Oxidative stress has been implicated in numerous degenerative diseases of aging, including
heart diseases. However, there is still a need to identify
biomarkers of oxidative stress-related events, such as
protein modification by the lipid peroxidation product
4-hydroxynonenal (HNE) in these diseases in humans. The objective of this study was to assess if circulating levels of HNE-
protein adducts (i) can be assessed with precision by GCMS and (ii) vary with
disease progression and aging in a model of
cardiomyopathy that displays enhanced oxidative stress, namely the spontaneously hypertensive rats (SHR). We modified a previously published
isotope dilution GCMS method that quantifies HNE and its inactive metabolite, 1,4-dihydroxynonene (DHN), bound to
thiol proteins following treatment with NaB(2)H(4) and Raney
nickel, to increase its sensitivity (20-fold), precision, and robustness. Levels of these adducts were measured in blood and plasma collected from SHR and control Wistar rats at 7, 15, 22, and 30 weeks of age. Levels of
protein-bound HNE, which were quantitated with good precision in the nanomolar range in blood, but not in plasma, were significantly increased by disease (SHR) and age (P < 0.0001 for both). Compared to Wistar rats, SHR showed greater blood levels of HNE-
protein adducts at 22 and 30 weeks. Levels of
protein-bound DHN, which were detected in blood and in plasma, were not affected by disease or age. Collectively, the results of this study conducted in an animal model of
cardiomyopathy demonstrate that changes in blood HNE-
protein thioether adducts with
disease progression and aging can be assessed with good precision by the described GCMS method. This method may prove to be useful in evaluating the occurrence and impact of oxidative stress-related events involving bioactive HNE in
heart diseases and aging in humans.