Published online Jun 28, 2017. doi: 10.3748/wjg.v23.i24.4354
Peer-review started: November 18, 2016
First decision: March 3, 2017
Revised: March 22, 2017
Accepted: June 1, 2017
Article in press: June 1, 2017
Published online: June 28, 2017
To study the molecular mechanisms involved in the hepatoprotective effects of naringenin (NAR) on carbon tetrachloride (CCl4)-induced liver fibrosis.
Thirty-two male Wistar rats (120-150 g) were randomly divided into four groups: (1) a control group (n = 8) that received 0.7% carboxy methyl-cellulose (NAR vehicle) 1 mL/daily p.o.; (2) a CCl4 group (n = 8) that received 400 mg of CCl4/kg body weight i.p. 3 times a week for 8 wk; (3) a CCl4 + NAR (n = 8) group that received 400 mg of CCl4/kg body weight i.p. 3 times a week for 8 wk and 100 mg of NAR/kg body weight daily for 8 wk p.o.; and (4) an NAR group (n = 8) that received 100 mg of NAR/kg body weight daily for 8 wk p.o. After the experimental period, animals were sacrificed under ketamine and xylazine anesthesia. Liver damage markers such as alanine aminotransferase (ALT), alkaline phosphatase (AP), γ-glutamyl transpeptidase (γ-GTP), reduced glutathione (GSH), glycogen content, lipid peroxidation (LPO) and collagen content were measured. The enzymatic activity of glutathione peroxidase (GPx) was assessed. Liver histopathology was performed utilizing Masson’s trichrome and hematoxylin-eosin stains. Zymography assays for MMP-9 and MMP-2 were carried out. Hepatic TGF-β, α-SMA, CTGF, Col-I, MMP-13, NF-κB, IL-1, IL-10, Smad7, Smad3, pSmad3 and pJNK proteins were detected via western blot.
NAR administration prevented increases in ALT, AP, γ-GTP, and GPx enzymatic activity; depletion of GSH and glycogen; and increases in LPO and collagen produced by chronic CCl4 intoxication (P < 0.05). Liver histopathology showed a decrease in collagen deposition when rats received NAR in addition to CCl4. Although zymography assays showed that CCl4 produced an increase in MMP-9 and MMP-2 gelatinase activity; interestingly, NAR administration was associated with normal MMP-9 and MMP-2 activity (P < 0.05). The anti-inflammatory, antinecrotic and antifibrotic effects of NAR may be attributed to its ability to prevent NF-κB activation and the subsequent production of IL-1 and IL-10 (P < 0.05). NAR completely prevented the increase in TGF-β, α-SMA, CTGF, Col-1, and MMP-13 proteins compared with the CCl4-treated group (P < 0.05). NAR prevented Smad3 phosphorylation in the linker region by JNK since this flavonoid blocked this kinase (P < 0.05).
NAR prevents CCl4 induced liver inflammation, necrosis and fibrosis, due to its antioxidant capacity as a free radical inhibitor and by inhibiting the NF-κB, TGF-β-Smad3 and JNK-Smad3 pathways.
Core tip: To study the effect of naringenin (NAR) on carbon tetrachloride (CCl4)-induced chronic liver fibrosis, male Wistar rats were administered 400 mg of CCl4/kg body weight and 100 mg of NAR/kg body weight for 8 wk. NAR prevented necrosis, cholestasis, oxidative stress, collagen accumulation and the increase in MMP activity caused by CCl4 administration. NAR completely prevented the increase in TGF-β, α-SMA, CTGF, Col-1, MMP-13, NF-κB, IL-1 and IL-10 protein levels caused by CCl4 administration and pSmad3 and pJNK activation. In conclusion, NAR prevents CCl4 induced liver fibrosis due to its antioxidant capacity and by inhibiting the TGF-β-Smad3, JNK-Smad3 and NF-κB pathways.