Hypoxia Induced by Cobalt Chloride Triggers Autophagic Apoptosis of Human and Mouse Drug-Resistant Glioblastoma Cells through Targeting the PI3K-AKT-mTOR Signaling Pathway

Figure 3

Effects of hypoxia induced by CoCl₂ on mitochondrial NADH dehydrogenase activity, ATP levels, cell proliferation, cell autophagy, and levels of light chain (LC)3-I and LC3-2 in human drug-resistant glioblastoma cells. Human TMZ-tolerant U87 MG-R cells were selected from TMZ-sensitive U87 MG cells. U87 MG-R glioblastoma cells were treated with hypoxia for 6, 12, and 24 h. (a) Activity of mitochondrial NADH dehydrogenase was assayed using a colorimetric method. (b) Levels of ATP were measured using a bioluminescence assay. (c) Cell proliferation was measured by a thymidine incorporation assay. (d) Autophagic cells with acidic vesicular organelles were observed and photographed using a fluorescent microscope (left panel) and quantified with flow cytometry (right panel). (e) Levels of LC3-I and LC3-II were immunodetected (top panels). β-Actin was measured as the internal control (bottom panels). (f) These protein bands were quantified and statistically analyzed. Mouse GL261-R glioblastoma cells were exposed to hypoxia for 24 h. (g) Autophagic cells were quantified. Data are expressed as the for . vs. control.

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