【作者】 李丽；

【导师】 黄起壬；

【作者基本信息】 南昌大学 ， 药理学， 2009， 硕士

【摘要】 目的:探讨阴离子交换蛋白2(AE2)在2型糖尿病大鼠胸主动脉内皮功能失常中的作用及其可能机制。方法:高脂高糖饮食加小剂量链脲佐菌素建立2型糖尿病大鼠模型,随机分为4组:正常(NC)组、糖尿病(DM)组、糖基化终末产物抑制剂(AG)组、ROS抑制剂(MTZ)组,处理8周后,观察以下指标:①免疫组化检测胸主动脉AE2蛋白表达;②乙酰胆碱(Ach)诱导的内皮依赖性血管舒张反应检测血管舒张功能;③RT-PCR检测胸主动脉eNOSmRNA表达;④硝酸还原酶法检测血清NO浓度。另外,在细胞水平利用RT-PCR检测HUVECs中AE2 mRNA的表达以及Western Blotting检测AE2蛋白进一步证实高糖诱导AE2蛋白上调的机制。结果:①DM组血糖(18.18±4.04mmol/L)、血清胰岛素(12.05±3.55mIU/L)、甘油三酯(1.26±0.53mmol/L)较NC组血糖(5.36±0.64mmol/L)、血清胰岛素(4.82±1.57 mIU/L)、甘油三酯(0.30±0.08mmol/L)均显著升高(P<0.05),提示模型制作成功。②DM组(0.672±0 .018)AE2蛋白表达较NC组(0.157±0 .020)明显上调(P<0.05),而AG组(0.321±0. 013)、MTZ组(0.284±0.024)较DM组表达下调(P<0.05)。③D M组血管舒张功能明显减弱(P <0.05),而AG组、MTZ组有部分恢复(P<0.05),且舒张功能与AE2蛋白表达量呈负相关。④DM组较NC组eNOS mRNA表达上调(P<0.01),血清NO浓度下降(P<0.01);在AG组、MTZ组eNOS mRNA表达进一步上调(P<0.01),NO明显升高(P<0.01)。⑤HUVECs AE2mRNA和AE2蛋白的表达: HG组较Control组AE2 mRNA转录显著上调(P <0.01),AE2蛋白表达上调(P <0.05);在处理组AG组及MTZ组两者表达均下调(P <0.05)。结论:AE2可介导糖尿病引起的内皮功能失常,抑制AE2表达及功能可改善血管内皮功能;AE2介导血管内皮功能失常的机制可能是eNOS/NO依赖的;糖尿病诱导AE2表达机制是AGEs/ROS依赖的。更多还原

【Abstract】 Objective: To study the effect of AE2 protein on arotic endothelium function in type 2 diabetic rats and its mechanism.Methods: Male SD rats were injected with low dose streptozotocin and fed with diets enriched in fat and sugar to form type 2 diabetic mode1.Then, the rats were dealed with in 4 groups, normal group(NC),diabetes group(DM),diabetes with drug aminoguanidine group(AG), diabetes with drug myxothiazole group (MTZ). After 8 weeks, the expression of AE2 protein and eNOSmRNA in the aorta, the Ach—dependent vasodilation response of isolated aorta,the synthesis of nitric oxide(NO) in serum were detected.We also used the method of RT-PCR and Western-Blotting to improve the mechanism that hight glucose upregulated the expression of AE2mRNA and protein in HUVEC.Results: (1)In DM group fast plasma glucose (18.18±4.04mmol/L), insulin (12.05±3.55 mIU/L),triglyceride(1.26±0.53mmol/L) levers were significantly higher than fast plasma glucose (5.36±0.64mmol/L), insulin (4.82±1.57 mIU/L), triglyceride (0.30±0.08mmol/L) in NC group (P<0.05). This suggested a perfect model made. (2) AE2 protein immnohistochemical expression in aorta endomembrane were significantly upregulated in DM group(0.672±0.018) compared with that in NC group (0.157±0.020), but in AG group(0.321±0.013), MTZ group(0.284±0.024) they were downregulated compared with that in DM group(P<0.05 ). (3) the maximum acetylcholine (Ach) -dependent vasodilation response (EDVRmax) of aorta decreased significantly in DM group compared with that in NC group(P<0.05); In AG,MTZ group EDVRmax recovered partly compared with that in DM group (P<0.05); Interesting, the EDVRmax was negatively correlated with AE2 protein(P<0.01). (4) In DM group, the eNOS mRNA expression were significantly higher and NO concentration were lower than that in NC group (P<0.01),and in AG、MTZ group the eNOS mRNA expression become more higher and NO concentration were higher also (P<0.01).(5) The expression of AE2 mRNA and AE2 protein in the HG group were higher than that in control group (P<0.05); But they decreased after dealed with in MTZ or AG group(P<0.05).Conclusion: AE2 protein induced endothelial disfunction in type 2 diabetes . Depressing the expression of AE2 protein and its function could get a improved EDVR .The mechanism of AE2 induced endothelial disfunction is probably eNOS/NO dependent. Hight glocuse upregulated the expression of AE2 protein though AGEs/ROS pathway.更多还原