Abstract 17462: Angiotensin II-Mediated Suppression of Skeletal Muscle Autophagy via mTOR and FoxO3a Pathways

Abstract

Background: Chronic skeletal muscle (SK.M) atrophy, cachexia, occurs in diseases such as congestive heart failure (CHF), which is associated with high levels of angiotensin II (Ang-II). We have shown that Ang-II infusion reproduces major aspects of CHF-induced SK.M atrophy and causes mitochondrial (MT) damage and dysfunction in SK.M. Ang-II also inhibits autophagy/mitophagy, and a combination of these effects likely causes accumulation of damaged MT, leading to SK.M atrophy. Hypothesis: Ang-II infusion suppresses autophagy in SK.M via activation of mTOR and inhibition of FoxO3a. Aim: To understand the molecular mechanisms whereby Ang-II inhibits autophagy/mitophagy in SK.M. Methods: 12 weeks old male FVB mice were infused with Ang-II (1.0 μg·kg -1 ·min -1 ) or saline via osmotic minipumps (ALZET 1007D) for 7 days (N=6). Expression of autophagic markers and FoxO3a target genes were analyzed in tibialis anterior muscles by western blotting (at d1 & d7) and qPCR (at d4), respectively. Colchicine treatment was included in the analyses to assess autophagic flux in vivo . For statistical analysis Two-way ANOVA and t-test were used. Results: In the presence of colchicine, LC3 lipidation was decreased in Ang-II infused mice, indicating that Ang-II inhibits autophagy at the autophagosome formation step. Furthermore, Ang-II increased p62 and PINK1 and decreased BNIP3, suggesting that Ang-II-mediated reduction of autophagy causes disruption of damaged MT clearance via mitophagy. Mechanistically, Ang-II likely inhibits autophagosome formation via two pathways, mTOR and FoxO3a: Ang-II increased phosphorylation of mTOR at Ser2448 (p<0.005) and its downstream target 4E-BP1 at Thr37/46 (p<0.0005). Lastly, Ang-II increased phosphorylation of FoxO3a at Thr32 (p<0.0005), associated with reduced mRNA expression of FoxO3a target genes, such as LC3B and BNIP3 (24% and 46% reductions, respectively). Conclusion: Ang-II inhibits autophagosome formation in SK.M via mTOR activation and FoxO3a inhibition. Suppression of autophagy via these pathways likely disrupts clearance of damaged MT, leading to SK.M atrophy. These results provide insights for future development of therapies for CHF-induced cachexia by targeting mTOR and FoxO3a pathways to restore autophagy.