Fig. 6
Effect of melatonin on mitochondrial function, ER stress, and AMPK signaling in response to H2O2-induced insults in BMSCs. BMSCs pretreated with melatonin at different concentrations (50 and 100 μM) for 6 h were exposed to 400 μM H2O2 for another 24 h. a ROS production, b mitochondrial superoxide levels, and c MMP levels were analyzed. d After pretreatment with melatonin at different concentrations, the cells subjected to H2O2 were stained with primary antibodies against Bip and CRT. Representative images of immunostaining in BMSCs are shown. Nuclei were labeled with DAPI. e Western blotting was used to measure and quantify the expression of cytochrome C (Cyto-C) translocation immunostaining in BMSCs. f After treatment with melatonin and H2O2, the levels of p-AMPK and proteins related to ER stress, such as p-PERK, Bip, p-eIF2α, and DDIT3, were determined using western blotting in BMSCs, and the values were normalized to that of β-actin. 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 the control group. One-way ANOVA followed by the LSD test was used to analyze the significance of differences. Scale bar = 50 μm. CRT, calreticulin; Cyto-C, Cytochrome C; Mel, melatonin; ROS, reactive oxygen species