| Type 2 Diabete Mellitus(T2DM)is defined as a pandemic metabolic disorder syndrome which has reached epidemic proportions worldwidly and become a serious public health concern. Insulin resistance is not only one of the main characters of T2DM but also the pathogenetic base of cardiovascular diseases and cerebrovascular diseases, so it becomes the warm spots. However, previous study showed that animal model which indicates pathogenetic feature and regularity of T2DM in human bodies really is very rare although there have been many literatures on researching the establishment of T2DM model. Therefore, it is very essential to establish the model of insulin resistance and turning into T2DM further, at the same time to clarify the main mechanism of insulin resistance and retroconversion, which will contribute to future studies on clinical diabetes pathogenesy and prevention and treatment.Our study was divided into two parts to research a new method of establishment of insulin resistance model, the relation between oxidative stress and the occurrence of insulin resistance and the effect of antioxidant on oxidative damage indexes of insulin resistance in experimental animals and correlated indexes of T2DM.1. Insulin resistance model in rat induced by high fat and high glucose diet and oxidant tBHP injection: Tert-Butyl Hydroperoxide (tBHP) plays an oxidative role by releasing hydrogen peroxide after the metabolism in vivo. A -(CH)3 group in tBHP molecular structure replace the group–OH in hydrogen peroxide which makes it more stable. tBHP, a kind of high-performance and lenteliberantes oxidant was used as injection medicine.40 Sprague–Dawley rats were randomly divided into 5 groups with 8 rats in each group: the control group (A), the double high diet group (B), the double high diet group with tBHP injection (C), the double high diet rats with tBHP injection and Vitamin E intragastric administration group (D) and the double high diet rats with tBHP injection and lipoic acid intragastric administration group (E). The control group was fed with normal diet. The double high diet group was fed with high fat and high glucose diet. The double high diet group with tBHP was fed with high fat and high glucose diet and given continual i.p. injection of 0.2mmol/l tBHP, 0.5ml/100g, everyday for a month. After one month the two medication administration teams were given intragastric administration of 30 mg/kg Vitamin E or 1 mg/kg lipoic acid everyday for two weeks. Other groups were given continual intraperitoneal injection (i.p) and intragastric administration of saline injection in the same volume. The weight, fasting blood glucose (FBG),fasting insulin (FINS),serum triglycerides (TG), serum cholesterol (TCH),serum free fatty acids (FFA) were observed at the end of four-week experiment. The calculation of insulin sensitivity index (ISI): ISI=In(1/FBG×FINS). The result indicated that we made use of high fat and high glucose diet and oxidant tBHP injection which produces a large mount of free radicals and reactive oxygen species to cause oxidative stress injury to decrease the anti-oxidative capability and increase lipid per-oxidation capability on the experimental subjects. Compared with that of group A, the weight of the B and C group changed obviously, the contents of fasting blood glucose and fasting insulin increased respectively, the contents of serum triglycerides and serum cholesterol and serum free fatty acids increased. The C group changed more obviously. Compared with that of group the contents of fasting blood glucose of group D and E decreased, fasting insulin decreased obviously, and insulin sensitivity increased. The results showed that the insulin resistance model can be established quickly and stably by high fat and high glucose diet in addition of oxidant tBHP injection and the occurrence of insulin resistance in experimental animals fed by high fat and high glucose diet was related to chronic oxidative stress closely.2. The research on pathogenic mechanism of the oxidative stress on the IR model and the effects of antioxidant intervention on oxidative damage index and T2DM correlated index: The animals were sacrificed after been fed with high fat and high glucose diet for 6 weeks. Serum and part of livers and muscles were collected for testing SOD, CAT, NOS, iNOS, MDA, GSH, GSSG, GPx activity. The livers and muscle mRNA level changes of CAT,PPARγwere observed by PCR. The changes of CAT,PPARγprotein level in livers and muscles were observed by Western Blot. Pancreas and livers were taken for pathologic evaluation. The result showed that Compared with that of control group, B and C groups could increase oxidative level, especially in livers. The activity of SOD and CAT in Serum and livers decreased, the activity of NOS and iNOS and the contents of MDA, GSSG and GPx in livers increased respectively, no obvious changes of SOD and CAT were observed in muscles. Compared with other groups, the activity of SOD and CAT in Serum, livers and muscles and the contents of GSH increased obviously, the contents of MDA, GSSG and GPx decreased, but no obvious changes of SOD and CAT were observed in muscles. The expression of CAT, PPARγmRNA in livers of group C decreased. However, that of CAT, PPARγmRNA in muscles increased significantly, the effect of E group was better than that of group D. But there was no obvious changes compared with group A and B. PPARγmRNA in muscles of group C decreased, that of group D was no obvious changes. Compared with the expression of PPARγmRNA of group B and C, group E increased. However, the expression of CAT mRNA of group C increased more obviously than that of group A. Compared with the expression of CAT mRNA of group A and B, only group D increased, no obvious changes were observed in other groups.These results indicated that the insulin resistance model can be established quickly and stably by high fat and high glucose diet in addition of oxidant tBHP injection, and it was a convenient and feasible method with high achievement ratio. And the occurrence of insulin resistance in experimental animals fed by high fat and high glucose diet was related closely to chronic oxidative stress. Hyperglycosemia can induce glycosylation of antioxidase, decrease the activity of antioxidase such as SOD, CAT, GPx, destroy the system of anti-oxidative buffering and reparation, which increased the anti-oxidative stress on bodies. After giving certain dose antioxidant intervention, it can decrease the anti-oxidative stress obviously and also clean free radicals in order to prevent from the oxidative stress injury efficiently by stimulating PPARγto up-regulate the activity of CAT. At the same time, it can decrease the contents of serum free fatty acid, inhibit formation of AGEs so as to improve insulin sensitivity. However, we cannot draw a definite conclusion that insulin resistance resulting from reduced antioxidant defences is the cause of the progression of diabetes as more direct evidence is required. And the research progress on exact pathogenic mechanism of the oxidative stress on the insulin resistance and Type 2 diabetes model is limited. So pathogenic mechanism of the oxidative stress on the insulin resistance enjoys a great potential for research and development, our results will provide important experimental evidence for studying the pathogenic mechanism on T2DM. It is also very significant for prevention and treatment on T2DM. |
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