氧化应激上调人神经母细胞瘤细胞内β-裂解酶的表达

Oxidative stress up-regulates the expression of β-Amyloid precursor protein cleavage enzyme 1 in SH-SY5Y human neuroblastoma cells

目的研究氧化应激对人神经母细胞瘤细胞(SH-SY5Y)β-裂解酶(BACE1)表达的影响及组蛋白乙酰化、DNA甲基化的改变。方法采用H2O2处理体外培养的SH-SY5Y,Western blot法检测细胞的BACE1表达及DNA甲基转移酶(DNMTs)和组蛋白去乙酰化酶(HDAC)的表达;实时定量PCR检测BACE1 mRNA的表达;吸光度值法检测组蛋白3(H3)和组蛋白4(H4)整体乙酰化水平。结果 SH-SY5Y细胞经H2O2处理1和72 h后BACE1 mRNA和蛋白表达均明显增多;H2O2处理72 h后DNMT1、DNMT3A表达均下降,分别是对照组的75%和65%(P<0.01);而组蛋白去乙酰化酶HDAC3的表达增高至对照组的1.6倍(P<0.01);同时,组蛋白H3整体乙酰化水平下降,但H4乙酰化水平无明显改变。结论氧化应激可能通过改变SH-SY5Y细胞内DNA甲基化水平及组蛋白乙酰化状态调节BACE1的表达,在阿尔茨海默病发病过程中发挥作用。

Objective To investigate the role of oxidative stress in the expression of β-Amyloid precursor protein cleavage enzyme 1（BACE1） and the changes DNA methylation and histone acetylation.Methods Cultured SH-SY5Y cells treated with H2O2 were used to test the expressions of BACE1,DNA methyltransferases 1,3A（DNMT1,DNMT3A） and histone deacetyltranferase（HDAC） by were examined by Western blot.The level of mRNA of BACE1 was assessed by RT-PCR.Acetylation level of histone H3 and H4 was examined by optical density assay.Results Both BACE1 mRNA and protein levels were up-regulated significantly after H2O2 treatment for 1 and 72 h;DNMT1 and DNMT3A expressions were decreased to 75% and 65% of control respectively after H2O2 treatment for 72 h;HDAC3 level was increased by 1.6 folds as compared with control;While the level of histone H3 acetylation was decreased and there was no change with histone H4 acetylation.Conclusions Oxidative stress may regulate BACE1 expression in SH-SY5Y through alteration of DNA methylation and histone acetylation which play a role in Alzheimer＇s disease（AD） pathogenesis.