443 Beta1-adrenergic receptor polymorphisms predict heart failure progression in idiopathic dilated cardiomyopathy

Abstract

Background: Mitochondria is known to represent the main intracellular source of ROS. However, the role that the mitochondrial KATP (mKATP) channel could play in the oxidative stress during myocardial hypoxiareoxygenation remains unclear. Methods: Spontaneously beating hearts were dissected from 4-day-old chick embryos, mounted in vitro and submitted successively to 45 min normoxia (21% O2), 30 min anoxia (0% O2) and 60 min reoxygenation at 37°C. The time-course of ROS production in the ventricle was determined by measuring changes in fluorescence resulting from oxidation of the intracellular probe DCFH (10 μM) and expressed as arbitrary units per second (a.u./s). In order to determine the contribution of the mitochondria-derived ROS and its dependence on O2 concentration during reoxygenation, hearts were reoxygenated with 21% or 98% O2 in the absence (control) or in the presence of the complex III inhibitor myxothiazol (10μM). The involvement of the mKATP channel in ROS production has been assessed by using an opener (diazoxide, 50μM) or a blocker (5-hydroxydecanoate, 500 μM) of this channel. Results: In control hearts, ROS production was 0.18±0.04 (n=6) a.u./s under steady normoxia (mean±SD), was suppressed by anoxia and peaked after 9±3 min of reoxygenation, reaching 1.02±0.37 (n=6) and 1.63±0.34 (n=3) a.u./s with 21 and 98% O2, respectively (mean±SD). In myxothiazol-treated hearts, the peak of ROS at reoxygenation was reduced to 0.29±0.18 (n=3) and 0.41±0.08 (n=3) a.u./s with 21 and 98% O2, respectively. With respect to control, 5-hydroxydecanoate (n=5) and diazoxide (n=5) had no effect on DCFH signal measured under normoxia. By contrast, the peak of ROS production during the phase of reoxygenation was doubled by diazoxide (p<0.03) whereas 5-hydroxydecanoate had no effect. Furthermore, although the mKATP channel opener was markedly prooxidant during reoxygenation, it did not significantly alter the chrono-, dromoand inotropic disturbances observed during oxygen deprivation and readmission. Conclusion: It appears that the mKATP channel of the developing myocardium remains in closed state throughout anoxia-reoxygenation, limiting the mitochondrial oxidative burst.