Effect Of Glucose Fluctuation On Endothelium-dependent Vasodilation In Diabetes And Its Mechanism

Objective:To establish the blood glucose fluctuation diabetic rat model by subcutaneous injecting insulin.Method:75 Male Sprague-Dawley(SD) rats were randomly assigened into two groups:normal control group rats (n=20) and model group rats (n=55). NC rats were fed with normal diet; model group rats were fed with high-sucrose-high-fat diet.After 4 weeks a low dose of Streptozotocin (STZ,35mg/Kg) was used to induce hyperglycemia. After that, the model group rats were randomly subdivided into two groups again:sustained high blood glucose group and blood glucose fluctuation group. The latter group were induced by Subcutaneous injecting insulin twice daily,while other group were given equal dose normal distilled water. We observed survival rate, body weight, water intake, urine output, and detected glucose, serum insulin, renal function,24 hours proteinuria and lipid.All rats were sacrificed after 3 months intervention.Results:1. Compared with NC group, survival rate and body weight were significantly lower, but of water intake and urine output were obviously higher in sustained high blood glucose group and blood glucose fluctuation (P<0.05). There were no significant difference between sustained high blood glucose group and blood glucose fluctuation group (P>0.05).2. Compared with normal control group, fasting glucose and insulin were increased, insulin sensitive index was decresed in diabetic group (P<0.05). 3. The average blood glucose levels (MBG), the standard deviation of daily average blood glucose (SDBG), the largest amplitude of glycemic excursions (LAGE) and M-value were significantly increased in sustained high blood glucose group and blood glucose fluctuation group.4. The 24 hours proteinuria, BUN and SCr of blood glucose fluctuation group were significantly higher than sustained high blood glucose group (P<0.05).5. Compared with NC group, HbA1c, TC, TG and LDL were significantly increased in sustained high blood glucose group and blood glucose fluctuation group (P<0.05), HDL was significantly decreased in sustained high blood glucose group and blood glucose fluctuation group (P<0.05). And there was no significant difference between both diabetic groups (P>0.05).Conclusions:The blood glucose fluctuation diabetic rat model can be established by twice-daily subcutaneous insulin injection method in Diabetic rats.Objective:To investigate the effects of blood glucose fluctuation on endothelium-dependent vasodilation function in diabetic rats and its mechanism.Method:Acetylcholine(Ach)-induced vasodilation response in isolated aortic rings of normal control group,sustained high blood glucose group and blood glucose fluctuation group rats were observed and aortic NO concentration were measured using Griess Reaction. eNOS activation were detected by immunohistochemical SP method. mRNA expression of PI3K, Akt and eNOS of aorta were measured by RT-PCR and the protein level were measured by Western blot.Result:1. Compared with NC group,endothelium-dependent vasodilation function and NO production were significantly decreased in blood glucose fluctuation group and sustained high blood glucose (P<0.05).2. Pathologic changes of aortic ultrastructure can be found in fluctuant high blood glucose and sustained high blood glucose.3. Compared with NC group, immunohistochemical positive expression of eNOS was decreased in blood glucose fluctuation group and sustained high blood glucose.4. Compared with NC group, expression of PI3K mRNA, P85 protein, PKB-Ser473 and eNOS-Serl 177 phosphorylation were decreased, and no alteration in expression of PKB and eNOS of rat aorta in blood glucose fluctuation group.Conclusion:These findings suggest that glucose fluctation attenuate endothelium-dependent vasodilation function, through inhibiting PI3K/Akt/eNOS pathway activationObjective:To investigate the effects of intermittent high glucose on the production of nitric oxide(NO) and the expression of phosphatidylinositol 3-kinase(PI3K)/protein kinase B(PKB)/the endothelial nitric oxide synthase(eNOS) in cultured human umbilical vein endothelial cells (HUVECs). Methods:Intermittent high glucose and constant high glucose were applied to human umbilical vein endothelial cells HUVEC for 7 days. Concentration of NO were measured using Griess Reaction in cell culture supernatants, expression of PI3K-P85,Akt,eNOS and phosphorylation of Akt-Ser473,eNOS-Ser1 177 were measured using Western Blot.Results:1. Compared with NC group, NO production was significantly decreased in intermittent high glucose and constant high glucose cultured HUVCE (P<0.05), the inhibiting effect was in intermittent high glucose (P<0.05).2. Compared with NC group, expression of P85, Akt-Ser473 and eNOS-Ser1177 phosphorylation were significantly decreased (P<0.05), and no alteration in expression of Akt and eNOS in intermittent high glucose group (P>0.05).Conclusion:glucose variability may be more deleterious to human umbilical vein endothelial cells compared with constant high glucose, which show in sysnhesis vasodilator NO, by inhibiting PI3K/PKB/eNOS pathway activation.