Table 1.
Primers used for real-time quantitative PCR.
Fig 1.
Pre-treatment with paeonol protected against APAP-induced liver injury.
Mice were administered with either vehicle (0.5% CMC-Na) or paeonol (25, 50, 100 mg/kg) by gavage for three days. At day 3, mice were intraperitoneally (i.p.) injected with either 400 mg/kg APAP or an equal volume of PBS. Liver tissues were collected after APAP administration for 6 h (A) and 24 h (B). Liver sections were stained with H&E. Magnification: 200X. (C) After APAP administration for 6 h, blood was collected and the levels of ALT and AST in the serum were determined by a commercially available kit. Data are shown as means ± S.E.M. *P<0.05, **P<0.01 v.s. APAP treatment (n = 8).
Fig 2.
Post-treatment with paeonol protected against APAP-induced liver injury.
Mice were intraperitoneally injected with either 400 mg/kg APAP or an equal volume of PBS. After 30 min, mice were administered with either vehicle (0.5% CMC-Na) or paeonol (25, 50, 100 mg/kg) by gavage. Liver tissue and blood were collected after paeonol treatment for 24 h. (A) Liver sections were stained with H&E. Magnification: 200X. (B) The levels of ALT and AST in the serum were determined by a commercially available kit. Data are shown as means ± S.E.M. *P<0.05, **P<0.01 v.s. APAP treatment (n = 8).
Fig 3.
Paeonol inhibited APAP-induced MAPK pathway activation.
The protein levels of total and phosphorylated p38, JNK and Erk1/2 in the liver were determined by western blot. β-actin was used as the endogenous control. The representative data are shown and bands were analyzed by densitometry. Data are shown as means ± S.E.M. *P<0.05, **P<0.01 v.s. APAP treatment.
Fig 4.
Paeonol reduced APAP-induced hepatic oxidative stress.
(A) Mice were administered with either vehicle (0.5% CMC-Na) or paeonol (25, 50, 100 mg/kg) by gavage for three days. At day 3, mice were intraperitoneally injected with either 400 mg/kg APAP or an equal volume of PBS. Liver was collected and hepatic homogenates were used for the determination of MDA, SOD, GSH and GSH-PX levels by using commercial kits. (B) Mice were administered with either vehicle (0.5% CMC-Na) or paeonol (25, 50, 100 mg/kg) by gavage for three days. At day 3, mice were co-treated with either 400 mg/kg APAP and 300 mg/kg NAC by intraperitoneal injection. The levels of ALT and AST in the serum were determined by a commercially available kit. (C) The protein levels of total and phosphorylated JNK in the liver were determined by western blot. β-actin was used as the endogenous control. The representative data are shown and bands were analyzed by densitometry. Data are shown as means ± S.E.M. *P<0.05, **P<0.01 v.s. APAP treatment (n = 8).
Fig 5.
Paeonol prevented against H2O2 or APAP-induced LDH releasing and ROS production in primary mouse hepatocytes.
Primary mouse hepatocytes were treated with paeonol (20, 40, 80 μM) in the absence or presence of 5 mM APAP or 250 μM H2O2 for 6 h. (A) LDH leakage percentage were determined by a commercial kit. (B) ROS formation was measured using a fluorescence microplate reader. Data are shown as means ± S.E.M. *P<0.05, **P<0.01 v.s. APAP treatment.
Fig 6.
Paeonol inhibited APAP-induced hepatic inflammation.
Total RNA from liver was isolated and hepatic mRNA levels of pro-inflammatory genes were determined by qPCR. GAPDH was used as the endogenous control. Data are shown as means ± S.E.M. *P <0.05, **P<0.01 v.s. APAP treatment (n = 8).
Fig 7.
Paeonol significantly inhibited IKKα/β, IκBα and p65 phosphorylation in APAP-treated liver.
The protein levels of total and phosphorylated IKKα/β, IκBα and p65 in the liver were determined by western blot. β-actin was used as the endogenous control. The representative data are shown and bands were analyzed by densitometry. Data are shown as means ± S.E.M. *P<0.05, **P<0.01 v.s. APAP treatment.