4-Hydroxy-2-nonenal induces calcium overload via NADPH oxidase-derived reactive oxygen species in isolated rat cardiac myocytes

Abstract

Background: Oxidative stress has been implicated in the pathogenesis of heart failure. Recently, we and other investigators have reported that the amount of 4-hydroxy-2-nonenal (HNE), which is a major lipid peroxidation product and a cytotoxic aldehyde, is increased in the failing myocardium. However, the effect of HNE as a mediator of oxidative stress in cardiomyocytes has not been assessed. This study was therefore designed to determine whether HNE has a pro-oxidant effect in cardiac myocytes and whether HNE increases intracellular Ca concentration ([Ca]i). Methods and results: HNE (10 to 100 Amol/L) induced the production of reactive oxygen species (ROS) within isolated rat ventricular myocytes in a dose-dependent manner, as assessed by 2V, 7V-dichlorofluorescin diacetate fluorescence. Diphenyleneiodonium (DPI) (100 Amol/L), which is an NADPH oxidase inhibitor, significantly inhibited the generation of ROS (P <0.0001). A high level of HNE (400 Amol/L), which is available in pathological conditions, increased [Ca]i as assessed by fura-2 ratio in a time-dependent manner. After 20 min of HNE exposure, [Ca]i was significantly increased compared with that in the control medium (P <0.0001) and hypercontracture was induced in 67% of HNE-treated cells. Catalase (100 U/mL), an antioxidative enzyme, significantly attenuated the increase in [Ca]i induced by HNE (P <0.0005) and completely inhibited HNE-induced hypercontracture. Furthermore, carvedilol (0.1 Amol/L), a beta-blocker with potent antioxidant activity, significantly attenuated the increase in [Ca]i induced by HNE (P=0.002) and completely inhibited HNE-induced hypercontracture, but propranolol (0.2 Amol/L) did neither (P=NS). Conclusion: HNE causes intracellular Ca overload via NADPH oxidase-derived ROS formation in cardiomyocytes.