Fig. 6
Melatonin inhibited MAMs formation and mitochondrial calcium overload in TS-treated hepatocytes by suppressing IP3R1 expression. A, B Representative blots and quantitative analysis of IP3R1, cleaved-caspase 3 (n = 4). C Relative activity of caspase 3. D The levels of ALT release (fold difference compared with TS + Ad-EV). E Representative confocal images of primary hepatocytes double-stained by mitotracker (red) and ER-localized virus (green) at × 600 magnification. F, G Statistical quantification of the colocalization area between mitochondria and ER. H Representative images and quantitative analysis of MitoSOX-stained mitochondria-derived superoxide production. I Quantification of fluorescence intensity normalized by TS + Ad-EV. J Representative traces of ionomycin-induced changes of mitochondrial Ca2+ concentration (n = 3). K Quantification of the relative increment of fluorescence intensity in mitochondrial Ca2+ concentration after ionomycin stimulation normalized by TS + Ad-EV. L Cell viability (percentage of TS + Ad-EV). M, N Oxygen consumption rate (OCR) measured by Seahorse and quantitative statistical analysis of OCR (n = 3). All of the values are shown as the means ± SEM. n = 6 in each group. *P < 0.05, **P < 0.01 vs TS + Ad-EV; ##P < 0.01 vs TS + Mel + Ad-EV