{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"indexed":{"date-parts":[[2025,6,17]],"date-time":"2025-06-17T21:21:54Z","timestamp":1750195314655,"version":"3.37.3"},"reference-count":75,"publisher":"Public Library of Science (PLoS)","issue":"12","license":[{"start":{"date-parts":[[2024,12,31]],"date-time":"2024-12-31T00:00:00Z","timestamp":1735603200000},"content-version":"vor","delay-in-days":0,"URL":"http:\/\/creativecommons.org\/licenses\/by\/4.0\/"}],"funder":[{"DOI":"10.13039\/100000066","name":"National Institute of Environmental Health Sciences","doi-asserted-by":"publisher","award":["T32 ES007126"],"award-info":[{"award-number":["T32 ES007126"]}],"id":[{"id":"10.13039\/100000066","id-type":"DOI","asserted-by":"publisher"}]},{"DOI":"10.13039\/100000066","name":"National Institute of Environmental Health Sciences","doi-asserted-by":"publisher","award":["T32 ES007126"],"award-info":[{"award-number":["T32 ES007126"]}],"id":[{"id":"10.13039\/100000066","id-type":"DOI","asserted-by":"publisher"}]},{"DOI":"10.13039\/100000066","name":"National Institute of Environmental Health Sciences","doi-asserted-by":"publisher","award":["R01 ES031173"],"award-info":[{"award-number":["R01 ES031173"]}],"id":[{"id":"10.13039\/100000066","id-type":"DOI","asserted-by":"publisher"}]},{"DOI":"10.13039\/100000066","name":"National Institute of Environmental Health Sciences","doi-asserted-by":"publisher","award":["R01 ES031173"],"award-info":[{"award-number":["R01 ES031173"]}],"id":[{"id":"10.13039\/100000066","id-type":"DOI","asserted-by":"publisher"}]},{"DOI":"10.13039\/100000066","name":"National Institute of Environmental Health Sciences","doi-asserted-by":"publisher","award":["R01 ES031173"],"award-info":[{"award-number":["R01 ES031173"]}],"id":[{"id":"10.13039\/100000066","id-type":"DOI","asserted-by":"publisher"}]}],"content-domain":{"domain":["www.plosone.org"],"crossmark-restriction":false},"short-container-title":["PLoS ONE"],"abstract":"<jats:p>Imbalance of airway proteases and antiproteases has been implicated in diseases such as COPD and environmental exposures including cigarette smoke and ozone. To initiate infection, endogenous proteases are commandeered by respiratory viruses upon encountering the airway epithelium. The airway proteolytic environment likely contains redundant antiproteases and proteases with diverse catalytic mechanisms, however a proteomic profile of these enzymes and inhibitors in airway samples has not been reported. The objective of this study was to first profile extracellular proteases and antiproteases using human airway epithelial cell cultures and <jats:italic>ex vivo<\/jats:italic> nasal epithelial lining fluid (NELF) samples. Secondly, we present an optimized method for probing the proteolytic environment of airway surface liquid samples (<jats:italic>in vitro<\/jats:italic> and <jats:italic>ex vivo<\/jats:italic>) using fluorogenic peptides modeling the cleavage sites of respiratory viruses. We detected 48 proteases in the apical wash of cultured human nasal epithelial cells (HNECs) (n = 6) and 57 in NELF (n = 13) samples from healthy human subjects using mass-spectrometry based proteomics. Additionally, we detected 29 and 48 antiproteases in the HNEC apical washes and NELF, respectively. We observed large interindividual variability in rate of cleavage of an Influenza H1 peptide in the <jats:italic>ex vivo<\/jats:italic> clinical samples. Since protease and antiprotease levels have been found to be altered in the airways of smokers, we compared proteolytic cleavage in <jats:italic>ex vivo<\/jats:italic> nasal lavage samples from male\/female smokers and non-smokers. There was a statistically significant increase in proteolysis of Influenza H1 in NLF from male smokers compared to female smokers. Furthermore, we measured cleavage of the S1\/S2 site of SARS-CoV, SARS-CoV-2, and SARS-CoV-2 Delta peptides in various airway samples, suggesting the method could be used for other viruses of public health relevance. This assay presents a direct and efficient method of evaluating the proteolytic environment of human airway samples in assessment of therapeutic treatment, exposure, or underlying disease.<\/jats:p>","DOI":"10.1371\/journal.pone.0306197","type":"journal-article","created":{"date-parts":[[2024,12,31]],"date-time":"2024-12-31T18:30:42Z","timestamp":1735669842000},"page":"e0306197","update-policy":"https:\/\/doi.org\/10.1371\/journal.pone.corrections_policy","source":"Crossref","is-referenced-by-count":1,"title":["Profiling endogenous airway proteases and antiproteases and modeling proteolytic activation of Influenza HA using in vitro and ex vivo human airway surface liquid samples"],"prefix":"10.1371","volume":"19","author":[{"given":"Stephanie A.","family":"Brocke","sequence":"first","affiliation":[],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Boris","family":"Reidel","sequence":"additional","affiliation":[],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Camille","family":"Ehre","sequence":"additional","affiliation":[],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Meghan E.","family":"Rebuli","sequence":"additional","affiliation":[],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Carole","family":"Robinette","sequence":"additional","affiliation":[],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Kevin D.","family":"Schichlein","sequence":"additional","affiliation":[],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Christian A.","family":"Brooks","sequence":"additional","affiliation":[],"role":[{"role":"author","vocabulary":"crossref"}]},{"ORCID":"https:\/\/orcid.org\/0000-0001-8728-0305","authenticated-orcid":true,"given":"Ilona","family":"Jaspers","sequence":"additional","affiliation":[],"role":[{"role":"author","vocabulary":"crossref"}]}],"member":"340","published-online":{"date-parts":[[2024,12,31]]},"reference":[{"issue":"D1","key":"pone.0306197.ref001","doi-asserted-by":"crossref","first-page":"D624","DOI":"10.1093\/nar\/gkx1134","article-title":"The MEROPS database of proteolytic enzymes, their substrates and inhibitors in 2017 and a comparison with peptidases in the PANTHER database","volume":"46","author":"ND Rawlings","year":"2017","journal-title":"Nucleic Acids Research"},{"issue":"6","key":"pone.0306197.ref002","doi-asserted-by":"crossref","first-page":"294","DOI":"10.1016\/j.tips.2005.04.006","article-title":"Cutting back on pro-protein convertases: the latest approaches to pharmacological inhibition","volume":"26","author":"M Fug\u00e8re","year":"2005","journal-title":"Trends Pharmacol Sci"},{"key":"pone.0306197.ref003","doi-asserted-by":"crossref","first-page":"650","DOI":"10.1016\/B978-0-12-394447-4.10076-8","article-title":"Extracellular: Plasma Membrane Proteases\u2014Serine Proteases.","author":"T.M. 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