Antifibrogenic effect of melatonin in rats with experimental liver cirrhosis induced by carbon tetrachloride

Abstract

Background and aim: Liver diseases are a major public health problem, accounting for a significant number of hospital visits and admissions and an increasing mortality rate. Melatonin (MLT) is a powerful antioxidant molecule that has been shown to be beneficial under various conditions. The objective was to evaluate the effect of MLT on experimental liver cirrhosis induced by carbon tetrachloride (CCl₄) in rats.

Methods: Twenty male Wistar rats (230-250 g) were divided into four groups. I: control group (CO); II: CO + MLT; III: CCl₄; and IV: CCl₄ + MLT. CCl₄ was administered intraperitoneally (i.p.) as follows: 10 doses every 5 days, 10 doses every 4 days, and 7 doses every 3 days. MLT was administered i.p. at a dose of 20 mg/kg from the 10th week to the end of the experiment (16th week).

Results: In the CCl₄ + MLT group, we found that MLT caused a decrease in the level of F2-isoprostanes and NQO1 expression. We also found that MLT reduced the inflammatory process as shown by decreased expressions of NF-KB/p65 and inducible nitric oxide synthase (iNOS) and a smaller amount of inflammatory infiltrate. MLT reduced the expression of transforming growth factor beta1 (TGF-β1), alpha-smooth muscle actin (α-SMA), and vascular endothelial growth factor (VEGF). Picrosirius staining showed that MLT decreases fibrosis.

Conclusion: MLT has a potent antifibrogenic effect, modulating the parameters of oxidative stress, angiogenesis, and inflammation.

Keywords: antioxidants; melatonin and liver cirrhosis; oxidative damage.

Figure 1

Level of F2‐isoprostanes in rat liver. Results are expressed as mean ± SD. *P < 0.05 carbon tetrachloride (CCl₄) versus other groups.

Figure 2

Western blot analysis of NQO1 and vascular endothelial growth factor (VEGF). (a) Cytoplasmic fractions were analyzed by WB with NAD(P)H:quinone oxidoreductase 1 (NQO1) and VEGF and glyceraldehyde phosphate dehydrogenase (GAPDH) antibodies. (b) Arbitrary values expressed as mean and SD. *P < 0.05 carbon tetrachloride (CCl₄) versus other groups. ^# P < 0.05 CCl₄ + melatonin (MLT) versus other groups. CO, CO + MLT, CCl₄, CCl₄ + MLT.

Figure 3

Histological analysis of liver sections by hematoxylin and eosin (10×), masson trichrome (10×), and picrosirius staining (10×).

Figure 4

Western blot analysis of p65 and inducible nitric oxide synthase (iNOS). (a) Nuclear fractions were analyzed by WB with NF‐KB/p65 and cytoplasmic iNOS and β‐actin antibodies. (b) Arbitrary values expressed as mean and SD. *P < 0.05 carbon tetrachloride (CCl₄) versus other groups. CO, CO + MLT, CCl₄, CCl₄ + MLT.

Figure 5

Western blot analysis of transforming growth factor beta (TGF‐β) and alpha‐smooth muscle actin (α‐SMA). (a) Cytoplasmic fractions were analyzed by WB with TGF‐β and α‐SMA and glyceraldehyde phosphate dehydrogenase (GAPDH) antibodies. (b) Arbitrary values expressed as mean and SD. *P < 0.05 carbon tetrachloride (CCl₄) versus other groups. ^# P < 0.05 CCl₄ + melatonin (MLT) versus other groups. CO, CO + MLT, CCl₄, CCl₄ + MLT.