Fig 2: Establishment of a mouse model with APAP-induced hepatic injury. (a) Schematic diagram of the experiment. BALB/c mice (n = 10, at each time point) were fasted for 16 h. At 0 h, the mice were intraperitoneally administered APAP, and sacrificed at 4, 8, 28, and 52 h for sample collection. (b) Serum ALT levels were quantified using Fuji Dri-Chem NX 500v. The levels are expressed as median values with the interquartile range from more than three independent experiments. The significant differences between APAP from untreated and treated samples are indicated using asterisks (**p < 0.01). (c) Hepatic histology (H&E staining) at 0, 4, 8, 28, and 52 h post APAP-administration are shown. Images are representative of the treated mice. Scale bar = 100 μm. (d) Serum ACAA2 (upper panel) and albumin (lower panel) were detected using western blotting with α-mouse ACAA2 mAb and α-mouse albumin pAb, respectively. Liver lysate sample (3 μg) was used as the control. (e) The serum levels of ACAA2 were determined using ELISA. The serum ACAA2 level represents the mean ± SD from more than three independent experiments. The significant differences between APAP from untreated and treated samples are indicated using asterisks (**p < 0.01). The dotted line (OD450 = 0.1) indicates the detection limit.

Fig 3: Complex formation between S16 and serum ACAA2. (a) Schematic diagram of the experiment. BALB/c mice (n = 6, in each group) were fasted for 16 h. At 0 h, the mice were intraperitoneally administered saline or APAP (300 mg/kg). At 4 h, the mice were injected intraperitoneally with saline or S16 (100 µg), and then sacrificed at 28 h. (b) Serum was collected from mice after 28 h of APAP administration, and complexes of S16 with ACAA2 in each serum sample were determined using sandwich ELISA. The values represent the mean ± SD from three independent experiments. Significant differences between groups are indicated using asterisks (**p < 0.01). The dotted line (OD450 = 0.1) indicates the detection limit.

Fig 4: A schematic showing the effects of QSG on FA and glucose metabolism in HF induced by AMI. QSG exerted a remarkable regulatory effect on lipid metabolism by lowering serum levels of TC, TG and LDL-C. QSG activated FAT/CD36-CPT1-FAO signaling through upregulating the expressional levels of FAT/CD36, CPT1A, ACADL, ACADM, ACAA2 and SCP2, which would lead to an increase of FA uptake, transportation into mitochondria and β-oxidation. QSG promoted FA metabolism to a large extent on the up-regulation of transcriptional regulator PPARα, RXRα, RXRβ, RXRγ and PGC-1α. LDHA and PDK4 involved in glycolysis and glucose oxidation were all down-regulated by treatment with QSG, indicating QSG inhibited uncoupling of glycolysis from glucose oxidation. QSG facilitated TAC and the transfer of ATP from mitochondria to cytoplasm by increasing the protein levels of CKMT2 and SUCLA2. Moreover, the mitochondrial function was enhanced with QSG administration proved by the increased PGC-1α and the decreased UCP2

Fig 5: Binding of S16 with ACAA2. HEK293 cells were transfected with vector alone or ACAA2-FLAG expressing vector. The ACAA2-FLAG expressed in the cells was purified. The whole cell lysate (2 μg) transfected with vector alone (Vector), ACAA2-FLAG, and purified ACAA2-Flag (0.1 and 0.5 μg) were separated using 12% SDS-PAGE. The gel was subjected to CBB staining (a) and western blotting using α-Flag M2 mAb (b) or S16 (c). Molecular weight markers are shown at the left of the gels (kDa). The images shown are representative results of three independent experiments.