Fig. 8
Regulatory effects of inactivated AMPK or ER stress on melatonin-mediated cellular protection against H2O2 damage. BMSCs were treated using the indicated agents for the indicated times to preinhibit AMPK and ER stress. a Cell viability, b ROS production, c mitochondrial superoxide levels, and d MMP levels are shown. e–g The TUNEL assay and C-Casp-3 immunofluorescence staining were conducted. The percentage of TUNEL+ cells and C-Casp-3 fluorescence intensity were used to determine the degree of apoptosis. Nuclei were labeled with DAPI. h, i Western blotting was used to measure and quantify the expression levels of p-AMPK, ER stress factors, apoptosis parameters, and mitochondrial function markers in BMSCs. p-PERK, Bip, p-eIF2α, and DDIT3 were the ER stress factors. Bcl2, Bax, and C-Casp-3 were the apoptosis parameters. Cyto-C served as the marker of mitochondrial function. Cells treated with PBS served as a control group. Values are expressed as the mean ± SD (n = 3 independent experiments). *p < 0.05 compared with different groups. One-way ANOVA followed by the LSD test was used to analyze the significance of differences. Scale bar = 50 μm. C-Casp-3 (C-C-3, C-C), cleaved Caspase-3; CpC, compound C dihydrochloride; Cyto-C, cytochrome C; Mel, melatonin; OD, optical density; ROS, reactive oxygen species; 4-PBA, 4-phenylbutyric acid