摘要
目的探讨百草枯(PQ)诱导下星形胶质细胞U87对SH-SY5Y细胞热休克蛋白表达的影响。方法U87细胞和SH-SY5Y细胞分别经0、100、200、300、400、500μmol/L PQ处理后,采用CCK8法检测各组细胞存活率。以Trans-well共培养方法建立U87和SH-SY5Y细胞共培养体系,细胞分为空白对照组(SH-SY5Y细胞无PQ处理)、模型对照组(SH-SY5Y细胞经300μmol/L PQ处理24 h)、共培养组(U87细胞和SH-SY5Y细胞共培养状态下经300μmol/L PQ处理24 h),采用CCK8法检测各组SH-SY5Y细胞的存活率,细胞免疫荧光化学染色法检测SH-SY5Y细胞内异常TAR DNA/RNA结合蛋白43(TDP43)聚集的定位表达,蛋白免疫印迹法检测SH-SY5Y细胞内热休克转录因子1(HSF1)、热休克蛋白70(HSP70)、热休克蛋白β8(HSPB8)、TDP43蛋白的表达水平。结果随着PQ浓度的增加,U87和SH-SY5Y细胞的存活率均逐渐降低(F=9.14、37.52,P<0.05);与对照组相比较,当PQ浓度为200μmol/L时,SH-SY5Y细胞的存活率显著降低(P<0.05),当PQ浓度为300μmol/L时,U87细胞的存活率显著降低(P<0.05)。空白对照组、模型对照组和共培养组之间的SH-SY5Y细胞存活率有显著性差异(F=22.69,P<0.01);与空白对照组相比,模型对照组SH-SY5Y细胞存活率显著降低(P<0.05);与模型对照组相比,共培养组SH-SY5Y细胞存活率显著降低(P<0.05)。空白对照组SH-SY5Y细胞中的TDP43蛋白在细胞核内表达,模型对照组SH-SY5Y细胞中的TDP43蛋白在细胞质中表达。空白对照组、模型对照组和共培养组之间的HSF1、HSP70、HSPB8和TDP43表达有显著性差异(F=6.42~28.19,P<0.05);与空白对照组相比,模型对照组SH-SY5Y细胞中HSP70、HSPB8以及TDP43表达均显著升高(P<0.05);与模型对照组相比,共培养组SH-SY5Y细胞中HSF1、HSPB8表达均显著降低(P<0.05)。结论PQ诱导下的星形胶质细胞可以抑制SH-SY5Y细胞内HSF1及HSPB8的表达,并促进SH-SY5Y细胞死亡。
Objective To investigate the effect of paraquat(PQ)-treated U87 astrocytes on the expression of heat shock protein in SH-SY5Y cells.Methods U87 cells and SH-SY5Y cells were treated with different concentrations of PQ(0,100,200,300,400,and 500μmol/L),and then CCK8 assay was used to measure the survival rate of cells.The Transwell co-culture method was used to establish the co-culture system of U87 cells and SH-SY5Y cells,and the cells were divided into blank control group(SH-SY5Y cells without PQ treatment),model control group(SH-SY5Y cells treated with 300μmol/L PQ for 24 h),and co-culture group(U87 cells and SH-SY5Y cells co-cultured with 300μmol/L PQ for 24 h).CCK8 assay was used to measure the survival rate of SH-SY5Y cells in each group;cellular immunofluorescence staining was used to measure the localization and expression of abnormal TAR DNA/RNA-binding protein-43(TDP-43)aggregation in SH-SY5Y cells;Western blotting was used to measure the protein expression of heat shock transcription factor 1(HSF1),heat shock protein 70(HSP70),heat shock protein beta-8(HSPB8),and TDP-43 in SH-SY5Y cells.Results The survival rates of U87 cells and SH-SY5Y cells gradually decreased with the increase in PQ concentration(F=9.14,37.52,P<0.01).Compared with those treated with control group,the SH-SY5Y cells treated with 200μmol/L PQ had a significant reduction in survival rate(P<0.05),and the U87 cells treated with 300μmol/L PQ had a significant reduction in survival rate(P<0.05).There was a significant difference in the survival rate of SH-SY5Y cells between the blank control group,the model control group,and the co-culture group(F=22.69,P<0.01).Compared with the blank control group,the model control group had a significant reduction in the survival rate of SH-SY5Y cells(F=22.69,P<0.01),and compared with the model control group,the co-culture group also had a significant reduction in the survival rate of SH-SY5Y cells(P<0.05).TDP-43 was expressed in the nucleus in SH-SY5Y cells in the blank control group and was expressed
作者
何晓燕
崔宝龙
李鹏飞
姚鲁清
赵芳
周畅
HE Xiaoyan;CUI Baolong;LI Pengfei;YAO Luqing;ZHAO Fang;ZHOU Chang(Department of Neurology, The Affiliated Hospital of Qingdao University, Qingdao 266003, China)
出处
《精准医学杂志》
2020年第4期339-342,346,共5页
Journal of Precision Medicine
基金
山东省自然科学基金资助项目(ZR2014HM064)。
