Fig. 6: BBR downregulates autophagy to enhance ROS/ferrous-mediated HSC ferroptosis.

A, B IHC for 4-HNE, Perls’-DAB for ferric ion and IF for FTH1 in TAA-induced hepatic fibrosis mouse. C HSC-LX2 was exposed to BBR (21 μM), CQ (10 μM) or 3-MA (5 mM) for 24 h; lipid ROS was investigated. D HSC-LX2 was treated with BBR (21 μM) or CQ (10 μM) for 24 h; markers of fibrosis, autophagy, and ferroptosis were assayed. E–H HSC-LX2 was exposed to BBR (21 μM), CQ (10 μM), or 3-MA (5 mM) for 24 h; cellular Fe²⁺, MDA, GSH, and cell viability were investigated. I HSC-LX2 cells were treated with BBR (21 μM) with or without CQ (10 μM) or Fer-1 (1 μM) for 24 h, and cell death was quantified by PI staining. J–L HSC-LX2 was exposed to BBR (21 μM) followed by trehalose (50 mM) for 12 h; lipid ROS, markers of autophagy, ferroptosis, cellular Fe²⁺, MDA, and cell viability were investigated. M–O HSC-LX2 cells were pretreated with Fer-1 (1 μM) for 1 h, followed by BBR (21 μM) for 24 h; lipid ROS, markers of autophagy, ferroptosis, cellular Fe²⁺, GSH, and MDA were investigated. P, Q HSC-LX2 was pretreated with DFO (25 µM) for 1 h, following treatment with BBR (21 μM) for 24 h; lipid ROS, cellular Fe²⁺, cell viability, and MDA were investigated. The results are expressed as mean ± SD. Data are representative of three independent experiments; n = 3–6 in every group; Scales: 100 μm. Representative photographs are shown. Compared with the control group, **P < 0.01; compared with the BBR group, ^#P < 0.05, ^##P < 0.01.