CARD2: Plasma Homocysteine and Oxidative Stress Markers May Predict the Stress Burden of Human Hearts Donated After Circulatory Death

Abstract

Background: Initial result of heart transplantation using donation after circulatory death (DCD) heart with warm ischemic time (WIT) less than 30 minutes was encouraging. However, little is known about how donor respond to WIT systemically and its impact on the heart with subsequent cold storage as used in normothermic regional perfusion(NRP) on the DCD hearts. Since elevated homocysteine (Hcy) and oxidative stress burden in cardiovascular system have been associated with ischemia, we investigated if circulating Hcy, oxidative stress and antioxidant were elevated in DCD blood and myocardium and if there is temporal relationship with the WIT and subsequent cold storage. Methods: We studied 12 human DCD hearts for research (WIT range:20-110 min). They were assigned to 3 groups of WIT [20 (n=4), 40 (n=5), ≥60 (n=3) min.]. Two time-point donor blood samples were collected from each donor at the baseline (prior to or at extubation) and at the cross-clamp. All hearts received del Nido cardioplegia before they were placed at cold storage for 6 hours. Left ventricular (LV) biopsies were performed at hour 0,2,4 and 6. Circulating Hcy, oxidative stress (oxidized low-density lipoprotein: OxLDL, malondialdehyde: MDA) and total antioxidant in blood were quantified. The MDA and GPx activity (glutathione peroxidase: antioxidant) were quantified in cardiac biopsies. Results: Change in plasma Hcy (Fig.1A), oxidative stress (Fig.1B,C) and antioxidant (Fig.1D) found to be comparable between the 3 groups at baseline; while significant differences were noticed in samples at cross-clamping; especially in DCD≥60 groups. The WIT found to be positively correlated Hcy and oxidative stress at cross-clamping; and negatively correlated with antioxidant (p<0.05,Table:1). The myocardial oxidative stress (MDA) found to be minimal at baseline (DCD20 and 40), and slightly elevated at 2 hrs. and maintained at similar levels at 4 and 6 hrs. of cold storage (Fig.1E); while a preexisting high oxidative stress at baseline in DCD≥60, an exponential increase during the cold storage indicating severity of stress burden. Temporal decrease in cardiac antioxidant (GPx) found to be more pronounced in DCD≥60 group, while the rates of change are comparable in DCD:20 and 40 groups (Fig.1F). The change of blood markers was found to be correlated with change of myocardial markers(Table:1). Conclusions: Systemic warm ischemia promoted hyperhomocystenemia and increased oxidative stress and might be used as initial screening biomarkers for the stress burden on the donor hearts. The correlation between blood and myocardial markers suggested the systemic markers can potentially be used in predicting stress burden of donor hearts. The duration of cold storage was related to progressive increase in myocardial oxidative stress and diminished antioxidant status. We recommend rejecting DCD hearts with WIT≥60 minutes, as both the blood and tissue data indicating maximum oxidative stress burden in this group.