电针对阿尔茨海默病模型幼鼠海马内源性神经干细胞增殖的影响

Effect of electroacupuncture on proliferation of hippocampal neural stem cells in young mice with Alzheimer′s disease

目的:观察电针对阿尔茨海默病(AD)模型幼鼠海马内源性神经干细胞增殖的影响,探讨电针防治AD的作用机制。方法:将40只1.5月龄APP/PS1转基因雄性幼鼠随机分为电针组和模型组,每组20只;20只同月龄C57BL/6J雄性幼鼠作为正常对照组。电针组于"百会""风府""肾俞"行电针干预,断续波,频率10 Hz,电流强度2 mA,每次20 min,每日1次,每周6 d,共干预16周。电针干预结束后,HE染色法观察各组小鼠海马齿状回神经元形态结构;免疫组化法检测海马CA1区5-溴脱氧尿嘧啶核苷(BrdU)阳性表达;免疫荧光法检测海马BrdU/神经元核抗原(NeuN)阳性表达;实时荧光定量PCR和Western blot法检测海马脑源性神经生长因子(BDNF)和巢蛋白(Nestin)mRNA和蛋白水平。结果:与正常对照组比较,模型组小鼠海马齿状回神经元结构不清晰,出现核固缩现象;海马CA1区BrdU阳性表达减少(P<0.01);海马BDNF和Nestin mRNA和蛋白水平均下降(P<0.01,P<0.05)。与模型组比较,电针组小鼠海马齿状回神经元轮廓较清晰,核结构较明显;海马CA1区BrdU阳性表达、BrdU/NeuN阳性表达均高于模型组(P<0.01,P<0.05);海马BDNF和Nestin mRNA和蛋白水平均高于模型组(P<0.05)。结论:电针可促进AD模型幼鼠海马内源性神经干细胞增殖,从而改善海马区神经元结构,其机制可能与促进BDNF的表达有关。

Objective To observe the effect of electroacupuncture(EA) on the proliferation of endogenous neural stem cells in the hippocampus of young mice with Alzheimer′s disease(AD), so as to explore its mechanisms underlying improvement of AD. Methods Forty 1.5-month-old APP/PS1 transgenic male mice were randomly divided into an EA group and a model group, 20 mice in each group, and other 20 C57 BL/6 J male mice of the same age were used as the normal control group. EA(intermittment wave 10 Hz, 2 mA) was applied to "Baihui"(GV 20), "Fengfu"(GV 16) and "Shenshu"(BL 23) for 20 min, once a day, 6 days a week for 16 weeks. H.E. staining was used to assess histopathological changes of neurons of the hippocampal dentate gyrus. Immunohistochemical stain was used to detect the expression of 5-bromodeoxyuridine(BrdU)-positive in the hippocampus, and immunofluorescence double-labeled technique was used to detect the number of proliferated positive neurons of hippocampal neural stem cells. The expression levels of brain derived neurotrophic factor(BDNF) and Nestin mRNA and protein were detected by using real-time PCR and Western blot, separately. Results The immunoactivity of BrdU, and the expression levels of BDNF and Nestin mRNA and protein in the hippocampus in the model group were significantly lower than in the normal control group(P<0.01, P<0.05), and considerably higher in the EA group than in the model group(P<0.01, P<0.05). The number of BrdU/NeuN dual labeled neurons was slightly increased in the model group than in the normal control group(P>0.05), and evidently increased in the EA group relevant to the model group(P<0.05), suggesting a proliferation of hippocampal neural stem cells. After modeling, the neurons of hippocampal dentate gyrus were arranged loosely and irregularly and their structure was fuzzy,with an appearance of different degrees of nuclear pyknosis, whereas in the EA group, the neuronal contour was clear and the nuclear structure was relatively distinct. Conclusion EA can activate the proliferation of neural stem cells in the hippocampus in AD mice, which may contribute to its function in improving the neuronal structure by upregulating the expression of BDNF.