铝暴露对大鼠PC12细胞tau蛋白异常磷酸化的影响  

Effect of aluminum exposure on abnormal phosphorylation of tau protein in PC12 cells of rats

作　　者：宋珊珊 徐旭 许杨丹 肖雯洁 杨晓娟[1,2] SONG Shanshan;XU Xu;XU Yangdan;XIAO Wenjie;YANG Xiaojuan(School of Public Health,Shanxi Medical University,Taiyuan,Shanxi 030001,China;The First Hospital of Shanxi Medical University,Taiyuan,Shanxi 030001,China)

机构地区：[1]山西医科大学公共卫生学院,山西太原030001 [2]山西医科大学第一医院,山西太原030001

出　　处：《预防医学》2023年第3期271-274,共4页CHINA PREVENTIVE MEDICINE JOURNAL

基　　金：山西省留学人员科技活动择优资助项目(20200008);山西省基础研究计划(202103021224398)。

摘　　要：目的分析铝暴露对大鼠肾上腺髓质嗜铬细胞瘤分化细胞(PC12细胞)中miR-497-5p、无翅型鼠类乳腺癌病毒整合位点家族成员3a(Wnt3a)、β连环蛋白(β-catenin)、糖原合成激酶-3β(GSK-3β)蛋白和tau蛋白表达的影响,为铝暴露致tau蛋白异常磷酸化的机制研究提供依据。方法体外培养的PC12细胞分别暴露于0、100、200和400μmol/L的Al(mal)3溶液24 h,采用超敏型细胞增殖检测试剂(CCK-8)法检测细胞活力;采用实时荧光定量PCR(RT-qPCR)法检测miR-497-5p和Wnt3a基因表达,采用免疫印迹法检测Wnt3a、β-catenin、GSK-3β、P-GSK-3β(Ser9)、tau和P-tau(Ser396)蛋白相对表达量,分析各剂量组目的基因和蛋白的表达趋势。结果PC12细胞活力随铝暴露剂量的增加呈下降趋势(F_(趋势)=323.473,P=0.001)。miR-497-5p相对表达量随铝暴露剂量的增加而上升(F_(趋势)=14.888,P=0.031);Wnt3a基因相对表达量随铝暴露剂量的增加而下降(F_(趋势)=165.934,P<0.001);miR-497-5p相对表达量与Wnt3a相对表达量呈负相关(r=-0.693,P=0.012)。Wnt3a、β-catenin和p-GSK-3β(Ser9)蛋白相对表达量随铝暴露剂量的增加而下降(F_(趋势)=357.656,P=0.001;F_(趋势)=208.750,P=0.001;F_(趋势)=512.583,P<0.001);GSK-3β、tau和p-tau(Ser396)蛋白相对表达量随铝暴露剂量的增加而上升(F_(趋势)=39.965,P<0.001;F_(趋势)=277.929,P=0.006;F_(趋势)=96.247,P=0.002)。结论miR-497-5p表达上升和Wnt3a、β-catenin表达下降,GSK-3β表达上升可能是铝暴露引起tau蛋白异常磷酸化的机制之一。Objective To investigate the effect of aluminum exposure on expression of miR-497-5p,wingless murine breast cancer virus integration site family member 3a(Wnt3a),β-catenin protein,glycogen synthase kinase-3β(GSK-3β)protein and tau protein in rat adrenal pheochromocytoma PC12 cells,so as to provide insight into unraveling the mechanisms underlying aluminum exposure-induced abnormal phosphorylation of tau protein.MethodsPC12 cells were exposed to Al(mal)3 at concentrations of 0,100,200,400μmol/L for 24 h.The viability of PC12 cells was measured using cell counting kit-8(CCK-8)assay.The relative expression of miR-497-5p and Wnt3a was detected using a realtime fluorescent quantitative PCR(RT-qPCR)assay,and the expression of Wnt3a,β-catenin,GSK-3β,P-GSK-3β(Ser9),tau and p-tau(Ser396)proteins were determined using Western blotting.ResultsThe viability of PC12 cells ap-peared a tendency towards a decline with the increase of aluminum dose(F_(trend)=323.473,P=0.001).RT-qPCR assay de-tected that the relative miR-497-5p expression appeared a tendency towards a rise with the increase of aluminum dose(F_(trend)=14.888,P=0.031),and the relative Wnt3a expression appeared a tendency towards a decline with the increase of aluminum dose(F_(trend)=165.934,P<0.001).The miR-497-5p expression negatively correlated with the relative Wnt3a ex-pression(r=-0.693,P=0.012).The expression of Wnt3a(F_(trend)=357.656,P=0.001),β-catenin(F_(trend)=208.750,P=0.001)and p-GSK-3β(Ser9)proteins(F_(trend)=512.583,P<0.001)appeared a tendency towards a decline with the increase of alu-minum dose,and the expression of GSK-3β(F_(trend)=39.965,P<0.001),tau(F_(trend)=277.929,P=0.006)and p-tau(Ser396)proteins(F_(trend)=96.247,P=0.002)appeared a tendency towards a rise with the increase of aluminum dose.Conclusion Up-regulation of miR-497-5p and GSK-3βexpression and down-regulation of Wnt3a andβ-catenin expression may be a mechanism underlying aluminum exposure-induced abnormal phosphorylation of tau protein.

关 键 词：无翅型鼠类乳腺癌病毒整合位点家族成员3a TAU蛋白 miR-497-5p 糖原合成激酶-3β Β连环蛋白 铝 阿尔茨海默病 

分 类 号：R114[医药卫生—卫生毒理学]