes of NOX2 mRNA expression in the airways, together with elevated urinary 8-iso-PGF2α in the smokers/ex-smokers group. Trial registration ClinicalTrials.gov-Identifier: NCT01976767.
氧化应激被认为是吸烟哮喘患者炎症的主要驱动因素。U-BIOPRED 项目招募了一组严重哮喘吸烟者/戒烟者（SAs/ex）和非吸烟者（SAn），他们具有广泛的临床和生物标志物信息，能够对这些受试者进行特征描述。我们通过分析严重哮喘患者的尿 8-异前列腺素 F2α和主要促氧化剂（NOX2；NOSs）和抗氧化剂（SODs；CAT；GPX1）在 SAs/ex 和 SAn 气道中的 mRNA 表达来研究严重哮喘患者的氧化应激。所有 U-BIOPRED 严重哮喘受试者都进一步分为当前吸烟者（CSA）、前吸烟者（ESA）和非吸烟者（NSA），以加深主动吸烟的影响。从严重哮喘患者中获得临床数据、尿液和痰液。在亚组患者中进行支气管镜检查以获得支气管活检和刷检。主要临床数据用于分析每个亚组的受试者（尿 8-iso-PGF2α；IS 转录组学；BB 转录组学；BBr 转录组学）。使用质谱法定量尿液 8-iso-PGF2α。处理痰液、支气管活检和支气管刷检，进行 mRNA 表达微阵列分析。与 SAn 相比，SAs/ex 的尿 8-iso-PGF2α 增加，中位数（IQR）=31.7（24.5-44.7）ng/mmol 肌酐，p<0.001，CSA 中位数（IQR）=34.25（24.4-47.7），与 ESA 相比，中位数（IQR）=29.4（22.3-40.5），与 NSA 相比，中位数（IQR）=26.5（19.6-16.6）ng/mmol 肌酐（p=0.004）。与 SAn 相比，SAs/ex 的痰中 NOX2 的 mRNA 表达增加（探针集 203922_PM_s_at 倍数变化=1.05，p=0.006；203923_PM_s_at 倍数变化=1.06，p=0.003；233538_PM_s_at 倍数变化=1.06，p=0.014）。两组严重哮喘患者在所有气道样本中的抗氧化酶 mRNA 表达相似。与 SAn 相比，SAs/ex 支气管刷检中的 NOS2 mRNA 表达降低（倍数变化=-1.10；p=0.029）。与 SAn 相比，NOS2 支气管刷检中的 mRNA 表达与 FeNO 相关（Kendal's Tau=0.535；p<0.001）。从临床和炎症分析来看，在所有分析的亚组受试者中，CSA 的 FeNO 低于 ESA（p<0.01），表明主动吸烟的影响。关于 FeNO 的结果表明其在严重哮喘活动吸烟者中的临床局限性，作为炎症生物标志物。这些数据提供了严重哮喘吸烟者中系统氧化应激更大的证据，这反映在气道中 NOX2 mRNA 表达的显著变化以及吸烟者/戒烟者组中尿液 8-iso-PGF2α 的升高。临床试验注册ClinicalTrials.gov-标识符：NCT01976767。
