Abstract 16025: Aldosterone Induces Autophagy via an ERK-JNK/Phospho-Bcl-2/Beclin-1 Dependent Pathway in Isolated Adult Rat Ventricular Myocytes
Background: Aldosterone induces left ventricular (LV) remodeling and heart failure (HF). Aldosterone increases reactive oxygen species (ROS) production and mediates its effects through JNK-MAPK signaling. Autophagy plays a role in the pathogenesis of LV remodeling and HF, but it is unknown whether aldosterone causes cardiac autophagy. We hypothesized that aldosterone induces cardiomyocyte autophagy in vitro.
Methods: Isolated adult rat ventricular myocytes (ARVMs) were exposed to aldosterone (1µM/18hrs). Autophagy related proteins and intracellular signaling molecules were measured by qRT-PCR and immunoblotting.
Results: Aldosterone increased LC3II expression by 39±4% (p<0.01 vs. control) and the LC3II:LC3I protein expression ratio by 39±6% (p<0.05 vs. control). Aldosterone increases ROS which is known to increase p62. p62 binds LC3 and is a selective substrate for autophagy and regulates the formation of autophagosomes. Aldosterone increased p62 (37±3%) and ATG4B (48±8%) expression (p<0.05 vs. control, for both). ATG4B is regulated by ROS. Spironolactone (100mM/30min) significantly decreased aldosterone-induced LC3II:LC3I ratio by 56±5% and ATG4B expression by 33±3% vs. aldosterone. We sought to examine the aldosterone-induced autophagy pathway. mTOR is a critical regulator of autophagy, with activated mTOR suppressing autophagy, and negative regulation of mTOR promoting it. Aldosterone did not inhibit p-mTOR expression in ARVMs. Co-incubation with rapamycin, an mTOR inhibitor, had no augmented effect on LC3II expression. Aldosterone however, increased p-Bcl-2 expression (56±8%) with a subsequent downstream increase in Beclin-1 expression (30±5%) as well as BNIP-3 expression by 4.6±0.9 fold (p<0.05 vs. control, for all). Co-incubation with the specific JNK inhibitor (SP600125) decreased aldosterone-induced p-Bcl-2 expression by 22±8% (p<0.05 vs. aldosterone), while also leading to a consistent decrease in LC3II expression. When co-incubated with SP600125 and the MEK inhibitor U0126, aldosterone-mediated LC3II expression was further decreased by 25±3% (p<0.01 vs. aldosterone).
Conclusion: Aldosterone induces cardiomyocyte autophagy independent of mTOR but via an ERK/JNK - pBcl-2 - Beclin-1 dependent pathway.
- © 2011 by American Heart Association, Inc.