二甲双胍对阿尔茨海默症模型大鼠认知功能障碍及PI3K/Akt通路的影响

doi:10.12300/j.issn.1674-5817.2020.185

摘要/Abstract

摘要： 目的探讨二甲双胍（metformin,Met）对阿尔茨海默症（Alzheimer's disease,AD）大鼠磷脂酰肌醇-3-激酶（phosphatidylinositol-3-kinase,PI3K）/蛋白激酶B（protein kinase B,Akt）通路及认知功能障碍的影响。方法取SD大鼠50只,用随机数字表法分为正常组（Normal组）、AD模型组（AD组）、Met低剂量（50 mg/kg）组、Met中剂量（100 mg/kg）组、Met高剂量（200 mg/kg）组,每组10只;除Normal组外,其余各组大鼠均在双侧脑室注射10 μL链脲霉素（3 mg/kg）建立AD模型。造模成功后开始给药,Met组灌胃给予相应剂量的Met溶液,Normal组和AD组灌胃给予等量生理盐水,各组连续给药14 d,1次/d。末次给药后用Morris水迷宫试验检测大鼠空间认知功能。处死大鼠,取海马组织,用ELISA法检测海马组织中β-淀粉样蛋白42（β-amyloid protein 42,Aβ42）与磷酸化Tau蛋白（p-tau）含量;用HE染色法观察海马组织中神经元病理变化;用免疫组织化学法检测PI3K阳性表达;用蛋白质印迹法检测海马组织中PI3K、磷酸化Akt（p-Akt）、胰岛素受体底物-1（insulin receptor substrate-1,IRS-1）、糖原合酶激酶-3（glycogen synthase kinase-3,GSK-3）蛋白相对表达水平。结果与Normal组相比,AD组大鼠海马可见神经元细胞排列紊乱、减少等病理损伤,大鼠穿越平台次数、原平台象限游泳距离与总距离的比值、游泳时间与总时间的比值均明显减少（P<0.05）,海马组织中PI3K、p-Akt和IRS-1蛋白表达水平均明显降低（P<0.05）,平均逃避潜伏期延长（P<0.05）,海马组织Aβ42及p-tau含量、GSK-3蛋白表达明显升高（P<0.05）;与AD组相比,Met低、中、高剂量组大鼠海马组织的病理损伤减轻,大鼠穿越平台次数、原平台象限游泳距离与总距离的比值、游泳时间与总时间的比值均明显增加（P<0.05）,海马组织PI3K、p-Akt、IRS-1蛋白表达水平明显升高（P<0.05）,平均逃避潜伏期缩短（P<0.05）,海马组织Aβ42及p-tau含量、GSK-3蛋白表达明显降低（P<0.05）,且Met各剂量组上述指标呈剂量依赖性。结论 Met可激活AD大鼠海马组织PI3K/Akt通路,降低海马组织p-tau、Aβ42含量,改善AD大鼠认知障碍。

关键词: 二甲双胍, 阿尔茨海默症, 磷脂酰肌醇-3-激酶, 蛋白激酶B, 认知功能障碍, 大鼠

Abstract: Objective To investigate the effects of metformin (Met) on the phosphatidylinositol-3-kinase (PI3K)/protein kinase B (Akt) pathway and cognitive dysfunction in Alzheimer's disease (AD) rats. Methods Fifty Sprague Dawley rats were selected and divided into the following groups: normal, AD model (AD group), Met low (50 mg/kg), Met medium (100 mg/kg) and Met high (200 mg/kg) according to the random number table method. Each group included 10 rats. With the exception of the normal group, AD models were established by injecting 10 μL streptozotocin (STZ, 3 mg/kg) into the bilateral ventricles. After the model was successfully established, Met at the corresponding dose was administered by gavage. The normal and AD groups were administered the same amount of normal saline by gavage. The medicine was administered once a dayfor 14 days. After the last administration, the Morris water maze test was used to detect the spatial cognitive function of rats. The rats were sacrificed, and the hippocampal tissues were collected. The contents of β-amyloid protein 42 (Aβ42) and phosphorylated tau protein (p-tau) in hippocampus were detected by enzyme-linked immunosorbent assay (ELISA). The pathological changes of neurons in hippocampus were observed by HE staining. The detection of PI3K positive expression was performed by immunohistochemistry. Western blots were used to detect the relative expression levels of PI3K, phosphorylated Akt (p-Akt), insulin receptor substrate-1 (IRS-1), and glycogen synthase kinase-3 (GSK-3) in hippocampus. Results Compared with the normal group, there were pathological damages, such as disordered arrangement and decrease of neurons in hippocampus of rats in the AD group. The number of times the platform was crossed, the ratio of swimming distance to the total distance in the original platform quadrant, the ratio of swimming time to the total time, and the protein expressions of PI3K, p-Akt and IRS-1 in hippocampus of the AD group were significantly low (P < 0.05). The average escape latency, the contents of Aβ42 and p-tau, and the protein expression of GSK-3 in hippocampus were high (P < 0.05). Compared with the AD group, the pathological damages of hippocampal tissue in low, medium, and high dose Met rats were alleviated. The number of times the platform was crossed, the ratio of swimming distance to the total distance in the original platform quadrant, the ratio of swimming time to the total time, and the protein expressions of PI3K, p-Akt and IRS-1 in hippocampus of the AD group were significantly high (P < 0.05). The average escape latency, the contents of Aβ42 and p-tau, and the protein expression of GSK-3 in hippocampus were low (P < 0.05). Moreover, the above indexes were dose-dependent in the Met groups. Conclusion Met can activate the PI3K/Akt pathway, decrease the levels of p-tau and Aβ42 in hippocampus, and improve cognitive impairment in AD rats.

Key words: Metformin, Alzheimer's disease, Phosphatidylinositol-3-kinase, Protein kinase B, Cognitive dysfunction, Rats

中图分类号:

Q95-33
R-332

王百乔, 林小茹, 韩敏, 刘煜, 唐超玲. 二甲双胍对阿尔茨海默症模型大鼠认知功能障碍及PI3K/Akt通路的影响[J]. 实验动物与比较医学, 2021, 41(4): 313-320.

WANG Baiqiao, LIN Xiaoru, HAN Min, LIU Yu, TANG Chaoling. Effects of Metformin on Cognitive Dysfunction and PI3K/Akt Pathway in Alzheimer's Disease Rats[J]. Laboratory Animal and Comparative Medicine, 2021, 41(4): 313-320.

图/表 5

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