| Objective:Non-alcoholic fatty liver disease (NAFLD) has become the most common liver disease, and the incidence rate is rising year by year. Unfortunately, the pathogenesis of the development from simple hepatic steatosis to steatohepatitis is incompletely understood. However the oxidative stress (OS) caused by reactive oxygen species has been considered as the major importance of the developing process of NAFLD. Our study aims to high-fat diet induced NAFLD rat model, then observe OS and the expression of matrix metalloproteinase in NAFLD rats, in order to explore the effect of probucol and to explore the role of matrix metalloproteinase in NAFLD rats, and the possible pathophysiological mechanisms.Methods:Forty health male SD rats were randomly divided into two groups:normal diet group (NG, n=16), which were common fed; high-fat diet group (HG, n=24), which were fed by high-fat diet. At the end of8th week,8rats were randomly selected from each group to identify the NAFLD model. These killed rats were named NG8W group and HG8W group individually. The HG was devided into two groups at the beginning of9th week:14-week high lipid group model group (HG14W) and probucol intervention group (PB group). These rats were continued fed by high-fat diet. PB group was given probucol gavage (500mg/d.Kg) while NG14W and HG14W suffered volumetric0.5%sodium carboxymethyl cellulose solution lavage. Rats in each group were sacrificed6weeks later than gavage, and determined body weight, liver index, triglycerides (TG), cholesterol (TC), alanine aminotransferase (ALT), aspartate aminotransferase (AST), tumor necrosis factor-alpha (TNF-α), interleukin prime6(IL-6). We also tested the lipoperoxidation makers and anti-oxidant substance in liver, such as MDA, SOD, GSH-Px. Expression of MMP-2and MMP-9in the liver were detected by immunohistochemistry. Conventional preparation of HE staining for liver histopathology. The pathological changes of liver were observed by light microscopy.Result:(1) Compared with the corresponding period of NG, the weight, liver index, levels of serum ALT, AST, TG, TC, TNF-α, IL-6and levels of liver homogenate MDA of HG8W and HG14W were higher, and GSH-Px and SOD decreased significantly, these differences were statistically significant (p<0.01). Under the light microscope, we can found vacuolar-like steatosis accompanied with inflammatory in HG8W rat liver, while severe fatty degeneration with massive infiltration of inflammatory cells and collagen fibers, coarse fiber spacing were obeserved in the HG14W liver cells.(2) Compared with HG14W rats, the liver index, levels of serum ALT, AST, TG, TC and levels of liver homogenate MDA in PB group were lower, while the GSH-Px and SOD significantly higher, these differences were statistically significant (p<0.01). When observing the liver tissue of PB group under light microscopy, the liver lobules and borders still exist, and the degree of steatosis and inflammatory cell infiltration were improved.(3) Immunohistochemistry of MMP-2and MMP-9showed that:Under lightscopy, there was little MMP-2and MMP-9positive liver cells in NG, while HG8W, HG14W have strong expression of MMP-2, MMP-9, whose performance of the cytoplasm and cell membrane staining was brown, tan, positive granules in different thickness and obviously coloring. Immunohistochemical score were statistically significant (p<0.01) when compared to NG. Expression of MMP-2, MMP-9in PB group is weaker than HG14W, with less positive granular coloring degree and positive cells, but still higher than NG14W, there were statistics differences (p<0.01).Conclusion:1. successfully constructed NAFLD model by high fat diet induced for8W, while the NASH model was constructed by high fat diet induced for14W.2. The NAFLD model showed hyperlipidemia, elevated serum transaminases, oxidative stress, and lipid peroxidation. All the above aggravated as time goes by.3. Formation and progression of NAFLD is associated with OS, lipid peroxidation, MMP-2, MMP-9.4. Probucol is an effective medicine for NAFLD treatment. The possible mechanism might be lipid-lowering, anti-inflammatory, and antioxidant effect. |
