补气开窍法对APP/PS1双转基因小鼠认知能力和β-淀粉样肽42的影响

Effect of Qi-tonifying and Resuscitation-inducing Therapy on Cognition and Aβ_(42) Expression in APP/PS1 Double Transgenic Mice

【目的】探讨补气开窍法对APP/PS1双转基因小鼠认知能力和β-淀粉样肽42(Aβ42)的影响及其自噬机制。【方法】将56只APP/PS1双转基因小鼠随机分为4组,雌雄各半,每组14只,即模型组、人参总皂苷组(剂量为75 mg/kg)、石菖蒲挥发油组(剂量为15 mg/kg)、补气开窍方组(剂量为石菖蒲挥发油15 mg/kg+人参总皂苷75 mg/kg)。另设正常组,C57BL/6品系的正常小鼠14只,雌雄各半。各组灌胃给药,模型组和正常组用等体积的蒸馏水代替,给药30 d。末次给药结束1 h后,先将各组小鼠进行行为学检测(水迷宫实验,避暗实验和跳台实验),再麻醉各组小鼠,其中4只灌注固定后的全脑用于免疫组织化学法检测Aβ42,其余小鼠在冰上迅速取出海马用于检测Aβ42,流式细胞术检测磷酸化的哺乳动物雷帕霉素蛋白(p-m TOR)和自噬相关基因(Beclin-1)。【结果】与正常组比较,模型组水迷宫实验的潜伏期显著延长(P<0.01),避暗实验和跳台实验的潜伏期显著缩短(P<0.01),Aβ42含量和Beclin-1表达显著增加(P<0.01),p-m TOR表达显著减少(P<0.01);与模型组、人参总皂苷组、石菖蒲挥发油组比较,补气开窍方组的水迷宫实验的潜伏期显著缩短(P<0.05),避暗实验和跳台实验的潜伏期显著延长(P<0.05),水迷宫实验、避暗实验和跳台实验的错误次数显著减少(P<0.05),Aβ42含量显著减少(P<0.05),此外,与模型组比较,补气开窍方组的p-m TOR表达显著增加(P<0.05),Beclin-1表达显著减少(P<0.05)。【结论】人参总皂苷联合石菖蒲挥发油具有改善APP/PS1双转基因模型小鼠的认知能力,减少Aβ42表达和调节自噬作用,且优于单独使用人参总皂苷组或石菖蒲挥发油组。

Objective To observe the influence of Qi-tonifying and resuscitation-inducing （QTRI） therapy on cognition and the level of β-amyloid 42 （Aβ42） in APP/PS1 double transgenic mice, and to explore its autophagy mechanism. Methods Fifty-six APP/PS1 double transgenic mice, male in half, were randomly divided into model group, total ginsenosides group（at the dosage of 75 mg/kg） , Rhizoma Acori Tatarinowii （Shichangpu） volatile oil group （at the dosage of 15 mg/kg） and QTRI group （15 mg/kg of Shichangpu volatile oil and 75 mg/kg of total ginsenosides）. Additionally, 14 normal C57BL/6 mice （male in half） were used as the normal group. The model group and the normal group were given the same volume of normal distilled water, and the medication groups were administrated with the corresponding drug for 30 days respectively. One hour after the last medication, the behavioral tests such as water maze test, step-through test and step-down test were carried out. After that, all the mice in various groups were killed, and then the whole brain tissues of 4 mice were used for the observation of Aβ42 expression by immunohistochemistry. The hippocampus of the other mice was separated for the detection of Aβ42 by enzyme-linked immunosorbent assay （ELISA） and the determination of phosphorylated mammalian target of rapamycin （p-mTOR） and Beclin-1 expression levels by flow cytometry. Results Compared with the normal group, the latent period of the model group in water maze test was prolonged （P 〈 0.01）, and that in step-through test and step-down test was shortened（P 〈 0.01 ）. Moreover, the expression levels of Aβ42 and Beclin-1 were increased （P 〈 0.01）, and the expression level ofp-mTOR was decreased （P 〈 0.01 ） in the model group. Compared with the model group, total ginsenosides group and Shichangpu volatile oil group, the latent period of QTRI group in water maze test was shortened （P〈 0.05） and that in step-through test and step-down test was prolonged （P〈 0.05）, the error times in water maze test, step-through test and step-down test were reduced （P 〈 0.05） , and the expression level of Aβ42 was decreased （P〈 0.05）. Additionally, the expression level of p-roTOR was increased（P 〈 0.01 ） , and that of Beclin-1 was decreased（P 〈 0.05 ） in QTRI group compared with those in the model group. Conclusion Total ginsenosides combined with Shichangpu volatile oil have an effect on improving cognition, decreasing the expression level of Aβ42 and regulating autophagy response in APP/PS1 double transgenic mice, and the effect is better than that of total ginsenosides or Shichangpu volatile oil alone.