title: S-glutathionylation of buccal cell proteins as biomarkers of exposure to hydrogen peroxide.
abstract: BACKGROUND
  Exogenous or endogenous hydrogen peroxide (H2O2) is a reactive oxygen species (ROS) that can lead to oxidation of cellular nucleophiles, particularly cysteines in proteins. Commercial mouthwashes containing H2O2 provide the opportunity to determine clinically whether changes in S-glutathionylation of susceptible proteins in buccal mucosa cells can be used as biomarkers of ROS exposure.
  
  
  METHODS
  Using an exploratory clinical protocol, 18 disease-free volunteers rinsed with a mouthwash containing 1.5% H2O2 (442 mM) over four consecutive days. Exfoliated buccal cell samples were collected prior and post-treatment and proteomics were used to identify S-glutathionylated proteins.
  
  
  RESULTS
  Four consecutive daily treatments with the H2O2-containing mouthwash induced significant dose and time-dependent increases in S-glutathionylation of buccal cell proteins, stable for at least 30 min following treatments. Elevated levels of S-glutathionylation were maintained with subsequent daily exposure. Increased S-glutathionylation preceded and correlated with transcriptional activation of ROS sensitive genes, such as ATF3, and with the presence of 8-hydroxy deoxyguanosine. Data from a human buccal cell line TR146 were consistent with the trial results. We identified twelve proteins that were S-glutathionylated following H2O2 exposure.
  
  
  CONCLUSIONS
  Buccal cells can predict exposure to ROS through increased levels of S-glutathionylation of proteins. These post-translationally modified proteins serve as biomarkers for the effects of H2O2 in the oral cavity and in the future, may be adaptable as extrapolated pharmacodynamic biomarkers for assessing the impact of other systemic drugs that cause ROS and/or impact redox homeostasis.
  
  
  CONCLUSIONS
  S-glutathionylation of buccal cell proteins can be used as a quantitative measure of exposure to ROS.
authors.affiliation: Department of Cell and Molecular Pharmacology and Experimental Therapeutics, Medical University of South Carolina, 70 President St., DD410, Charleston, SC 29425, USA.
authors.forename: Christina L
authors.initials: CL
authors.lastname: Grek
authors.affiliation: Department of Cell and Molecular Pharmacology and Experimental Therapeutics, Medical University of South Carolina, 70 President St., DD410, Charleston, SC 29425, USA.
authors.forename: Leticia
authors.initials: L
authors.lastname: Reyes
authors.affiliation: Department of Pharmaceutical and Biomedical Sciences, Medical University of SC, 274 Calhoun Street MSC 141, Charleston, SC 29425-1410, USA.
authors.forename: Danyelle M
authors.initials: DM
authors.lastname: Townsend
authors.affiliation: Department of Cell and Molecular Pharmacology and Experimental Therapeutics, Medical University of South Carolina, 70 President St., DD410, Charleston, SC 29425, USA.
authors.forename: Kenneth D
authors.initials: KD
authors.lastname: Tew
keywords: Buccal cells; Cysteine; Glutathione; Glutathione S-transferase; Hydrogen peroxide; Kinase signaling; Reactive oxygen species; S-glutathionylation