Figure 4.

The functions of lipotoxic HC-exosomal miR-27a in PINK1-related mitophagy and activation in HSCs

The functions of activated LX2 cells or PHSCs incubated with Exo-CN/Exo-PA derived from in-NC/in-miR27a LO2 cells were detected. (A) PINK1, Parkin, p62, and LC3B protein levels in activated LX2 cells were examined via western blotting. (B) Mitochondrial respiration of activated LX2 cells was assessed using Seahorse. (C) LC3B (red IF) and COX4 (green IF) proteins, mtSOX red, and mtCMXRos red in activated LX2 cells were observed via IF staining. Scale bar, 25 μm. (D) Representative images and statistical analysis of cell morphology changes in PHSCs at day 7 are shown. Scale bar, 25 μm. (E) Fibrosis-related (α-SMA), proliferation-related (cyclin D1 and PCNA) proteins were detected in activated LX2 cells via western blotting. All quantifications are presented as the mean ± SD, and p values were calculated using an unpaired Student's t test. Statistical significance: ∗p < 0.05, compared with the (B and C) Exo-CN group from in-NC LO2 cells, (D) Exo-CN group from in-NC LO2 cells among day 5 or 7 PHSCs.