8-hydroxy-2--deoxyguanosine and Cystic-Fibrosis

The DNA lesion 8-oxo-7,8-dihydro-2'-deoxyguanosine (8-oxodG), a urinary marker of oxidative stress, is produced from reactions of reactive oxygen species with host DNA 2'-deoxyribonucleotides. The current gold-standard assessment is by complex chromatographic methods using HPLC or LC-MS/MS. Several studies have reported that commercial 8-oxodG ELISA kits correlate sufficiently with chromatographic techniques to be an easier alternative for laboratories without access to gold-standard techniques. However, the assumption that significant correlation translates into a similar ability to differentiate disease categories or treatment groups is yet to be tested. Using LC-MS/MS and two variants of a commercial ELISA, we measured urinary 8-oxodG and creatinine concentrations in young children with cystic fibrosis, a disease associated with oxidative stress, and age-matched controls. We show that, despite significant correlation, both ELISAs overestimate the levels of 8-oxodG, and neither ELISA accurately depicted the difference in group means that was observed by gold-standard LC-MS/MS. The implications of these findings for study outcomes add further support for chromatographic techniques, despite their cost and complexity, to remain the gold standard in urinary 8-oxodG assessment.

Topics: 8-Hydroxy-2'-Deoxyguanosine; Biomarkers; Child; Child, Preschool; Chromatography, Liquid; Cystic Fibrosis; Deoxyguanosine; DNA Damage; Enzyme-Linked Immunosorbent Assay; False Positive Reactions; Female; Humans; Infant; Male; Oxidative Stress; Reproducibility of Results; Tandem Mass Spectrometry

Oxidative stress caused by chronic lung inflammation in patients with cystic fibrosis (CF) and chronic lung infection with Pseudomonas aeruginosa is characterized by the reactive oxygen species (ROS) liberated by polymorphonuclear leukocytes (PMNs). We formulated the hypothesis that oxidation of the bacterial DNA by ROS presents an increased risk for the occurrence of hypermutable P. aeruginosa. The occurrence of hypermutable P. aeruginosa isolates was investigated directly in the sputum of 79 CF patients and among 141 isolates collected from 11 CF patients (10 to 15 isolates/patient) collected from the 1st and up to the 25th year of their chronic lung infection. The level of oxidized guanine moiety 8-oxo-2'-deoxyguanosine (8-oxodG), which is a frequently investigated DNA oxidative lesion, was measured. Hypermutable P. aeruginosa isolates were found in the sputum bacterial population of 54.4% of the CF patients. The earliest mutator P. aeruginosa isolates were found after 5 years from the onset of the chronic lung infection, and once they were present in the CF lung, the prevalence increased with time. The hypermutable isolates were significantly more resistant to antipseudomonal antibiotics than nonhypermutable isolates (P

Topics: 8-Hydroxy-2'-Deoxyguanosine; Adult; Anti-Bacterial Agents; Chronic Disease; Cross-Sectional Studies; Cystic Fibrosis; Deoxyguanosine; Drug Resistance, Bacterial; Humans; Inflammation; Lung Diseases; Mutation; Oxidative Stress; Pseudomonas aeruginosa; Pseudomonas Infections; Sputum