Inhibition of Dystrophin Breakdown and Endothelial Nitric-Oxide Synthase Uncoupling Accounts for Cytoprotection by 3-[2-[4-(3-Chloro-2-methylphenyl)-1-piperazinyl]ethyl]-5,6-dimethoxy-1-(4-imidazolylmethyl)-1H-indazole Dihydrochloride 3.5 Hydrate (DY-9760e) in Left Ventricular Hypertrophied Mice

Abstract

Using a heart ischemia/reperfusion model in rats, we recently demonstrated that 3-[2-[4-(3-chloro-2-methylphenyl)-1-piperazinyl]ethyl]-5,6-dimethoxy-1-(4-imidazolylmethyl)-1H-indazole dihydrochloride 3.5 hydrate (DY-9760e), a calmodulin inhibitor, is a cardioprotective drug. Here, we examined cardioprotective mechanisms of DY-9760e in hypertrophy and heart failure using a mouse transverse aortic constriction (TAC) model. Mice were subjected to TAC and 2 weeks later they were administered DY-9760e for another 6 weeks (at 10 or 20 mg/kg/day p.o.). Chronic administration inhibited TAC-induced increased heart-to-body weight ratio dose-dependently. Consistent with inhibition of hypertrophy, fraction shortening, an indicator of heart contractile function, assessed by echocardiography was completely restored by DY-9760e (20 mg/kg/day) administration. Inhibition of TAC-induced atrial natriuretic peptide (ANP) up-regulation further confirmed an antihypertrophic effect of DY-9760e. It is noteworthy that we found that breakdown of dystrophin and spectrin by calpain was associated with heart failure in TAC mice. Caveolin-3 breakdown was closely associated with endothelial nitric-oxide synthase (eNOS) dissociation from the plasma membrane and its subsequent uncoupling. Uncoupled monomeric eNOS formation was associated with increased protein tyrosine nitration, suggesting peroxynitrite production and NO and superoxide formation. It is important to note that 6 weeks of DY-9760e treatment significantly blocked hypertrophic responses, such as increased heart weight and ANP induction. Overall, we show that inhibition of both dystrophin/spectrin breakdown and uncoupling of eNOS probably underlies the cardioprotective mechanisms of DY-9760e. The observed protection of sarcolemmal proteins and eNOS by DY-9760e during pressure overload suggests a novel therapeutic strategy to rescue the heart from hypertrophy-induced failure.