The Effect And Mechanism Of Probucol Treatment On High-fat Diet Induced IR Rat Research

Objective:Establish rat obesity-induced insulin resistance model with high-fat diet. To study the effects of insulin resistance(IR), c-Jun N-terminal kinase-1(JNK1), inflammatory reaction and oxidative stress on the pathogenesis of IR. To observe the effects of probucol on IR and the expression of JNK1in liver. To explore the mechanisms of probucol on IR.Method:forty male SD rats were randomly grouped into normal diet group (NG, n=16), which were fed normal diet, and high-fat diet group (HG, n=24), which were fed high-fat diet (normal diet adding1%cholesterol and14%lard). After8weeks,8rats were randomly selected from each group to identify the IR model, and these rats were labeled NG8W and HG8W. started from week9, all rats of HG were randomly grouped into two groups:HG14W (n=8) and probucol group (PG, n=8), these rats were continued on high-fat diet fed. PB rats were administered probucol500mg/(Kg.d)(5%turbid liquid of probucol made with0.5%sodium carboxymethylcellulose) for6weeks, and while the NG14W and HG14W rats were administered5%sodium carboxymethylcellulose lml/100g weight for6weeks. After6weeks, the rats were fasted for12h hours and weighed before sacrificed. The level of serum alanine aminotransferase (ALT), aspartate aminotransferase (AST), triglyceride (TG), total cholesterol (TC), low density lipoprotein (LDL), fasting blood glucose (FBS), fasting insulin (FINS), free fatty acids (FFAs), tumor necrosis factor-a (TNF-α), and adiponectin (Adp) were measured, then HOMA-IR was calculated. The liver was dissociated and weighed, then Liver index was calculated. The liver tissues were prepared into10%liver homogenate for measuring the levels of liver MDA and SOD. The expression of JNK1in liver tissue was determined using the immunohistochemistry method.Result:(1) The body weight, liver index, serum level of ALT, AST, TG, TC, LDL, FINS, FFAs, TNF-a, and HOMA-IR, as well as the liver tissue MDA level of HG8W and HG14W were respectively higher than NG8W and NG14W; meanwhile, the serum level of Adp, and the liver tissue SOD level were lower. There were statistical differences between these groups (p<0.05). There was no statistic significant for serum level of FBS between these groups.(2) Compared to HG14W rats, the liver index,the weight,liver index,levels of serum ALT, AST, TG, TC, LDL, FINS, FFAs, TNF-α, HOMA-IR and levels of liver homogenate MDA of PB were lower and levels of serum Adp,liver homogenate SOD were higher, and there were significant statistics differences (p<0.05). Except FINS, Adp and MDA level, all other indexes showed statistic significant between PB and NG14W.(3) The immunohistochemistry study showed that liver cells with JNK1-positive were brown stained under the light microscope. NG had no significant JNKl-positive cells. Compared to the corresponding period of NG, HG have a strong expression of JNKl which statistically significant (p<0.001). The positive particles were mainly located in the nucleus and cytoplasm of liver cell, disclosed as diffuse brown granular.Expression of JNK1-positive cell in PB is weaker compare with HG14W with statistically significant (p<0.001), disclosed as reduced grade of staining and amount of JNK1-positive cells. Nevertheless, the expression of JNK1-positive cells in PB is still stronger than NG14W and there were statistics differences (p<0.001).(4) HOMA-IR had positive correlation with the expression of JNK1in liver. The expression of JNK1in liver had positive correlation with serum level of TNF-a, FFAs, and liver tissue homogenate MDA and negative correlation with serum level of Adp、 liver homogenate SOD. TNF-a and FFAs had positive correlation, but they both had negative correlation with Adp. TNF-a had positive correlation with liver homogenate MDA and negative correlation with liver homogenate SOD.Conclusion:1. The SD rat IR model was successfully established by feeding high-fat diet for8weeks. This model metabolic characteristic was coupled with hyperlipidemia, and increment of serum transaminase.2. The high-fat diet could induce lipid metabolism abnormality, increased the level of TNF-α and FFAs, lead to oxidative stress and lipid peroxidation. All these factors play an interaction in the JNK signal pathway by activate the expression of JNKl protein and reduced insulin sensitivity, and thus causing IR.3. Administered Probucol for6weeks can improve IR in high-fat diet-induced IR rat; the mechanisms involved probably through the lipid metabolism improvement, inflammatory reaction reduction, anti-oxidative stress and reduced of liver JNK-1expression.