目的探讨Aβ_(25-35)介导SH-SY5Y细胞Bcl-2、Bax基因表达改变是否通过基因启动子区甲基化的机制。方法不同浓度Aβ_(25-35)(0、25、50μmol·L~(-1))分别作用于体外培养的SH-SY5Y细胞48、72 h,MTT法确定Aβ_(25-35)诱导SH-SY5Y细胞凋亡的最佳浓度和时间。Western blot检测不同药物处理组细胞凋亡相关蛋白Bcl-2、Bax表达变化,Real time PCR检测DNA甲基化酶DNMT1、DNMT3a、DNMT3b、Me CP2的mRNA水平。Methylation specific PCR(MSP)法分析Aβ_(25-35)介导的Bcl-2、Bax基因启动子区甲基化水平的变化。结果 25μmol·L~(-1)Aβ_(25-35)暴露SH-SY5Y细胞72 h后,MTT检测细胞存活率达(68.49±9.83)%,与对照组比较明显降低(P<0.05),表明成功建立了AD细胞凋亡模型。Western blot结果显示,Aβ_(25-35)药物处理组与空白组比较,Bcl-2表达明显减少,Bax表达明显增加。Real-time PCR结果显示,不同浓度...
【英文摘要】
Aim To investigate whether the effect of Aβ_(25-35) on Bcl-2 and Bax gene transcription through DNA methylation in SH-SY5 Y cell. Methods Different concentrations of Aβ_(25-35)( 0,25,50 μmol·L~(-1))were treated with SH-SY5 Y cells for 48 h or 72 h in vitro. The optimal concentration and time of Aβ_(25-35) induced SH-SY5 Y apoptosis were determined by MTT method. Protein expression levels of Bcl-2 and Bax of Aβ_(25-35)-treated groups were determined by Western blot. Real time PCR was used to detect the mRNA ...