Study On Protection Of Mulberry (Morus Alba Var. Multicaulis (Perrott.) Loud) Leaf Polysaccharide MLP On Oxidative Stress On Pancreas Islet β Cells Of Diabetic Rat And Its Mechanism

Diabetes Mellitus(DM), caused by the joint action of genetic and environmental factors, has become the third largest disease around the world after cardiovascular disease and cancer, and type 2 diabetes accounts for 85-95%. Studies have shown that oxidative stress is the important reason for the occurrence and development of diabetes. Oxidative stress reflects the imbalance between the production of reactive oxygen species(ROS) and reactive nitrogen species(RNS) and the biological system’s ability to readily detoxify the reactive intermediates and resulting the damage of proteins and nucleic acids of organism. Oxidative stress not only can disturb the transmission of insulin signal, induce insulin resistance, but also injure the islet β cells, result the decrease of insulin secret and β cells apoptosis. Therefore, it is an important approach to cure diabetes that decrease the oxidative stress of body.Mulberry(Morus alba L., family of Moraceae) leaf can be used not only as silkworm feed, but also in traditional Chinese medicine to treat diabetes certificate. “Compendium of Materia Medica” records: mulberry fried juice and moxibustion replace of tea can prevent thirst quencher. Modern pharmacological studies have shown that mulberry leaf polysaccharide, which is one of the bioactive components in the mulberry leaf, has significant hypoglycemic effects. In early studies, we extracted and purified a homogeneous polysaccharide constituent named MLP, and proved it possess the function of improving glucose metabolism and insulin signaling and promoting insulin secretion. Furthermore, mulberry leaf polysaccharide MLP has a significant effect of antioxidant, but it is uncertainty about the mechanism underlying the antioxidant effect.In view of this, the present study deeply investigated the protective effect of MLP on pancreas islet β cells of rat and its mechanism in vivo and in vitro. The main results are as follows:(1) Effects of mulberry leaf polysaccharide MLP on oxidative stress in type 2 diabetic rats. MLP therapeutic intervention in type 2 diabetic rats were measured by MLP on the impacts of biomarkers of oxidative stress in serum and tissues, the morphology and function of tissues and the activity of anti-oxidase in tissues, in order to investigate the protective effect of MLP on oxidative stress in type 2 diabetic rats. The results showed that MLP could effectively decrease the content of 8-Ohd G and MDA in serum, pancreas, liver and muscle, improve the morphology of pancreas, liver and muscle, promote insulin secretion, enhance the activity of GS in liver and promote the translocation of GLUT4 in muscle. Furthermore, MLP could significantly improve the activity of SOD, CAT and GPx in pancreas, liver and muscle of the type 2 diabetic rats. Moreover, the results also shown that the protective effect of MLP on pancreas is the most obvious in type 2 diabetic rats.(2) The protective effect of MLP on oxidative stress in INS-1 cells cultured with high glucose in vitro. MLP therapeutic intervention in INS-1 cells cultured with high glucose were measured by MLP on the impacts of biomarkers of oxidative stress in cells, the morphology of cells, the activity of anti-oxidase and the biomarkers of oxidative and activity of antioxidase in mitochondrial of cells, in order to investigate the protective effect of MLP on oxidative stress in INS-1 cells cultured with high glucose. The results shown that MLP could effectively decrease the content of 8-Ohd G and MDA in cells, improve the morphology of cells, the activity of SOD, CAT and GPx in cells. Furthermore, the content of 8-Ohd G and MDA in mitochondrial was significantly decreased, the activity of SOD, CAT and GPx in mitochondrial was obviously increased.(3) The studies on the mechanism of improvement effect on energy metabolism of MLP on INS-1 cells cultured with high glucose in vitro. Immunofluorescence staining, western blotting and RT-PCR were used to detect the effect of MLP on mitochondrial energy metabolism-related factors of INS-1 cells cultured with high glucose. The results shown that MLP could effectively increase the activity of mitochondrial electron transport chain complex Ⅰ-Ⅳ, decrease the protein and m RNA expression levels of UCP-2, and increase the synthesis of ATP in cells.(4) The studies on the mechanism of suppression on cell apoptosis of MLP on INS-1 cells cultured with high glucose in vitro. Flow cytometry, immunofluorescence staining, western blotting and RT-PCR were used to detect the effect of MLP on cell apoptosis-related factors of INS-1 cells cultured with high glucose. The results shown that MLP could effectively increase mitochondrial membrane potential, decrease cytochrome C(Cyt-C) released from the mitochondria to the cytoplasm, thereby inhibit cell apoptosis.(5) The studies on the mechanism of promotion on insulin secretion of MLP on INS-1 cells cultured with high glucose in vitro. Immunofluorescence staining, western blotting and RT-PCR were used to detect the effect of MLP on insulin secretion-related factors of INS-1 cells cultured with high glucose. The results shown that MLP could effectively increase the expression of PDX-1 in the nucleus and the expression of insulin.This paper systematically studied the protective effect of MLP on oxidative stress in pancreas islet β cells of rat, and its specific mechanism has also been discussed in depth from organization, protein and gene levels. It has provided a theoretical basis for the further development of the MLPⅡthat used in the prevention of type 2 diabetes.