Qiweizhigan granule ameliorates metabolic dysfunction-associated steatohepatitis by modulating the galectin 3/tumor necrosis factor receptor-associated factor 6-mediated ferroptosis

Figure 6 Galectin 3 mediates ferroptosis. A: The levels of ferroptosis-related genes [achaete-scute family bHLH transcription factor 4 (ACSL4), transferrin receptor, glutathione peroxidase 4 (GPX4), solute carrier family 7 member 11] in RAW264.7 cells after galectin 3 (LGALS3) overexpression were verified; B: The levels of GPX4, ACSL4, and solute carrier family 7 member 11 in RAW264.7 cells were verified; C: Transmission electron microscopy observation of ferroptosis in RAW264.7 cells following LGALS3 overexpression was performed; D: LGALS3 knockdown efficiency and alterations of ferroptosis proteins (GPX4, ACSL4) in RAW264.7 cells were performed; E: Measurement of ferrous iron (Fe²⁺) content in RAW264.7 cells after LGALS3 knockdown; F: Measurement of total iron content in RAW264.7 cells after LGALS3 knockdown; G-I: The levels of malondialdehyde, superoxide dismutase, and glutathione were performed among the three groups; J: Quantification of ferrous iron (Fe²⁺) levels in liver tissues among the normal, choline-deficient, L-amino acid-defined high-fat diet (CDAHFD), and Qiweizhigan granule (QWZG) groups; K: Enhanced Prussian blue staining with DAB amplification showing iron deposition in liver sections among the normal, CDAHFD, and QWZG groups; L: GPX4 protein expression in liver tissues among the normal, CDAHFD, and QWZG groups was performed. Data were presented as means ± SEM. ^aP < 0.05, ^bP < 0.01, ^cP < 0.001. ACSL4: Achaete-scute family bHLH transcription factor 4; CDAHFD: Choline-deficient, L-amino acid-defined high-fat diet; GPX4: Glutathione peroxidase 4; GSH: Glutathione; LGALS3: Galectin 3; MDA: Malondialdehyde; QWZG: Qiweizhigan granule; SLC7A11: Solute carrier family 7 member 11; SOD: Superoxide dismutase; Tfrc: Transferrin receptor.