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Assessment of protein modifications in liver of rats under chronic treatment with paracetamol (acetaminophen) using two complementary mass spectrometry-based metabolomic approaches
- Mast, Carole, Lyan, Bernard, Joly, Charlotte, Centeno, Delphine, Giacomoni, Franck, Martin, Jean-François, Mosoni, Laurent, Dardevet, Dominique, Pujos-Guillot, Estelle, Papet, Isabelle
- Journal of proteomics 2015 v.120 pp. 194-203
- acetaminophen, cysteine, detection limit, diet, glutathione, hepatotoxicity, hydrolysates, hydrolysis, liver, liver protein, mass spectrometry, metabolomics, oxidation, proline, protein hydrolysates, protein synthesis, proteolysis, rats, thiols, tryptophan, tyrosine
- Liver protein can be altered under paracetamol (APAP) treatment. APAP-protein adducts and other protein modifications (oxidation/nitration, expression) play a role in hepatotoxicity induced by acute overdoses, but it is unknown whether liver protein modifications occur during long-term treatment with non-toxic doses of APAP. We quantified APAP-protein adducts and assessed other protein modifications in the liver from rats under chronic (17 days) treatment with two APAP doses (0.5% or 1% of APAP in the diet w/w). A targeted metabolomic method was validated and used to quantify APAP-protein adducts as APAP-cysteine adducts following proteolytic hydrolysis. The limit of detection was found to be 7ng APAP–cysteine/mL hydrolysate i.e. an APAP–Cys to tyrosine ratio of 0.016‰. Other protein modifications were assessed on the same protein hydrolysate by untargeted metabolomics including a new strategy to process the data and identify discriminant molecules. These two complementary mass spectrometry (MS)-based metabolic approaches enabled the assessment of a wide range of protein modifications induced by chronic treatment with APAP.APAP-protein adducts were detected even in the absence of glutathione depletion and hepatotoxicity, i.e. in the 0.5% APAP group, and increased by 218% in the 1% APAP group compared to the 0.5% APAP group. At the same time, the untargeted metabolomic method revealed a decrease in the binding of cysteine, cysteinyl–glycine and GSH to thiol groups of protein cysteine residues, an increase in the oxidation of tryptophan and proline residues and a modification in protein expression. This wide range of modifications in liver proteins occurred in rats under chronic treatment with APAP that did not induce hepatotoxicity.