Figure 6.

Erythropoietin (EPO) enhances the hypoxia‐induced autophagy in an AMP‐activated protein kinase‐dependent manner in PC12 cells. PC12 cells were cultured under a conventional atmosphere for 24 h followed by an oxygen‐free atmosphere of 95% N₂ and 5% CO₂. Prior to hypoxia, experimental wells were treated with EPO (50 U/mL), or combined with compound C (5 μmol/L), or compound C (5 μmol/L) alone for 2 h. A, The cell viability was determined by CCK8 assay. B,The protein levels of LC3II/I, Beclin 1, and p62 were determined by Western blotting. β‐actin was used as the loading control. C‐D, Histograms of the relative expression of LC3‐II/I, Beclin 1, and p62. E, Representative fluorescent images of PC12 cells incubated with tandem sensor RFP‐GFP‐LC3B kit and treated as indicated. F, The numbers of yellow puncta (autophagosomes) and red puncta (autolysosomes) in the merged images were counted. Data are mean ± SD, n = 4 per group. **P < .01, *P < .05 vs hypoxia group, ^# P < .05 vs the hypoxia + EPO group