Figure 4
Autophagy impairment and autophagy induction under chronic hypoxia. (a) Neuro-2a cells were exposed to hypoxia and/or high glucose for 48 h, and the protein levels of P-AMPK, AMPK, P-mTOR and mTOR were measured by immunoblot. (b) The ImageJ densitometric analysis showed that chronic hypoxia upregulated the ratio of P-AMPK/AMPK and lowered the ratio of P-mTOR/ mTOR when compared with those under normoxia. High glucose decreased the ratio of P-AMPK/AMPK. “C” represented control conditions (25 mM glucose), “G” represented high glucose conditions (75 mM glucose). The results are expressed as the mean ± SEM from three independent experiments. Two-way ANOVA, #P < 0.05 High Glucose vs. Control; *P < 0.05 Hypoxia vs. Normoxia. (c) Neuro-2a cells were submitted to hypoxia and/or high glucose for up to 42 h and then subjected to 10 nM BAF for an additional 6 h. Immunoblots were performed to determine the levels of P62 and LC3B-II proteins in Neuro-2a cells. (d) The ImageJ densitometric analysis showed that BAF treatment increased the protein levels of LC3-II and P62 in Neuro-2a cells under normoxia but did not further change the expression of LC3-II and P62 under hypoxia. The results are expressed as the mean ± SEM from three independent experiments. Two-way ANOVA, **p < 0.01 treated vs. untreated. Full-length blots are presented in Supplementary Fig. S2.
