SH-SY5Y神经细胞中α3尼古丁受体基因表达沉默对β分泌酶蛋白表达的影响

Effects of α3 nicotinic receptor gene silence in SH-SY5Y cells on the expression of β-secretase protein

目的研究α3神经型尼古丁受体(nAChR)基因沉默对β分泌酶(BACE)蛋白表达的影响。方法稳定转染α3 nAChR siRNA表达质粒及空质粒的SH-SY5Y神经细胞与正常对照组细胞用Real time-PCR和Western印迹法检测α3 nAChR mRNA表达和蛋白水平表达,以评价RNA的干扰效率;用蛋白质印迹方法分别检测细胞中β分泌酶两个亚型(BACE1和BACE2)蛋白表达水平的变化。结果稳定转染α3 nAChR siRNA表达质粒的细胞克隆株,与空质粒和正常对照组相比BACE1蛋白表达水平增加,BACE2蛋白表达水平减少。结论α3 nAChR可以通过增加BACE2蛋白表达水平及减少BACE1蛋白表达水平,影响Aβ生成从而减少Aβ的神经毒性作用,发挥神经保护作用。

Objective To examine the effects of α3 nicotinic acetylcholine receptor（nAChR） gene silence on the expression of β-secretase protein.Methods Neuroblastoma（SH-SY5Y） cells were transfected with small interference RNAs（siRNAs） that target specifically towards α3 nAChR.Real-time PCR and Western-blotting were used to detect whether the expression plasmid ofα3 nAChR siRNA had already transfected SH-SY5Y stably.Western-blotting was used to identify the protein expression changes of BACE1 and BACE2 which are two subtypes of β-secretase.Results Compared to controls and negative controls,SH-SY5Y cells transfected with siRNA expressed decreasing levels of α3 nAChR mRNA and protein;BACE1 was increased and BACE2 was decreased on the level of protein.Conclusions On account of reducing BACE1 and increasing BACE2 protein levels,α3 nAChR reduces the concentration of Aβ,which indicates α3 nAChR may play a significant neuroprotective role.