Autophagy is promoted as a response to such environmental stress conditions as ATP depletion and excessive accumulation of reactive oxygen species (ROS). Multiple signaling pathways, including AMP-activated protein kinase (AMPK), are indicated to promote autophagy in ischemic/hypoxic heart. However, it's far more to clarify the orchestrated crosstalk between AMPK and other signaling pathways in the autophagy. In the present study, we investigated the autophagy induction by hypoxia in Rat H9C2 cardiomyocytes with LC3-EGFP reporter, electron microscopy and western blot analysis. Then we examined the promotion of Forkhead box O (FOXO) 3, one member of FOXO transcriptional protein family, by hypoxia in Rat H9C2 cells and determined the mediation of FOXO 3 in the hypoxia-induced autophagy in H9C2 cells. In addition, we investigated the role of AMPK signaling in the FOXO3-mediated, hypoxia-induced autophagy in H9C2 cells. It was demonstrated that hypoxia induced significant autophagy in H9C2 cells, via promoting autophagic vesicles, inducing the conversion of LC3-I to LC3-II and upregulating autophagy-related markers. Moreover, FOXO3 was upregulated by the hypoxia in H9C2 cells; and the knockdown of FOXO3 significantly reduced the hypoxia-induced autophagy. In addition, AMPK signaling was significantly promoted by hypoxia in H9C2 cells, and the chemical manipulation of AMPK exerted significant influence on the hypoxia-induced autophagy and on the FOXO3 level. In conclusion, FOXO3 regulated the hypoxia-induced autophagy in cardiomyocytes, and AMPK mediated the FOXO3 promotion during the autophagy induction by hypoxia, implying the key regulatory role of FOXO3 and AMPK signaling in the hypoxia-induced autophagy in cardiomyocytes.
- Forkhead box O (FOXO) 3
- AMP-activated protein kinase (AMPK)
- ©2016 The Author(s)
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