Substrate-dependent metabolomic signatures of myeloperoxidase activity in airway epithelial cells: Implications for early cystic fibrosis lung disease

Abstract

Myeloperoxidase (MPO) is released by neutrophils in inflamed tissues. MPO oxidizes chloride, bromide, and thiocyanate to produce hypochlorous acid (HOCl), hypobromous acid (HOBr), and hypothiocyanous acid (HOSCN), respectively. These oxidants are toxic to pathogens, but may also react with host cells to elicit biological activity and potential toxicity. In cystic fibrosis (CF) and related diseases, increased neutrophil inflammation leads to increased airway MPO and airway epithelial cell (AEC) exposure to its oxidants. In this study, we investigated how equal dose-rate exposures of MPO-derived oxidants differentially impact the metabolome of human AECs (BEAS-2B cells). We utilized enzymatic oxidant production with rate-limiting glucose oxidase (GOX) coupled to MPO, and chloride, bromide (Br^-), or thiocyanate (SCN^-) as substrates. AECs exposed to GOX/MPO/SCN^- (favoring HOSCN) were viable after 24 h, while exposure to GOX/MPO (favoring HOCl) or GOX/MPO/Br^- (favoring HOBr) developed cytotoxicity after 6 h. Cell glutathione and peroxiredoxin-3 oxidation were insufficient to explain these differences. However, untargeted metabolomics revealed GOX/MPO and GOX/MPO/Br^- diverged significantly from GOX/MPO/SCN^- for dozens of metabolites. We noted methionine sulfoxide and dehydromethionine were significantly increased in GOX/MPO- or GOX/MPO/Br^--treated cells, and analyzed them as potential biomarkers of lung damage in bronchoalveolar lavage fluid from 5-year-olds with CF (n = 27). Both metabolites were associated with increasing bronchiectasis, neutrophils, and MPO activity. This suggests MPO production of HOCl and/or HOBr may contribute to inflammatory lung damage in early CF. In summary, our in vitro model enabled unbiased identification of exposure-specific metabolite products which may serve as biomarkers of lung damage in vivo. Continued research with this exposure model may yield additional oxidant-specific biomarkers and reveal explicit mechanisms of oxidant byproduct formation and cellular redox signaling.

Keywords: Cystic fibrosis; Dehydromethionine; Hypothiocyanite; Metabolomics; Myeloperoxidase; Oxidative stress.