Hydrogen (H₂) Inhibits Isoproterenol-Induced Cardiac Hypertrophy via Antioxidative Pathways

Background and Purpose: Hydrogen (H₂) has been shown to have a strong antioxidant effect on preventing oxidative stress-related diseases. The goal of the present study is to determine the pharmacodynamics of H₂ in a model of isoproterenol (ISO)-induced cardiac hypertrophy. Methods: Mice (C57BL/6J; 8-10 weeks of age) were randomly assigned to four groups: Control group (n = 10), ISO group (n = 12), ISO plus H₂ group (n = 12), and H₂ group (n = 12). Mice received H₂ (1 ml/100g/day, intraperitoneal injection) for 7 days before ISO (0.5 mg/100g/day, subcutaneous injection) infusion, and then received ISO with or without H₂ for another 7 days. Then, cardiac function was evaluated by echocardiography. Cardiac hypertrophy was reflected by heart weight/body weight, gross morphology of hearts, and heart sections stained with hematoxylin and eosin, and relative atrial natriuretic peptide (ANP) and B-type natriuretic peptide (BNP) mRNA levels. Cardiac reactive oxygen species (ROS), 3-nitrotyrosine and p67 (phox) levels were analyzed by dihydroethidium staining, immunohistochemistry and Western blotting, respectively. For in vitro study, H9c2 cardiomyocytes were pretreated with H₂-rich medium for 30 min, and then treated with ISO (10 μM) for the indicated time. The medium and ISO were re-changed every 24 h. Cardiomyocyte surface areas, relative ANP and BNP mRNA levels, the expression of 3-nitrotyrosine, and the dissipation of mitochondrial membrane potential (MMP) were examined. Moreover, the expression of extracellular signal-regulated kinase1/2 (ERK1/2), p-ERK1/2, p38, p-p38, c-Jun NH2-terminal kinase (JNK), and p-JNK were measured by Western blotting both in vivo and in vitro. Results: Intraperitoneal injection of H₂ prevented cardiac hypertrophy and improved cardiac function in ISO-infused mice. H₂-rich medium blocked ISO-mediated cardiomyocytes hypertrophy in vitro. H₂ blocked the excessive expression of NADPH oxidase and the accumulation of ROS, attenuated the decrease of MMP, and inhibited ROS-sensitive ERK1/2, p38, and JNK signaling pathways. Conclusion: H₂ inhibits ISO-induced cardiac/cardiomyocytes hypertrophy both in vivo and in vitro, and improves the impaired left ventricular function. H₂ exerts its protective effects partially through blocking ROS-sensitive ERK1/2, p38, and JNK signaling pathways.