酸枣仁皂苷缓解阿尔茨海默病模型小鼠的机制研究

Polygonum perfoliatum L.extract mechanisms in relieving an Alzheimer’s disease model in mice

目的探讨酸枣仁皂苷(ZSS)缓解七氟烷麻醉9月龄APP/PS1/tau三转基因阿尔茨海默病模型小鼠的机制研究。方法选取9月龄雄性APPSwe/PS1M146 V/tauP301 L 3xTg-AD 3xTg-AD小鼠和其背景C57BL/6J小鼠,按照随机分配原则,分为四组,每组12只,分别为:野生对照组Con(生理盐水)、三转模型对照组Model Con(生理盐水)、三转模型麻醉组Group 1(生理盐水+七氟烷)、三转模型给药组Group 2(生理盐水+七氟烷+ZSS)。所有小鼠,经侧脑室分别注射等容生理盐水或0.1μg/4μL的酸枣仁皂苷,连续10 d。对照组小鼠吸入2 h氧,麻醉组和给药组吸入2 h七氟烷。麻醉结束后24 h,每组取6只小鼠应用Morris水迷宫检测小鼠的空间学习及其记忆能力。其它小鼠通过断头方式取新鲜的海马组织,制备成匀浆,通过蛋白印迹法(Western blot)检测b淀粉样肽(Aβ)聚集(Aβ、Aβ1-40、Aβ1-42)情况,同时还检测了Tau蛋白磷酸化及其位点(丝氨酸396、丝氨酸262、苏氨酸231)的磷酸化水平变化情况,并通过Real-time PCR技术检测了线粒体凋亡信号通路基因,包括B淋巴细胞瘤-2基因(BcL-2)、BcL-2-相关的X蛋白(Bax)和胱天蛋白酶(Caspase-3)表达的变化情况。结果Morris水迷宫实验显示,七氟烷对小鼠学习记忆能力没有影响,但是对小鼠会产生潜在的神经性毒性;酸枣仁皂苷能降低三转模型小鼠中Aβ聚集和Tau蛋白磷酸化;与三转模型麻醉组相比,三转模型给药组中BcL-2表达减少,Bax和Caspase-3表达增强。结论酸枣仁皂苷能缓解七氟烷麻醉活化线粒体相关的凋亡信号通路,减少Aβ聚集,降低Tau蛋白磷酸化,抑制小鼠海马神经元的凋亡。

Objective We investigated mechanisms associated with Polygonum perfoliatum L.extract in relieving a 9-month-old APP/PS1/tau three-transgenic Alzheimer’s disease model in mice.Methods Male APPSwe/PS1M146 V/tauP301 L 3xTg-AD 3xTg-AD mice and their background C57BL/6J mice were randomly divided into four groups,with 12 mice in eachper group,respectively:a control group(normal saline),a three-turn model control group(normal saline),a three-turn model anesthesia group(normal saline+sevoflurane),and a three-turn model administration group(normal saline+sevoflurane+Polygonum perfoliatum L.extract).All mice were injected with isovolumic saline or 0.1μg/4μL of Polygonum perfoliatum L.extract through the lateral ventricle for 10 consecutive days.The control group inhaled two hours of oxygen,and the anesthetized group and the administration group inhaled two hours of sevoflurane.At 24 hours after the end of anesthesia,six mice in each group were tested for spatial learning and memory in a Morris water maze.Fresh hippocampal tissue was obtained by decapitation from the remaining mice,prepared into a homogenate,and b-amyloid peptide(Aβ)aggregation(Aβ,Aβ1-40,Aβ1-42)was detected by Western blot and by tau protein phosphorylation.The phosphorylation levels of all associated loci(Serine 396,Serine 262 and Threonine 231)were checked as well.The expression of mitochondrial apoptosis signaling pathway genes,including B-cell lymphoma-2(BcL-2),BcL-2-associated X protein(Bax)and Cysteine-requiring Aspartate Protease(Caspase-3)was detected by Real-time PCR.Results Morris water maze experiments show that sevoflurane has no effect on learning and memory ability in mice,but it does have potential neurotoxicity in mice.Extracts of Polygonum perfoliatum L.can reduce Aβaggregation and tau protein phosphorylation states in three-transformation mice.Compared with the three-turn model anesthesia group,BcL-2 expression was decreased and Bax and Caspase-3 expression was enhanced in the three-turn model administration group.Conclusions Polygonum perfoliatum L.extract can alleviate mitochondria-associated apoptotic signaling pathways induced by sevoflurane anesthesia,as well as reduce Aβaggregation,decrease tau protein phosphorylation,and inhibit the apoptosis of mouse hippocampal neurons.