α7尼古丁受体mRNA表达抑制对SH-SY5Y细胞氧化应激水平的影响

Influence of inhibiting α7 neuronal nicotinic acetylcholine receptor mRNA on lipid peroxidation in SH-SY5Y cells

目的探讨通过RNA干扰技术抑制神经母细胞瘤细胞(SH-SY5Y细胞)中α7尼古丁乙酰胆碱受体(nAChR)基因表达后细胞氧化应激水平的变化,以了解该受体的神经保护作用。方法 SH-SY5Y细胞转染针对α7 nAChR的小分子干扰RNA,Real-time PCR法检测转染细胞中α7 nAChR mRNA水平蛋白质印迹方法检测蛋白表达水平;并用Aβ1～42处理培养细胞,比色法测定细胞脂质过氧化产物水平、超氧化物歧化酶(SOD)和谷胱甘肽过氧化物酶(GSH-PX)活性。结果转染siRNA后,α7 nAChR mRNA水平和蛋白表达水平分别降低了81.7%和76.9%;细胞脂质过氧化水平升高38.4%;SOD和GSH-PX活性分别降低30.3%和21.1%。用Aβ1～42处理细胞后氧化应激水平升高,且转染siRNA后能增强Aβ的神经毒性作用。结论α7 nAChR基因表达抑制后能增强细胞氧化应激水平,同时增强Aβ的神经毒性作用。

Objective To investigate the influence of inhibiting the α7 nicotinic receptor （nAChR） by small interference RNA on lipid peroxidation in SH-SY5Y cells and reveal the connection of these changes with the pathogenesis of neuroprotective function. Methods The siRNAs of α7 nAChR were transfected into SH-SY5Y cells and the levels of α7 nAChR mRNA and protein were studied by real-time PCR and Western blotting, respectively. After the SH-SYSY cells were transfected with siRNA, the cells were exposed again to 1 μmol/L Aβ1 -42, the lipid peroxidation, activities of SOD and GSH-PX were measured by spectrophotometry methods. Results Compared with con- trois, the expressions of mRNA and protein in the SH-SY5Y ceils transfected with the α7 nAChR siRNA were decreased by 81.7% and 76. 9%, respectively. The product of lipid peroxidation was increased by 38.4%, the activities of SOD and GSH-PX were decreased by 30. 3% and 21.1%, respectively. Inhibited gene expression of α7 nAChR enhanced the toxicity induced by Aβ. Conclusions The inhibi- ted α7 nAChR mRNA induced by siRNA could stimulate the level of oxidative stress and enhance the neurotoxic effects of Aβ.