DNA SINGLE-STRAND BREAKS AND ADENINE-NUCLEOTIDE DEPLETION AS INDEXES OF OXIDANT EFFECTS ON HUMAN LUNG-CELLS

Citation
Trj. Ollikainen et al., DNA SINGLE-STRAND BREAKS AND ADENINE-NUCLEOTIDE DEPLETION AS INDEXES OF OXIDANT EFFECTS ON HUMAN LUNG-CELLS, Free radical biology & medicine, 24(7-8), 1998, pp. 1088-1096
Citations number
28
Categorie Soggetti
Endocrynology & Metabolism",Biology
ISSN journal
08915849
Volume
24
Issue
7-8
Year of publication
1998
Pages
1088 - 1096
Database
ISI
SICI code
0891-5849(1998)24:7-8<1088:DSBAAD>2.0.ZU;2-A
Abstract
The comet assay (single cell gel electrophoresis) is a novel method to assess DNA strand breaks in single cells. We studied the oxidant sens itivity of cultured primary and transformed (MeT-5A) human pleural mes othelial cells, as well as primary and transformed (BEAS 2B) human bro nchial epithelial cells, and compared the results obtained with the Co met assay to other markers of oxidant effects on cells, such as deplet ion of intracellular high-energy nucleotides (ATP, ADP, AMP), accumula tion of products of nucleotide catabolism (xanthine, hypoxanthine, uri c acid), and release of lactate dehydrogenase (LDH). The cells were ex posed for 5 min to 4 h to 50-500 mu M H2O2 or to 5-50 mu M menadione. Significant tail moment increase, which is a marker of DNA strand brea ks in the Comet assay, and intracellular nucleotide depletion occurred simultaneously in MeT-5A and BEAS 2B cells during the first 30-60 min of exposure to H2O2 and menadione, In the Comet assay variation betwe en the individual cells could be detected. LDH release, a marker of ce ll injury, showed that mesothelial cells were far more sensitive than epithelial cells to oxidant-induced lytic cell injury. MeT-5A and BEAS 2B cells contained similar intracellular antioxidant enzyme activitie s, which may explain their similar oxidant sensitivity in the Comet as say. A significant increase (164%) in the tail moment was delectable i n MeT-5A cells exposed to 50 mu M H2O2 for 30 min. This returned to co ntrol level during the 4 h of continuing exposure. A 30 min exposure t o 25 mu M menadione caused a 61% increase in the mean toil moment but, unlike with H2O2, the change was irreversible during the following 4 h incubation. We conclude that human pleural mesothelial cells and bro nchial epithelial cells show similar oxidant sensitivity when assessed by the Comet assay, but various oxidants differ in their potency in c ausing DNA breaks in these cells. (C) 1998 Elsevier Science Inc.