title: An improved method for the isolation of rat alveolar type II lung cells: Use in the Comet assay to determine DNA damage induced by cigarette smoke.
abstract: Smoking is a cause of serious diseases, including lung cancer, emphysema, chronic bronchitis and heart disease. DNA damage is thought to be one of the mechanisms by which cigarette smoke (CS) initiates disease in the lung. Indeed, CS induced DNA damage can be measured in vitro and in vivo. The potential of the Comet assay to measure DNA damage in isolated rat lung alveolar type II epithelial cells (AEC II) was explored as a means to include a genotoxicity end-point in rodent sub-chronic inhalation studies. In this study, published AEC II isolation methods were improved to yield viable cells suitable for use in the Comet assay. The improved method reduced the level of basal DNA damage and DNA repair in isolated AEC II. CS induced DNA damage could also be quantified in isolated cells following a single or 5 days CS exposure. In conclusion, the Comet assay has the potential to determine CS or other aerosol induced DNA damage in AEC II isolated from rodents used in sub-chronic inhalation studies.
authors.affiliation: British American Tobacco, Group R&D, Southampton, Hampshire SO15 8TL, UK. Electronic address: Annette_Dalrymple@bat.com.
authors.forename: Annette
authors.initials: A
authors.lastname: Dalrymple
authors.affiliation: Vivotecnia Research S.L., Parque Científico de Madrid, C/Santiago Grisolía, 2 (PTM), 28760 Tres Cantos, Madrid, Spain.
authors.forename: Patricia
authors.initials: P
authors.lastname: Ordoñez
authors.affiliation: British American Tobacco, Group R&D, Southampton, Hampshire SO15 8TL, UK.
authors.forename: David
authors.initials: D
authors.lastname: Thorne
authors.affiliation: British American Tobacco, Group R&D, Southampton, Hampshire SO15 8TL, UK.
authors.forename: Debbie
authors.initials: D
authors.lastname: Dillon
authors.affiliation: British American Tobacco, Group R&D, Southampton, Hampshire SO15 8TL, UK.
authors.forename: Clive
authors.initials: C
authors.lastname: Meredith
mesh_terms: D000818:Animals; D002469:Cell Separation / Q000379:methods*; D020552:Comet Assay / Q000379:methods; D004249:DNA Damage / Q000187:drug effects*; D004260:DNA Repair / Q000187:drug effects; D005260:Female; D008168:Lung / Q000166:cytology / Q000187:drug effects; D011650:Pulmonary Alveoli / Q000166:cytology* / Q000187:drug effects*; D051381:Rats; D017207:Rats, Sprague-Dawley; D012906:Smoke / Q000009:adverse effects*; D012907:Smoking / Q000009:adverse effects; D062789:Tobacco Products / Q000009:adverse effects*
chemical_list: D012906:Smoke
keywords: 3R4F reference cigarette; Alveolar type II cells (AEC II); Cigarette smoke; Comet assay; DNA damage; Genotoxicity endpoint; Rat