Ischemic preconditioning is a cardioprotective phenomenon involving adaptive changes in cells and molecules that occurs in a biphasic pattern: an early phase (1-2 h), and a late phase (12-24 h). Reactive oxygen species (ROS) are strongly involved in triggering ischemic preconditioning, but it is not clear if they play a major role in the early or late phase of preconditioning and which are the molecular mechanisms involved and the cellular effects generated. In this study we investigated the mechanisms behind H2O2 induced cardioprotection in cultured rat cardiomyocytes demonstrating the induction of phase II enzymes and the involvement of Nuclear factor E2-related factor 1 (Nrf1) and Nrf2. Rat neonatal cardiomyocytes were grown until confluence. Preconditioning was simulated with 100 µM H2O2 for 10 min and after different times oxidative stress was generated by 100 µM H2O2 for 30 min. Cell viability was evaluated by MTT and ROS production by 2’,7’-dichlorofluorescein-diacetate. Phase II enzyme expression and Nrf1 and Nrf2 nuclear translocation were determined by immunoblotting. Moreover, Nrf1 and Nrf2 were silenced with Nrf1-siRNA and Nrf2-siRNA, to evaluate the involvement in phase II enzyme induction. H2O2-preconditioning showed a higher ability to counteract oxidative stress in the late than in the early phase of adaptation. H2O2-preconditioning led to a decrease in intracellular ROS production related to the induction of glutathione reductase, catalase, thioredoxin reductase, and NADPH quinone oxidoreductase 1 expression. H2O2-preconditioning induced the nuclear translocation of both Nrf1 and Nrf2. The down-regulation of Nrf1 and Nrf2 by Nrf1-siRNA and Nrf2-siRNA inhibited the enzyme induction. These findings demonstrate that, both Nrf1 nd Nrf2 are involved in the induction of phase II enzymes by preconditioning, thus contributing to the cardioprotection elicited by these important antioxidant/detoxifying enzymes.
Fabbri D., Angeloni C., Motori E., Leoncini E. , Malaguti M., Lorenzini A., et al. (2011). Preconditioning modulates phase II enzymes in cultured cardiomyocytes by Nrf1 and Nrf2 activation.. Bologna : Società italiana di Ricerche Cardiovascolari.
Preconditioning modulates phase II enzymes in cultured cardiomyocytes by Nrf1 and Nrf2 activation.
FABBRI, DANIELE;ANGELONI, CRISTINA;LEONCINI, EMANUELA;MALAGUTI, MARCO;LORENZINI, ANTONELLO;HRELIA, SILVANA
2011
Abstract
Ischemic preconditioning is a cardioprotective phenomenon involving adaptive changes in cells and molecules that occurs in a biphasic pattern: an early phase (1-2 h), and a late phase (12-24 h). Reactive oxygen species (ROS) are strongly involved in triggering ischemic preconditioning, but it is not clear if they play a major role in the early or late phase of preconditioning and which are the molecular mechanisms involved and the cellular effects generated. In this study we investigated the mechanisms behind H2O2 induced cardioprotection in cultured rat cardiomyocytes demonstrating the induction of phase II enzymes and the involvement of Nuclear factor E2-related factor 1 (Nrf1) and Nrf2. Rat neonatal cardiomyocytes were grown until confluence. Preconditioning was simulated with 100 µM H2O2 for 10 min and after different times oxidative stress was generated by 100 µM H2O2 for 30 min. Cell viability was evaluated by MTT and ROS production by 2’,7’-dichlorofluorescein-diacetate. Phase II enzyme expression and Nrf1 and Nrf2 nuclear translocation were determined by immunoblotting. Moreover, Nrf1 and Nrf2 were silenced with Nrf1-siRNA and Nrf2-siRNA, to evaluate the involvement in phase II enzyme induction. H2O2-preconditioning showed a higher ability to counteract oxidative stress in the late than in the early phase of adaptation. H2O2-preconditioning led to a decrease in intracellular ROS production related to the induction of glutathione reductase, catalase, thioredoxin reductase, and NADPH quinone oxidoreductase 1 expression. H2O2-preconditioning induced the nuclear translocation of both Nrf1 and Nrf2. The down-regulation of Nrf1 and Nrf2 by Nrf1-siRNA and Nrf2-siRNA inhibited the enzyme induction. These findings demonstrate that, both Nrf1 nd Nrf2 are involved in the induction of phase II enzymes by preconditioning, thus contributing to the cardioprotection elicited by these important antioxidant/detoxifying enzymes.I documenti in IRIS sono protetti da copyright e tutti i diritti sono riservati, salvo diversa indicazione.
