Quantitative gas chromatographic-mass spectrometric assay of 4-hydroxynonenal bound to thiol proteins in ischemic/reperfused rat hearts.
Increasing evidence indicates that protein-aldehyde adducts involving mostly 4-hydroxynonenal could be causally involved in both pathophysiological and adaptive events following an oxidative stress insult such as ischemia/reperfusion. The goal of this study was to assess if isotope dilution chromatography-mass spectrometry can be used to quantitate changes in the cardiac levels of 4-hydroxynonenal and 1,4-dihydroxynonene, one of its major metabolites, bound to thiol proteins during ischemia/reperfusion. For this purpose, we modified a previously published method to include treatment with Raney Nickel, which specifically cleaves thioether linkages. Our study model was the isolated Langendorff-perfused rat heart subjected to various ischemia/reperfusion protocols. Hearts perfused under normoxia contained small amounts of protein-bound 4-hydroxynonenal and 1,4-dihydroxynonene (1.38 +/- 0.29 and 2.69 +/- 0.17 nmol/g wet weight, respectively). The accumulation of these adducts after global ischemia depended on the severity of the ischemic insult up to a plateau and was not exacerbated by reperfusion. In conclusion, our method allows the quantification of time-dependent changes in 4-hydroxynonenal and 1,4-dihydroxynonene bound to proteins via thioether linkage in ischemic/reperfused heart tissues. The presence of protein-bound 1,4-dihydroxynonene in heart tissues suggests that this organ can detoxify protein-bound 4-hydroxynonenal.
Véronneau
Marc
M
Department of Nutrition, Université de Montréal, Montréal, Québec, Canada.
Comte
Blandine
B
Des Rosiers
Christine
C
D000447:Aldehydes / Q000032:analysis*; D000818:Animals; D005609:Free Radicals; D008401:Gas Chromatography-Mass Spectrometry; D006321:Heart / Q000502:physiology*; D008297:Male; D009206:Myocardium / Q000378:metabolism*; D010100:Oxygen / Q000378:metabolism; D051381:Rats; D017207:Rats, Sprague-Dawley; D015427:Reperfusion Injury; D013438:Sulfhydryl Compounds / Q000032:analysis*; D013440:Sulfides; D013696:Temperature; D013997:Time Factors
D000447:Aldehydes; D005609:Free Radicals; D013438:Sulfhydryl Compounds; D013440:Sulfides; C027576:4-hydroxy-2-nonenal; D010100:Oxygen