4-Hydroxy-2-nonenal Induces Calcium Overload via the Generation of Reactive Oxygen Species in Isolated Rat Cardiac Myocytes

Background: It has been reported that that the amount of 4-hydroxy-2-nonenal (HNE), which is a major lipid peroxidation product and a cytotoxic aldehyde, is increased in the human failing myocardium. This study was designed to determine whether HNE has a pro-oxidant effect in cardiac myocytes and whether HNE causes Ca²⁺ overload. Methods and Results: Exposure to HNE for 10 minutes in the presence of ferric nitrilotriacetate induced the production of hydroxyl radical (·OH) in the rat myocardium as assessed by electron spin resonance spectroscopy, and HNE induced the generation of reactive oxygen species (ROS) in a dose-dependent manner as assessed by 2′, 7′-dichlorofluorescein diacetate fluorescence. HNE increased intracellular Ca²⁺ concentration ([Ca²⁺]_i) as assessed by fura-2 ratio in a dose- and time-dependent manner. After 20 minutes of HNE (400 μmol/L) exposure, hypercontracture was induced in 67% of the cells. Catalase, an antioxidative enzyme that can decompose hydrogen peroxide (H₂O₂), significantly attenuated the increase in [Ca²⁺]_i and completely inhibited hypercontracture. Carvedilol, a β-blocker with potent antioxidant activity, also significantly attenuated the increase in [Ca²⁺]_i and completely inhibited hypercontracture, but propranolol had no effect on either [Ca²⁺]_i increase or hypercontracture. Conclusions: HNE induces the formation of ROS, especially H₂O₂ and ·OH, in cardiomyocytes and subsequently ROS cause intracellular Ca²⁺ overload. HNE formation may play an important role as a mediator of oxidative stress in heart failure.