Figure 4.

D-GalN-induced mitochondrial dysfunction and ROS overgeneration in HepG2 cells are improved by hPH treatment. HepG2 cells were stimulated with D-GalN (50 mM, 24 h) in the presence or absence of 5% hPH. (A) HepG2 cells post-hPH treatment were immunostained with anti-Tomm20 antibody, followed by the addition of Cy3-conjugated secondary antibody. Nuclei were identified using DAPI staining (scale bar=5 µm). (B) MitoTracker fluorescence signals for mitochondrial mass were measured. (C) ΔΨm was measured under the indicated conditions using the ΔΨm-sensitive fluorochrome MitoTracker Red CMXRos. (D) Cells were stained with 2′,7′-dichlorofluorescin diacetate for 30 min to measure intracellular hydrogen peroxide. (E) Cells were stained with MitoSOX for 30 min to measure mitochondrial ROS. Bar graphs show relative analysis. All data are presented as the mean ± standard error of the mean. ^**P<0.01, vs. D-GalN group. hPH, human placental hydrolysate; D-GalN, D-galactosamine; Con, control; MMP/ΔΨm, mitochondrial membrane potential; ROS, reactive oxygen species.