Abstract P046: Hsp20 Promotes Cardiac Autophagy via Interaction with Beclin-1
Hsp20 has been shown to prevent stress-triggered cardiac injury. However, it remains obscure whether Hsp20-elicited cardioprotection is associated with the activation of autophagy. We first assessed cardiac autophagy in a transgenic (TG) mouse model with 10-fold overexpression of Hsp20 by immunoblotting. Levels of two autophagy markers (LC3II and Beclin1) were increased by 1.6- and 2.2-fold, respectively, in Hsp20-hearts, compared to WTs. To examine whether Hsp20 activates autophagic flux in the heart, mice were i.p. injected with chloroquine (CQ, 50mg/kg), an inhibitor for autophagosome-lysosome fusion, for 4h. Cadaverine dye-binding analysis showed that autophagy levels were increased by 1.5-fold in CQ-treated Hsp20-hearts compared to saline controls. To examine whether increased autophagy levels contribute to Hsp20-induced cardioprotection, 3-methyladenine (3-MA, an inhibitor for autophgy) was i.p. injected into TG mice (30mg/kg). One hour later, hearts were subjected to global no-flow ischemia/reperfusion (I/R: 45min/1h), and showed that contractile function (+dP/dt) was recovered by 93α 5 % in saline-treated Hsp20-hearts, comparable to 74 α 6 % recovery in 3-MA-treated TGs (n=6, p<0.05), indicating that the activation of autophagy by Hsp20 is beneficial for hearts upon I/R. To exclude in vivo compensatory effects, Hsp20 was delivered into isolated adult cardiomyocytes by adenoviral vector. Western-blotting analysis indicated that acute expression of Hsp20 increased the levels of LC3II and Beclin1 by ∼2-fold, compared to GFP control. Accordingly, overexpression of Hsp20 augmented myocyte survival upon addition of H2O2, as related to GFP-control. Furthermore, co-infection with AdBeclin1-siRNA attenuated the protective effects of Hsp20 in myocytes upon H2O2 stress. Excitingly, Hsp20 was found to localize at the LC3-labeled autophagosome. Indeed, co-immunoprecipitation results showed an interaction of Hsp20 with Beclin-1 in the heart. Moreover, a competitive ELISA assay revealed that Hsp20 dose-dependently suppressed the interaction of Beclin1/Bcl-2, a known complex negatively regulating autophagy. In summary, Hsp20 activates cardiac autophagic flux via interaction with Beclin1, thereby confering cadioprotection.
- © 2011 by American Heart Association, Inc.