Abstract:
:Oxidative stress in biological systems can result in radical-induced lipid peroxidation (LPO), which can lead to the production of secondary reactive by-products such as 4-hydroxy-2-nonenal (HNE), malondialdehyde (MDA), acrolein, and acetaldehyde. These deleterious compounds are known to react with and concomitantly modify nucleophilic amino acid residues on proteins. Oxidative stress induced by cigarette smoke (CS) has been put forth as a major mechanism for tobacco-induced pathologies. At present, there are few reliable biomarkers for measuring the extent of oxidatively-induced damage resulting from CS exposure in vivo. This study has utilized a previously reported CS exposure system to expose cultured cells in-vitro to whole CS and determine the extent of LPO resulting from CS exposure by quantifying the increase in HNE within the exposure media versus controls via gas chromatography mass spectrometry. Additionally, we obtained protein enriched cell lysate post-CS exposures and measured the fluorescent signal obtained via direct injection fluorescent analysis at 375 nm ex./415 nm em. This study determined that the fluorescent signal intensity was directly proportional to the quantity increase of HNE in CS exposed media. It further tested this correlation by performing HNE titration addition experiments to cultured cells and Western blot analysis on proteins obtained from cell lysates. Finally, the fluorescent signal increase from authentic BSA solutions incubated with increasing concentrations of HNE was measured. It is proposed that the fluorescent signal observed from the protein lysate of CS exposed cultured cells corresponds to the extent of biological damage resulting from secondary reactive by-products formed from LPO induced via CS exposure and represented by HNE. The fluorescent signals increased in intensity upon increasing CS dose up to 20 min and remained elevated over 24 h after cessation of CS exposure.
journal_name
Toxicol Mech Methodsjournal_title
Toxicology mechanisms and methodsauthors
Rainey RP,Gillman IG,Shi X,Cheng T,Stinson A,Gietl D,Albino APdoi
10.1080/15376510903104224subject
Has Abstractpub_date
2009-09-01 00:00:00pages
401-9issue
6-7eissn
1537-6516issn
1537-6524journal_volume
19pub_type
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