氟暴露下SH-SY5Y细胞ERK5信号转导通路的改变  

Changes of ERK5 signal transduction pathway in SH-SY5Y cells exposed to fluoride

作　　者：邱志伟 高峰[1,2] 陈从山 张远洋[1] 赵乐 刘艳洁[2] QIU Zhiwei;GAO Feng;CHEN Congshan;ZHANG Yuanyang;ZHAO Le;LIU Yanjie(Rehabilitation Center,Shiyan Taihe Hospital,Shiyan 442000,Hubei,China;Pathology Teaching and Research Division,School of Clinical Medicine,Guizhou Medical University,Guiyang 550004,Guizhou,China)

机构地区：[1]十堰市太和医院康复中心,湖北十堰442000 [2]贵州医科大学临床医学院病理学教研室,贵州贵阳550004

出　　处：《贵州医科大学学报》2023年第6期661-667,共7页Journal of Guizhou Medical University

基　　金：国家自然科学基金(81960572)。

摘　　要：目的检测ERK5信号通路中磷酸化的细胞外信号调节激酶5(P-ERK5)及P-ERK5在氟暴露下SH-SY5Y细胞中的表达,探讨ERK5信号通路与氟暴露下SH-SY5Y神经损伤机制的关系。方法用不同剂量氟化钠(NaF,0.000、0.100、0.200、0.300、0.400、0.500、1.000、2.000、3.000、4.000及5.000 mmol/L)处理SH-SY5Y细胞24 h及48 h,采用CCK8实验筛选出低和高剂量氟中毒浓度;再以正常组SH-SY5Y细胞为参照,用低、高剂量氟化钠剂量分别处理SH-SY5Y细胞24 h和48 h,采用实时荧光定量PCR检测处理48 h时各组细胞的ERK5 mRNA水平,用Western blot检测P-ERK5、ERK5蛋白表达水平。结果染氟处理后,0.500、1.000 mmol/L NaF处理SH-SY5Y细胞24 h及48 h细胞的存活率均低于正常对照组(P<0.05),可作为低剂量组和高剂量组的染氟剂量;染氟48 h时,对照组、低剂量组和高剂量组SH-SY5Y细胞中ERK5的mRNA表达水平分别为1.00±0.00、0.85±0.08、0.94±0.21,组间比较均无统计学意义。染氟24 h时,P-ERK5蛋白高氟组较低氟组和正常组表达升高,差异有统计学意义(P<0.05),而染氟48 h时,染氟组P-ERK5蛋白表达升高,各组间比较均有统计学意义(P<0.05),染氟24 h和48 h正常组、低氟组、高氟组P-ERK5蛋白表达水平分别为(0.20±0.05、0.28±0.04、0.50±0.18)、(0.28±0.01、0.46±0.02、0.61±0.08);染氟24 h后,总ERK5蛋白升高,高氟组较低氟组、正常组有意义(F=14.731,P<0.05),而染氟48 h后,染氟组较正常组升高有统计学意义(F=17.877,P<0.05),染氟24 h和48 h总ERK蛋白表达水平分别为(0.34±0.04、0.36±0.05、0.67±0.12),(0.55±0.02、0.72±0.03、0.78±0.08)。结论ERK5信号通路中P-ERK5蛋白及ERK5蛋白在慢性氟中毒时被激活,随着染氟时间与染氟剂量增加其表达水平出现逐渐升高,其可能参与代偿性神经保护机制对抗氟的神经毒性作用。Objective To investigate the expression of phosphorylated extracellular signal regulated kinase 5(P-ERK5)in ERK5 signal transduction pathway and extracellular signal regulated kinase 5 in SH-SY5Y cells exposed to fluoride,and to investigate the mechanism between ERK5 signal transduction pathway and nerve injury in SH-SY5Y cells exposed to fluoride.Methods SH-SY5Y cells were treated with different doses of sodium fluoride(NaF,0.000,0.100,0.200,0.300,0.400,0.500,1.000,2.000,3.000,4.000,and 5.000 mmol/L)for 24 and 48 hours respectively.The low and high doses of fluoride intoxication concentration were screened by CCK8 experiment;SH-SY5Y cells in normal group were used as reference,SH-SY5Y cells were treated with low and high doses of sodium fluoride for 24 h and 48 h respectively,ERK5 mRNA level was detected by real-time fluorescent quantitative PCR on each group at 48 h;P-ERK5 and ERK5 protein expression level was detected by Western blot.Results After fluoride treatment,the 24h and 48h survival rates of SH-SY5Y cells treated with 0.500 and 1.000 mmol/L sodium fluoride were lower than those of the normal control group(P<0.05),which could be used as the fluoride staining dose of low-dose and high-dose group.After 48 hours of fluoride exposure,the mRNA expression levels of ERK5 in three groups were 1.00±0.00,0.85±0.08,and 0.94±0.21,respectively,with no statistical significance between groups.After 24 hours of fluoride exposure,the expression of P-ERK5 protein in the high fluoride group increased more than that of low fluoride group and the normal group,difference was statistically significant(P<0.05).After 48 hours of fluoride exposure,the expression of P-ERK5 protein in the fluoride group increased,comparison among groups showed statistical significance(P<0.05).The expression levels of P-ERK5 protein in normal group,low fluoride group and the high fluoride group were(0.20±0.05,0.28±0.04,0.50±0.18;0.28±0.01,0.46±0.02,0.61±0.08)respectively after 24 and 48 hours of fluoride exposure.After 24 hours of fluoride

关 键 词：氟 暴露 SH-SY5Y ERK5 信号转导通路 改变 

分 类 号：R361[医药卫生—病理学]