Abstract 16498: AMP-Activated Protein Kinase Plays an Essential Role in Sildenafil-Induced Cardioprotection against Ischemia-Reperfusion Injury
Background: Sildenafil (SIL), a inhibitor of phosphodiesterase-5 induces powerful protection against myocardial ischemia/reperfusion (I/R) injury through nitric oxide (NO) and protein kinase G (PKG). AMP-activated protein kinase (AMPK) is a key regulator of myocardial energy metabolism and plays a protective role in ischemic myocardium. Since the catalytic subunit α2 of AMPK is highly expressed in heart and AMPK interacts with both NO and PKG, we tested the hypothesis that AMPK mediates sildenafil-induced cardioprotection against I/R injury, using the transgenic mice with cardiac-specific overexpression of a dominant-negative AMPKα2 subunit (dn-AMPKα2).
Methods & Results: Adult male dn-AMPKα2 and wild-type (WT) mice were pretreated with saline or SIL (0.7 mg/kg i.p.). One hour later, the hearts were subjected to I/R by occlusion of coronary artery for 30 min followed by reperfusion for 24 hours. As shown in Fig. A, myocardial infarct size was significantly reduced in SIL-treated WT mice as compared to saline treated mice (P<0.05, n=6/group). The infarct-limiting effect of SIL was abolished in dn-AMPKα2 mice (Fig. A). At the cellular level, ventricular cardiomyocytes were isolated from dn-AMPKα2 and WT mice and subjected to 40 min of simulated ischemia (SI) with/without 1 hr pre-incubation with SIL (1 µM). Myocyte necrosis, apoptosis and mitochondrial membrane potential were determined after reoxygenation (RO) using trypan blue, TUNEL assay and JC-1 staining, respectively. Sildenafil protected WT cardiomyocytes against necrosis (Fig. B) and apoptosis (Fig. C) after SI-RO as compared to control and also attenuated the loss of JC-1 fluorescence (Fig. D). However, these protective effects of SIL were completely abrogated in dn-AMPKα2 cardiomyocytes (Fig. B, C, D).
Conclusion: The loss of protective effects of SIL in the dn-AMPKα2 mice strongly suggests that activation AMPK is essential for SIL-induced cardioprotection against I/R injury.
- © 2012 by American Heart Association, Inc.