Anogeissus leiocarpus (DC.) Guill. & Perr. extract-protected rats against CCl4-induced liver fibrosis

Abstract

Objective: Anogeissus leiocarpus (DC.) Guill. & Perr. is locally used for the treatment of hepatic dysfunctions in Nigeria. This study investigated the hepatoprotective impact of A. leiocarpus stem bark hydromethanolic extract (AL) in carbon tetrachloride (CCl₄)-induced hepatic fibrosis in Wistar rats.

Methods: Thirty-five male rats were randomly assigned to seven groups. Group I (control) received distilled water and olive oil, and Group II (negative control) received 30 % CCl₄ intraperitoneally in olive oil three times a week for 8 weeks. In addition to CCl₄, rats in groups III-VI were orally given silymarin (positive control) and graded doses (125, 250, and 500 mg/kg body weight) of AL. Group VII (follow-up group) received CCl₄ only, but was spared for another 2 weeks before necropsy. After 8 weeks, the animals were necropsied, and various biochemical, histological, and immunohistochemical assessments were conducted.

Results: High-performance liquid chromatography analysis of AL detected 13 compounds, including gallic acid and castalagin. CCl₄ exposure led to a significant increase in liver index relative to the control, but the increased liver index improved following AL administration. AL elevated the antioxidant parameters, including the levels of superoxide dismutase, glutathione peroxidase, and glutathione peroxidase, with a corresponding decline in malondialdehyde levels. Furthermore, AL caused a decline in the plasma level of tumor necrosis factor alpha and interleukin 6 relative to the CCl₄ group. AL treatment moderately thinned the hepatocyte ballooning, while reducing the expression of hepatic alpha smooth muscle actin (α-SMA) and transforming growth factor beta (TGF-β) compared with CCl₄. These findings indicate that the antifibrotic action of AL might involve the downregulation of α-SMA and TGF-β expression. Furthermore, AL appreciably improved liver function indices (alanine transaminase, aspartate transaminase, and alkaline phosphatase) in a manner comparable to that of the control and CCl₄ + silymarin treatment groups.

Conclusion: Collectively, AL exhibited anti-inflammatory, hepatoprotective, and antifibrotic effects and thus has prospects for further exploration as a potential therapy in liver diseases.

Keywords: Alpha smooth muscle actin; Liver disease; Phytomedicine; Toxicology; Transforming growth factor beta (TGF-β) signaling.

Figure 1

HPLC-UV chromatogram of AL.

Figure 2

Effect of AL on the liver index in CCl₄-induced liver fibrosis in rats. Data are expressed as an average of five replicates ± the SEM. Values denoted by ‘#' indicate statistically significant differences compared to the control group, whereas values marked with ‘∗' indicate statistically significant differences relative to the CCl₄-treated group. ,∗, ∗∗, ∗∗∗∗, and #### p < 0.05.

Figure 3

Effect of AL on the plasma protein, albumin, and globulin levels in CCl₄-induced liver fibrosis in rats. (a) Plasma protein; (b) Plasma albumin; and (c) Plasma globulin. Data are expressed as an average of five replicates ± the SEM.

Figure 4

Effect of AL on liver function parameters in CCl₄-induced liver fibrosis in rats. (a) ALT; (b) AST; and (c) ALP. Data are expressed as an average of five replicates ± the SEM. Values denoted by ‘#’ indicate statistically significant differences compared to the control group, whereas values marked with ‘∗' indicate statistically significant differences relative to the CCl₄-treated group. ∗, ∗∗, ∗∗∗, ∗∗∗∗, #, and ##### p < 0.05.

Figure 5

Effect of AL on kidney function markers in CCl₄-induced liver fibrosis in rats. (a) Urea and (b) Creatinine. Data are expressed as an average of five replicates ± the SEM. Values denoted by ‘#' indicate statistically significant differences compared to the control group, whereas values marked with ‘∗' indicate statistically significant differences relative to the CCl₄-treated group. #, ∗∗, ∗∗∗, and ∗∗∗∗ p < 0.05.

Figure 6

Effect of AL on the lipid profile in CCl₄-induced liver fibrosis in rats. (a) Plasma cholesterol; (b) Plasma triglyceride; (c) Plasma HDL; and (d) Plasma LDL. Data are expressed as an average of five replicates ± the SEM.

Figure 7

Effect of AL on the hepatic redox balance and lipid peroxidation in CCl₄-induced liver fibrosis in rats. (a) Hepatic superoxide dismutase; (b) Hepatic catalase; (c) Hepatic glutathione peroxidase; (d) Hepatic glutathione; and (e) Hepatic malondialdehyde. Data are expressed as an average of five replicates ± the SEM. Values denoted by ‘#' indicate statistically significant differences compared to the control group, whereas values marked with ‘∗' indicate statistically significant differences relative to the CCl₄-treated group. #, ####, ∗, ∗∗, ∗∗∗, and ∗∗∗∗ p < 0.05.

Figure 8

Effect of AL on hepatic TNF-α and IL-6 in CCl₄-induced liver fibrosis in rats. (a) Hepatic TNF and (b) hepatic IL-6. Data are expressed as an average of five replicates ± the SEM. Values denoted by ‘#' indicate statistically significant differences compared to the control group, whereas values marked with ‘∗' indicate statistically significant differences relative to the CCl₄-treated group. ##, ####, ∗∗, ∗∗∗, and ∗∗∗∗ p < 0.05.

Figure 9

Photomicrographs showing the effect of AL on hepatic histology in CCl₄-induced liver fibrosis in rats. (40 × ; Scale bar = 50 μm).

Figure 10

Photomicrograph of immunohistochemically stained rat liver sections showing the qualitative effect of AL on the expression of α-SMA (40 × ; scale bar = 50 μm).

Figure 11

Photomicrograph of immunohistochemically stained rat liver sections showing the qualitative effect of AL on the expression of TGF-β (40 × ; scale bar = 50 μm).