【作者】 刘英；

【导师】 邹伟；

【作者基本信息】 辽宁师范大学 ， 动物学， 2009， 硕士

【摘要】 糖尿病是一种常见的代谢类疾病,近年来,发病率逐步升高,其临床表现主要有血糖升高、胰岛素抵抗以及糖尿病引起的各种并发症,如高血压、神经病变、增生性视网膜病和肾病综合症等,其中糖尿病肾病综合症(diabeticnephropathy,DN)是常见的微血管并发症,严重威胁糖尿病患者的生命及生活质量。DN的确切发病机制不明,目前认为受多方面因素的影响,包括代谢因素、血流动力学因素、氧化应激和遗传因素等,因此研究DN的发病机制对糖尿病。肾病的治疗具有重要意义。钒是人体必需的微量元素之一。研究表明,无机钒化合物和有机钒化合物均可改善糖尿病大鼠的高血糖症状,促进葡萄糖转运和糖原合成。但无机钒化合物的胃肠道吸收率低,服用高剂量化合物后,实验动物常出现腹泻、精神萎靡、死亡等现象。与无机钒化合物相比,有机钒化合物能提高钒在体内吸收率及生物利用率,并且有机钒氧配合物的降糖效果更为明显,毒副作用也较低,其作用机制并不完全清楚。本论文采用实验室前期筛选出来的低毒有机钒氧配合物NVC-T2对四氧嘧啶诱导的糖尿病小鼠进行给药干预,检测有机钒氧配合物干预后各组小鼠血糖、血清肌酐、血尿素氮、谷丙转氨酶和谷草转氨酶等的变化,评价其对糖尿病小鼠肾功能的影响。蛋白质免疫印迹检测NVC-T2给药干预后糖尿病小鼠。肾皮层及不同浓度葡萄糖刺激的肾小球系膜细胞中胰岛素受体信号转导通路相关蛋白的表达,进一步探讨NVC-T2类胰岛素作用的信号转导机制,旨在为有机钒化合物成为糖尿病肾病治疗药物的研发提供一定的理论基础。实验结果:(1)有机钒氧配合物NVC-T2给药干预后,糖尿病组小鼠血糖、血清肌酐、血尿素氮含量明显下降,谷丙转氨酶和谷草转氨酶的比值升高。同时,通过制作组织切片观察糖尿病小鼠肾小球,发现糖尿病小鼠肾小球发生了明显的形态学变化。(2)与对照组小鼠相比,糖尿病小鼠肾皮层中Cav-1、MAPK和p-MAPK表达量均上调,经NVC-T2给药干预后下调。糖尿病小鼠肾皮层中p-Akt蛋白表达量下调,NVC-T2给药干预后上调。总Akt表达量和对照组小鼠基本一致。(3)与低糖对照组肾小球系膜细胞相比,高糖刺激的肾小球系膜细胞中Cav-1、MAPK和p-MAPK蛋白表达量显著上调,p-Akt蛋白表达量显著下调。Akt蛋白表达量上调,但与对照组相比,无显著性差异。NVC-T2刺激后,Cav-1、MAPK、p-MAPK和Akt表达量均下调,p-Akt蛋白表达量上调。结论:1.有机钒氧配合物NVC-T2具有明显的降血糖效果,可以改善糖尿病小鼠的肝肾功能。2.NVC-T2可明显上调糖尿病小鼠肾皮层和高糖刺激的肾小球系膜细胞中p-Akt蛋白的表达,下调MAPK蛋白和p-MAPK蛋白的表达,提示其对糖尿病小鼠肾功能的影响可能与胰岛素受体起始的PI3K/Akt通路及Ras/MAPK通路有关。3.糖尿病小鼠肾皮层及高糖刺激的肾小球系膜细胞中Cav-1蛋白的表达量升高,Ras/MAPK通路被激活,而PI3K/Akt通路被抑制,提示在糖尿病肾病的发展过程中Cav-1蛋白可能是通过调节胰岛素信号受体信号转到而起重要的作用。更多还原

【Abstract】 Diabetes is a common metabolic disease, recently, the incidence of diabetes was escalate in China, the clinical manifestations of diabetes were shown as hyperglycemia, insulin resistance and complications induced by diabetes, such as hypertension, neuropathy, proliferative retinopathy and diabetic nephropathy, DN. DN is a kind of microvascular complications which seriously affect the quality of life of diabetic patients. The pathogenesis of DN is unclear. It is considered that multiple factors can induced DN include metabolic factors, hemodynamic factors, oxidative stress and genetic factors. So there is important significance to research the pathogenesis of DN for the treatment of DN.Vanadium is one of necessary trace elements for human body. The results show that both inorganic vanadium complexes and organic vanadium complexes can increase glucose level of diabetic rats, promote glucose transport and glycogen synthesis. But the absorption rate of gastrointestinal tract of inorganic vanadium complexes is lower than organic vanadium complexes. Inorganic vanadium complexes often result in diarrhea and death of experimental animals. Organic vanadium complexes can improve the absorption rate and bioavailability of vanadium, and its hypoglycemic effect is more significant, they have low toxicity. But its mechanism is still unclear.In this study, we use the organic vanadium complex NVC-T2 selected with low toxicity to intervene the diabetic mice induced by alloxan, detect the change of glucose level, serum Cr, BUN, AST and ALT, evaluate its effect on the renal function of diabetic mice. Western blot detects the expression of associated proteins of insulin receptor signal transduction pathway in the renal cortex of diabetic mice and glomerular mesangial cells stimulated by high glucose, investigate the signal transduction mechanism of insulin like activity of NVC-T2. Our results showed that:(1) After the intervention of NVC-T2, the glucose level, serum Cr and BUN of diabetic mice decreased, AST/ALT rates increased. Observe the glomerular by histological sections, we found that the glomerular of diabetic mice occurred significant histopathologic changes.(2) Compared with control mice, the expression of Cav-1, MAPK, p-MAPK, increased significantly in the renal cortex of diabetic mice. After the intervention of NVC-T2, their expression decreased. The expression of p-Akt decreased, after the intervention of NVC-T2, its expression increased. The expression of Akt was similar with control mice. (3) Compared with control glomerular mesangial cells, the expression of Cav-1, MAPK, p-MAPK, increased significantly in glomerular mesangial cells stimulated by high glucose. The expression of p-Akt decreased significantly. The expression of Akt increased, but there was no significant difference compared with control glomerular mesangial cells. After the stimulation of NVC-T2, the expression of Cav-1, MAPK, p-MAPK, decreased, p-Akt increased.Conclusion:1. Organic vanadium complex NVC-T2 has obvious hypoglycemic effect and can improve hepatic and renal function.2. After the intervention of NVC-T2, the expression of p-Akt increased, the expression of MAPK, p-MAPK, decreased in the renal cortex of diabetic mice and glomerular mesangial cells stimulated by high glucose. Indicate that the effect of NVC-T2 on the renal function of diabetic mice may have relationship with PI3K/Akt pathway and Ras/MAPK pathway.3. The expression of Cav-1 increased in the renal cortex of diabetic mice and glomerular mesangial cells stimulated by high glucose. Ras/MAPK pathway was activated ,and PI3K/Akt pathway was inhibited, indicate that Cav-1 may play an importmant rolein the development of DN by regulating insulin receptor signal transduction pathway更多还原