Neferine prevents autophagy induced by hypoxia through activation of Akt/mTOR pathway and Nrf2 in muscle cells.

We have previously reported the prevention of Hypoxia mediated apoptosis by Neferine, an alkaloid from Nelumbo nucifera. The present study was designed to assess the role of neferine in modulation of hypoxia induced autophagy in muscle cells. Cytoprotective dose of neferine in muscle cells (Rhabdomyosarcoma cells) exposed to hypoxia was determined by MTT assay. Hypoxia induced oxidative stress in muscle cells by enhancing lipid peroxidation and depleting cellular antioxidant enzymes. The inhibition of PI3K/Akt/mTOR cell survival signaling acts as a trigger for the hypoxia induced Autophagy. Hypoxia exposure in muscle cells resulted in acidic vesicular formation, as studied by acridine orange staining which serves as an indicator of autophagosome formation. Pretreatment with neferine inhibited autophagy induction by activating Akt/mTOR pathway and down regulating Beclin 1, PI3KCIII and LC3B-II in cells exposed to hypoxia. Further, Neferine also modulated hypoxia mediated changes in the cellular antioxidant levels by Nrf2 nuclear translocation. Collectively, results of the study suggest the role of neferine in preventing hypoxia induced autophagy in muscle cells.