Emodin Improve Kkay Mechanism Study Of Insulin Resistance In Diabetic Mice

Purpose:After emodin was given to the type2diabetic KKAy mice for8weeks, we use biochemical method, RT-PCR method, immunohistocemistry method and Western-blot method to test the mechanism of how can emodin improve KKAy diabetes mouse liver, muscle and adipose ti ssue insulin sensitivity and lowering blood sugar. And we study how can emodin affect PI3K/AKt insulin signal transduction pathway. In order to provide the experimental basis of emodin in treatment of diabetes mellitus.Materials and Methods:32female KKAy mice of Specific pathogen free (SPF) were divided randomly into four groups:model(DM) group, emodin-low dose group(EL), emodin-high dose group (EH), and pioglitazone group (PI) according to plasma glucose level. We selected10C57BL/6J mice as normal control group(NC)8weeks later, all animals were tested fasting plasma glucose, fasting insulin level for caculating Insulin Sensitivity Index, total cholesterol, total triacylglycerol, high-density lipoprotein cholesterol, low-density lipoprotein cholesterol, floating fatty acid, high sensitive creaetor protein, tumor necrosis factor-α. The expression of PPARγ and GluT-2in hepatic tissue were determined by immunohistochemistry and RT-PCR. The expression of PPARγ and GluT-4in musle and adipose tissue were determined by immunohistochemistry and RT-PCR. The expressions of IRS-1, PI3K, AktSer473and Fox01in hepatic tissue, musle tissue and adipose tissue were determined by Western-blot. The experimental results were analyzed by statistical softwareResults:8weeks later, the experiment was over. The mice were fasting for8hours, and then we used the serum to test biochemical datas. The mice were put to death, we take out partial liver, musle and adipose speedly into the liquid nitrogen for RT-PCR and Western-blot.The other partial liver,musle and adipose wre fixed by paraformaldehyde. Paraffin embedding for Immunohistochemistry.1. Compared with normal control group, DM group showed higher fasting plasma glucose, total cholesterol, total triacylglycerol, low-density lipoprotein cholesterol, floating fatty acid, high sensitive creaetor protein, and tumor necrosis factor-α (P<0.05), lower ISI(P<0.05). Compared with DM group, emodin could reduce fasting plasma glucose, total cholesterol, total triacylglycerol, low-density lipoprotein cholesterol, floating fatty acid, high sensitive creaetor protein, and tumor necrosis factor-α (P<0.05). In addition, emodin could improve insulin sensitivity in KKAy diabetic mice(P<0.05). emodin showed dose dependent.2. By RT-PCR products and gel electrophoresis results of the observation optical density scanning system integration analysis, the expression of PPARγmRNA and GluT-2mRNA in hepatic tissue of DM group was decreased as compared with NC group (p<0.05). The expression of PPAR y mRNA and GluT-4mRNA in musle tissue and adipose tissue of DM group was decreased too as compared with NC group (p<0.05). Compared with DM group, the expression of PPARγmRNA and GluT-2mRNA in hepatic tissue of EL、EH、PI group was up-regulated (p<0.05) the expression of PPARγmRNA and GluT-4mRNA in musle tissue and adipose tissue of EL、EH、PI group was also up-regulated(p<0.05). Immunohistochemical results showed lower protein expression of PPARγ and GluT-2in hepatic tissue of DM group as compared with normal control group (p<0.05), and showed lower protein expression of PPARγ and GluT-4in musle tissue and adipose tissue of DM group as compared with normal control group (p<0.05). Compared with DM group, the protein expression of PPAR Y and GluT-2in hepatic tissue of EL、EH、PI group was up-regulated (p<0.05); the protein expression of PPARγand GluT-4in musle tissue and adipose tissue of EL、EH、PI group was also up-regulated (p <0.05).3.By the results of Western blot analysis of absorbance points, the expression of IRS-1、PI3K、AktSer473in hepatic tissue, musle tissue and adipose tissue of DM group was decreased; the protein expression of Fox01was up-regulated as compared with NC group (p<0.05). Compared with DM group, the protein expression of IRS-1、PI3K、AktSer473in hepatic tissue, musle tissue and adipose tissue of EL、EHgroup was up-regulated (p<0.05); the protein expression of Fox01was decreased (p<0.05).Conclusion:1.10weeks old KKAy mice have appeared high blood sugar. After8weeks high-fat diet feeding, KKAy mice existed typical insulin resistance. Fasting plasma Glucose, fasting insulin level, total cholesterol, total triacylglycerol, low-density lipoprotein cholesterol, floating fatty acid, high sensitive creaetor protein, tumor necrosis factor-α. increased significantly and the blood insulin sensitivity index. decreased significantly.2. emodin can decrease fasting plasma glucose, fasting insulin level, total cholesterol, total triacylglycerol,, low-density lipoprotein cholesterol, floating fatty acid, high sensitive creaetor protein, tumor necrosis factor-α and increase insulin sensitivity.The role of high dose emodin outstripped low dose.3. In model group mice, the expression of PPAR γ mRNA and GluT-2mRNA in hepatic tissue are decreased obviously.. Emodin can up-regulate the expression of PPAR γ mRNA and GluT-2mRNA in hepatic tissue of KKAy mice. Emodin can improve liver glucose utilization of inadequate and excessive release and enhance insulin sensitivity and improve insulin resistance.4. In model group mice, the expression of PPARγ mRNA and GluT-4mRNA in skeletal muscle and adipose tissue are decreased obviously.Emodin can up-regulate the expression of PPARγ mRNA and GluT-4mRNA in skeletal muscle and adipose tissue of KKAy mice In order to promote skeletal muscle and adipose tissue uptake of glucose and enhance insulin sensitivity and improve insulin resistance.5. In model group mice, the protein expression of IRS-1, PI3K and Akt phosphorylation of Ser473in hepatic, skeletal muscle and adipose tissue are decreased obviously. Emodin can up-regulate the protein expression of IRS-1, PI3K and Akt phosphorylation of Ser473and down-regulate the protein expression of Fox01in hepatic, skeletal muscle and adipose tissue of KKAy mice. Emodin can improve insulin resistance from PI3K insulin signal transduction pathway.