铝对PC12细胞淀粉样前体蛋白β位点裂解酶-1蛋白及基因表达的影响
Effects of Aluminum on the Expression of BACE1 Proteins and Genes in PC12 Cells
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摘要:
目的 探讨铝对PC12细胞淀粉样前体蛋白(APP)β位点裂解酶-1(beta-site amyloid precursor proteincleaving enzyme-1,BACE1)蛋白及基因表达的影响。 方法 采用PC12细胞进行培养及麦芽酚铝Al (mal)3染毒,将细胞分为6组,分别为:200 μmol/L生理盐水组;0、50、100、200和400 μmol/L Al(mal)3组。分别采用酶联免疫吸附测定法(enzyme linked immunosorbent assay,ELISA)、荧光实时定量聚合酶链反应(quantitative real-time polymerase chainreaction,qRT-PCR)于染毒12、24和48 h后测定BACE1蛋白含量及基因表达。 结果 细胞计数试剂盒-8(CCK-8)细胞活力测定结果显示,不同浓度Al (mal)3染毒PC12细胞,可使其细胞活力呈现随染毒时间延长而逐渐下降的趋势。qRT-PCR结果显示,100 μmol/L Al(mal)3组在染毒48 h后BACE1基因表达明显高于生理盐水组、0 μmol/L Al(mal)3组,差异有统计学意义(P<0.05);200、400 μmol/L Al(mal)3组染毒12、24、48 h后BACE1基因表达明显高于生理盐水组和0μmol/L Al(mal)3组,差异均有统计学意义(P<0.05)。ELISA测定结果显示,染毒12h后400μmol/L Al(mal)3组BACE1表达量明显高于生理盐水组和0μmol/L Al(mal)3组,差异均有统计学意义(P<0.05);染毒24h后200、400μmol/L Al(mal)3组BACE1表达量明显高于生理盐水组和0μmol/L Al(mal)3组,差异有统计学意义(P<0.05);染毒48h后400μmol/L Al(mal)3组的BACE1表达量明显高于生理盐水组和0 μmol/L Al(mal)3组,差异有统计学意义(P<0.05)。 结论 Al(mal)3对PC12细胞具有明显毒性作用,该作用可能与麦芽酚铝致PC12细胞BACE1蛋白和基因表达增强有关。 Abstract:Objective To explore the effect of aluminum on the expression of beta-site amyloid precursor protein cleaving enzyme-1 (BACE1) proteins and genes in PC12 cells. Methods Cultured PC12 cells were randomly divided into 6 groups:200 μmol/L saline group, and 0, 50, 100, 200 and 400 μmol/L Al(mal)3 groups. The protein and gene expression of BACE1 was detected by enzyme linked immunosorbent assay (ELISA) and quantitative real-time polymerase chain reaction (qRT-PCR) at 12, 24 and 48h after treatment. Results The viability of PC12 cells was decreased in a time-dependent manner in different dose groups. qRT-PCR results showed that the expression of BACE1 mRNA in 100 μmol/L Al(mal)3 group increased significantly compared with that in 0 μmol/L Al(mal)3 group and saline group after 48 h exposure (P<0.05); and that in 200 and 400 μmol/L Al(mal)3 groups in creased significantly compared with that in 0 μmol/L Al(mal)3 group and saline group after 12, 24 and 48 h exposure (P<0.05). ELISA detection results showed that, compared with the 0 μmol/L Al(mal)3 group and the saline group, the expression of BACE1 in 200 μmol/L Al(mal)3 group increased significantly after 24 h exposure (P<0.05); and that in 200 and 400 μmol/L Al(mal)3 groups in creased significantly after 12, 24 and 48 h exposure (P<0.05). Conclusion The obvious toxicity of Al(mal)3 on PC12 cells may be related to the enhancing expression of BACE1 protein and gene in PC12 cells.
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