title: Free and protein-associated nitrotyrosine formation following rat liver preservation and transplantation.
abstract: Nitrotyrosine in human and animal tissues has been associated with pathologic conditions such as atherosclerosis, renal failure, and acute lung disease. In this study, free and protein-associated nitrotyrosine were determined in plasma and tissue samples using a dual-channel electrochemical detection method. Free nitrotyrosine was quantified in acetonitrile-extracted samples while protein-associated nitrotyrosine was determined in proteinase K-digested samples. In human plasma, total nitrotyrosine increased from 2.3 to 4.3 and 13.2 mumol/mol Tyr following addition of 0, 0.5, and 1 mM ONOO-. To determine if nitrotyrosine was produced during ex vivo hypothermic preservation, rat livers were stored in University of Wisconsin solution (UW) for 0, 6, or 8 h and reperfused for 3 h. Total nitro-tyrosine increased 359 and 908% after 6 and 8 h preservation compared to 0 h. To determine if nitrotyrosine was produced in vivo following hepatic ischemia, a rat preservation-transplantation model was utilized in which livers were flushed with cold UW (0-h group) or transplanted following 6 h hypothermic preservation in UW. Free nitrotyrosine increased from 15.7 +/- 0.3 in the 0-h group to 23.6 +/- 2.5 mumol/mol Tyr, 24 h posttransplant of 6-h preserved livers. Protein-associated nitrotyrosine increased from 9.5 +/- 1.1 in the 0-h group to 27.5 +/- 0.7 mumol/mol Tyr in the 6-h preservation-transplantation group. Protein-associated nitrotyrosine provides an integrative determination of nitration. Detection of free and protein-associated nitrotyrosine in biologic samples may allow insight into the role of .NO-derived oxidants in tissue injury associated with various pathologic conditions.
authors.affiliation: Department of Anesthesiology, University of Alabama at Birmingham 35233, USA.
authors.forename: K A
authors.initials: KA
authors.lastname: Skinner
authors.forename: J P
authors.initials: JP
authors.lastname: Crow
authors.forename: H B
authors.initials: HB
authors.lastname: Skinner
authors.forename: R T
authors.initials: RT
authors.lastname: Chandler
authors.forename: J A
authors.initials: JA
authors.lastname: Thompson
authors.forename: D A
authors.initials: DA
authors.lastname: Parks
mesh_terms: D000818:Animals; D002851:Chromatography, High Pressure Liquid; D004563:Electrochemistry; D005609:Free Radicals / Q000378:metabolism / Q000494:pharmacology; D006801:Humans; D008099:Liver / Q000378:metabolism; D016031:Liver Transplantation; D009566:Nitrates / Q000378:metabolism; D009926:Organ Preservation; D011485:Protein Binding; D051381:Rats; D017207:Rats, Sprague-Dawley; D013997:Time Factors; D014443:Tyrosine / Q000031:analogs & derivatives* / Q000097:blood / Q000378:metabolism
chemical_list: D005609:Free Radicals; D009566:Nitrates; C039642:peroxynitric acid; C002744:3-nitrotyrosine; D014443:Tyrosine