Silymarin protects against acute ethanol-induced hepatotoxicity in mice

Abstract

Background: Accumulated evidence has demonstrated that both oxidative stress and abnormal cytokine production, especially tumor necrosis factor-alpha (TNF), play important etiological roles in the pathogenesis of alcoholic liver disease (ALD). Agents that have both antioxidant and anti-inflammation properties, particularly anti-TNF production, represent promising therapeutic interventions for ALD. We investigated the effects and the possible mechanism(s) of silymarin on liver injury induced by acute ethanol (EtOH) administration.

Methods: Nine-week-old mice were divided into 4 groups, control, silymarin treatment, EtOH treatment, and silymarin/EtOH treatment, with 6 mice in each group. Because control and silymarin values were virtually identical, only control treatment is shown for ease of viewing. Ethanol-treated mice received EtOH [5 g/kg body weight (BW)] by gavage every 12 hours for a total of 3 doses. Control mice received an isocalorical maltose solution. In the silymarin/EtOH group, silymarin was dissolved in the EtOH and gavaged simultaneously with EtOH at a dose of 200 mg/kg BW. At 4 hours after the last dosing, the mice were anesthetized and subsequent serum alanine aminotransferase (ALT) level, hepatic lipid peroxidation, enzymatic activity of hepatic cytochrome P450 2E1, hepatic TNF-alpha, and glutathione (GSH) levels were measured. Histopathological change was assessed by hematoxylin and eosin staining.

Results: Acute EtOH administration caused prominent hepatic microvesicular steatosis with mild necrosis and an elevation of serum ALT activity, induced a significant decrease in hepatic GSH in conjunction with enhanced lipid peroxidation, and increased hepatic TNF production. Supplementation with a standardized silymarin attenuated these adverse changes induced by acute EtOH administration.

Conclusions: Silymarin protects against the liver injury caused by acute EtOH administration. In view of its nontoxic nature, it may be developed as an effective therapeutic agent for alcohol-induced liver disease by its antioxidative stress and anti-inflammatory features.

Fig. 1

High-performance liquid chromatography fingerprint of standarized silymarin.

Fig. 2

Serum alanine aminotransferase (ALT) activity in different groups. N = 6 for each group. Values are means ± SD. Values in bars that do not share a letter differ (p < 0.05).

Fig. 3

Histopathological changes in the liver of different groups. (A) Control mice. (B) Ethanol (EtOH) treatment induced prominent microvesicular steatosis (arrows) along with necrosis (arrowheads) in the liver. The necrotic hepatocytes are characterized by cell enlargement and nuclear dissolution. (C) Livers from silymarin supplemental group showed only microvesicular steatosis, which was less extensive than in livers from mice receiving EtOH alone. CV, central vein. ×40.

Fig. 4

Hepatic triglyceride content in different groups. N = 6 for each group. Values are means ± SD. Values in bars that do not share a letter differ (p < 0.05).

Fig. 5

Hepatic lipid peroxidation (TBARS content) in different groups. N = 6 for each group. Values are means ± SD. Values in bars that do not share a letter differ (p < 0.05).

Fig. 6

Heptic glutathione (GSH) concentrations in different groups. N = 6 for each group. Values are means ± SD. Values in bars that do not share a letter differ (p < 0.05).

Fig. 7

Changes of cytochrome P450 2E1 (CYP2E1) enzymatic activity in different groups. N = 6 for each group. The CYP2E1 activity in control is 1.643 ± 0.476 µmol · min/mg protein. Values are means ± SD. Values in bars that do not share a letter differ (p < 0.05).

Fig. 8

Hepatic tumor necrosis factor (TNF) levels in different groups. N = 6 for each group. Values are means ± SD. Values in bars that do not share a letter differ (p < 0.05).