Supplementary Material for: Comparison of Exhaled Endogenous Particles from Smokers and Non-Smokers Using Multivariate Analysis

<b><i>Background:</i></b> Smoking, along with many respiratory diseases, has been shown to induce airway inflammation and alter the composition of the respiratory tract lining fluid (RTLF). We have previously shown that the phospholipid and protein composition of particles in exhaled air (PEx) reflects that of RTLF. In this study, we hypothesized that the composition of PEx differs between smokers and non-smokers, reflecting inflammation in the airways. <b><i>Objective:</i></b> It was the aim of this study to identify differences in the phospholipid composition of PEx from smokers and non-smokers. <b><i>Methods:</i></b> PEx from 12 smokers and 13 non-smokers was collected using a system developed in-house. PEx was analysed using time-of-flight secondary ion mass spectrometry, and the mass spectral data were evaluated using multivariate analysis. Orthogonal partial least squares (OPLS) was used to relate smoking status, lung function and pack years to the chemical composition of RTLF. The discriminating ions identified by OPLS were then used as explanatory variables in traditional regression analysis. <b><i>Results:</i></b> There was a clear discrimination between smokers and non-smokers according to the chemical composition, where phospholipids from smokers were protonated and sodiated to a larger extent. Poor lung function showed a strong association with higher response from all molecular phosphatidylcholine species in the samples. Furthermore, the accumulated amount of tobacco consumed was associated with variations in mass spectra, indicating a dose-response relationship. <b><i>Conclusion:</i></b> The chemical composition of PEx differs between smokers and non-smokers, reflecting differences in the RTLF. The results from this study may suggest that the composition of RTLF is affected by smoking and may be of importance for lung function.

<b><i>背景：</i></b> 吸烟与多种呼吸系统疾病一样，可诱发气道炎症并改变呼吸道衬液（respiratory tract lining fluid, RTLF）的组成。此前我们的研究已证实，呼出气微粒（exhaled particles, PEx）中的磷脂与蛋白质组成可反映呼吸道衬液的组成情况。本研究假设，吸烟者与非吸烟者的呼出气微粒组成存在差异，该差异可反映气道炎症状态。<b><i>目标：</i></b> 本研究旨在明确吸烟者与非吸烟者呼出气微粒的磷脂组成差异。<b><i>方法：</i></b> 本研究采用自研装置收集了12名吸烟者与13名非吸烟者的呼出气微粒样本。采用飞行时间二次离子质谱（time-of-flight secondary ion mass spectrometry）对样本进行分析，并通过多变量分析处理质谱数据。使用正交偏最小二乘（Orthogonal partial least squares, OPLS）模型将吸烟状态、肺功能及吸烟包年数与呼吸道衬液的化学组成进行关联分析。随后将OPLS模型识别出的差异离子作为解释变量，应用传统回归分析进行验证。<b><i>结果：</i></b> 基于化学组成可清晰区分吸烟者与非吸烟者：吸烟者样本中的磷脂更易发生质子化与钠结合。肺功能不佳与样本中所有磷脂酰胆碱分子的信号响应增强呈显著关联。此外，烟草累积摄入量与质谱信号的变化存在关联，提示存在剂量-反应关系。<b><i>结论：</i></b> 吸烟者与非吸烟者的呼出气微粒化学组成存在差异，该差异可反映呼吸道衬液的组成变化。本研究结果提示，吸烟可影响呼吸道衬液的组成，且该组成变化可能与肺功能状态密切相关。