摘要目的 通过RNA干扰技术抑制神经母细胞瘤细胞(SH-SY5Y细胞)中α7神经型尼古丁乙酰胆碱受体(nAChR)的基因表达,以了解α7 nAChR对β淀粉样蛋白(Aβ)的影响,进一步探讨阿尔茨海默病的发病机制.方法 SH-SY5Y细胞转染α7 nAChR的小干扰RNA(siRNA),用real-time PCR法和蛋白免疫印迹法分别测定细胞中α7 nAChR、β分泌酶两个亚型(BACE1和BACE2)在mRNA和蛋白表达水平的变化；用双抗体夹心酶联免疫吸附法测定Aβ1-42含量的变化.结果 转染α7 nAChR siRNA后,与对照组相比,α7 nAChR mRNA及蛋白表达水平分别降低了84％和79％(P＜0.01)；BACE1 mRNA及蛋白表达水平分别升高了527％和71％(P＜0.01)；BACE2 mRNA及蛋白表达水平分别降低了58％和75％ (P＜0.01)；Aβ1-42含量升高了208％(P＜0.01).结论 α7 nAChR基因表达抑制后Aβ含量明显升高,其机制可能与BACE1表达上升和BACE2表达抑制有关,这种改变可能与阿尔茨海默病的发病有一定的关系.

abstractsObjective To investigate the influence of inhibited α7 neuronal nicotinic acetylcholine receptor (nAChR) by small interference RNA (siRNA) in SH-SY5Y cells and to explore the connection of these changes with the β-amyloid precursor protein (APP) metabolism and the pathogenesis of Alzheimer's disease (AD).Methods The siRNA of α7 nAChR was transfected into SH-SY5Y cells,and the expression of α7 nAChR and two subtypes of β-secretases (BACE1 and BACE2) at mRNA and protein levels was studied by real-time PCR and Western blot,respectively.The variation of Aβ1-42 content was detected by ELISA.Results As compared with controls,the expression of α7 nAChR at mRNA and protein levels in the SH-SY5Y cells transfected with the α7 nAChR siRNA were decreased by 84％ and 79％ (P ＜0.01),respectively.The expressions of BACE1 mRNA and protein levels was increased by 527％ and 71％ (P ＜ 0.01),respectively,while the expression of BACE2 decreased by 58％ and 75％ (P ＜ 0.01),respectively.The Aβ1-42 content increased by 208％ (P ＜ 0.01).Conclusions An inhibited α7 nAChR mRNA induced by siRNA may markedly stimulate the production of Aβ through the mechanism of increased expression of BACE1 and inhibited expression of BACE2,which may be related to the pathogenesis of AD.