Establishment of an Animal Model of Thioacetamide(TAA)-Induced Liver Fibrosis and Investigation of the Molecular Mechanism of Silymarin in Liver Fibrosis Prevention

Liver fibrosis and cirrhosis are chronic and progressive liver diseases that most often caused by chronic liver injuries, including alcoholism, hepatitis infection and other illnesses. Damage to the liver causes the development of scar tissue in the healthy liver tissue. If the underlying cause of liver fibrosis or cirrhosis continues, liver fibrosis and cirrhosis will progress and liver function will diminish. Cirrhosis is irreversible, but it is demonstrated liver fibrosis in animals and humans can be restored when collagen synthesis is inhibited. For this reason, the aim of this study is to establish an animal model of liver fibrosis and then find out the therapy pathway to cure liver fibrosis.　The liver damage and fibrosis of animals were induced by intraperitoneal administration of thioacetamide (TAA), associated with increased oxidative stress and activation of hepatic stellate cells. Male ICR mice, BALB/c mice and Wistar rats were used to test which species may be more suitable to develop liver fibrosis. Preliminary results showed that intraperitoneal administration of 100 mg/kg bw TAA dosage would cause obvious liver fibrosis according to Masson’s Trichrome staining and increase liver weight (1.3 - 1.5 times), serum AST (1.6 - 2.4 times) and ALT (3 - 4 times) levels compared to those without TAA. This animal model successfully induced liver damage and fibrosis. Male and female ICR mice were also compared in this study for testing the gender differences on the development of iver fibrosis.　In order to compare the therapy ability on liver fibrosis, curcumin, resveratrol and silymarin were used to evaluate the beneficial effects in this animal model. These phytochemicals were all reporterd with anti-inflammatory, anti-oxidant and anti-cancer properties. Our results showed that curcumin, resveratrol and silymarin all had beneficial effects on liver injury and fibrosis. Besides, silymarin had the most beneficial effects compared to curcumin and resveratrol, and it normalize liver weight, serum AST and ALT levels to those of animals without TAA treatments.　In the late stage of liver fibrosis in this animal model, the mRNA levels of NF-κB, activator protein-1 (AP-1) and Kruppel-like factor 6 (KLF6) were detected by semi-quantitative RT-PCR. It showed that TAA treatments increased the expression of these 3 transcription factors. Taking of silymarin decreased the mRNA expression of AP-1 and KLF6, but not NF-κB, where KLF-6 would up-regulate the expression of TGF-β1 in hepatocyte. TGF-β1 then stimulated the activation of hepatic stellae cell and then caused extra cellular matrix accumulation in the liver. These results showed that silymarin may prevent liver fibrosis and reduce KLF6 pathway in mRNA levels, and then inhibit the activation of hepatic stellate cell and minimize hepatic fibrosis.　In conclusion, the animal models of liver fibrosis were successfully established in ICR mice, BALB/c mice and Wistar rats. These models were helpful for further studies to test whether the health food have a hepatic protection or not. Curcumin, resveratrol and silymarin all had the beneficial effects in mouse model of liver fibrosis. Silymarin can prevent liver fibrosis and reduce KLF6 pathway. So far, this is the first finding in silymarin associated with KLF6 pathway, but the detailed pathway needs a further study to clarify.