Amelioration of CCl4-induced oxidative stress and hepatotoxicity by Ganoderma lucidum in long evans rats

Abstract

Liver disease is a serious health problem affecting people worldwide at an alarming rate. The present study aimed to investigate the protective effects of Ganoderma lucidum against CCl₄-induced liver toxicity in rats. The experimental Long Evans rats were divided into five groups, of which four groups were treated with carbon tetrachloride (CCl₄). Among the CCl₄ treated groups, one of the groups was treated with silymarin and two of them with ethanolic extract of G. lucidum at 100 and 200 mg/Kg body weight. The oxidative stress parameters and endogenous antioxidant enzyme concentrations were assessed by biochemical tests. Liver enzymes ALT, AST, and ALP were determined spectrophotometrically. Histopathological examinations were carried out to assess hepatic tissue damage and fibrosis. Reverse transcription PCR (RT-PCR) was performed to determine the expression of IL-1β, IL-6, IL-10, TNF-α, and TGF-β genes. Gas Chromatography-Mass Spectroscopy (GC-MS) analysis revealed that G. lucidum is rich in several phytochemicals including 6-Octadecanoic acid (55.81%), l-( +)-Ascorbic acid 2,6-dihexadecanoate (18.72%), Cis-11-Eicosenamide (5.76%), and Octadecanoic acid (5.26%). Treatment with the G. lucidum extract reduced the elevated ALT, AST, ALP levels, and cellular oxidative stress markers and increased the endogenous antioxidant levels. Histopathology observations revealed that the inflammation, infiltration of immune cells, and aberration of collagen fibers in the hepatocytes were altered by the G. lucidum treatment. The increased expression of inflammatory cytokines TNF-α, TGF-β, IL-1 β, and IL-6 were markedly suppressed by G. lucidum extract treatment. G. lucidum also prevented the suppression of protective IL-10 expression by CCl₄. This study strongly suggests that G. lucidum extract possesses significant hepatoprotective activity as evidenced by reduced oxidative stress and inflammation mediated by suppression in inflammatory cytokine expression and increased protective IL-10 cytokine expression.

Figure 1

GC chromatogram of the ethanolic extract of G. lucidum powder.

Figure 2

Effect of G. lucidum extract supplementation on the body weight (a) and the organ wet weight (b–d) in different groups. Statistical analysis was carried out by one‐way ANOVA followed by Newman–Keul’s post hoc test. Statistical significance was considered at *p ≤ 0.05 and **p ≤ 0.01, ns not significant. HD high dose (200 mg/kg), LD low dose (100 mg/kg).

Figure 3

Effect of G. lucidum extract supplementation on ALP (a), ALT (b), and AST (c) enzyme activities in plasma of CCl₄ administered rats. Statistical analysis was carried out by one‐way ANOVA followed by Newman–Keul’s post hoc test. Statistical significance was considered at *p ≤ 0.05 and **p ≤ 0.01, ns not significant. HD high dose (200 mg/kg), LD low dose (100 mg/kg).

Figure 4

Effect G. lucidum powder extract on MDA (a, b), NO (c, d), and AOPP (e, f) activities in liver and plasma of CCl₄ administered rats. Statistical significance was considered at *p ≤ 0.05, **p ≤ 0.01. HD high dose (200 mg/kg), LD low dose (100 mg/kg).

Figure 5

Effect of G. lucidum extract supplementation on SOD (a, b), GSH (c, d) and CAT (e, f) activities in liver and plasma of CCl₄ administered rats. Statistical significance was considered at *p ≤ 0.05, **p ≤ 0.01. HD high dose (200 mg/kg), LD low dose (100 mg/kg).

Figure 6

Histopathological observations showing the effects of CCl₄-induced hepatotoxicity and the hepatoprotective effect of G. lucidum extract (Low dose: 100 mg/kg and High dose: 200 mg/kg). The upper panel (a–e) shows hematoxylin/eosin staining and the lower panel (f–j) shows Sirius Red staining. Arrowheads indicate infiltrating cells (b) and arrows indicate collagen deposition (g, h).

Figure 7

Expression of a housekeeping gene (GAPDH) and cytokine genes (IL-1β, IL-6, Il-10, TNF-α, TGF-β) in liver tissues collected from different groups.

Figure 8

The inclusion and exclusion of the test animals and the experimentation groups.