Identification of oxidized phospholipids in bronchoalveolar lavage exposed to low ozone levels using multivariate analysis

Abstract

Chemical reactions with unsaturated phospholipids in the respiratory tract lining fluid have been identified as one of the first important steps in the mechanisms mediating environmental ozone toxicity. As a consequence of these reactions, complex mixtures of oxidized lipids are generated in the presence of mixtures of non-oxidized naturally occurring phospholipid molecular species, which challenge methods of analysis. Untargeted mass spectrometry and statistical methods were employed to approach these complex spectra. Human bronchoalveolar lavage (BAL) was exposed to low levels of ozone and samples, with and without derivatization of aldehydes, were analyzed by liquid chromatography electrospray ionization tandem mass spectrometry. Data processing was carried out using principal component analysis (PCA). Resulting PCA score plots indicated an ozone dose-dependent increase, with apparent separation between BAL samples exposed to 60 ppb ozone and non-exposed BAL samples, and a clear separation between ozonized samples before and after derivatization. Corresponding loadings plots revealed that more than 30 phosphatidylcholine (PC) species decreased due to ozonation. A total of 13 PC and 6 phosphatidylglycerol oxidation products were identified with the majority being structurally characterized as chain-shortened aldehyde products. This method exemplifies an approach for comprehensive detection of low abundance, yet important, components in complex lipid samples.

Ozone-derived Oxysterols Affect Liver X Receptor (LXR) Signaling

“…Although formation of oxysterols following exposure to O 3 has been shown before (28,35), whether and how these O 3 -derived oxysterols can form oxysterol-protein adducts in the context of O 3 exposure in the human airway has not been examined. Assays based on "click" cyclo-addition methods make it possible to visualize protein-lipid adducts via immunostaining and to determine protein adductions with specific signaling molecules, thus providing novel insight into the mechanisms behind O 3 -induced adverse health effects (39).…”

“…Rodent exposure studies and in vitro studies using human pulmonary surfactant have demonstrated the formation of O 3 -derived oxidized cholesterol products in various chemical compositions (5,6,10,27,28). Fig.…”

Surfactant Lipids at the Host–Environment Interface. Metabolic Sensors, Suppressors, and Effectors of Inflammatory Lung Disease

“…Several pro-oxidant exposures have now been shown to drive the formation of oxPLs in pulmonary surfactant (Figure 2). In vitro exposure of human BALF to ozone directly induces multiple PC and PG oxidation products (4). In vivo exposure of mice to a wide variety of airway challenges, including cigarette smoke, particulate matter 2.5, acid, H1N1 influenza A virus, and H5N1 avian influenza virus also increases oxPAPC species and 1-palmitoyl-2-(9'-oxo-nonanoyl)glycerophosphocholine (PON-GPC) in airspace fluid (73)(74)(75)(76)(77).…”

Oxidative Stress Mechanisms in the Pathogenesis of Environmental Lung Diseases

“…A large body of evidence has reported elevated levels of MDA, 4HNE or Ox-PLs or their corresponding protein adducts in the bronchoalveolar lavage fluid, lung tissue and/or serum of patients with various lung diseases such as asthma, COPD, ARDS and IPF [43,[64][65][66]. Immunohistochemical analysis revealed greater accumulation of 4HNE in the airways, alveolar epithelium and inflammatory cells of the lungs of COPD patient when compared to smoker non-COPD patient with similar smoking history [67].…”